Butantan Institute, Viral Immunology Laboratory, Av Vital Brasil n degrees 1500, CEP 05503-900, Butantã, São Paulo, SP, Brazil.
BMC Infect Dis. 2010 Feb 23;10:36. doi: 10.1186/1471-2334-10-36.
The quasispecies nature of HCV may have important implications for viral persistence, pathogenicity and resistance to antiviral agents. The variability of one of the viral proteins, NS5A, is believed to be related to the response to IFN therapy, the standard treatment for infection. In this study we analyzed the quasispecies composition of NS5A protein in patients infected with HCV genotype 3a, before IFN therapy.
Viral RNA was isolated from samples of 12 patients: four sustained virological responders (SVR), four non-responders (NR), and four end-of-treatment responders (ETR). cDNA was synthesized, the NS5A region was amplified and the fragments obtained were cloned. Fifteen clones from each patient were sequenced with eight primers, generating 179 contigs.
Higher values for substitution (either synonymous or non-synonymous) and for distance were found in the SVR group. However, the NR group showed relatively more non-synonymous mutations than the other groups, owing to the higher values of dN/dS in complete NS5A and most specific regions. Overall, NS5A protein is undergoing purifying selection, since all dN/dS ratios values are below 0.5.
Our study provides an overview of the genetic variability of complete NS5A protein in HCV genotype 3a.
HCV 的准种性质可能对病毒的持续性、致病性以及对抗病毒药物的耐药性具有重要影响。人们认为一种病毒蛋白 NS5A 的变异性与 IFN 治疗的反应（HCV 的标准治疗方法）有关。在这项研究中，我们分析了感染 HCV 基因型 3a 的患者在 IFN 治疗前 NS5A 蛋白的准种组成。
从 12 名患者的样本中提取病毒 RNA：4 名持续病毒学应答者（SVR）、4 名无应答者（NR）和 4 名治疗结束时应答者（ETR）。合成 cDNA，扩增 NS5A 区域，并对获得的片段进行克隆。对每个患者的 15 个克隆进行 8 个引物测序，生成 179 个连续序列。
SVR 组的替换（同义或非同义）和距离值较高。然而，NR 组的非同义突变相对较多，这归因于完整 NS5A 和大多数特定区域的 dN/dS 值较高。总体而言，NS5A 蛋白正在经历纯化选择，因为所有 dN/dS 比值均低于 0.5。
我们的研究提供了 HCV 基因型 3a 中完整 NS5A 蛋白遗传变异性的概述。
