1 Research Laboratory of Clinical Virology, Ruijin Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai, China.
2 Department of Infectious Diseases, Ruijin Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai, China.
Microb Drug Resist. 2019 Jul/Aug;25(6):944-950. doi: 10.1089/mdr.2018.0360. Epub 2019 Jan 31.
The presence of pre-existing hepatitis C virus (HCV) resistance-associated substitutions (RASs) could attenuate viral susceptibility to direct-acting antiviral agents. The aim of this study was to better understand the differences among HCV RASs over time. We compared the prevalence and characteristics of naturally occurring HCV RASs in the NS3, NS5A, and NS5B genes between 2008 and 2016 in Chinese patients chronically infected with HCV genotypes (GT) 1b, 2a, 3a, 3b, and 6a. HCV RNA was extracted after serum samples were collected from 242 patients at treatment baseline, including 120 samples in 2008 and 122 samples in 2016. Reverse transcription and nested PCR were performed, and the PCR products of the NS3, NS5A, and NS5B regions were sequenced using the Sanger sequencing method. Finally, RASs were identified from the different viral strains. In GT1b, the overall frequency of NS5A RASs in 2016 was significantly higher than that in 2008 (42.0% vs. 18.4%; = 0.002). Among NS5A RASs, the most frequently detected RAS was Y93H (5.3% in 2008 vs. 15.9% in 2016; = 0.035), which confers medium- to high-level resistance to the NS5A inhibitors: daclatasvir (DCV), ledipasvir (LDV), ombitasvir (OMV), and elbasvir. The frequency of NS5A L28 (low-level resistance to DCV/LDV/OMV) in 2016 was also higher than that in 2008 (11.6% vs. 1.3%; = 0.027). In addition, the highest frequency of clinically relevant NS3 RASs was S122G/A/T (69.7% in 2008 and 72.5% in 2016) in HCV GT1b isolates, which had medium-level resistance to simeprevir and asunaprevir, followed by Y56F (7.9% in 2008 and 14.5% in 2016), which confers resistance to paritaprevir. Although NS5B C316N had the highest substitution rate in GT1b (80.2% in 2008 and 91.3% in 2016), it was associated with low-level resistance to sofosbuvir and dasabuvir. However, HCV RASs were rarely detectable at baseline in other genotypes or subtypes except GT1b in this study. The frequency of NS5A RASs in 2016 was significantly higher than that in 2008, especially at the L28 and Y93 substitution positions, which may be due to their better fitness compared with wild-type viruses.
本研究旨在更好地了解 HCV RAS 随时间推移的差异。我们比较了 2008 年至 2016 年期间中国慢性 HCV 基因型（GT）1b、2a、3a、3b 和 6a 感染患者中 NS3、NS5A 和 NS5B 基因中天然存在的 HCV RAS 的流行率和特征。在治疗基线时从 242 名患者采集血清样本后提取 HCV RNA，包括 2008 年的 120 个样本和 2016 年的 122 个样本。进行逆转录和嵌套 PCR，然后使用 Sanger 测序法对 NS3、NS5A 和 NS5B 区域的 PCR 产物进行测序。最后，从不同病毒株中鉴定出 RAS。在 GT1b 中，2016 年 NS5A RAS 的总体频率明显高于 2008 年（42.0%比 18.4%；=0.002）。在 NS5A RAS 中，最常检测到的 RAS 是 Y93H（2008 年为 5.3%，2016 年为 15.9%；=0.035），它对 NS5A 抑制剂：达卡他韦（DCV）、雷迪帕韦（LDV）、奥比他韦（OMV）和艾尔巴韦具有中至高耐药性。2016 年 NS5A L28（对 DCV/LDV/OMV 的低水平耐药性）的频率也高于 2008 年（11.6%比 1.3%；=0.027）。此外，GT1b 中临床相关 NS3 RAS 的最高频率是 S122G/A/T（2008 年为 69.7%，2016 年为 72.5%），对西米普韦和阿那普韦具有中等水平的耐药性，其次是 Y56F（2008 年为 7.9%，2016 年为 14.5%），对帕立瑞韦具有耐药性。尽管 GT1b 中 NS5B C316N 的取代率最高（2008 年为 80.2%，2016 年为 91.3%），但它与索非布韦和达沙布韦的低水平耐药性相关。然而，除 GT1b 外，本研究中其他基因型或亚型在基线时很少检测到 HCV RAS。2016 年 NS5A RAS 的频率明显高于 2008 年，尤其是在 L28 和 Y93 取代位置，这可能是由于它们与野生型病毒相比具有更好的适应性。
