Evaluation and Determination of Quantitative Hepatitis B Surface Antigen Diagnostic Performance in Chronic Hepatitis B Virus-Infected Patients.

Background Hepatitis B virus DNA (HBV-DNA) assessment is recommended for diagnosing and monitoring chronic hepatitis B (CHB) patients. Quantitative hepatitis B surface antigen (qHBsAg) estimation adjunct to HBV-DNA is vital for assessing HBV chronicity and therapeutic prognosis. This study aimed to estimate the qHBsAg and compare its diagnostic performance with that of the HBV-DNA levels in CHB patients from Bangladesh. Methodology A total of 148 CHB patients were enrolled in this study. qHBsAg and hepatitis B e-antigen (HBeAg) were estimated using chemiluminescent and enzyme immunoassays, respectively, and HBV-DNA was quantified using real-time polymerase chain reaction. The parameters and diagnostic performances were analyzed by receiver operating characteristic (ROC) curve analysis. Results The overall levels (mean ± SD) of qHBsAg, HBV-DNA, and alanine aminotransferase (ALT) among the total population (n = 148) were 3.45 ± 0.80 log10 IU/mL, 4.40 ± 2.44 log10 IU/mL, and 86.17 ± 39.06 IU/L, respectively. Significant differences were observed in the levels of both qHBsAg (p = 0.004) and HBV-DNA (p < 0.0001) in cases with HBeAg positivity. qHBsAg levels showed a weak positive correlation with the levels of HBV-DNA and ALT in HBeAg-positive CHB patients, but no such relationship was observed in HBeAg-negative CHB patients. ROC curve analysis showed that the area under the curve for the qHBsAg level to distinguish high HBV-DNA levels (>5 log10 IU/mL) was 0.653 (p = 0.002), which indicated an acceptable diagnostic performance. The best cut-off of qHBsAg for predicting high HBV-DNA levels was 3.469 log10 IU/mL. Conclusions Our results indicated that qHBsAg might be a useful marker for monitoring HBV-DNA in CHB patients throughout treatment and follow-up.

Identification of a novel variant of hepatitis B virus isolated from patient co-infected with hepatitis C virus.

Hepatitis B virus (HBV) is a major public health concern worldwide. Co-infection of hepatitis B patients with other pathogens intensifies the severity of the disease. We report a novel variant of hepatitis B virus (HBV) in Bangladesh isolated from a patient co-infected with hepatitis C virus (HCV) who exhibited liver cirrhosis. From 150 collected plasma samples, we sequenced HBV complete genome from one HBV-HCV co-infected patient. The complete genome was analysed using bioinformatics tools, NCBI BLAST, Geno2Pheno, and SnapGene software. The strain belongs to genotype A and subgenotype A1. Upon analysing the complete genome of this strain, we found a frameshift deletion of 54 nucleotides at the pre-S2 region, a functional regulator of HBV surface protein. Furthermore, we observed a Y126H mutation in the polymerase protein of this strain. This is the first report with such an unusual pre-S deletion event of the HBV genome in an HCV-co-infected patient associated with liver cirrhosis. These findings may inform scientists about genomic modifications in the HBV genome associated with HCV co-infection.

Co-circulation of Multiple SARS-CoV-2 Variants of Concern in Dhaka, Bangladesh, during the Second Wave of the COVID-19 Pandemic.

Thirty partial coding DNA sequences (CDSs) of the spike gene of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were obtained from nasopharyngeal swab samples collected in March to July 2021 in Dhaka, Bangladesh, during the second wave of the coronavirus disease 2019 (COVID-19) pandemic, using Sanger sequencing technology. Sequence analysis showed the presence of multiple WHO-designated variants of concern (VOCs), including Alpha, Beta, Gamma, and Delta, with predominant circulation of Delta variants during that period.

Genome Sequencing of Omicron Variants of SARS-CoV-2 Circulating in Bangladesh during the Third Wave of the COVID-19 Pandemic.

The coding-complete genome sequence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was obtained from 39 nasopharyngeal swab samples collected in January 2022 from Dhaka, Bangladesh, during the 3rd wave of the COVID-19 pandemic, using Illumina MiniSeq sequencing technology. Sequence analysis showed that all of them belonged to the WHO-designated variant of concern (VOC) Omicron. The presence of different sublineages of Omicron was noted, among which sublineage BA.2 (Nextstrain clade 21L) was the most prevalent one.

Genomic Surveillance of SARS-CoV-2 Viruses Collected during the Ending Phase of the First Wave of the COVID-19 Pandemic in Bangladesh.

Mutations, deletions, and the emergence of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may pose a serious health threat. Here, we report the genome sequences of SARS-CoV-2 viruses that were collected from SARS-CoV-2-infected patients during the end phase of the first wave of the COVID-19 pandemic in Dhaka, Bangladesh.

Prevalent HBeAg-negative HBV DNA-positive Chronic Hepatitis B Individuals in Bangladesh.

How to cite this article: Rashed Ul Islam SM, Shahera U, Jahan M, et al. Prevalent HBeAg-negative HBV DNA-positive Chronic Hepatitis B Individuals in Bangladesh. Euroasian J Hepato-Gastroenterol 2021;11(1):49-50.

Challenges in the establishment of a biosafety testing laboratory for COVID-19 in Bangladesh.

At the beginning of the coronavirus disease 2019 (COVID-19) pandemic in Bangladesh, there was a scarcity of ideal biocontainment facilities to detect the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a risk group of 3 organisms. Molecular detection of SARS-CoV-2 must be performed in a BSL-2 laboratory with BSL-3-equivalent infection prevention and control practices. Establishing these facilities within a short timeframe proved to be an enormous challenge, including locating a remote space distant from the university campus to establish a laboratory, motivating the laboratory staff to work with a novel pathogen without any prior experience, allocation of funds for essential equipment and accessories, and arrangement of a safe waste management system for environmental hazard reduction. This report also highlights several limitations, such as the facility's architectural design that did not follow the biosafety guidelines, lack of continuous flow of funds, and an inadequate number of laboratory personnel. This article describes various efforts taken to overcome the challenges during the establishment of this facility that may be adopted to create similar facilities in other regions of the country. Establishing a BSL-2 laboratory with BSL-3-equivalent infection prevention and control practices will aid in the early detection of a large number of cases, thereby isolating persons with COVID-19, limiting the transmission of SARS-CoV-2, and promoting a robust public health response to contain the pandemic.

Role of molecular biomarker human papilloma virus (HPV) E6 oncoprotein in cervical cancer screening.

OBJECTIVE: The Onco E6™ Cervical Test, based on detection of the E6 oncoprotein of HPV 16 and 18 genotypes is evaluated as a screen for the early detection cervical neoplasia in resource-limited countries. METHODS: This prospective study from June 2018 to June 2019 evaluated 235 women aged 21-65 years, who came to Gynaecological Oncology Outpatient Department by VIA, cytology, E6 oncoprotein test and by colposcopy. Screen-positive women by any of the tests or women with suspicious findings were further evaluated by biopsy at colposcopy. The McNemar test was used to compare the performance of E6 oncoprotein test with other screening tests. RESULTS: The E6 oncoprotein positivity rate was 6.8% (n = 16) with 81.25% HPV 16 positive and 18.75% HPV 18 positive. Among VIA positive cases (n = 100), E6 oncoprotein was positive in 9% (p < .001). In histopathology confirmed chronic cervicitis, CIN I, CIN II, CIN III and invasive cervical cancer, E6 test was positive for 2.8%, 4.7%, 25%, 50% and 100% respectively. E6 oncoprotein test had the highest specificity and Positive Predictive Value (PPV; 97% and 75%) compared to VIA (42% and 18%), cytology (95% and 46%) and colposcopy (94% and 59%). Sensitivity of the E6 oncoprotein test for detection of CIN3+ was significantly higher than that of cytology (52% VS 25%) but lower than that of VIA (52% VS 74%). CONCLUSIONS: The HPV E6 oncoprotein test is highly specific and is an effective triage test to reduce colposcopy referrals for the large number of false positive test outcomes seen with VIA.

Response to First-Line Antiretroviral Therapy Among PLHIV from a High-Risk, Low-Prevalence Setting.

The study reports the response of first-line antiretroviral therapy (ART) by assessing CD4 and CD8 T-lymphocyte and viral load (VL) among Bangladeshi people living with HIV (PLHIV). This observational approach was conducted on 100 PLHIVs, grouped into therapy naive (n = 33), therapy initiators with CD4 T-cell count of <350 cells/µL (n = 33), and therapy receivers for >1 year prior to the study period (n = 34). Therapy initiators who continued the study (n = 20) were followed up after 12 and 24 weeks of therapy initiation. The CD4 and CD8 T-lymphocyte count estimation and (VL) were quantified. The mean CD4 T-lymphocyte count was significantly reduced among the therapy initiators in comparison to therapy naive and therapy receivers. Similar findings were observed for CD8 T-lymphocyte count among the study groups. The mean HIV-1 RNA VL among therapy initiators showed a significant decrease after 12 and 24 weeks, and 85% patients in this group obtained undetectable VL status indicating the good therapeutic outcome.

Analysis of the complete genome of hepatitis B virus subgenotype C2 isolate NHB17965 from a HBV infected patient.

The burden of chronic hepatitis B virus (HBV) infections is increasingly detected nowadays. Herein, we report a complete genome of HBV subgenotype C2 (HBV/C2) from a HBV infected patient. Complete genome analysis revealed that the isolated strain was a non-recombinant wild type and had several regular substitutions in the reverse transcriptase domain and small surface proteins of HBV. This study may help clinicians and scientists gain in-depth knowledge on the current substitutions of HBV/C2 genome and to identify potential therapies against HBV infections.

Identification of a novel tri-genotypic recombinant Hepatitis B virus in Bangladesh.

We report a novel tri-genotypic recombinant Hepatitis B virus (HBV) strain circulating in Bangladesh. The strain is recombinant with the genotypes D, C and E, of which, genotype E was not reported before in Bangladesh. Additionally, the complete genome has a frameshift deletion of nine nucleotides from overlapping Surface and Polymerase genes, and a vaccine escape mutation, A128 V, in the surface protein. This is the first report with such unusual recombination event responsible for rapid liver cirrhosis in a 13 year old patient in Bangladesh. This report may alert the clinicians to take the measure to prevent an upcoming outbreak of recombinant HBV.

Complete Genome Sequence of a Circulating Hepatitis B Virus Genotype C Strain Isolated from a Chronically Infected Patient Identified at an Outdoor Hospital in Bangladesh.

Hepatitis B virus (HBV) causes significant global health problems despite the presence of a potential vaccine. HBV chronic cases are increasing rapidly in developing countries like Bangladesh. Here, we report the complete genome sequence of an HBV genotype C strain isolated from a chronic patient identified at an outdoor hospital section.

Molecular characterization of hepatitis B virus in Bangladesh reveals a highly recombinant population.

The natural history and treatment outcome of hepatitis B viruses (HBV) infection is largely dependent on genotype, subgenotype, and the presence or absence of virulence associated mutations. We have studied the prevalence of genotype and subgenotype as well as virulence and drug resistance associated mutations and prevalence of recombinant among HBV from Bangladesh. A prospective cross-sectional study was conducted among treatment naïve chronic HBV patients attending at Bangabandhu Sheikh Mujib Medical University, Dhaka, Bangladesh for HBV viral load assessment between June and August 2015. Systematical selected 50% of HBV DNA positive patients (every second patient) were enrolled. Biochemical and serological markers for HBV infection and whole genome sequencing (WGS) was performed on virus positive sample. Genotype, subgenotype, virulence, nucleos(t)ide analogue (NA) resistance (NAr) mutations, and the prevalence of recombinant isolates were determined. Among 114 HBV DNA positive patients, 57 were enrolled in the study and 53 HBV WGS were generated for downstream analysis. Overall, 38% (22/57) and 62% (35/57) of patients had acute and chronic HBV infections, respectively. The prevalence of genotypes A, C, and D was 18.9% (10/53), 45.3% (24/53), and 35.8% (19/53), respectively. Among genotype A, C and D isolates subgenotype A1 (90%; 9/10), C1 (87.5%; 21/24) and D2 (78.9%; 15/19) predominates. The acute infection, virulence associated mutations, and viral load was higher in the genotype D isolates. Evidence of recombination was identified in 22.6% (12/53) of the HBV isolates including 20.0% (2/10), and 16.7% (4/24) and 31.6% (6/19) of genotype A, C and D isolates, respectively. The prevalence of recombination was higher in chronic HVB patients (32.2%; 10/31 versus 9.1%; 2/22); p<0.05. NAr mutations were identified in 47.2% (25/53) of the isolates including 33.9% novel mutations (18/53). HBV genotype C and D predominated in this population in Bangladesh; a comparatively high prevalence of recombinant HBV are circulating in this setting.

Anti-HBc Screening of Blood Donors in Bangladesh: Relevance to Containment of HBV Propagation.

OBJECTIVES: To avoid further transmission of hepatitis B virus (HBV) infection, blood is tested for hepatitis B surface antigen (HBsAg) before transfusion. However, post-transfusion hepatitis B has been detected in clinics after transfusion of HBsAg-negative blood. The study presented here was undertaken to assess if HBsAg-negative blood is free from HBV or not. METHODS: Sera were collected from 398 blood donors who were negative for HBsAg. Out of 398 blood samples, antibody to hepatitis B core antigen (ant-HBc) was detected in 82 sera samples. HBV DNA was evaluated in HBsAg-negative, anti-HBc-positive sera. HBsAg, hepatitis B e antigen (HBeAg), antibody to HBeAg (anti-HBe), and anti-HBc in the sera were measured by an enzyme-linked immunosorbent assay (ELISA). HBV DNA was quantified by a real time polymerase chain reaction (PCR). RESULTS: Out of 82 HBsAg-negative, anti-HBc-positive sera samples, HBV DNA were detected in the sera of 7 voluntary blood donors. Out of these 7 subjects, all were negative for HBeAg. The levels of ALT were more than 30 IU/L in 6 of 7 HBVDNA-positive subjects and it was above upper limit of normal (>42 IU/ml) in one subject. CONCLUSIONS: The present recommendation about blood transfusion of HBsAg-negative blood system is not capable of blocking HBV transmission to blood recipients. Although advanced countries have adopted nucleic acid testing (NAT) for preventing HBV transmission, developing countries may apply anti-HBc testing and ALT estimation before blood transmission.

IP-10, p53, and Foxp3 Expression in Hepatocytes of Chronic Hepatitis B Patients with Cirrhosis and Hepatocellular Carcinoma.

AIM: Elucidating differences in gene expression may be useful in understanding the molecular pathogenesis and for developing specific markers for the outcome of hepatitis B virus (HBV) infection. In the present study, expressions of host gene interferon gamma-inducible protein (IP-10), p53, and Foxp3 were studied in hepatocytes of patients with chronic HBV infection to determine a possible link between selected host gene expression and the outcome of HBV infection. MATERIALS AND METHODS: The study was conducted in 60 patients with chronic HBV infection and they were divided into four groups: HBV-positive cirrhosis (n = 15), HBV-negative cirrhosis (n = 15), HBV-positive hepatocellular carcinoma (HCC) (n = 15) and HBV-negative HCC (n = 15). Total messenger ribonucleic acid (mRNA) extraction was done followed by complementary deoxyribonucleic acid (cDNA) synthesis, and finally gene expression was performed using real-time polymerase chain reaction (PCR) technique. RESULTS: IP-10 and p53 gene expressions were lower in HBV-positive cirrhosis, and Foxp3 gene expression was upregulated in HBV-positive cirrhosis in comparison to HBV-negative cirrhosis. The expressions of all the three genes were upregulated among HBV-positive HCC in comparison to HBV-negative HCC. The expression of IP-10, p53, and Foxp3 genes was upregulated in HBV-positive HCC in comparison to HBV-positive cirrhosis. CONCLUSION: This study indicates that there are variations in the expression of the selected genes among cirrhosis and HCC patients with or without HBV. All the three selected genes were more or less upregulated in HBV-positive HCC patients, but only Foxp3 expression was upregulated in HBV-positive cirrhosis. These three particular genes may have a role in the molecular pathogenesis and clinical outcome of HBV-positive cirrhosis and HCC patients. These aspects need further evaluation by studies with larger numbers of cirrhosis and HCC patients. HOW TO CITE THIS ARTICLE: Shahera U, Munshi S, Jahan M, Nessa A, Alam S, Tabassum S. IP-10, p53, and Foxp3 Expression in Hepatocytes of Chronic Hepatitis B Patients with Cirrhosis and Hepatocellular Carcinoma. Euroasian J Hepato-Gastroenterol 2016;6(2):149-153.

Novel insights into immunotherapy for hepatitis B patients.

The possible use of immunotherapy for hepatitis B has emerged for two major reasons: (1) chronic hepatitis B (CHB) is an immune-mediated pathological condition, and (2) commercially available antiviral drugs are of limited efficacy. Although various immunomodulatory agents have been used to treat patients with CHB during the last three decades, there is currently no consensus among physicians and hepatologists regarding the suitability of immunotherapy for patients with CHB. However, new insights into immunotherapy for CHB have emerged; these may facilitate design of effective and tolerable immunotherapy regimens for these patients. This review provides a comprehensive overview of immunotherapy for CHB.

Evaluation of a Rapid One-step Real-time PCR Method as a High-throughput Screening for Quantification of Hepatitis B Virus DNA in a Resource-limited Setting.

Virological monitoring is the best predictor for the management of chronic hepatitis B virus (HBV) infections. Consequently, it is important to use the most efficient, rapid and cost-effective testing systems for HBV DNA quantification. The present study compared the performance characteristics of a one-step HBV polymerase chain reaction (PCR) vs the two-step HBV PCR method for quantification of HBV DNA from clinical samples. A total of 100 samples consisting of 85 randomly selected samples from patients with chronic hepatitis B (CHB) and 15 samples from apparently healthy individuals were enrolled in this study. Of the 85 CHB clinical samples tested, HBV DNA was detected from 81% samples by one-step PCR method with median HBV DNA viral load (VL) of 7.50 × 103 lU/ml. In contrast, 72% samples were detected by the two-step PCR system with median HBV DNA of 3.71 × 103 lU/ml. The one-step method showed strong linear correlation with two-step PCR method (r = 0.89; p < 0.0001). Both methods showed good agreement at Bland-Altman plot, with a mean difference of 0.61 log10 IU/ml and limits of agreement of -1.82 to 3.03 log10 IU/ml. The intra-assay and interassay coefficients of variation (CV%) of plasma samples (4-7 log10 IU/ml) for the one-step PCR method ranged between 0.33 to 0.59 and 0.28 to 0.48 respectively, thus demonstrating a high level of concordance between the two methods. Moreover, elimination of the DNA extraction step in the one-step PCR kit allowed time-efficient and significant labor and cost savings for the quantification of HBV DNA in a resource limited setting. HOW TO CITE THIS ARTICLE: Rashed-Ul Islam SM, Jahan M, Tabassum S. Evaluation of a Rapid One-step Real-time PCR Method as a High-throughput Screening for Quantification of Hepatitis B Virus DNA in a Resource-limited Setting. Euroasian J Hepato-Gastroenterol 2015;5(1):11-15.

Hepatitis B Core Antigen in Hepatocytes of Chronic Hepatitis B: Comparison between Indirect Immunofluorescence and Immunoperoxidase Method.

BACKGROUND: Hepatitis B virus (HBV) infection has many faces. Precore and core promoter mutants resemble inactive carrier status. The identification of hepatitis B core antigen (HBcAg) in hepatocytes may have variable clinical significance. The present study was undertaken to detect HBcAg in chronic hepatitis B (CHB) patients and to assess the efficacy of detection system by indirect immunofluorescence (IIF) and indirect immunoperoxidase (IIP). MATERIALS AND METHODS: The study was done in 70 chronic HBV-infected patients. Out of 70 patients, eight (11.4%) were hepatitis B e antigen (HBeAg) positive and 62 (88.57%) were HBeAg negative. Hepatitis B core antigen was detected by indirect immunofluorescence (IIF) and indirect immunoperoxidase (IIP) methods in liver tissue. RESULTS: All HBeAg positive patients expressed HBcAg by both IIF and IIP methods. Out of 62 patients with HBeAg-negative CHB, HBcAg was detected by IIF in 55 (88.7%) patients and by IIP in 51 (82.26%) patients. A positive relation among viral load and HBcAg detection was also found. This was more evident in the case of HBeAg negative patients and showed a positive relation with HBV DNA levels. CONCLUSION: Hepatitis B core antigen can be detected using the IIF from formalin fixed paraffin block preparation and also by IIP method. This seems to reflect the magnitudes of HBV replication in CHB. HOW TO CITE THIS ARTICLE: Raihan R, Tabassum S, Al-Mahtab M, Nessa A, Jahan M, Kabir CMS, Kamal M, Aguilar JC. Hepatitis B Core Antigen in Hepatocytes of Chronic Hepatitis B: Comparison between Indirect Immunofluorescence and Immunoperoxidase Method. Euroasian J Hepato-Gastroenterol 2015;5(1):7-10.

Role of the HPV DNA test in follow-up of treated cervical intraepithelial neoplasia in Bangladesh.

BACKGROUND: Cervical cancer is a major public health problem in Bangladesh. Persistence of high risk human papillomavirus (HRHPV) influences the progression of the disease, with an important role in follow- up for cervical intraepithelial neoplasia (CIN). OBJECTIVE: To establish application of high risk HPV DNA test in the follow-up of women after treatment of CIN. MATERIALS AND METHODS: This cross-sectional and hospital based study was carried out among 145 CIN treated women during the previous six months to three years at the colposcopy clinic of Bangabandhu Sheikh Mujib Medical University, Dhaka, between January 2011 and June 2012. Pap smear and HPV samples were collected and colposcopy was performed to find out the persistence of the disease. Cervical samples obtained were tested for HPV DNA using the Hybrid Capture II (HC-II) test. A cervical biopsy was collected whenever necessary. The results were compared to assess the efficacy of different methods during follow up such as Pap smear, HPV test and colposcopy. RESULTS: Mean age of the recruited women (n=145) was 33.6 (± 7.6), mean age of marriage was 16.8 (±2.9) and mean age of 1st delivery was 18.8 (±3.5) years. More than half had high grade CIN before treatment and 115 (79.3%) women were managed by LEEP and 20.7% were managed by cold coagulation. Among the 145 treated women, 139 were negative for HPV DNA and six of them (4.1%) were HPV positive. Sensitivity of Pap smear (40.0) and HPV DNA test (40.0) was poor, but specificity was quite satisfactory (>93.0) for all the tests. CONCLUSIONS: The high risk HPV DNA test can be an effective method of identifying residual disease. It can be added to colposcopy and this should be applied to all treated women attending for their first or second post-treatment follow-up visit at 6 months to one year, irrespective of the grade of treated CIN.

Epidemiology of Hepatitis E Virus in an Urban Community in Dhaka City.

INTRODUCTION: Hepatitis E virus (HEV) is endemic in Bangladesh and sporadic and epidemic outbreaks of acute hepatitis E occur in this country almost regularly. Although the real magnitude of HEV prevalence has not been documented in Bangladesh, HEV infections and HEV-related acute hepatitis of Bangladeshi origin have been reported from different parts of the world. METHODS: The study was conducted in Mirpur area of Dhaka city, which is a major residential area of the capital of Bangladesh. Three hundred adults were randomly included in the study. None had any history of jaundice or complains of liver diseases. RESULTS: The study revealed 30% prevalence of HEV in this population. The prevalence increased with age, but there was no gender difference. CONCLUSION: HEV is a highly prevalent disease in Bangladesh as elsewhere in the developing world. Since there is no specific treatment for HEV, improvement of personal hygiene and ensuring supply of safe food and drinking water remain most important approach to sustain the virus.How to cite this article: Rahman S, Mahtab MA, Jahan M, Tabassum S, Akbar SMF. Epidemiology of Hepatitis E Virus in an Urban Community in Dhaka City. Euroasian J Hepato-Gastroenterol 2014; 4(1):4-6.