哈尔滨地区男男性行为人群HIV-1感染未治疗者耐药基因变异分析

Virus subtype and drug resistance mutations among HIV-1 infected men who have sex with men in Harbin

目的了解2016年至2017年哈尔滨地区男男性行为人群(men who have sex with men,MSM)流行的人类免疫缺陷病毒1型(human immunodeficiency virus type 1,HIV-1)的亚型分布及原发耐药流行特点。方法收集哈尔滨市传染病院和哈尔滨医科大学附属第四医院60的新诊断的MSM途径HIV-1感染者未经抗逆转录病毒治疗(antiretroviral therapy,ART)的外周血样本,提取血浆中RNA,逆转录PCR和巢式PCR扩增HIV-1的gag、pol、int片段并纯化和测序。应用Mega 6.0软件构建系统进化树并分析病毒亚型。利用美国斯坦福大学HIV耐药检测系统分析耐药突变的种类和频率。结果成功获得50条gag基因序列、56条pol基因序列和44条int基因序列。亚型分析结果显示:CRF01_AE、CRF07_BC、B亚型和独特重组型(unique recombinant form,URF)分别占比为59.7%(34/57)、26.3%(15/57)、3.5%(2/57)和10.5%(6/57)。本研究共发现7份(12.5%)样本存在耐药突变,其中针对非核苷类逆转录抑制剂的耐药位点V179D/E/T频率最高为8.75%(5/56),此外还发现蛋白酶抑制剂耐药位点Q58E(1/56)和M46I(1/56)突变。未发现针对核苷类逆转录酶抑制剂和整合酶抑制剂的耐药突变位点。结论哈尔滨地区部分新诊断的MSM途径HIV-1感染者中以CRF01_AE亚型比例最多,其次为CRF07_BC和CRF01_AE与其他亚型重组株,耐药株所占比例较高,未发现整合酶抑制剂耐药突变。

Objective To investigate the distribution of subtypes of human immunodeficiency virus type 1(HIV-1)epidemic strains and the prevalence of primary drug resistance in the HIV infected individuals among men who have sex with men(MSM)population in Harbin from 2016 to 2017.Methods Peripheral blood samples with the antiretroviral therapy(ART)-na?ve were collected from 60 newly diagnosed HIV-1 infected individuals in Harbin Infectious Diseases Hospital and the Fourth Affiliated Hospital of Harbin Medical University.RNA was extracted from the plasma.HIV-1 gag,pol,and int gene fragments were amplified by reverse transcription PCR and nested PCR,purified,and sequenced.Phylogenetic tree and subtype analysis were performed using Mega 6.0 software.The type and frequency of drug resistance mutations were analyzed using the Stanford HIV Drug Resistance Database.Results 50 gag gene sequences,56 pol gene sequences,and 44 int gene sequences were successfully amplified.The subtype analysis showed that CRF01_AE,CRF07_BC,B subtypes,and unique recombinant forms(URF)accounted for 59.7%(34/57),26.3%(15/57),3.5%(2/57),and 10.5%(6/57),respectively.The drug resistance mutations were found in total of 7 samples(12.5%),among them the highest frequency of V179D/E/T was 8.75%(5/56)for non-nucleoside reverse transcription inhibitors.In addition,Q58E(1/56)and M46I(1/56)mutations which were resistance to protease inhibitors were also found.No resistance mutation sites to nucleoside reverse transcriptase inhibitors and integrase inhibitors were found.Conclusions Among the newly diagnosed HIV-1 infected individuals via MSM transmission route,the proportion of CRF01_AE subtypes is the highest,followed by CRF07_BC and other URF recombinant with CRF01_AE.The proportion of drug-resistant strains is higher and no drug resistance(DR)mutations to the integrase inhibitors was found.