辽宁省99例人类免疫缺陷病毒1感染者gag-pol区基因序列分析

目的 研究辽宁省人类免疫缺陷病毒／艾滋病（HIV／AIDS）患者gag-pol区的基因序列特征。方法采集辽宁省99例HIV-1感染者外周静脉血,提取前病毒DNA,经巢式PCR方法扩增gag—pol区2．6kb片段,与HIVS序列数据库中亚型参考序列共同构建系统进化树以区分亚型,并计算gag—pol区不同编码区域的基因离散率和Ks／Ka比值。结果 在辽宁省99例HIV-1感染者中发现A、B’、B、C、G5种亚型,循环重组型CRF01_AE、CRF03_AB、CRF06_cpx、CRF07_BC、CRF08_BC5种循环重组型和循环重组型间的重组ICR07／08。以B’亚型最多见,其次为CRF01_AE。在gag—pol区内,p17区、p2-p6区基因离散率高于p24区、蛋白酶及逆转录酶编码区。但B亚型p24区基因离散率处于较高水平。CRF07_BC和CRF08_BC在gag-pol各区基因离散率及同义替换、错义替换普遍处于较低水平。各亚型毒株的p2-p6区的Ks／Ka值最低,其均值〈1,而CRF01_AE毒株RT区的Ks／Ka值高达10．45。结论 辽宁省流行的HIV-1 gag—pol区亚型十分复杂。p24、蛋白酶及逆转录酶编码区是更为保守的区域。B亚型毒株的p24区承受较大的正向选择压力,CRF01_AE毒株RT区承受了较大的负向选择压力。CRF07_BC和CRF08_BC传入辽宁省的时间较短,奠基者效应明显。

Study on gag-pol region characteristics of HIV-1 strains epidemic in Liaoning province

OBJECTIVE: To investigate the genetic subtypes and sequence characteristics of gag-pol region of HIV-1 epidemic in Liaoning province. METHODS: 99 whole blood samples of HIV-1 infected individuals of Liaoning province were collected. Provirus DNA was extracted from anti-coagulated peripheral whole blood, and then the sequences of gag-pol region were amplified through nested PCR and then sequenced. Genetic subtypes were identified by phylogenitic analysis, controlled with new version subtype reference sequences of HIV sequence database. The genetic distances of different coding regions were calculated by MEGA2.0 with Kimura 2-parameter model. The synonymous (Ks) and antonymous (Ka) as well as the ratio of Ks/Ka were analyzed by SNAP tool in HIV sequence database. RESULTS: There are many HIV-1subtypes epidemic in Liaoning province. We found five subtype: A, Thai B, B, C and G, five CRFs: CRF01, CRF03, CRF06, CRF07, CRF08, and one inter CRF recomtinant ICR07/08 in 99 HIV-1 infected individuals of Liaoning province. Thai B subtype was the main subtype epidemic in Liaoning, next were CRF01_AE. In gag-pol region, the genetic distances of p17, p2-p6 were higher than p24, protease and RT coding region. The genetic distance of p24 region in B subtype strains were relative high compared with other subtypes. The genetic distance and the value of Ks and Ka were rather low in each region of CRF07 and CRF08. The ratio of Ks/Ka of p2-p6 region was the lowest in each subtype, the mean value was below 1. The ratio of Ks/Ka of RT region was as high as 10.45. CONCLUSION: The HIV-1 genetic subtypes in Liaoning province were rather complex. The p24, protease and RT coding region coding region were more conserved in gag-pol region. P24 region of B subtype strains received more positive selective pressure, while the RT region of CRF01_AE strains received more negative selective pressure. CRF07_BC and CRF08_BC were the youngest strains transmitted to Liaoning province and showed strong founder effect.