| Objective To carry out molecular epidemiological survey to AIDS virus-1, which is epidemic in northern Guangxi , to provide the basis for making strategy for prevention and control of AIDS ,to provide valuable information to the work of prevention and control of AIDS in northern Guangxi ,even in the whole country.Methods 80 HIV-1 infected people /AIDS patients are investigated by questionnaires in Guilin, Laibin city of Guangxi Autonomous Region. To collect their epidemiological data, such as their demographic data, time of infection and affirmation and collect their anticoagulant whole blood sample 10ml, separate PBMC and extract DNA , amplify the C2-V3 region of env gene of HIV-1 virus using nested polymerase chain reaction, then conduct purification and recovery to the amplified products, sequence kits in a circular manner using fluorescence labelling extremity endings of ABI company , sequence usingD3 primer by 377-type DNA sequenator of PE company, then edit and proofread measured sequence using SeqEd and DNA software, use Wisconsin GCG package to conduct the arrangement, comparison and homologous analysis of the sequence.Results 1. Specimen source and amplification: The experiment collect 80 blood samples of HIV-positive infected people. After extracting provirus DNA and amplifying C2-V3 region of env gene of HIV-1 virus by nested PCR, C2-V3 region sequence of env gene of 55 specimens are gained in 80 specimens (amplification rate of 68.75%). In 55 specimens, 16 specimens have intravenous drug use history, 34 specimens have licentious history, 2 have illegal blood donation history, the route of infection of 3 specimens remains unknown.2. Gene subtype analysis: In 55 C2-V3 region sequences of env gene of HIV-1 virus, CRF01-AE recombinant strains occupy 14.55%(8/55), CRF08-BC recombinant strains occupy 41.82% (23/55), C subtype occupy 7.27%(4/55),CRF07-BC recombinant strains occupy 34.55% (19/55), B subtype occupy 1.82% ( 1/55). 8 CRF01-AE recombinant strains are close to representative strains of Thailand THCM240 and the average dispersion ratio is 7.68%. 23 CRF08-BC recombinant strains are close to 08-BC.CN.97.97CN GF-7F and the average dispersion ratio is 6.29%. 4 C subtype strains are close to C.95IN21068 and the average dispersion ratio is 8.69%. 19 CRF07-BC recombinant strains are close to CRF07-BC. CN97.CN54 and the average dispersion ratio is 6.92%. The average dispersion ratio among them and standard reference strains of other subtypes>20 %. Phylogenetic tree analysis further confirm the relationship among each strain .3. The analysis of dispersion ratio of C2-V3 region of env gene of HIV-1virus: The average dispersion ratio of C2-V3 region gene and its adjacent area,C2 region and V3 region in 55 strain samples are calculated respectively . The results show that the average dispersion ratio of 4 subtypes of C2 gene region are greater than V3 area. The difference has statistical significance between them(P<0.05),which indicate variation of C2 region is greater than V3 region4. Amino acid sequence variation analysis of C2-V3 region and its adjacent region: 8 CRF01-AE recombinant subtypes, 23 CRF08-BC recombinant subtypes, 19 CRF07-BC recombinant subtypes ,which are contained in C2-V3 region sequence, are translated respectively into amino acid sequences and generate consensus sequences. The results show that the top tetrapeptide of V3 loop in CRF01-AE recombinant strains have 4 types: GPGQ, GPGR, GPGH and GRGE, mainly in GPGQ; amino acid variation in V3 region of CRF08-BC recombinant strains is relatively less, only GPGQ; there are two kinds of amino acid variation in V3 region of CRF07-BC: GPGQ and GRGE.Conclusion 1. Trough determining C2-V3 region of env gene of provirus DNA in PBMC and nucleotide sequence of both sides from 55 HIV-1 infected people or AIDS patients, which are amplified by nested PCR in northern Guangxi, we find four subtypes : CRF01-AE, CRF-BC(CRF07-BC, CRF08-BC),C and B, indicating the diversity of HIV-1 strain subtypes.2. According the results of studying, CRF08-BC and CRF07-BC appear in sexual transmission infected people. We infer that the infective trend has spread from special population to general population and intravenous drug infected people may be the infective bridge to general population.3.Through the analysis of CRF01-AE recombinant strains in northern Guangxi, we find that the top tetrapeptide of V3 loop shows 4 types: GPGQ, GPGR, GPGH and GRGE, mainly in GPGQ; the result also shows two kinds of the top tetrapeptide of V3 loop in CRF07-BC recombinant strains: GPGQ and GRGE, which provide references for research and development of new tests and HIV-1 vaccine design.4. From the analysis of gene dispersion ratio, the gene dispersion ratio of CRF08-BC, CRF07-BC, CRF01AE and C is not too great, but with international standards, the dispersion ratio is relatively larger. According to the calculation of popular variation rate, the three subtypes have been popular for a long period of time in northern Guangxi. Currently, the diversity of popular strains in northern Guangxi may come into being because of the difference of the source of infection, which will obviously require further investigation.5. The study confirms that at the molecular level, there are subtypes diversity of HIV-1 strains in northern Guangxi people and have various routes of infection. |
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