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Nucleotide identity and variability among different Pakistani hepatitis C virus isolates.

ABSTRACT: BACKGROUND: The variability within the hepatitis C virus (HCV) genome has formed the basis for several genotyping methods and used widely for HCV genotyping worldwide. AIM: The aim of the present study was to determine percent nucleotide identity and variability in HCV isolates prevalent in different geographical regions of Pakistan. METHODS: Sequencing analysis of the 5'noncoding region (5'-NCR) of 100 HCV RNA-positive patients representing all the four provinces of Pakistan were carried out using ABI PRISM 3100 Genetic Analyzer. RESULTS: The results showed that type 3 is the predominant genotypes circulating in Pakistan, with an overall prevalence of 50%. Types 1 and 4 viruses were 9% and 6% respectively. The overall nucleotide similarity among different Pakistani isolates was 92.50% +/- 0.50%. Pakistani isolates from different areas showed 7.5% +/- 0.50% nucleotide variability in 5'NCR region. The percent nucleotide identity (PNI) was 98.11% +/- 0.50% within Pakistani type 1 sequences, 98.10% +/- 0.60% for type 3 sequences, and 99.80% +/- 0.20% for type 4 sequences. The PNI between different genotypes was 93.90% +/- 0.20% for type 1 and type 3, 94.80% +/- 0.12% for type 1 and type 4, and 94.40% +/- 0.22% for type 3 and type 4. CONCLUSION: Genotype 3 is the most prevalent HCV genotype in Pakistan. Minimum and maximum percent nucleotide divergences were noted between genotype 1 and 4 and 1 and 3 respectively.

SUBMITTER: Idrees M

PROVIDER: S-EPMC2737312 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Nucleotide identity and variability among different Pakistani hepatitis C virus isolates.

Idrees Muhammad M Butt Sadia S Awan Zunaira Z Aftab Mahwish M Khubaib Bushra B Rehman Irshad-Ur IU Akram Madiha M Manzoor Sobia S Akbar Haji H Rafiqe Shazia S Riazuddin Sheikh S

Virology journal 20090824

<h4>Background</h4>The variability within the hepatitis C virus (HCV) genome has formed the basis for several genotyping methods and used widely for HCV genotyping worldwide.<h4>Aim</h4>The aim of the present study was to determine percent nucleotide identity and variability in HCV isolates prevalent in different geographical regions of Pakistan.<h4>Methods</h4>Sequencing analysis of the 5'noncoding region (5'-NCR) of 100 HCV RNA-positive patients representing all the four provinces of Pakistan ...[more]

PMID: 19698187

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Project description:BACKGROUND:Hepatitis C virus (HCV) is the cause of high morbidity and mortality worldwide, inflicting around one million people in Pakistan alone. The HCV genomic RNA harbors conserved structural elements that are indispensable for its replication. The 3' untranslated region (UTR) contains several of these elements essentially involved in regulating the major steps of the viral life cycle. OBJECTIVES:Differences in regulatory elements of HCV may contribute towards differential infectivity of local isolates. The present study explicates sequence analysis and secondary structure prediction of HCV 3'UTR region of subtype 3a from Pakistan to characterize this particular region. PATIENTS AND METHODS:HCV 3'UTR region was amplified, cloned and sequenced from five different patients. Sequence and structural analysis was performed and phylogenetic analysis was carried out using the 3'UTR sequence reported in NCBI nucleotide data base (http://www.ncbi.nlm.nih.gov/nuccore) by other studies. RESULTS:Sequence analysis of the amplified fragment from five patients indicated that the 3'UTR is composed of 214-235 nts. Its sequence contains a type-specific variable region followed by a poly U/UC region and a highly conserved X-tail of 98 nts. The variable region reported here has 26 nts and one stem loop at the secondary structure that differentiate it from HCV genotype 1a ( GT1a) 3'UTR which contains additional 14 nts and two stem loops. The poly U/UC region varied in length (100-79 nts) and nucleotide sequence within the Pakistani isolates, and among different genotypes. Some substitutions found in the X-tail do not affect secondary structure of this element suggesting that this region might play an important role in replication, stabilization and packaging of HCV genome. Additionally, U residues are not present at the end of the X-tail in Pakistani 3a isolates as otherwise reported for the variants of genotype 1b. CONCLUSIONS:Sequence and structural diversity of the 3'UTR variable region and Poly U/UC region found in the local isolates indicate specificity in the regulating elements of 3'UTR that might be associated with differential replication efficacy of the HCV Pakistani isolates. The study necessitates functional characterization of these regulating elements to elucidate variable viral efficiency and pathogenicity associated with inter-geographical isolates.

Project description:BackgroundSince it is currently not possible to eradicate hepatitis B virus (HBV) infection with existing treatments, research continues to uncover new therapeutic strategies. HBV core protein, encoded by the HBV core gene (HBC), intervenes in both structural and functional processes, and is a key protein in the HBV life cycle. For this reason, both the protein and the gene could be valuable targets for new therapeutic and diagnostic strategies. Moreover, alterations in the protein sequence could serve as potential markers of disease progression.AimTo detect, by next-generation sequencing, HBC hyper-conserved regions that could potentially be prognostic factors and targets for new therapies.MethodsThirty-eight of 45 patients with chronic HBV initially selected were included and grouped according to liver disease stage [chronic hepatitis B infection without liver damage (CHB, n = 16), liver cirrhosis (LC, n = 5), and hepatocellular carcinoma (HCC, n = 17)]. HBV DNA was extracted from patients' plasma. A region between nucleotide (nt) 1863 and 2483, which includes HBC, was amplified and analyzed by next-generation sequencing (Illumina MiSeq platform). Sequences were genotyped by distance-based discriminant analysis. General and intergroup nt and amino acid (aa) conservation was determined by sliding window analysis. The presence of nt insertion and deletions and/or aa substitutions in the different groups was determined by aligning the sequences with genotype-specific consensus sequences.ResultsThree nt (nt 1900-1929, 2249-2284, 2364-2398) and 2 aa (aa 117-120, 159-167) hyper-conserved regions were shared by all the clinical groups. All groups showed a similar pattern of conservation, except for five nt regions (nt 1946-1992, 2060-2095, 2145-2175, 2230-2250, 2270-2293) and one aa region (aa 140-160), where CHB and LC, respectively, were less conserved (P < 0.05). Some group-specific conserved regions were also observed at both nt (2306-2334 in CHB and 1935-1976 and 2402-2435 in LC) and aa (between aa 98-103 in CHB and 28-30 and 51-54 in LC) levels. No differences in insertion and deletions frequencies were observed. An aa substitution (P79Q) was observed in the HCC group with a median (interquartile range) frequency of 15.82 (0-78.88) vs 0 (0-0) in the other groups (P < 0.05 vs CHB group).ConclusionThe differentially conserved HBC and HBV core protein regions and the P79Q substitution could be involved in disease progression. The hyper-conserved regions detected could be targets for future therapeutic and diagnostic strategies.

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Nucleotide identity and variability among different Pakistani hepatitis C virus isolates.

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Nucleotide identity and variability among different Pakistani hepatitis C virus isolates.

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