In this article, the serological and molecular diagnosis of HBV has been reviewed.
Introduction
Hepatitis B virus (HBV) is a DNA virus and is transmitted through parenteral, sexual and vertical route.
Hepatitis B virus (HBV) infection is a major global health problems leading to severe liver disease such as cirrhosis and hepatocellular carcinoma (HCC). Serological markers for HBV infection
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
The recognition of serological markers helps to:
Identify patients with HBV infection.
Elucidate the natural course of chronic hepatitis B (CHB).
To assess the clinical phases of infection.
Monitor antiviral therapy.
HBsAg : appears in serum within 1 to 10 weeks of infection.
If persist > 6 months → chronic infection.
The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
Monitoring of it’s levels predicts treatment response to INF and disease progression.
Anti-HBs: a neutralizing antibody, and confers long-term immunity.
The only serological marker detected in serum after vaccination.
In past HBV infection, it is present in concurrence with anti-HBc IgG.
In most cases patients with anti-HBs antibodies are regarded as carriers of HBV. HBeAg to anti-HBe seroconversion is related to
the remission of hepatic disease
Active viral replication is sustained in some patients with HBe
seroconversion). HBcAg: is not identified in the serum.
Anti-HBc IgM and IgG: emerges 1–2 weeks
after the presence of HBsAg
After 6 months of acute infection, anti-HBc IgM wears off.
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs.
If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV
exposure.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg. Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load.
Detection of HBV DNA is a reliable marker of replication activity, and
higher titers are related to the more rapid disease progression and higher incidence of HCC.
It is useful in routine clinical setting. Principles of techniques to identify and quantify HBV DNA:
Signal amplification.
Target amplification (Real-time PCR the standard method to detect
and quantify HBV DNA in clinical setting).
HBV genotyping
HBV has a high genetic heterogeneity (ten genotype labelled A–J with distinct geographic distribution).
Can predict HBV disease progression and determine therapy.
Can be confrmed using diverse methods. Diagnosis of hepatitis B infection Acute infection:
Is a clinical diagnosis based on:
Detection of HBsAg
Symptoms
High serum aminotransferases.
Usually anti-HBc IgM and HBV DNA and HBeAg can be detected.
Chronic infection:
Is based on the persistence of HBsAg for more than 6 months.
A laboratory based diagnosis.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc. Occult infection:
Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
A serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
The detection of HBV DNA in the liver is the gold standard of diagnosis.
HBV DNA testing.
Tests for occult HBV infection are considered in:
Patients with cryptogenic liver disease
Patients considering immunosuppression therapy or
chemotherapy.
Solid organ transplantation donors. Conclusions
Serological markers for detecting antigens and antibodies against HBV is the first step for diagnosis.
Diagnosis of HBV infection is an important tool to identify
acute, chronic and occult cases of infection.
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg is the serological hallmark of HBV infection which appear after an acute exposure to HBV, within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity . In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum. In the past HBV infection, it is present in concurrence with anti-HBc IgG. Occasionally, the simultaneous appearance of HBsAg and anti-HBs has been reported in patients with HBsAg positive . In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease , however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg .
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB. Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive. It can be seen in three conditions.
1- it can be predominantly seen as IgM class during the window period of acute phase.
2- after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
3- after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg .
Molecular methods for HBV infection HBV include DNA which is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection. The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC . Furthermore, HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment (.
Hepatitis B virus (HBV) is a member of the Hepadnaviridae family and its genome consists of a circular, partially double-stranded DNA. This viral infection is responsible for the majority of chronic liver diseases worldwide and can be transmitted through various routes, including parenteral, sexual, and vertical transmission. HBV can be classified into ten genotypes, designated as A to J, with genotypes A and D being the most prevalent in Africa and Europe. Acute infections caused by genotypes A and D are more likely to progress to chronicity. Genotype C is often associated with an increased risk of perinatal transmission and is linked to severe liver diseases such as cirrhosis and hepatocellular carcinoma (HCC). When it comes to interferon therapy, patients infected with genotypes A and B generally exhibit better treatment responses compared to those infected with genotypes C and D.
Serological markers are used to diagnose and monitor hepatitis B virus (HBV) infection. One of the primary markers is HBsAg, which appears in the serum within 1 to 10 weeks after infection. If HBsAg persists for more than 6 months, it indicates chronic HBV infection. On the other hand, the presence of anti-HBs, a neutralizing antibody, signifies long-term immunity. In individuals who have acquired immunity through vaccination, only anti-HBs is detected in the serum. In cases of past HBV infection, anti-HBs is present alongside anti-HBc IgG.
HBeAg and anti-HBe markers were previously used to assess infectivity and viral replication, but they have been replaced by HBV DNA testing. Another set of markers, anti-HBc IgM and IgG, emerge within 1-2 weeks after the presence of HBsAg, along with elevated serum aminotransferase levels and symptoms. After 6 months of acute infection, anti-HBc IgM disappears, but anti-HBc IgG continues to be detected in individuals with resolved HBV infection.
HBV DNA is a direct measurement of the viral load and serves as a reliable marker of replication activity. Higher titers of HBV DNA are associated with more rapid disease progression and an increased risk of hepatocellular carcinoma (HCC). HBV DNA testing is also used to monitor the response to antiviral treatment. By tracking changes in HBV DNA levels, healthcare providers can assess the effectiveness of antiviral therapy in controlling the infection.
Hepatitis B infection can be diagnosed based on different clinical scenarios. In the case of acute hepatitis B, diagnosis is made by detecting the presence of HBsAg, along with symptoms and elevated serum aminotransferases. Additionally, anti-HBc IgM antibodies can usually be detected, and HBV DNA is present in the bloodstream. Chronic hepatitis B is diagnosed when HBsAg persists for more than 6 months. This indicates an ongoing infection and the need for long-term management and monitoring.Past hepatitis B infection is identified by the coexistence of anti-HBs (antibodies against HBsAg) and anti-HBc IgG (antibodies against core antigen). This indicates prior exposure to the virus and the development of immunity. Occult HBV infection is a unique scenario characterized by the persistence of a low level of intrahepatic HBV DNA and the presence of isolated anti-HBc antibodies, with the absence of both HBsAg and anti-HBs antibodies in the blood. The gold standard for diagnosing occult HBV infection is detecting HBV DNA in liver tissue samples. However, serum HBV DNA testing using real-time PCR has become the preferred method for diagnosis.
HBV is a double stranded DNA with different serological markers and is one of the major health problem globally leading to liver cirrhosis and hepatocellular carcinoma HCC. There are variable presentation of hepatitis B -acute, chronic and occult cases. Sexual, parenteral, or vertical route are the modes of transmission.Early diagnosis and treatment will help in eradicating the virus. North American, and Western Europe have the lowest prevalence of hepatitis B Infection-around 0.5 -2% while highest prevalence countries are China, and South East Asia-affecting around > 8%.10 genotypes of hepatitis are known- A- J. Serological Markers
Earliest serological marker to appear in serum is HBsAg (1-2 weeks of exposure).Chronic hepatitis B infection is labeled as having HbsAg in serum for more than 6 months. anti-HBs – marker of long term immunity and is a neutralizing antibody .HBeAg and anti-HBe –marker of infectivity and viral replication. Anti-HBc IgM and IgG –IgM appears in blood after 1-2 week of appearance of HbsAg and disappears after 06 months. However, IgG appears in blood life long even after resolved HBV infection and chronic hepatitis B. Molecular Methods for HBV Infection:
HBV PCR DNA (quantitative)-detected by Signal amplification,or Target amplification-indicates active infection and quantitative test is better than qualitative test
Acute hepatitis B: HBsAg +, HBV DNA+, anti-HBc IgM and HBeAg +/-ve
Chronic infection: HBsAg + for more than 6 months.
Past HBV infection: anti-HBs +, IgG anti-HBc+ HBsAg-ve , and HBV DNA PCR-ve.
Occult HBV infection : HBV DNA +ve ,HBsAg –ve , anti-HBc IgG +ve CONCLUSION:
This article deals with the diagnostic strategies both serological and molecular which ill help in differentiating between acute, chronic and occult infection.
Its review article was published in the Annals of Translational Medicine 2016 Diagnosis of HBV 1- Serological markers for HBV infection (antigen or antibodies screening in the serum). 2- Molecular methods for HBV infection by PCR 3- HBV genotyping Serological markers for HBV infection HBsAg: It can be detected in serum within 1 to 10 weeks after infection and usually disappears after 6 months but if it persists for more than 6 months indicates chronic HBV infection Anti-HBs: It is a neutralizing antibody that indicates long-term immunity. After vaccination, it is the only serological marker detected in serum but after past exposure to HBV infection, it will be associated with anti-HBc IgG. HBeAg and anti-HBe: it’s related to infectivity and viral replication. HBcAg: It is an intracellular antigen and not seen in the blood. anti-HBc IgM and IgG: after acute infection, anti-HBc IgM and IgG were seen 1–2 weeks after HBsAg. anti-HBc IgM starts to wear off after 6 months on the other hand Anti-HBc IgG continues to detect in both patients with resolved HBV infection and chronic hepatitis B (never got by vaccination). Molecular Methods for HBV Infection HBV DNA reveals the replication activity of the virus. It can be detectable during the first-month post-infection then gradually it decreases in chronic infection or completely disappears if recovery occurred. HBV genotyping HBV genotyping: can be done by Reverse hybridization, Genotype specific PCR and Sequence analysis. It can be divided into ten genotypes, labeled A–J. Genotype B and C are only seen in Oceania and Asia, whereas genotype A and D are the most common genotypes in Africa and Europe with higher rate of chronicity but higher response to interferon therapy. Genotype C is generally associated with perinatal infection and is more aggressive in the form of cirrhosis and HCC. Diagnosis of hepatitis B infection Acute hepatitis B: HBsAg positive, HBV DNA is positive, anti-HBc IgM and HBeAg can also be positive. chronic infection: HBsAg positive and HBV DNA PCR (for more than 6 months) Past HBV infection: anti-HBs positive, IgG anti-HBc positive HBsAg negative, and HBV DNA PCR negative. Occult HBV infection: low level of intrahepatic HBV DNA (gold standard but invasive), HBsAg negative, anti-HBc IgG positive Occult HBV infection has some clinical importance. 1- First, risk of transmission by solid organ transplantation (especially liver). 2- Second, risk of reactivation of HBV infection after starting immunosuppression. 3- Third, risk of LCF and HCC. Liver tissue (biopsy) PCR is indicated if all the following are present
Hepatic dysfunction of unknown cause.
Isolated anti-HBc Ab positivity with negative blood PCR.
Summarize the article;; Introduction;
HBV infection is the most common cause of the liver failure, and a major global health problem in developing countries. Serologic marker for HBV infection;
How differentiate serologic markers, and different stages of infectivity and activity, like,
HBs Ag, marker for infection, will positive within 8-10 weeks, and persist for 6 months if converts into chronicity, being used as marker for treatment monitoring.
Anti-HBs, shows immunity or previous infection, vaccination. It’s known as neutralizing antibody.
HBeAg, active infection.
Anti-HBe antibody,
Anti-HBc IgM and IgG. Molecular methods for diagnosis of Hepatitis B infection;
HBV DNA is a direct measurement of the viral load, shows the active infection, detected in early phase of infection.
Two principles 1, Signal amplification,
2, Target amplification.
Standard method for detection is HBV DNA quantification. Genotyping;
There are ten genotypes.
Labeled as A- J. done to predict the treatment. Diagnosis of Hepatitis B infection;
Acute hepatitis is a clinical diagnosis, and confirmed by serological testing.
For diagnosis of chronic infection if HBVs-Ag persist for more than 6 months.
Past infection HBV-s Ab, and HBVc- Ab.
HBV DNA is gold standard for detection of infection. Conclusion;
In this article the aim was to give guidance for detection, diagnosis, to differentiate B/W acute and chronic and when to take action.
Summarize the article; Introduction;
HBV infection is the most common cause of the liver failure, and a major global health problem in developing countries. Serological markers for HBV infection;
How differentiate serologic markers, and different stages of infectivity and activity, like,
HBs Ag, marker for infection, will positive within 8-10 weeks, and persist for 6 months if converts into chronicity, being used as marker for treatment monitoring.
Anti-HBs, shows immunity or previous infection, vaccination. It’s known as neutralizing antibody.
HBeAg, active infection.
Anti-HBe antibody,
Anti-HBc IgM and IgG. Molecular methods for Diagnosis;
HBV DNA is a direct measurement of the viral load, shows the active infection, detected in early phase of infection.
Two principles 1, Signal amplification,
2, Target amplification.
Standard method for detection is HBV DNA quantification. Genotyping;
There are ten genotypes.
Labeled as A- J. done to predict the treatment. Diagnosis of hepatitis B infection;
Acute hepatitis is a clinical diagnosis, and confirmed by serological testing.
For diagnosis of chronic infection if HBVs-Ag persist for more than 6 months.
Past infection HBV-s Ab, and HBVc- Ab.
HBV DNA is gold standard for detection of infection. Conclusions;
In this article the aim was to give guidance for detection, diagnosis, to differentiate B/W acute and chronic and when to take action.
The article discusses the diagnosis of acute, chronic, and occult hepatitis B virus (HBV) infections through serological and molecular methods. Key points from the article include:
HBV is a member of the Hepadnaviridae family, and its genome consists of four open reading frames overlapped, including surface proteins, hepatitis B e antigen, polymerase including reverse transcriptase, and a transcriptional transactivator factor.
The covalently closed circular DNA (cccDNA) is the transcriptional template of HBV and stays inside the hepatocyte nucleus as a mini-chromosome.
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG, which can identify patients with HBV infection, elucidate the natural course of chronic hepatitis B, assess the clinical phases of infection, and monitor antiviral therapy.
HBsAg is the serological hallmark of HBV infection, and its persistence for more than 6 months implies chronic HBV infection.
Anti-HBs is a neutralizing antibody, confers long-term immunity, and is the only serological marker detected in serum in patients with acquired immunity through vaccination.
HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum, and anti-HBc IgM and IgG can emerge during acute infection and wear off after 6 months.
PCR-based molecular methods have greatly improved HBV detection sensitivity and quantification and are useful in diagnosing occult HBV infection.
Introduction:
Hepatitis B virus (HBV) infection is a major global health problems leading to severe liver disease such as cirrhosis and hepatocellular carcinoma.
Significant method to diminish the burden of this disease is timely diagnosis of acute, chronic and occult cases of HBV. Serological markers for HBV infection:
Consist of HBsAg, Anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG. HBsAg:
Is the serological hallmark of HBV infection.
Appears in serum within 1 to 10 weeks.
Persistence of this marker for more than 6 months implies chronic HBV infection. Anti-HBs:
Is known as a neutralizing antibody, and confers long-term immunity.
Vaccination, anti-HBs is the only serological marker detected in serum.
Past HBV infection, it is present in concurrence with anti-HBc IgG.
As carriers of HBV : HBsAg and anti-H HBsAg has been reported in patients. HBeAg and anti-HBe: Know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay. HBcAg :
Is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. Anti-HBc IgM and IgG:
Emerges in Acute infection, 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB. Isolated anti-HBc positive: negative HBsAg and HBs ab, seen in three conditions:
First, as IgM class during the window period of acute phase.
Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
Anti-HBc IgM should be done and HBV DNA checked. Molecular methods for HBV infection:
One month after HBV infection.
A reliable marker of replication activity,
Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment. HBV genotyping:
HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution.
The HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C .Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
The interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
Genotype B or C had a lower opportunity to gain serologic response to tenofovir. Diagnosis of hepatitis B infection: Acute hepatitis B:
Symptoms, high serum aminotransferases. HBsAg, POSTIVE, anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase. Chronic infection:
The persistence of HBsAg for more than 6 month. Past HBV infection:
AntiHBs AB and IgG anti-HBc. Occult HBV infection:
HBV DNA without detectable HBsAg.
Isolated anti-HBc with the absence of HBsAg and antiHBs antibody. Occult HBV infection has some clinical importance.
First, it can be transmitted via transfusion, solid organ transplantation including orthotropic liver transplantation, or hemodialysis
. Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
. Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
Forth, it appears to be a risk factor for HCC. Occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease, especially when having anti-HBc in serum;
Patients considering immunosuppression therapy or chemotherapy;
In solid organ transplantation donors.
Serological markers for HBV infection consist of HBsAg anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection(CHB).The serum HBsAg titers are higher in patients with HBeAgpositive CHB than in HBeAg-negative CHB . Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAgnegative CHB patients with normal serum alanine aminotransferase levels
.Anti-HBs is known as a neutralizing antibody, and confers long-term immunity.In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum. In the past HBV infection, it is present in concurrence with anti-HBc IgG
.
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease . however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB. HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg .
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.As the prevalence of serologically negative HBV infection (HBeAg-negative CHB and occult HBV infection) has increased, HBV-DNA detection has obtained more awareness in clinical medicine The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC .
There are two principles of techniques to identify and quantify HBV DNA: signal amplification such as hybrid capture and branched DNA technology; target amplifcation such as polymerase chain reaction (PCR) (19,22). Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting.
HBV genotyping
According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution .Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy. HBV genotyping can be confrmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fuorescence polarization assay.
Occult HBV infections–
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg . It is a serological situation defned by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody . Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases; thus, HBV DNA testing is widely used to diagnose occult HBV infection.
Occult HBV infection has some clinical importance. It can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis . Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state . Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis,Tests for occult HBV infection are considered in the following conditions: in patients with cryptogenic liver disease, especially when having anti-HBc in serum; in patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission.
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family. It shows a diameter of 30–42 nm and consists of outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein.
Viral capsid bears viral genome and DNA polymerase that has reverse transcriptase activity. HBV genomes comprise a circular, partly double-stranded DNA and has four open reading frames overlapped: (I) S that encodes for surface proteins (HBsAg); (II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg); (III) P for polymerase including reverse transcriptase; (IV) X that encodes for a transcriptional transactivator factor (HBxAg)
Serological markers for HBV infection
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG. The identification of serological markers allows: to identify patients with HBV infection; to elucidate the natural course of chronic hepatitis B (CHB); to assess the clinical phases of infection; and to monitor antiviral therapy.
HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
Several studies have reported the association between transcription activity of cccDNA in the liver and serum HBsAg levels.
Differences in the serum HBsAg levels during the different phases of infection indicate the distribution of cccDNA during the respective phases of the disease. The serum HBsAg titers are higher in patients with HBeAg- positive CHB than in HBeAg-negative CHB.
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg- negative CHB patients with normal serum alanine aminotransferase levels.
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
HBV genotyping
HBV has a high genetic heterogeneity because it reproduces via a reverse transcriptase that has insufficient proofreading.
capability. According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution
Genotype B and C are restricted to Oceania and Asia, whereas genotype A and Dare omnipresent but most common in Africa and Europe
Genotype I is unusual and can be observed in Vietnam, Laos, India and China, while genotype J has been reported in Japan and Ryukyu
Other genotypes such as E, F, G, and H are also occasionally found in Asia.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually, anti-HBc IgM can be detected, and HBV DNA is present. HBeAg can also be identified in most acute phase of infections but has little clinical importance. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
Occult HBV infection has some clinical importance. First, it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis.
Conclusions
In this article, we aimed to give informations about HBV serological and molecular diagnosis. First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus. In order to verify first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used. Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
Introduction
HBV belongs hepadnaviridae family.
The reverse transcriptase responsible for viral replication is susceptible to errors thus high mutation rate.
HBV is transmitted parenteral, sexual and vertical route.
HBV is divided into three endemicity: high, intermediate and low; with high endemicity in sub saharan Africa and south east Asia. Serological markers
HBsAg is the serological hallmark of HBV infection.
After acute infection it appears within 1-10 weeks and persistence beyond 6 months indicates chronic infection.
Anti-HBs is a neutralising antibody, confers longterm immunity.
Vaccinated patients only have anti-HBs.
Anti-HBs may appear simultaneous with HBsAg in carriers.
HBeAg and anti-HBe prior used to indicate replication and infectivity however has been replaced by HBV DNA.
HBeAg seroconversion to Anti-HBe indicates remission.
HBcAg is an intracellular antigen thus not detected in serum.
After acute infection IgM Anti-HBc appears within 1-2 weeks, and wears off in 6 months.
IgG Anti-HBc indicates chronic and resolved infection.
Anti-HBc may occur isolated without HBsAg and in the absence of AntiHBs.
Molecular methods
HBV DNA indicates the replication of the virus.
Detected 1 month after acute infection, peaks in 3 months then declines in chronic infection.
High titers are associated with more rapid progression and development of hepatocellular carcinoma.
Useful to determine patients who are in need of antiviral therapy and to monitor response to treatment.
HBV genotype
There are 10 genotypes A-J with distinct geographical distribution.
Genotype A and D associated with higher rates of chronicity than B and C.
Genotype A and C have better response to interferon than B and D. Diagnosis
Acute hepatitis B infection is a clinical diagnosis based by the presence of HBsAg detection, raised serum aminotransferase and symptoms.
IgM anti-HBc may be present.
HBV DNA may be detectable.
Detection of HBeAg has no clinical significance.
Chronic hepatitis detected by persistence of HBsAg beyond 6 months.
Past infection identified by Anti-HBs and IgG anti-HBc antibodies.
Occult infection defined by detection of HBV DNA intra-hepatic.
Presence of Anti-HBc without HBsAg or anti-HBs.
Can be transmitted via solid organ transplantation, hemodialysis, or transfusion.
Testing should be done in patients with cryptogenic liver disease with anti-HBc, and donors for solid organ transplant.
Summarise the Article Diagnosis of hepatitis B Introduction Hepatitis B virus (HBV) infection is a major global health problem leading to severe liver disease, cirrhosis and hepatocellular carcinoma (HCC). It is transmitted by sexual, parenteral and vertical route, infecting 240 million people worldwide. Depending on prevalence in general population and transmission rate, geographically categorised in to 3 endemic zones – high endemic areas (>8% prevalence – China, SE Asia, Indonesia, sub-Saharan Africa); intermediate endemic regions (2-7% prevalence – South America, South West Asia, Eastern and Southern Europe); low endemic regions (0.5-2% prevalence – North America and Western Europe). One significant method to diminish the burden of this disease is timely diagnosis of acute, chronic and occult cases of HBV.
HBV diagnosis involves screening using serological markers detecting antigens and antibodies, which is then verified by molecular tests like DNA-PCR, viral load quantification and genotyping.
The serological and molecular tests for diagnosis of HBV infection are reviewed in this article. HBV structure: circular, double-stranded DNA virus, has 4 open reading frames of genes overlapped, which code for various protein antigens. (I) S encodes for surface proteins (HBsAg) (II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg) (III) P for polymerase including reverse transcriptase (IV) X that encodes for a transcriptional trans-activator factor (HBxAg) Serological markers for HBV infection Hepatitis B surface antigen (HBsAg) and anti-HBs, anti-HBc IgM and IgG, and hepatitis B e antigen (HBeAg) and anti-HBe – these antigens and antibodies appear in serum of infected individual in a particular time frame and can be used as serological markers. HBsAg – hallmark of HBV infection, appears in serum in 1-10 weeks – Its persistence for more than 6 months implies chronic HBV infection. HBsAg titres are higher in patients with HBeAg + CHB – Its levels predict treatment response and progression in HBeAg-negative CHB patients Anti-HBs – indicates long-term immunity, and will be the only marker detected in serum post vaccination. – anti-HBs antibodies are unable to neutralize the circulating viruses (HBV carriers). HBcAg is intracellular, present in infected hepatocyte, thus not detected in serum. HBeAg to anti-HBe seroconversion is related to remission of HBV infection. In case of mutations in the pre-core and core region active viral replication is sustained in spite of seroconversion.
During acute infection, anti-HBc (IgM > IgG) rise 1–2 weeks after the appearance of HBsAg, associated with raised serum aminotransferase and clinical symptoms. IgM anti-HBc disappear after 6 months of acute infection, but IgG Anti-HBc continues to detect in both patients with resolved HBV infection and CHB. Isolated Anti-HBc antibody positivity can be seen 1) during the window period of acute phase
2) after the end of acute infection
3) after many years of chronic HBV infection, when HBsAg may be undetectable, indicating a previous infection
Molecular methods for HBV Detection – HBV-DNA detection by PCR – detectable 1 month after infection, increases to peak level by 3 months quantification of viral load reveals rate of viral replication – higher titre indicates rapid disease progression, associated with higher incidence of HCC. – HBV DNA titre can be used as guide for antiviral therapy and monitoring response HBV genotyping – 10 genotypes labelled A–J, are identified.
Genotype B & C – are restricted to Oceania and Asia.
Genotype A & D – are omnipresent but most common in Africa and Europe.
Genotypes A & D – more liable to chronicity than genotypes B and C.
Genotype C has risk perinatal infection, and related to severe liver disease, including cirrhosis and HCC.
Genotypes A & B have better response to Interferon than genotypes C & D.
Patients with genotype B &C shows lower serological response to tenofovir.
Diagnosis of hepatitis B infection Acute hepatitis: HBsAg positive + clinical symptoms + elevated AST & ALT Usually, anti-HBc IgM can be detected and HBV-DNA is present. Chronic hepatitis-B: persistence of HBsAg for more than 6 months.
Diagnosed by laboratory means but not by clinical presentations.
Past HBV infection: defined by coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection: – persistence of low level of intrahepatic HBV DNA without detectable HBsAg. – defined as presence of isolated anti-HBc with absence of HBsAg, anti-HBs antibody. – HBV-DNA PCR can be used for diagnosis of occult HBV infection. The importance of occult HBV infection: – transmitted via transfusion, solid organ transplantation, or haemodialysis – reactivation of HBV infection may occur in patients receiving chemotherapy, immunocompromised state – it may accelerate liver injury, lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection – it could be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis Indications for testing occult HBV infection: – Patients with cryptogenic liver disease, mainly those with + anti-HBc in serum – in patients on chemotherapy or immunosuppression therapy – in solid organ transplantation donors, due to the possibilities for transmission
Introduction:
It is a double strand DNA virus and belong to e Hepadnaviridae family.
Viral capsid bears viral genome and DNA polymerase that has reverse transcriptase activity.
HBV genome comprise a circular, partly double-stranded DNA and has four open reading frames overlapped:
1) S that encodes for surface proteins (HBsAg);
2) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg);
3) P for polymerase including reverse transcriptase;
4) X that encodes for a transcriptional trans activator factor (HBxAg)
It has high, intermediate and low endemicity.
Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection. Serological markers for HBV infection:
Consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG
The identification of serological markers allows: to identify patients with HBV infection; to elucidate the natural course of chronic hepatitis B (CHB); to assess the clinical phases of infection; and to monitor antiviral therapy. HBsAg:
It appears in the serum within 1 to 10 weeks and indicates acute infection and persist up to 6 months and indicates chronic infection and its titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB. Anti-HBs:
Is known as a neutralizing antibody, and confers long-term immunity and in acquired immunity through vaccination, it is the only serological marker detected in serum.
In the past HBV infection, it is present in concurrence with anti-HBc IgG.
In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, so considered as carriers of HBV. HBeAg and anti-HBe:
In the past used to know infectivity and viral replication, but nowadays replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion. HBcAg:
It is not detected in serum because it intracellular. Anti-HBc IgM and IgG:
Appears 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms and it wear off after 6 months of acute infection.
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Isolated anti-HBc positive is considered as HBsAg-negative and positive anti-HBc IgG without anti-HBs and for that anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
In chronic liver disease patients HBV DNA should be tested to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg. Molecular methods for HBV infection:
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level at 3 months after the exposure to HBV and then gradually decrease in chronic infection or disappears at the recovery from HBV infection.
The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
There are two principles of techniques to identify and quantify HBV DNA:
1) Signal amplification such as hybrid capture and ranched DNA technology.
2) Target amplification such as (PCR). Real-time PCR is standard method to detect and quantify HBV DNA in clinical setting HBV genotyping:
It has a high genetic heterogeneity and divided into ten genotypes, labelled(A-J) which have distinct geographic distribution.
Acute infection with genotypes A and D leads to higher rate of chronicity
than genotypes B and C. Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
HBV genotyping can be confirmed using diverse methods:
1) Reverse hybridization.
2) Genotype-specific PCR assays.
3) Real-time PCR.
4) Restriction fragment-length polymorphism.
5) Sequence analysis.
6) Microarray (DNAChip).
7) Fluorescence polarization assay. Diagnosis of hepatitis B infection: Acute infection:
Identified by:
1) The detection of HBsAg, symptoms, high serum aminotransferases.
2) anti-HBc IgM Anti-HBc IgM can be detected and HBV DNA is present
3) HBeAg can also be identified in most acute phase of infections, but has little Chronic infection: Characterized by:
1) Persistence of HBsAg for more than 6 months.
2) Patients diagnosed by laboratory means but not by clinical presentations.
3) Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection: characterized by:
1. Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
2. Presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
3. The detection of HBV DNA in the liver is the gold standard of diagnosis
4. Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases
Occult HBV infection has some clinical importance:
A. Can be transmitted via transfusion, solid organ transplantation.
B. Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
C. Accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
D. Appears to be a risk factor for HCC.
Tests for occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease; in patients considering IS therapy or chemotherapy and in SOT donors.
The aim of this study: to review serological and molecular diagnosis of HBV.
# Introduction:
*Hepatitis B virus (HBV) belongs to the Hepadnaviridae family and it is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
*HBV is a circular, partly double-stranded DNA virus with various serological markers.
*Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection. # Serological markers for HBV infection:
*Mainly consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
*The identification of serological markers allows:
1) Identify patients with HBV infection.
2) Elucidate the natural course of chronic hepatitis B.
3) Assess the clinical phases of infection.
4) Monitor antiviral therapy. HBsAg:
*Is the serological hallmark of HBV infection, in acute infection it is appears in serum within 1 to 10 weeks.
*Persistence of HBsAg for more than 6 months indicates chronic infection.
*The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
*Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels Anti-HBs:
*Known as a neutralizing antibody, and confers long-term immunity.
* In acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
*In the past HBV infection, it is present in concurrence with anti-HBc IgG.
*In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, so considered as carriers of HBV. HBeAg and anti-HBe:
*Previously used to know infectivity and viral replication, but recently replaced by HBV DNA assay. *HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion. HBcAg:
*Is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. Anti-HBc IgM and IgG:
*It is emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
* After 6 months of acute infection, anti-HBc IgM wears off.
* Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
*In HBsAg-negative individuals and positive anti-HBc IgG without anti-HBs, considered isolated anti-HBc positive.
*If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
*HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg. # Molecular methods for HBV infection
*HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
*It is detectable at the early stage of infection and increases up to peak level at 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
*The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
*HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
*There are two principles of techniques to identify and quantify HBV DNA:
1) Signal amplification such as hybrid capture and ranched DNA technology.
2) Target amplification such as (PCR). Real-time PCR is standard method to detect and quantify HBV DNA in clinical setting # HBV genotyping:
*HBV has a high genetic heterogeneity.HBV can be divided into ten genotypes, labelled A–J: they have
distinct geographic distribution
*Acute infection with genotypes A and D leads to higher rate of chronicity
than genotypes B and C. Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
*HBV genotyping can be confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray and fluorescence polarization assay. # Diagnosis of hepatitis B infection: Acute infection
*By the detection of HBsAg, symptoms, high serum aminotransferases.
*Anti-HBc IgM can be detected and HBV DNA is present.
* HBeAg can also be identified in most acute phase of infections, but has little clinical importance. Chronic infection:
*Persistence of HBsAg for more than 6 months.
*Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
* Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc. Occult infection:
*Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
*Presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
*The detection of HBV DNA in the liver is the gold standard of diagnosis
*Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases
*Occult HBV infection has some clinical importance.
1) Can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
2) Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
3) Accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4) Appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic
inflammation and fibrosis.
*Tests for occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease; in patients considering IS therapy or chemotherapy and in SOT donors.
● Viral capsid bears viral genome and DNA polymerase that has reverse transcriptase activity.
● HBV genome comprise a circular, partly double-stranded DNA and has four open reading frames overlapped:
(I) S that encodes for surface proteins (HBsAg)
(II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg)
(III) P for polymerase including reverse transcriptase
(IV) X that encodes for a transcriptional transactivator factor (HBxAg)
● HBV infection is transmitted through parenteral, sexual and vertical route.
● About 240 million people are chronically infected by HBV, so have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC).
● HBV endemicity is divided into three categories;
☆ high
China, South East Asia, Indonesia, and sub-Saharan Africa
☆ intermediate
South America, South West Asia, Eastern and Southern Europe.
☆ low.
North America and Western Europe
● HBsAg is the serological hallmark of HBV infection.
● After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks.
● Persistence of this marker for more than 6 months implies chronic HBV infection
● The serum HBsAg titers are higher in patients with HBeAg-positive CHB than in HBeAg-negative CHB
● Anti-HBs is known as a neutralizing antibody, and confers long-term immunity
● Anti-HBs is the only serological marker detected in serum In patients with acquired immunity through vaccination
● The simultaneous appearance of HBsAg
and anti-HBs has been reported in patients with HBsAg positive these patients are regarded as carriers of HBV.
● HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease
● During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
● HBsAg-negative with positive HBc IgG without anti-HBs It can be seen in :
☆ The window period of acute phase
☆ Decreased anti-HBs below cutoff level of detection after acute infection had ended
☆ After several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
● So anti-HBc IgM should be checked to assess the possibility of recent HBV
● Occult HBV infection : detectable HBV DNA without serum HBsAg
● Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
● Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 10^7–10^8 IU/mL).
HBV genotyping
● HBV can be divided into ten genotypes, labelled A–J
● Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
● Genotype C generally is considered as a risk factor for perinatal infection and related cirrhosis and HCC
● Genotypes A and B have better therapy response than genotypes C and D
● Genotype B or C had a lower opportunity to gain serological response to tenofovir
Diagnosis of hepatitis B infection
● Acute hepatitis B is identified:
☆ HBsAg symptoms, high serum ALT .
☆ Anti-HBc IgM can be detected and HBV DNA is present.
☆ HBeAg has little clinical importance.
● Chronic infection is based on :
☆ Persistence of HBsAg for more than 6 M
● Past HBV infection is defined : coexistence of anti-HBs and IgG anti-HBc.
● Occult HBV infection is defined :
☆ persistence of low level of intrahepatic HBV DNA without detectable HBsAg
☆ Presence of isolated anti-HBc with the absence of HBsAg and anti-HBs antibody
● Clinical importance of Occult HBVi :
☆ It can be transmitted via transfusion, SOT including orthotopic liver transplantation or hemodialysis
☆ Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
☆ It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
☆ it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fbrosis
● Tests for occult HBV infection are considered in :
☆ Patients with cryptogenic liver disease, especially when having anti-HBc in serum
☆ patients considering immunosuppression therapy or chemotherapy
☆ solid organ transplantation donors, due to the possibilities for transmission
Diagnosis of hepatitis B Introduction
· HBV genome comprise a circular, partly double-stranded DNA and has four open reading frames overlapped:
(I) S that encodes for surface proteins (HBsAg);
(II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg);
(III) P for polymerase including reverse transcriptase;
(IV) X that encodes for a transcriptional transactivator factor (HBxAg)
· HBV infection is the cause of most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
· 240 million people are chronically infected by HBV.
· high endemic areas for HBV are: China, South East Asia, Indonesia, and sub-Saharan Africa
· Low endemic regions: North America and Western Europe
Serological markers for HBV infection
· HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
· HBsAg is the hallmark of HBV infection, appears in serum within 1 to 10 weeks, and its persistence for more than 6 months implies chronic HBV infection.
· HBsAg titers are higher in patients with HBeAg + CHB, and its levels predicts treatment response and progression in HBeAg-negative CHB patients
· Anti-HBs indicates long-term immunity, and will be the only marker detected in serum post vaccination.
· In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses (HBV carriers).
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but in case of mutations in the pre-core and core region active viral replication is sustained in spite of seroconversion.
· HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
· During acute infection, anti-HBc IgM and IgG occurs 1–2 weeks after the appearance of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM disappears, but Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
· Isolated anti-HBc positive can be seen in:
1) during the window period of acute phase
2) after the end of acute infection
3) after many years of chronic HBV infection, HBsAg may became undetectable
Molecular methods for HBV infection
· HBV DNA is measures the viral load, which reveals the replication activity of the virus.
· Can be detectable 1 month after HBV infection and increases up to peak level nearly 3 months after the exposure to HBV.
· HBV-DNA detection is a reliable marker of replication activity, and higher titers indicates rapid disease progression and higher incidence of HCC.
· HBV DNA testing is an indication to determine patients who need antiviral therapy and monitor them for suitable treatment.
HBV genotyping
· HBV can be divided into ten genotypes, labelled A–J.
· Genotype B and C are restricted to Oceania and Asia.
· Genotype A and D are omnipresent but most common in Africa and Europe.
· Genotype I is unusual and can be observed in Vietnam, Laos, India and China, while genotype J has been reported in Japan and Ryukyu.
· Other genotypes such as E, F, G, and H are also occasionally found in Asia.
· Acute infection with genotypes A and D leads more liable to chronicity than genotypes B and C.
· Genotype C has a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
· Patients with genotype B or C had a lower opportunity to gain serological response to tenofovir.
Diagnosis of hepatitis B infection Acute hepatitis B
· Diagnosed by + HBsAg, symptoms, high serum aminotransferases.
· Usually, anti-HBc IgM can be detected and HBV DNA is present. Chronic hepatitis B
· Diagnosed by the persistence of HBsAg for more than 6 months.
· Diagnosed by laboratory means but not by clinical presentations. Past HBV infection
· defined by the coexistence of anti-HBs and IgG anti-HBc. Occult HBV infection
· defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
· defned by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
· Since hepatic HBV DNA is invasive PCR for serum HBV DNA can be used for diagnosis of occult HBV infection.
The importance of occult HBV infection:
1) it can be transmitted via transfusion, solid organ transplantation, or hemodialysis
2) reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3) it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4) it could be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis
Indications for occult HBV infection testing:
1) patients with cryptogenic liver disease, mainly those with + anti-HBc in serum;
2) in patients’ pre- immunosuppression therapy or chemotherapy
3) in solid organ transplantation donors, due to the possibilities for transmission
# The aim of this study: to review serological and molecular diagnosis of HBV.
# Introduction:
*Hepatitis B virus (HBV) belongs to the Hepadnaviridae family and it is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
*HBV is a circular, partly double-stranded DNA virus with various serological markers.
*Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection.
# Serological markers for HBV infection:
*Mainly consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
*The identification of serological markers allows:
1) Identify patients with HBV infection.
2) Elucidate the natural course of chronic hepatitis B.
3) Assess the clinical phases of infection.
4) Monitor antiviral therapy.
HBsAg:
*Is the serological hallmark of HBV infection, in acute infection it is appears in serum within 1 to 10 weeks.
*Persistence of HBsAg for more than 6 months indicates chronic infection.
*The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
*Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs:
*Known as a neutralizing antibody, and confers long-term immunity.
* In acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
*In the past HBV infection, it is present in concurrence with anti-HBc IgG.
*In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, so considered as carriers of HBV.
HBeAg and anti-HBe:
*Previously used to know infectivity and viral replication, but recently replaced by HBV DNA assay. *HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion.
HBcAg:
*Is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
Anti-HBc IgM and IgG:
*It is emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
* After 6 months of acute infection, anti-HBc IgM wears off.
* Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
*In HBsAg-negative individuals and positive anti-HBc IgG without anti-HBs, considered isolated anti-HBc positive.
*If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
*HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
# Molecular methods for HBV infection
*HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
*It is detectable at the early stage of infection and increases up to peak level at 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
*The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
*HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
*There are two principles of techniques to identify and quantify HBV DNA:
1) Signal amplification such as hybrid capture and ranched DNA technology.
2) Target amplification such as (PCR). Real-time PCR is standard method to detect and quantify HBV DNA in clinical setting
# HBV genotyping:
*HBV has a high genetic heterogeneity.HBV can be divided into ten genotypes, labelled A–J: they have
distinct geographic distribution
*Acute infection with genotypes A and D leads to higher rate of chronicity
than genotypes B and C. Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
*HBV genotyping can be confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray and fluorescence polarization assay.
# Diagnosis of hepatitis B infection: Acute infection
*By the detection of HBsAg, symptoms, high serum aminotransferases.
*Anti-HBc IgM can be detected and HBV DNA is present.
* HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
Chronic infection:
*Persistence of HBsAg for more than 6 months.
*Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
* Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection:
*Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
*Presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
*The detection of HBV DNA in the liver is the gold standard of diagnosis
*Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases
*Occult HBV infection has some clinical importance.
1) Can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
2) Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
3) Accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4) Appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic
inflammation and fibrosis.
*Tests for occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease; in patients considering IS therapy or chemotherapy and in SOT donors.
DIAGNOSIS OF HBV
SEROLOGY
HBV is commonest cause of chronic hepatitis in world
prevalance – high S-E asia, china , africa, intermediate – europe , low – Northamericca and western europe
markers
HBsAg,anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg – main marker
positive after 8-10 week
if persistant after 6 montsh – chronic hepatitis
HBsAg titre can be used in monitoring treatment
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity
post vaccination – only HBs AB is positive
past HBV infection, Anti-HBs is present in concurrence with anti-HBc IgG
HBeAg and anti-HBe is measure or viral replication
HBV DNA is a better measure of replication at present
HBcAg is an intracellular presence in infected
hepatocyte, thus it is not identified in the serum
IT IS NOT A MARKER
anti HBc AB
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and
CHB.
ONLY Anti HBc AB positive and rest is negative is possible in special situations
MOLECULAR DIAGNOSIS
HBV DNA is a direct measurement of the viral load, which
reveals the replication activity of the virus
Real-time PCR can detect wide dynamic range of viral load and it is STANDARD OF CARE
GENOTYPING
HBV genotyping is significant to predict HBV disease progression and
determine appropriate antiviral therapy.
ACTIVE HEPATITIS
HBsAg positive and symptoms are present with abnormal labs
CHRONIC HEPATITIS
HBsAg positive for more than 6 months
PAST INFECTION
coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low
level of intrahepatic HBV DNA without detectable HBsAg
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG. The identification of serological markers allows: to identify patients with HBV infection; to elucidate the natural course of chronic hepatitis B (CHB); to assess the clinical phases of infection; and to monitor antiviral therapy .
HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
Differences in the serum HBsAg levels during the different phases of infection indicate the distribution of cccDNA during the respective phases of the disease.
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity .In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
three conditions. First, it can be predominantly seen as IgM class during the window period of acute phase. Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels. If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure. HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 10 8 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
Furthermore, HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase of infections, but has little clinical importance. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
Occult HBV infection has some clinical importance.
First, it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation ,or hemodialysis .
Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .
Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection .
Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis .
Tests for occult HBV infection are considered in the following conditions: in patients with cryptogenic liver disease, especially when having anti-HBc in serum; in patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission.
Conclusions
In this article, we aimed to give informations about HBV serological and molecular diagnosis. First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus. In order to verify first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used. Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
This is a very useful article. The main concern of this article is knowing the differences between acute, chronic and occult HBV.
The above presentation can be identified from the HBV serological type, HBVs Ag is denoting for infectivity, and it appears. In the blood 1 to 10 weeks after acute infections. If it remains in circulation for over six months, it means chronic hepatitis. In some situations, we may not detect HBVs Ag in the blood in the early post-infective period and in this time, we may find Anti HBVc Ab only.
In diagnosing HBV infection we rely mainly on HBV DNA assay but sometimes in the case of occult HBV we may. need to have a hepatic sample to detect the viruses.
In general, now is no significant implication for HBVcAg to detect the virus.
The presence of Anti HBc and anti-HBS AB ing indicate occult infections without the presence of HBsAg
HBV genotyping is significant in predicting HBV disease progression. · Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C. · Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
1.HBs Ag
After exposure to HBV, appears 1 to 10 weeks.Persistent > 6 months indicate chronic infectionHigher in HBe Ag positive vs negativeMonitoring quantitative HBs Ag levels predict treatment response and disease progression2.anti-HBs
Neutralizing antibodyConfer long-term immunityanti-HBs positive (others were negative) –> vaccinationanti-HBs positive + HBc IgG positive (others were negative) –> past infection3.HBe Ag
Active viral replicationanti-HBe4.HBc IgG
Isolated positive in:After the acute infection has endedPast infectionOccult infection (positive HBV DNA)5.HBc IgM
Raised or positive during acute infectionDisappearance after 6 months6.HBV DNA
Replication activityHigher titre related to more rapid replicationHigher incidence of HCC7.HBV genotyping
Diagnosis of Hepatitis B infection
Acute hepatitis
HBsAg positiveanti-HepBc IgMHBV DNA detectedElevated ALT, ASTChronic hepatitis
HBsAg positive > 6 monthsPast HBV infection
anti-HBsanti-HBc IgG positiveOccult HBV infection
HBV DNA detectedHBs Ag not detectedHepBc IgG positive
Summary of the article Diagnosis of hepatitis B HBV is dsDNA virus ( Hepadnaviridae family).
Transmission of HBV via parenteral route, sexual activity, and sometimes vertical routes. HBV serological markers:
1. Hepatitis B surface antigen (HBsAg): encodes for surface proteins.
– appears within 1 to 10 weeks after infection.
· Persistence for more than 6 months indicates chronic HBV infection.
· The serum HBsAg titers are higher in patients with HBeAg- positive CHB than in HBeAg-negative CHB. But Quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg- negative CHB patients with normal serum alanine aminotransferase levels.
2. Anti-HBs: · confers long-term immunity.
· Positive HBs-Ab denotes acquired immunity through vaccination.
· Positive HBs-Ab in concurrence with anti-HBc IgG indicates past HBV infection.
· Positive HBs-Ab and positive HBsAg: denotes that anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
3. Anti-HBc IgM and IgG:
· HBcAg indicates infected hepatocyte, thus virus is not identified in the serum.
· anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM weans off.
· Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Isolated anti-HBc positive: when HBsAg and anti-HBs are not detected. This can be seen in 3 conditions:
a) During the window period its seen predominantly as IgM class.
b) After acute infection HBsAg has decreased below the cutoff level of detection.
c) After several years of chronic HBV infection: HBsAg has diminished to undetectable levels.
· If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
4. HBeAg and Anti-HBe:
· HBeAg and anti-HBe had been used in past to know infectivity (presently replaced by HBV DNA assay).
· HBeAg to anti-HBe seroconversion is associated with remission of hepatic disease.
· Active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
Molecular methods for HBV infection
1.HBV DNA is a direct measurement of the viral load which reveals the replication activity of the virus. It is a reliable marker of replication activity. Detects early stage of infection (1 month after HBV infection). Higher titers of HBV DNA are related to the more disease progression and higher incidence of HCC. It is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
2. HBV DNA techniques: there are two principles of techniques to identify and quantify HBV DNA:
a) Signal amplification b) Target amplification.
Real time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107–108 IU/mL). Real-time PCR is the standard method to detect and quantify HBV DNA in clinical setting. 3.HBV genotyping
1.HBV have ten genotypes, labelled A–J: have distinct geographic distribution.
· Genotype B and C: are restricted in Oceania and Asia.
· Genotype A and D: are omnipresent but most common in Africa and Europe.
· Genotype I is unusual and can be observed in Vietnam, Laos, India and China.
· Genotype J has been reported in Japan and Ryukyu.
· Other genotypes such as E, F, G, and H are also occasionally found in Asia.
2. HBV genotyping is significant in predicting HBV disease progression.
· Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
· Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
· Genotype B or C had a lower opportunity to gain serological response to tenofovir.
3. HBV genotyping can be confirmed using:
reverse hybridization,
genotype-specific PCR assays,
real-time PCR,
restriction fragment-length polymorphism,
sequence analysis, microarray (DNA Chip) and fluorescence polarization assay. Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, and high serum aminotransferases. Usually, anti-HBc IgM is detected with HBV DNA presence. HBeAg can also be identified in most acute phase of infections but has little clinical importance. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months. Past HBV infection is defined by the coexistence of anti-HBs and or IgG anti-HBc. Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. Serologically there is presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection. Occult HBV infection has clinical importance because it can lead to Transmation in solid organ transplantation.
Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state, it may accelerate liver injury leading to hepatic fibrosis and is risk factor for HCC. Tests for occult HBV infection are considered in the following conditions:
· in patients with cryptogenic liver disease (when having anti-HBc in serum).
· in patients considering immunosuppression therapy or chemotherapy.
· in solid organ transplantation donors, due to the possibilities for transmission.
indications for screening for occult HBV infection: –
patients with cryptogenic liver disease, especially with a positive anti-HBc in serum;
patients considering immunosuppression therapy or chemotherapy;
in SOT donors due to the risk of transmission
Conclusion
– the 1st step of HBV diagnosis is use of serological markers to detect antigens and antibodies against HBV
– qualitative and quantitative molecular tests are used to verify the 1st step of diagnosis by quantifying the viral load and identifying the genotypes
– it is important to establish a diagnosis of HBV infection, this helps determine acute, chronic and occult cases of infection so as to institute preventive remedies and initiate antiviral therapy
-HBV belongs to hepadnaviridae family, approx. 240000000 ppl are chronically infected and are at risk of cirrhosis and HCC
SEROLOGICAL MARKERS FOR HBV INFECTION.
HBsAg-Hallmark of infection, acute/chronic. Higher titers in active vs chronic.
Anti HBs – Neutralizing antibodies and gives immunity. can be seen post vaccination. In carriers of Hep B it occurs at the same time with HBsAg.
HBeAg and Anti HBe -Markers of infectivity. Replaced by HBV DNA.
HBcAg -Marker of infectivity. In those with HBsAg -VE,they are considered isolated anti HBc positive. Possible explanations for isolated anti HBc +VE;
IgM class in window period of acute phase.
At the end of acute infection and anti HBs has declined to undetectable levels.
Several yrs of chronic HBV infection with HBsAg being undetectable.
MOLECULAR METHODS FOR HBV INFECTION.
HBV DNA – Directly estimates VL/replication of virus, seen 1/12 post infection, peaks(10*8 copies/ml) ~ 3/12 and declines in chronic states or on recovery. It is identified by signal amplification(hybrid/branched DNA) or Target amplification(Real time PCR)
HBV GENOTYPING – HBV divided into 10 genotypes ;A-J,A & D have more chronicity than B & C. Confirmed via specific PCR assay, Real time PCR, Micro assay and Fluorescence polarization assay.
DIAGNOSIS OF HEP B INFECTION.
>Acute Hep B ; HBsAg, symptoms, High serum aminotransferin, HBV DNA +/- anti HBc IgM and HBeAg.
>Chronic Hep ; Persistent HBsAg >6/12,No symptoms.
>Past HBV infection -Anti HBs and IgG anti HBc.
>Occult HBV infection – low HBV DNA without HBsAG and anti HBs antibody ,+VE anti HBc
>Vaccinated -anti HBs +VE
Importance of Occult HBV infection;
Can be transmitted via transfusion, SOT.
Can reactivate in immunosuppressed states.
Can accelerate liver injury and lead to fibrosis in CLD pts.
– HBsAg titers are higher in HBeAg positive CHB patients than in HBeAg negative CHB patients
– monitoring quantitative HBsAg levels predicts response to treatment and disease progression in HBeAg negative CHB patients with normal ALT levels
– anti-HBs (HBsAb) is a neutralizing antibody which confers long-term immunity
– a positive HBsAb implies acquired immunity through vaccination
– previously HBeAg and HBeAb (anti-HBe) were used as markers for infectivity and viral replication but this has now been replaced by HBV DNA
– seroconversion from HBeAg to HBeAb indicates remission of hepatic disease
– HBcAg is not identified in serum
– during an acute infection, anti-HBc IgM and IgG appear 1-2 weeks after presence of HBsAg alongside elevated ALT and AST and symptoms
– anti-HBc IgM wears off after 6 months of acute infection
– anti-HBc IgG remains detectable in both patients with resolved HBV infection and CHB infection
– isolated anti-HBc positive: refers to presence of positive anti-HBc IgG, negative HBsAg, negative anti-HBs
– in cases of isolated positive anti-HBc, check anti-HBc IgM so as to assess for the possibility of a recent HBV exposure
– occult HBV infection is characterised by existence of detectable HBV DNA without HBsAg
Molecular methods for HBV infection
– HBV DNA: is a direct measurement of the viral load, it reveals the virus replication activity
– it is detectable in the early stage of infection i.e., one month following HBV infection
– it increases to peak levels (>10⁸ copies/ml) within 3 months post HBV exposure then gradually decreases in chronic infection or disappears at recovery from HBV infection
– detection of HBV DNA: is a reliable marker of replication activity, higher HBV DNA titers are associated with a more rapid disease progression and higher HCC incidence
– HBV DNA testing helps determine which patients require antiviral therapy and monitor them for suitable treatment
HBV genotyping
– HBV reproduces via reverse transcriptase which has insufficient proofreading capability hence has a high genetic heterogeneity
– 10 HBV genotypes, labelled A-J have been described, they have distinct geographic distribution
– acute infection with genotypes A and D leads to a higher chronicity rate than genotypes B and C
Diagnosis of hepatitis B infection
– acute HBV infection diagnosis is identified by detection of HBsAg, symptoms and high AST and ALT
– usually, anti-HBc IgM can be detected and HBV DNA is present
– HBeAg can be detected in most acute phase infections
– persistence of HBsAg for >6months suggests a diagnosis of chronic HBV infection
– patients with chronic HBV infection are mostly diagnosed via lab tests as opposed to clinical presentation
– coexistence of anti-HBs and IgG anti-HBc is suggestive of past HBV infection
– occult HBV infection is defined by: –
persistence of low intrahepatic HBV DNA levels without detectable HBsAg,
presence of isolated anti-HBc in the absence of HBsAg and anti-HBs
– the gold standard of diagnosis for occult HBV infection is detection of HBV DNA in the liver, but this is an invasive procedure
– HBV DNA PCR has adequate sensitivity to identify occult HBV infection
– clinical relevance of occult HBV infection: –
it can be transmitted via transfusion, hemodialysis, solid organ transplantation
HBV infection reactivation may occur in immunocompromised patients or in patients on chemotherapy
it may accelerate liver injury leading to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis
– indications for screening for occult HBV infection: –
patients with cryptogenic liver disease, especially with a positive anti-HBc in serum;
patients considering immunosuppression therapy or chemotherapy;
in SOT donors due to the risk of transmission
Conclusion
– the 1st step of HBV diagnosis is use of serological markers to detect antigens and antibodies against HBV
– qualitative and quantitative molecular tests are used to verify the 1st step of diagnosis by quantifying the viral load and identifying the genotypes
– it is important to establish a diagnosis of HBV infection, this helps determine acute, chronic and occult cases of infection so as to institute preventive remedies and initiate antiviral therapy
DIAGNOSIS OF HBV
HBV id DNA virus and commonest cause of hepititis
High endemic 8% prevalence CHINA AND S-E ASIA, SUBSAHARIAN AFRICA
intermediate 2-7%
low- Less than 2%
Why molecular marker are important ?
diagnosis\
to know natural history
to monitor anti-viral treatment
to know the clinical phase of disease
What is actually tested ?
HBsAg
anti HBs
anti-HBc
HBe AG
anti HBE – Ig M IN EARLY PERIOD, Ig G in late phase
DNA copies of HBV– more important in seronegative patients
real time PCR is best
Genome – A B C D – specific to geographical area , like C is common in INDIA
can predict disease progression, genome C is linked to HCC and cirrhosis
can determine appropriate anti viral therapy A and B RESPOND BETTER TO INTERFERON
How to interpret?
ACUTE INFECTION
HBsAg present
raised enzymes
symptoms
HBV DNA present
anti HBC may be present
CHRONIC INFECTION
more than 6 months
only lab test positive
no symptoms
anti HBs and Ig G anti HBc present
OCCULT INFECTION
anti HBc present
HBs Ag and anti HBs both are absent
intrahepatic HBV DNA present
VACCINATED
anti HBs present
more than 10 IU/ML is good titre
what occult infection can cause ?
it can be transmitted through organ donation
increases risk of fibrosis with HCV
reactivation is possible with IS
risk factor for HCC
Summarise this article Introduction
-Hepatitis B virus (HBV) belongs to the Hepadnaviridae
family.HBV genome comprise a circular, partly double-stranded DNA.
-HBV infection is transmitted through parenteral, sexual and vertical route and they have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC) .
-Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection. Serological markers for HBV infection
-They consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
-HBsAg is the serological hallmark of HBV infection.Persistence of this marker for more than 6 months implies chronic HBV infection .
Monitoring of quantitative HBsAg levels predicts treatment
response to interferon and disease progression in HBe Ag negative
CHB patients with normal serum alanine aminotransferase levels .
-Anti-HBs is known as a neutralizing antibody, and confers long-term immunity . In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
– In the past HBV infection, it is present in concurrence with anti-HBc IgG.
-In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
– HBe Ag to anti-HBe seroconversion is related to the remission of hepatic disease , however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg .
-HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
-During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks
after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
– Some HBsAg-negative individuals are positive for
anti-HBc IgG without anti-HBs in three conditions: First, it can be predominantly seen as IgM class during the window period of acute phase. Second, after acute infection had ended, anti-HBs has decreased
below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
-HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg . Molecular methods for HBV infection
– The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC .
– It is detectable at 1 month after HBV infection and increases up to peak level approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
-HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor patients. HBV genotyping
-HBV can be divided into ten genotypes, labelled A–J.
-Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C and genotype C generally is
considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC .
-In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D. Diagnosis of hepatitis B infection
-Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases.
-Anti-HBc IgM can be detected and HBV DNA is present.
-The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
-Patients with chronic HBV infection are commonly diagnosed by
laboratory means but not by clinical presentations.
-Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
-Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody .
– The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection .
– Occult HBV infection has some clinical importance : First, it can be transmitted via transfusion, solid organ transplantation or hemodialysis . Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .Third, it accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic HCV infection . Forth, it a risk factor for HCC by its carcinogenic effect .
Diagnosis of HBV
Review article Introduction
Hepatitis B viral infection consider one of the public health problems worldwide as it can lead to progressive liver disease including liver cirrhosis and hepatocellular carcinoma if not treated
HBV is a circular DNA virus with a wide range of serological assays that can determine the acute and chronic clinical course of the disease, usually transmitted through contaminated blood and blood products, sexual contact, and vertical transmission
HBsAg positive referred to acute infection usually appear between 1-10 weeks, if persists more than 6 months indicate chronic HBV infection while AntiHBs AB is called neutralizing AB and indicates a long immune response, and alone indicate acquired immunity from vaccination.HBsAg negative with antiHBc indicates previous exposure
historical, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use has generally been replaced by HBV DNA
isolated anti-HBc positive. It can be seen in 3 conditions
1. acute HBV infection window with antiHBc IgM
2. Past HBV infection with antiHBc IgG
3. occult HBV infection HBV DNA assays should be tested in chronic
liver disease patients to find out occult HBV infection categorized by the presence of detectable HBV DNA without
serum HBsAg. Different molecular and genotypes assay is available for the diagnosis of HBV infectious pattern
HBV DNA is one of the direct molecular testings for the viral load by real-time -PCR and reflects the viral replication activity and can be positive as early as the first month of acute HBV infection with the peak level in 3 months after viral infection followed by subsequent reduction, its important for the diagnosis as well as monitoring the response e for antiviral therapy
HBV Genotyping
HBV can be alienated into ten genotypes, labeled A–J: they have distinct geographic distributions (24). Genotypes B and C are
restricted to Oceania and Asia, whereas genotype A and D are omnipresent but most common in Africa and Europe. HBV genotyping
is significant to predict HBV disease progression and determine appropriate antiviral therapy
Acute infection with genotypes A and D leads to a higher rate of chronicity than with genotypes B and C
Genotype c and B have more vertical transmission and are associated with fast progression to liver cirrhosis and HCC with less response to interferon-gamma compared to genotypes A&D
Occult HBV infection is defined by the persistence of a low level of intrahepatic HBV DNA without detectable HBsAg only isolated anti-HBc with the absence of HBsAg and anti-HBs antibody
And the detection of HBV DNA is the gold standard for the diagnosis of OBI and is widely used in clinical practice
Conclusion
This narrative review gives us a summary of HBV epidemiology and its serological testing along with the different molecular and genotyping assays that help in the early diagnosis and determination of the different clinical aspects of HBV infection
This is the review article on diagnosis of hepatitis b virus. Hepatitis B virus (HBV) belongs to the Hepadnaviridae family with a diameter of 30–42 nm and consists of outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein.
HBV is transmitted through parenteral, sexual and vertical route. China, South East Asia, Indonesia, and sub-Saharan Africa are regarded as highly endemic areas.
Serological markers for HBV infection
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG
After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
In the past HBV infection, it is present in concurrence with anti-HBc IgG.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms
After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection
HBV genotyping
HBV genotyping can be confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNA Chip) and fluorescence polarization assay.
According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution
Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
Diagnosis of hepatitis B infection Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually, anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase of infections
The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg . It is a serological situation isolated anti-HBc +ve with the absence of HBsAg and antiHBs antibody
Clinical significance of Occult HBV infection
Can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
It appears to be a risk factor for HCC by its carcinogenic effect and leading to continuous hepatic inflammation and fibrosis
Tests for occult HBV infection are considered in
Cryptogenic liver disease, especially when having anti-HBc in serum;
Immunosuppression therapy or chemotherapy patients; and
Solid organ transplantation donors
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
Serological markers for HBV infection
HBsAg is the serological hallmark of HBV infection. appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBe Ag negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity.
In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive. It can be seen in three conditions. First, it can be predominantly seen as IgM class during the window period of acute phase. Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. There are two principles of techniques to identify and quantify HBV DNA: signal amplification such as hybrid capture and branched DNA technology; target amplification such as polymerase chain reaction (PCR)
HBV genotyping
HBV has a high genetic heterogeneity because it reproduces via a reverse transcriptase that has insufficient proofreading capability. According to the sequence divergence, HBV
can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases chronic infection is based on the persistence of HBsAg for more than 6 months.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAgThe detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in
the serum.
Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
IntroductionHepatitis B (HBV) belongs to the hepadnaviridae family. It consists of an outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein. The viral capsid contains the viral genome and DNA polymerase. The HBV genome consists of a circular partly double stranded DNA. The covalently closed circular DNA (cccDNA) is the transcriptional template of HBV and stays inside the hepatocyte nucleus. HBV infection is responsible for many chronic liver diseases, and can lead to the development of liver cirrhosis and hepatocellular carcinoma (HCC). It can be transmitted via the parenteral, sexual and vertical routes. The endemicity can be classified into three categories: high (infection is reported as more than 8% of the population), intermediate (Infection rate between 2-7%) and low (Infection rate less than 2%). China, South East Asia, Indonesia and sub-Saharan Africa are regarded as high endemic areas. Anti-HBV vaccinations and diagnosis of the infection are significant methods that aid in reducing the burden of this disease. This article reviewed the serological and molecular diagnosis of HBV.
Serological markers for HBV infection The serological markers of HBV infection include HBsAg, anti-HBs, HBeAg, and anti-HBc IgM and IgG.
HBsAg is the serological hallmark of HBV infection. It appears in the serum within 1 to 10 weeks after exposure to HBV.
If it persists more than 6 months, it implies a chronic HBV infection. The differences in the serum HBsAg levels during the different phases of the infection indicate the distribution of cccDNA during the respective phases of the disease.
HBV DNA assay should be tested in chronic liver disease patients to find out occult HBV infection characterized by the existence of detectable HBV DNA without serum HBsAg. Anti-HBs is a neutralizing antibody and confers long-term immunity. If patients have acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
Molecular methods for Detecting HBV infection HBV DNA is a direct measurement of the viral load. It is detectable at the early stages of infection and increases to peak level approximately 3 months after the exposure to HBV. It then gradually decreases in chronic infection or disappears at the recovery from HBV infection. The viral load reveals the replication activity for the virus, and high titers of HBV DNA are related to the more rapid disease progression and a higher incidence of HCC.
HBV genotyping HBV has a high genetic heterogeneity as it reproduces via reverse transcriptase that does not have sufficient proofreading capability. HBV can be divided into 10 genotypes, labelled A to J, and they have distinct geographic distribution. Acute infections with genotype A and D lead to higher rates of chronicity than genotype B and C. genotype C is considered a risk factor for perinatal infection and is related to sever liver disease, including liver cirrhosis and HCC.
Patients with genotype A and B have been shown to respond better to interferon therapy, compared to genotypes C and D. HBV genotyping can be confirmed using reverse hybridization, genotype- specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fluorescence polarization assay.
Diagnosis of hepatitis B infection Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferase levels. The diagnosis of chronic HBV infection is based on the persistence of HBsAg for more than 6 months. Occult HBV infection is defined by the persistence of low level of intrahepatic HBV DNA without detectable HBsAg. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, unfortunately the procedure for obtaining samples is invasive. Occult HBV infection can be transmitted via transfusion, transplantation or hemodialysis.
The reactivation of the infection may occur in patients receiving chemotherapy, or patients in an immunocompromised state. Occult HBV infection may also lead to hepatic fibrosis in patients with chronic liver disease, including chronic hepatitis C infection. It also appears to be a risk factor for HCC.
Tests for occult HBV infection should be considered in patients with cryptogenic liver disease, in patients considering immunosuppression therapy or chemotherapy and in solid organ transplantation donors.
Conclusion The aim of the article was to provide information regarding the serological and molecular testing of HBV infection. Diagnosis is an important tool in order to establish preventative remedies and to initiate treatment.
Introduction
· HBV has an outer lipid envelope containing HBsAg and an icosahedral capsid with viral genome and DNA polymerase.
· HBV genome has four open reading frames, encoding surface proteins, polymerases, reverse transcriptases, and transcriptional transactivator factors.
· Covalently closed circular DNA (cccDNA) is the transcriptional template of HBV, making it error-prone and high in mutation rate.
· HBV infection is responsible for most chronic liver diseases worldwide and is transmitted through parenteral, sexual, and vertical routes.
· Diagnosis of acute, chronic, and occult HBV is essential for reducing its burden.
Serological markers for HBV infection
· Serological markers for HBV infection include HBsAg, anti-HBs, HBeAg, and anti-HBc IgM and IgG, which are used to identify patients with HBV infection, elucidate the natural course of chronic hepatitis B (CHB), assess clinical phases, and monitor antiviral therapy.
· Anti-HBs is a neutralizing antibody that confers long-term immunity in patients with acquired immunity.
· HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use has mostly been replaced by HBV DNA assay.
· HBeAg is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
· During acute infection, anti-HBc IgM and IgG emerge 1-2 weeks after the presence of HBsAg and wear off after 6 months.
· After chronic HBV infection, HBsAg has diminished to undetectable levels.
· HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by the existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
· HBV DNA is a reliable marker of replication activity and is useful in routine clinical settings to determine patients who need antiviral therapy and monitor them for suitable treatment.
· Real-time PCR is the standard method to detect and quantify HBV DNA in clinical settings, as it is fully automated and does not generate carry-over contamination.
HBV genotyping
· HBV has a high genetic heterogeneity, divided into ten genotypes with distinct geographic distributions.
· Acute infection with genotypes A and D leads to a higher rate of chronicity than genotypes B and C, while genotype C is a risk factor for perinatal infection and severe liver disease.
· Recent studies have shown that patients infected with genotypes B or C have a lower opportunity to gain serological response to tenofovir.
Diagnosis of hepatitis B infection
· Acute hepatitis B is identified by HBsAg, symptoms, and high serum aminotransferases, while chronic infection is based on the persistence of HBsAg for more than 6 months.
· HBV DNA testing is widely used to diagnose occult HBV infection, as (cccDNA) remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
· Occult HBV infection can be transmitted via transfusion, solid organ transplantation, or hemodialysis, and can accelerate liver injury and lead to hepatic fibrosis.
· Tests for occult HBV infection are recommended in cryptogenic liver disease, immunosuppression therapy, chemotherapy, and solid organ donation donors.
Conclusion
· First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus
· Diagnosis of HBV is important for establishing preventive remedies and initiating antiviral treatment.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your well-structured reply that is a very clinically oriented summary.
This article talks about the HBV virus, its prevalence, and how to diagnose it.
Intruduction :
HBV is a double-strand DNA virus responsible for most chronic liver diseases worldwide.
Endemic areas are :
China, South East Asia, Indonesia, and sub-Saharan Africa
Diagnosis :
Serological markers for HBV infection:
1-HBS ag:appears in the serum
within 1 to 10 weeks.
Persistence of this marker for more
than 6 months implies chronic HBV infection
.
2-Anti-HBs is known as a neutralizing antibody, and
confers long-term immunity.
3-HBeAg and anti-HBe had been used to
know infectivity and viral replication, but their use for
this purpose has mostly been replaced by HBV DNA
assay.
4-HBcAg is an intracellular presence in infected
hepatocyte, thus it is not identified in the serum. During
acute infection, anti-HBc IgM and IgG emerges 1–2 weeks
after the presence of HBsAg along with raised serum
aminotransferase and symptoms.
5-HBV DNA : is a direct measurement of the viral load.
reveals the replication activity of the virus.
Higher titers of HBV DNA are related to the more rapid
disease progression and higher incidence of HCC.
There are tow type to detect :
signal amplification such as hybrid
capture and branched DNA technology; target amplifcation
such as polymerase chain reaction (PCR).
HBV genotyping:
the HBV genotyping
is significant to predict HBV disease progression and
determine appropriate antiviral therapy.
There are multigenotyping form A to F
Occult HBV infection is defined by persistence of low
level of intrahepatic HBV DNA without detectable HBsAg.
☆Diagnosis of hepatitis B
In this article, the serological and molecular diagnosis of HBV have been reviewed. ◇ Introduction
▪︎Hepatitis B virus (HBV) is a DNA virus. It can cause an infection with a major global health problems leading to severe liver disease. ◇ Serological markers for HBV infection
▪︎HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
▪︎The recognition of serological markers helps to:
1. Identify patients with HBV infection.
2. Elucidate the natural course of chronic hepatitis B (CHB).
3. Assess the clinical phases of infection.
4. Monitor antiviral therapy.
HBsAg:
▪︎Appears in serum within 1 to 10 weeks of infection.
▪︎If persist > 6 months → chronic infection.
▪︎The serum titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
▪︎Monitoring of it’s levels predicts treatment response to INF and disease progression.
Anti-HBs:
▪︎A neutralizing antibody that confers long-term immunity.
▪︎The only serological marker detected in serum after vaccination.
▪︎In past HBV infection, it is present in concurrence with anti-HBc IgG.
▪︎In most cases patients with anti-HBs antibodies are regarded as carriers of HBV.
HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease
▪︎Active viral replication is sustained in some patients with HBe seroconversion.
HBcAg: is not identified in the serum.
Anti-HBc IgM and IgG:
▪︎Emerges 1–2 weeks after the presence of HBsAg.
▪︎After 6 months of acute infection, anti-HBc IgM wears off.
▪︎Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
▪︎Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs.
▪︎If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
HBV DNA assays
▪︎Should be tested in chronic liver disease patients to find out occult HBV infection, which is characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
▪︎HBV DNA is a direct measurement of the viral load.
▪︎Detection of HBV DNA is a reliable marker of replication activity, and higher titers are related to the more rapid disease progression and higher incidence of HCC.
▪︎It is useful in routine clinical setting.
Principles of techniques to identify and quantify HBV DNA:
1. Signal amplification.
2. Target amplification (Real-time PCR the standard method to detect
and quantify HBV DNA in clinical setting). HBV genotyping
▪︎HBV has a high genetic heterogeneity (ten genotype labelled A–J with distinct geographic distribution).
▪︎Can predict HBV disease progression and determine therapy.
▪︎Can be confrmed using diverse methods. Diagnosis of hepatitis B infection Acute infection:
▪︎Is a clinical diagnosis based on:
1. Detection of HBsAg
2. Symptoms
3. High serum aminotransferases.
▪︎Usually anti-HBc IgM and HBV DNA and HBeAg can be detected.
Chronic infection:
▪︎Is based on the persistence of HBsAg for more than 6 months.
▪︎ A laboratory based diagnosis.
▪︎Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection:
▪︎Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
▪︎A serological situation, defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
▪︎The detection of HBV DNA in the liver is the gold standard of diagnosis.
▪︎Tests for occult HBV infection are considered in:
1. Patients with cryptogenic liver disease
2. Patients considering immunosuppression therapy or
chemotherapy.
3. Solid organ transplantation donors.
Conclusions
▪︎The first step for diagnosis of HBV infection is the serological markers for detecting Ags and Abs against HBV.
▪︎Diagnosis of HBV infection is an important tool to identify acute, chronic and occult cases of infection.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections
I. Diagnosis of hepatitis B Summarise this article HBV serological markers:
HBsAg & anti-HBs
Anti-HBc IgM & IgG
HBeAg & anti-HBe. Serological diagnosis of HBV HBsAg:
It is the serological indicator of HBV infection.
It appears in serum 1 to 10 weeks after an acute exposure to HBV.
Its persistence for > 6 months indicates chronic HBV infection.
Serum levels are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB. Anti-HBs:
Is a neutralizing antibody.
Confers long-lasting immunity.
The sole serological marker found in patients who have acquired immunity by vaccination.
Present together with anti-HBc IgG in the past HBV infection.
It fails to neutralize the circulating viruses in most cases_ HBV carrier status. HBeAg & anti-HBe:
Their use to indicate activity has been replaced by HBV DNA assay.
HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
Pre-core & core region mutations can cause sustained viral replication in some patients with HBe seroconversion. HBcAg:
Not identified in the serum (intracellular in infected hepatocyte).
Anti-HBc IgM & IgG appears 1–2 weeks, together with raised aminotransferase & symptoms, after the presence of HBsAg during acute infection.
Anti-HBc IgM disappears 6 months after acute infection. Anti-HBc IgG persists in resolved HBV infection & CHB.
If isolated anti-HBc positive is detected, anti-HBc IgM should be done to assess a recent HBV exposure.
HBV DNA assays should be tested in CLD patients to detect occult HBV infection (detectable HBV DNA without serum HBsAg). Molecular diagnosis of HBV HBV DNA:
Directly measures the viral load.
A marker of replicative activity.
Detected early (1 month) after HBV infection & increases to peak level 3 months after the exposure & then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
Higher titers indicate:
More rapid disease progression
Higher incidence of HCC
The need for antiviral therapy
Techniques to identify & quantify HBV DNA:
Signal amplification (hybrid capture & branched DNA technology)
Target amplification (PCR):
The standard method
Detects wide range of viral load (10–15 IU/mL to 107–108 IU/mL). HBV genotyping
HBV can be divided into ten genotypes (A–J).
Genotype B & C are restricted to Oceania & Asia.
A & D are most common in Africa and Europe.
A & D linked to higher rate of chronicity than B & C.
C is a risk factor for perinatal infection & linked to severe liver disease (cirrhosis & HCC).
A & B responds better to interferon than C & D.
B & C had lower serological response to tenofovir. Methods to confirm HBV genotyping:
Reverse hybridization
Genotype-specific PCR assays
Real time PCR
Restriction fragment-length polymorphism
Sequence analysis
Microarray (DNAChip)
Fluorescence polarization assay. Diagnosis of hepatitis B infection Acute hepatitis B:
Detection of HBsAg,
Symptomatology
High serum aminotransferases.
Anti-HBc IgM & HBV DNA can be present.
HBeAg in acute phase of infections has little clinical.
Significance. Chronic infection:
Persistence of HBsAg for > 6 months.
Coexistence of anti-HBs & IgG anti-HBc defines past HBV infection. Occult HBV infection:
Persistence of low level intra-hepatic HBV DNA without detectable HBsAg.
Isolated anti-HBc in the absence of HBsAg & anti-HBs. HBV DNA in the liver is the gold standard of diagnosis; however, it is invasive & not pravtical.
Real-time PCR for serum HBV DNA can identify occult HBV infection in many cases.
Significance of occult HBV infection:
Transmitted via transfusion, SOT, or HD.
Reactivation after chemotherapy or immunocompromised state.
May accelerate liver injury & lead to hepatic fibrosis in CLD.
A risk factor for HCC by its carcinogenic effect.
Diagnosis of hepatitis B
Hepatitis B virus (HBV) carry risk of chronic hepatitis and hepatocellular carcinoma. It’s transmitted by parenteral rote and sexual activity and vertical rote. This article focus on serological and molecular diagnosis of HBV. Serological markers for HBV infection:
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg is appear after acute exposure to HBV within 1-10 weeks and if persist more than 6 months is indicated chronic exposure to HBV.
Quantity of HBsAg is a marker for treatment response to interferon and disease outcome.
Presence of anti-HBs antibodies indicates patients are carrier for HBV.
HBeAg and anti-HBe indicated infection stage but virama know is detected by HBV DNA and HBeAg to anti-HBe seroconversion is used as a marker for remission of hepatic disease.
HBcAg is an intracellular presence in infected hepatocyte and not found in the serum. During acute infection, anti-HBc IgM and IgG appear within 1–2 weeks after the presence of HBsAg and associated with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM disappeared and Anti-HBc IgG indicates chronicity and continues to detect in both patients with resolved HBV infection and CHB. Molecular methods for HBV infection:
HBV DNA test used for viral load and viral replication. It is detectable at the early stage of infection approximately one month after HBV infection and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
Presence of high titer HBV DNA is associated with viral replication and activity of virus and more aggressive disease and high risk HCC and it’s a time for antiviral therapy for eradication of virus. There are 2 technical procedures to detect HBV DNA:
Hybrid capture depend on PCR
Branched DNA technology
HBV genotyping:
It’s reproduced reverse transcription
It’s 10 subtypes from (A to J).
HBV genotyping is confirmed by diverse methods:
reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fluorescence polarization assay. Diagnosis of hepatitis B infection: Acute hepatitis B:
Detection of HBsAg
Symptoms of low grade fever and jaundice
High serum aminotransferases
Anti- HBc IgM can be detected and HBV DNA is present.
HBeAg positive
chronic infection:
Persistent of HBsAg for more than 6 months.
Past HBV infection is presence of anti-HBs and and IgG anti-HBc.
Occult HBV infection is presence of low level of intrahepatic HBV DNA without detectable HBsAg.
Detection of HBV DNA in the liver is the gold standard diagnosis of occult HBV infection.
Real-time PCR for serum HBV DNA is used for diagnosis of occult HBV infection.
Clinical Importance of Occult HBV infection:
First: it can be transmitted by blood transfusion
Solid organ transplantation including orthotopic liver transplantation
Hemodialysis
Second: reactivation of HBV infection occur in patients receiving chemotherapy or immunocompromised state.
Third: it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
Forth: it’s a risk factor for HCC.
Tests for occult HBV infection are considered in the following conditions:
in patients with cryptogenic liver disease, especially patients with anti-HBc in serum.
in patients on immunosuppression therapy or chemotherapy.
in solid organ transplantation donors (donor derived infection).
Hepatitis B virus HBV;
Is continuing to be a significant infection with escalated risk of morbidity and mortality. Its infection is associated with higher risk of Hepato-cellular carcinoma HCC.
Its way of transmission isessentially sexual, vertical and parentral .Its a acircular virus with double stranded DNA genome and containing Surface antigen, core antigen and e antigen. Its the most common cause of chronic liver disease infecting more than 240 millions over all the world. Incidence of HBV infection ranged from 2-7% of population. HBV vaccine represent a glorious bounce in defeating the spread of virus and its consequent dismal prognosis of HCC.
Meticulous survey for all varieties of HBV infection including: Acute HBV infection which is the presence of HBV surface antigen for 6 month. Chronic HBV is the persistence of of HBV surface antigen for more than 6 months. Occult HBV infection which feature positive HBV DNA in the setting of absent HB surface antigen. Serological markers of HBV infection:
HBV surface antigen HBsAg and HBV surface antibodies HBsAb.
HBeAg and HBeAb
HBcAb:
Its reflecting infection with HBV, its usually detected within 1- 2 weeks post exposure as IgM antibodies. which will vanished and replaced with IgG HBcAb to continue to be detected with chronic HBV infection or recovered infection.
HBsAg reflect infection with HBV and when pesists for more than 6 months it indicate Chronic HBV infection. HBsAb are neutralizing antibodies that bolster a long term immunity.
In the specific circumstances of detecting both HBsAg and HBsAb its indicative of a Carrier state .
I like your summary. I make note of your interpretation of serological testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections.
The following tests can be used for the diagnosis and evaluation of HBV infection:
HBs Ag
The presence of HBs Ag indicates infection. Occur 1 to 10 weeks following an infection. If infection persists for more than six months, it is considered chronic. HBs Ag levels using PCR can be used to predict treatment response.
Anti-Hbs Antibodies
It is known as a neutralizing antibody, and it provides immunity that lasts for a long time.
Anti-HBs is the only serological marker that can be found in the serum of patients who have acquired immunity as a result of vaccination.
If It is present at the same time with anti-HBc IgG, indicates a previous infection with HBV.
HBeAg and anti-HBe Abs
HBeAg and anti-HBe were formerly employed to determine infectivity, however currently viral tests for HBV are more common. The transformation of HBeAg into anti-HBe is diagnostic of remission.
Hbc Ag
Due to the fact that it is already present in infected hepatocytes, HBcAg cannot be found in serum. In acute infections, anti-HBc IgM and IgG antibodies begin to show up one to two weeks after the appearance of HBsAg alongside elevated liver enzymes. Anti-HBc IgM antibodies are no longer detectable in patients with cured HBV infection or CHB six months after the onset of acute infection, although anti-HBc IgG antibodies continue to be found in these patients.
Molecular methods for HBV infection
HBV DNA can be identified one month after HBV infection, reaches its maximal level three months after exposure to HBV, and then gradually lowers in chronic infection or vanishes upon recovery. HBV DNA can be detected one month after HBV infection. In addition to indicating a quick course of the disease and a higher incidence of HCC, the presence of HBV DNA can help determine which patients require antiviral therapy and track how well they are responding to treatment.
Both the presence of HBV DNA and its quantity can be determined by the use of signal amplification techniques, such as hybrid capture and branched DNA technologies, and target amplification techniques, such as PCR. In the meanwhile, the HBV PCR test is the gold standard.
Hepatitis B genotyping
The ten genotypes of HBV, denoted by the letters A through J, each correspond to a different region of the world.
The progression of HBV disease can be predicted by HBV genotyping, which can also point to the most effective antiviral treatment. An acute infection with the genotypes A and D has a higher rate of chronicity than an acute infection with the genotypes B and C, whereas the genotype C is a risk factor for perinatal infection and severe liver disease. A and B have a better response to the therapy with interferon than C and D.
Tenofovir elicits a weaker response from strains B and C. Reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip), and fluorescence polarization assay are some of the methods that can be used to detect HBV genotyping. Other methods include restriction fragment-length polymorphism, real-time PCR, and restriction fragment-length polymorphism.
I like your detailed well-structured summary well supported by references. I note that you have explained how natural history of disease is affected by genotype of HBV.
SEROLOGICAL MARKERS FOR HBV INECTION ;
———————————————————————————–
1-HBsAg;
is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
2-Anti-HBs;
is known as a neutralizing antibody, and confers long-term immunity.
3-HBeAg and anti-HBe ;
had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
4-HBcAg;
is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
SOME HBsAg-negative INDIVIDUALS are positive for anti-HBc IgG without anti-HBs;
——————————————————————————————
In this situation, it should be considered isolated anti-HBc positive. It can be seen inthree conditions;
A- It can be predominantly seen as IgM class during the window period of acute phase.
B- After acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
C- After several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure. HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
MPLECULAR METHOD for HBV INFECTION ;
———————————————————————————————
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
There are two principles of techniques to identify and quantify HBV DNA:
1-Signal amplification such as hybrid capture and branched DNA technology; target amplification such as polymerase chain reaction (PCR) .
2- Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 10 7 –10 8 IU/mL).
HBV GENOTYPING ;
————————————————-
HBV can be divided into ten genotypes, labelled A–J.
Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C .
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D .
Recent studies reported that patients infected with genotype B or C had a lower opportunity to gain serologicaL response to tenofovir.
DIAGNOSIS of HEPATITIS B INFECTION ;
——————————————————————-
1-Acute hepatitis B;
is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
2-The diagnosis of chronic infection ;
is based on the persistence of HBsAg for more than 6 months. Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations. Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
3-Occult HBV infection;
It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody .
The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
However, gaining hepatic HBV DNA is difficult in clinical setting since the procedure is invasive. Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases; thus, HBV DNA testing is widely used to diagnose occult HBV infection .
OCCULT HBV INFECTION has some CLINICAL IMPORTANCE ;
—————————————————————–
1-It can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis .
2-Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .
3- It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection .
4- It appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis .
TESTS for OCCULT HBV INFECTION are CONSIDERED in the FOLLOWING CONDITIONS:
——————————————————————————————–
1-In patients with cryptogenic liver disease, especially when having anti-HBc in serum .
2-In patients considering immunosuppression therapy or chemotherapy.
3- In solid organ transplantation donors, due to the possibilities for transmission .
.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your well-structured reply that is a very clinically oriented summary.
However, there are few spelling errors such as MPLECULAR
Serological markers for HBV infection HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG. 1-HBsAg
· HBsAg is the serological hallmark of HBV infection.
· HBsAg appears in serum within 1 to 10 weeks after acute exposure
· Persistence for more than 6 months indicate chronic HBV infection .
· occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg
2- Anti-HBs
· known as a neutralizing antibody ,long-term immunity
· In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
· In the past HBV infection, it is present in concurrence with anti-HBc IgG. 3-HBeAg and anti-Hbe
· HBeAg and anti-HBe had been used to know infectivity and viral replication,
· but their use for this purpose has mostly been replaced by HBV DNA assay.
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease 4-HbcAg is an intracellular presence in infected hepatocyte, it is not identified in the serum. 5- anti-HBc IgM and IgG
· emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
· some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, (isolated anti-HBc positive)
It can be seen in -IgM class during the window period of acute phase. -after acute infection had ended, anti-HBs has decreased below the cutoff level of detection. -after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
–If isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess thepossibility of recent HBV exposure. Molecular methods for HBV infection
· HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
· It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection
· higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
· Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107–108 IU/mL). HBV genotyping
HBV has a high genetic heterogeneity because it reproduces via a reverse transcriptase that has insufficient proofreading capability.
According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution Diagnosis of hepatitis B infection Acute hepatitis B is a clinical diagnosis identified by
· the detection of HBsAg,
· symptoms,
· high serum aminotransferases.
· Usually anti-HBc IgM can be detected and HBV DNA is present.
· HBeAg can also be identified in most acute phase of infections, but has little clinical importance. The diagnosis of chronic infection is based on
· the persistence of HBsAg for more than 6 months.
· Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations. Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc. Occult HBV infection is defined by
· persistence of low level of intrahepatic HBV DNA without detectable HbsAg
· It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody
· The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection,
· Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases; clinical importanceof Occult HBV infection
1- it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis
2- reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3- it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4- it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis Tests for occult HBV infection are considered in the following conditions
1. in patients with cryptogenic liver disease, especially when having anti-HBc in serum;
2. in patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission Conclusions
· First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus. thenqualitative or quantitative molecular tests are used.
· Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive measures and to initiate antiviral treatment.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your well-structured reply that is a very clinically oriented summary. Typing whole sentence in bold amounts to shouting
Diagnosis and evaluation of HBV infection can be done through the following investigations
HBs Ag
The presence of HBs Ag means infection
If persist > 6 months that means chronic infection
Appear after 1-10 weeks of infection
Monitoring of quantitative HBsAg levels can be used to predicts response to treatment
anti-HBs Ab
It is neutralizing antibody, that provide immunity for the host
Isolated anti-HBs Ab is seen in vaccinated patients
Sometimes HBs Ag is found simultaneously with anti-HBs Ab this is known as carrier state
HBeAg, anti-HBe
Generally, the presence of HBeAg is associated with viral replication and infectivity, and the presence of a however, active
But occasionally viral replication still occur with HBe seroconversion due to mutations that inhibit the production of HBeAg
Its rule is limited and replaced by PCR
HBcAg
It is present in hepatocytes, and cannot be tested in the serum
anti-HBc IgM and IgG.
Both appear after 1-2 weeks of appearance of HBs Ag
After 6 months of infection, anti-HBc IgM disappear and anti-HBc IgG persists in both resolved and chronic HBV infection
So the presence of anti-HBc IgM denotes recent infection
Patient with anti-HBc IgG denotes previous or chronic infection
Isolated anti-HBc positivity (HBsAg-negative, anti-HBs-negative) may be seen in case of early infection (window period, it will be IgM), occult infection (PCR will be positive), or resolved infection
Elevation of liver enzymes and symptoms
Occur usually after 1-2 weeks of appearance of HBs Ag in the serum
More evident in acute rather than chronic infections
PCR
Can be detected as early as 1 month after infection, reach its peak affet 3 months and start to wean off gradually in chronic infection and disappear in resolved infection
Positive PCR can be seen in overt and occult infection
Used to asses viral replication better than HBe Ag
Used to monitor treatment
Diagnosis
Vaccination : HBs Ag negative, anti-HBs Ab positive, antiHbc IgM negative, anti-HBc IgG negative
Acute infection : HBs Ag positive < 6 months, anti-HBs Ab negative, antiHbc IgM positive, anti-HBc IgG may be positive, PCR positive
Previous infection without immunity : HBs Ag negative, anti-HBs Ab negative, antiHbc IgM negative, anti-HBc IgG positive, PCR negative
Previous infection with immunity : HBs Ag negative, anti-HBs Ab positive, antiHbc IgM negative, anti-HBc IgG positive, PCR negative
Chronic infection : HBs Ag positive> 6 months, anti-HBs Ab negative, antiHbc IgM negative, anti-HBc IgG positive, PCR negative
Occult infection : HBs Ag negative, anti-HBs Ab positive, antiHbc IgM negative, anti-HBc IgG positive, PCR positive in blood or in the liver
Occult HBV infection is important to diagnose because of the following
It can be transmitted by blood transfusion ond organ donation
Can be reactivated once the patient is immunosuppressed
Can accelerate liver injury in case of chronic liver disease
It is a risk factor for HCC
Liver tissue (biopsy) PCR is indicated if all the following are present
Hepatic dysfunction of unknown cause
Isolated anti-HBc Ab positivity with negative blood PCR
In high risk patients including those on immunosuppression, dialysis patients
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
I note that you have explained how a donor with evidence of past infection of HBV in relation to the decision -making necessary for transplantation.
(II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg);
(III) P for polymerase including reverse transcriptase;
(IV) X that encodes for a transcriptional transactivator factor (HBxAg
HBV endemicity is divided into three categories;
high, intermediate, low.
Diagnosis of HBV
1- Serological markers for HBV infection
2- Molecular methods for HBV infection
3- HBV genotyping
Serological markers for HBV infection
1- anti-HBs,
2- HBeAg,
3- anti-HBe,
4- anti-HBc IgM and IgG.
HBsAg
It is the serological hallmark of HBV infection.
It appears in serum within 1 to 10 weeks.
its Persistence for more than 6 months implies chronic HBV infection
Monitoring of quantitative HBsAg levels predicts treatment response to interferon in HBeAg negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs
It is known as a neutralizing antibody
confers long-term immunity
1- In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
2- In the past HBV infection, it is present in concurrence with anti-HBc IgG.
HBeAg and anti-HBe
It had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
HBcAg
It is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
anti-HBc IgM and IgG
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg .
After 6 months of acute infection, anti-HBc IgM wears off.
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Anti-Hbc positive can be seen in three conditions.
1- First, it can be predominantly seen as IgM class during the window period of acute phase.
2- Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
3- Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
Molecular methods for HBV infection
HBV DNA reveals the replication activity of the virus.
It is detectable at the early stage of infection (1 month after HBV infection)
it increases up to peak level approximately 3 months after the exposure to HBV
then gradually it diminishes in chronic infection or disappears at the recovery from HBV infection.
There are some techniques to identify and quantify HBV DNA:
1- Semi-automated qPCR
2 Semi-automated real-time PCR
3 Automated real-time PCR
4 Branched DNA.
Real-time PCR has come to be the standard method to detect and quantify HBV DNA in clinical setting.
Furthermore, it can be fully automated and does not generate carry-over contamination
HBV genotyping
can be divided into ten genotypes, labelled A–J.
Genotype B and C are restricted to Oceania and Asia,
whereas genotype A and D are omnipresent but most common in Africa and Europe
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
Methods of HBV genotyping
1- RFLP
2 Reverse hybridization
3- Genotype specific PCR
4 Sequence analysis
Diagnosis of hepatitis B infection Acute hepatitis B
1- HBsAg pos
2- anti-HBc IgM can be pos
3 HBV DNA is pos
4- HBeAg can also be pos , but has little clinical importance.
chronic infection
HBsAg pos (for more than 6 months.)
Past HBV infection
1- anti-HBs pos
2- IgG anti-HBc pos
Occult HBV infection
1- low level of intrahepatic HBV DNA (gold standard but invasive )
2- HBsAg neg
3 anti-HBc IgG pos
4- HBsAg neg
5- antiHBs antibody neg
Occult HBV infection has some clinical importance.
1- First, it can be transmitted via solid organ transplantation, or hemodialysis.
2- Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
3- Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection .
4- Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis
Tests for occult HBV infection are considered in the following conditions:
1- in patients with cryptogenic liver disease, especially when having anti-HBc in serum;
2- in patients considering immunosuppression therapy or chemotherapy;
3- in solid organ transplantation donors, due to the possibilities for transmission
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary. I note that you have explained how a donor with evidence of past infection of HBV in relation to the decision -making necessary for transplantation.
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family. It shows a diameter of 30–42 nm and consists of outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein.
Serological markers for HBV infection consist of:
v HBsAg
v anti-HBs
v HBeAg
v anti-HBe
v anti-HBc IgM and IgG.
The identification of serological markers allows:
To identify patients with HBV infection.
To elucidate the natural course of chronic hepatitis B (CHB).
To assess the clinical phases of infection; and to monitor antiviral therapy.
Molecular methods for HBV infection:
ü Semi-automatedq PCR
ü Semi-automated real-time PCR
ü Automated real-time PCR
ü Branched DNA
HBV genotyping:
v RFLP
v Reverse hybridization
v Genotype specific PCR
v Sequence analysis
Diagnosis of hepatitis B infection:
Acute phase of infections:
Ø HBsAg.
Ø High serum aminotransferases.
Ø Anti-HBc IgM .
Ø HBV DNA
Ø HBeAg Chronic infection:
Ø The persistence of HBsAg for more than 6 months
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
HBV is dsDNA virus belongs to Hepadnaviridae family. Transmission of HBV via sexual activity, parenteral and vertical routes.
HBV serological markers:
1. Hepatitis B surface antigen (HBsAg): encodes for surface proteins and appears within 1 to 10 weeks after infection.
· Persistence of this marker for more than 6 months implies chronic HBV infection.
· The serum HBsAg titers are higher in patients with HBeAg- positive CHB than in HBeAg-negative CHB.
· Quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg- negative CHB patients with normal serum alanine aminotransferase levels.
· HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HbsAg.
2. Anti-HBs:
· A neutralizing antibody and confers long-term immunity.
· Positive HBs-Ab denotes acquired immunity through vaccination.
· Positive HBs-Ab in concurrence with anti-HBc IgG indicates past HBV infection.
· Positive HBs-Ab/positive HBsAg: denotes that anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
3. Anti-HBc IgM and IgG:
· HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
· anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM wears off.
· Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
· Isolated anti-HBc positive: when HBsAg and anti-HBs are not detected. This can be seen in 3 conditions:
a) During the window period of acute phase: seen predominantly as IgM class.
b) After acute infection had ended: anti-HBs has decreased below the cutoff level of detection.
c) After several years of chronic HBV infection: HBsAg has diminished to undetectable levels.
· If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
4. Hepatitis B e antigen (HBeAg) and Anti-HBe:
· HBeAg and anti-HBe had been used to know infectivity and viral replication(has mostly been replaced by HBV DNA assay).
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
· Active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
Molecular methods for HBV infection 1. HBV DNA is a direct measurement of the viral load which reveals the replication activity of the virus. · It is a reliable marker of replication activity. · It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL). · higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC. · It is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment. 2. HBV DNA techniques: there are two principles of techniques to identify and quantify HBV DNA: a) Signal amplification such as hybrid capture and branched DNA technology; b) Target amplification such as polymerase chain reaction (PCR). Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107–108 IU/mL). Real-time PCR is the standard method to detect and quantify HBV DNA in clinical setting. It can be fully automated and does not generate carry-over contamination. HBV genotyping
1. HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution.
· Genotype B and C: are restricted to Oceania and Asia.
· Genotype A and D: are omnipresent but most common in Africa and Europe.
· Genotype I is unusual and can be observed in Vietnam, Laos, India and China.
· Genotype J has been reported in Japan and Ryukyu.
· Other genotypes such as E, F, G, and H are also occasionally found in Asia.
2. HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
· Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
· Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
· Genotype B or C had a lower opportunity to gain serologicalresponse to tenofovir.
3. HBV genotyping can be confirmed using diverse methods:reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fluorescence polarization assay.
Diagnosis of hepatitis B infection
1. Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases.
2. Usually anti-HBc IgM can be detected and HBV DNA is present.
3. HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
4. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
5. Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
6. Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
7. Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
8. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
9. Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases.
10.Occult HBV infection has some clinical importance:
· it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis.
· reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
· it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
· it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis.
11.Tests for occult HBV infection are considered in the following conditions:
· in patients with cryptogenic liver disease (when having anti-HBc in serum).
· in patients considering immunosuppression therapy or chemotherapy.
· in solid organ transplantation donors, due to the possibilities for transmission.
Please summarise this article. Introduction Hepatitis B virus (HBV) infection can lead to severe liver disease such as cirrhosis and hepatocellular carcinoma (HCC). HBV is a circular, partly double-stranded DNA virus Serological markers include : hepatitis B surface antigen (HBsAg) and anti-HBs, anti-HBc IgM and IgG, and hepatitis B e antigen (HBeAg) and anti-HBe. Transmitted is by sexual, parenteral and vertical route. High endemic areas are China, South East Asia, Indonesia, and sub-Saharan Africa South America, South West Asia, Eastern and Southern Europe Intermediate areas. North America and Western Europe are grouped as low endemic regions with prevalence rates range from 0.5% to 2%.
Serological markers for HBV infection
HBsAg– HBsAg is the serological hallmark of HBV infection. It appears in serum within 1 to 10 weeks. Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression
Anti-HBs – It is known as a neutralizing antibody, and confers long-term immunity
HBeAg and anti-HBe– HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease
HBcAg– It is intracellular and not identified in serum.
Anti HBc-The presence of anti-HBc indicates previous or ongoing infection with hepatitis B virus.
Molecular methods for HBV infection HBV DNA is a direct measurement of the viral load It is detectable at the early stage of infection 1-3 months HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC Two techniques to identify and quantify HBV DNA: Signal amplification such as hybrid capture and branched DNA technology Target amplifcation such as polymerase chain reaction (PCR) HBV genotyping Ten genotypes, labelled A–J Genotype B and C are restricted to Oceania and Asia Genotype A and D are omnipresent but most common in Africa and Europe Genotype I is unusual and can be observed in Vietnam, Laos, India and China Genotype J has been reported in Japan and Ryukyu Other genotypes such as E, F, G, and H are also occasionally found in Asia. Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Diagnosis of hepatitis B infection Acute hepatitis B – identified by the detection of HBsAg, symptoms, high serum aminotransferases. HBeAg can also be identified in most acute phase of infections Chronic infection is based on the persistence of HBsAg for more than 6 months Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc. Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg It is defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody Detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV
Importance of Occult HBV Infection · Transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation · Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised. · It can accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection · It is a risk factor for HCC by its carcinogenic effect
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family.
HBV genome comprise a circular, partly double-stranded DNA.
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
HBV can be divided into ten genotypes, labelled A–J, genotype A and D are most common in Africa and Europe.
Acute infection with genotypes A and D leads to higher rate of chronicity.
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
Serological markers: 1- HBsAg:
HBsAg appears in serum within 1 to 10 weeks after infection .
Persistence of this marker for more than 6 months implies chronic HBV infection.
2- Anti-HBs (a neutralizing antibody), and confers long-term immunity .
In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
In the past HBV infection, it is present in concurrence with anti-HBc IgG.
3- HBeAg and anti-HBe:
had been used to know infectivity and viral replication but has been replaced by HBV DNA
4- anti-HBc IgM and IgG:
emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
After 6 months of acute infection, anti-HBc IgM wears off.
Anti-HBc IgG continues to detect in both patients with resolved HBV.
5- HBV DNA: is a direct measurement of the viral load
The detection of HBV DNA is a reliable marker of replication activity
Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
can be used to monitor response to antiviral treatment.
Diagnosis of hepatitis B infection 1- Acute hepatitis B:
By the detection of HBsAg, symptoms, high serum aminotransferases.
Usually anti-HBc IgM can be detected and HBV DNA is present
2- chronic hepatitis:
persistence of HBsAg for more than 6 months.
3- Past infection:
the coexistence of anti-HBs and anti-HBc Ig G.
4- Occult HBV infection:
defined by persistence of low level of intrahepatic HBV DNA and presence of isolated anti-HBc with the absence of HBs Ag and anti-HBs antibody.
The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection. But no is replaced by serum HBV DNA by real time PCR.
Clinical significance of occult hepatitis B:
1- It can be transmitted via transfusion, solid organ transplantation.
2- Reactivation of HBV infection may occur in patients receiving chemotherapy or
immunocompromised state.
3- It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver
disease including chronic hepatitis C infection.
4- A risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic
inflammation and fibrosis
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
Diagnosis of hepatitis B. Introduction.
HBV is one of the most common cause of chronic liver disease worldwide, especially in high endemic area, HBV vaccination is available worldwide and considered one of the important tools for preventing HBV spreading, and herein this article reviewed the serological and molecular diagnosis of HBV. Serological markers for HBV infection.
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG which are used to assess course of chronic hepatitis B (CHB), the clinical phases of infection; and to monitor antiviral therapy.
HBsAg appears in serum within 1 to 10 weeks of exposure and if persist for more than 6 months implies chronic HBV infection, the serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB, and is better marker for monitoring in CHB patients with HBeAg negative and normal serum alanine aminotransferase levels.
In vaccinated person, anti-HBs is the only serological marker detected in serum but if associated with anti-HBc IgG indicates past infection.
HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
HBcAg is an intracellular and not detected in serum and Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Isolated anti-HBc positive: considered when HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs which occurs in the window period of acute phase, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection, HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection with negative HBsAg. Molecular methods for HBV infection.
HBV DNA increase up with acute infection and decrease later on with CHB and disappear after recovery.
HBV DNA are related to the more rapid disease progression and higher incidence of HCC. Furthermore, HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment. HBV genotyping.
HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution, for example genotype B and C are restricted to Oceania and Asia, also each genotype has its character such as acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C. Diagnosis of hepatitis B infection. Acute hepatitis B: diagnosed by detection of HBsAg, symptoms, high serum aminotransferases, anti-HBc IgM can be detected and HBV DNA is present. Chronic infection :the persistence of HBsAg for more than 6 months. Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc. Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg and it is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody, thus HBV DNA testing is widely used to diagnose occult HBV infection. Occult HBV infection diagnosis is challenging and so tests should be considered in following conditions:
1-Patients with cryptogenic liver. disease, especially when having anti-HBc in serum.
2-Patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission. Clinical significance of occult HBV infection in the following conditions:
1-It can be transmitted via transfusion, solid organ transplantation including orthotropic liver transplantation or hemodialysis.
2-Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .
3-Accelerate liver injury and lead to hepatic fibrosis in patients with
chronic liver disease including chronic hepatitis C infection.
4-It appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis. Conclusion:
Diagnosis of HBV is important to reduce its infection specially in immunocompromised patients , diagnosis usually started with serology and then amplification of viral DNA to detect active/chronic and occult infection and used also to follow up treatment.
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections.
Introduction
HBV belongs to the Hepadnaviridae family. It a circular, partly double-stranded DNA virus
HBV infection is responsible for the most of chronic liver diseases worldwide
Mode of transmission: parenteral, sexual and vertical route
HBsAg prevalence:
1. High endemic areas (China, South East Asia, Indonesia, and sub-Saharan Africa). > 8% of the population are affected
2. Intermediate endemic areas (South America, South West Asia, Eastern and Southern Europe). chronic HBV infection rate 2%-7%
3. Low endemic areas (developed countries). Prevalence is 0.5%-2%
Aim of the study: review serological and molecular tests for diagnosis of HBV infection
Serological markers for HBV infection
HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG
HBsAg:
o Appears in serum within 1 to 10 weeks after exposture
o Persistence more than 6 months implies chronic HBV infection
o Titers are higher in patients with HBeA gpositive chronic hepatitis B (CHB) than in HBeAg-negative CHB
Anti-HBs (neutralizing antibody):
o Confers long-term immunity
o Are unable to neutralize the circulating viruses in most cases (so regarded as carriers of HBV)
o It is the only serological marker for vaccination
o Simultaneous appearance of HBsAg and anti-HBs may occur in patients with HBsAg positive
HBeAg and anti-HBe:
o For Infectivity and viral replication (replaced by HBV DNA)
o Seroconversion is related to the remission of hepatic disease
o Active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg
HBcAg:
o Intracellular in infected hepatocyte and is not identified in the serum
anti-HBc:
o During acute infection, anti-HBc IgM and IgG appears 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms
o Anti-HBc IgM wears off after 6 months of acute infection
o Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB
o Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs (isolated anti-HBc positive)
o Isolated anti-HBc positive seen in 3 conditions:
1. seen as IgM class during the window period of acute phase
2. after acute infection ended, anti-HBs has decreased below the cutoff level of detection
3. after several years of chronic HBV infection, HBsAg has diminished to undetectable levels
o In isolated anti-HBc positive, anti-HBc IgM should be checked to assess the possibility of recent HBV exposure
o HBV DNA should be tested in chronic liver disease patients to find out occult HBV infection (existence of detectable HBV DNA without serum HBsAg)
Molecular methods for HBV infection
o HBV DNA (replication activity)
o Detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 10*8 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection
o Also useful in serologically negative HBV infection (HBeAg-negative CHB and occult HBV infection)
o Detection is a reliable marker of replication activity, and higher titers are related to the more rapid disease progression and higher incidence of HCC
o Useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment
Two principles of techniques to identify and quantify HBV DNA:
1. signal amplification such as hybrid capture and branched DNA technology
2. target amplification such as polymerase chain reaction (PCR)
Quantitative methods for HBV DNA:
1. Semi-automated Qpcr
2. Semi-automated real-time PCR
3. Automated real-time PCR
4. Branched DNA
HBV genotyping
According to the sequence divergence, HBV can be divided into ten genotypes (A–J): they have distinct geographic distribution
Genotype B and C are restricted to Oceania and Asia
Genotype A and D are omnipresent but most common in Africa and Europe
Genotype I is unusual: Vietnam, Laos, India and China
Genotype J (Japan and Ryukyu)
Genotypes E, F, G, and H are also occasionally found in Asia
HBV genotyping is useful in predicting HBV disease progression and determine appropriate antiviral therapy
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
Patients infected with genotype B or C had a lower opportunity to gain serological response to tenofovir (recent study)
HBV genotyping can be confirmed using diverse methods:
1. reverse hybridization: high sensitivity, automated systems but relatively high cost
2. genotype-specific PCR assays: high sensitivity, automated systems, easy to perform, and suitable for detecting mixed genotype infections but high cost
3. real-time PCR
4. restriction fragment-length polymorphism (RFLP): easily done, low cost, simple, rapid but low sensitivity for typing samples with low HBV
5. sequence analysis: gold standard method for genotyping, identification of patients infected with recombinant genotypes but it is time consuming, technically demanded
6. microarray (DNAChip)
7. fluorescence polarization assay
Diagnosis of hepatitis B infection Acute hepatitis B:
o Detection of HBsAg, symptoms, high serum aminotransferases
o Usually anti-HBc IgM can be detected and HBV DNA is present
o HBeAg can be identified in most cases (of little clinical importance)
chronic HB infection:
o Persistence of HBsAg for more than 6 months
o Commonly diagnosed by laboratory means but not by clinical presentations
Past HBV infection:
o Coexistence of anti-HBs and IgG anti-HBc
Occult HBV infection:
o Persistence of low level of intrahepatic HBV DNA without detectable HBsAg
o It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody
o Detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection (invasive procedture), since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum
o Real-time PCR for serum HBV DNA detection have a adequate sensitivity to identify occult HBV infection in many cases (HBV DNA testing is widely used to diagnose occult HBV infection)
o Importance of occult HBV infection:
1. it can be transmitted via transfusion, SOT including orthotopic liver transplantation, or hemodialysis
2. reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3. it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4. it is a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis
o Tests for occult HBV infection in:
1. patients with cryptogenic liver disease, especially when having anti-HBc in serum
2. in patients considering immunosuppression therapy or chemotherapy
3. in solid organ transplantation donors, due to the possibilities for transmission
Conclusions
o Serological markers for detecting antigens and antibodies are the first step of HBV diagnosis
o To prove first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used
o Diagnosis of HBV infection is important for determining acute, chronic and occult cases of infection in order to establish preventive measures and to start antiviral treatment
Diagnosis of Hepatitis B ===> Summary of This Article:
Introduction:
Hepatitis B infection is a one of the major problems worldwide that leads to liver cirrhosis and hepatocellular carcinoma, Globally, about 240 million people are living with a chronic state of the virus which could progress to liver cirrhosis or HCC, HBV is a double-stranded virus that belongs to the family Hepadnaviridae, The presence of vaccination is active method to decrease the burden of the disease. Mode of transmission:
Parenteral route Vertical route Sexual route HBV Endemicity:
High endemicity, affects more than 8% population – China, Southeast Asia, Indonesia, Sub-Saharan African
Intermediate, affects between 2-7% population – South American, Southwest Asia, Eastern, and Sothern Europe
Low endemicity, the prevalence of 0.5 -2% – North American, Western Europe
The identification of serological markers allows:
To identify patient with HBV infection
To know the natural course of chronic HBV infection
To assess the clinical phases of infection
To assess antiviral therapy
Serological Markers for HBV infection:
HBsAg – serological hallmark of HBV infection, After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks, its persistence in the serum more than 6 months denotes chronic infection, it is elevated in the patient positive HBeAg chronic HBV CHB more than in CHB with negative HBeAg.
Anti-HBs – In vaccinated patient, neutralizing antibody that provide long-life immunity. In past infection, anti-HBs is present together with anti-HBc IgG
HBsAg and Anti-HBs when present together denotes carrier status of HBV, as antibodies are unable to neutralize the virus.
HBeAg – use for viral infectivity.
Anti-HBe – use for viral infectivity.
Anti-HBc IgM and IgG
HBV DNA replace HBeAg as a measure for infectivity.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
After 6 months of acute infection, anti-HBc IgM disappeared and Anti-HBc IgG is detected in both patients with resolved HBV infection and CHB
Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive, in the following situations: – Predominantly seen as IgM class during the window period of acute phase. – After acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
– After several years of chronic HBV infection, HBsAg has diminished to undetectable levels. – In isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure. – HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection:
HBV DNA is the direct measurement of viral load (replication activity). DNA can be detected early in a month after infection and reach peak at 3 months. Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC. HBV DNA is a useful marker for starting the treatment. DNA quantified by PCR (standard method). HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment. Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting. Furthermore, it can be fully automated and does not generate carry-over contamination. HBV genotyping:
It has 10 genotypes with distinct geographical distribution, and significantly predict disease progression and determine the suitable antiviral therapy.
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC. In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
Patients with genotype B or C had a lower opportunity to gain serological response to tenofovir. Diagnosis of hepatitis B infection:
A- Acute hepatitis B is a clinical diagnosis identified by : the detection of HBsAg, symptoms, high serum aminotransferases, Usually anti-HBc IgM can be detectedHBV DNA is present.
HBeAg can also be identified in most acute phase of infections
B- Chronic HBV is based on the persistence of HBsAg for more than 6 months, Anti HBs
Ant HBc IgG
C- Past infection diagnosed by presence of anti-HBs and anti-HBc IgG antibodies.
D- Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg, by serological method, isolated anti-HBc is present with the absence of HBsAg and antiHBs antibody. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, means virus remains and detected in the liver not in the serum.
E- Occult HBV infection – it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation, or hemodialysis. – Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
– it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
– it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis.
Tests for occult HBV infection are considered in the following conditions:
– In patients with cryptogenic liver disease, especially when having anti-HBc in serum.
– In patients considering immunosuppression therapy or chemotherapy.
– In solid organ transplantation donors, due to the possibilities for transmission.
Occult HBV infection: Persistent low level of intrahepatic HBV DNA but absent HBsAg and anti-HBs, Can be transmitted by blood transfusion of SOT HBV can be reactivated in patient on immunosuppressive it can accelerate liver injury and can lead to HCC Conclusion:
Both quantitative (HBV DNA for viral load) & qualitative molecular tests are needed to identify viral load and identify HBV genotypes, All these will help to establish if the infection is acute or chronic & to establish preventive methods and effective antiviral treatment and and monitor responses to antiviral agent
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
Introduction: Hepatitis B Virus (HBV) isa 30-42 nm diameter hepadnaciridae family member having an icosahedral capsid core and outer lipid envelope containing hepatitis B surface antigen (HbsAg). It has circular, partly double-stranded DNA with 4 overlapped open reading frames (S, pre-C/C, P, and X). Mutation rate is high due to error susceptibility of the reverse transcriptase involved in HBV replication. HBV causes most of the CLD (chronic liver diseases: cirrhosis and hepatocellular carcinoma, HCC) worldwide, and is spread through sexual, parenteral, and vertical routes. HBV endemicity can be high (>8%: China and South-East Asia), intermediate (2-7%: South America, Eastern and Southern Europe), and low (0.5-2%: North America and Western Europe). Early diagnosis of HBV infection, and vaccination against HBV can be helpful in decreasing the burden of HBV infection.
Serological markers of HBV infection: These include HBsAg, Anti-HBs antibody, HBeAg, AntiHBe antibody, AntiHBc antibody (IgG and IgM).
HBsAg: hallmark of HBV infection, appears within 1-10 weeks of exposure to HBV infection. Its presence for more than 6 months signifies chronic HBV infection. Its titers are higher in HBeAg positive patients.
AntiHBs antibody: It can be acquired either post-vaccination, or post-HBV infection, and provides long-term immunity. Presence of both HBsAg and AntiHBs antibody signifies chronic HBV carrier state.
HBeAg and AntiHBe antibody: Were useful to indicate infectivity and viral replication (not used nowadays due to availability of HBV DNA assays). HBeAg to Anti HBe antibody conversion signifies remission of hepatic disease.
HBcAg: It is intracellular, hence not seen in serum.
AntiHBc antibodies (IgG and IgM): They are seen 1-2 weeks after HBsAg (in conjunction with elevated hepatic enzymes), with disappearance of IgM after 6 months while the IgG remains. AntiHBc antibody alone is seen in window period of acute phase (IgM alone), in post-acute phase where AntiHBs antibody becomes undetectable, and in chronic HBV infection after many years when HBsAg levels become undetectable.
Molecular methods for HBV infection: HBV DNA assays should be done in CLD patients without HBsAg to find out occult HBV infection.
HBV DNA can be detected within 1 month of infection, with peak levels at 3 months reduces. High titres signify more rapid disease progression and increased risk of HCC. The 2 principles of techniques used to identify and quantify HBV DNA include signal amplification (hybrid capture and branched DNA technology) and target amplification (polymerase chain reaction, PCR).
HBV genotyping: There are 10 different genotypes of HBV (A-J) due to high genetic heterogeneity. Genotype A and D are present everywhere; B &C are present in Asia and Oceania; E, F, G & H are occasionally present in Asia; I is unusual, present in Vietnam, India, Laos, & China; J is seen in Japan and Ryukyu.
Genotype A & D have increased rates of chronicity; A & B have better response to interferon treatment; B & C have lower response to Tenofovir; Genotype C s associated with perinatal infection as well as severe liver disease (cirrhosis and HCC).
Diagnosis of HBV infection:
Acute hepatitis B: Symptoms with HBsAg positive, and high serum aminotransferases. Anti HBc IgM and HBV DNA are also present usually.
Chronic infection: Persistent HBsAg for more than 6 months.
Past HBV infection: Presence of AntiHBs antibody and Anti HBc IgG antibody.
Occult HBV infection: HBsAg and Anti HBs antibody negative, low level intrahepatic HBV DNA positive, Serum HBV DNA PCR positive, and positive Anti HBc antibody. It can be transmitted via blood transfusion, solid organ transplantation, or hemodialysis. Chemotherapy and immunosuppression can lead to its reactivation. It can accelerate hepatic disease progression, as well as increase risk for HCC. It should be especially looked for in patients with cryptogenic liver disease, in patients being considered for immunosuppression, and in prospective solid organ donors.
Conclusion: HBV infection is common, and has grave implications for liver. HBV diagnosis involves serological testing followed by use of molecular tests to verify the diagnosis, which will help in preventive and therapeutic tool use to reduce the adverse effects of HBV infection.
Introduction
· HBV is partly double-stranded DNA with a high mutation rate.
· HBV infection is responsible for the most of chronic liver diseases worldwide.
· In addition to HBV vaccination, the diagnosis of acute, chronic and occult HBV infection, diminish the burden of this disease. Serological markers for HBV infection HBsAg
· It is the serological hallmark of HBV infection.
· It appears in serum within 1 to 10 weeks after an acute exposure to HBV.
· If detected for more than 6 months, it implies chronic HBV infection.
· Monitoring of quantitative HBsAg levels predicts treatment response to interferon. Anti-HBs
· Anti-HBs confers long-term immunity.
· After vaccination, anti-HBs is the only serological marker detected in serum.
· If present with anti-HBc IgG, this indicates past HBV infection.
· If anti-HBs antibodies are unable to neutralize the circulating viruses, these patients are regarded as carriers of HBV. HBeAg and anti-HBe
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
· In the past were used to detect infectivity and viral replication, but now replaced by HBV PCR.
HBcAg
· It is present intracellulary, so it is not identified in the serum.
· After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB. Molecular methods for HBV infection
HBV DNA
· A direct measurement of the viral load.
· It is detectable at the early as stage 1 month after HBV infection.
· It disappears at the recovery from HBV infection.
· HBV-DNA can help detection of infection in serologically negative HBV infection, HBeAg-negative CHB and occult HBV infection.
· Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
· HBV DNA testing is useful to determine patients who need antiviral therapy and monitor them for treatment
· Real-time PCR is the standard method to detect and quantify HBV DNA. HBV genotyping
· HBV has a high genetic heterogeneity.
· HBV can be divided into ten genotypes (labelled A–J).
· HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy
· Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
· Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
Diagnosis of hepatitis B infection
· Acute hepatitis B is identified by the detection of HBsAg, symptoms and high serum aminotransferases.
· Usually anti-HBc IgM can be detected and HBV DNA is present in the acute state
· The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
· Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
· Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. Serologically there is isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
· HBV DNA testing is widely used to diagnose occult HBV infection.
· Tests for occult HBV infection are considered in patients with cryptogenic liver disease, in patients considering immunosuppression therapy or chemotherapy and in solid organ transplantation donors.
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
Serological markers for HBV infection consist of
HBsAg,anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection
Differences in the serum HBsAg levels during the different phases of infection indicate the distribution of cccDNA during the respective phases of the disease.
The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs confers long-term immunity -acquired immunity through vaccination,
the simultaneous appearance of HBsAg and anti-HBs has been reported in patients with HBsAg positive In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
isolated anti-HBc positive.
1-it can be predominantly seen as IgM class during the window period of acute phase.
2-after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
3- after several years of chronic HBV infection, HBsAg has diminished to undetectable levels. If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg
Molecular methods for HBV infection HBV DNA – a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
– higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
– HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment
principles of techniques to identify and quantify HBV DNA:
signal amplification such as hybrid capture and branched DNA technology; target amplifcation such as polymerase chain reaction (PCR)
Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting.
HBV can be divided into ten genotypes, labelled A–J: geographic distribution
Genotype B and C are restricted to Oceania and Asia,
genotype A and D most common in Africa and Europe .
Genotype I is unusual and can be observed in Vietnam, Laos, India and China
genotype J has been reported in Japan and Ryukyu
Other genotypes such as E, F, G, and H are also occasionally found in Asia.
Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy. Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D . Recent studies reported that patients infected with genotype B or C had a lower opportunity to gain serological response to tenofovir
The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identifed in the liver but not in the serum. However, gaining hepatic HBV DNA is diffcult in clinical setting since the procedure is invasive.
Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases; thus, HBV DNA testing is widely used to diagnose occult HBV infection
Occult HBV infection clinical importance.
1- it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis
2- reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3- it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4- it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fbrosis
Summary Introduction
HBV belongs hepadnaviridae family.
The reverse transcriptase responsible for viral replication is susceptible to errors thus high mutation rate.
HBV is transmitted parenteral, sexual and vertical route.
HBV is divided into three endemicity: high, intermediate and low; with high endemicity in sub saharan Africa and south east Asia.
Serological markers
HBsAg is the serological hallmark of HBV infection.
After acute infection it appears within 1-10 weeks and persistence beyond 6 months indicates chronic infection.
Anti-HBs is a neutralising antibody, confers longterm immunity.
Vaccinated patients only have anti-HBs.
Anti-HBs may appear simultaneous with HBsAg in carriers.
HBeAg and anti-HBe prior used to indicate replication and infectivity however has been replaced by HBV DNA.
HBeAg seroconversion to Anti-HBe indicates remission.
HBcAg is an intracellular antigen thus not detected in serum.
After acute infection IgM Anti-HBc appears within 1-2 weeks, and wears off in 6 months.
IgG Anti-HBc indicates chronic and resolved infection.
Anti-HBc may occur isolated without HBsAg and in the absence of AntiHBs.
Molecular methods
HBV DNA indicates the replication of the virus.
Detected 1 month after acute infection, peaks in 3 months then declines in chronic infection.
High titers are associated with more rapid progression and development of hepatocellular carcinoma.
Useful to determine patients who are in need of antiviral therapy and to monitor response to treatment.
HBV genotype
There are 10 genotypes A-J with distinct geographical distribution.
Genotype A and D associated with higher rates of chronicity than B and C.
Genotype A and C have better response to interferon than B and D.
Diagnosis
Acute hepatitis B infection is a clinical diagnosis based by the presence of HBsAg detection, raised serum aminotransferase and symptoms.
IgM anti-HBc may be present.
HBV DNA may be detectable.
Detection of HBeAg has no clinical significance.
Chronic hepatitis detected by persistence of HBsAg beyond 6 months.
Past infection identified by Anti-HBs and IgG anti-HBc antibodies.
Occult infection defined by detection of HBV DNA intra-hepatic.
Presence of Anti-HBc without HBsAg or anti-HBs.
Can be transmitted via solid organ transplantation, hemodialysis, or transfusion.
Testing should be done in patients with cryptogenic liver disease with anti-HBc, and donors for solid organ transplant.
1- HBV is one of the major problem worldwide that leads to liver cirrhosis and hepatocellular carcinoma HCC
2- It is double stranded DNA, has different serological markers.
3- Infection can be acute, chronic and occult cases.
4- First step to diagnose it is by serological marker then to quantify the load of the virus.
5- The presence of vaccination is active method to decrease the burden of the disease.
Serological markers for HBV infection
1- HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks, its persistence in the serum more than 6 months denotes chronic infection, it is elevated in the patient positive HBeAg chronic HBV CHB more than in CHB with negative HBeAg.
2- Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels ALT.
3- Anti-HBs it is neutralizing antibody, and provide long-life immunity
4- In vaccinated patient, the only marker appeared is Anti-HBs
5- In past infection, anti-HBs is present together with anti-HBc IgG
6- HBsAg and anti-HBs when present together denotes carrier status of HBV, as antibodies are unable to neutralize the virus.
7- HBeAg means infectivity.
8- Anti-HBeAg means seroconversion or remission of hepatic disease
9- HBV DNA replace HBeAg as a measure for infectivity
10- HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum
11- During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
12- After 6 months of acute infection, anti-HBc IgM disappeared and Anti-HBc IgG is detected in both patients with resolved HBV infection and CHB
13- Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive, in the following situations:
a- predominantly seen as IgM class during the window period of acute phase.
b- after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
c- after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
14- In isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
15- HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
1- HBV DNA is a direct measure to replication and infectivity of the virus, it started to appear after 1 month of acute infection, and reach peak after 3 months of acute infection, then decreased in chronic stage and disappear in the recovery stage.
2- higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
3- HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
4- Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting. Furthermore, it can be fully automated and does not generate carry-over contamination.
HBV genotyping
1- It has 10 genotypes with distinct geographical distribution, and significantly predict disease progression and determine the suitable antiviral therapy.
2- Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
3- Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC
4- In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
5- Patients with genotype B or C had a lower opportunity to gain serological response to tenofovir.
Diagnosis of hepatitis B infection
1- Acute hepatitis B is a clinical diagnosis identified by :
a- the detection of HBsAg
b- symptoms
c- high serum aminotransferases
d- Usually anti-HBc IgM can be detected
e- HBV DNA is present.
f- HBeAg can also be identified in most acute phase of infections
2- Chronic infection is based on the persistence of HBsAg for more than 6 months.
3- Past infection diagnosed by presence of anti-HBs and anti-HBc IgG antibodies.
4- Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg, by serological method, isolated anti-HBc is present with the absence of HBsAg and antiHBs antibody . The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, means virus remains and detected in the liver not in the serum.
5- Occult HBV infection
a- it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis.
b- reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
c- it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
d- it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis.
6- Tests for occult HBV infection are considered in the following conditions:
a- in patients with cryptogenic liver disease, especially when having anti-HBc in serum
b- in patients considering immunosuppression therapy or chemotherapy
c- in solid organ transplantation donors, due to the possibilities for transmission
I make note of your interpretation of virology reports in relation to timings of test and viral load pertaining to HBV infections I like your detailed well-structured summary.
This article is focussed on the diagnosis of hepatitis B infection. Hepatitis B is a serious infection that can lead to significant liver damage, cirrhosis and hepatocellular carcinoma. This infection is caused by a double stranded DNA virus with different serological markers. Disease transmission is via sexual, parenteral, or vertical route. The way to fight the infection is early diagnosis and treatment.
Discussion
Hepatitis B identification of serological markers is significant for effective management of disease. This allows the physician to make suer that the natural course of infection is dealt with appropriately, while monitoring the clinical phases and response to antiviral therapy.
HBsAg is a serological marker of HBV infection and appears in serum 1 to 10 weeks of exposure. If the marker is still seen in serum after 6 months, it means that the patient has chronic HBV infection.
Another serological marker that is significant to determine infection is anti-HBs. It is the only marker that can be seen in patients who have been vaccinated against Hepatitis B.
During acute infection, IgM and IgG emerge 1-2 weeks after HBsAg is seen in serum, along with raised serum aminotransferase and symptoms. After 6 months, IgM starts reducing. IgG continues even after infection is resolved.
Occult infection is characterized by existence of detectable HBV DNA without serum HBsAg. HBV DNA assay is to be done to identify if chronic liver disease patients have occult HBV infection.
Two methods of detection of HBV DNA in any patient include methods that quantify HBV DNA. These include :
signal amplification method – hybrid capture, branched DNA tech
target amplification – PCR method which can detect a wide range of viral load. PCR is the standard method to detect and quantify HBV DNA.
Conclusion
Both quantitative and qualitative molecular tests are needed to identify viral load and identify HBV genotypes In order to establish preventive methods and effective antiviral treatment.
Hepatitis B infection is a public health concern in view of its grave complication and lack of treatment to cure for now even though vaccination is available. Globally, about 240 million people are living with a chronic state of the virus which could progress to liver cirrhosis or HCC. HBV is a double-stranded virus that belongs to the family Hepadnaviridae.
Mode of transmission
Parenteral route
Vertical route
Sexual route
HBV Endemicity
High endemicity, affects> 8% population – China, South East Asia, Indonesia, Sub-saharan African
Intermediate, affects between 2-7% population – South American, South West Asia, Eastern, and Sothern Europe
Low endemicity, the prevalence of 0.5 -2% – North American, Western Europe
Serological Markers for HBV infection
HBsAg – serological hallmark of HBV infection
anti-HBs – neutralizing antibody that confers long term immunity
HBeAg – use for viral infectivity
anti-HBe – use for viral infectivity
anti-HBc IgM and IgG
The identification of serological markers allows:
to identify patient with HBV infection
to know the natural course of chronic HBV infection
to assess the clinical phases of infection
to assess antiviral therapy
Molecular methods for HBV Infection
HBV DNA
usually detected at early stage of infection x 1 month and gets to pick at 3 months
detection in the blood is reliable marker of virus replication
very useful in patient that needs antiviral treatments and to monitor drug effect
Diagnosis of HBV infection
A) Acute hepatitis is diagnosed by
detection of HBsAg
high serum aminotransferases
detection of anti-HBc IgM
B) Chronic HBV
persistent HBsAg > 6 months
anti HBs
ant HBc IgG
Occult HBV infection
persistent low level of intrahepatic HBV DNA but absent HBsAg and anti-HBs
Can be transmitted by blood transfusion of SOT
HBV can be reactivated in patient on immunosuppressive
it can accelerate liver injury and can lead to HCC
Conclusion
The total care for HBV infection is not to use serological markers to make diagnosis, but the use of HBV DNA for viral load, and identification of the genotype is paramount. All these will help to establish if the infection is acute or chronic and to also administer and monitor responses to antiviral agent use for treatment.
I make note of your interpretation of virology reports in relation to timings of test and viral load pertaining to HBV infections I like your detailed well-structured summary.
Hepatitis B virus (HBV) infection is major public health concerns as it causes liver diseases in form of acute, chronic, cirrhosis & hepatocellular carcinoma.
HBV is a member of hepadnaviridae family
HBV structure composed of outer lipid envelope (hepatitis B surface antigen) and icosahedral caspid core (Viral genome & DNA)
HBV DNA is circular, and partly double stranded DNA
Around 240 million people are affected world wide
Prevalence is high (> 8% of population) in China, South East Asia, Indonesia, and sub-sharahan Africa. Intermediate (between 2 to 7%) in South America, South West Asia, Eastern & Southern Europe. Low (0.5 to 2%) in North America & Western Europe.
Serological markers for HBV infection
1.HBSAg
Serological hallmark of HBV infection
Appear 1 to 10 weeks after exposure to HBV
HBSAg +ve > 6 months is consistent with chronic HBV
Titers are higher in those with HBeAg +ve compared to HBeAg -ve
2.Anti-HBsAb
Neutralizing antibody and confer long lasting immunity
Used to be marker of infectivity & viral replication
HBeAg to anti-HBeAb seroconversion = remission
4.HBcAg
It is present in the infected liver cells (hepatocytes)
Both anti-HBcIgM & anti-HBcIgG appears in the blood within 1 to 2 weeks after the presence of HBsAg
Anti-HBIgM disappeared after 6 months
Molecular diagnosis
Positive HBV DNA is a reliable marker of viral replications
Elevated levels are associated with progressive liver disease and risk of hepatocellular carcinoma
Two techniques: signal amplification & target amplification
HBV genotyping
Ten genotype: A to J distributed world wide
It is important to predict disease progression & make therapeutic decisions e.g., genotype A/D associated with chronic liver disease, genotype B/C are less likely to respond to tenofovir
Chronic HBV infection: HBsAg > 6 months, may be asymptomatic
Past / Previous HBV infection: Anti-HBsAg + Anti-HBcIgG
Occult HBV infection
Low HBV DNA in the liver + anti-HBcIgG + negative HBsAg
May be diagnosed by serum HBV DNA due to challenges of getting the hepatic HBV DNA (invasive)
Clinical significance of occult HBV infection:
Can be transmitted to other people by blood, organ transplantation or hemodialysis
It may reactivate in immune-compromised host or cancer chemotherapy
It may deteriorates liver disease in the presence of co-infections e.g., HCV
It may predispose to HCC
Conclusion
The early diagnosis of HBV infection is of paramount importance to prevent or reduced the morbidity and mortality associated with this infection by therapeutic interventions.
Diagnosis of Hepatitis B Summary of the Article Introduction A DNA virus contains the following
Viral capsid and DNA genome which includes the following
S that encodes for surface antigen (HBsAg).
Pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg).
P for polymerase including reverse transcriptase.
X that encodes for a transcriptional transactivator factor HBxAg).
2. The covalently closed circular DNA (cccDNA) is the transcriptional template of HBV. 3. Reverse transcriptase is involved in the replication of HBV. HBV is responsible for most liver diseases worldwide and about 240 million people are infected by chronic HBV. According to the HBsAGs prevalence, HBV endemicity is divided into three categories;
High endemic areas; (China, South East Asia, Indonesia, and Sub-Saharan Africa) with a prevalence of HBV is >8% of the general population.
Intermediate area; (South America, South West Asia, Eastern, and Southern Europe) with an incidence between 2-7%.
Low endemic region; (North America, Western Europe) with the prevalence of about 0.5-2%.
Serological Marker of HBV infection
HBsAg.
anti-HBs.
HBeAg.
anti-HBc IgM and IgG.
The identification of the marker alow;
To diagnose the infection.
To address the natural course of chronic hepatitis B.
To assess the clinical phases of the infection.
To monitor anti-viral therapy.
HBsAg is the hollmarker of HBV infection, HBsAg appear on the serum within 1-10 days, and the infection considered chronic if the NBsAg detected in the serum for than 6 months, so;
Several studies reported association between HBsAglevel and the cccDNA transcription activity.
Serum HBsAg is higher in HBeAg-positive CHB than in HBeAg-negative.
Serum HBsAg levels used to monitor treatment response and disease progression in HBsAg-negative CHB patients with normal ALT.
Anti-HBs is the only serlogical marker that indicate acquired immunity. HBeAg and anti-HBe seroconversion is related to remission of the hepatic disease. HBV DNA is used for viral activity and replication. HBcAg denote hepaticyte infection, and detection of both HBcAg IgM and IgG denote that te infection is during acute phase, and presence of only IgG denote either resolved infection or CHB. In isolatedanti-HBc positive with HBsAg and HBs-negative, a serum IgM should be detected to diagnose the occurrence or not of acute infection. Isolated cases of HBc-positive can denote the following;
During the window period of acute infection as IgM class.
After resolving of acute infection, that anti-HBs can not be detected (low titre).
After several years of HB infection.
Molecular methods for HBV infection
HBV DNA is the measurement of the viral load, it can measured as early as 1 month and reach the peak at 3 months and it disappear in chronic phase or after recovery.
It is an indicators of viral replication and it higher load related to te more rapid progressive disease and higher incidence of HCC.
Used for monitoring therapy and to determine who is for antiviral therapy.
There are several quantitative methods;
Semi-automated qPCR.
Semi-automated real-time PCR.
Automated real time PCR.
Branched DNA.
There are 10 genotype HBV, labelled as A-J;
G (A and D) in Africa and Europe.
G (B and C) in Oceania and Asia.
G (I) in Vietnam, Laos, India, and China.
G (J) in Japan and Ryukyu.
G (E, F, G, and H) in Asia.
G C is generally considered as a risk factor of perinatal infection and elated to a sever liver disease including cirrhosis and HCC.
G A and B have better treatment response with interferon therapy than G C and D.
Recent studies reported that G B and C infection had a lower opportunity to gain serological response to tenofovir.
Diagnosis of HBV infection Acute infection;
Clinical symptoms.
Detection of HBsAg.
High ALT.
anti-HBc IgM can be detected.
HBV DNA is present.
HBeAg can be detected of most acute infection.
CHB;
Presence of HBsAg for more than 6 months.
No clinical presentation.
Past HBV infection;
Coexistance of anti-HBs and IgG anti-HBc.
Occult HBV infection
persistence of low level of intrahepatic HBV DNA with no detectable HBsAg, but it is an invasive procedure, but can identified by real-time PCR .
It has many clinical importance
it can be transmitted via;
Blood transfusion.
Solid organ transplantation.
Hemodialysis.
2.Reactivation of HBV infection in immunocompromised patients.
3.It can accerelate liver injury hepatic fibrosis in patient with CLL. 4.Risk factor of HCC.
Testing for occult infection should be considered in the following conditions;
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family.
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
According to HBsAg prevalence, HBV endemicity is divided into three categories; high, intermediate, low.the high edamic area are China, South East Asia, Indonesia, and sub-Saharan Africa which represent about 8% of general population.
South America, South West Asia, Eastern and Southern Europe which is intermediate about 2% to 7%.
North America and Western Europe which is lowest area of endemicity around .0.5 % to 2 %.
Vaccination is the way to decrease the HBV infection. Serological markers for HBV infection.
HBsAg
is the serological hallmark of HBV infection, after acute infection appears in serum within 1 to 10 weeks.
If remaining for more than 6 month patient diagnosed as chronic HBV infection .
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative chronic hepatitis B patients with normal serum alanine aminotransferase levels. anti-HBsg
called as neutralizing antibody, and confers long-term immunity, and confers long-term immunity. HBcAg
is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. HBeAg
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels.
anti-HBc IgM and IgG
anti-HBc IgM become negative after 6 month of acute HBV infection .
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB. Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
It is appear at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
PCR is test that to detect and quantify HBV DNA which is fully automated . HBV genotyping
HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.genotyping is important to determine response to treatment . Diagnosis of hepatitis B infection
Diagnosis of HBV infection by, symptoms, high serum aminotransferases and the detection of HBsAg.
To diagnose chronic HBV infection is based on the persistence of HBsAg for more than 6 months,in absence of any symptoms . Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
Occult HBV infection is diagnosed by detection of HBV DNA in the liver( invasive ).
occult HBV infection diagnosis also by Real-time PCR for serum HBV DNA( sensitive)
It is important because occult HBV infection transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation and hemodialysis . Conclusion
Hepatitis B virus infection is a major health problems and causing severe liver disease such as cirrhosis and hepatocellular carcinoma .
Vaccination is the important to limited infection of HBV infection .
This article define what is HBV ,how to diagnose ,occult HBV infection .
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family and has a diameter of 30-42 nm and an icosahedral capsid core composed of protein.
It is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical routes.
About 240 million people are chronically infected, and HBV endemicity is divided into three categories: high, intermediate, low.
Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection.
Serological markers for HBV infection
Serological markers for HBV infection include HBsAganti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg is the serological hallmark of HBV infection, and persistence of this marker for more than 6 months implies chronic HBV infection.
Anti-HBs is a neutralizing antibody that confers long term immunity and is the only serum marker detected in patients with acquired immunity through vaccination.
It is present in concurrence with anti-HBc IgG, but is unable to neutralize the circulating viruses.
HBcAg is an intracellular presence in infected hepatocyte, and anti-HBc IgM and IgG emerge 1-2 weeks after the presence of HBsAg and wear off after 6 months of acute infection.
HBV DNA assays should be tested in chronic liver disease patients to detect occult HBV infection.
Molecular methods for HBV infection
HBV DNA is a reliable marker of replication activity and is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
There are two principles of techniques to identify and quantify HBV DNA: signal amplification and target amplification.
PCR is the standard method to detect and quantify HBV DNA in clinical setting, as it is fully automated and does not generate carry over contamination.
HBV genotyping
HBV genotyping is important to predict disease progression and determine appropriate antiviral therapy.
It can be confirmed using diverse methods such as reverse hybridization, genotype-specific PCR assays, real time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fluorescence polarization assay.
Diagnosis of hepatitis B infection
Diagnosis of hepatitis B is based on HBsAg, symptoms, and high serum AMTransferases, while chronic infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is characterized by low levels of intrahepatic HBV DNA without detectable HBsAg and anti-HBs antibody.
HBV DNA testing is used to diagnose it.
Occult HBV infection can be transmitted via transfusion, solid organ transplantation, chemotherapy, or hemodialysis, and can accelerate liver injury and lead to hepatic fibrosis.
Tests for occult HBV infection are recommended in cryptogenic liver disease, immunosuppression therapy, and solid organ transplantation donors.
Conclusions
Diagnosis of HBV is an important tool to establish preventive remedies and initiate antiviral treatments.
HBV is a virus from hepadnaviridae family, with a circular, partly double stranded DNA, the covalently closed circular DNA (cccDNA) is the transcriptional template of HBV and stays inside the hepatocyte nucleus as a mini-chromosome.
HBV infection is the most common cause of chronic liver diseases worldwide, transmitted through parenteral, sexual and vertical route, and about 240 million populations with chronic HBV infection, making them prone to HCC and cirrhosis.
Serological markers for HBV infection:
HBsAg is the serological hallmark of HBV infection, HBsAg appears in serum within 1 to 10 weeks. Persistence for more than 6 months indicates chronic HBV infection. q
Quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels.
Anti-HBs Ab did not neutralize the circulating viruses, thus these patients are regarded as carriers of HBV, or immunized.
HBcAg = not detected in serum, indicates intracellular presence in infected hepatocyte.
Anti-HBc IgM and IgGrise in 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms during acute infection. After 6 months of acute infection. anti-HBc IgM becomes negative, and Anti-HBc IgG positive in resolved HBV infection and Chronic HB.
Isolated anti-HBc Abs, can be seen in three conditions:
During the window period of acute phase, predominantly IgM.
At the end of acute infection, with very low anti-HBs Ab.
After several years of chronic HBV infection, HBsAg has diminished.
Molecular methods for HBV infection:
There are two principles of techniques to identify and quantify HBV DNA:
Signal amplification such as hybrid capture and branched DNA technology.
Target amplifcation such as polymerase chain reaction (PCR).
Real-time PCR is the standard of care to detect and quantify HBV DNA in clinical setting.
Higher titers of HBV DNA, related to disease progression and higher incidence of HCC. HBV DNA can detect occult HBV infection in the absence of HBsAg. HBV genotyping:
Genotyping done the following methods:
RFLP (restriction fragment length polypmorphism) Easily done, low cost, simple, rapid Low sensitivity for typing samples with low HBV.
Reverse hybridization High sensitivity, automated systems Relatively high cost.
Genotype specific PCR High sensitivity, automated systems, easy to perform, suitable for detecting mixed genotype infections High cost.
Sequence analysis Gold standard method for genotyping, identification of patients infected with recombinant genotypes Time consuming, technically demanded.
Utility of genotyping:
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
Genotype B or C had a lower opportunity to gain serological response to tenofovir.
Interferon therapy, has better therapeutic response ingenotypes A and B, than genotypes C and D.
Diagnosis of hepatitis B infection:
Acute HBV infection= HBsAg, symptoms and elevated transaminases.
Chronic HBV infection= persistence of HBsAg> 6 months+ anti HBs+ anti HBc Ab.
Occult HBV infection= persistence of low level of intra-hepatic HBV DNA without detectable HBsAg. It is a serological situation defined by +ve anti-HBc Ab and -ve both HBsAg and antiHBs Ab.
Clinical importance of Occult HBV infection are:
Can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation, or hemodialysis.
Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
Accelerates liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
A risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis.
Tests for occult HBV infection are considered in the following conditions:
in patients with cryptogenic liver disease, especially when having anti-HBc in serum.
in patients considering immunosuppression therapy or chemotherapy.
in solid organ transplantation donors, due to the possibilities for transmission
Conclusion: The article identify the structure of HBV and tools used for quantitative and qualitative (gentypes) testing of the virus, and defining and better understanding the acute, chronic and occult HBV infection, thus; help to prevent and treat the HBV.
Introduction:
-Hepatitis B virus (HBV) infection is a global health problems and responsible for majority of chronic liver disease.
-Its prevalence varies according to geographic distribution, and can be divided into high, intermediate, low.
-HBV is a circular, partly double-stranded DNA virus with various serological markers.
-Hepatitis B vaccination, accurate diagnosis and treatment methods to decrease disease burden.
Serological markers for HBV infection:
-Markers allow to identify infected patients; to elucidate the natural course of infection; to assess the clinical phases of infection; and to monitor antiviral therapy.
-Serological markers for HBV infection consist of:
HBsAg:
– It is the serological hallmark of HBV infection. Appears in serum within 1 to 10 weeks.
– Persistence elevation > 6 months implies chronic infection.
– Patients with HBeAg positive CHB have higher HBsAg level.
– Its monitoring predicts treatment response and disease progression in HBeAg negative CHB patients.
Anti-HBs
– It is neutralizing antibody, and confers immunity.
– Concurrence with anti-HBc IgG ; indicate past HBV infection.
– Carriers of HBV have it simultaneously with HBsAg, when virus was not neutralize completely.
HBeAg and anti-HBe
– Previously used to indicate infectivity and viral replication, but HBV DNA their taking this role currently.
– Seroconversion from HBeAg to anti-HBe indicates remission, however, active viral replication can still occurs due to mutation inhibit or decrease HBeAg.
HBcAg; (intracellular) not present in the serum.
Anti-HBc (IgM &IgG)
– In acute infection emerges 1–2 weeks after the presence of HBsAg and IgM weaned off after 6 months.
– Anti-HBc IgG; detect in both resolved HBV infection and CHB.
– Isolated anti-HBc can be seen in:
– IgM class during the window period of acute phase.
– Anti-HBs below the cutoff level of detection ( after acute infection).
– HBsAg has diminished to undetectable levels (chronic HBV infection)
Molecular methods for HBV infection HBV DNA
-Measure directly viral load and indicated replication activity and disease progression.
-Detected early (1 mo), peaked at (3 mo) then gradually diminishes in chronic infection or disappearsat the recovery.
– Real-time PCR; standard method to detect and quantify HBV DNA.
HBV genotyping:
– It hashigh genetic heterogeneity, 10 genotype A-J.
– Genotyping predict disease progression and treatment response. – Genotype A and D leads to higher rate of chronicity than B,C
– Genotype C is risk for perinatal infection , cirrhosis and HCC. – Response to interferon therapy better in type A&B than C&D.
-Tenofovir response less in type B&C.
Diagnosis of hepatitis B infection
– Acute hepatitis B: is a clinical diagnosis with HBsAg + symptoms+ high aminotransferases. anti-HBc IgM and HBV DNA is present.
– Chronic infection; the persistence of HBsAg for > 6 months.
– Past HBV infection; both anti-HBs and IgG anti-HBc.
– Occult HBV infection; persistence of low HBV DNA without detectable HBsAg.
-Clinical importance of occult HBV infection:
– Viral transmission; via transfusion, organ transplantation or hemodialysis.
– Reactivation of HBV infection in immunocompromised
– Accelerate liver injury and lead to hepatic fibrosis (
– Risk factor for HCC.
-Tests for occult HBV infection are indicated in cryptogenic liver disease, in considering immunosuppression therapy, and in SOT donors,
I make note of your interpretation of virology reports in relation to timings of test and viral load pertaining to HBV infections
I like your detailed well-structured summary.
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family and has a diameter of 30-42 nm consisting of a lipid envelope containing HBsAg that surrounds an inner nucleocapsid composed of hepatitis B core antigen (HBcAg) complexed with virally encoded polymerase and the viral DNA genome. HBV infection is a common cause of chronic liver diseases worldwide, the mode of transmission mainly through reproductive, sexual and vertical routes.
Serological markers for HBV infection:
There are five serological markers including (HBsAg), (HBeAg), (anti-HBs), (anti-HBe) and (anti-HBc), which are valuable for the diagnosis and monitoring of hepatitis B infection and vaccination efficacy. Of these markers HBsAg is considered as a serological marker of HBV infection; it appears early in the serum within 1 to 10 weeks .Anti-HBs may confer long-term immunity only in some cases .
HBV DNA assays have replaced HBeAg and anti-HBe seroconversion to measure infectivity and viral replication . HBcAg is an intracellular presence in infected hepatocytes, hence it is not detected in patients serum and emerges as IgM and IgG 1-2 weeks after the presence of HBsAg. Usually anti-HBc IgM disappears after 6 months of acute infection ,and its presence after that indicates resolved HBV infection or CHB. After several years of chronic HBV infection, however, HBsAg has diminished to undetectable levels.
Molecular methods for HBV infection:
HBV DNA is a reliable marker of replication activity and is useful in routine clinical settings to determine patients who need antiviral therapy and monitor them for suitable treatment. There are two principles of techniques to identify and quantify HBV DNA: signal amplification and target amplification. PCR can detect a wide dynamic range of viral load and is fully automated and does not generate carry-over contamination.
HBV genotyping:
HBV is divided into ten genotypes, A-J, with B and C restricted to Oceania and Asia. Acute infection with genotypes A and D leads to higher rate of chronicity, while C is a risk factor for perinatal infection and severe liver disease. HBV genotyping can be used to predict disease progression and determine appropriate antiviral therapy.
Diagnosis of hepatitis B infection:
Acute hepatitis : Is a clinical diagnosis identified by the detection of HBsAg, symptoms, and high serumaminotransferases. Chronic HBV infection: Is defined by the coexistence of anti-HBc IgM and HBV DNA. Occult HBV : Is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. HBV DNA testing is widely used to diagnose occult HBV infection, which has some clinical importance due to its transmission via transfusion, solid organ transplantation, or hemodialysis. It may also accelerate liver injury and lead to hepatic fibrosis in patients with chronic hepatitis C infection.
Conclusions: Diagnosis of HBV infection play an important role in determining acute, chronic and occult cases, establish preventive remedies, and initiate antiviral treatment. Serological markers are used to detect antigens and antibodies, and molecular tests are used to quantify viral load and identify genotypes.
Parenteral, sexual, and vertical HBV infection causes most chronic liver disorders globally. 240 million HBV-infected persons are at risk of liver cirrhosis and hepatocellular cancer (HCC). HBsAg prevalence determines HBV endemicity: high, middle, or low.
China, Southeast Asia, Indonesia, and sub-Saharan Africa are highly endemic because over 8% of the population has chronic HBV infection.
South America, South West Asia, and Eastern, and Southern Europe have 2%–7% chronic HBV infection rates.
HBV prevalence is 0.5%–2% in low-endemic regions like North America and Western Europe.
Serological markers for HBV infection:
HBsAg indicates HBV infection.
Serum HBsAg occurs 1–10 weeks after acute HBV exposure. This marker indicates persistent HBV infection after 6 months. Many studies have linked liver cccDNA transcription to blood HBsAg levels. Serum HBsAg values reflect cccDNA distribution during infection stages. HBeAg-positive CHB patients had higher HBsAg titers.
Quantitative HBsAg levels predict interferon response and illness progression in HBeAg-negative CHB patients with normal serum alanine aminotransferase.
HBV molecular techniques:
HBV DNA directly measures viral load, revealing virus replication. It is detected at the early stage of infection (1 month after HBV infection) and grows to a peak level (more than 108 copies/mL) roughly 3 months after HBV exposure. It subsequently progressively decreases in chronic infection or vanishes upon recovery.
HBV-DNA detection has grown in clinical medicine as serologically negative HBV infection (HBeAg-negative CHB and occult HBV infection) has risen. Higher HBV DNA titers indicate replication activity and faster disease progression and HCC development.
HBV DNA testing also helps clinicians identify individuals who require antiviral medication and evaluate their progress.
HBV genotyping:
HBV’s insufficient proofreading of reverse transcriptase causes genetic heterogeneity. Sequence divergence divides HBV into 10 genotypes, A–J, with varied geographic distributions. Genotypes B and C are limited to Oceania and Asia, whereas genotypes A and D are widespread but most frequent in Africa and Europe.
Genotype I is rare in Vietnam, Laos, India, and China, but genotype J is found in Japan and Ryukyu. Asia also sometimes has the genotypes E, F, G, and H.
HBV infection diagnosis:
HBsAg, symptoms, and elevated aminotransferases indicate acute hepatitis B. HBV DNA and anti-HBc IgM are usually found. HBeAg is seen in most acute infections but is not clinically significant. HBsAg persistence beyond 6 months indicates persistent infection. Laboratory tests confirm persistent HBV infection, not clinical symptoms. Anti-HBs and IgG anti-HBc antibodies indicate HBV infection.
Occult HBV infection is low intrahepatic HBV DNA without HBsAg. Isolated anti-HBc without HBsAg or anti-HBs antibodies defines it serologically. As cccDNA persists in the hepatocytes and HBV DNA is sometimes found in the liver but not in the blood, liver HBV DNA testing is the gold standard for detecting occult HBV infection.
Conclusions:
We explained HBV serological and molecular diagnostics in this article. HBV diagnosis begins with serological indicators for antigens and antibodies. Qualitative or quantitative molecular testing validates the diagnosis, measures the viral load, and identifies genotypes.
To prevent and cure acute, chronic, and hidden HBV infections, diagnosis is crucial.
I make note of your interpretation of reports pertaining to HBV infections. I like your summary.I appreciate that you mention genotypes, A–J, with varied geographic distributions.
Diagnosis of hepatitis B: Introduction;
-HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
-About 240 million people are chronically infected by HBV, so have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC).
-According to HBsAg prevalence, HBV endemicity is divided into three categories; high, intermediate, low.
-China, South East Asia, Indonesia, and sub-Saharan Africa are regarded as highly endemic areas because chronic HBV infection is reported in more than 8% of the population. HBV genotyping HBV;
-HBV can be divided into ten genotypes, labelled A–J; according to the sequence, variation, and they have distinct geographic distribution.
-Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
-Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
-Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
-In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
-Recent studies reported that patients infected with genotype B or C had a lower opportunity to gain serological response to tenofovir. Methodology for HBV genotyping;
-It is confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNA Chip) and fluorescence polarization assay. Serological markers for HBV infection;
-It consist of HBsAg , anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
-HBsAg is the serological hallmark of HBV infection.
-After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks & persistence of this marker for more than 6 months implies chronic HBV infection.
-HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
-During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
-After 6 months of acute infection, anti-HBc IgM wears off & anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
-In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
-Isolated anti-HBc positive can be seen in three conditions.
*First, it can be predominantly seen as IgM class during the window period of acute phase.
*Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
*Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
-In these conditions, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure. Molecular methods for HBV infection;
-HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
-HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
-It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
-HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg. Diagnosis of hepatitis B infection; Acute hepatitis B infection;
-Identified by the detection of HBsAg, symptoms, high serum aminotransferases.
-Usually anti-HBc IgM can be detected and HBV DNA is present.
-HBeAg can also be identified in most acute phase of infections, but has little clinical importance. Chronic hepatitis B infection;
-Based on the persistence of HBsAg for more than 6 months.
-Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations. Past HBV infection;
-Defined by the coexistence of anti-HBs and IgG anti-HBc. Occult HBV infection;
-Defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
-It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti-HBs antibody.
-The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, however, gaining hepatic HBV DNA is difficult in clinical setting since the procedure is invasive. Importance of diagnosis of occult HBV infection;
-It can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis.
-Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
-It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
-It appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis. Tests for occult HBV infection are considered in the following conditions:
*In patients with cryptogenic liver disease, especially when having anti-HBc in serum,
*In patients considering immunosuppression therapy or chemotherapy,
*In solid organ transplantation donors, due to the possibilities for transmission. Conclusions;
-Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
-This Review Articles on Diagnosis of Hepatitis B with (LOE V)
Introduction
HBV is a virus of Hepadnaviridae family. It has an outer lipid layer with hepatitis B surface antigen (HBsAg) and a capsid core composed of viral genome made of partly double-stranded DNA with 4 overlapping frames including
· S encodes for HBsAg
· pre-C/C for HBeAg and HBcAg
· P for polymerase including reverse transcriptase;
· X that encodes for a transcriptional transactivator factor HBxAg
The covalently closed circular DNA (cccDNA) as a mini chromosome inside the hepatocyte nucleus .
Reverse transcriptase is liable to errors
Methods of HBV infection transfer include parenteral, sexual and vertical routes.
Chronic HBV infection can lead to liver cirrhosis and hepatocellular carcinoma.
High HBV endemic areas as China and Sub saharan Africa.
Intermediate HBV endemic areas as South America, South West Asia
Low HBV endemic areas as North America and Western
Europe . Serological markers for HBV infection
Include HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG to identify the viral infection phase and follow antiviral treatment. HBsAg appears in serum within 1 -10 weeks.
It’s persistence for more than 6 months indicates chronic HBV infection. HBsAg titer monitoring reflects response to interferon and disease progression in HBe Ag negative Chronic Hepatitis B patients with normal serum ALT levels. Anti-HBs
o on it’s own indicates vaccination
o With HBc IgG indicates past infection
o HBsAg and anti-HBs can be detected in HBsAg positive patients
o indicates carriers of HBV if unable to neutralize the virus . HBeAg and anti-HBe
o Used to indicate infectivity but now HBV viral assays are used
o HBeAg conversion to anti-HBe signifies remission HBcAg
is not detected in serum because it is present in infected
hepatocyte.
In acute infection, anti-HBc IgM and IgG appears 1–2 weeks
After HBsAg occurrence with raised liver enzymes.
After 6 months of acute infection, anti-HBc IgM disappears and Anti-HBc IgG continues to be detected in resolved HBV infection and
CHB. HBsAg
o Can be seen during the window period of acute phase with Ig M.
o after acute infection had ended.
o after several years of chronic HBV infection. Molecular methods for HBV infection
HBV DNA can be detected 1 month after HBV infection and reaches peak level 3 months after the exposure to HBV and
Then decreases gradually in chronic infection or disappears on recovery .
HBV DNA indicates rapid disease progression and higher incidence of HCC also it can aid in determining those in need of antiviral therapy and monitor them for the response.
HBV DNA is detected and quantified by signal amplification such as hybrid capture and branched DNA technology; and by target amplification such as PCR.
Meanwhile HBV PCR is the standard test . HBV genotyping
HBV is divided into ten genotypes, labelled A–J differs according to geographic areas.
HBV genotyping can predict HBV disease progression and
Points to proper antiviral therapy. Acute infection with genotypes A and D have higher rate of chronicity than genotypes B and C while C is a risk factor for perinatal infection and severe liver disease.
A and B responds to interferon treatment better than C and D.
B and C have less response to tenofovir.
HBV genotyping is detected by different methods including reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fuorescence polarization assay HBV infection diagnosis In acute hepatitis B infection , HBsAg, anti-HBc IgM, HBV DNA and HBeAg can be detected along with clinical diagnosis and elevated liver enzymes.
In chronic infection , HBsAg persists for more than 6 months without clinical manifestations In past HBV infection, anti-HBs and IgG anti-HBc can be co detected In occult HBV infection, anti-HBc is detected in serum without HBsAg and antiHBs antibody. HBV DNA in the liver is the gold standard of diagnosis but Real-time PCR for serum HBV DNA detection is a sensitive test to diagnose occult HBV infection.
Occult HBV infection detection is essential due to multiple causes including
– transmission of HBV infection through transfusion, solid organ
Transplantation or haemodialysis .
– reactivation of HBV infection particularly withimmunocompromised status .
-worsen liver injury with co infection of HCV infection
-a risk of HCC Conclusion
HBV is diagnosed by serological markers to detect antigens and antibodies . In order to diagnose and quantify viral load and to identify genotypes, qualitative or quantitative molecular tests.
The Hepadnaviridae family of viruses includes the hepatitis B virus (HBV).
It has a diameter of 30-42 nm and is made up of an icosahedral capsid core made of protein and an outer lipid envelope bearing hepatitis B surface antigen (HBsAg).
The viral genome and DNA polymerase with reverse transcriptase activity are both contained in the viral capsid.
Four open reading frames overlap in the circular, partially double-stranded DNA that makes up the HBV genome. (I) S that encodes for surface proteins (HBsAg); (II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg); (III) P for polymerase including reverse transcriptase (IV) X that encodes for a transcriptional transactivator factor (HBxAg). The covalently closed circular DNA (cccDNA) is the transcriptional template of HBV and stays inside the hepatocyte nucleus as a mini-chromosome.
Reverse transcriptase involved in the replication of HBV is error susceptible, the mutation rate is high, observed in retroviruses and RNA viruses.
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
About 240 million people are chronically infected by HBV, so have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC) According to HBsAg prevalence, HBV endemicity is divided into three categories; high, intermediate, low.
South east Asia, Indonasia and sub-Saharan Africa are regarded as highly endemic areas because chronic HBV infection is reported in more than 8% of the population .
Europe are grouped as low endemic regions; in these areas, HBV prevalence rates range from 0.5% to 2% Objectives
We aimed to give informations about HBV
Methods
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level.
HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
There are two principles of techniques to identify and quantify HBV DNA: signal amplification such as hybrid capture and branched DNA technology; target amplification such as polymerase chain reaction (PCR)
Genotype specific PCR High sensitivity, automated systems, easy to perform, suitable for detecting mixed genotype infections High cost.
HBV, hepatitis B virus; RFLP, restriction fragment length polypmorphism; PCR Findings
Infection is reported in more than 8% of the population Conclusion
We aimed to give information about HBV serological and molecular diagnosis.
First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus.
In order to verify first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used.
Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment
Together with a clinical diagnosis and high liver enzyme levels, HBsAg, anti-HBc IgM, HBV DNA, and HBeAg can all be found in acute hepatitis B infections.
HBsAg persists in chronic infection for longer than 6 months without showing any clinical symptoms.
Anti-HBs and IgG anti-HBc can both be found in people with prior HBV infection.
Anti-HBs and IgG anti-HBc can both be found in people with prior HBV infection.
Anti-HBc is seen in serum after occult HBV infection even while HBsAg and antiHBs antibodies are absent.
Nonetheless, Real-time PCR for blood HBV DNA detection is a sensitive test to determine occult HBV infection. HBV DNA in the liver is the gold standard for diagnosis.
Hepatitis B virus ~ Hepadnaviridae family. 30-40 nm outer lipid envelope contains Hbsag icosahedral capsid core – viral genome and DNA polymerase. viral genome – ds DNA Contains S- HbSAg pre- c/c – Hbe Ag and HbcAg P- DNA polymerase x- HBx Ag
HBV infection occurs through Parenteral Sexual Vertical
Serological markers for HBV infection HbsAg – appears in Serum within 1 to 10 weeks after infection persists> 6months – Then chronic infection Higher Hbsag levels in Hbe Ag positive anti – HBs – long term immunity only anti- Hbs positive – Post Vaccination H Be Ag – infectivity anti HBC IgG IgM- Past HBV infection – anti Hbs and anti Hbc IgG positive.
Course Acute Infection Hbsag + 1-2 weeks later anti HBc Igm IgG + after 6months anti Hbc IgM wears off anti Hbc IgG remains+ in CHB and resolved HBV infection.
Some pts HbsAg – anti -Hbs- anti – HBc Ig G +
Isolate Anti – HBc seen in- During window phase of acute infection after acute infection has ended , anti- H Bs has decreased after several years of chronic HBV infection when Hbsag has diminished
So if only anti-HBc + check anti – HBc IgM and HBV DNA
Molecular methods HBV DNA – viral load increases from 1month of infection peak levels at 3months gradually diminishes in chronic infection Or disappears at recovery.
HBV DNA by PCR – standard method hybrid capture and branched DNA technology.
HBV genotyping 10 genotypes A-J genotype A and D infection leads to high rate of chronicity. genotype C – risk factor for perinatal infection also related to severe liver disease like cirrhosis and HCC.
With interferon treatment genotype Aand B do well. genotype B and C – lower response to tenofovir.
Diagnosis of Hepatitis B infection Acute Hep B Hbs Ag + symptoms high aminotransferase anti HBc Ig M + HBV DNA present HbeAg can be positive
chronic Hep B Hbsag persists > 6 months
Past HBV infection anti HBs + anti HBc IgG +
Occult HBV infection H BS Ag – low level of intrahepatic HBV DNA on liver biopsy – gold standard isolated anti- H Bc + anti – Hbs –
o Can be transmitted by – Transfusion hemodialysis Organ donation
o Reactivation may Occur in those receiving chemotherapy.
o may accelerate liver injury
o Risk factor for HCC
Test for occult HBV infection – in solid organ Transplantation tation donors patients with cryptogenic liver disease in patients considering chemotherapy or immunosuppression
Diagnosis of hepatitis B
In this article, the serological and molecular diagnosis of HBV has been reviewed.
Introduction
Hepatitis B virus (HBV) is a DNA virus and is transmitted through parenteral, sexual and vertical route.
Hepatitis B virus (HBV) infection is a major global health problems leading to severe liver disease such as cirrhosis and hepatocellular carcinoma (HCC).
Serological markers for HBV infection
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
The recognition of serological markers helps to:
HBsAg : appears in serum within 1 to 10 weeks of infection.
If persist > 6 months → chronic infection.
The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
Monitoring of it’s levels predicts treatment response to INF and disease progression.
Anti-HBs: a neutralizing antibody, and confers long-term immunity.
The only serological marker detected in serum after vaccination.
In past HBV infection, it is present in concurrence with anti-HBc IgG.
In most cases patients with anti-HBs antibodies are regarded as carriers of HBV.
HBeAg to anti-HBe seroconversion is related to
the remission of hepatic disease
Active viral replication is sustained in some patients with HBe
seroconversion).
HBcAg: is not identified in the serum.
Anti-HBc IgM and IgG: emerges 1–2 weeks
after the presence of HBsAg
After 6 months of acute infection, anti-HBc IgM wears off.
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs.
If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV
exposure.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load.
Detection of HBV DNA is a reliable marker of replication activity, and
higher titers are related to the more rapid disease progression and higher incidence of HCC.
It is useful in routine clinical setting.
Principles of techniques to identify and quantify HBV DNA:
and quantify HBV DNA in clinical setting).
HBV genotyping
HBV has a high genetic heterogeneity (ten genotype labelled A–J with distinct geographic distribution).
Can predict HBV disease progression and determine therapy.
Can be confrmed using diverse methods.
Diagnosis of hepatitis B infection
Acute infection:
Is a clinical diagnosis based on:
Usually anti-HBc IgM and HBV DNA and HBeAg can be detected.
Chronic infection:
Is based on the persistence of HBsAg for more than 6 months.
A laboratory based diagnosis.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection:
Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
A serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
The detection of HBV DNA in the liver is the gold standard of diagnosis.
HBV DNA testing.
Tests for occult HBV infection are considered in:
chemotherapy.
Conclusions
Serological markers for detecting antigens and antibodies against HBV is the first step for diagnosis.
Diagnosis of HBV infection is an important tool to identify
acute, chronic and occult cases of infection.
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg is the serological hallmark of HBV infection which appear after an acute exposure to HBV, within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity . In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum. In the past HBV infection, it is present in concurrence with anti-HBc IgG. Occasionally, the simultaneous appearance of HBsAg and anti-HBs has been reported in patients with HBsAg positive . In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease , however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg .
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB. Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive. It can be seen in three conditions.
1- it can be predominantly seen as IgM class during the window period of acute phase.
2- after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
3- after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg .
Molecular methods for HBV infection HBV include DNA which is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection. The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC . Furthermore, HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment (.
Hepatitis B virus (HBV) is a member of the Hepadnaviridae family and its genome consists of a circular, partially double-stranded DNA. This viral infection is responsible for the majority of chronic liver diseases worldwide and can be transmitted through various routes, including parenteral, sexual, and vertical transmission. HBV can be classified into ten genotypes, designated as A to J, with genotypes A and D being the most prevalent in Africa and Europe. Acute infections caused by genotypes A and D are more likely to progress to chronicity. Genotype C is often associated with an increased risk of perinatal transmission and is linked to severe liver diseases such as cirrhosis and hepatocellular carcinoma (HCC). When it comes to interferon therapy, patients infected with genotypes A and B generally exhibit better treatment responses compared to those infected with genotypes C and D.
Serological markers are used to diagnose and monitor hepatitis B virus (HBV) infection. One of the primary markers is HBsAg, which appears in the serum within 1 to 10 weeks after infection. If HBsAg persists for more than 6 months, it indicates chronic HBV infection. On the other hand, the presence of anti-HBs, a neutralizing antibody, signifies long-term immunity. In individuals who have acquired immunity through vaccination, only anti-HBs is detected in the serum. In cases of past HBV infection, anti-HBs is present alongside anti-HBc IgG.
HBeAg and anti-HBe markers were previously used to assess infectivity and viral replication, but they have been replaced by HBV DNA testing. Another set of markers, anti-HBc IgM and IgG, emerge within 1-2 weeks after the presence of HBsAg, along with elevated serum aminotransferase levels and symptoms. After 6 months of acute infection, anti-HBc IgM disappears, but anti-HBc IgG continues to be detected in individuals with resolved HBV infection.
HBV DNA is a direct measurement of the viral load and serves as a reliable marker of replication activity. Higher titers of HBV DNA are associated with more rapid disease progression and an increased risk of hepatocellular carcinoma (HCC). HBV DNA testing is also used to monitor the response to antiviral treatment. By tracking changes in HBV DNA levels, healthcare providers can assess the effectiveness of antiviral therapy in controlling the infection.
Hepatitis B infection can be diagnosed based on different clinical scenarios. In the case of acute hepatitis B, diagnosis is made by detecting the presence of HBsAg, along with symptoms and elevated serum aminotransferases. Additionally, anti-HBc IgM antibodies can usually be detected, and HBV DNA is present in the bloodstream. Chronic hepatitis B is diagnosed when HBsAg persists for more than 6 months. This indicates an ongoing infection and the need for long-term management and monitoring.Past hepatitis B infection is identified by the coexistence of anti-HBs (antibodies against HBsAg) and anti-HBc IgG (antibodies against core antigen). This indicates prior exposure to the virus and the development of immunity. Occult HBV infection is a unique scenario characterized by the persistence of a low level of intrahepatic HBV DNA and the presence of isolated anti-HBc antibodies, with the absence of both HBsAg and anti-HBs antibodies in the blood. The gold standard for diagnosing occult HBV infection is detecting HBV DNA in liver tissue samples. However, serum HBV DNA testing using real-time PCR has become the preferred method for diagnosis.
HBV is a double stranded DNA with different serological markers and is one of the major health problem globally leading to liver cirrhosis and hepatocellular carcinoma HCC. There are variable presentation of hepatitis B -acute, chronic and occult cases. Sexual, parenteral, or vertical route are the modes of transmission.Early diagnosis and treatment will help in eradicating the virus. North American, and Western Europe have the lowest prevalence of hepatitis B Infection-around 0.5 -2% while highest prevalence countries are China, and South East Asia-affecting around > 8%.10 genotypes of hepatitis are known- A- J. Serological Markers
Earliest serological marker to appear in serum is HBsAg (1-2 weeks of exposure).Chronic hepatitis B infection is labeled as having HbsAg in serum for more than 6 months. anti-HBs – marker of long term immunity and is a neutralizing antibody .HBeAg and anti-HBe –marker of infectivity and viral replication. Anti-HBc IgM and IgG –IgM appears in blood after 1-2 week of appearance of HbsAg and disappears after 06 months. However, IgG appears in blood life long even after resolved HBV infection and chronic hepatitis B.
Molecular Methods for HBV Infection:
HBV PCR DNA (quantitative)-detected by Signal amplification,or Target amplification-indicates active infection and quantitative test is better than qualitative test
Acute hepatitis B: HBsAg +, HBV DNA+, anti-HBc IgM and HBeAg +/-ve
Chronic infection: HBsAg + for more than 6 months.
Past HBV infection: anti-HBs +, IgG anti-HBc+ HBsAg-ve , and HBV DNA PCR-ve.
Occult HBV infection : HBV DNA +ve ,HBsAg –ve , anti-HBc IgG +ve
CONCLUSION:
This article deals with the diagnostic strategies both serological and molecular which ill help in differentiating between acute, chronic and occult infection.
Diagnosis of hepatitis B
Its review article was published in the Annals of Translational Medicine 2016
Diagnosis of HBV
1- Serological markers for HBV infection (antigen or antibodies screening in the serum).
2- Molecular methods for HBV infection by PCR
3- HBV genotyping
Serological markers for HBV infection
HBsAg: It can be detected in serum within 1 to 10 weeks after infection and usually disappears after 6 months but if it persists for more than 6 months indicates chronic HBV infection
Anti-HBs: It is a neutralizing antibody that indicates long-term immunity. After vaccination, it is the only serological marker detected in serum but after past exposure to HBV infection, it will be associated with anti-HBc IgG.
HBeAg and anti-HBe: it’s related to infectivity and viral replication.
HBcAg: It is an intracellular antigen and not seen in the blood.
anti-HBc IgM and IgG: after acute infection, anti-HBc IgM and IgG were seen 1–2 weeks after HBsAg. anti-HBc IgM starts to wear off after 6 months on the other hand Anti-HBc IgG continues to detect in both patients with resolved HBV infection and chronic hepatitis B (never got by vaccination).
Molecular Methods for HBV Infection
HBV DNA reveals the replication activity of the virus.
It can be detectable during the first-month post-infection then gradually it decreases in chronic infection or completely disappears if recovery occurred.
HBV genotyping
HBV genotyping: can be done by Reverse hybridization, Genotype specific PCR and Sequence analysis. It can be divided into ten genotypes, labeled A–J.
Genotype B and C are only seen in Oceania and Asia, whereas genotype A and D are the most common genotypes in Africa and Europe with higher rate of chronicity but higher response to interferon therapy.
Genotype C is generally associated with perinatal infection and is more aggressive in the form of cirrhosis and HCC.
Diagnosis of hepatitis B infection
Acute hepatitis B: HBsAg positive, HBV DNA is positive, anti-HBc IgM and HBeAg can also be positive.
chronic infection: HBsAg positive and HBV DNA PCR (for more than 6 months)
Past HBV infection: anti-HBs positive, IgG anti-HBc positive HBsAg negative, and HBV DNA PCR negative.
Occult HBV infection: low level of intrahepatic HBV DNA (gold standard but invasive), HBsAg negative, anti-HBc IgG positive
Occult HBV infection has some clinical importance.
1- First, risk of transmission by solid organ transplantation (especially liver).
2- Second, risk of reactivation of HBV infection after starting immunosuppression.
3- Third, risk of LCF and HCC.
Liver tissue (biopsy) PCR is indicated if all the following are present
Summarize the article;;
Introduction;
HBV infection is the most common cause of the liver failure, and a major global health problem in developing countries.
Serologic marker for HBV infection;
How differentiate serologic markers, and different stages of infectivity and activity, like,
HBs Ag, marker for infection, will positive within 8-10 weeks, and persist for 6 months if converts into chronicity, being used as marker for treatment monitoring.
Anti-HBs, shows immunity or previous infection, vaccination. It’s known as neutralizing antibody.
HBeAg, active infection.
Anti-HBe antibody,
Anti-HBc IgM and IgG.
Molecular methods for diagnosis of Hepatitis B infection;
HBV DNA is a direct measurement of the viral load, shows the active infection, detected in early phase of infection.
Two principles 1, Signal amplification,
2, Target amplification.
Standard method for detection is HBV DNA quantification.
Genotyping;
There are ten genotypes.
Labeled as A- J. done to predict the treatment.
Diagnosis of Hepatitis B infection;
Acute hepatitis is a clinical diagnosis, and confirmed by serological testing.
For diagnosis of chronic infection if HBVs-Ag persist for more than 6 months.
Past infection HBV-s Ab, and HBVc- Ab.
HBV DNA is gold standard for detection of infection.
Conclusion;
In this article the aim was to give guidance for detection, diagnosis, to differentiate B/W acute and chronic and when to take action.
Summarize the article;
Introduction;
HBV infection is the most common cause of the liver failure, and a major global health problem in developing countries.
Serological markers for HBV infection;
How differentiate serologic markers, and different stages of infectivity and activity, like,
HBs Ag, marker for infection, will positive within 8-10 weeks, and persist for 6 months if converts into chronicity, being used as marker for treatment monitoring.
Anti-HBs, shows immunity or previous infection, vaccination. It’s known as neutralizing antibody.
HBeAg, active infection.
Anti-HBe antibody,
Anti-HBc IgM and IgG.
Molecular methods for Diagnosis;
HBV DNA is a direct measurement of the viral load, shows the active infection, detected in early phase of infection.
Two principles 1, Signal amplification,
2, Target amplification.
Standard method for detection is HBV DNA quantification.
Genotyping;
There are ten genotypes.
Labeled as A- J. done to predict the treatment.
Diagnosis of hepatitis B infection;
Acute hepatitis is a clinical diagnosis, and confirmed by serological testing.
For diagnosis of chronic infection if HBVs-Ag persist for more than 6 months.
Past infection HBV-s Ab, and HBVc- Ab.
HBV DNA is gold standard for detection of infection.
Conclusions;
In this article the aim was to give guidance for detection, diagnosis, to differentiate B/W acute and chronic and when to take action.
The article discusses the diagnosis of acute, chronic, and occult hepatitis B virus (HBV) infections through serological and molecular methods. Key points from the article include:
Introduction:
Hepatitis B virus (HBV) infection is a major global health problems leading to severe liver disease such as cirrhosis and hepatocellular carcinoma.
Significant method to diminish the burden of this disease is timely diagnosis of acute, chronic and occult cases of HBV.
Serological markers for HBV infection:
Consist of HBsAg, Anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg:
Is the serological hallmark of HBV infection.
Appears in serum within 1 to 10 weeks.
Persistence of this marker for more than 6 months implies chronic HBV infection.
Anti-HBs:
Is known as a neutralizing antibody, and confers long-term immunity.
Vaccination, anti-HBs is the only serological marker detected in serum.
Past HBV infection, it is present in concurrence with anti-HBc IgG.
As carriers of HBV : HBsAg and anti-H HBsAg has been reported in patients.
HBeAg and anti-HBe:
Know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
HBcAg :
Is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
Anti-HBc IgM and IgG:
Emerges in Acute infection, 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Isolated anti-HBc positive: negative HBsAg and HBs ab, seen in three conditions:
First, as IgM class during the window period of acute phase.
Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
Anti-HBc IgM should be done and HBV DNA checked.
Molecular methods for HBV infection:
One month after HBV infection.
A reliable marker of replication activity,
Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
HBV genotyping:
HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution.
The HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C .Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
The interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
Genotype B or C had a lower opportunity to gain serologic response to tenofovir.
Diagnosis of hepatitis B infection:
Acute hepatitis B:
Symptoms, high serum aminotransferases. HBsAg, POSTIVE, anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase.
Chronic infection:
The persistence of HBsAg for more than 6 month.
Past HBV infection:
AntiHBs AB and IgG anti-HBc.
Occult HBV infection:
HBV DNA without detectable HBsAg.
Isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
Occult HBV infection has some clinical importance.
First, it can be transmitted via transfusion, solid organ transplantation including orthotropic liver transplantation, or hemodialysis
. Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
. Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
Forth, it appears to be a risk factor for HCC.
Occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease, especially when having anti-HBc in serum;
Patients considering immunosuppression therapy or chemotherapy;
In solid organ transplantation donors.
Serological markers for HBV infection
Serological markers for HBV infection consist of HBsAg anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection(CHB).The serum HBsAg titers are higher in patients with HBeAgpositive CHB than in HBeAg-negative CHB . Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAgnegative CHB patients with normal serum alanine aminotransferase levels
.Anti-HBs is known as a neutralizing antibody, and confers long-term immunity.In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum. In the past HBV infection, it is present in concurrence with anti-HBc IgG
.
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease . however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB. HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg .
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.As the prevalence of serologically negative HBV infection (HBeAg-negative CHB and occult HBV infection) has increased, HBV-DNA detection has obtained more awareness in clinical medicine The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
.
There are two principles of techniques to identify and quantify HBV DNA: signal amplification such as hybrid capture and branched DNA technology; target amplifcation such as polymerase chain reaction (PCR) (19,22). Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting.
HBV genotyping
According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution .Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy. HBV genotyping can be confrmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fuorescence polarization assay.
Occult HBV infections–
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg . It is a serological situation defned by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody . Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases; thus, HBV DNA testing is widely used to diagnose occult HBV infection.
Occult HBV infection has some clinical importance. It can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis . Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state . Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis,Tests for occult HBV infection are considered in the following conditions: in patients with cryptogenic liver disease, especially when having anti-HBc in serum; in patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission.
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family. It shows a diameter of 30–42 nm and consists of outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein.
Viral capsid bears viral genome and DNA polymerase that has reverse transcriptase activity. HBV genomes comprise a circular, partly double-stranded DNA and has four open reading frames overlapped: (I) S that encodes for surface proteins (HBsAg); (II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg); (III) P for polymerase including reverse transcriptase; (IV) X that encodes for a transcriptional transactivator factor (HBxAg)
Serological markers for HBV infection
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG. The identification of serological markers allows: to identify patients with HBV infection; to elucidate the natural course of chronic hepatitis B (CHB); to assess the clinical phases of infection; and to monitor antiviral therapy.
HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
Several studies have reported the association between transcription activity of cccDNA in the liver and serum HBsAg levels.
Differences in the serum HBsAg levels during the different phases of infection indicate the distribution of cccDNA during the respective phases of the disease. The serum HBsAg titers are higher in patients with HBeAg- positive CHB than in HBeAg-negative CHB.
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg- negative CHB patients with normal serum alanine aminotransferase levels.
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
HBV genotyping
HBV has a high genetic heterogeneity because it reproduces via a reverse transcriptase that has insufficient proofreading.
capability. According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution
Genotype B and C are restricted to Oceania and Asia, whereas genotype A and Dare omnipresent but most common in Africa and Europe
Genotype I is unusual and can be observed in Vietnam, Laos, India and China, while genotype J has been reported in Japan and Ryukyu
Other genotypes such as E, F, G, and H are also occasionally found in Asia.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually, anti-HBc IgM can be detected, and HBV DNA is present. HBeAg can also be identified in most acute phase of infections but has little clinical importance. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
Occult HBV infection has some clinical importance. First, it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis.
Conclusions
In this article, we aimed to give informations about HBV serological and molecular diagnosis. First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus. In order to verify first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used. Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
Summarise this article
Introduction
HBV belongs hepadnaviridae family.
The reverse transcriptase responsible for viral replication is susceptible to errors thus high mutation rate.
HBV is transmitted parenteral, sexual and vertical route.
HBV is divided into three endemicity: high, intermediate and low; with high endemicity in sub saharan Africa and south east Asia.
Serological markers
HBsAg is the serological hallmark of HBV infection.
Anti-HBs is a neutralising antibody, confers longterm immunity.
HBeAg and anti-HBe prior used to indicate replication and infectivity however has been replaced by HBV DNA.
HBcAg is an intracellular antigen thus not detected in serum.
Molecular methods
HBV DNA indicates the replication of the virus.
HBV genotype
There are 10 genotypes A-J with distinct geographical distribution.
Genotype A and D associated with higher rates of chronicity than B and C.
Genotype A and C have better response to interferon than B and D.
Diagnosis
Acute hepatitis B infection is a clinical diagnosis based by the presence of HBsAg detection, raised serum aminotransferase and symptoms.
Chronic hepatitis detected by persistence of HBsAg beyond 6 months.
Past infection identified by Anti-HBs and IgG anti-HBc antibodies.
Occult infection defined by detection of HBV DNA intra-hepatic.
Summarise the Article
Diagnosis of hepatitis B
Introduction
Hepatitis B virus (HBV) infection is a major global health problem leading to severe liver disease, cirrhosis and hepatocellular carcinoma (HCC).
It is transmitted by sexual, parenteral and vertical route, infecting 240 million people worldwide.
Depending on prevalence in general population and transmission rate, geographically categorised in to 3 endemic zones – high endemic areas (>8% prevalence – China, SE Asia, Indonesia, sub-Saharan Africa); intermediate endemic regions (2-7% prevalence – South America, South West Asia, Eastern and Southern Europe); low endemic regions (0.5-2% prevalence – North America and Western Europe).
One significant method to diminish the burden of this disease is timely diagnosis of acute, chronic and occult cases of HBV.
HBV diagnosis involves screening using serological markers detecting antigens and antibodies, which is then verified by molecular tests like DNA-PCR, viral load quantification and genotyping.
The serological and molecular tests for diagnosis of HBV infection are reviewed in this article.
HBV structure: circular, double-stranded DNA virus, has 4 open reading frames of genes overlapped, which code for various protein antigens.
(I) S encodes for surface proteins (HBsAg)
(II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg)
(III) P for polymerase including reverse transcriptase
(IV) X that encodes for a transcriptional trans-activator factor (HBxAg)
Serological markers for HBV infection
Hepatitis B surface antigen (HBsAg) and anti-HBs, anti-HBc IgM and IgG, and hepatitis B e antigen (HBeAg) and anti-HBe – these antigens and antibodies appear in serum of infected individual in a particular time frame and can be used as serological markers.
HBsAg
– hallmark of HBV infection, appears in serum in 1-10 weeks
– Its persistence for more than 6 months implies chronic HBV infection.
HBsAg titres are higher in patients with HBeAg + CHB
– Its levels predict treatment response and progression in HBeAg-negative CHB patients
Anti-HBs
– indicates long-term immunity, and will be the only marker detected in serum post vaccination.
– anti-HBs antibodies are unable to neutralize the circulating viruses (HBV carriers).
HBcAg is intracellular, present in infected hepatocyte, thus not detected in serum.
HBeAg to anti-HBe seroconversion is related to remission of HBV infection. In case of mutations in the pre-core and core region active viral replication is sustained in spite of seroconversion.
During acute infection, anti-HBc (IgM > IgG) rise 1–2 weeks after the appearance of HBsAg, associated with raised serum aminotransferase and clinical symptoms.
IgM anti-HBc disappear after 6 months of acute infection, but IgG Anti-HBc continues to detect in both patients with resolved HBV infection and CHB.
Isolated Anti-HBc antibody positivity can be seen
1) during the window period of acute phase
2) after the end of acute infection
3) after many years of chronic HBV infection, when HBsAg may be undetectable, indicating a previous infection
Molecular methods for HBV Detection –
HBV-DNA detection by PCR
– detectable 1 month after infection, increases to peak level by 3 months quantification of viral load reveals rate of viral replication
– higher titre indicates rapid disease progression, associated with higher incidence of HCC.
– HBV DNA titre can be used as guide for antiviral therapy and monitoring response
HBV genotyping – 10 genotypes labelled A–J, are identified.
Genotype B & C – are restricted to Oceania and Asia.
Genotype A & D – are omnipresent but most common in Africa and Europe.
Genotypes A & D – more liable to chronicity than genotypes B and C.
Genotype C has risk perinatal infection, and related to severe liver disease, including cirrhosis and HCC.
Genotypes A & B have better response to Interferon than genotypes C & D.
Patients with genotype B &C shows lower serological response to tenofovir.
Diagnosis of hepatitis B infection
Acute hepatitis: HBsAg positive + clinical symptoms + elevated AST & ALT
Usually, anti-HBc IgM can be detected and HBV-DNA is present.
Chronic hepatitis-B: persistence of HBsAg for more than 6 months.
Diagnosed by laboratory means but not by clinical presentations.
Past HBV infection: defined by coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection:
– persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
– defined as presence of isolated anti-HBc with absence of HBsAg, anti-HBs antibody.
– HBV-DNA PCR can be used for diagnosis of occult HBV infection.
The importance of occult HBV infection:
– transmitted via transfusion, solid organ transplantation, or haemodialysis
– reactivation of HBV infection may occur in patients receiving chemotherapy, immunocompromised state
– it may accelerate liver injury, lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
– it could be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis
Indications for testing occult HBV infection:
– Patients with cryptogenic liver disease, mainly those with + anti-HBc in serum
– in patients on chemotherapy or immunosuppression therapy
– in solid organ transplantation donors, due to the possibilities for transmission
Introduction:
It is a double strand DNA virus and belong to e Hepadnaviridae family.
Viral capsid bears viral genome and DNA polymerase that has reverse transcriptase activity.
HBV genome comprise a circular, partly double-stranded DNA and has four open reading frames overlapped:
1) S that encodes for surface proteins (HBsAg);
2) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg);
3) P for polymerase including reverse transcriptase;
4) X that encodes for a transcriptional trans activator factor (HBxAg)
It has high, intermediate and low endemicity.
Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection.
Serological markers for HBV infection:
Consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG
The identification of serological markers allows: to identify patients with HBV infection; to elucidate the natural course of chronic hepatitis B (CHB); to assess the clinical phases of infection; and to monitor antiviral therapy.
HBsAg:
It appears in the serum within 1 to 10 weeks and indicates acute infection and persist up to 6 months and indicates chronic infection and its titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
Anti-HBs:
Is known as a neutralizing antibody, and confers long-term immunity and in acquired immunity through vaccination, it is the only serological marker detected in serum.
In the past HBV infection, it is present in concurrence with anti-HBc IgG.
In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, so considered as carriers of HBV.
HBeAg and anti-HBe:
In the past used to know infectivity and viral replication, but nowadays replaced by HBV DNA assay. HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion.
HBcAg:
It is not detected in serum because it intracellular.
Anti-HBc IgM and IgG:
Appears 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms and it wear off after 6 months of acute infection.
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Isolated anti-HBc positive is considered as HBsAg-negative and positive anti-HBc IgG without anti-HBs and for that anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
In chronic liver disease patients HBV DNA should be tested to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection:
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level at 3 months after the exposure to HBV and then gradually decrease in chronic infection or disappears at the recovery from HBV infection.
The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
There are two principles of techniques to identify and quantify HBV DNA:
1) Signal amplification such as hybrid capture and ranched DNA technology.
2) Target amplification such as (PCR). Real-time PCR is standard method to detect and quantify HBV DNA in clinical setting
HBV genotyping:
It has a high genetic heterogeneity and divided into ten genotypes, labelled(A-J) which have distinct geographic distribution.
Acute infection with genotypes A and D leads to higher rate of chronicity
than genotypes B and C. Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
HBV genotyping can be confirmed using diverse methods:
1) Reverse hybridization.
2) Genotype-specific PCR assays.
3) Real-time PCR.
4) Restriction fragment-length polymorphism.
5) Sequence analysis.
6) Microarray (DNAChip).
7) Fluorescence polarization assay.
Diagnosis of hepatitis B infection:
Acute infection:
Identified by:
1) The detection of HBsAg, symptoms, high serum aminotransferases.
2) anti-HBc IgM Anti-HBc IgM can be detected and HBV DNA is present
3) HBeAg can also be identified in most acute phase of infections, but has little
Chronic infection:
Characterized by:
1) Persistence of HBsAg for more than 6 months.
2) Patients diagnosed by laboratory means but not by clinical presentations.
3) Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection: characterized by:
1. Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
2. Presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
3. The detection of HBV DNA in the liver is the gold standard of diagnosis
4. Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases
Occult HBV infection has some clinical importance:
A. Can be transmitted via transfusion, solid organ transplantation.
B. Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
C. Accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
D. Appears to be a risk factor for HCC.
Tests for occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease; in patients considering IS therapy or chemotherapy and in SOT donors.
The aim of this study: to review serological and molecular diagnosis of HBV.
# Introduction:
*Hepatitis B virus (HBV) belongs to the Hepadnaviridae family and it is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
*HBV is a circular, partly double-stranded DNA virus with various serological markers.
*Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection.
# Serological markers for HBV infection:
*Mainly consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
*The identification of serological markers allows:
1) Identify patients with HBV infection.
2) Elucidate the natural course of chronic hepatitis B.
3) Assess the clinical phases of infection.
4) Monitor antiviral therapy.
HBsAg:
*Is the serological hallmark of HBV infection, in acute infection it is appears in serum within 1 to 10 weeks.
*Persistence of HBsAg for more than 6 months indicates chronic infection.
*The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
*Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs:
*Known as a neutralizing antibody, and confers long-term immunity.
* In acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
*In the past HBV infection, it is present in concurrence with anti-HBc IgG.
*In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, so considered as carriers of HBV.
HBeAg and anti-HBe:
*Previously used to know infectivity and viral replication, but recently replaced by HBV DNA assay. *HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion.
HBcAg:
*Is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
Anti-HBc IgM and IgG:
*It is emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
* After 6 months of acute infection, anti-HBc IgM wears off.
* Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
*In HBsAg-negative individuals and positive anti-HBc IgG without anti-HBs, considered isolated anti-HBc positive.
*If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
*HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
# Molecular methods for HBV infection
*HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
*It is detectable at the early stage of infection and increases up to peak level at 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
*The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
*HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
*There are two principles of techniques to identify and quantify HBV DNA:
1) Signal amplification such as hybrid capture and ranched DNA technology.
2) Target amplification such as (PCR). Real-time PCR is standard method to detect and quantify HBV DNA in clinical setting
# HBV genotyping:
*HBV has a high genetic heterogeneity.HBV can be divided into ten genotypes, labelled A–J: they have
distinct geographic distribution
*Acute infection with genotypes A and D leads to higher rate of chronicity
than genotypes B and C. Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
*HBV genotyping can be confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray and fluorescence polarization assay.
# Diagnosis of hepatitis B infection:
Acute infection
*By the detection of HBsAg, symptoms, high serum aminotransferases.
*Anti-HBc IgM can be detected and HBV DNA is present.
* HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
Chronic infection:
*Persistence of HBsAg for more than 6 months.
*Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
* Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection:
*Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
*Presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
*The detection of HBV DNA in the liver is the gold standard of diagnosis
*Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases
*Occult HBV infection has some clinical importance.
1) Can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
2) Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
3) Accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4) Appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic
inflammation and fibrosis.
*Tests for occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease; in patients considering IS therapy or chemotherapy and in SOT donors.
● Viral capsid bears viral genome and DNA polymerase that has reverse transcriptase activity.
● HBV genome comprise a circular, partly double-stranded DNA and has four open reading frames overlapped:
(I) S that encodes for surface proteins (HBsAg)
(II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg)
(III) P for polymerase including reverse transcriptase
(IV) X that encodes for a transcriptional transactivator factor (HBxAg)
● HBV infection is transmitted through parenteral, sexual and vertical route.
● About 240 million people are chronically infected by HBV, so have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC).
● HBV endemicity is divided into three categories;
☆ high
China, South East Asia, Indonesia, and sub-Saharan Africa
☆ intermediate
South America, South West Asia, Eastern and Southern Europe.
☆ low.
North America and Western Europe
Serological markers for HBV infection
☆ HBsAg
☆ anti-HBs
☆ HBeAg
☆ anti-HBe
☆ anti-HBc IgM and IgG.
● HBsAg is the serological hallmark of HBV infection.
● After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks.
● Persistence of this marker for more than 6 months implies chronic HBV infection
● The serum HBsAg titers are higher in patients with HBeAg-positive CHB than in HBeAg-negative CHB
● Anti-HBs is known as a neutralizing antibody, and confers long-term immunity
● Anti-HBs is the only serological marker detected in serum In patients with acquired immunity through vaccination
● The simultaneous appearance of HBsAg
and anti-HBs has been reported in patients with HBsAg positive these patients are regarded as carriers of HBV.
● HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease
● During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
● HBsAg-negative with positive HBc IgG without anti-HBs It can be seen in :
☆ The window period of acute phase
☆ Decreased anti-HBs below cutoff level of detection after acute infection had ended
☆ After several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
● So anti-HBc IgM should be checked to assess the possibility of recent HBV
● Occult HBV infection : detectable HBV DNA without serum HBsAg
● Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
● Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 10^7–10^8 IU/mL).
HBV genotyping
● HBV can be divided into ten genotypes, labelled A–J
● Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
● Genotype C generally is considered as a risk factor for perinatal infection and related cirrhosis and HCC
● Genotypes A and B have better therapy response than genotypes C and D
● Genotype B or C had a lower opportunity to gain serological response to tenofovir
Diagnosis of hepatitis B infection
● Acute hepatitis B is identified:
☆ HBsAg symptoms, high serum ALT .
☆ Anti-HBc IgM can be detected and HBV DNA is present.
☆ HBeAg has little clinical importance.
● Chronic infection is based on :
☆ Persistence of HBsAg for more than 6 M
● Past HBV infection is defined : coexistence of anti-HBs and IgG anti-HBc.
● Occult HBV infection is defined :
☆ persistence of low level of intrahepatic HBV DNA without detectable HBsAg
☆ Presence of isolated anti-HBc with the absence of HBsAg and anti-HBs antibody
● Clinical importance of Occult HBVi :
☆ It can be transmitted via transfusion, SOT including orthotopic liver transplantation or hemodialysis
☆ Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
☆ It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
☆ it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fbrosis
● Tests for occult HBV infection are considered in :
☆ Patients with cryptogenic liver disease, especially when having anti-HBc in serum
☆ patients considering immunosuppression therapy or chemotherapy
☆ solid organ transplantation donors, due to the possibilities for transmission
Diagnosis of hepatitis B
Introduction
· HBV genome comprise a circular, partly double-stranded DNA and has four open reading frames overlapped:
(I) S that encodes for surface proteins (HBsAg);
(II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg);
(III) P for polymerase including reverse transcriptase;
(IV) X that encodes for a transcriptional transactivator factor (HBxAg)
· HBV infection is the cause of most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
· 240 million people are chronically infected by HBV.
· high endemic areas for HBV are: China, South East Asia, Indonesia, and sub-Saharan Africa
· Low endemic regions: North America and Western Europe
Serological markers for HBV infection
· HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
· HBsAg is the hallmark of HBV infection, appears in serum within 1 to 10 weeks, and its persistence for more than 6 months implies chronic HBV infection.
· HBsAg titers are higher in patients with HBeAg + CHB, and its levels predicts treatment response and progression in HBeAg-negative CHB patients
· Anti-HBs indicates long-term immunity, and will be the only marker detected in serum post vaccination.
· In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses (HBV carriers).
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but in case of mutations in the pre-core and core region active viral replication is sustained in spite of seroconversion.
· HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
· During acute infection, anti-HBc IgM and IgG occurs 1–2 weeks after the appearance of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM disappears, but Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
· Isolated anti-HBc positive can be seen in:
1) during the window period of acute phase
2) after the end of acute infection
3) after many years of chronic HBV infection, HBsAg may became undetectable
Molecular methods for HBV infection
· HBV DNA is measures the viral load, which reveals the replication activity of the virus.
· Can be detectable 1 month after HBV infection and increases up to peak level nearly 3 months after the exposure to HBV.
· HBV-DNA detection is a reliable marker of replication activity, and higher titers indicates rapid disease progression and higher incidence of HCC.
· HBV DNA testing is an indication to determine patients who need antiviral therapy and monitor them for suitable treatment.
HBV genotyping
· HBV can be divided into ten genotypes, labelled A–J.
· Genotype B and C are restricted to Oceania and Asia.
· Genotype A and D are omnipresent but most common in Africa and Europe.
· Genotype I is unusual and can be observed in Vietnam, Laos, India and China, while genotype J has been reported in Japan and Ryukyu.
· Other genotypes such as E, F, G, and H are also occasionally found in Asia.
· Acute infection with genotypes A and D leads more liable to chronicity than genotypes B and C.
· Genotype C has a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
· Patients with genotype B or C had a lower opportunity to gain serological response to tenofovir.
Diagnosis of hepatitis B infection
Acute hepatitis B
· Diagnosed by + HBsAg, symptoms, high serum aminotransferases.
· Usually, anti-HBc IgM can be detected and HBV DNA is present.
Chronic hepatitis B
· Diagnosed by the persistence of HBsAg for more than 6 months.
· Diagnosed by laboratory means but not by clinical presentations.
Past HBV infection
· defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection
· defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
· defned by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
· Since hepatic HBV DNA is invasive PCR for serum HBV DNA can be used for diagnosis of occult HBV infection.
The importance of occult HBV infection:
1) it can be transmitted via transfusion, solid organ transplantation, or hemodialysis
2) reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3) it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4) it could be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis
Indications for occult HBV infection testing:
1) patients with cryptogenic liver disease, mainly those with + anti-HBc in serum;
2) in patients’ pre- immunosuppression therapy or chemotherapy
3) in solid organ transplantation donors, due to the possibilities for transmission
# The aim of this study: to review serological and molecular diagnosis of HBV.
# Introduction:
*Hepatitis B virus (HBV) belongs to the Hepadnaviridae family and it is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
*HBV is a circular, partly double-stranded DNA virus with various serological markers.
*Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection.
# Serological markers for HBV infection:
*Mainly consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
*The identification of serological markers allows:
1) Identify patients with HBV infection.
2) Elucidate the natural course of chronic hepatitis B.
3) Assess the clinical phases of infection.
4) Monitor antiviral therapy.
HBsAg:
*Is the serological hallmark of HBV infection, in acute infection it is appears in serum within 1 to 10 weeks.
*Persistence of HBsAg for more than 6 months indicates chronic infection.
*The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
*Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs:
*Known as a neutralizing antibody, and confers long-term immunity.
* In acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
*In the past HBV infection, it is present in concurrence with anti-HBc IgG.
*In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, so considered as carriers of HBV.
HBeAg and anti-HBe:
*Previously used to know infectivity and viral replication, but recently replaced by HBV DNA assay. *HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion.
HBcAg:
*Is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
Anti-HBc IgM and IgG:
*It is emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
* After 6 months of acute infection, anti-HBc IgM wears off.
* Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
*In HBsAg-negative individuals and positive anti-HBc IgG without anti-HBs, considered isolated anti-HBc positive.
*If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
*HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
# Molecular methods for HBV infection
*HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
*It is detectable at the early stage of infection and increases up to peak level at 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
*The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
*HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
*There are two principles of techniques to identify and quantify HBV DNA:
1) Signal amplification such as hybrid capture and ranched DNA technology.
2) Target amplification such as (PCR). Real-time PCR is standard method to detect and quantify HBV DNA in clinical setting
# HBV genotyping:
*HBV has a high genetic heterogeneity.HBV can be divided into ten genotypes, labelled A–J: they have
distinct geographic distribution
*Acute infection with genotypes A and D leads to higher rate of chronicity
than genotypes B and C. Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
*HBV genotyping can be confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray and fluorescence polarization assay.
# Diagnosis of hepatitis B infection:
Acute infection
*By the detection of HBsAg, symptoms, high serum aminotransferases.
*Anti-HBc IgM can be detected and HBV DNA is present.
* HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
Chronic infection:
*Persistence of HBsAg for more than 6 months.
*Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
* Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection:
*Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
*Presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
*The detection of HBV DNA in the liver is the gold standard of diagnosis
*Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases
*Occult HBV infection has some clinical importance.
1) Can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
2) Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
3) Accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4) Appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic
inflammation and fibrosis.
*Tests for occult HBV infection are considered in the following conditions:
Patients with cryptogenic liver disease; in patients considering IS therapy or chemotherapy and in SOT donors.
DIAGNOSIS OF HBV
SEROLOGY
HBV is commonest cause of chronic hepatitis in world
prevalance – high S-E asia, china , africa, intermediate – europe , low – Northamericca and western europe
markers
HBsAg,anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg – main marker
positive after 8-10 week
if persistant after 6 montsh – chronic hepatitis
HBsAg titre can be used in monitoring treatment
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity
post vaccination – only HBs AB is positive
past HBV infection, Anti-HBs is present in concurrence with anti-HBc IgG
HBeAg and anti-HBe is measure or viral replication
HBV DNA is a better measure of replication at present
HBcAg is an intracellular presence in infected
hepatocyte, thus it is not identified in the serum
IT IS NOT A MARKER
anti HBc AB
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and
CHB.
ONLY Anti HBc AB positive and rest is negative is possible in special situations
MOLECULAR DIAGNOSIS
HBV DNA is a direct measurement of the viral load, which
reveals the replication activity of the virus
Real-time PCR can detect wide dynamic range of viral load and it is STANDARD OF CARE
GENOTYPING
HBV genotyping is significant to predict HBV disease progression and
determine appropriate antiviral therapy.
ACTIVE HEPATITIS
HBsAg positive and symptoms are present with abnormal labs
CHRONIC HEPATITIS
HBsAg positive for more than 6 months
PAST INFECTION
coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low
level of intrahepatic HBV DNA without detectable HBsAg
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG. The identification of serological markers allows: to identify patients with HBV infection; to elucidate the natural course of chronic hepatitis B (CHB); to assess the clinical phases of infection; and to monitor antiviral therapy .
HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
Differences in the serum HBsAg levels during the different phases of infection indicate the distribution of cccDNA during the respective phases of the disease.
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity .In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
three conditions. First, it can be predominantly seen as IgM class during the window period of acute phase. Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels. If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure. HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 10 8 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
Furthermore, HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase of infections, but has little clinical importance. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
Occult HBV infection has some clinical importance.
First, it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation ,or hemodialysis .
Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .
Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection .
Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis .
Tests for occult HBV infection are considered in the following conditions: in patients with cryptogenic liver disease, especially when having anti-HBc in serum; in patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission.
Conclusions
In this article, we aimed to give informations about HBV serological and molecular diagnosis. First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus. In order to verify first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used. Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
This is a very useful article. The main concern of this article is knowing the differences between acute, chronic and occult HBV.
The above presentation can be identified from the HBV serological type, HBVs Ag is denoting for infectivity, and it appears. In the blood 1 to 10 weeks after acute infections. If it remains in circulation for over six months, it means chronic hepatitis. In some situations, we may not detect HBVs Ag in the blood in the early post-infective period and in this time, we may find Anti HBVc Ab only.
In diagnosing HBV infection we rely mainly on HBV DNA assay but sometimes in the case of occult HBV we may. need to have a hepatic sample to detect the viruses.
In general, now is no significant implication for HBVcAg to detect the virus.
The presence of Anti HBc and anti-HBS AB ing indicate occult infections without the presence of HBsAg
HBV genotyping is significant in predicting HBV disease progression.
· Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
· Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
Diagnosis of Hepatitis B
Serological markers and the timeline
1.HBs Ag
After exposure to HBV, appears 1 to 10 weeks.Persistent > 6 months indicate chronic infectionHigher in HBe Ag positive vs negativeMonitoring quantitative HBs Ag levels predict treatment response and disease progression2.anti-HBs
Neutralizing antibodyConfer long-term immunityanti-HBs positive (others were negative) –> vaccinationanti-HBs positive + HBc IgG positive (others were negative) –> past infection3.HBe Ag
Active viral replicationanti-HBe4.HBc IgG
Isolated positive in:After the acute infection has endedPast infectionOccult infection (positive HBV DNA)5.HBc IgM
Raised or positive during acute infectionDisappearance after 6 months6.HBV DNA
Replication activityHigher titre related to more rapid replicationHigher incidence of HCC7.HBV genotyping
Diagnosis of Hepatitis B infection
Acute hepatitis
HBsAg positiveanti-HepBc IgMHBV DNA detectedElevated ALT, ASTChronic hepatitis
HBsAg positive > 6 monthsPast HBV infection
anti-HBsanti-HBc IgG positiveOccult HBV infection
HBV DNA detectedHBs Ag not detectedHepBc IgG positive
Summary of the article
Diagnosis of hepatitis B HBV is dsDNA virus ( Hepadnaviridae family).
Transmission of HBV via parenteral route, sexual activity, and sometimes vertical routes.
HBV serological markers:
1. Hepatitis B surface antigen (HBsAg): encodes for surface proteins.
– appears within 1 to 10 weeks after infection.
· Persistence for more than 6 months indicates chronic HBV infection.
· The serum HBsAg titers are higher in patients with HBeAg- positive CHB than in HBeAg-negative CHB. But Quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg- negative CHB patients with normal serum alanine aminotransferase levels.
2. Anti-HBs: · confers long-term immunity.
· Positive HBs-Ab denotes acquired immunity through vaccination.
· Positive HBs-Ab in concurrence with anti-HBc IgG indicates past HBV infection.
· Positive HBs-Ab and positive HBsAg: denotes that anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
3. Anti-HBc IgM and IgG:
· HBcAg indicates infected hepatocyte, thus virus is not identified in the serum.
· anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM weans off.
· Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Isolated anti-HBc positive: when HBsAg and anti-HBs are not detected. This can be seen in 3 conditions:
a) During the window period its seen predominantly as IgM class.
b) After acute infection HBsAg has decreased below the cutoff level of detection.
c) After several years of chronic HBV infection: HBsAg has diminished to undetectable levels.
· If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
4. HBeAg and Anti-HBe:
· HBeAg and anti-HBe had been used in past to know infectivity (presently replaced by HBV DNA assay).
· HBeAg to anti-HBe seroconversion is associated with remission of hepatic disease.
· Active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
Molecular methods for HBV infection
1.HBV DNA is a direct measurement of the viral load which reveals the replication activity of the virus. It is a reliable marker of replication activity. Detects early stage of infection (1 month after HBV infection). Higher titers of HBV DNA are related to the more disease progression and higher incidence of HCC. It is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
2. HBV DNA techniques: there are two principles of techniques to identify and quantify HBV DNA:
a) Signal amplification b) Target amplification.
Real time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107–108 IU/mL). Real-time PCR is the standard method to detect and quantify HBV DNA in clinical setting.
3. HBV genotyping
1.HBV have ten genotypes, labelled A–J: have distinct geographic distribution.
· Genotype B and C: are restricted in Oceania and Asia.
· Genotype A and D: are omnipresent but most common in Africa and Europe.
· Genotype I is unusual and can be observed in Vietnam, Laos, India and China.
· Genotype J has been reported in Japan and Ryukyu.
· Other genotypes such as E, F, G, and H are also occasionally found in Asia.
2. HBV genotyping is significant in predicting HBV disease progression.
· Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
· Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
· Genotype B or C had a lower opportunity to gain serological response to tenofovir.
3. HBV genotyping can be confirmed using:
reverse hybridization,
genotype-specific PCR assays,
real-time PCR,
restriction fragment-length polymorphism,
sequence analysis, microarray (DNA Chip) and fluorescence polarization assay.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, and high serum aminotransferases. Usually, anti-HBc IgM is detected with HBV DNA presence. HBeAg can also be identified in most acute phase of infections but has little clinical importance. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months. Past HBV infection is defined by the coexistence of anti-HBs and or IgG anti-HBc. Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. Serologically there is presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection. Occult HBV infection has clinical importance because it can lead to Transmation in solid organ transplantation.
Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state, it may accelerate liver injury leading to hepatic fibrosis and is risk factor for HCC. Tests for occult HBV infection are considered in the following conditions:
· in patients with cryptogenic liver disease (when having anti-HBc in serum).
· in patients considering immunosuppression therapy or chemotherapy.
· in solid organ transplantation donors, due to the possibilities for transmission.
indications for screening for occult HBV infection: –
Conclusion
– the 1st step of HBV diagnosis is use of serological markers to detect antigens and antibodies against HBV
– qualitative and quantitative molecular tests are used to verify the 1st step of diagnosis by quantifying the viral load and identifying the genotypes
– it is important to establish a diagnosis of HBV infection, this helps determine acute, chronic and occult cases of infection so as to institute preventive remedies and initiate antiviral therapy
DIAGNOSIS OF HEP B.
INTRODUCTION.
-HBV belongs to hepadnaviridae family, approx. 240000000 ppl are chronically infected and are at risk of cirrhosis and HCC
SEROLOGICAL MARKERS FOR HBV INFECTION.
MOLECULAR METHODS FOR HBV INFECTION.
HBV DNA – Directly estimates VL/replication of virus, seen 1/12 post infection, peaks(10*8 copies/ml) ~ 3/12 and declines in chronic states or on recovery. It is identified by signal amplification(hybrid/branched DNA) or Target amplification(Real time PCR)
HBV GENOTYPING – HBV divided into 10 genotypes ;A-J,A & D have more chronicity than B & C. Confirmed via specific PCR assay, Real time PCR, Micro assay and Fluorescence polarization assay.
DIAGNOSIS OF HEP B INFECTION.
>Acute Hep B ; HBsAg, symptoms, High serum aminotransferin, HBV DNA +/- anti HBc IgM and HBeAg.
>Chronic Hep ; Persistent HBsAg >6/12,No symptoms.
>Past HBV infection -Anti HBs and IgG anti HBc.
>Occult HBV infection – low HBV DNA without HBsAG and anti HBs antibody ,+VE anti HBc
>Vaccinated -anti HBs +VE
Importance of Occult HBV infection;
I. Diagnosis of hepatitis B
Summarise this article
Introduction
– HBV is a major global health burden, it causes severe liver disease i.e., liver cirrhosis and hepatocellular carcinoma
– Transmission: sexual, parenteral, vertical route
Serological markers for HBV infection
– HBV has various serological markers: HBsAg, HBsAb, HBcAb IgM and IgG, HBeAg, HBeAb
– these serological markers help to identify patients with HBV infection, assess the clinical phases of the infection and monitor antiviral therapy
– HBsAg is the serological hallmark of HBV infection
– following an acute exposure to HBV, HBsAg appears in serum within 1-10 weeks
– persistence of HBsAg beyond 6 months implies chronic HBV infection
– HBsAg titers are higher in HBeAg positive CHB patients than in HBeAg negative CHB patients
– monitoring quantitative HBsAg levels predicts response to treatment and disease progression in HBeAg negative CHB patients with normal ALT levels
– anti-HBs (HBsAb) is a neutralizing antibody which confers long-term immunity
– a positive HBsAb implies acquired immunity through vaccination
– previously HBeAg and HBeAb (anti-HBe) were used as markers for infectivity and viral replication but this has now been replaced by HBV DNA
– seroconversion from HBeAg to HBeAb indicates remission of hepatic disease
– HBcAg is not identified in serum
– during an acute infection, anti-HBc IgM and IgG appear 1-2 weeks after presence of HBsAg alongside elevated ALT and AST and symptoms
– anti-HBc IgM wears off after 6 months of acute infection
– anti-HBc IgG remains detectable in both patients with resolved HBV infection and CHB infection
– isolated anti-HBc positive: refers to presence of positive anti-HBc IgG, negative HBsAg, negative anti-HBs
– in cases of isolated positive anti-HBc, check anti-HBc IgM so as to assess for the possibility of a recent HBV exposure
– occult HBV infection is characterised by existence of detectable HBV DNA without HBsAg
Molecular methods for HBV infection
– HBV DNA: is a direct measurement of the viral load, it reveals the virus replication activity
– it is detectable in the early stage of infection i.e., one month following HBV infection
– it increases to peak levels (>10⁸ copies/ml) within 3 months post HBV exposure then gradually decreases in chronic infection or disappears at recovery from HBV infection
– detection of HBV DNA: is a reliable marker of replication activity, higher HBV DNA titers are associated with a more rapid disease progression and higher HCC incidence
– HBV DNA testing helps determine which patients require antiviral therapy and monitor them for suitable treatment
HBV genotyping
– HBV reproduces via reverse transcriptase which has insufficient proofreading capability hence has a high genetic heterogeneity
– 10 HBV genotypes, labelled A-J have been described, they have distinct geographic distribution
– HBV genotyping helps predict HBV disease progression and determine appropriate antiviral therapy
– acute infection with genotypes A and D leads to a higher chronicity rate than genotypes B and C
Diagnosis of hepatitis B infection
– acute HBV infection diagnosis is identified by detection of HBsAg, symptoms and high AST and ALT
– usually, anti-HBc IgM can be detected and HBV DNA is present
– HBeAg can be detected in most acute phase infections
– persistence of HBsAg for >6months suggests a diagnosis of chronic HBV infection
– patients with chronic HBV infection are mostly diagnosed via lab tests as opposed to clinical presentation
– coexistence of anti-HBs and IgG anti-HBc is suggestive of past HBV infection
– occult HBV infection is defined by: –
– the gold standard of diagnosis for occult HBV infection is detection of HBV DNA in the liver, but this is an invasive procedure
– HBV DNA PCR has adequate sensitivity to identify occult HBV infection
– clinical relevance of occult HBV infection: –
– indications for screening for occult HBV infection: –
Conclusion
– the 1st step of HBV diagnosis is use of serological markers to detect antigens and antibodies against HBV
– qualitative and quantitative molecular tests are used to verify the 1st step of diagnosis by quantifying the viral load and identifying the genotypes
– it is important to establish a diagnosis of HBV infection, this helps determine acute, chronic and occult cases of infection so as to institute preventive remedies and initiate antiviral therapy
DIAGNOSIS OF HBV
HBV id DNA virus and commonest cause of hepititis
High endemic 8% prevalence CHINA AND S-E ASIA, SUBSAHARIAN AFRICA
intermediate 2-7%
low- Less than 2%
Why molecular marker are important ?
diagnosis\
to know natural history
to monitor anti-viral treatment
to know the clinical phase of disease
What is actually tested ?
HBsAg
anti HBs
anti-HBc
HBe AG
anti HBE – Ig M IN EARLY PERIOD, Ig G in late phase
DNA copies of HBV– more important in seronegative patients
real time PCR is best
Genome – A B C D – specific to geographical area , like C is common in INDIA
can predict disease progression, genome C is linked to HCC and cirrhosis
can determine appropriate anti viral therapy A and B RESPOND BETTER TO INTERFERON
How to interpret?
ACUTE INFECTION
HBsAg present
raised enzymes
symptoms
HBV DNA present
anti HBC may be present
CHRONIC INFECTION
more than 6 months
only lab test positive
no symptoms
anti HBs and Ig G anti HBc present
OCCULT INFECTION
anti HBc present
HBs Ag and anti HBs both are absent
intrahepatic HBV DNA present
VACCINATED
anti HBs present
more than 10 IU/ML is good titre
what occult infection can cause ?
it can be transmitted through organ donation
increases risk of fibrosis with HCV
reactivation is possible with IS
risk factor for HCC
Summarise this article
Introduction
-Hepatitis B virus (HBV) belongs to the Hepadnaviridae
family.HBV genome comprise a circular, partly double-stranded DNA.
-HBV infection is transmitted through parenteral, sexual and vertical route and they have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC) .
-Along with active anti-HBV vaccination, one significant method to diminish the burden of this disease is the diagnosis of acute, chronic and occult HBV infection.
Serological markers for HBV infection
-They consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
-HBsAg is the serological hallmark of HBV infection.Persistence of this marker for more than 6 months implies chronic HBV infection .
Monitoring of quantitative HBsAg levels predicts treatment
response to interferon and disease progression in HBe Ag negative
CHB patients with normal serum alanine aminotransferase levels .
-Anti-HBs is known as a neutralizing antibody, and confers long-term immunity . In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
– In the past HBV infection, it is present in concurrence with anti-HBc IgG.
-In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
– HBe Ag to anti-HBe seroconversion is related to the remission of hepatic disease , however, active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg .
-HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
-During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks
after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
– Some HBsAg-negative individuals are positive for
anti-HBc IgG without anti-HBs in three conditions: First, it can be predominantly seen as IgM class during the window period of acute phase. Second, after acute infection had ended, anti-HBs has decreased
below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
-HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg .
Molecular methods for HBV infection
– The detection of HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC .
– It is detectable at 1 month after HBV infection and increases up to peak level approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
-HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor patients.
HBV genotyping
-HBV can be divided into ten genotypes, labelled A–J.
-Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C and genotype C generally is
considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC .
-In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
Diagnosis of hepatitis B infection
-Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases.
-Anti-HBc IgM can be detected and HBV DNA is present.
-The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
-Patients with chronic HBV infection are commonly diagnosed by
laboratory means but not by clinical presentations.
-Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
-Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody .
– The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection .
– Occult HBV infection has some clinical importance : First, it can be transmitted via transfusion, solid organ transplantation or hemodialysis . Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .Third, it accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic HCV infection . Forth, it a risk factor for HCC by its carcinogenic effect .
Diagnosis of HBV
Review article
Introduction
Hepatitis B viral infection consider one of the public health problems worldwide as it can lead to progressive liver disease including liver cirrhosis and hepatocellular carcinoma if not treated
HBV is a circular DNA virus with a wide range of serological assays that can determine the acute and chronic clinical course of the disease, usually transmitted through contaminated blood and blood products, sexual contact, and vertical transmission
HBsAg positive referred to acute infection usually appear between 1-10 weeks, if persists more than 6 months indicate chronic HBV infection while AntiHBs AB is called neutralizing AB and indicates a long immune response, and alone indicate acquired immunity from vaccination.HBsAg negative with antiHBc indicates previous exposure
historical, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use has generally been replaced by HBV DNA
isolated anti-HBc positive. It can be seen in 3 conditions
1. acute HBV infection window with antiHBc IgM
2. Past HBV infection with antiHBc IgG
3. occult HBV infection HBV DNA assays should be tested in chronic
liver disease patients to find out occult HBV infection categorized by the presence of detectable HBV DNA without
serum HBsAg.
Different molecular and genotypes assay is available for the diagnosis of HBV infectious pattern
HBV DNA is one of the direct molecular testings for the viral load by real-time -PCR and reflects the viral replication activity and can be positive as early as the first month of acute HBV infection with the peak level in 3 months after viral infection followed by subsequent reduction, its important for the diagnosis as well as monitoring the response e for antiviral therapy
HBV Genotyping
HBV can be alienated into ten genotypes, labeled A–J: they have distinct geographic distributions (24). Genotypes B and C are
restricted to Oceania and Asia, whereas genotype A and D are omnipresent but most common in Africa and Europe. HBV genotyping
is significant to predict HBV disease progression and determine appropriate antiviral therapy
Acute infection with genotypes A and D leads to a higher rate of chronicity than with genotypes B and C
Genotype c and B have more vertical transmission and are associated with fast progression to liver cirrhosis and HCC with less response to interferon-gamma compared to genotypes A&D
Occult HBV infection is defined by the persistence of a low level of intrahepatic HBV DNA without detectable HBsAg only isolated anti-HBc with the absence of HBsAg and anti-HBs antibody
And the detection of HBV DNA is the gold standard for the diagnosis of OBI and is widely used in clinical practice
Conclusion
This narrative review gives us a summary of HBV epidemiology and its serological testing along with the different molecular and genotyping assays that help in the early diagnosis and determination of the different clinical aspects of HBV infection
Summary
This is the review article on diagnosis of hepatitis b virus. Hepatitis B virus (HBV) belongs to the Hepadnaviridae family with a diameter of 30–42 nm and consists of outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein.
HBV is transmitted through parenteral, sexual and vertical route. China, South East Asia, Indonesia, and sub-Saharan Africa are regarded as highly endemic areas.
Serological markers for HBV infection
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG
After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
In the past HBV infection, it is present in concurrence with anti-HBc IgG.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms
After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection
HBV genotyping
HBV genotyping can be confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNA Chip) and fluorescence polarization assay.
According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution
Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually, anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase of infections
The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg . It is a serological situation isolated anti-HBc +ve with the absence of HBsAg and antiHBs antibody
Clinical significance of Occult HBV infection
Can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis.
Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
It appears to be a risk factor for HCC by its carcinogenic effect and leading to continuous hepatic inflammation and fibrosis
Tests for occult HBV infection are considered in
Cryptogenic liver disease, especially when having anti-HBc in serum;
Immunosuppression therapy or chemotherapy patients; and
Solid organ transplantation donors
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
Serological markers for HBV infection
HBsAg is the serological hallmark of HBV infection. appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBe Ag negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs is known as a neutralizing antibody, and confers long-term immunity.
In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive. It can be seen in three conditions. First, it can be predominantly seen as IgM class during the window period of acute phase. Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection. Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus. There are two principles of techniques to identify and quantify HBV DNA: signal amplification such as hybrid capture and branched DNA technology; target amplification such as polymerase chain reaction (PCR)
HBV genotyping
HBV has a high genetic heterogeneity because it reproduces via a reverse transcriptase that has insufficient proofreading capability. According to the sequence divergence, HBV
can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution.
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases
chronic infection is based on the persistence of HBsAg for more than 6 months.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAgThe detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in
the serum.
Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
Introduction Hepatitis B (HBV) belongs to the hepadnaviridae family. It consists of an outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein. The viral capsid contains the viral genome and DNA polymerase. The HBV genome consists of a circular partly double stranded DNA. The covalently closed circular DNA (cccDNA) is the transcriptional template of HBV and stays inside the hepatocyte nucleus. HBV infection is responsible for many chronic liver diseases, and can lead to the development of liver cirrhosis and hepatocellular carcinoma (HCC). It can be transmitted via the parenteral, sexual and vertical routes. The endemicity can be classified into three categories: high (infection is reported as more than 8% of the population), intermediate (Infection rate between 2-7%) and low (Infection rate less than 2%). China, South East Asia, Indonesia and sub-Saharan Africa are regarded as high endemic areas. Anti-HBV vaccinations and diagnosis of the infection are significant methods that aid in reducing the burden of this disease. This article reviewed the serological and molecular diagnosis of HBV.
Serological markers for HBV infection The serological markers of HBV infection include HBsAg, anti-HBs, HBeAg, and anti-HBc IgM and IgG.
HBsAg is the serological hallmark of HBV infection. It appears in the serum within 1 to 10 weeks after exposure to HBV.
If it persists more than 6 months, it implies a chronic HBV infection. The differences in the serum HBsAg levels during the different phases of the infection indicate the distribution of cccDNA during the respective phases of the disease.
HBV DNA assay should be tested in chronic liver disease patients to find out occult HBV infection characterized by the existence of detectable HBV DNA without serum HBsAg. Anti-HBs is a neutralizing antibody and confers long-term immunity. If patients have acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
Molecular methods for Detecting HBV infection HBV DNA is a direct measurement of the viral load. It is detectable at the early stages of infection and increases to peak level approximately 3 months after the exposure to HBV. It then gradually decreases in chronic infection or disappears at the recovery from HBV infection. The viral load reveals the replication activity for the virus, and high titers of HBV DNA are related to the more rapid disease progression and a higher incidence of HCC.
HBV genotyping HBV has a high genetic heterogeneity as it reproduces via reverse transcriptase that does not have sufficient proofreading capability. HBV can be divided into 10 genotypes, labelled A to J, and they have distinct geographic distribution. Acute infections with genotype A and D lead to higher rates of chronicity than genotype B and C. genotype C is considered a risk factor for perinatal infection and is related to sever liver disease, including liver cirrhosis and HCC.
Patients with genotype A and B have been shown to respond better to interferon therapy, compared to genotypes C and D. HBV genotyping can be confirmed using reverse hybridization, genotype- specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fluorescence polarization assay.
Diagnosis of hepatitis B infection Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferase levels. The diagnosis of chronic HBV infection is based on the persistence of HBsAg for more than 6 months. Occult HBV infection is defined by the persistence of low level of intrahepatic HBV DNA without detectable HBsAg. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, unfortunately the procedure for obtaining samples is invasive. Occult HBV infection can be transmitted via transfusion, transplantation or hemodialysis.
The reactivation of the infection may occur in patients receiving chemotherapy, or patients in an immunocompromised state. Occult HBV infection may also lead to hepatic fibrosis in patients with chronic liver disease, including chronic hepatitis C infection. It also appears to be a risk factor for HCC.
Tests for occult HBV infection should be considered in patients with cryptogenic liver disease, in patients considering immunosuppression therapy or chemotherapy and in solid organ transplantation donors.
Conclusion The aim of the article was to provide information regarding the serological and molecular testing of HBV infection. Diagnosis is an important tool in order to establish preventative remedies and to initiate treatment.
Introduction
· HBV has an outer lipid envelope containing HBsAg and an icosahedral capsid with viral genome and DNA polymerase.
· HBV genome has four open reading frames, encoding surface proteins, polymerases, reverse transcriptases, and transcriptional transactivator factors.
· Covalently closed circular DNA (cccDNA) is the transcriptional template of HBV, making it error-prone and high in mutation rate.
· HBV infection is responsible for most chronic liver diseases worldwide and is transmitted through parenteral, sexual, and vertical routes.
· Diagnosis of acute, chronic, and occult HBV is essential for reducing its burden.
Serological markers for HBV infection
· Serological markers for HBV infection include HBsAg, anti-HBs, HBeAg, and anti-HBc IgM and IgG, which are used to identify patients with HBV infection, elucidate the natural course of chronic hepatitis B (CHB), assess clinical phases, and monitor antiviral therapy.
· Anti-HBs is a neutralizing antibody that confers long-term immunity in patients with acquired immunity.
· HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use has mostly been replaced by HBV DNA assay.
· HBeAg is related to the remission of hepatic disease, but active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
· During acute infection, anti-HBc IgM and IgG emerge 1-2 weeks after the presence of HBsAg and wear off after 6 months.
· After chronic HBV infection, HBsAg has diminished to undetectable levels.
· HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by the existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
· HBV DNA is a reliable marker of replication activity and is useful in routine clinical settings to determine patients who need antiviral therapy and monitor them for suitable treatment.
· Real-time PCR is the standard method to detect and quantify HBV DNA in clinical settings, as it is fully automated and does not generate carry-over contamination.
HBV genotyping
· HBV has a high genetic heterogeneity, divided into ten genotypes with distinct geographic distributions.
· Acute infection with genotypes A and D leads to a higher rate of chronicity than genotypes B and C, while genotype C is a risk factor for perinatal infection and severe liver disease.
· Recent studies have shown that patients infected with genotypes B or C have a lower opportunity to gain serological response to tenofovir.
Diagnosis of hepatitis B infection
· Acute hepatitis B is identified by HBsAg, symptoms, and high serum aminotransferases, while chronic infection is based on the persistence of HBsAg for more than 6 months.
· HBV DNA testing is widely used to diagnose occult HBV infection, as (cccDNA) remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
· Occult HBV infection can be transmitted via transfusion, solid organ transplantation, or hemodialysis, and can accelerate liver injury and lead to hepatic fibrosis.
· Tests for occult HBV infection are recommended in cryptogenic liver disease, immunosuppression therapy, chemotherapy, and solid organ donation donors.
Conclusion
· First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus
· Diagnosis of HBV is important for establishing preventive remedies and initiating antiviral treatment.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your well-structured reply that is a very clinically oriented summary.
Thanks a lot Professor.
This article talks about the HBV virus, its prevalence, and how to diagnose it.
Intruduction :
HBV is a double-strand DNA virus responsible for most chronic liver diseases worldwide.
Endemic areas are :
China, South East Asia, Indonesia, and sub-Saharan Africa
Diagnosis :
Serological markers for HBV infection:
1-HBS ag:appears in the serum
within 1 to 10 weeks.
Persistence of this marker for more
than 6 months implies chronic HBV infection
.
2-Anti-HBs is known as a neutralizing antibody, and
confers long-term immunity.
3-HBeAg and anti-HBe had been used to
know infectivity and viral replication, but their use for
this purpose has mostly been replaced by HBV DNA
assay.
4-HBcAg is an intracellular presence in infected
hepatocyte, thus it is not identified in the serum. During
acute infection, anti-HBc IgM and IgG emerges 1–2 weeks
after the presence of HBsAg along with raised serum
aminotransferase and symptoms.
5-HBV DNA : is a direct measurement of the viral load.
reveals the replication activity of the virus.
Higher titers of HBV DNA are related to the more rapid
disease progression and higher incidence of HCC.
There are tow type to detect :
signal amplification such as hybrid
capture and branched DNA technology; target amplifcation
such as polymerase chain reaction (PCR).
HBV genotyping:
the HBV genotyping
is significant to predict HBV disease progression and
determine appropriate antiviral therapy.
There are multigenotyping form A to F
Occult HBV infection is defined by persistence of low
level of intrahepatic HBV DNA without detectable HBsAg.
I make note of your superb summary of interpretation of serological reports pertaining to HBV infections.
☆Diagnosis of hepatitis B
In this article, the serological and molecular diagnosis of HBV have been reviewed.
◇ Introduction
▪︎Hepatitis B virus (HBV) is a DNA virus. It can cause an infection with a major global health problems leading to severe liver disease.
◇ Serological markers for HBV infection
▪︎HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
▪︎The recognition of serological markers helps to:
1. Identify patients with HBV infection.
2. Elucidate the natural course of chronic hepatitis B (CHB).
3. Assess the clinical phases of infection.
4. Monitor antiviral therapy.
HBsAg:
▪︎Appears in serum within 1 to 10 weeks of infection.
▪︎If persist > 6 months → chronic infection.
▪︎The serum titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
▪︎Monitoring of it’s levels predicts treatment response to INF and disease progression.
Anti-HBs:
▪︎A neutralizing antibody that confers long-term immunity.
▪︎The only serological marker detected in serum after vaccination.
▪︎In past HBV infection, it is present in concurrence with anti-HBc IgG.
▪︎In most cases patients with anti-HBs antibodies are regarded as carriers of HBV.
HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease
▪︎Active viral replication is sustained in some patients with HBe seroconversion.
HBcAg: is not identified in the serum.
Anti-HBc IgM and IgG:
▪︎Emerges 1–2 weeks after the presence of HBsAg.
▪︎After 6 months of acute infection, anti-HBc IgM wears off.
▪︎Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
▪︎Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs.
▪︎If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
HBV DNA assays
▪︎Should be tested in chronic liver disease patients to find out occult HBV infection, which is characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
▪︎HBV DNA is a direct measurement of the viral load.
▪︎Detection of HBV DNA is a reliable marker of replication activity, and higher titers are related to the more rapid disease progression and higher incidence of HCC.
▪︎It is useful in routine clinical setting.
Principles of techniques to identify and quantify HBV DNA:
1. Signal amplification.
2. Target amplification (Real-time PCR the standard method to detect
and quantify HBV DNA in clinical setting).
HBV genotyping
▪︎HBV has a high genetic heterogeneity (ten genotype labelled A–J with distinct geographic distribution).
▪︎Can predict HBV disease progression and determine therapy.
▪︎Can be confrmed using diverse methods.
Diagnosis of hepatitis B infection
Acute infection:
▪︎Is a clinical diagnosis based on:
1. Detection of HBsAg
2. Symptoms
3. High serum aminotransferases.
▪︎Usually anti-HBc IgM and HBV DNA and HBeAg can be detected.
Chronic infection:
▪︎Is based on the persistence of HBsAg for more than 6 months.
▪︎ A laboratory based diagnosis.
▪︎Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult infection:
▪︎Persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
▪︎A serological situation, defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
▪︎The detection of HBV DNA in the liver is the gold standard of diagnosis.
▪︎Tests for occult HBV infection are considered in:
1. Patients with cryptogenic liver disease
2. Patients considering immunosuppression therapy or
chemotherapy.
3. Solid organ transplantation donors.
Conclusions
▪︎The first step for diagnosis of HBV infection is the serological markers for detecting Ags and Abs against HBV.
▪︎Diagnosis of HBV infection is an important tool to identify acute, chronic and occult cases of infection.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections
I. Diagnosis of hepatitis B
Summarise this article
HBV serological markers:
HBsAg & anti-HBs
Anti-HBc IgM & IgG
HBeAg & anti-HBe.
Serological diagnosis of HBV
HBsAg:
It is the serological indicator of HBV infection.
It appears in serum 1 to 10 weeks after an acute exposure to HBV.
Its persistence for > 6 months indicates chronic HBV infection.
Serum levels are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB.
Anti-HBs:
Is a neutralizing antibody.
Confers long-lasting immunity.
The sole serological marker found in patients who have acquired immunity by vaccination.
Present together with anti-HBc IgG in the past HBV infection.
It fails to neutralize the circulating viruses in most cases_ HBV carrier status.
HBeAg & anti-HBe:
Their use to indicate activity has been replaced by HBV DNA assay.
HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
Pre-core & core region mutations can cause sustained viral replication in some patients with HBe seroconversion.
HBcAg:
Not identified in the serum (intracellular in infected hepatocyte).
Anti-HBc IgM & IgG appears 1–2 weeks, together with raised aminotransferase & symptoms, after the presence of HBsAg during acute infection.
Anti-HBc IgM disappears 6 months after acute infection. Anti-HBc IgG persists in resolved HBV infection & CHB.
If isolated anti-HBc positive is detected, anti-HBc IgM should be done to assess a recent HBV exposure.
HBV DNA assays should be tested in CLD patients to detect occult HBV infection (detectable HBV DNA without serum HBsAg).
Molecular diagnosis of HBV
HBV DNA:
Directly measures the viral load.
A marker of replicative activity.
Detected early (1 month) after HBV infection & increases to peak level 3 months after the exposure & then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
Higher titers indicate:
More rapid disease progression
Higher incidence of HCC
The need for antiviral therapy
Techniques to identify & quantify HBV DNA:
Signal amplification (hybrid capture & branched DNA technology)
Target amplification (PCR):
The standard method
Detects wide range of viral load (10–15 IU/mL to 107–108 IU/mL).
HBV genotyping
HBV can be divided into ten genotypes (A–J).
Genotype B & C are restricted to Oceania & Asia.
A & D are most common in Africa and Europe.
A & D linked to higher rate of chronicity than B & C.
C is a risk factor for perinatal infection & linked to severe liver disease (cirrhosis & HCC).
A & B responds better to interferon than C & D.
B & C had lower serological response to tenofovir.
Methods to confirm HBV genotyping:
Reverse hybridization
Genotype-specific PCR assays
Real time PCR
Restriction fragment-length polymorphism
Sequence analysis
Microarray (DNAChip)
Fluorescence polarization assay.
Diagnosis of hepatitis B infection
Acute hepatitis B:
Detection of HBsAg,
Symptomatology
High serum aminotransferases.
Anti-HBc IgM & HBV DNA can be present.
HBeAg in acute phase of infections has little clinical.
Significance.
Chronic infection:
Persistence of HBsAg for > 6 months.
Coexistence of anti-HBs & IgG anti-HBc defines past HBV infection.
Occult HBV infection:
Persistence of low level intra-hepatic HBV DNA without detectable HBsAg.
Isolated anti-HBc in the absence of HBsAg & anti-HBs. HBV DNA in the liver is the gold standard of diagnosis; however, it is invasive & not pravtical.
Real-time PCR for serum HBV DNA can identify occult HBV infection in many cases.
Significance of occult HBV infection:
Transmitted via transfusion, SOT, or HD.
Reactivation after chemotherapy or immunocompromised state.
May accelerate liver injury & lead to hepatic fibrosis in CLD.
A risk factor for HCC by its carcinogenic effect.
I appreciate that you mention genotypes, A–J, with varied geographic distributions and differences of outcomes. I like your summary.
Diagnosis of hepatitis B
Hepatitis B virus (HBV) carry risk of chronic hepatitis and hepatocellular carcinoma. It’s transmitted by parenteral rote and sexual activity and vertical rote. This article focus on serological and molecular diagnosis of HBV.
Serological markers for HBV infection:
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg is appear after acute exposure to HBV within 1-10 weeks and if persist more than 6 months is indicated chronic exposure to HBV.
Quantity of HBsAg is a marker for treatment response to interferon and disease outcome.
Presence of anti-HBs antibodies indicates patients are carrier for HBV.
HBeAg and anti-HBe indicated infection stage but virama know is detected by HBV DNA and HBeAg to anti-HBe seroconversion is used as a marker for remission of hepatic disease.
HBcAg is an intracellular presence in infected hepatocyte and not found in the serum. During acute infection, anti-HBc IgM and IgG appear within 1–2 weeks after the presence of HBsAg and associated with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM disappeared and Anti-HBc IgG indicates chronicity and continues to detect in both patients with resolved HBV infection and CHB.
Molecular methods for HBV infection:
HBV DNA test used for viral load and viral replication. It is detectable at the early stage of infection approximately one month after HBV infection and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
Presence of high titer HBV DNA is associated with viral replication and activity of virus and more aggressive disease and high risk HCC and it’s a time for antiviral therapy for eradication of virus.
There are 2 technical procedures to detect HBV DNA:
HBV genotyping:
It’s reproduced reverse transcription
It’s 10 subtypes from (A to J).
HBV genotyping is confirmed by diverse methods:
reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fluorescence polarization assay.
Diagnosis of hepatitis B infection:
Acute hepatitis B:
chronic infection:
Clinical Importance of Occult HBV infection:
Tests for occult HBV infection are considered in the following conditions:
Hepatitis B virus HBV;
Is continuing to be a significant infection with escalated risk of morbidity and mortality. Its infection is associated with higher risk of Hepato-cellular carcinoma HCC.
Its way of transmission is essentially sexual, vertical and parentral .Its a acircular virus with double stranded DNA genome and containing Surface antigen, core antigen and e antigen. Its the most common cause of chronic liver disease infecting more than 240 millions over all the world. Incidence of HBV infection ranged from 2-7% of population. HBV vaccine represent a glorious bounce in defeating the spread of virus and its consequent dismal prognosis of HCC.
Meticulous survey for all varieties of HBV infection including:
Acute HBV infection which is the presence of HBV surface antigen for 6 month.
Chronic HBV is the persistence of of HBV surface antigen for more than 6 months.
Occult HBV infection which feature positive HBV DNA in the setting of absent HB surface antigen.
Serological markers of HBV infection:
HBV surface antigen HBsAg and HBV surface antibodies HBsAb.
HBeAg and HBeAb
HBcAb:
Its reflecting infection with HBV, its usually detected within 1- 2 weeks post exposure as IgM antibodies. which will vanished and replaced with IgG HBcAb to continue to be detected with chronic HBV infection or recovered infection.
HBsAg reflect infection with HBV and when pesists for more than 6 months it indicate Chronic HBV infection. HBsAb are neutralizing antibodies that bolster a long term immunity.
In the specific circumstances of detecting both HBsAg and HBsAb its indicative of a Carrier state .
I like your summary. I make note of your interpretation of serological testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections.
The following tests can be used for the diagnosis and evaluation of HBV infection:
HBs Ag
The presence of HBs Ag indicates infection. Occur 1 to 10 weeks following an infection. If infection persists for more than six months, it is considered chronic. HBs Ag levels using PCR can be used to predict treatment response.
Anti-Hbs Antibodies
It is known as a neutralizing antibody, and it provides immunity that lasts for a long time.
Anti-HBs is the only serological marker that can be found in the serum of patients who have acquired immunity as a result of vaccination.
If It is present at the same time with anti-HBc IgG, indicates a previous infection with HBV.
HBeAg and anti-HBe Abs
HBeAg and anti-HBe were formerly employed to determine infectivity, however currently viral tests for HBV are more common. The transformation of HBeAg into anti-HBe is diagnostic of remission.
Hbc Ag
Due to the fact that it is already present in infected hepatocytes, HBcAg cannot be found in serum. In acute infections, anti-HBc IgM and IgG antibodies begin to show up one to two weeks after the appearance of HBsAg alongside elevated liver enzymes. Anti-HBc IgM antibodies are no longer detectable in patients with cured HBV infection or CHB six months after the onset of acute infection, although anti-HBc IgG antibodies continue to be found in these patients.
Molecular methods for HBV infection
HBV DNA can be identified one month after HBV infection, reaches its maximal level three months after exposure to HBV, and then gradually lowers in chronic infection or vanishes upon recovery. HBV DNA can be detected one month after HBV infection. In addition to indicating a quick course of the disease and a higher incidence of HCC, the presence of HBV DNA can help determine which patients require antiviral therapy and track how well they are responding to treatment.
Both the presence of HBV DNA and its quantity can be determined by the use of signal amplification techniques, such as hybrid capture and branched DNA technologies, and target amplification techniques, such as PCR. In the meanwhile, the HBV PCR test is the gold standard.
Hepatitis B genotyping
The ten genotypes of HBV, denoted by the letters A through J, each correspond to a different region of the world.
The progression of HBV disease can be predicted by HBV genotyping, which can also point to the most effective antiviral treatment. An acute infection with the genotypes A and D has a higher rate of chronicity than an acute infection with the genotypes B and C, whereas the genotype C is a risk factor for perinatal infection and severe liver disease. A and B have a better response to the therapy with interferon than C and D.
Tenofovir elicits a weaker response from strains B and C. Reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip), and fluorescence polarization assay are some of the methods that can be used to detect HBV genotyping. Other methods include restriction fragment-length polymorphism, real-time PCR, and restriction fragment-length polymorphism.
I like your detailed well-structured summary well supported by references.
I note that you have explained how natural history of disease is affected by genotype of HBV.
SEROLOGICAL MARKERS FOR HBV INECTION ;
———————————————————————————–
1-HBsAg;
is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection.
2-Anti-HBs;
is known as a neutralizing antibody, and confers long-term immunity.
3-HBeAg and anti-HBe ;
had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
4-HBcAg;
is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
SOME HBsAg-negative INDIVIDUALS are positive for anti-HBc IgG without anti-HBs;
——————————————————————————————
In this situation, it should be considered isolated anti-HBc positive. It can be seen inthree conditions;
A- It can be predominantly seen as IgM class during the window period of acute phase.
B- After acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
C- After several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure. HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
MPLECULAR METHOD for HBV INFECTION ;
———————————————————————————————
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
There are two principles of techniques to identify and quantify HBV DNA:
1-Signal amplification such as hybrid capture and branched DNA technology; target amplification such as polymerase chain reaction (PCR) .
2- Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 10 7 –10 8 IU/mL).
HBV GENOTYPING ;
————————————————-
HBV can be divided into ten genotypes, labelled A–J.
Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C .
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D .
Recent studies reported that patients infected with genotype B or C had a lower opportunity to gain serologicaL response to tenofovir.
DIAGNOSIS of HEPATITIS B INFECTION ;
——————————————————————-
1-Acute hepatitis B;
is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases. Usually anti-HBc IgM can be detected and HBV DNA is present. HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
2-The diagnosis of chronic infection ;
is based on the persistence of HBsAg for more than 6 months. Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations. Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
3-Occult HBV infection;
It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody .
The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
However, gaining hepatic HBV DNA is difficult in clinical setting since the procedure is invasive. Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases; thus, HBV DNA testing is widely used to diagnose occult HBV infection .
OCCULT HBV INFECTION has some CLINICAL IMPORTANCE ;
—————————————————————–
1-It can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis .
2-Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .
3- It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection .
4- It appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis .
TESTS for OCCULT HBV INFECTION are CONSIDERED in the FOLLOWING CONDITIONS:
——————————————————————————————–
1-In patients with cryptogenic liver disease, especially when having anti-HBc in serum .
2-In patients considering immunosuppression therapy or chemotherapy.
3- In solid organ transplantation donors, due to the possibilities for transmission .
.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your well-structured reply that is a very clinically oriented summary.
However, there are few spelling errors such as MPLECULAR
Serological markers for HBV infection
HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
1-HBsAg
· HBsAg is the serological hallmark of HBV infection.
· HBsAg appears in serum within 1 to 10 weeks after acute exposure
· Persistence for more than 6 months indicate chronic HBV infection .
· occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg
2- Anti-HBs
· known as a neutralizing antibody ,long-term immunity
· In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
· In the past HBV infection, it is present in concurrence with anti-HBc IgG.
3-HBeAg and anti-Hbe
· HBeAg and anti-HBe had been used to know infectivity and viral replication,
· but their use for this purpose has mostly been replaced by HBV DNA assay.
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease
4-HbcAg
is an intracellular presence in infected hepatocyte, it is not identified in the serum.
5- anti-HBc IgM and IgG
· emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
· some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, (isolated anti-HBc positive)
It can be seen in
-IgM class during the window period of acute phase.
-after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
-after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
–If isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
Molecular methods for HBV infection
· HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
· It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection
· higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
· Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107–108 IU/mL).
HBV genotyping
HBV has a high genetic heterogeneity because it reproduces via a reverse transcriptase that has insufficient proofreading capability.
According to the sequence divergence, HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution
Diagnosis of hepatitis B infection
Acute hepatitis B is a clinical diagnosis identified by
· the detection of HBsAg,
· symptoms,
· high serum aminotransferases.
· Usually anti-HBc IgM can be detected and HBV DNA is present.
· HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
The diagnosis of chronic infection is based on
· the persistence of HBsAg for more than 6 months.
· Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by
· persistence of low level of intrahepatic HBV DNA without detectable HbsAg
· It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody
· The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection,
· Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases;
clinical importance of Occult HBV infection
1- it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis
2- reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3- it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4- it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis
Tests for occult HBV infection are considered in the following conditions
1. in patients with cryptogenic liver disease, especially when having anti-HBc in serum;
2. in patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission
Conclusions
· First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus. thenqualitative or quantitative molecular tests are used.
· Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive measures and to initiate antiviral treatment.
I make note of your interpretation of serological and genotyping testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your well-structured reply that is a very clinically oriented summary.
Typing whole sentence in bold amounts to shouting
Diagnosis of hepatitis B
Summary
· HBV is a major health problem and important cause of chronic liver disease.
· Mode of transmission: parenteral, sexual, vertical and liver transplantation.
· Reactivation of HBV infection may occur in immunocompromised patients as in SOT.
· Early diagnosis is essential for management.
· Diagnosis has 2 steps;
o Serology: (hepatitis B surface antigen (HBsAg). anti-HBs antibodies, anti-HBc IgM and IgG, HBeAg and anti-HBe.
o PCR to detect viral DNA (quantitative or qualitative).
· HBV endemicity is divided according to HBsAg postivity, into three categories; high (>7%), intermediate (2-7%), low (<2% of population).
· HBsAg:
o It appears in serum within 1 – 10 weeks.
o If it persists > 6 months, it means chronic HBV infection
o It may correlate with the viral replication in the liver
o It predicts disease progression and t response to interferon treatment.
· Anti-HBs (immunity either natural or vaccination)
o It is a neutralizing antibody, that represents long-term immunity.
o In vaccinated individuals, anti-HBs is the only positive serological marker.
o In the past HBV infection, it is present together with anti-HBc IgG.
· HBeAg and anti-HBe:
o Indicate active viral replication and high infectivity (but its role is undertaken by HBV PCR).
o Appearance of antibodies indicates remission of hepatic disease.
· HBcAg:
o only intracellular (infected hepatocyte) and not identified in the serum.
· HBcAb:
o During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg.
o Then anti-HBc IgM disappears 6 months of acute infection, and Anti-HBc IgG continues to be positive in both resolved HBV infection and CHB.
· Causes of isolated anti-HBc IgG positivity;
o Window period of acute phase.
o After the end of acute infection (anti-HBs became undetectable).
o Several years of chronic HBV infection (undetectable HBsAg).
· HBV PCR:
o Real time PCR (either semiautomated or automated).
o It appears in serum 1 month after HBV infection.
o Peak level 3 months after the exposure to HBV (more than 108 copies/mL).
o Gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
o Higher titer means rapid progression and increased risk of HCC.
o It determines the need and the response to antiviral therapy.
· Genotyping:
o 10 genotypes, labelled A–J.
o A and D (increased chronicity) than B and C.
o Genotype C (high risk factor for perinatal and severe liver disease, including cirrhosis and HCC).
o Genotypes A and B have better treatment response to interferon.
o Genotype B or C may have decreased serological response to tenofovir.
· Acute HBV infection;
o Symptomatic, Jaundice, elevated liver enzymes.
o Positive HBsAg, anti-HBc IgM and HBV DNA.
· Chronic HBV infection;
o mostly asymptomatic.
o Persistent positive HBsAg for > 6 months.
· Past HBV infection;
o coexistence of anti-HBs IgG + anti-HBc.
· Occult HBV infection: detectable only by PCR (HBV DNA without HBsAg in the serum).
I appreciate that you mention genotypes, A–J, with varied differences in behaviour and patient outcomes. I like your well-structured summary.
Diagnosis and evaluation of HBV infection can be done through the following investigations
HBs Ag
anti-HBs Ab
HBeAg, anti-HBe
HBcAg
anti-HBc IgM and IgG.
Elevation of liver enzymes and symptoms
PCR
Diagnosis
Occult HBV infection is important to diagnose because of the following
Liver tissue (biopsy) PCR is indicated if all the following are present
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
I note that you have explained how a donor with evidence of past infection of HBV in relation to the decision -making necessary for transplantation.
This is a review article on treatment for HBV
HBV has 4 reading frames
(I) S that encodes for surface proteins (HBsAg);
(II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg);
(III) P for polymerase including reverse transcriptase;
(IV) X that encodes for a transcriptional transactivator factor (HBxAg
HBV endemicity is divided into three categories;
high, intermediate, low.
Diagnosis of HBV
1- Serological markers for HBV infection
2- Molecular methods for HBV infection
3- HBV genotyping
Serological markers for HBV infection
1- anti-HBs,
2- HBeAg,
3- anti-HBe,
4- anti-HBc IgM and IgG.
HBsAg
It is the serological hallmark of HBV infection.
It appears in serum within 1 to 10 weeks.
its Persistence for more than 6 months implies chronic HBV infection
Monitoring of quantitative HBsAg levels predicts treatment response to interferon in HBeAg negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs
It is known as a neutralizing antibody
confers long-term immunity
1- In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
2- In the past HBV infection, it is present in concurrence with anti-HBc IgG.
HBeAg and anti-HBe
It had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
HBcAg
It is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
anti-HBc IgM and IgG
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg .
After 6 months of acute infection, anti-HBc IgM wears off.
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Anti-Hbc positive can be seen in three conditions.
1- First, it can be predominantly seen as IgM class during the window period of acute phase.
2- Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
3- Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
Molecular methods for HBV infection
HBV DNA reveals the replication activity of the virus.
It is detectable at the early stage of infection (1 month after HBV infection)
it increases up to peak level approximately 3 months after the exposure to HBV
then gradually it diminishes in chronic infection or disappears at the recovery from HBV infection.
There are some techniques to identify and quantify HBV DNA:
1- Semi-automated qPCR
2 Semi-automated real-time PCR
3 Automated real-time PCR
4 Branched DNA.
Real-time PCR has come to be the standard method to detect and quantify HBV DNA in clinical setting.
Furthermore, it can be fully automated and does not generate carry-over contamination
HBV genotyping
can be divided into ten genotypes, labelled A–J.
Genotype B and C are restricted to Oceania and Asia,
whereas genotype A and D are omnipresent but most common in Africa and Europe
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
Methods of HBV genotyping
1- RFLP
2 Reverse hybridization
3- Genotype specific PCR
4 Sequence analysis
Diagnosis of hepatitis B infection
Acute hepatitis B
1- HBsAg pos
2- anti-HBc IgM can be pos
3 HBV DNA is pos
4- HBeAg can also be pos , but has little clinical importance.
chronic infection
HBsAg pos (for more than 6 months.)
Past HBV infection
1- anti-HBs pos
2- IgG anti-HBc pos
Occult HBV infection
1- low level of intrahepatic HBV DNA (gold standard but invasive )
2- HBsAg neg
3 anti-HBc IgG pos
4- HBsAg neg
5- antiHBs antibody neg
Occult HBV infection has some clinical importance.
1- First, it can be transmitted via solid organ transplantation, or hemodialysis.
2- Second, reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
3- Third, it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection .
4- Forth, it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis
Tests for occult HBV infection are considered in the following conditions:
1- in patients with cryptogenic liver disease, especially when having anti-HBc in serum;
2- in patients considering immunosuppression therapy or chemotherapy;
3- in solid organ transplantation donors, due to the possibilities for transmission
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
I note that you have explained how a donor with evidence of past infection of HBV in relation to the decision -making necessary for transplantation.
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family. It shows a diameter of 30–42 nm and consists of outer lipid envelope containing hepatitis B surface antigen (HBsAg) and an icosahedral capsid core composed of protein.
Serological markers for HBV infection consist of:
v HBsAg
v anti-HBs
v HBeAg
v anti-HBe
v anti-HBc IgM and IgG.
The identification of serological markers allows:
To identify patients with HBV infection.
To elucidate the natural course of chronic hepatitis B (CHB).
To assess the clinical phases of infection; and to monitor antiviral therapy.
Molecular methods for HBV infection:
ü Semi-automatedq PCR
ü Semi-automated real-time PCR
ü Automated real-time PCR
ü Branched DNA
HBV genotyping:
v RFLP
v Reverse hybridization
v Genotype specific PCR
v Sequence analysis
Diagnosis of hepatitis B infection:
Acute phase of infections:
Ø HBsAg.
Ø High serum aminotransferases.
Ø Anti-HBc IgM .
Ø HBV DNA
Ø HBeAg
Chronic infection:
Ø The persistence of HBsAg for more than 6 months
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
Summary of the article
Diagnosis of hepatitis B
HBV is dsDNA virus belongs to Hepadnaviridae family. Transmission of HBV via sexual activity, parenteral and vertical routes.
HBV serological markers:
1. Hepatitis B surface antigen (HBsAg): encodes for surface proteins and appears within 1 to 10 weeks after infection.
· Persistence of this marker for more than 6 months implies chronic HBV infection.
· The serum HBsAg titers are higher in patients with HBeAg- positive CHB than in HBeAg-negative CHB.
· Quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg- negative CHB patients with normal serum alanine aminotransferase levels.
· HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HbsAg.
2. Anti-HBs:
· A neutralizing antibody and confers long-term immunity.
· Positive HBs-Ab denotes acquired immunity through vaccination.
· Positive HBs-Ab in concurrence with anti-HBc IgG indicates past HBV infection.
· Positive HBs-Ab/positive HBsAg: denotes that anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV.
3. Anti-HBc IgM and IgG:
· HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
· anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
· After 6 months of acute infection, anti-HBc IgM wears off.
· Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
· Isolated anti-HBc positive: when HBsAg and anti-HBs are not detected. This can be seen in 3 conditions:
a) During the window period of acute phase: seen predominantly as IgM class.
b) After acute infection had ended: anti-HBs has decreased below the cutoff level of detection.
c) After several years of chronic HBV infection: HBsAg has diminished to undetectable levels.
· If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
4. Hepatitis B e antigen (HBeAg) and Anti-HBe:
· HBeAg and anti-HBe had been used to know infectivity and viral replication(has mostly been replaced by HBV DNA assay).
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
· Active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg.
Molecular methods for HBV infection
1. HBV DNA is a direct measurement of the viral load which reveals the replication activity of the virus.
· It is a reliable marker of replication activity.
· It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL).
· higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
· It is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
2. HBV DNA techniques: there are two principles of techniques to identify and quantify HBV DNA:
a) Signal amplification such as hybrid capture and branched DNA technology;
b) Target amplification such as polymerase chain reaction (PCR). Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107–108 IU/mL). Real-time PCR is the standard method to detect and quantify HBV DNA in clinical setting. It can be fully automated and does not generate carry-over contamination.
HBV genotyping
1. HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution.
· Genotype B and C: are restricted to Oceania and Asia.
· Genotype A and D: are omnipresent but most common in Africa and Europe.
· Genotype I is unusual and can be observed in Vietnam, Laos, India and China.
· Genotype J has been reported in Japan and Ryukyu.
· Other genotypes such as E, F, G, and H are also occasionally found in Asia.
2. HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
· Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
· Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
· Genotype B or C had a lower opportunity to gain serological response to tenofovir.
3. HBV genotyping can be confirmed using diverse methods:reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fluorescence polarization assay.
Diagnosis of hepatitis B infection
1. Acute hepatitis B is a clinical diagnosis identified by the detection of HBsAg, symptoms, high serum aminotransferases.
2. Usually anti-HBc IgM can be detected and HBV DNA is present.
3. HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
4. The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
5. Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
6. Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
7. Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody.
8. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum.
9. Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases.
10.Occult HBV infection has some clinical importance:
· it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis.
· reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
· it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
· it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis.
11.Tests for occult HBV infection are considered in the following conditions:
· in patients with cryptogenic liver disease (when having anti-HBc in serum).
· in patients considering immunosuppression therapy or chemotherapy.
· in solid organ transplantation donors, due to the possibilities for transmission.
. I like your summary, level of evidence, analysis and conclusions.
I appreciate
your understanding of a range of genotyping tests in HBV infection.
Please summarise this article.
Introduction
Hepatitis B virus (HBV) infection can lead to severe liver disease such as cirrhosis and hepatocellular carcinoma (HCC).
HBV is a circular, partly double-stranded DNA virus
Serological markers include : hepatitis B surface antigen (HBsAg) and anti-HBs, anti-HBc IgM and IgG, and hepatitis B e antigen (HBeAg) and anti-HBe.
Transmitted is by sexual, parenteral and vertical route.
High endemic areas are China, South East Asia, Indonesia, and sub-Saharan Africa
South America, South West Asia, Eastern and Southern Europe Intermediate areas.
North America and Western Europe are grouped as low endemic regions with prevalence rates range from 0.5% to 2%.
Serological markers for HBV infection
HBsAg– HBsAg is the serological hallmark of HBV infection. It appears in serum within 1 to 10 weeks.
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression
Anti-HBs – It is known as a neutralizing antibody, and confers long-term immunity
HBeAg and anti-HBe– HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease
HBcAg– It is intracellular and not identified in serum.
Anti HBc-The presence of anti-HBc indicates previous or ongoing infection with hepatitis B virus.
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load
It is detectable at the early stage of infection 1-3 months
HBV DNA is a reliable marker of replication activity, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
Two techniques to identify and quantify HBV DNA:
Signal amplification such as hybrid capture and branched DNA technology
Target amplifcation such as polymerase chain reaction (PCR)
HBV genotyping
Ten genotypes, labelled A–J
Genotype B and C are restricted to Oceania and Asia
Genotype A and D are omnipresent but most common in Africa and Europe
Genotype I is unusual and can be observed in Vietnam, Laos, India and China
Genotype J has been reported in Japan and Ryukyu
Other genotypes such as E, F, G, and H are also occasionally found in Asia.
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Diagnosis of hepatitis B infection
Acute hepatitis B – identified by the detection of HBsAg, symptoms, high serum aminotransferases. HBeAg can also be identified in most acute phase of infections
Chronic infection is based on the persistence of HBsAg for more than 6 months
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg
It is defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody
Detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV
Importance of Occult HBV Infection
· Transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation
· Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised.
· It can accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
· It is a risk factor for HCC by its carcinogenic effect
I appreciate
that you mention genotypes, A–J, with varied geographic distributions.
Serological markers:
1- HBsAg:
2- Anti-HBs (a neutralizing antibody), and confers long-term immunity .
3- HBeAg and anti-HBe:
4- anti-HBc IgM and IgG:
5- HBV DNA: is a direct measurement of the viral load
Diagnosis of hepatitis B infection
1- Acute hepatitis B:
2- chronic hepatitis:
3- Past infection:
4- Occult HBV infection:
Clinical significance of occult hepatitis B:
1- It can be transmitted via transfusion, solid organ transplantation.
2- Reactivation of HBV infection may occur in patients receiving chemotherapy or
immunocompromised state.
3- It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver
disease including chronic hepatitis C infection.
4- A risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic
inflammation and fibrosis
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
Diagnosis of hepatitis B.
Introduction.
HBV is one of the most common cause of chronic liver disease worldwide, especially in high endemic area, HBV vaccination is available worldwide and considered one of the important tools for preventing HBV spreading, and herein this article reviewed the serological and molecular diagnosis of HBV.
Serological markers for HBV infection.
Serological markers for HBV infection consist of HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG which are used to assess course of chronic hepatitis B (CHB), the clinical phases of infection; and to monitor antiviral therapy.
HBsAg appears in serum within 1 to 10 weeks of exposure and if persist for more than 6 months implies chronic HBV infection, the serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB, and is better marker for monitoring in CHB patients with HBeAg negative and normal serum alanine aminotransferase levels.
In vaccinated person, anti-HBs is the only serological marker detected in serum but if associated with anti-HBc IgG indicates past infection.
HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
HBcAg is an intracellular and not detected in serum and Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Isolated anti-HBc positive: considered when HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs which occurs in the window period of acute phase, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection, HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection with negative HBsAg.
Molecular methods for HBV infection.
HBV DNA increase up with acute infection and decrease later on with CHB and disappear after recovery.
HBV DNA are related to the more rapid disease progression and higher incidence of HCC. Furthermore, HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
HBV genotyping.
HBV can be divided into ten genotypes, labelled A–J: they have distinct geographic distribution, for example genotype B and C are restricted to Oceania and Asia, also each genotype has its character such as acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
Diagnosis of hepatitis B infection.
Acute hepatitis B: diagnosed by detection of HBsAg, symptoms, high serum aminotransferases, anti-HBc IgM can be detected and HBV DNA is present.
Chronic infection :the persistence of HBsAg for more than 6 months.
Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg and it is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti- HBs antibody, thus HBV DNA testing is widely used to diagnose occult HBV infection.
Occult HBV infection diagnosis is challenging and so tests should be considered in following conditions:
1-Patients with cryptogenic liver. disease, especially when having anti-HBc in serum.
2-Patients considering immunosuppression therapy or chemotherapy; and in solid organ transplantation donors, due to the possibilities for transmission.
Clinical significance of occult HBV infection in the following conditions:
1-It can be transmitted via transfusion, solid organ transplantation including orthotropic liver transplantation or hemodialysis.
2-Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state .
3-Accelerate liver injury and lead to hepatic fibrosis in patients with
chronic liver disease including chronic hepatitis C infection.
4-It appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis.
Conclusion:
Diagnosis of HBV is important to reduce its infection specially in immunocompromised patients , diagnosis usually started with serology and then amplification of viral DNA to detect active/chronic and occult infection and used also to follow up treatment.
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections.
Introduction
HBV belongs to the Hepadnaviridae family. It a circular, partly double-stranded DNA virus
HBV infection is responsible for the most of chronic liver diseases worldwide
Mode of transmission: parenteral, sexual and vertical route
HBsAg prevalence:
1. High endemic areas (China, South East Asia, Indonesia, and sub-Saharan Africa). > 8% of the population are affected
2. Intermediate endemic areas (South America, South West Asia, Eastern and Southern Europe). chronic HBV infection rate 2%-7%
3. Low endemic areas (developed countries). Prevalence is 0.5%-2%
Aim of the study: review serological and molecular tests for diagnosis of HBV infection
Serological markers for HBV infection
HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG
HBsAg:
o Appears in serum within 1 to 10 weeks after exposture
o Persistence more than 6 months implies chronic HBV infection
o Titers are higher in patients with HBeA gpositive chronic hepatitis B (CHB) than in HBeAg-negative CHB
Anti-HBs (neutralizing antibody):
o Confers long-term immunity
o Are unable to neutralize the circulating viruses in most cases (so regarded as carriers of HBV)
o It is the only serological marker for vaccination
o Simultaneous appearance of HBsAg and anti-HBs may occur in patients with HBsAg positive
HBeAg and anti-HBe:
o For Infectivity and viral replication (replaced by HBV DNA)
o Seroconversion is related to the remission of hepatic disease
o Active viral replication is sustained in some patients with HBe seroconversion due to mutations in the pre-core and core region that inhibit or decrease the production of HBeAg
HBcAg:
o Intracellular in infected hepatocyte and is not identified in the serum
anti-HBc:
o During acute infection, anti-HBc IgM and IgG appears 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms
o Anti-HBc IgM wears off after 6 months of acute infection
o Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB
o Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs (isolated anti-HBc positive)
o Isolated anti-HBc positive seen in 3 conditions:
1. seen as IgM class during the window period of acute phase
2. after acute infection ended, anti-HBs has decreased below the cutoff level of detection
3. after several years of chronic HBV infection, HBsAg has diminished to undetectable levels
o In isolated anti-HBc positive, anti-HBc IgM should be checked to assess the possibility of recent HBV exposure
o HBV DNA should be tested in chronic liver disease patients to find out occult HBV infection (existence of detectable HBV DNA without serum HBsAg)
Molecular methods for HBV infection
o HBV DNA (replication activity)
o Detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 10*8 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection
o Also useful in serologically negative HBV infection (HBeAg-negative CHB and occult HBV infection)
o Detection is a reliable marker of replication activity, and higher titers are related to the more rapid disease progression and higher incidence of HCC
o Useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment
Two principles of techniques to identify and quantify HBV DNA:
1. signal amplification such as hybrid capture and branched DNA technology
2. target amplification such as polymerase chain reaction (PCR)
Quantitative methods for HBV DNA:
1. Semi-automated Qpcr
2. Semi-automated real-time PCR
3. Automated real-time PCR
4. Branched DNA
HBV genotyping
According to the sequence divergence, HBV can be divided into ten genotypes (A–J): they have distinct geographic distribution
Genotype B and C are restricted to Oceania and Asia
Genotype A and D are omnipresent but most common in Africa and Europe
Genotype I is unusual: Vietnam, Laos, India and China
Genotype J (Japan and Ryukyu)
Genotypes E, F, G, and H are also occasionally found in Asia
HBV genotyping is useful in predicting HBV disease progression and determine appropriate antiviral therapy
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
Patients infected with genotype B or C had a lower opportunity to gain serological response to tenofovir (recent study)
HBV genotyping can be confirmed using diverse methods:
1. reverse hybridization: high sensitivity, automated systems but relatively high cost
2. genotype-specific PCR assays: high sensitivity, automated systems, easy to perform, and suitable for detecting mixed genotype infections but high cost
3. real-time PCR
4. restriction fragment-length polymorphism (RFLP): easily done, low cost, simple, rapid but low sensitivity for typing samples with low HBV
5. sequence analysis: gold standard method for genotyping, identification of patients infected with recombinant genotypes but it is time consuming, technically demanded
6. microarray (DNAChip)
7. fluorescence polarization assay
Diagnosis of hepatitis B infection
Acute hepatitis B:
o Detection of HBsAg, symptoms, high serum aminotransferases
o Usually anti-HBc IgM can be detected and HBV DNA is present
o HBeAg can be identified in most cases (of little clinical importance)
chronic HB infection:
o Persistence of HBsAg for more than 6 months
o Commonly diagnosed by laboratory means but not by clinical presentations
Past HBV infection:
o Coexistence of anti-HBs and IgG anti-HBc
Occult HBV infection:
o Persistence of low level of intrahepatic HBV DNA without detectable HBsAg
o It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and antiHBs antibody
o Detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection (invasive procedture), since cccDNA remains in the hepatocytes and HBV DNA is occasionally identified in the liver but not in the serum
o Real-time PCR for serum HBV DNA detection have a adequate sensitivity to identify occult HBV infection in many cases (HBV DNA testing is widely used to diagnose occult HBV infection)
o Importance of occult HBV infection:
1. it can be transmitted via transfusion, SOT including orthotopic liver transplantation, or hemodialysis
2. reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3. it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4. it is a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis
o Tests for occult HBV infection in:
1. patients with cryptogenic liver disease, especially when having anti-HBc in serum
2. in patients considering immunosuppression therapy or chemotherapy
3. in solid organ transplantation donors, due to the possibilities for transmission
Conclusions
o Serological markers for detecting antigens and antibodies are the first step of HBV diagnosis
o To prove first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used
o Diagnosis of HBV infection is important for determining acute, chronic and occult cases of infection in order to establish preventive measures and to start antiviral treatment
I appreciate that you mention genotypes, A–J, with varied geographic distributions.
Diagnosis of Hepatitis B ===> Summary of This Article:
Introduction:
Hepatitis B infection is a one of the major problems worldwide that leads to liver cirrhosis and hepatocellular carcinoma, Globally, about 240 million people are living with a chronic state of the virus which could progress to liver cirrhosis or HCC, HBV is a double-stranded virus that belongs to the family Hepadnaviridae, The presence of vaccination is active method to decrease the burden of the disease.
Mode of transmission:
Parenteral route
Vertical route
Sexual route
HBV Endemicity:
High endemicity, affects more than 8% population – China, Southeast Asia, Indonesia, Sub-Saharan African
Intermediate, affects between 2-7% population – South American, Southwest Asia, Eastern, and Sothern Europe
Low endemicity, the prevalence of 0.5 -2% – North American, Western Europe
The identification of serological markers allows:
To identify patient with HBV infection
To know the natural course of chronic HBV infection
To assess the clinical phases of infection
To assess antiviral therapy
Serological Markers for HBV infection:
HBsAg – serological hallmark of HBV infection, After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks, its persistence in the serum more than 6 months denotes chronic infection, it is elevated in the patient positive HBeAg chronic HBV CHB more than in CHB with negative HBeAg.
Anti-HBs – In vaccinated patient, neutralizing antibody that provide long-life immunity.
In past infection, anti-HBs is present together with anti-HBc IgG
HBsAg and Anti-HBs when present together denotes carrier status of HBV, as antibodies are unable to neutralize the virus.
HBeAg – use for viral infectivity.
Anti-HBe – use for viral infectivity.
Anti-HBc IgM and IgG
HBV DNA replace HBeAg as a measure for infectivity.
During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
After 6 months of acute infection, anti-HBc IgM disappeared and Anti-HBc IgG is detected in both patients with resolved HBV infection and CHB
Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive, in the following situations:
– Predominantly seen as IgM class during the window period of acute phase.
– After acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
– After several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
– In isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
– HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection:
HBV DNA is the direct measurement of viral load (replication activity).
DNA can be detected early in a month after infection and reach peak at 3 months.
Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
HBV DNA is a useful marker for starting the treatment.
DNA quantified by PCR (standard method).
HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting. Furthermore, it can be fully automated and does not generate carry-over contamination.
HBV genotyping:
It has 10 genotypes with distinct geographical distribution, and significantly predict disease progression and determine the suitable antiviral therapy.
Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
Patients with genotype B or C had a lower opportunity to gain serological response to tenofovir.
Diagnosis of hepatitis B infection:
A- Acute hepatitis B is a clinical diagnosis identified by :
the detection of HBsAg, symptoms, high serum aminotransferases, Usually anti-HBc IgM can be detectedHBV DNA is present.
HBeAg can also be identified in most acute phase of infections
B- Chronic HBV is based on the persistence of HBsAg for more than 6 months, Anti HBs
Ant HBc IgG
C- Past infection diagnosed by presence of anti-HBs and anti-HBc IgG antibodies.
D- Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg, by serological method, isolated anti-HBc is present with the absence of HBsAg and antiHBs antibody. The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, means virus remains and detected in the liver not in the serum.
E- Occult HBV infection
– it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation, or hemodialysis.
– Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
– it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
– it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis.
Tests for occult HBV infection are considered in the following conditions:
– In patients with cryptogenic liver disease, especially when having anti-HBc in serum.
– In patients considering immunosuppression therapy or chemotherapy.
– In solid organ transplantation donors, due to the possibilities for transmission.
Occult HBV infection:
Persistent low level of intrahepatic HBV DNA but absent HBsAg and anti-HBs, Can be transmitted by blood transfusion of SOT
HBV can be reactivated in patient on immunosuppressive
it can accelerate liver injury and can lead to HCC
Conclusion:
Both quantitative (HBV DNA for viral load) & qualitative molecular tests are needed to identify viral load and identify HBV genotypes, All these will help to establish if the infection is acute or chronic & to establish preventive methods and effective antiviral treatment and and monitor responses to antiviral agent
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
Introduction: Hepatitis B Virus (HBV) isa 30-42 nm diameter hepadnaciridae family member having an icosahedral capsid core and outer lipid envelope containing hepatitis B surface antigen (HbsAg). It has circular, partly double-stranded DNA with 4 overlapped open reading frames (S, pre-C/C, P, and X). Mutation rate is high due to error susceptibility of the reverse transcriptase involved in HBV replication. HBV causes most of the CLD (chronic liver diseases: cirrhosis and hepatocellular carcinoma, HCC) worldwide, and is spread through sexual, parenteral, and vertical routes. HBV endemicity can be high (>8%: China and South-East Asia), intermediate (2-7%: South America, Eastern and Southern Europe), and low (0.5-2%: North America and Western Europe). Early diagnosis of HBV infection, and vaccination against HBV can be helpful in decreasing the burden of HBV infection.
Serological markers of HBV infection: These include HBsAg, Anti-HBs antibody, HBeAg, AntiHBe antibody, AntiHBc antibody (IgG and IgM).
Molecular methods for HBV infection: HBV DNA assays should be done in CLD patients without HBsAg to find out occult HBV infection.
HBV DNA can be detected within 1 month of infection, with peak levels at 3 months reduces. High titres signify more rapid disease progression and increased risk of HCC. The 2 principles of techniques used to identify and quantify HBV DNA include signal amplification (hybrid capture and branched DNA technology) and target amplification (polymerase chain reaction, PCR).
HBV genotyping: There are 10 different genotypes of HBV (A-J) due to high genetic heterogeneity. Genotype A and D are present everywhere; B &C are present in Asia and Oceania; E, F, G & H are occasionally present in Asia; I is unusual, present in Vietnam, India, Laos, & China; J is seen in Japan and Ryukyu.
Genotype A & D have increased rates of chronicity; A & B have better response to interferon treatment; B & C have lower response to Tenofovir; Genotype C s associated with perinatal infection as well as severe liver disease (cirrhosis and HCC).
Diagnosis of HBV infection:
Conclusion: HBV infection is common, and has grave implications for liver. HBV diagnosis involves serological testing followed by use of molecular tests to verify the diagnosis, which will help in preventive and therapeutic tool use to reduce the adverse effects of HBV infection.
I appreciate that you mention genotypes, A–J, with varied geographic distributions.
Diagnosis of hepatitis B
Introduction
· HBV is partly double-stranded DNA with a high mutation rate.
· HBV infection is responsible for the most of chronic liver diseases worldwide.
· In addition to HBV vaccination, the diagnosis of acute, chronic and occult HBV infection, diminish the burden of this disease.
Serological markers for HBV infection
HBsAg
· It is the serological hallmark of HBV infection.
· It appears in serum within 1 to 10 weeks after an acute exposure to HBV.
· If detected for more than 6 months, it implies chronic HBV infection.
· Monitoring of quantitative HBsAg levels predicts treatment response to interferon.
Anti-HBs
· Anti-HBs confers long-term immunity.
· After vaccination, anti-HBs is the only serological marker detected in serum.
· If present with anti-HBc IgG, this indicates past HBV infection.
· If anti-HBs antibodies are unable to neutralize the circulating viruses, these patients are regarded as carriers of HBV.
HBeAg and anti-HBe
· HBeAg to anti-HBe seroconversion is related to the remission of hepatic disease.
· In the past were used to detect infectivity and viral replication, but now replaced by HBV PCR.
HBcAg
· It is present intracellulary, so it is not identified in the serum.
· After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Molecular methods for HBV infection
HBV DNA
· A direct measurement of the viral load.
· It is detectable at the early as stage 1 month after HBV infection.
· It disappears at the recovery from HBV infection.
· HBV-DNA can help detection of infection in serologically negative HBV infection, HBeAg-negative CHB and occult HBV infection.
· Higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
· HBV DNA testing is useful to determine patients who need antiviral therapy and monitor them for treatment
· Real-time PCR is the standard method to detect and quantify HBV DNA.
HBV genotyping
· HBV has a high genetic heterogeneity.
· HBV can be divided into ten genotypes (labelled A–J).
· HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy
· Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
· Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
· In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
Diagnosis of hepatitis B infection
· Acute hepatitis B is identified by the detection of HBsAg, symptoms and high serum aminotransferases.
· Usually anti-HBc IgM can be detected and HBV DNA is present in the acute state
· The diagnosis of chronic infection is based on the persistence of HBsAg for more than 6 months.
· Past HBV infection is defined by the coexistence of anti-HBs and IgG anti-HBc.
· Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. Serologically there is isolated anti-HBc with the absence of HBsAg and antiHBs antibody.
· HBV DNA testing is widely used to diagnose occult HBV infection.
· Tests for occult HBV infection are considered in patients with cryptogenic liver disease, in patients considering immunosuppression therapy or chemotherapy and in solid organ transplantation donors.
I make note of your interpretation of serological and molecular testing reports pertaining to HBV infections in relation to timings in the natural history HBV infections. I like your summary.
Serological markers for HBV infection consist of
HBsAg,anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
HBsAg After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks. Persistence of this marker for more than 6 months implies chronic HBV infection
Differences in the serum HBsAg levels during the different phases of infection indicate the distribution of cccDNA during the respective phases of the disease.
The serum HBsAg titers are higher in patients with HBeAg positive CHB than in HBeAg-negative CHB
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels
Anti-HBs confers long-term immunity -acquired immunity through vaccination,
the simultaneous appearance of HBsAg and anti-HBs has been reported in patients with HBsAg positive In most cases, anti-HBs antibodies are unable to neutralize the circulating viruses, thus these patients are regarded as carriers of HBV
In the past, HBeAg and anti-HBe had been used to know infectivity and viral replication, but their use for this purpose has mostly been replaced by HBV DNA assay.
HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms. After 6 months of acute infection, anti-HBc IgM wears off. Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
isolated anti-HBc positive.
1-it can be predominantly seen as IgM class during the window period of acute phase.
2-after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
3- after several years of chronic HBV infection, HBsAg has diminished to undetectable levels. If the result of serological markers shows isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg
Molecular methods for HBV infection HBV DNA
– a direct measurement of the viral load, which reveals the replication activity of the virus. It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
– higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC
– HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment
principles of techniques to identify and quantify HBV DNA:
signal amplification such as hybrid capture and branched DNA technology; target amplifcation such as polymerase chain reaction (PCR)
Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting.
HBV can be divided into ten genotypes, labelled A–J:
geographic distribution
Genotype B and C are restricted to Oceania and Asia,
genotype A and D most common in Africa and Europe .
Genotype I is unusual and can be observed in Vietnam, Laos, India and China
genotype J has been reported in Japan and Ryukyu
Other genotypes such as E, F, G, and H are also occasionally found in Asia.
Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy. Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC
In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D . Recent studies reported that patients infected with genotype B or C had a lower opportunity to gain serological response to tenofovir
The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, since cccDNA remains in the hepatocytes and HBV DNA is occasionally identifed in the liver but not in the serum. However, gaining hepatic HBV DNA is diffcult in clinical setting since the procedure is invasive.
Real-time PCR for serum HBV DNA detection have been shown with adequate sensitivity to identify occult HBV infection in many cases; thus, HBV DNA testing is widely used to diagnose occult HBV infection
Occult HBV infection clinical importance.
1- it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation or hemodialysis
2- reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state
3- it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection
4- it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fbrosis
I appreciate that you mention genotypes, A–J, with varied geographic distributions.
. I like your summary, analysis and conclusions.
Summary
Introduction
HBV belongs hepadnaviridae family.
The reverse transcriptase responsible for viral replication is susceptible to errors thus high mutation rate.
HBV is transmitted parenteral, sexual and vertical route.
HBV is divided into three endemicity: high, intermediate and low; with high endemicity in sub saharan Africa and south east Asia.
Serological markers
HBsAg is the serological hallmark of HBV infection.
Anti-HBs is a neutralising antibody, confers longterm immunity.
HBeAg and anti-HBe prior used to indicate replication and infectivity however has been replaced by HBV DNA.
HBcAg is an intracellular antigen thus not detected in serum.
Molecular methods
HBV DNA indicates the replication of the virus.
HBV genotype
There are 10 genotypes A-J with distinct geographical distribution.
Genotype A and D associated with higher rates of chronicity than B and C.
Genotype A and C have better response to interferon than B and D.
Diagnosis
Acute hepatitis B infection is a clinical diagnosis based by the presence of HBsAg detection, raised serum aminotransferase and symptoms.
Chronic hepatitis detected by persistence of HBsAg beyond 6 months.
Past infection identified by Anti-HBs and IgG anti-HBc antibodies.
Occult infection defined by detection of HBV DNA intra-hepatic.
I appreciate that you mention genotypes, A–J, with varied geographic distributions.
Introduction
1- HBV is one of the major problem worldwide that leads to liver cirrhosis and hepatocellular carcinoma HCC
2- It is double stranded DNA, has different serological markers.
3- Infection can be acute, chronic and occult cases.
4- First step to diagnose it is by serological marker then to quantify the load of the virus.
5- The presence of vaccination is active method to decrease the burden of the disease.
Serological markers for HBV infection
1- HBsAg is the serological hallmark of HBV infection. After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks, its persistence in the serum more than 6 months denotes chronic infection, it is elevated in the patient positive HBeAg chronic HBV CHB more than in CHB with negative HBeAg.
2- Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels ALT.
3- Anti-HBs it is neutralizing antibody, and provide long-life immunity
4- In vaccinated patient, the only marker appeared is Anti-HBs
5- In past infection, anti-HBs is present together with anti-HBc IgG
6- HBsAg and anti-HBs when present together denotes carrier status of HBV, as antibodies are unable to neutralize the virus.
7- HBeAg means infectivity.
8- Anti-HBeAg means seroconversion or remission of hepatic disease
9- HBV DNA replace HBeAg as a measure for infectivity
10- HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum
11- During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
12- After 6 months of acute infection, anti-HBc IgM disappeared and Anti-HBc IgG is detected in both patients with resolved HBV infection and CHB
13- Some HBsAg-negative individuals are positive for anti-HBc IgG without anti-HBs, in this situation, it should be considered isolated anti-HBc positive, in the following situations:
a- predominantly seen as IgM class during the window period of acute phase.
b- after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
c- after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
14- In isolated anti-HBc positive, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
15- HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Molecular methods for HBV infection
1- HBV DNA is a direct measure to replication and infectivity of the virus, it started to appear after 1 month of acute infection, and reach peak after 3 months of acute infection, then decreased in chronic stage and disappear in the recovery stage.
2- higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
3- HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
4- Real-time PCR can detect wide dynamic range of viral load (lower range, 10–15 IU/mL; upper range, 107 –108 IU/mL). For this reason, it has come to be the standard method to detect and quantify HBV DNA in clinical setting. Furthermore, it can be fully automated and does not generate carry-over contamination.
HBV genotyping
1- It has 10 genotypes with distinct geographical distribution, and significantly predict disease progression and determine the suitable antiviral therapy.
2- Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C
3- Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC
4- In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D
5- Patients with genotype B or C had a lower opportunity to gain serological response to tenofovir.
Diagnosis of hepatitis B infection
1- Acute hepatitis B is a clinical diagnosis identified by :
a- the detection of HBsAg
b- symptoms
c- high serum aminotransferases
d- Usually anti-HBc IgM can be detected
e- HBV DNA is present.
f- HBeAg can also be identified in most acute phase of infections
2- Chronic infection is based on the persistence of HBsAg for more than 6 months.
3- Past infection diagnosed by presence of anti-HBs and anti-HBc IgG antibodies.
4- Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg, by serological method, isolated anti-HBc is present with the absence of HBsAg and antiHBs antibody . The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, means virus remains and detected in the liver not in the serum.
5- Occult HBV infection
a- it can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis.
b- reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
c- it may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
d- it appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic infammation and fibrosis.
6- Tests for occult HBV infection are considered in the following conditions:
a- in patients with cryptogenic liver disease, especially when having anti-HBc in serum
b- in patients considering immunosuppression therapy or chemotherapy
c- in solid organ transplantation donors, due to the possibilities for transmission
I make note of your interpretation of virology reports in relation to timings of test and viral load pertaining to HBV infections
I like your detailed well-structured summary.
Summary
Introduction
This article is focussed on the diagnosis of hepatitis B infection. Hepatitis B is a serious infection that can lead to significant liver damage, cirrhosis and hepatocellular carcinoma. This infection is caused by a double stranded DNA virus with different serological markers. Disease transmission is via sexual, parenteral, or vertical route. The way to fight the infection is early diagnosis and treatment.
Discussion
Hepatitis B identification of serological markers is significant for effective management of disease. This allows the physician to make suer that the natural course of infection is dealt with appropriately, while monitoring the clinical phases and response to antiviral therapy.
HBsAg is a serological marker of HBV infection and appears in serum 1 to 10 weeks of exposure. If the marker is still seen in serum after 6 months, it means that the patient has chronic HBV infection.
Another serological marker that is significant to determine infection is anti-HBs. It is the only marker that can be seen in patients who have been vaccinated against Hepatitis B.
During acute infection, IgM and IgG emerge 1-2 weeks after HBsAg is seen in serum, along with raised serum aminotransferase and symptoms. After 6 months, IgM starts reducing. IgG continues even after infection is resolved.
Occult infection is characterized by existence of detectable HBV DNA without serum HBsAg. HBV DNA assay is to be done to identify if chronic liver disease patients have occult HBV infection.
Two methods of detection of HBV DNA in any patient include methods that quantify HBV DNA. These include :
Conclusion
Both quantitative and qualitative molecular tests are needed to identify viral load and identify HBV genotypes In order to establish preventive methods and effective antiviral treatment.
. I like your summary.
I appreciate your understanding of a range of genotyping tests in HBV infection.
SUMMARY
Introduction
Hepatitis B infection is a public health concern in view of its grave complication and lack of treatment to cure for now even though vaccination is available. Globally, about 240 million people are living with a chronic state of the virus which could progress to liver cirrhosis or HCC. HBV is a double-stranded virus that belongs to the family Hepadnaviridae.
Mode of transmission
HBV Endemicity
Serological Markers for HBV infection
The identification of serological markers allows:
Molecular methods for HBV Infection
HBV DNA
Diagnosis of HBV infection
A) Acute hepatitis is diagnosed by
B) Chronic HBV
Occult HBV infection
Conclusion
The total care for HBV infection is not to use serological markers to make diagnosis, but the use of HBV DNA for viral load, and identification of the genotype is paramount. All these will help to establish if the infection is acute or chronic and to also administer and monitor responses to antiviral agent use for treatment.
I make note of your interpretation of virology reports in relation to timings of test and viral load pertaining to HBV infections I like your detailed well-structured summary.
Introduction
Serological markers for HBV infection
1.HBSAg
2.Anti-HBsAb
3.HBeAg & anti-HBeAb
4.HBcAg
Molecular diagnosis
HBV genotyping
Diagnosis
Occult HBV infection
Clinical significance of occult HBV infection:
Conclusion
I make note of your interpretation of serological reports pertaining to HBV infections. I like your summary.
Thnxs, prof
Diagnosis of Hepatitis B
Summary of the Article
Introduction
A DNA virus contains the following
2. The covalently closed circular DNA (cccDNA) is the transcriptional template of HBV.
3. Reverse transcriptase is involved in the replication of HBV.
HBV is responsible for most liver diseases worldwide and about 240 million people are infected by chronic HBV.
According to the HBsAGs prevalence, HBV endemicity is divided into three categories;
Serological Marker of HBV infection
The identification of the marker alow;
HBsAg is the hollmarker of HBV infection, HBsAg appear on the serum within 1-10 days, and the infection considered chronic if the NBsAg detected in the serum for than 6 months, so;
Anti-HBs is the only serlogical marker that indicate acquired immunity.
HBeAg and anti-HBe seroconversion is related to remission of the hepatic disease.
HBV DNA is used for viral activity and replication.
HBcAg denote hepaticyte infection, and detection of both HBcAg IgM and IgG denote that te infection is during acute phase, and presence of only IgG denote either resolved infection or CHB.
In isolated anti-HBc positive with HBsAg and HBs-negative, a serum IgM should be detected to diagnose the occurrence or not of acute infection.
Isolated cases of HBc-positive can denote the following;
Molecular methods for HBV infection
Diagnosis of HBV infection
Acute infection;
CHB;
Past HBV infection;
Occult HBV infection
2.Reactivation of HBV infection in immunocompromised patients.
3.It can accerelate liver injury hepatic fibrosis in patient with CLL.
4.Risk factor of HCC.
I appreciate that you mention genotypes, A–J, with varied geographic distributions and differences of outcomes. I like your summary.
Summarise this article
Introduction
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family.
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
According to HBsAg prevalence, HBV endemicity is divided into three categories; high, intermediate, low.the high edamic area are China, South East Asia, Indonesia, and sub-Saharan Africa which represent about 8% of general population.
South America, South West Asia, Eastern and Southern Europe which is intermediate about 2% to 7%.
North America and Western Europe which is lowest area of endemicity around .0.5 % to 2 %.
Vaccination is the way to decrease the HBV infection.
Serological markers for HBV infection.
HBsAg
is the serological hallmark of HBV infection, after acute infection appears in serum within 1 to 10 weeks.
If remaining for more than 6 month patient diagnosed as chronic HBV infection .
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative chronic hepatitis B patients with normal serum alanine aminotransferase levels.
anti-HBsg
called as neutralizing antibody, and confers long-term immunity, and confers long-term immunity.
HBcAg
is an intracellular presence in infected hepatocyte, thus it is not identified in the serum. During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
HBeAg
Monitoring of quantitative HBsAg levels predicts treatment response to interferon and disease progression in HBeAg negative CHB patients with normal serum alanine aminotransferase levels.
anti-HBc IgM and IgG
anti-HBc IgM become negative after 6 month of acute HBV infection .
Anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
Molecular methods for HBV infection
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
It is appear at the early stage of infection (1 month after HBV infection) and increases up to peak level (more than 108 copies/mL) approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
PCR is test that to detect and quantify HBV DNA which is fully automated .
HBV genotyping
HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.genotyping is important to determine response to treatment .
Diagnosis of hepatitis B infection
Diagnosis of HBV infection by, symptoms, high serum aminotransferases and the detection of HBsAg.
To diagnose chronic HBV infection is based on the persistence of HBsAg for more than 6 months,in absence of any symptoms .
Occult HBV infection is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
Occult HBV infection is diagnosed by detection of HBV DNA in the liver( invasive ).
occult HBV infection diagnosis also by Real-time PCR for serum HBV DNA( sensitive)
It is important because occult HBV infection transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation and hemodialysis .
Conclusion
Hepatitis B virus infection is a major health problems and causing severe liver disease such as cirrhosis and hepatocellular carcinoma .
Vaccination is the important to limited infection of HBV infection .
This article define what is HBV ,how to diagnose ,occult HBV infection .
I make note of your interpretation of serological reports pertaining to HBV infections. I like your summary.
I. Diagnosis of hepatitis B
====================================================================
Introduction
Serological markers for HBV infection
Molecular methods for HBV infection
HBV genotyping
Diagnosis of hepatitis B infection
Conclusions
. I like your summary, level of evidence, analysis and conclusions.
I appreciate your understanding of a range of genotyping tests in HBV infection.
Typing whole sentence in bold or typing in capitals amounts to shouting.
Thanks alot for you Prof.Sharma
Summarise this article
Introduction:
Serological markers for HBV infection:
Molecular methods for HBV infection:
There are two principles of techniques to identify and quantify HBV DNA:
Real-time PCR is the standard of care to detect and quantify HBV DNA in clinical setting.
Higher titers of HBV DNA, related to disease progression and higher incidence of HCC.
HBV DNA can detect occult HBV infection in the absence of HBsAg.
HBV genotyping:
Genotyping done the following methods:
Utility of genotyping:
Diagnosis of hepatitis B infection:
Clinical importance of Occult HBV infection are:
Tests for occult HBV infection are considered in the following conditions:
Conclusion:
The article identify the structure of HBV and tools used for quantitative and qualitative (gentypes) testing of the virus, and defining and better understanding the acute, chronic and occult HBV infection, thus; help to prevent and treat the HBV.
Well done
Introduction:
-Hepatitis B virus (HBV) infection is a global health problems and responsible for majority of chronic liver disease.
-Its prevalence varies according to geographic distribution, and can be divided into high, intermediate, low.
-HBV is a circular, partly double-stranded DNA virus with various serological markers.
-Hepatitis B vaccination, accurate diagnosis and treatment methods to decrease disease burden.
Serological markers for HBV infection:
-Markers allow to identify infected patients; to elucidate the natural course of infection; to assess the clinical phases of infection; and to monitor antiviral therapy.
-Serological markers for HBV infection consist of:
HBsAg:
– It is the serological hallmark of HBV infection. Appears in serum within 1 to 10 weeks.
– Persistence elevation > 6 months implies chronic infection.
– Patients with HBeAg positive CHB have higher HBsAg level.
– Its monitoring predicts treatment response and disease progression in HBeAg negative CHB patients.
Anti-HBs
– It is neutralizing antibody, and confers immunity.
– Concurrence with anti-HBc IgG ; indicate past HBV infection.
– Carriers of HBV have it simultaneously with HBsAg, when virus was not neutralize completely.
HBeAg and anti-HBe
– Previously used to indicate infectivity and viral replication, but HBV DNA their taking this role currently.
– Seroconversion from HBeAg to anti-HBe indicates remission, however, active viral replication can still occurs due to mutation inhibit or decrease HBeAg.
HBcAg; (intracellular) not present in the serum.
Anti-HBc (IgM &IgG)
– In acute infection emerges 1–2 weeks after the presence of HBsAg and IgM weaned off after 6 months.
– Anti-HBc IgG; detect in both resolved HBV infection and CHB.
– Isolated anti-HBc can be seen in:
– IgM class during the window period of acute phase.
– Anti-HBs below the cutoff level of detection ( after acute infection).
– HBsAg has diminished to undetectable levels (chronic HBV infection)
Molecular methods for HBV infection
HBV DNA
-Measure directly viral load and indicated replication activity and disease progression.
-Detected early (1 mo), peaked at (3 mo) then gradually diminishes in chronic infection or disappears at the recovery.
– Real-time PCR; standard method to detect and quantify HBV DNA.
HBV genotyping:
– It has high genetic heterogeneity, 10 genotype A-J.
– Genotyping predict disease progression and treatment response.
– Genotype A and D leads to higher rate of chronicity than B,C
– Genotype C is risk for perinatal infection , cirrhosis and HCC.
– Response to interferon therapy better in type A&B than C&D.
-Tenofovir response less in type B&C.
Diagnosis of hepatitis B infection
– Acute hepatitis B: is a clinical diagnosis with HBsAg + symptoms+ high aminotransferases. anti-HBc IgM and HBV DNA is present.
– Chronic infection; the persistence of HBsAg for > 6 months.
– Past HBV infection; both anti-HBs and IgG anti-HBc.
– Occult HBV infection; persistence of low HBV DNA without detectable HBsAg.
-Clinical importance of occult HBV infection:
– Viral transmission; via transfusion, organ transplantation or hemodialysis.
– Reactivation of HBV infection in immunocompromised
– Accelerate liver injury and lead to hepatic fibrosis (
– Risk factor for HCC.
-Tests for occult HBV infection are indicated in cryptogenic liver disease, in considering immunosuppression therapy, and in SOT donors,
Level of evidence: 5 narrative review.
Thankyou, well done
Introduction:
Serological markers of HBV infection:
Molecular methods for HBV infection:
HBV genotyping:
Diagnosis of HBV infection:
Very well written summary
Thank you Prof.
Diagnosis of hepatitis B
HBsAg
Anti-HBs
HBcAg
HBV DNA
I make note of your interpretation of virology reports in relation to timings of test and viral load pertaining to HBV infections
I like your detailed well-structured summary.
Introduction:
Hepatitis B virus (HBV) belongs to the Hepadnaviridae family and has a diameter of 30-42 nm consisting of a lipid envelope containing HBsAg that surrounds an inner nucleocapsid composed of hepatitis B core antigen (HBcAg) complexed with virally encoded polymerase and the viral DNA genome. HBV infection is a common cause of chronic liver diseases worldwide, the mode of transmission mainly through reproductive, sexual and vertical routes.
Serological markers for HBV infection:
There are five serological markers including (HBsAg), (HBeAg), (anti-HBs), (anti-HBe) and (anti-HBc), which are valuable for the diagnosis and monitoring of hepatitis B infection and vaccination efficacy. Of these markers HBsAg is considered as a serological marker of HBV infection; it appears early in the serum within 1 to 10 weeks .Anti-HBs may confer long-term immunity only in some cases .
HBV DNA assays have replaced HBeAg and anti-HBe seroconversion to measure infectivity and viral replication .
HBcAg is an intracellular presence in infected hepatocytes, hence it is not detected in patients serum and emerges as IgM and IgG 1-2 weeks after the presence of HBsAg.
Usually anti-HBc IgM disappears after 6 months of acute infection ,and its presence after that indicates resolved HBV infection or CHB. After several years of chronic HBV infection, however, HBsAg has diminished to undetectable levels.
Molecular methods for HBV infection:
HBV DNA is a reliable marker of replication activity and is useful in routine clinical settings to determine patients who need antiviral therapy and monitor them for suitable treatment.
There are two principles of techniques to identify and quantify HBV DNA: signal amplification and target amplification. PCR can detect a wide dynamic range of viral load and is fully automated and does not generate carry-over contamination.
HBV genotyping:
HBV is divided into ten genotypes, A-J, with B and C restricted to Oceania and Asia. Acute infection with genotypes A and D leads to higher rate of chronicity, while C is a risk factor for perinatal infection and severe liver disease. HBV genotyping can be used to predict disease progression and determine appropriate antiviral therapy.
Diagnosis of hepatitis B infection:
Acute hepatitis : Is a clinical diagnosis identified by the detection of HBsAg, symptoms, and high serumaminotransferases.
Chronic HBV infection: Is defined by the coexistence of anti-HBc IgM and HBV DNA.
Occult HBV : Is defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg. HBV DNA testing is widely used to diagnose occult HBV infection, which has some clinical importance due to its transmission via transfusion, solid organ transplantation, or hemodialysis. It may also accelerate liver injury and lead to hepatic fibrosis in patients with chronic hepatitis C infection.
Conclusions:
Diagnosis of HBV infection play an important role in determining acute, chronic and occult cases, establish preventive remedies, and initiate antiviral treatment. Serological markers are used to detect antigens and antibodies, and molecular tests are used to quantify viral load and identify genotypes.
I make note of your interpretation of serological reports pertaining to HBV infections. I like your summary,
Introduction:
Parenteral, sexual, and vertical HBV infection causes most chronic liver disorders globally. 240 million HBV-infected persons are at risk of liver cirrhosis and hepatocellular cancer (HCC). HBsAg prevalence determines HBV endemicity: high, middle, or low.
China, Southeast Asia, Indonesia, and sub-Saharan Africa are highly endemic because over 8% of the population has chronic HBV infection.
South America, South West Asia, and Eastern, and Southern Europe have 2%–7% chronic HBV infection rates.
HBV prevalence is 0.5%–2% in low-endemic regions like North America and Western Europe.
Serological markers for HBV infection:
HBsAg indicates HBV infection.
Serum HBsAg occurs 1–10 weeks after acute HBV exposure. This marker indicates persistent HBV infection after 6 months. Many studies have linked liver cccDNA transcription to blood HBsAg levels. Serum HBsAg values reflect cccDNA distribution during infection stages. HBeAg-positive CHB patients had higher HBsAg titers.
Quantitative HBsAg levels predict interferon response and illness progression in HBeAg-negative CHB patients with normal serum alanine aminotransferase.
HBV molecular techniques:
HBV DNA directly measures viral load, revealing virus replication. It is detected at the early stage of infection (1 month after HBV infection) and grows to a peak level (more than 108 copies/mL) roughly 3 months after HBV exposure. It subsequently progressively decreases in chronic infection or vanishes upon recovery.
HBV-DNA detection has grown in clinical medicine as serologically negative HBV infection (HBeAg-negative CHB and occult HBV infection) has risen. Higher HBV DNA titers indicate replication activity and faster disease progression and HCC development.
HBV DNA testing also helps clinicians identify individuals who require antiviral medication and evaluate their progress.
HBV genotyping:
HBV’s insufficient proofreading of reverse transcriptase causes genetic heterogeneity. Sequence divergence divides HBV into 10 genotypes, A–J, with varied geographic distributions. Genotypes B and C are limited to Oceania and Asia, whereas genotypes A and D are widespread but most frequent in Africa and Europe.
Genotype I is rare in Vietnam, Laos, India, and China, but genotype J is found in Japan and Ryukyu. Asia also sometimes has the genotypes E, F, G, and H.
HBV infection diagnosis:
HBsAg, symptoms, and elevated aminotransferases indicate acute hepatitis B. HBV DNA and anti-HBc IgM are usually found. HBeAg is seen in most acute infections but is not clinically significant. HBsAg persistence beyond 6 months indicates persistent infection. Laboratory tests confirm persistent HBV infection, not clinical symptoms. Anti-HBs and IgG anti-HBc antibodies indicate HBV infection.
Occult HBV infection is low intrahepatic HBV DNA without HBsAg. Isolated anti-HBc without HBsAg or anti-HBs antibodies defines it serologically. As cccDNA persists in the hepatocytes and HBV DNA is sometimes found in the liver but not in the blood, liver HBV DNA testing is the gold standard for detecting occult HBV infection.
Conclusions:
We explained HBV serological and molecular diagnostics in this article. HBV diagnosis begins with serological indicators for antigens and antibodies. Qualitative or quantitative molecular testing validates the diagnosis, measures the viral load, and identifies genotypes.
To prevent and cure acute, chronic, and hidden HBV infections, diagnosis is crucial.
I make note of your interpretation of reports pertaining to HBV infections. I like your summary.I appreciate that you mention genotypes, A–J, with varied geographic distributions.
Diagnosis of hepatitis B:
Introduction;
-HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
-About 240 million people are chronically infected by HBV, so have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC).
-According to HBsAg prevalence, HBV endemicity is divided into three categories; high, intermediate, low.
-China, South East Asia, Indonesia, and sub-Saharan Africa are regarded as highly endemic areas because chronic HBV infection is reported in more than 8% of the population.
HBV genotyping HBV;
-HBV can be divided into ten genotypes, labelled A–J; according to the sequence, variation, and they have distinct geographic distribution.
-Evidences increasingly suggest that the HBV genotyping is significant to predict HBV disease progression and determine appropriate antiviral therapy.
-Acute infection with genotypes A and D leads to higher rate of chronicity than genotypes B and C.
-Genotype C generally is considered as a risk factor for perinatal infection and related to severe liver disease, including cirrhosis and HCC.
-In the interferon therapy, patients with genotypes A and B have better treatment response than genotypes C and D.
-Recent studies reported that patients infected with genotype B or C had a lower opportunity to gain serological response to tenofovir.
Methodology for HBV genotyping;
-It is confirmed using diverse methods: reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNA Chip) and fluorescence polarization assay.
Serological markers for HBV infection;
-It consist of HBsAg , anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG.
-HBsAg is the serological hallmark of HBV infection.
-After an acute exposure to HBV, HBsAg appears in serum within 1 to 10 weeks & persistence of this marker for more than 6 months implies chronic HBV infection.
-HBcAg is an intracellular presence in infected hepatocyte, thus it is not identified in the serum.
-During acute infection, anti-HBc IgM and IgG emerges 1–2 weeks after the presence of HBsAg along with raised serum aminotransferase and symptoms.
-After 6 months of acute infection, anti-HBc IgM wears off & anti-HBc IgG continues to detect in both patients with resolved HBV infection and CHB.
-In patients with acquired immunity through vaccination, anti-HBs is the only serological marker detected in serum.
-Isolated anti-HBc positive can be seen in three conditions.
*First, it can be predominantly seen as IgM class during the window period of acute phase.
*Second, after acute infection had ended, anti-HBs has decreased below the cutoff level of detection.
*Third, after several years of chronic HBV infection, HBsAg has diminished to undetectable levels.
-In these conditions, anti-HBc IgM should be checked in order to assess the possibility of recent HBV exposure.
Molecular methods for HBV infection;
-HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus, and higher titers of HBV DNA are related to the more rapid disease progression and higher incidence of HCC.
-HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
-It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level approximately 3 months after the exposure to HBV and then gradually diminishes in chronic infection or disappears at the recovery from HBV infection.
-HBV DNA assays should be tested in chronic liver disease patients to find out occult HBV infection characterized by existence of detectable HBV DNA without serum HBsAg.
Diagnosis of hepatitis B infection;
Acute hepatitis B infection;
-Identified by the detection of HBsAg, symptoms, high serum aminotransferases.
-Usually anti-HBc IgM can be detected and HBV DNA is present.
-HBeAg can also be identified in most acute phase of infections, but has little clinical importance.
Chronic hepatitis B infection;
-Based on the persistence of HBsAg for more than 6 months.
-Patients with chronic HBV infection are commonly diagnosed by laboratory means but not by clinical presentations.
Past HBV infection;
-Defined by the coexistence of anti-HBs and IgG anti-HBc.
Occult HBV infection;
-Defined by persistence of low level of intrahepatic HBV DNA without detectable HBsAg.
-It is a serological situation defined by the presence of isolated anti-HBc with the absence of HBsAg and anti-HBs antibody.
-The detection of HBV DNA in the liver is the gold standard of diagnosis for occult HBV infection, however, gaining hepatic HBV DNA is difficult in clinical setting since the procedure is invasive.
Importance of diagnosis of occult HBV infection;
-It can be transmitted via transfusion, solid organ transplantation including orthotopic liver transplantation , or hemodialysis.
-Reactivation of HBV infection may occur in patients receiving chemotherapy or immunocompromised state.
-It may accelerate liver injury and lead to hepatic fibrosis in patients with chronic liver disease including chronic hepatitis C infection.
-It appears to be a risk factor for HCC by its carcinogenic effect and by leading to continuous hepatic inflammation and fibrosis.
Tests for occult HBV infection are considered in the following conditions:
*In patients with cryptogenic liver disease, especially when having anti-HBc in serum,
*In patients considering immunosuppression therapy or chemotherapy,
*In solid organ transplantation donors, due to the possibilities for transmission.
Conclusions;
-Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment.
-This Review Articles on Diagnosis of Hepatitis B with (LOE V)
I make note of your interpretation of reports pertaining to HBV infections. I like your summary, level of evidence, analysis and take home messages.
Thanks so much our Prof. I appreciate your note and I really get benefit from your comment.
Introduction
HBV is a virus of Hepadnaviridae family. It has an outer lipid layer with hepatitis B surface antigen (HBsAg) and a capsid core composed of viral genome made of partly double-stranded DNA with 4 overlapping frames including
· S encodes for HBsAg
· pre-C/C for HBeAg and HBcAg
· P for polymerase including reverse transcriptase;
· X that encodes for a transcriptional transactivator factor HBxAg
The covalently closed circular DNA (cccDNA) as a mini chromosome inside the hepatocyte nucleus .
Reverse transcriptase is liable to errors
Methods of HBV infection transfer include parenteral, sexual and vertical routes.
Chronic HBV infection can lead to liver cirrhosis and hepatocellular carcinoma.
High HBV endemic areas as China and Sub saharan Africa.
Intermediate HBV endemic areas as South America, South West Asia
Low HBV endemic areas as North America and Western
Europe .
Serological markers for HBV infection
Include HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc IgM and IgG to identify the viral infection phase and follow antiviral treatment.
HBsAg appears in serum within 1 -10 weeks.
It’s persistence for more than 6 months indicates chronic HBV infection. HBsAg titer monitoring reflects response to interferon and disease progression in HBe Ag negative Chronic Hepatitis B patients with normal serum ALT levels.
Anti-HBs
o on it’s own indicates vaccination
o With HBc IgG indicates past infection
o HBsAg and anti-HBs can be detected in HBsAg positive patients
o indicates carriers of HBV if unable to neutralize the virus .
HBeAg and anti-HBe
o Used to indicate infectivity but now HBV viral assays are used
o HBeAg conversion to anti-HBe signifies remission
HBcAg
is not detected in serum because it is present in infected
hepatocyte.
In acute infection, anti-HBc IgM and IgG appears 1–2 weeks
After HBsAg occurrence with raised liver enzymes.
After 6 months of acute infection, anti-HBc IgM disappears and Anti-HBc IgG continues to be detected in resolved HBV infection and
CHB.
HBsAg
o Can be seen during the window period of acute phase with Ig M.
o after acute infection had ended.
o after several years of chronic HBV infection.
Molecular methods for HBV infection
HBV DNA can be detected 1 month after HBV infection and reaches peak level 3 months after the exposure to HBV and
Then decreases gradually in chronic infection or disappears on recovery .
HBV DNA indicates rapid disease progression and higher incidence of HCC also it can aid in determining those in need of antiviral therapy and monitor them for the response.
HBV DNA is detected and quantified by signal amplification such as hybrid capture and branched DNA technology; and by target amplification such as PCR.
Meanwhile HBV PCR is the standard test .
HBV genotyping
HBV is divided into ten genotypes, labelled A–J differs according to geographic areas.
HBV genotyping can predict HBV disease progression and
Points to proper antiviral therapy. Acute infection with genotypes A and D have higher rate of chronicity than genotypes B and C while C is a risk factor for perinatal infection and severe liver disease.
A and B responds to interferon treatment better than C and D.
B and C have less response to tenofovir.
HBV genotyping is detected by different methods including reverse hybridization, genotype-specific PCR assays, real-time PCR, restriction fragment-length polymorphism, sequence analysis, microarray (DNAChip) and fuorescence polarization assay
HBV infection diagnosis
In acute hepatitis B infection , HBsAg, anti-HBc IgM, HBV DNA and HBeAg can be detected along with clinical diagnosis and elevated liver enzymes.
In chronic infection , HBsAg persists for more than 6 months without clinical manifestations
In past HBV infection, anti-HBs and IgG anti-HBc can be co detected
In occult HBV infection, anti-HBc is detected in serum without HBsAg and antiHBs antibody. HBV DNA in the liver is the gold standard of diagnosis but Real-time PCR for serum HBV DNA detection is a sensitive test to diagnose occult HBV infection.
Occult HBV infection detection is essential due to multiple causes including
– transmission of HBV infection through transfusion, solid organ
Transplantation or haemodialysis .
– reactivation of HBV infection particularly withimmunocompromised status .
-worsen liver injury with co infection of HCV infection
-a risk of HCC
Conclusion
HBV is diagnosed by serological markers to detect antigens and antibodies . In order to diagnose and quantify viral load and to identify genotypes, qualitative or quantitative molecular tests.
Thankyou well done
Introduction
The Hepadnaviridae family of viruses includes the hepatitis B virus (HBV).
It has a diameter of 30-42 nm and is made up of an icosahedral capsid core made of protein and an outer lipid envelope bearing hepatitis B surface antigen (HBsAg).
The viral genome and DNA polymerase with reverse transcriptase activity are both contained in the viral capsid.
Four open reading frames overlap in the circular, partially double-stranded DNA that makes up the HBV genome. (I) S that encodes for surface proteins (HBsAg); (II) pre-C/C for hepatitis B e antigen (HBeAg) and core protein (HBcAg); (III) P for polymerase including reverse transcriptase (IV) X that encodes for a transcriptional transactivator factor (HBxAg). The covalently closed circular DNA (cccDNA) is the transcriptional template of HBV and stays inside the hepatocyte nucleus as a mini-chromosome.
Reverse transcriptase involved in the replication of HBV is error susceptible, the mutation rate is high, observed in retroviruses and RNA viruses.
HBV infection is responsible for the most of chronic liver diseases worldwide and is transmitted through parenteral, sexual and vertical route.
About 240 million people are chronically infected by HBV, so have the risk of developing of liver cirrhosis and hepatocellular carcinoma (HCC) According to HBsAg prevalence, HBV endemicity is divided into three categories; high, intermediate, low.
South east Asia, Indonasia and sub-Saharan Africa are regarded as highly endemic areas because chronic HBV infection is reported in more than 8% of the population .
Europe are grouped as low endemic regions; in these areas, HBV prevalence rates range from 0.5% to 2%
Objectives
We aimed to give informations about HBV
Methods
HBV DNA is a direct measurement of the viral load, which reveals the replication activity of the virus.
It is detectable at the early stage of infection (1 month after HBV infection) and increases up to peak level.
HBV DNA testing is useful in routine clinical setting to determine patients who need antiviral therapy and monitor them for suitable treatment.
There are two principles of techniques to identify and quantify HBV DNA: signal amplification such as hybrid capture and branched DNA technology; target amplification such as polymerase chain reaction (PCR)
Genotype specific PCR High sensitivity, automated systems, easy to perform, suitable for detecting mixed genotype infections High cost.
HBV, hepatitis B virus; RFLP, restriction fragment length polypmorphism; PCR
Findings
Infection is reported in more than 8% of the population
Conclusion
We aimed to give information about HBV serological and molecular diagnosis.
First step of HBV diagnosis is achieved by using serological markers for detecting antigens and antibodies against this virus.
In order to verify first step of diagnosis, to quantify viral load and to identify genotypes, qualitative or quantitative molecular tests are used.
Diagnosis of HBV infection is an important tool to determine acute, chronic and occult cases of infection in order to establish preventive remedies and to initiate antiviral treatment
More specific identification of clinical variants are needed.
Together with a clinical diagnosis and high liver enzyme levels, HBsAg, anti-HBc IgM, HBV DNA, and HBeAg can all be found in acute hepatitis B infections.
HBsAg persists in chronic infection for longer than 6 months without showing any clinical symptoms.
Anti-HBs and IgG anti-HBc can both be found in people with prior HBV infection.
Anti-HBs and IgG anti-HBc can both be found in people with prior HBV infection.
Anti-HBc is seen in serum after occult HBV infection even while HBsAg and antiHBs antibodies are absent.
Nonetheless, Real-time PCR for blood HBV DNA detection is a sensitive test to determine occult HBV infection. HBV DNA in the liver is the gold standard for diagnosis.
Yes, I make note of your interpretation of reports pertaining to HBV infections.
Hepatitis B virus ~ Hepadnaviridae family.
30-40 nm
outer lipid envelope contains Hbsag
icosahedral capsid core – viral genome and DNA polymerase.
viral genome – ds DNA
Contains
S- HbSAg
pre- c/c – Hbe Ag and HbcAg
P- DNA polymerase
x- HBx Ag
HBV infection occurs through
Parenteral
Sexual
Vertical
Serological markers for HBV infection
HbsAg – appears in Serum within 1 to 10 weeks after infection
persists> 6months – Then chronic infection
Higher Hbsag levels in Hbe Ag positive
anti – HBs – long term immunity
only anti- Hbs positive – Post Vaccination
H Be Ag – infectivity
anti HBC IgG IgM-
Past HBV infection – anti Hbs and anti Hbc IgG positive.
Course
Acute Infection
Hbsag +
1-2 weeks later anti HBc Igm IgG +
after 6months anti Hbc IgM wears off
anti Hbc IgG remains+ in CHB and resolved HBV infection.
Some pts
HbsAg –
anti -Hbs-
anti – HBc Ig G +
Isolate Anti – HBc seen in-
During window phase of acute infection
after acute infection has ended , anti- H Bs has decreased
after several years of chronic HBV infection when Hbsag has diminished
So if only anti-HBc +
check anti – HBc IgM and HBV DNA
Molecular methods
HBV DNA – viral load
increases from 1month of infection
peak levels at 3months
gradually diminishes in chronic infection Or disappears at recovery.
HBV DNA by
PCR – standard method
hybrid capture and branched DNA technology.
HBV genotyping
10 genotypes A-J
genotype A and D infection leads to high rate of chronicity.
genotype C – risk factor for perinatal infection
also related to severe liver disease like cirrhosis and HCC.
With interferon treatment genotype Aand B do well.
genotype B and C – lower response to tenofovir.
Diagnosis of Hepatitis B infection
Acute Hep B
Hbs Ag +
symptoms
high aminotransferase
anti HBc Ig M +
HBV DNA present
HbeAg can be positive
chronic Hep B
Hbsag persists > 6 months
Past HBV infection
anti HBs +
anti HBc IgG +
Occult HBV infection
H BS Ag –
low level of intrahepatic HBV DNA on liver biopsy – gold standard
isolated anti- H Bc +
anti – Hbs –
o Can be transmitted by –
Transfusion
hemodialysis
Organ donation
o Reactivation may Occur in those receiving chemotherapy.
o may accelerate liver injury
o Risk factor for HCC
Test for occult HBV infection –
in solid organ Transplantation tation donors
patients with cryptogenic liver disease
in patients considering chemotherapy or immunosuppression