Detectable HBV DNA Research Articles

A quantitative real time polymerase chain reaction for detection of HBV covalently closed circular DNA in livers of the HBV infected patients

To establish and optimize a sensitive and specific quantitative real-time polymerase chain reaction (PCR) method for detection of hepatitis B virus covalently closed circular DNA (HBV cccDNA) in liver tissue. Specific primers and probes were designed to detect HBV DNA (tDNA) and cccDNA. A series of plasmids (3.44 × 10(0) - 3.44 × 10(9) copies/µl) containing a full double-stranded copies of HBV genome (genotype C) were used to establish the standard curve of real-time PCR. Liver samples of 33 patients with HBV related hepatocellular carcinoma (HCC), 13 Chronic hepatitis B patients (CHB) and 10 non-HBV patients were collected to verify the sensitivity and specificity of the assay. A fraction of extracted DNA was digested with a Plasmid-Safe ATP-dependent Dnase (PSAD) for HBV cccDNA detection and the remaining was used for tDNA and β-globin detection. The amount (copies/cell) of HBV cccDNA and tDNA were measured by a real-time PCR, using β-globin housekeeping gene as a quantitation standard. The standard curves of real-time PCR with a linear range of 3.44 × 10(0) to 3.44 × 10(9) copies/µl were established for detecting HBV cccDNA and tDNA, and both of the lowest detection limits of HBV cccDNA and tDNA were 3.44 × 10(0) copies/µl. The lowest quantitation levels of HBV cccDNA in liver tissues tested in 33 HBV related HCC patients and 13 CHB patients were 0.003 copies/cell and 0.031 copies/cell, respectively. HBV cccDNA and tDNA in liver tissue of 10 non-HBV patient appeared to be negative. The true positive rate was increasing through the digestion of HBV DNA by PSAD, and the analytic specificity of cccDNA detection improved by 7.24 × 10(2) times. Liver tissues of 2 patients were retested 5 times in the PCR for detecting cccDNA and the coefficient of variations on cycle threshold (Ct) were between 0.224% - 0.609%. A highly sensitive and specific quantitative real time PCR method for the detection of HBV cccDNA in liver tissue was established and could be used for clinical and epidemiological studies.

High Rate of Recurrent Hepatitis B Viremia Following Treatment-Induced Hepatitis B E Antigen (HBeAg) Seroconversion

Purpose: In patients with HBeAgpositive chronic hepatitis B (CHB), one of the primary endpoints of therapy is HBeAg seroconversion; however, there is inconsistent data on the durability of HBeAg seroconversion following consolidation therapy. Our goal is to investigate the rate of recurrent HBV viremia in CHB patients after HBeAg seroconversion. Methods: We retrospectively studied 88 consecutive CHB patients who achieved treatment-induced HBeAg seroconversion among the 458 HBeAg-positive patients treated with various antiviral regimens at 3 GI and liver clinics in the U.S. between 3/1998 and 9/2010. HBeAg seroconversion was defined as loss of HBeAg and development of hepatitis B e antibody (anti-HBe). Recurrent HBV viremia was defined as reappearance of detectable HBV DNA (>100 IU/mL) from nadir in two consecutive laboratory tests. Results: Patients were stratified into two groups: Group I 49 patients who remained on therapy, and Group II 39 patients who discontinued therapy following HBeAg seroconversion. In both groups, the majority of patients were Asian (94-95%) and male (62-69%). Antiviral medications in which patients achieved HBeAg seroconversion included: lamivudine (23%), adefovir (34%), entecavir (36%), tenofovir (4%), adefovir+lamivudine (1%), and entecavir+tenofovir (2%). Median treatment duration before HBeAg seroconversion was similar in Group I and Group II (18 [1-86] vs. 21 [5-63] months, p=0.99). At the time of HBeAg seroconversion, compared to Group I, Group II was younger (37±10 vs. 41±12 years), had lower mean HBV DNA levels (0.27±0.78 vs. 0.95±1.69 log10 IU/mL), and median ALT levels (24 [8-60] vs. 30 [593] U/L). In Group II, the median duration of consolidation therapy before treatment discontinuation was 12 months (range, 1-55), in which 10 completed <6 months, 10 completed 6-11 months, and 19 completed ≥12 months. At time of treatment discontinuation, all patients in Group II had undetectable HBV DNA. The rate of recurrent HBV viremia was significantly higher in Group II compared to Group I (90% vs. 0%, p<0.0001). After a median of 3 (1-42) months off therapy, the mean HBV DNA level detected at time of recurrent HBV viremia was 4.11±1.92 log10 IU/mL. Conclusion: The majority of patients (90%) who discontinued therapy after achieving HBeAg seroconversion with undetectable HBV DNA by PCR experienced recurrent HBV viremia. Patients who remained on therapy achieved and maintained undetectable level of HBV DNA. Patients should be monitored closely for recurrent HBVwhen therapy is discontinued, despite achieving HBeAg seroconversion.

Markers of Microbial Translocation Are Elevated in Hepatitis B (HBV) and Hepatitis C (HCV) Infection; A Window Into Biology and Surrogate Markers for Detecting Disease Progression

Purpose: In patients with HBeAgpositive chronic hepatitis B (CHB), one of the primary endpoints of therapy is HBeAg seroconversion; however, there is inconsistent data on the durability of HBeAg seroconversion following consolidation therapy. Our goal is to investigate the rate of recurrent HBV viremia in CHB patients after HBeAg seroconversion. Methods: We retrospectively studied 88 consecutive CHB patients who achieved treatment-induced HBeAg seroconversion among the 458 HBeAg-positive patients treated with various antiviral regimens at 3 GI and liver clinics in the U.S. between 3/1998 and 9/2010. HBeAg seroconversion was defined as loss of HBeAg and development of hepatitis B e antibody (anti-HBe). Recurrent HBV viremia was defined as reappearance of detectable HBV DNA (>100 IU/mL) from nadir in two consecutive laboratory tests. Results: Patients were stratified into two groups: Group I 49 patients who remained on therapy, and Group II 39 patients who discontinued therapy following HBeAg seroconversion. In both groups, the majority of patients were Asian (94-95%) and male (62-69%). Antiviral medications in which patients achieved HBeAg seroconversion included: lamivudine (23%), adefovir (34%), entecavir (36%), tenofovir (4%), adefovir+lamivudine (1%), and entecavir+tenofovir (2%). Median treatment duration before HBeAg seroconversion was similar in Group I and Group II (18 [1-86] vs. 21 [5-63] months, p=0.99). At the time of HBeAg seroconversion, compared to Group I, Group II was younger (37±10 vs. 41±12 years), had lower mean HBV DNA levels (0.27±0.78 vs. 0.95±1.69 log10 IU/mL), and median ALT levels (24 [8-60] vs. 30 [593] U/L). In Group II, the median duration of consolidation therapy before treatment discontinuation was 12 months (range, 1-55), in which 10 completed <6 months, 10 completed 6-11 months, and 19 completed ≥12 months. At time of treatment discontinuation, all patients in Group II had undetectable HBV DNA. The rate of recurrent HBV viremia was significantly higher in Group II compared to Group I (90% vs. 0%, p<0.0001). After a median of 3 (1-42) months off therapy, the mean HBV DNA level detected at time of recurrent HBV viremia was 4.11±1.92 log10 IU/mL. Conclusion: The majority of patients (90%) who discontinued therapy after achieving HBeAg seroconversion with undetectable HBV DNA by PCR experienced recurrent HBV viremia. Patients who remained on therapy achieved and maintained undetectable level of HBV DNA. Patients should be monitored closely for recurrent HBVwhen therapy is discontinued, despite achieving HBeAg seroconversion.

Évolutions technologiques en qualification biologique du don et leur impact sur le risque résiduel transfusionnel

During the two last decades, the risk of viral transfusion transmission for some infectious agents (HIV, HCV, HBV and HTLV) has been markedly reduced by improved donor screening, improvements of serological assays and the implementation of minipool nucleic acid testing for HIV-1 and HCV viruses. However, implementation during the year 2010 of nucleic acid testing for the detection of HIV RNA, HCV RNA and HBV DNA in a single triplex assay may provide additional safety, especially after acute infection during the window period. New Procleix(®) Tigris(®) technology (Novartis) allow the French blood screening laboratories to answer their actual requirements in terms of security and throughput and to implement nucleic acid testing in case of emergent risks requiring a direct detection of viruses. Furthermore, Tigris(®) is a fully automated system with high level of security during the analytical process, reducing the number of invalid or non-available results observed with the previous semi-automated technologies. Moreover, renewal of material by fully automated and secure systems, especially for the critical step of sample pipeting, may provide additional safety in blood screening laboratories.

The correlation and clinical significance of HBV infection makers

Objective To explore the internal relations of HBV markers and its clinical significance.Methods HBV Pre S1-Ag、HBV-M(HBsAg、HBsAb、HBeAg、HBeAb、HBcAb)and HBV DNA of 106 patients were detected by ELISA and FQ-PCR respectively.Results The total positive rates of Pre-S1 antigen and HBV DNA in 106 cases were 81.1%and 84.9%respectively.In41 HBeAg-positive cases.Pre S1-Ag and HBV DNA detection rate was 95.1%.90.2%.And HBeAg-negative 2 groups Pre S1-Ag and HBV DNA detection rate was also higher.Conclusion HBV Per S1-Ag Was a very valuable serum marker in terms of direct HBV replication alone can not HBeAg-positive to determine whether the replication of the virus. Key words: Biological narkers; DNA virus Infections

The estimation of traceability and uncertainty of measurement on the result of HBV DNA with different detecting system

Objective To study the uncertainty and traceability of HBV DNA assays and discuss the comparability of among different detection systems. Methods Different detecting systems were used to detect HBV DNA using the national standard substance as control The uncertainty of the was evaluated referring Guidelines for estimating and reporting measurement uncerTAinty of chemical test results of NATA The were traced back to the national standard substance. According to the CLSI document EP9-A2, the were analyzed and subjected to bias estimation with the t(0.05sv) √u2b1+ u2b2 as the criterion clinically accepted to investigate the comparability of different detecting systems. Results The means (-y) measured by 3 HBV DNA assay systems were 6.15,5.88,and 6.31 lg(kIU/L) respectively. Except system A,both the biases of system B and C had statistical significance (all P ＜ 0. 05) and expanded uncertainty of three detection systems was varied, but the difference was within the maximum acceptable range (± 0. 5) of the external quality assessment by National Center for Clinical Laboratory. Being traceable to national standard substance, the of HBV DNA of the three detecting systems were (5.45 ± 1.23), (5.55 ± 1.32) and (5.42 ± 1.25) lg(kIU/L), respectively.There was significant difference among three systems (F = 5.63, P ＜ 0. 05). Comparing system A and B,there was significant difference in statistic (q = 5. 12, P ＜ 0. 05) and the difference between system B and C also had statistically significant (q = 6. 85, P ＜ 0. 05), but the between system A and C had no statistical difference (q = 1.85,P ＞ 0. 05). Among these three systems, the difference of any two detection systems had no statistical significance (all P ＞ 0. 05). It showed that system bias was acceptable in clinical application and the between different systems were comparable. Conclusions It is necessary to estimate the uncertainty and traceability when comparing the HBV DNA assay among the different labs. It also needs to estimate the bias of different systems and evaluate the clinical acceptability to ensure the accuracy and comparability of the results. Key words: Uncertainty; Hepatitis B virus; DNA,viral; Polymerase chain reaction

Occult HBV infection among Egyptian hepatocellular carcinoma patients

BackgroundOccult HBV infection accelerates the progression of liver fibrosis, cirrhosis, and finally leading to hepatocellular carcinoma (HCC). This study analyzed the occult HBV-genotypes in HCC patients.MethodsTo achieve our objective, matched serum and tissue samples were collected from 40 HCC patients. Three sets of primers were used for the HBV-DNA detection by nested-PCR, which cover the HBV-genome; Core, Surface and X genes. Genotyping system based on PCR using type-specific primers was applied on HBV-DNA positive samples.ResultsIntrahepatic occult HBV-DNA was detected in 62.5%, whereas; Serum occult HBV-DNA were detected in only 22.5% of HCC patients. In patients' positive for both anti-HBs and anti-HBc, 10% had occult HBV in serum. In serologically negative HCV patients, 63% had intrahepatic HBV-DNA, and 21% had HBV-DNA in serum samples. HBV-genotype D (32%) and B (24%) attributed predominantly to intrahepatic HBV infections in HCC patients, whereas HBV-genotype A (4%) and C (8%) infections were the least observed.ConclusionThis is the first study to show the genotypes of occult HBV infection in HCC Patients. We suggest that B or D may influence the outcome of HBV infection which may lead to the development of HCC.

Safety of complete and sustained prophylaxis withdrawal in patients liver-transplanted for HBV-related cirrhosis at low risk of HBV recurrence

HBV reactivation after liver transplantation may be related to persistence of covalently closed circular (ccc) DNA. We investigated the safety of HBV prophylaxis withdrawal in selected HBV transplanted patients. Thirty patients transplanted 64-195months earlier (23 males, median age 56yrs), HBsAg-positive, HBeAg, and HBV-DNA negative at transplant (43% HCV/HDV co-infected), with undetectable intrahepatic total and ccc-DNA were enrolled. All patients underwent HBIg withdrawal and continued lamivudine with monthly HBsAg and HBV-DNA monitoring and sequential liver biopsies. Those with confirmed intrahepatic total and ccc-DNA undetectability 24weeks after stopping HBIg, also underwent lamivudine withdrawal and were followed-up without prophylaxis. Twenty-five patients did not exhibit signs of HBV recurrence after prophylaxis withdrawal (median follow-up 28.7months, range 22-42). Five patients became HBsAg-positive: one early after HBIg withdrawal, the other four after HBIG and lamivudine withdrawal. None of these patients experienced clinically relevant events. In the first patient, HBIg were reinstituted with prompt HBsAg negativization. Of the other four, one remained HBsAg-positive with detectable HBV-DNA and mild ALT elevation and was successfully treated with tenofovir. In the remaining three, HBsAg positivity was transient and followed by anti-HBs seroconversion, thus no antiviral treatment was needed. Patients with undetectable HBV viremia at transplant and no evidence of intrahepatic total and cccDNA may safely undergo cautious weaning of prophylaxis, showing low rate of HBV recurrence after a 2 year follow-up. Undetectability of intrahepatic ccc-DNA may help to identify patients at low-risk of recurrence, yet studies with longer follow-up are needed.

Role of occult hepatitis B virus in chronic hepatitis C patients with flare of liver enzymes

Background Occult HBV infection is defined by detection of HBV DNA in the serum or liver tissue of patients who test negative for HBsAg. The prevalence of occult HBV is higher in hepatitis C virus (HCV) positive patients than HCV negative patients and may have an impact on their clinical outcome. In this study, we evaluated the role of occult hepatitis B virus infection in chronic hepatitis C patients with ALT flare. Methods Sixty HBsAg negative patients with chronic hepatitis C virus infection were included. Patients were divided into 2 groups according to their ALT level: 30 patients with normal or slightly high ALT and 30 patients with ALT flare (≥ 5 times normal values). Patients in both groups were examined for the detection of anti-HBs, anti-HBc IgM, and anti-HBc IgG. HBV DNA was detected using semi-nested PCR technique. Results In patients with normal or slightly high ALT, HBV DNA was detected in 4 (13.3%) patients, while in those with ALT flare, HBV DNA was detected in 19 (63.3%) patients (p < 0.001). No association was found between the presence of HBV DNA and various serology markers of HBV infection. Conclusion Presence of occult hepatitis B, with its added deleterious effect, must always be considered in chronic hepatitis C patients especially those with flare in liver enzymes; HBsAg should not be used alone for the diagnosis of HBV infection.

Detection of hepatitis B virus genotype A3 and primary drug resistance mutations in African immigrants with chronic hepatitis B in Spain

Universal vaccination and antiviral therapy have reduced chronic hepatitis B virus (HBV) in natives in the Western world. However, immigration from high HBV endemic areas continues to maintain a relatively stable prevalence of chronic hepatitis B in most developed countries. All foreigners attending a referral infectious diseases department in Madrid, Spain, from January 2007 to December 2008, were evaluated for serum HBV surface antigen (HBsAg). Positive cases underwent further virological characterization. A total of 1718 foreigners were examined, of whom 1322 (77%) were sub-Saharan Africans. Serum HBsAg was positive in 121 (7%), HIV in 135 (7.9%) and hepatitis C virus antibodies in 212 (12.3%). HBV subgenotype A3, which so far had only been reported in people originating from Cameroon, was found in nearly half (14/29) of the tested specimens with detectable serum HBV-DNA. Interestingly, the lamivudine resistance mutation rtM204V was found in two Africans (6.9%), one infected with HBV-A3 and the other with HBV-E. Lack of prior exposure to antiviral therapy in these two patients was confirmed retrospectively. Circulation of uncommon HBV variants, including strains with primary drug resistance, may follow large immigrant flows from HBV endemic regions to Western countries. Close surveillance of this population is warranted, as early diagnosis and early antiviral therapy may reduce transmission and prevent clinical complications.

Occult hepatitis B infection in egyptian chronic hepatitis C patients: prevalence, impact on pegylated interferon/ribavirin therapy

BackgroundChronic HCV infection combined with occult hepatitis B infection has been associated with liver enzymes flare, advanced hepatic fibrosis and cirrhosis, poor response to standard interferon-α, and increased risk of HCC. This study aimed to elucidate the prevalence of occult hepatitis B infection in Egyptian chronic HCV patients, and to clarify its role in non-response of those patients to pegylated interferon/ribavirin therapy. This study enrolled 155 consecutive chronic HCV patients under pegylated interferon/ribavirin therapy. All patients were exposed to clinical assessment, biochemical, histological and virological examinations. HBV parameters (HBV DNA, anti-HBc, anti-HBs) and patients' response status to the combination therapy were determined.ResultsIn this study, occult hepatitis B infection occurs in 3.9% of Egyptian chronic HCV patients; tends to affect younger age patients, associated with higher base line HCV viral load, less hepatic fibrosis than monoinfected patients. This occult hepatitis B infection is not a statistically significant cause of non-response to pegylated interferon/ribavirin therapy. Anti-HBs was not associated with any biochemical, histological or virological abnormalities in those patients, contrary to low response rate to therapy and higher HCV viral load that was observed with anti-HBc.ConclusionsDetection of HBV DNA in HBsAg negative chronic HCV patients plays a non significant role in non-response of Egyptian patients to pegylated interferon/ribavirin therapy.

Improvement of the analytical sensitivity of domestic HBV DNA kit based on magnetic beads nucleic acid extraction method

Objective To establish a method of nucleic acid extraction and enrichment based on magnetic nanoparticle as medium for elevating the analytical sensitivity of domestic HBV real-time PCR kit and detection of the trace amount HBV DNA. Methods After receiving antiviral treatment, the serum samples of 50 hepatitis B patients with HBV DNA concentration ≤1×104 IU/ml were collected. The WHO HBV DNA calibrator was used as the standard material. Nanometer magnetic beads were used to adsorb and enrich the HBV nucleic acid and increase the concentration of the extracted HBV nucleic acid template. Compared with Roche HBV DNA detection reagent and four domestic reagent with conventional nucleic acid extraction and detection method, the improvement effect of this method on domestic nucleic acid detection reagent was evaluated. Results After application of nanometer magnetic extraction method to domestic regent, the analytical sensitivities of the domestic reagent reached 10 and 50 IU/ml, respectively,which was about the same detection level to 12 IU/ml of the imported Roche reagent. The positive rates of the detection of serum trace amount HBV DNA of hepatitis B patients with four kinds of domestic extraction reagent were 64% ( 32 ), 56% ( 28 ), 62% ( 31 ) and 58% (29), respectively. There were significant statistical differences between Roche reagent and four domestic extraction reagent kits(x2 = 7. 895, 12. 698,9. 013 and 11. 416 ,P ＜0. 05 ). With nanometer magnetic extraction method combined with domestic reagent kits, the detection rates were 88% (44), 88% (44), 88% (44) and 86% (43) ,respectively. There was no significant difference compared with the imported Roche reagent (x2 = 0. 000, 0. 000, 0. 000 and 0. 088,P ＞0. 05). Moreover, when the HBV nucleic acid concentration was 101-103 IU/ml, the logarithm value of viral nucleic acid concentration was in reverse correlation to Ct value, but the correlation decreased in the concentration range of 103-106 IU/ml. Conclusions The nucleic acid extraction method based on magnetic nanoparticle as medium can significantly improve the analytical sensitivity of domestic HBV DNA detection reagent, which can be used to monitor the trace amounts HBV DNA in the sera of the hepatitis B patients. Key words: Hepatitis B virus; Magnetic nanoparticle; Nucleic acid test; Analysis sensitivity

Are isolated anti-HBc blood donors in high risk group? The detection of HBV DNA in isolated anti-HBc cases with nucleic acid amplification test (NAT) based on transcription-mediated amplification (TMA) and HBV discrimination

AimHepatitis B virus (HBV) can be transmitted by blood transfusions even so using serological tests having high sensitivity and specificity. We aimed to detect HBV DNA in isolated Anti-HBc donors and show if they have transfusion risk or not. MethodWe investigated Anti-Hbc and Anti-HBs in serum samples of 12858 HBs-Ag negative blood donors who were applied to the Turkish Redcrescent between June 2007 and October 2008 by the Micro ELISA kit (Hepanostica ultra HBsAg, Bio Meriux, France). Anti-HBc and Anti-Hbs positive cases were omitted. We used Procleix ultrio (Chiro, USA) test kit (Chiron Tigris automated instrument was used) based TMA (Transcription-Mediated Amplification) for NAT study in Anti-HBc positive and Anti-HBs negative plasma samples. The discrimination of HBV in NAT positive samples were performed by Procleix Discrimination (Chiro, USA) test. Results2748 (21.4%) Anti-HBc seropositivity were detected in 12852 HBsAg(−) serum samples. 23.5% Anti-HBs negativity was detected in 2748 Anti-HBc positive serum samples. On the other hand, 5.1% isolated Anti-Hbc positivity [HBsAg(−), Anti-HBc(+), Anti-HBs(−)] were detected in 12852 HBsAg(−) serum samples. 0.091% and 0.047% HBV-DNA positivity were detected in isolated Anti-HBc positive plasma samples and HBsAg(−) plasma samples, respectively. ConclusionAs a result, even we have detected one HBV transmission in every 2142 blood transfusion by HBsAg screening tests; we suggest that it is not necessary to add additional tests to detect isolated Anti-HBc for routine purposes in Blood Banking. The reasons are higher negativity rates (99%) of isolated Anti-HBc serum samples and the rejection of blood donors with Anti-HBc positivity.

Virologic Relapse following Treatment Induced Hepatitis B e Antigen Seroconversion in Chronic Hepatitis B (CHB) Patients

Purpose: The treatment goal for hepatitis B e antigen (HBeAg) positive patients is HBeAg seroconversion; however, this endpoint may not be durable following discontinuation of antiviral therapy. Our goal was to examine the rate of relapse in these patients. Methods: We retrospectively studied 52 consecutive CHB patients who achieved treatment-induced seroconversion (out of 396 patients treated for a median duration of 18 months). Our cohort consists of patients who maintained on therapy (n=28) and who discontinued therapy (n=24) between January 2001 and November 2009 at 2 community U.S. GI clinics. HBeAg seroconversion is defined as conversion of HBeAg-positive to HBeAg-negative and development of hepatitis B e antibody (anti-HBe). Virologic relapse is defined as reappearance of detectable HBV DNA by PCR testing (>60-100 IU/mL). Results: All patients were Asians and predominantly male (67%). Antiviral medication in which patients achieved seroconversion were lamivudine (17%), adefovir dipivoxil (40%), entecavir (35%), tenofovir (4%), and entecavir+tenofovir (4%). Median treatment duration after seroconversion to treatment discontinuation was 12 (1-41) months, in which all patients had undetectable serum HBV DNA (<60-100 IU/mL) and normal ALT (≤40 U/L). Following seroconversion, 71% of patients received another 12 months or more of therapy prior to discontinuation, 17% with another 6-12 months of therapy, and only 12% having less than 6 months of consolidation therapy. At time of seroconversion, patients who were maintained on therapy had significantly higher HBV DNA (1.36±1.97 vs. 0.33±0.86, p=0.03) and median ALT (33 [11-85] vs. 24 [8-60], p=0.04) compared to patients who discontinued therapy. Median age at seroconversion was similar in both cohorts (39 [19-58] vs. 38 [13-68], p=0.53). Virologic relapse occurred in 23 of 24 patients (96%) who discontinued therapy compared to 0 of 28 patients (0%) who maintained on treatment, p<0.0001. For patients who ceased treatment, the time to relapse was 3 (1-42) months. Conclusion: After achieving HBeAg seroconversion, the majority of patients who discontinued antiviral therapy relapsed with reappearance of viremia. Patients should be monitored closely for relapse when therapy is discontinued, even after HBeAg seroconversion.

Prevalence and characteristics of hepatitis B and C virus infections in treatment-naïve HIV-infected patients

In HIV-infected treatment-naïve patients, we analyzed risk factors for either chronic hepatitis B (HBV) infection, occult HBV infection (OHBV) or a positive hepatitis C (HCV) serostatus. A total of 918 patients of the RESINA-cohort in Germany were included in this study. Before initiating antiretroviral therapy, clinical parameters were collected and blood samples were analyzed for antibodies against HIV, HBV and HCV, HBs antigen and viral nucleic acids for HIV and HBV. Present or past HBV infection (i.e. HBsAg and/or anti-HBc) was found in 43.4% of patients. HBsAg was detected in 4.5% (41/918) and HBV DNA in 6.1% (34/554), resulting in OHBV infection in 2.9% (16/554) of patients. OHBV infection could not be ruled out by the presence of anti-HBs (50.1%) or the absence of all HBV seromarkers (25%). A HCV-positive serostatus was associated with the IVDU transmission route, non-African ethnicity, elevated liver parameters (ASL or GGT) and low HIV viral load. Replicative HBV infection and HCV-positive serostatus both correlated with HIV resistance mutations (P=0.001 and P=0.028). HBV and HCV infection are frequent co-infections in HIV treatment-naive patients. These co-infections influence viral evolution, clinical parameters and serological markers. Consequently, HIV patients should routinely be tested for HBV and HCV infection before initiating HIV treatment. OHBV infection constituted almost half of all HBV infections with detectable HBV DNA. Due to a lack of risk factors indicating OHBV infection, HBV diagnosis should not only include serological markers but also the detection of HBV DNA.

Detection of HBV-DNA in Dried Bloodstains on Filter Paper by Nested Polymerase Chain Reaction

To describe the technical performance of nested PCR for identifying hepatitis B viral (HBV) DNA. Hepatitis B viral DNA was extracted from a dried bloodstain on filter paper by a Chelex-100 method. Then the DNA fragment was amplified by nested polymerase chain reaction (PCR). The sensitivity and specificity of this method were also analyzed. The positive rate of the nested PCR-based method was compared with that of the enzyme-linked immunosorbent assays (ELISA) method. The lowest detection limit of the test was 5 copies of HBV DNA per μL by this method. McNemar’s test showed that the difference between the positive rates of these 2 methods was not statistically significant (P=0.289, P>0.05). Hepatitis B viral test results showed a good concordance between these 2 methods (kappa=0.727). A very small amount of the dried blood sample is required for the detection, which could overcome the shortage of the normal ELISA method that requires a relatively large amount of fresh blood samples.

Hepatitis B pre-S1 antigen in the Diagnosis

Objective To study the serum of patients with different disease Hepatitis B antigen pre-S1 (HBVpre-S1) levels in patients with HBV serum markers, HBV-DNA of the clinical relevance, evaluation HB-Vpre-S1 the detection of hepatitis diagnosis value. Methods Used enzyme linked immunosorbent assay (ELISA) were conducted on 268 cases of the five serum HBV serum markers, Pre-S1 test. Automatic biochemical analyzer Lai liver function was detected, anddetected by fluorescence quantitative PCR HBV-DNA quantity. Results 196 patients with chronic hepatitis B (CH) patients, HBeAg (+) group, and HBeAg (-) group, Pre-S1(+)and HBVDNA(+)detection rate and the relative ratio of both high Pre-S1/HBsAg in HBeAg(-) group,the difference was significant,P ＜0. 01 ;and in HBeAg(-) group. HBeAb(-) group and HBeAb(+)group compared to, Pre-S1/HBsAg relative ratio was also significantly higher than HBeAb(+)group,P ＜ 0. 05. In CH patients, mild,moderate and severe among the three groups of Pre-S1 (+)rate, HBVDNA and Pre-S1/HBsAg relative ratio,the difference was not statistically significant, and the relative ratio Pre-S1/HBsAg levels and liver inflammation grade. In the 72 cases of AH patients,Pre-S1/HBsAg the relative ratio and the severity of liver acute injury related to the ratio of AH relative decline in patients with earlier Pre-S1/HBsAg faster, shorter course of disease,better prognosis. Conclusion Pre-S1 can be used as a marker of HBV replication,and Pre-S1/HBsAg the relative ratio of height,in the diagnosis of hepatitis, AH has important prognostic value. Detection Pre-S1 can be combined to complement and improve the HBV less than five detection, particularly for HBeAg (-)or a variation of HBV infection to better reflect the state of viral replication and infectious. Key words: Hepatitis B virus; Before the S1 antigen; Odds ratio; HBV-DNA; Clinical laboratory techniques

The prevalence of “anti‐HBc alone” and HBV DNA detection among anti‐HBc alone in Korea

The "anti-HBc alone" is a frequent serological finding in clinical laboratories, making it difficult to determine whether the HBV infection has resolved. The objectives of this study were to investigate the prevalence of anti-HBc alone and HBV DNA detection (occult HBV infection) among anti-HBc alone, and to describe the demographic and clinical characteristics of anti-HBc alone. A total of 17,677 sera referred from the Health Promotion Center (HPC group, 4,014 sera) as well as all the hospital clinical departments (Patient group, 13,663 sera) were tested for HBs Ag, anti-HBc, and anti-HBs. HBV DNA test using real-time PCR was performed on 230 anti-HBc alone. The prevalence of anti-HBc alone was 8.9%, significantly higher in the Patient group than in the HPC group. The prevalence of anti-HBc was higher in men than women and was increased with age. Very low levels of HBV DNA were found in only 4 (1.7%) out of 230 subjects with anti-HBc alone. They were patients with conditions unrelated to chronic liver disease. Considering the high prevalence of anti-HBc alone, the frequency of occult HBV infection among anti-HBc alone was unexpectedly low. In addition, HBV viral load was low in these patients. Further studies are required to determine the clinical significance and infectivity of anti-HBc alone, in conjunction with very low levels of HBV DNA and to standardize the detection methodology for both anti-HBc alone and HBV DNA.

Main mutations in the hepatitis B virus basic core promoter (A1762T/G1764A) before HBeAg loss are markers that identify patients who will require long‐term treatment

Some patients continue to have detectable HBV-DNA levels with liver disease progression after hepatitis B e antigen (HBeAg) loss. It is important to identify these patients, candidates for long-term treatment. To evaluate hepatitis B virus (HBV) genotype and the main mutations in the basic core promoter (BCP, A1762T/G1764A) and precore (G1896A) sequences as markers of persistent HBV-DNA after HBeAg loss. We analysed 60 serum samples from 20 Caucasian, HBeAg-positive, chronic hepatitis B patients, who lost HBeAg and were followed-up longitudinally. HBV genotype and precore and BCP mutations were determined before, at the time of, and after HBeAg loss. After HBeAg loss, eight (40%) patients continued to have undetectable HBV-DNA and 12 (60%) had persistent HBV-DNA (median level 4.7 log(10) copies/mL). The presence of BCP mutations prior to therapy was the only variable associated with persistently detectable viraemia (P = 0.017). Four patients with genotype A and no mutations in the BCP region experienced hepatitis B surface antigen (HBsAg) loss after a mean period of 35 months from baseline. Main BCP mutations in HBeAg-positive patients are useful markers to identify patients who will not have sustained virological suppression after HBeAg loss and therapy discontinuation and could benefit from long-term treatment.

Occult Hepatitis B : A Hidden Threat

Occult hepatitis B (OHB) is defined as the presence of HBV-DNA in liver with or without detectable HBV DNA in serum in patients who are HBsAg negative in currently available assays. Diagnosis of occult HBV infections requires sensitive HBV-DNA PCR assay. Though reports of occult hepatitis B infections are coming from almost every corner of the world, the exact prevalence of this hidden menace is still not know. Several aspects of occult hepatitis B infection are still not resolved including its clinical significance relating to the molecular basis, risk of transmission, reactivation and progression to chronic liver disease. In this review article, we will try to explore the current concepts and clinical implications of occult hepatitis B virus infection.&#x0D; TAJ 2010; 23(1): 109-113