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1.
新型HCV EIA诊断试剂盒的研制 总被引:3,自引:0,他引:3
丙型肝炎病毒(HCV)基因组结构区核壳蛋白(C)区抗原、膜蛋白E1和E2区抗原,以及非结构区NS3-NS5区抗原的区段,已经在原核细胞中获得有效的表达。同时,相应区段中的优势抗原表位肽也经化学合成法大规模地制备。HCV基因组上各区段抗原性的分析发现,由C区和NS3区分别编码的C抗原和C33c抗原是HCV基因组上两个优势抗原区段。其相应的抗体出现早(感染后6周可检出抗C33c抗体),阳转率高(约99%阳性检出率),特异性和重复性均优于其它区段抗原。以中国人HCV的C33c重组蛋白和分支状合成肽MAP-C-19为复合抗原,研制了适合我国抗HCV抗体检测的新型丙型肝炎病毒酶免疫测定(HCVELA)诊断试剂盒。它同当代美国Abbott/UBIHCVELA诊断试剂的符合率约98%,同加拿大YES公司HCVEIA诊断试剂的符合率约97.8%,阳性检出率提高了约2%,3次重复性达100%,表明其特异性、敏感性和重复性均达到了当代第二代JCVELA诊断试剂的水平。我国人群中抗HCV抗体的分布情况为:正常人群的检出率1%-2%;外科类住院病人检出率约28.8%;肝炎患者抗HCV阳性率为34.4%,慢活肝、肝硬化和重症肝炎患者 相似文献
2.
S. Rahamathulla B. S. V. V. Ratnagiri M. Manickam S. Sultana D. M. Mamatha O. Magisetty R. Nagarapu S. P. D. Ponamgi 《Russian Journal of Genetics》2018,54(9):1013-1024
Since last few decades hepatitis C virus (HCV) has been a major cause of death due to the involvement of acute and chronic type of liver diseases throughout the world. Genotype variability and mutations occurring at different regions of HCV genome provides a critical parameter for the study of sustained virological response (SVR) against mono and combinational therapies. Most of these mutations occurring in E2 and NS5A-ISDR regions in HCV genotypes play a significant role in SVR against Interferon-monotherapy and combination therapy. Therefore, the aim of the present study is to evaluate the SVR in various genotypes and the role of mutations in specific regions. In line with this, the NS5A and E2 proteins of HCV genotype 1 were found to suppress the double-stranded (ds) RNA-dependent protein kinase (PKR), which in turn is entailed in the cellular antiviral response stimulated by interferon (IFN). The response to IFN therapy varies between genotypes, with response rates among patients infected with types 2 and 3 nearly two-three-fold greater than in patients infected with type 1. Surprisingly, a considerable percentage of HCV genotype 3a infected patients do not react to treatment at all. In Japan, a link was observed between the numbers of mutations in an “interferon sensitivity determining region” (ISDR) and the result of interferon treatment in genotype 1b infected patients. Therefore, we published data on E2 (PePHD) and NS5A-ISDR regions including our data on SVR of different HCV genotypes, the relationship between the number and patterns of mutations in the E2 (PePHD) and NS5A-ISDR regions and responsiveness to IFN therapy. 相似文献
3.
利用逆转录套式PCR扩增Ⅲ型中国株HCVE2/NS1基因片段,将其克隆到pcDNA3载体上.采用双脱氧链终止法测定插入片段的核苷酸序列.并与已知分离株的相应区域进行同源性比较.首次克隆出Ⅲ型中国株HCVE2/NS1基因(HC-W14),其核苷酸序列与Ⅲ型日本株HCV(HC-J6)该区域同源性为88.37%,其推定的氨基酸同源性为89.29%.而与已知的非Ⅲ型株HCV该区域相比,核苷酸及氨基酸的同源性均相对较低.Ⅲ型中国株HCV与Ⅱ型中国株HCV在E2/NS1区域有较大的变异,揭示研制我国的HCV疫苗应该考虑这种基因型之间的变异性. 相似文献
4.
Lodrini S Bagaglio S Canducci F De Mitri MS Andreone P Loggi E Lazzarin A Clementi M Morsica G 《Journal of biological regulators and homeostatic agents》2003,17(2):198-204
The amino terminal region of the non structural gene 3 (NS3) of hepatitis C virus (HCV) is a chymotripsinlike serine-protease responsible for cleavage of the non structural proteins of Hepatitis C virus (HCV). In order to investigate the genetic variation of this region, we developed a nested PCR to obtain NS3 protease sequences from 54 patients chronically infected with HCV genotypes 1a, 1b and 3, respectively. Comparison of nucleotide and amino acids sequences of NS3 protease domain with consensus sequence obtained within the same genotype, showed 3.73% nucleotide divergence and 1.64% amino acid divergence in isolates of genotype 3a, whereas isolates 1a exhibited 4.45% nucleotide and 4% amino acid change, respectively. Finally, NS3 sequence from 1b isolates revealed 6.47% nucleotide and 3.5 % aa changes. Comparison of consensus amino acid sequences derived from isolates 1a, 1b and 3, with the HCV prototypes showed a low amino acid sequence diversity. However, the consensus sequence of HCV genotype 3 isolates showed an amino acid changed from the prototype, that was located within a region important for enzyme structure and activity. These results indicated that the NS3 protease gene is highly conserved within the same HCV genotype. The domains involved in enzyme function were highly conserved in 1a and 1b strains, whereas consensus sequence of isolates 3a showed that the majority of these strains were not perfectly conserved in one of such regions. These findings altogether suggested that the NS3 protease enzyme of HCV may constitute an important target for antiviral therapy, but the NS3 protease variability of isolates 3 within a region that is a potential target for antiviral therapy could pose a problem for structure based drug development. 相似文献
5.
猪瘟病毒兔化弱毒株cDNA片段的克隆及序列分析 总被引:2,自引:0,他引:2
猪瘟是猪最重要的传染病之一,往往给养猪业造成重大经济损失,猪瘟的病原为猪瘟病毒(HCV),属黄病毒科,瘟病毒属成员,其基因组为单股正链RNA,长度为123kb,仅含有一个大的开放阅读框架,编码一个含3898个氨基酸残基(AA)的多聚前体蛋白[1,2]。目前已经定位的蛋白有5种,即Npro、C、E0、E1和E2,它们均由HCVRNA5′端所编码,除Npro外,其它4种均为HCV的结构蛋白[3]。Npro为具有自我催化功能的蛋白水解酶,也是多聚蛋白N端的第一个蛋白水解酶,分子量为23kD,C为构成… 相似文献
6.
Suppression of hepatitis C virus replicon by RNA interference directed against the NS3 and NS5B regions of the viral genome 总被引:17,自引:0,他引:17
Takigawa Y Nagano-Fujii M Deng L Hidajat R Tanaka M Mizuta H Hotta H 《Microbiology and immunology》2004,48(8):591-598
RNA interference (RNAi) is a phenomenon in which small interfering RNA (siRNA), an RNA duplex 21 to 23 nucleotides (nt) long, or short hairpin RNA (shRNA) resembling siRNA, mediates degradation of the target RNA molecule in a sequence-specific manner. RNAi is now expected to be a useful therapeutic strategy for hepatitis C virus (HCV) infection. In the present study we compared the efficacy of a number of shRNAs directed against different target regions of the HCV genome, such as 5'-untranslated region (5'UTR) (nt 286 to 304), Core (nt 371 to 389), NS3-1 (nt 2052 to 2060), NS3-2 (nt 2104 to 2122), and NS5B (nt 7326 to 7344), all of which except for NS5B are conserved among most, if not all, HCV subtype 1b (HCV-1b) isolates in Japan. We utilized two methods to express shRNAs, one utilizing an expression plasmid (pAVU6+27) and the other utilizing a recombinant lentivirus harboring the pAVU6+27-derived expression cassette. Although 5'UTR has been considered to be the most suitable region for therapeutic siRNA and/or shRNA because of its extremely high degree of sequence conservation, we observed only a faint suppression of an HCV subgenomic replicon by shRNA against 5'UTR. In both plasmid-and lentivirus-mediated expression systems, shRNAs against NS3-1 and NS5B suppressed most efficiently the replication of the HCV replicon without suppressing host cellular gene expression. Synthetic siRNA against NS3-1 also inhibited replication of the HCV replicon in a dose-dependent manner. Taken together, the present results imply the possibility that the recombinant lentivirus expressing shRNA against NS3-1 would be a useful tool to inhibit HCV-1b infection. 相似文献
7.
Ping Qiu Xiao-Yan Cai Wei Ding Qing Zhang Ellie D Norris Jonathan R Greene 《Journal of biomedical science》2009,16(1):62-9
The genotype of Hepatitis C Virus (HCV) strains is an important determinant of the severity and aggressiveness of liver infection
as well as patient response to antiviral therapy. Fast and accurate determination of viral genotype could provide direction
in the clinical management of patients with chronic HCV infections. Using publicly available HCV nucleotide sequences, we
built a global Position Weight Matrix (PWM) for the HCV genome. Based on the PWM, a set of genotype specific nucleotide sequence
"signatures" were selected from the 5' NCR, CORE, E1, and NS5B regions of the HCV genome. We evaluated the predictive power
of these signatures for predicting the most common HCV genotypes and subtypes. We observed that nucleotide sequence signatures
selected from NS5B and E1 regions generally demonstrated stronger discriminant power in differentiating major HCV genotypes
and subtypes than that from 5' NCR and CORE regions. Two discriminant methods were used to build predictive models. Through
10 fold cross validation, over 99% prediction accuracy was achieved using both support vector machine (SVM) and random forest
based classification methods in a dataset of 1134 sequences for NS5B and 947 sequences for E1. Prediction accuracy for each
genotype is also reported. 相似文献
8.
9.
Next-Generation Sequencing Reveals Frequent Opportunities for Exposure to Hepatitis C Virus in Ghana
Joseph C. Forbi Jennifer E. Layden Richard O. Phillips Nallely Mora Guo-liang Xia David S. Campo Michael A. Purdy Zoya E. Dimitrova Dorcas O. Owusu Lili T. Punkova Pavel Skums Shirley Owusu-Ofori Fred Stephen Sarfo Gilberto Vaughan Hajung Roh Ohene K. Opare-Sem Richard S. Cooper Yury E. Khudyakov 《PloS one》2015,10(12)
Globally, hepatitis C Virus (HCV) infection is responsible for a large proportion of persons with liver disease, including cancer. The infection is highly prevalent in sub-Saharan Africa. West Africa was identified as a geographic origin of two HCV genotypes. However, little is known about the genetic composition of HCV populations in many countries of the region. Using conventional and next-generation sequencing (NGS), we identified and genetically characterized 65 HCV strains circulating among HCV-positive blood donors in Kumasi, Ghana. Phylogenetic analysis using consensus sequences derived from 3 genomic regions of the HCV genome, 5''-untranslated region, hypervariable region 1 (HVR1) and NS5B gene, consistently classified the HCV variants (n = 65) into genotypes 1 (HCV-1, 15%) and genotype 2 (HCV-2, 85%). The Ghanaian and West African HCV-2 NS5B sequences were found completely intermixed in the phylogenetic tree, indicating a substantial genetic heterogeneity of HCV-2 in Ghana. Analysis of HVR1 sequences from intra-host HCV variants obtained by NGS showed that three donors were infected with >1 HCV strain, including infections with 2 genotypes. Two other donors share an HCV strain, indicating HCV transmission between them. The HCV-2 strain sampled from one donor was replaced with another HCV-2 strain after only 2 months of observation, indicating rapid strain switching. Bayesian analysis estimated that the HCV-2 strains in Ghana were expanding since the 16th century. The blood donors in Kumasi, Ghana, are infected with a very heterogeneous HCV population of HCV-1 and HCV-2, with HCV-2 being prevalent. The detection of three cases of co- or super-infections and transmission linkage between 2 cases suggests frequent opportunities for HCV exposure among the blood donors and is consistent with the reported high HCV prevalence. The conditions for effective HCV-2 transmission existed for ~ 3–4 centuries, indicating a long epidemic history of HCV-2 in Ghana. 相似文献
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