新疆地区人类免疫缺陷病毒-1感染者CRF07-BC流行毒株gag基因遗传特性

Genetic dynamics of gag gene of subtype CRF07_ BC in human immunodeficiency virus type-1 infected patients in Xinjiang Uygur Autonomous Region

目的分析新疆地区HIV-1感染者中流行的单-亚型CRF07-BC毒株的gag基因遗传特性。方法对35例CRF07-BC亚型的HIV-1感染者分别于2011年9月（T1时间点）、2012年5月（T2时间点）、2013年2月（T3时间点）和2013年10月（T4时间点）4个时间点采集全血标本。为排除不同程度免疫功能对HIV-1基因变异的影响，使用流式细胞仪进行CD4＋T淋巴细胞计数，并按照CD4’T淋巴细胞〈200／pL（重度免疫抑制）、200～500／pL（中度免疫抑制）和〉500／pL（无免疫抑制）分为不同免疫功能的3组。抽提标本RNA，套式PCR扩增gag基因片段，最终将3组研究对象得到的4次序列进行基因离散率和进化树分析，并使用BEAST软件估算该地区CRF07-BC流行毒株的最近共同祖先株形成时间和平均进化速率。为排除HARRT对HIV-1基因变异影响，对HARRT组和非HARRT组进行基因离散率分析。结果3组不同免疫功能的研究对象均呈现出在T1、T2、T3、T4时间点的组内和组间基因离散率依次升高的趋势。HARRT组和非HARRT组也均呈现出在T1、T2、T3、T4时间点的组内和组间基因离散率依次升高的趋势。进化树分析显示，同一样本的几次序列在进化树上聚集成一簇，大部分毒株均在T4时间点上独立分出一支；其最近共同祖先株形成时间为10．9年E95％最高后验密度（HPD）：2．O～22．2年]，平均进化速率为1．34×10。替换·位点-1·年-1（95％HPD：5．16×10-4-2．23×10。替换·位点-1·年-1）。结论新疆地区HIV-1感染者CRF07-BC亚型毒株会随着在一个地区流行时间的延长，加大该地区同一亚型内的基因离散率；CRF07-BC单-亚型内病毒株的gag基因进化速率相对文献报道的略慢。

Objective To analyze the genetic dynamics of prevailing single subtype CRF07_BC in human immunodeficiency virus-1 （HIV-1） infected patients in Xinjiang. Methods Blood samples were collected at four time points, including in 2011 September （T1 time point）, in 2012 May （T2 time point）, in 2013 February （T3 time point） and in 2013 October （T4 time point）, from 35 HIV-1 infected patients of subtype CRF07_BC in Xinjiang. CD4＋ T lymphocyte count was conducted by using flow cytometry to eliminate the effect of immune function on HIV-1 gene mutation. All cases were divided into three groups according to CD4＋T lymphocyte counts〈200/μL （severe immune suppression）,200--500/μL （moderate immune suppression） and 〉 500/μL （nonimmuno suppression）. Then HIV-1 RNA was extracted and HIV-1 gag gene was amplified by nested polymerase chain reaction （nPCR）. The gag gene was sequenced by gene distance and phylogenetic analyses. The time to the formulation of the most recent common ancestor （tMRCA） and evolutionary rate of subtype CRF07 BC in Xinjiang were estimated by software Bayesian evolutionary analysis by sampling trees （BEAST）. Analysis of gene distance was conducted in highly active antiretroviral therapy （HAART） group and nonHAART group to eliminate the effect of HAART on the HIV-1 mutation. Results The gene distance within 3 groups at T1 time point, T2 time point, T3 time point and T4 time point showed an increasing trend respectively. Gene distance in HAART group and nonHAART group also showed the similar trend. Phylogenetic tree showed a cluster of sequences of the same samples and the branches were split into a new one at T4 time point, tMRCA was 10.9 years （95% highest posterior density [-HPD] ：2.0--22.2 years）. Evolutionary rate was estimated to be 1.34 X 10.3 substitutions site-1 year-1 （95% HPD： 5.16 X 10.4 --2.23 X 10.3 substitutions, site-1 year-1） substitutions per site per year. Conclusions With the viruses prevailing in Xinjiang, the gene distance increases in the single subtype CRF07_BC in different immune function groups. Evolutionary rate of gag gene of subtype CRF07_BC is slower than that of reference.