Isolation And Sequence Analysis Of HIV-1 Strains Circulating In Some Regions Of China

Isolations and sequences analyses of HIV-1 strains circulating in China afford a solid basis to build the HIV strains stocks, acquire their biological features and analyze the rules of pandemic, diversities and recombinations.Full-length genomes sequencing play a important role in clarifying evolving characteristics of different clades, and supply a complete, clear genetics and molecular background to the virus epidemiology characters.In the research descried this article, we isoalted and cultured several HIV-1 strains from partional regions in China and analyze their important structural genes sequences such as gag, pol, env, in particular, full-length genome sequences analyses were also performed on the rare circulating isolates .Objective To isolate HIV-1 strains circulating in some regions of China, then learn their biological and sequence feature.Methods Primary HIV-1 strains were isolated from HIV-1 infected persons using PBMCs coculture method, then their growth dynamics feature and capacity of inducing syncytium were identified. Meanwhile, RNA was prepared and gag,pol,env gene was amplified by RT-PCR. Then the sequence was determined and sequence was analyzed by Clustal,BioEdit version,MEGA3.1 .The DNA of NX2 and SH52 were prepared and the full-length sequence was amplified by nested-PCR, sequence was analyzed by SimPlot.Results In 67 HIV-1 strains, 37 virus were subculturied steadily. 58 primary strains were isolated from 270 HIV-1 infected persons. It showed us three types growth dynamics that were rapid/high,slow/high,slow/low. It suggested that strains in samples of higher viral loads get a higher isolating rat(eH_C=78.959,χ²_(1)0.01=6.63.P<0.01),and the proportion of rapid/high strains in samples of higher viral loads were bigger than that in samples of lower viral loads. The sequence of 52 strains turned out to be 4 subtypes of B,CRF01_AE,CRF_BC and G. The strains of subtype B existed in the widest area. The subtype CRF01_AE strains mostly existed in Guangxi.Subtype G was found in shanghai.V3 loop central motif-GPGR mostly existed in the subtype B and subtype CRF01_AE The number of net charge in V3 loop in the subtype CRF01_AE is the higher in the subtype B.Genic diversity in env is bigger than gag and pol. Genic diversity in pol and env in subtype CRF01_AE is the biggest in three subtypes.Majority of the isolated strains were predicted to be M-tropism and CCR5 using strains.The full-length sequence of NX2 (subtype CRF_BC) and SH52(subtype G)was amplified by PCR.NX2 CRF08_BC showed three breakpoints located at positions at 1000 in gag,2600 in pol and 9200 in nef.Conclusion 95 HIV-1 strains circulating in some regions of China were subculturied and isolated; The replication dynamics of the isolated strains varied and related to HIV-1 viral loads in plasma; There existed many kinds of subtype as B,CRF01_AE,CRF07_BC and G, and the distribution of the subtype varied according to different regions and different transmission path; Genic diversity in env is bigger than gag and pol. Majority of the isolated strains were predicted to be M-tropism and NSI strains; The sequence variation of V3 regions were apparent.Two full-length sequence of subtype 08BC and G were amplified.