Department of Biology, UNESP-São Paulo State University, IBILCE-Institute of Bioscience, Language & Literature and Exact Science, São José do Rio Preto, São Paulo, Brazil.
PLoS One. 2013 Apr 25;8(4):e62393. doi: 10.1371/journal.pone.0062393. Print 2013.
Hepatitis C is a disease spread throughout the world. Hepatitis C virus (HCV), the etiological agent of this disease, is a single-stranded positive RNA virus. Its genome encodes a single precursor protein that yields ten proteins after processing. NS5A, one of the non-structural viral proteins, is most associated with interferon-based therapy response, the approved treatment for hepatitis C in Brazil. HCV has a high mutation rate and therefore high variability, which may be important for evading the immune system and response to therapy. The aim of this study was to analyze the evolution of NS5A quasispecies before, during, and after treatment in patients infected with HCV genotype 3a who presented different therapy responses.
Viral RNA was extracted, cDNA was synthesized, the NS5A region was amplified and cloned, and 15 clones from each time-point were sequenced. The sequences were analyzed for evolutionary history, genetic diversity and selection.
This analysis shows that the viral population that persists after treatment for most non-responder patients is present in before-treatment samples, suggesting it is adapted to evade treatment. In contrast, the population found in before treatment samples from most end-of-treatment responder patients either are selected out or appears in low frequency after relapse, therefore changing the population structure. The exceptions illustrate the uniqueness of the evolutionary process, and therefore the treatment resistance process, in each patient.
Although evolutionary behavior throughout treatment showed that each patient presented different population dynamics unrelated to therapy outcome, it seems that the viral population from non-responders that resists the treatment already had strains that could evade therapy before it started.
丙型肝炎是一种在全球范围内传播的疾病。丙型肝炎病毒（HCV）是该病的病原体，是一种单链正链 RNA 病毒。其基因组编码一种单一的前体蛋白，经加工后产生十种蛋白。非结构病毒蛋白之一的 NS5A 与基于干扰素的治疗反应最相关，这是巴西批准的丙型肝炎治疗方法。HCV 具有很高的突变率，因此具有很高的变异性，这可能对逃避免疫系统和对治疗的反应很重要。本研究旨在分析感染 HCV 基因型 3a 的患者在治疗前、治疗中和治疗后 NS5A 准种的进化，这些患者对治疗有不同的反应。
提取病毒 RNA，合成 cDNA，扩增和克隆 NS5A 区，并对每个时间点的 15 个克隆进行测序。对序列进行进化史、遗传多样性和选择分析。
该分析表明，大多数无应答患者治疗后持续存在的病毒种群存在于治疗前的样本中，表明它已适应逃避治疗。相比之下，大多数治疗结束时应答患者的治疗前样本中发现的种群要么被选择淘汰，要么在复发后频率较低，从而改变了种群结构。例外情况说明了每个患者的进化过程的独特性，因此也说明了治疗耐药性的过程在每个患者中都是不同的。
尽管整个治疗过程中的进化行为表明每个患者都表现出与治疗结果无关的不同种群动态，但似乎对治疗产生抗药性的非应答者的病毒种群在开始治疗之前就已经存在能够逃避治疗的菌株。
