云南省德宏和昆明地区高效抗逆转录病毒治疗后HIV-1流行毒株pol区基因变异分析

目的 调查云南省德宏和昆明两个地区HIV-1流行毒株pol区基因在接受HAART后的变异情况.方法 选择接受HAART 1年以上血浆HIV-1病毒载量＞1 000拷贝/ml的139例HIV/AIDS患者,提取血浆中HIV-1 RNA,采用RT-nested-PCR扩增HIV-1的pol区基因蛋白酶(PR)和逆转录酶(RT)基因片段,测定DNA序列.经拼接整理后与美国斯坦福大学网站提供的HIV耐药数据库(Stanford HIV Drug Resistance Database)进行比对,分析耐药突变位点,采用MEGA 4.1构建系统进化树以确定HIV的亚型,结合HIV/AIDS患者流行病学资料进行综合分析.结果 昆明地区患者HIV以T215F/NY/I、M41L/M和T69G/N/I/S/A/D突变较多,突变率分别为39%(24/62)、27%(17/62)和27%(17/62),高于德宏地区患者HIV的相应突变率[分别为16%(11/69)、13%(9/69)和9%(6/69)],差异有统计学意义(x2值分别为8.646、4.242和7.909,P均＜0.05).德宏地区HIV-1流行毒株pol区基因亚型主要为CRF01_AE亚型为32%(22/69)、C亚型为25%(17/69)、B亚型19%(13/69).而昆明地区主要是08_BC亚型60%(37/62)、CRF01_AE亚型21%(13/62)和07_BC亚型15%(9/62).结论 云南省德宏、昆明两地区HIV-1流行毒株pol区基因PR、RT基因突变频率存在部分差异,昆明地区T215F/N/Y/I、M41L/M和T69G/N/I/S/A/D突变率高于德宏地区.两地共检出HIV-1流行毒株pol区基因CRF01_AE、B、C、BC、08_BC和07_BC共6种基因亚型.

Abstract：

Objective To investigate the variations in the pol region of HIV-1 strain in treatment failed patients in Yunnan province's Dehong prefecture and Kunming. Methods Blood samples were collected from 139 patients who experienced treatment failure ( HAART treatment ＞ 1 years and HIV-1 RNA Viral load ＞ 1 000 copies/ml). HIV-1 RNA was extracted from plasma, and nested-PCR was performed for amplification of PR and RT genes on the HIV-1 pol region. The PCR products were then sequenced and submitted to Stanford HIV Drug Resistance Database for comparison. The evolution tree was built up with MEGA 4. 1 system, combined with patients' demographics. Results The most prevalent mutation in Kunming patients were T215F/N/Y/I, M41L/M, and T69G/N/I/S/A/D, the mutation rates were 39%(24/62), 27% (17/62) and 27% (17/62) , respectively, which were higher than the corresponding mutations in the Dehong prefecture [16% ( 11/69), 13% (9/69) and 9% (6/69)]. The rate differences were statistically significant ( x2 = 8.646, 4.242 and 7. 909, all P ＜ 0.05 ). The most common HIV-1 pol region subtype in the Dehong patients were CRF01_AE subtype (32%, 22/69), followed by C subtype (25% ,17/69), and B subtype ( 19%, 13/69). Major subtypes in Kunming patients were 08_BC (60%,37/62 ), CRF01_AE subtype(21% , 13/62 ) and 07_BC ( 15% ,9/62). Conclusions Partial differences of the point mutations of the HIV-1 strain pol region and frequency of their occurrences exist among Dehong and Kunming patients, HIV-1 strains in Dehong prefecture for the NNRTIs mutations at the T215 Y/N/T, M41L and T69G/N/I/S/A/D are significantly higher than those in Kunming. Six isoforms are found respectively:CRF01_AE, B, C, BC, 08_BC and 07_BC from the epidemic strains of HIV-1 pol region subtype in Dehong and Kunming areas.