Hepatitis C Virus Core Region Research Articles

Impact of amino acid substitutions in hepatitis C virus core region on the severe oxidative stress

Hepatitis C virus (HCV) infection is a major cause of chronic liver disease, leading to liver steatosis, fibrosis, and hepatocellular carcinoma (HCC). Despite the accumulation of clinical data showing the impact of amino acid substitutions at positions 70 (R70Q/H) and/or 91 (L91M) in the HCV core protein in progressive liver diseases, including HCC, the underlying mechanisms have not been elucidated.We analyzed 72 liver biopsy specimens from patients with chronic HCV genotype 1b (HCV-1b) infection prior to antiviral treatment. Levels of 8-hydroxy-2′-deoxyguanosine (8-OHdG) and nuclear factor erythroid 2-related factor 2 (NRF2) in the nucleus were quantified using liver tissue immunohistochemistry. The effects of amino acid substitutions in the HCV core region on hepatocellular oxidative stress were investigated using wild-type or double-mutant (R70Q/H+L91M) HCV-1b core transfection and stable expression in human hepatoma HuH-7 cells.Overall, 24, 19, 11, and 18 patients had the wild-type, R70Q/H, L91M, and R70Q/H+L91M genotypes, respectively, in the HCV core. A significantly higher accumulation of hepatocellular 8-OHdG and a lower NRF2/8-OHdG ratio were observed in patients with R70Q/H+L91M than in those with the wild-type disease. Increased levels of intracellular superoxide and hydrogen peroxide in the cytoplasm and mitochondria, mRNA expression of enzymes generating oxidative stress, and nuclear expression of nicotinamide adenine dinucleotide phosphate oxidase 4 were augmented in cells treated with R70Q+L91M.HCV core proteins harboring either or both substitutions of R70Q/H or L91M enhanced hepatocellular oxidative stress in vivo and in vitro. These amino acid substitutions may affect HCC development by enhancing hepatic oxidative stress in patients with chronic HCV-1b infection.

Variability of the HCV core region and host genetic and epigenetic factors can predict the response to pegylated interferon/ribavirin therapy in genotype 1b hepatitis C patients from Serbia

Variations in the hepatitis C virus (HCV) core sequence have been related to disease progression and response to antiviral therapy. Previously we showed that the methylation status of RASSF1A and p16 genes, and IL28B genotypes affects the response to pegylated interferon/ribavirin (PEG-IFN/RBV) therapy. Herein we investigated whether amino acid (aa) substitutions in the HCV core region alone or in combination with IL28B genotypes and RASSF1A/p16 methylation affect the response to PEG-IFN/RBV therapy and liver disease progression. Among 29 examined patients, we found no association between single aa substitutions and response to therapy. However, we observed that patients with the HCV core aa substitution at position 75 and CT/TT IL28B genotypes were non-responders (NR), (P=0.023). Moreover, these patients had unmethylated RASSF1A. In contrast, most patients (75%) with aa substitutions at position 91 and CC IL28B genotype achieved sustained virologic response (SVR), (P=0.030), and 70% of them had methylated RASSF1A gene. Our results suggest that combined analysis of aa substitutions in the core protein, the IL28B rs12979860 polymorphism, and the methylation status of the RASSF1A gene may help in predicting treatment response to PEG-IFN/RBV in genotype 1b chronic hepatitis C patients.

Inhibition of hepatitis C virus genotype 4 replication using siRNA targeted to the viral core region and the CD81 cellular receptor.

Hepatitis C virus (HCV) is one of the most important causative agents of hepatitis worldwide. The current study aimed to evaluate the silencing effect of the small interference RNA (siRNA) molecules designed against the core region of HCV genotype 4 (HCV-4) and the CD81 gene, which is the cellular receptor for HCV in the human hepatocytes. RT-PCR was used to measure the changes in both the viral HCV core and the cellular CD81 genes induced by the specific siRNA molecules. Additionally, the fluctuations in either the viral or the cellular proteins of the target regions were tested by flow cytometry and immunofluorescence. The results showed the effectiveness of the used siRNA molecules against the target genes in either RNA or protein levels. The effect of 100nM of siCD81 and 40nM of siCore was more evident at 24 and 48h post-transfection. The combination of the two siRNA molecules resulted in an extra inhibitory effect of the HCV core at both the RNA (85.6%) and protein (98.5%) levels. The current study suggested that targeting of the CD81 cellular receptor and/or the viral HCV core region by the small interference molecules might be a suitable choice in the suppression of HCV-4 replication. This might assist the development of new antiviral medications and provides a new alternative strategy for the targeting and treatment of HCV genotype 4.

Core encoding sequences of Hepatitis C virus in Ghanaian blood donors are predominantly mosaics of different genotype 2 strains and cannot distinguish subtypes

BackgroundDistribution of Hepatitis C virus (HCV) genotypes varies significantly worldwide. Genomic diversity between genotypes has implications for treatment, vaccine development and optimal design of HCV diagnostic assays. Molecular characterization of HCV in different geographical areas is therefore very essential for management and public health control of HCV infection. This study investigated the molecular epidemiology and characteristics of HCV genotypes in healthy individuals in Accra, Ghana.MethodsAn experimental study was carried out on blood samples obtained from voluntary blood donors. Two hundred samples were initially screened for HCV antibodies and infection was confirmed by RNA detection through RT-PCR of the 5′-untranslated region (5’UTR). The core gene sequences were analysed for HCV genotype determination by genotype-specific PCR; and then by cloning and direct sequencing followed by phylogenetic analysis. The sequences were further analysed in detail by similarity plotting.ResultsMolecular diagnosis confirmed the presence of HCV RNA in 2 out of 200 (1%) blood donors. Initial genotyping by genotype-specific PCR identified all two infections as subtypes 2a and 2b of genotype 2. Extensive evolutionary and genetic analyses indicated two epidemiological profiles. First, phylogenetic tree topologies clearly showed that, collectively, the core sequences of the Ghanaian HCV isolates belong to a single, distinct genetic group within HCV genotype 2 cluster, with high genetic similarity and rapid sequence variation in a single individual. Second, the sequences are mosaics comprising 2e and other genotype 2 subtype fragments. The analyses underscore a unique and complex HCV genotype 2 core sequence profile of the Ghanaian isolates.ConclusionsAnalysis of HCV core encoding sequences from Ghanaian blood donors in Accra confirmed predominance of genotype 2 HCV among healthy individuals. However, the isolates could not be classified into subtypes, possibly due to their complex sequence pattern that might suggest high mutability of the prevailing genotype. The core region of Ghanaian HCV therefore may not be suitable for distinguishing subtypes. These findings extend those from previous studies and thus underscore the need to search for subtype-informative region of Ghanaian HCV to elucidate the genetic diversity and factors determining outcome of HCV infections in Ghana.

Mixed HCV Infection of Genotype 1B and Other Genotypes Influences Non-response during Daclatasvir + Asunaprevir Combination Therapy.

Daclatasvir (DCV) + asunaprevir (ASV) combination therapy has become available for patients with hepatitis C virus (HCV) serogroup 1 infection. We studied the efficacy of this therapy by focusing on the factors associated with sustained virological responses (SVR) including resistance-associated variants (RAVs) and mixed infection of different HCV genotypes. We enrolled 951 HCV serogroup 1-positive patients who received this combination therapy at our hospital or affiliated hospitals. The presence of RAVs in non-structural (NS) regions 3 and 5A was analyzed by direct sequencing. HCV genotypes were determined by PCR with genotype-specific primers targeting HCV core and NS5B regions. SVR was achieved in 91.1% of patients. Female sex, age > 70 years, and RAVs were significantly associated with non-SVR (p<0.01 for all). Propensity score-matching results among the patients without RAVs regarding sex, age, and fibrosis revealed that mixed HCV infection determined by HCV NS5B genotyping showed significantly lower SVR rates than 1B-mono infection (p=0.02). Female sex and RAVs were significant factors associated with treatment failure of this combination therapy for patients with HCV serogroup 1 infection. Mixed HCV infection other than 1B-mono infection would be useful for predicting treatment failure.

Molecular characterization and clinical epidemiology of HCV in District Dir (Lower), Pakistan.

With about 200 million infections globally, Hepatitis C virus (HCV) infection is a major global health threat. The relative prevalence of hepatitis and HCV genotypes/subtypes varies among different geographic regions. Therefore, the present study was conducted to determine the prevalence of hepatitis C and HCV genotypes/subtypes in District Dir (Lower), Pakistan. Blood samples from HCV positive patients were genotyped through multiplex PCR using specific primers for HCV core region. A structured questionnaire was used to obtain information from the patients and data was statistically analyzed for different epidemiological parameters. The molecular evaluation results suggested the prevalence of genotype 3 in this study. The frequency of hepatitis C was found higher in males (P < 0.01). Present study suggests injections received at local clinics as a highly significant mode of HCV transmission to these patients (P < 0.002). These findings might be helpful for clinicians and related health care personnel to assess status of this important disease and highlight the need for more detailed evaluation of this devastating disease in order to frame better treatment strategies.

Lack of Association between Hepatitis C Virus core Gene Variation 70/91aa and Insulin Resistance.

The role of hepatitis C virus (HCV) in insulin resistance (IR) is not fully understood. The aim of this study was to determine the impact of amino acid (aa) substitutions in the core region of HCV according to IR and to identify clinical and laboratory associations. Ninety-two treatment-naive HCV patients were recruited to determine laboratory data and blood cell count. IR was determined using Homeostasis Model Assessment (HOMA) index where IR was defined as HOMA ≥2. HCV RNA load and genotype were determined by Abbott Real time HCV. HCV core region was determined by direct nucleotide sequencing. Bivariate analysis was conducted using HOMA IR ≥2 as a dependent factor. IR prevalence was 43.5% (n = 40), vitamin D sufficiency was found in 76.1% (n = 70) and 72.8% (n = 67) had advanced liver fibrosis. In the bivariate analyses, elevated values of γGT (p = 0.024) and fibrosis staging (p = 0.004) were associated with IR, but IR was not related to core mutations. The presence of glutamine in position 70 was associated with low vitamin D concentration (p = 0.005). In the multivariate analysis, no variable was independently associated with HOMA-IR. In conclusion, lack of association between IR and HCV core mutations in positions 70 and 91 suggests that genetic variability of this region has little impact on IR.

Discrepancy between Hepatitis C Virus Genotypes and NS4-Based Serotypes: Association with Their Subgenomic Sequences

Determination of hepatitis C virus (HCV) genotypes plays an important role in the direct-acting agent era. Discrepancies between HCV genotyping and serotyping assays are occasionally observed. Eighteen samples with discrepant results between genotyping and serotyping methods were analyzed. HCV serotyping and genotyping were based on the HCV nonstructural 4 (NS4) region and 5′-untranslated region (5′-UTR), respectively. HCV core and NS4 regions were chosen to be sequenced and were compared with the genotyping and serotyping results. Deep sequencing was also performed for the corresponding HCV NS4 regions. Seventeen out of 18 discrepant samples could be sequenced by the Sanger method. Both HCV core and NS4 sequences were concordant with that of genotyping in the 5′-UTR in all 17 samples. In cloning analysis of the HCV NS4 region, there were several amino acid variations, but each sequence was much closer to the peptide with the same genotype. Deep sequencing revealed that minor clones with different subgenotypes existed in two of the 17 samples. Genotyping by genome amplification showed high consistency, while several false reactions were detected by serotyping. The deep sequencing method also provides accurate genotyping results and may be useful for analyzing discrepant cases. HCV genotyping should be correctly determined before antiviral treatment.

夫婦間感染が明らかとなったC型急性肝炎の一例

夫婦間感染が明らかとなったC型急性肝炎の一例

Comparison of Three Different Hepatitis C Virus Genotyping Methods: 5'NCR PCR-RFLP, Core Type-Specific PCR, and NS5b Sequencing in a Tertiary Care Hospital in South India.

Based on genetic heterogeneity, hepatitis C virus (HCV) is classified into seven major genotypes and 64 subtypes. In spite of the sequence heterogeneity, all genotypes share an identical complement of colinear genes within the large open reading frame. The genetic interrelationships between these genes are consistent among genotypes. Due to this property, complete sequencing of the HCV genome is not required. HCV genotypes along with subtypes are critical for planning antiviral therapy. Certain genotypes are also associated with higher progression to liver cirrhosis. In this study, 100 blood samples were collected from individuals who came for routine HCV genotype identification. These samples were used for the comparison of two different genotyping methods (5'NCR PCR-RFLP and HCV core type-specific PCR) with NS5b sequencing. Of the 100 samples genotyped using 5'NCR PCR-RFLP and HCV core type-specific PCR, 90% (κ = 0.913, P < 0.00) and 96% (κ = 0.794, P < 0.00) correlated with NS5b sequencing, respectively. Sixty percent and 75% of discordant samples by 5'NCR PCR-RFLP and HCV core type-specific PCR, respectively, belonged to genotype 6. All the HCV genotype 1 subtypes were classified accurately by both the methods. This study shows that the 5'NCR-based PCR-RFLP and the HCV core type-specific PCR-based assays correctly identified HCV genotypes except genotype 6 from this region. Direct sequencing of the HCV core region was able to identify all the genotype 6 from this region and serves as an alternative to NS5b sequencing.

Distribution of hepatitis C virus genotypes in volunteer blood donors from Chengdu, China.

Hepatitis C virus (HCV) is a significant pathogen of global concern. The virus is usually spread through blood contact, such as transfusion, hemodialysis and injection of illegal drugs. HCV genotypes have a geographic distribution in different areas. In this paper, we focus on the distribution of HCV genotypes from volunteer blood donors in Chengdu. The prevalence of genotypes was analyzed using phylogenetic analysis. Phylogenetic trees were constructed based on the HCV core and NS5B regions from 313 sequences. HCV sequences were classified into six subtypes, and HCV genotypes were determined with the following results: 1b in 283, 2a in 14, 3b in seven, 3a in three, 6a in five and 6u in one. Subtype 1b was the most common and accounted for approximately 90.41% (283/313), and a virus of subtype 6u was isolated for the first time from the Chengdu area. Genotypes 4 and 5 were not detected.

Molecular characterization of hepatitis C virus core region in moroccan intravenous drug users.

Intravenous drug users (IDUs) represent a highly-infected reservoir for Hepatitis C virus (HCV) worldwide, harboring some of the most elevated prevalences and majority of the epidemic in developed nations. Studies aimed at sequencing regions of the viral genome uncovered amino acid mutations, some of which have been implicated in resistance to standard of care pegylated interferon/Ribavirin double therapy. Using the nested PCR method on the Core region of HCV strains in Moroccan IDUs living in the Tangier region this study sought to identify genotype-specific amino acid mutations, followed by Phylogenetic methods in order to compare them with international strains so as to identify sequences of highest homology. Genotyping was confirmed and recombination events excluded by line-probe assay. Italy was found most homologous for genotypes 1a and 3a, Iran for genotype 1a and Egypt for genotype 4a. Amino Acid Mutation analysis revealed the following novel genotype 3a-specific mutations: N16I, L36V, T49A, P71S, T75S, and T110N. The outcome of this work describes the HCV genetic heterogeneity in high-risk intravenous drug users, and it gives clues to the global migratory flow of genotypes as they cross geographical boundaries between various IDU populations and identifies "signature" amino acid mutations traceable to HCV genotype 3a. Identification of key amino acid positions in the HCV Core region with higher rates of mutations paves the way for eventual clinical trials seeking to establish a link between these recurrent mutations and response to standard of care Interferon and Ribavirin antiviral therapy. J. Med. Virol. 88:1376-1383, 2016. © 2016 Wiley Periodicals, Inc.

The Influence of Hepatitis C Virus Genetic Region on Phylogenetic Clustering Analysis

Sequencing is important for understanding the molecular epidemiology and viral evolution of hepatitis C virus (HCV) infection. To date, there is little standardisation among sequencing protocols, in-part due to the high genetic diversity that is observed within HCV. This study aimed to develop a novel, practical sequencing protocol that covered both conserved and variable regions of the viral genome and assess the influence of each subregion, sequence concatenation and unrelated reference sequences on phylogenetic clustering analysis. The Core to the hypervariable region 1 (HVR1) of envelope-2 (E2) and non-structural-5B (NS5B) regions of the HCV genome were amplified and sequenced from participants from the Australian Trial in Acute Hepatitis C (ATAHC), a prospective study of the natural history and treatment of recent HCV infection. Phylogenetic trees were constructed using a general time-reversible substitution model and sensitivity analyses were completed for every subregion. Pairwise distance, genetic distance and bootstrap support were computed to assess the impact of HCV region on clustering results as measured by the identification and percentage of participants falling within all clusters, cluster size, average patristic distance, and bootstrap value. The Robinson-Foulds metrics was also used to compare phylogenetic trees among the different HCV regions. Our results demonstrated that the genomic region of HCV analysed influenced phylogenetic tree topology and clustering results. The HCV Core region alone was not suitable for clustering analysis; NS5B concatenation, the inclusion of reference sequences and removal of HVR1 all influenced clustering outcome. The Core-E2 region, which represented the highest genetic diversity and longest sequence length in this study, provides an ideal method for clustering analysis to address a range of molecular epidemiological questions.

Associations between responses to interferon therapy and genetic variation in interleukin-28B and the core region of hepatitis C virus genotype 3a.

The single-nucleotide polymorphism (SNP) of interleukin-28B (IL-28B) and mutations in the core region of hepatitis C virus (HCV) genotype 1b have been associated with response to interferon (IFN) therapy. However, whether this IL-28B SNP affects responses to INF therapy for HCV genotype 3a is not known. The aim of this study is to investigate whether this IL-28B SNP (rs8099917) and specific missense mutations in the HCV core region affect the response to IFN therapy for HCV genotype 3a. Patients (n = 19; median age 44.5) infected with HCV genotype 3a who received IFN therapy were studied. Of the 19 patients, 12 (63.1%) achieved sustained virological response. Of those 12 patients, 11 had the TT genotype (11/16; 68.7%), and one had the TG genotype (1/3; 33.3%). The difference in the sustained virological response rate between IL-28B genotype groups was not significant (P = 0.5232). HCV core region was well conserved; however, polymorphisms at position 72 were identified. Of the 19 HCV samples; 15 carried a glutamic acid at position 72, and these were defined as E type; the others (4/19) were defined as non-E type. Notably, there was a significant difference in the sustained virological response rate between E type and non-E-type; 12 of the 15 patients with E-type achieved sustained virological response, but none of the four patients with non-E-type achieved sustained virological response (P = 0.009). A glutamic acid at position 72 in the core region of HCV genotype 3a was associated with a good response to IFN therapy. J. Med. Virol. 87:1361-1367, 2015. © 2015 Wiley Periodicals, Inc.

Computational models of liver fibrosis progression for hepatitis C virus chronic infection.

BackgroundChronic infection with hepatitis C virus (HCV) is a risk factor for liver diseases such as fibrosis, cirrhosis and hepatocellular carcinoma. HCV genetic heterogeneity was hypothesized to be associated with severity of liver disease. However, no reliable viral markers predicting disease severity have been identified. Here, we report the utility of sequences from 3 HCV 1b genomic regions, Core, NS3 and NS5b, to identify viral genetic markers associated with fast and slow rate of fibrosis progression (RFP) among patients with and without liver transplantation (n = 42).MethodsA correlation-based feature selection (CFS) method was used to detect and identify RFP-relevant viral markers. Machine-learning techniques, linear projection (LP) and Bayesian Networks (BN), were used to assess and identify associations between the HCV sequences and RFP.ResultsBoth clustering of HCV sequences in LP graphs using physicochemical properties of nucleotides and BN analysis using polymorphic sites showed similarities among HCV variants sampled from patients with a similar RFP, while distinct HCV genetic properties were found associated with fast or slow RFP. Several RFP-relevant HCV sites were identified. Computational models parameterized using the identified sites accurately associated HCV strains with RFP in 70/30 split cross-validation (90-95% accuracy) and in validation tests (85-90% accuracy). Validation tests of the models constructed for patients with or without liver transplantation suggest that the RFP-relevant genetic markers identified in the HCV Core, NS3 and NS5b genomic regions may be useful for the prediction of RFP regardless of transplant status of patients.ConclusionsThe apparent strong genetic association to RFP suggests that HCV genetic heterogeneity has a quantifiable effect on severity of liver disease, thus presenting opportunity for developing genetic assays for measuring virulence of HCV strains in clinical and public health settings.

Alternate Reading Frame Protein (F Protein) of Hepatitis C Virus: Paradoxical Effects of Activation and Apoptosis on Human Dendritic Cells Lead to Stimulation of T Cells

Hepatitis C virus (HCV) leads to chronic infection in the majority of infected individuals due to lack, failure, or inefficiency of generated adaptive immune responses. In a minority of patients, acute infection is followed by viral clearance. The immune correlates of viral clearance are not clear yet but have been extensively investigated, suggesting that multispecific and multifunctional cellular immunity is involved. The generation of cellular immunity is highly dependent upon how antigen presenting cells (APCs) process and present various viral antigens. Various structural and non-structural HCV proteins derived from the open reading frame (ORF) have been implicated in modulation of dendritic cells (DCs) and APCs. Besides the major ORF proteins, the HCV core region also encodes an alternate reading frame protein (ARFP or F), whose function in viral pathogenesis is not clear. In the current studies, we sought to determine the role of HCV-derived ARFP in modulating dendritic cells and stimulation of T cell responses. Recombinant adenovirus vectors containing F or core protein derived from HCV (genotype 1a) were prepared and used to endogenously express these proteins in dendritic cells. We made an intriguing observation that endogenous expression of F protein in human DCs leads to contrasting effects on activation and apoptosis of DCs, allowing activated DCs to efficiently internalize apoptotic DCs. These in turn result in efficient ability of DCs to process and present antigen and to prime and stimulate F protein derived peptide-specific T cells from HCV-naive individuals. Taken together, our findings suggest important aspects of F protein in modulating DC function and stimulating T cell responses in humans.

Amino Acid 70 Substitution in the Core Region of Hepatitis C Virus in Serum Lipid Markers of Patients with Chronic Hepatitis C Genotype 1b

Introduction: Previously, we reported that Hepatitis C Virus (HCV) infection had effects for lipid metabolism in patients with Chronic Hepatitis C (CHC). Recently, HCV with substitutions in amino acids (aa) 70 and/or 91 in the core region of HCV (HCV-C) has been reported to be more difficult to treat than HCV without aa 70 and 91 substitutions. The aim of this study was to clarify whether aa 70 or 91 substitution in the HCV-C influenced serum cholesterol fractions in patients with CHC genotype 1b and high viral load. Patients and Methods: Twenty-two patients infected with genotype 1b and high viral loads, whose serum samples taken before the start of therapy had been stored at -80°centigrade, and in whom aa 70 and 91 substitutions in the HCV-C could be detected, were selected. Patients without aa 70 and 91 substitutions in the HCV-C were assigned to wild (n=12), those with aa 70 substitution in the HCV-C assigned to mutant-70 (n=6), and those with aa 91 substitution in the HCV-C were assigned to mutant-91 (n=4). All patients received interferon (IFN)- based therapy. Fasting serum total cholesterol (C) and its fractions were compared before starting IFN therapy and at 24 weeks after the End of Therapy (EOT). When serum HCV-RNA was negative at 24 weeks after EOT, the patient was defined as having SVR. RESULTS: The SVR rates were 42% (5/12) in the wild, and 17i¼Â… (1/6) in the mutant-70, and 0% (0/4) in the mutant-91. Serum levels of LDL-C were significantly lower and those of HDL-C were significantly higher in the mutant-70 patients than in the wild patients before starting therapy. Only serum level of VLDL-C increased significantly at 24 weeks after EOT than before starting therapy in the wild patients. Conclusions: It was clarified that the mutant-70 influenced serum cholesterol fractions before starting therapy.

Deep-Sequencing Analysis of the Association between the Quasispecies Nature of the Hepatitis C Virus Core Region and Disease Progression

Variation of core amino acid (aa) 70 of hepatitis C virus (HCV) has been shown recently to be closely correlated with liver disease progression, suggesting that the core region might be present as a quasispecies during persistent infection and that this quasispecies nature might have an influence on the progression of disease. In our investigation, the subjects were 79 patients infected with HCV genotype 1b (25 with chronic hepatitis [CH], 29 with liver cirrhosis [LC], and 25 with hepatocellular carcinoma [HCC]). Deep sequencing of the HCV core region was carried out on their sera by using a Roche 454 GS Junior pyrosequencer. Based on a plasmid containing a cloned HCV sequence (pCV-J4L6S), the background error rate associated with pyrosequencing, including the PCR procedure, was calculated as 0.092 ± 0.005/base. Deep sequencing of the core region in the clinical samples showed a mixture of "mutant-type" Q/H and "wild-type" R at the core aa 70 position in most cases (71/79 [89.9%]), and the ratio of mutant residues to R in the mixture increased as liver disease advanced to LC and HCC. Meanwhile, phylogenetic analysis of the almost-complete core region revealed that the HCV isolates differed genetically depending on the mutation status at core aa 70. We conclude that the core aa 70 mixture ratio, determined by deep sequencing, reflected the status of liver disease, demonstrating a significant association between core aa 70 and disease progression in CH patients infected with HCV genotype 1b.

Characteristics of short double stranded RNA against hepatitis C virus: a literature-based analysis

ObjectiveTo describe the characteristics of short interfering double stranded RNA (dsRNA) against hepatitis C virus (HCV) and to find out the determining factors in design for desirable inhibitory efficacy. MethodsThe data were collected and analyzed by retrieval of 229 published short dsRNAs designed for degradation of HCV RNA. ResultsStatistical analyses showed that the most frequently involved short dsRNAs were directing against 5'NTR/core and genotype 1b, accounting for 64.2% and 69.9%, respectively. Inhibitory efficacy varied with the structural characteristics of short dsRNAs, of which the most potential were those directed against HCV core region with inhibitory efficacy of 70.2%. Moreover, the mean inhibitory efficacy of short dsRNAs with GC contents from 30% to 52% was higher than that of those with GC contents out of this range. ConclusionBased on this pooled data in a relatively large sample, the present results provided clues to design for short dsRNAs with more potent inhibitory efficacy.

Hepatitis C virus infections in the Democratic Republic of Congo exhibit a cohort effect

The prevalence and genetic diversity of hepatitis C virus (HCV) and human pegivirus (HPgV) in many regions of sub-Saharan Africa is poorly characterized, including in the Democratic Republic of Congo – the largest country in the region and one of the most populous. To address this situation we conducted a molecular epidemiological survey of HCV and HPgV (previously named GB Virus C or hepatitis G virus) in samples collected in 2007 from 299 males from the DRC, whose ages ranged from 21 to 71years old. Samples were tested for the presence of HCV antibodies by ELISA and reactive samples were subsequently tested for HCV RNA using RT-PCR in which both the HCV Core and NS5B genome regions were amplified. Remaining samples were tested for HPgV RNA and the HPgV NS3 genome region of positive samples was amplified. For HCV, 13.7% of the samples were seropositive (41/299) but only 3.7% were viremic (11/299). HPgV RNA was found in 12.7% (33/259) of samples. HCV viremia was strongly associated with age; the percentage of samples that contained detectable HCV RNA was ∼0.5% in those younger than 50 and 13% in those older than 50. Our study represents the first systematic survey of HCV genetic diversity in the DRC. HCV sequences obtained belonged to diverse lineages of genotype 4, including subtypes 4c, 4k, 4l and 4r, plus one unclassified lineage that may constitute a new subtype. These data suggest that HCV in the DRC exhibits an age ‘cohort effect’, as has been recently reported in neighbouring countries, and are consistent with the hypothesis that HCV transmission rates were higher in the mid-twentieth century, possibly as a result of parenteral, iatrogenic, or other unidentified factors. Different HCV subtypes were associated with individuals of different ages, implying that HCV infection in the DRC may have arisen through multiple separate HCV epidemics with different causes.