scholarly journals A CRISPR-Cas12b–Based Platform for Ultrasensitive, Rapid, and Highly Specific Detection of Hepatitis B Virus Genotypes B and C in Clinical Application

Author(s):  
Xu Chen ◽  
Yan Tan ◽  
Shuoshi Wang ◽  
Xueli Wu ◽  
Rui Liu ◽  
...  

Hepatitis B virus (HBV) is one of the most dangerous and prevalent agents that causes acute and chronic liver diseases in humans. Genotyping plays an important role in determining clinical outcomes and response to antiviral treatment in HBV–infected patients. Here, we first devised a CRISPR–based testing platform, termed “CRISPR-HBV,” for ultrasensitive, highly specific, and rapid detection of two major HBV genotypes (HBV-B and HBV-C) in clinical application. The CRISPR-HBV employed multiple cross displacement amplification (MCDA) for rapid preamplification and then Cas12b–based detection for decoding the targets. Finally, the detection result was read out with real-time fluorescence and a lateral flow biosensor. The sensitivity of CRISPR-HBV was 10 copies per test. The specificity was one hundred percent, and no cross reactions were observed in other HBV genotypes and pathogens. The whole detection process, including DNA template extraction (15 min), preamplification reaction of MCDA (30 min at 65°C), CRISPR-Cas12b–based detection (5 min at 37°C), and results readout (∼2 min), could be completed within 1 h. The feasibility of the CRISPR-HBV assay for genotyping HBV-B and -C as successfully validated with clinical samples. Hence, the CRISPR-HBV assay has remarkable potential to develop a point-of-care testing for identifying and distinguishing HBV genotypes B and C in clinical settings, especially in resource-scarcity countries.

2017 ◽  
Vol 89 ◽  
pp. 14-21 ◽  
Author(s):  
Vincent Thibault ◽  
Annabelle Servant-Delmas ◽  
Thoai Duong Ly ◽  
Anne-Marie Roque-Afonso ◽  
Syria Laperche

2019 ◽  
Vol 11 (03) ◽  
pp. 240-243 ◽  
Author(s):  
Iram Masood ◽  
Usman Waheed ◽  
Muhammad Arshad ◽  
Muhammad Saeed ◽  
Ahmad Farooq ◽  
...  

Abstract BACKGROUND: Hepatitis B virus (HBV) is a major causative agent of early, severe and prolonged liver infection that subsequently leads to cirrhosis of liver and hepatocellular carcinoma. The aim of this study was to evaluate the molecular epidemiology of hepatitis B virus (HBV) genotypes and comparison of serological assay performance versus polymerase chain reaction (PCR) in HBV screening. METHODS: Blood samples of 8517 healthy blood donors were collected during the period of January to June 2017 from Blood Bank of Shaheed Zulfiqar Ali Bhutto Medical University, Islamabad. Samples were screened for HBsAg assay using technique of chemiluminescence immunoassay. PCR of positive samples was carried out using already reported genotype-specific primers by Naito et al. (2001). The results were confirmed by visualizing genotype bands. RESULTS: The study confirmed the presence of HBV in 2.5% of blood donors, and PCR confirmed the presence of HBV-DNA in 92 samples. The genotyping was done by PCR using type-specific primer sequences. PCR was dogged to check six genotypes, i.e., A, B, C, D, E, and F. The results of this study show high levels of Genotype D is this region, i.e., 52.17% with less dominating Genotype C, which is 16.30% with decreasing ratio of Genotype E (14.13%), Genotype A and B (9.78%), and mixed D + E (2.17%). The presence of coinfection is found at lowest rate. Due to the high percentage of HBV/D, it is concluded that D genotype is common in our population. CONCLUSION: The most prevalent HBV genotype in ICT region was genotype D, which is responsible for liver cirrhosis and hepatocellular carcinoma. Efficacy of drugs varies with variation in genotypes of hepatitis B virus and also with geographical distribution.


2010 ◽  
Vol 2010 ◽  
pp. 1-6 ◽  
Author(s):  
Gaetano Scotto ◽  
Domenico Martinelli ◽  
Rocco Di Tullio ◽  
Vincenzina Fazio

Background/aims. This study aims to determine the distribution and clinical features of HBV-genotypes in a population of immigrants affected by HBV-infection. Methods. Between 01/2003 and 03/2009, 1623 immigrants were tested for HBV-infection. Biochemical and virological activities were determined in HBsAg-positive patients; HBV-genotypes were determined, by the INNO-LiPA HBV Genotyping, in the subjects with HBV DNA detectable. In every patient we evaluated the stage and classified the infection as inactive carrier, mild or moderate/severe chronic hepatitis, cirrhosis, and/or HCC. Results. Among the tested subjects, 191 (11.7%) resulted HBsAg-positive, and in 144/191 (75.4%) serum HBV-DNA was detectable. The genotype distribution was as follows: 45,13% genotype E, 18,1% genotype D, 15,3% genotype B, 13,2% genotype C, 4,9% genotype A, 3,5% mixed genotypes (A–D). The evaluation of liver disease degree showed that 24.6% patients were inactive carriers of HBV infection, 19.4% presented a immunotolerance phase, 34.5% had mild chronic hepatitis, 13.6% had a moderate/severe chronic hepatitis, 6.3% had cirrhosis, and 1.6% presented HCC. Conclusions. Our study evidences a high prevalence of HBV-infection in immigrants, and the potentiality of migratory flow in the introduction of genotype non-D hepatitis B virus. The Hepatitis B virus genotypes presented significant differences in epidemiological and clinical characteristics.


2019 ◽  
Author(s):  
Yi Yi Kyaw ◽  
Aye Aye Lwin ◽  
Khin Saw Aye ◽  
Hlaing Myat Thu ◽  
Moh Moh Htun ◽  
...  

Abstract Background Hepatitis B virus (HBV) infection is a severe health concern worldwide. HBV is a DNA virus with a rapid rate of mutation. Based on the heterogeneity of the HBV nucleotide sequence, the HBV strains are divided into ten genotypes, A to J, with a characteristic geographical distribution. Identifying and tracking the changes of HBV genotypes is important in epidemiological and transmission studies, predicting the risk for the development of severe liver disease and response to antiviral treatment. The present study was conducted to detect HBV genotypes and sub-genotypes in general population of different states and regions in Myanmar. Methods A total of 5,547 general adult population who residing at seven states, seven regions and Nay Pyi Taw Union Territory were screened for Hepatitis B Surface antigen (HBsAg) by Immunochromatgraphic test (ICT) in 2015. Of 353 HBsAg positive samples, HBV DNA were detected by using polymerase chain reactions (PCR) targeting the DNA sequences encoding the Pre-S region. A total of 153 PCR positive samples were preceded for genotyping by partial genome sequencing of both directions. The resulting sequences were then edited, aligned and compared with reference sequences using National Centre for Biotechnology Information (NCBI) web based genotyping tool. Results Three HBV genotypes; HBV/ C, HBV/ D and HBV/ B were detected in Myanmar, in which genotype HBV/ C (66.7%) was the most prevalent genotype followed by HBV/ D (32%) and HBV/ B (1.3%) respectively. Sub-genotyping revealed a total of 7 sub-genotypes within genotypes B, C and D: two (B4 and B5) in HBV/ B, three (C1, C5 and C7) in HBV/C and two (D3 and D6) in HBV/ D. Conclusion Genotype HBV/C, sub-genotype C1 was the most predominant genotype distributed in all states and regions of Myanmar. This study was first report on Nation- wide distribution of HBV genotype and sub-genotypes in Myanmar and the findings will be a huge support for hepatitis disease surveillance programme which is the one of the National Priority Diseases in Myanmar.


2020 ◽  
Author(s):  
Yi Yi Kyaw ◽  
Aye Aye Lwin ◽  
Khin Saw Aye ◽  
Hlaing Myat Thu ◽  
Moh Moh Htun ◽  
...  

Abstract Background Hepatitis B virus (HBV) infection is a severe health concern worldwide. HBV is a DNA virus with a rapid rate of mutation. Based on the heterogeneity of the HBV nucleotide sequence, the HBV strains are divided into ten genotypes, A to J, with a characteristic geographical distribution. Identifying and tracking the changes of HBV genotypes is important in epidemiological and transmission studies, predicting the risk for the development of severe liver disease and response to antiviral treatment. The present study was conducted to detect HBV genotypes and sub-genotypes in general population of different states and regions in Myanmar. Methods A total of 5,547 general adult population who residing at seven states, seven regions and Nay Pyi Taw Union Territory were screened for Hepatitis B Surface antigen (HBsAg) by Immunochromatgraphic test (ICT) in 2015. Of 353 HBsAg positive samples, HBV DNA were detected by using polymerase chain reactions (PCR) targeting the DNA sequences encoding the Pre-S region. A total of 153 PCR positive samples were preceded for genotyping by partial genome sequencing of both directions. The resulting sequences were then edited, aligned and compared with reference sequences using National Centre for Biotechnology Information (NCBI) web based genotyping tool. Results Three HBV genotypes; HBV/ C, HBV/ D and HBV/ B were detected in Myanmar, in which genotype HBV/ C (66.7%) was the most prevalent genotype followed by HBV/ D (32%) and HBV/ B (1.3%) respectively. Sub-genotyping revealed a total of 7 sub-genotypes within genotypes B, C and D: two (B4 and B5) in HBV/ B, three (C1, C5 and C7) in HBV/C and two (D3 and D6) in HBV/ D. Conclusion Genotype HBV/C, sub-genotype C1 was the most predominant genotype distributed in all states and regions of Myanmar. This study was first report on Nation- wide distribution of HBV genotype and sub-genotypes in Myanmar and the findings will be a huge support for hepatitis disease surveillance programme which is the one of the National Priority Diseases in Myanmar.


Author(s):  
Eman Mohammad Al-Shawkany ◽  
Abdul-Al-Raoof Mohammad AlShawkany ◽  
Saleh S Bahaj ◽  
Arwa Mohammed Othman ◽  
Hassan A Al-Shamahy ◽  
...  

Background and aims: Hepatitis B virus infection is a significant public health crisis global. Hepatitis B virus genotyping is an important tool in epidemiological studies to determine the category and extent of treatment and to predict the outcome of chronic infections, for instance hepatocellular carcinoma and cirrhosis. The study designed to determine the prevalence of hepatitis B virus genotypes among Yemeni patients with chronic hepatitis B (CHB) and to evaluate some of the associated risk factors. Methods: Fifty patients (38 males, 12 females) with chronic hepatitis B from Al-Thawra Modern General Hospital, Al-Kuwait University Hospital, and AL-Gomhoria Hospital were included. HBV DNA was first detected by conventional PCR then HBV genotypes were determined using nested and multiplex PCR.  Results: Mixed HBV genotypes (A+B+C+D+E), (A+B+C+D+E+F), and  (A+B+C+D)  were found to be the most prevalent (60 %), it is followed by genotype D (16 %), genotype B (16%) and genotype A (8%), whereas C, E, and F genotype were not found individually among the study population. Blood transfusion was associated with mixed infection (χ2=13.06; p= 0.005). Conclusions: In assumption, this study demonstrates the general prevalence of hepatitis B virus genotypes among HBV-infected Yemeni hepatitis B patients who request medical consideration in a hospital. In mono-genotype HBV infection, genotype B and D were the most prevalent genotypes. In HBV mixed genotype infection, the A/B/C/D/E genotype was the most prevalent in the study area. In the future, based on genotype, clinical trials and treatment regimens must be individually assumed to efficiently manage chronic HBV infection. To this end, a prospective nationwide population study of HBV genotype spreading and clinical outcomes is suggested.                    Peer Review History: Received: 15 May 2021; Revised: 11 June; Accepted: 27 June, Available online: 15 July 2021 Academic Editor:  Ahmad Najib, Universitas Muslim Indonesia, Makassar, Indonesia, [email protected] UJPR follows the most transparent and toughest ‘Advanced OPEN peer review’ system. The identity of the authors and, reviewers will be known to each other. This transparent process will help to eradicate any possible malicious/purposeful interference by any person (publishing staff, reviewer, editor, author, etc) during peer review. As a result of this unique system, all reviewers will get their due recognition and respect, once their names are published in the papers. We expect that, by publishing peer review reports with published papers, will be helpful to many authors for drafting their article according to the specifications. Auhors will remove any error of their article and they will improve their article(s) according to the previous reports displayed with published article(s). The main purpose of it is ‘to improve the quality of a candidate manuscript’. Our reviewers check the ‘strength and weakness of a manuscript honestly’. There will increase in the perfection, and transparency.  Received file:                Reviewer's Comments: Average Peer review marks at initial stage: 6.5/10 Average Peer review marks at publication stage: 7.5/10 Reviewer(s) detail: Dr. Salfarina Ramli,  Department of Pharmacology and Pharmaceutical Chemistry, Faculty of Pharmacy, Universiti Teknologi MARA, 42300 Puncak Alam, Selangor, Malaysia. [email protected]   Dr. Asia Selman Abdullah, University of Basrah, Iraq, [email protected] Similar Articles: EXPLOSION OF HEPATITIS B AND C VIRUSES AMONG HEMODIALYSIS PATIENTS AS A RESULT OF HEMODIALYSIS CRISIS IN YEMEN PREVALENCE AND GENOTYPING OF HEPATITIS C VIRUS IN HEMODIALYSIS PATIENTS AND EVALUATION OF HCV-CORE ANTIGEN TEST IN SCREENING PATIENTS FOR DIALYSIS IN SANA'A CITY, YEMEN PREVALENCE OF HBV AND HCV; AND THEIR ASSOCIATED RISK FACTORS AMONG PUBLIC HEALTH CENTER CLEANERS AT SELECTED PUBLIC HEALTH CENTERS IN SANA'A CITY-YEMEN


1998 ◽  
Vol 36 (11) ◽  
pp. 3362-3365 ◽  
Author(s):  
V. A. Mbayed ◽  
J. L. López ◽  
P. F. S. Telenta ◽  
G. Palacios ◽  
I. Badía ◽  
...  

Differences in pathogenesis and the probability of becoming a chronic carrier depend on the age at which hepatitis B virus (HBV) infection is acquired, ranging from 82% in infants less than 6 months of age to 15 to 30% in older children. HBV genotypes from 22 pediatric patients from two areas that differ in prevalence have been determined. Phylogenetic analysis shows a clear difference between the genotype distribution in Buenos Aires, a low-prevalence area, and that found in Gualeguay, Entre Rı́os, a high-prevalence area. While the analysis allocated the sequences in the Buenos Aires group to genotypes A (36%), D (9%), and F (55%), the Gualeguay group presented exclusively genotype A isolates with very low nucleotide divergence, which suggests a strong founder viral population. The high prevalence of genotype F in the Buenos Aires group and its high intragroup heterogeneity agree with the American origin of this genotype.


2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Chun Shen Lim ◽  
Vitina Sozzi ◽  
Margaret Littlejohn ◽  
Lilly K.W. Yuen ◽  
Nadia Warner ◽  
...  

Hepatitis B virus (HBV) is a major human pathogen that causes liver diseases. The main HBV RNAs are unspliced transcripts that encode the key viral proteins. Recent studies have shown that some of the HBV spliced transcript isoforms are predictive of liver cancer, yet the roles of these spliced transcripts remain elusive. Furthermore, there are nine major HBV genotypes common in different regions of the world, these genotypes may express different spliced transcript isoforms. To systematically study the HBV splice variants, we transfected human hepatoma cells, Huh7, with four HBV genotypes (A2, B2, C2 and D3), followed by deep RNA-sequencing. We found that 13–28 % of HBV RNAs were splice variants, which were reproducibly detected across independent biological replicates. These comprised 6 novel and 10 previously identified splice variants. In particular, a novel, singly spliced transcript was detected in genotypes A2 and D3 at high levels. The biological relevance of these splice variants was supported by their identification in HBV-positive liver biopsy and serum samples, and in HBV-infected primary human hepatocytes. Interestingly the levels of HBV splice variants varied across the genotypes, but the spliced pregenomic RNA SP1 and SP9 were the two most abundant splice variants. Counterintuitively, these singly spliced SP1 and SP9 variants had a suboptimal 5′ splice site, supporting the idea that splicing of HBV RNAs is tightly controlled by the viral post-transcriptional regulatory RNA element.


2009 ◽  
Vol 81 (10) ◽  
pp. 1716-1720 ◽  
Author(s):  
Andreas Erhardt ◽  
Thomas Göbel ◽  
Amei Ludwig ◽  
George K.K. Lau ◽  
Patrick Marcellin ◽  
...  

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