The First CD4 Counts And Co-receptor Tropism Of Main HIV Strains Of Newly Reported HIV Infections In Some Regions Of Guangxi

Background and Objective:Since the first case of local human immunodeficiency virus infection was reported in the Guangxi Zhuang Autonomous Region in the 1990s,the epidemic and spread of HIV have been about the following stages.In the initial stage,the population was mainly intravenous drug users,and the virus gene subtypes were mainly B subtype and C subtype,and there were signs of subtype recombination.Then,the infected population gradually changed,and unique CRF08_BC and CRF07 BC subtypes appeared.Subsequently,the proportion of CRF01_AE subtype strains gradually surpassed other subtypes,and heterosexual transmission surpassed drug transmission as the most important transmission route.The emergence of a new second-generation recombinant strain CRF55₀1B in the recent stage indicates that new mutations and recombination of the HIV strain have presented new challenges to the prevention and treatment of AIDS.The number of CD4+T lymphocytes in an infected person is an important indicator for judging immunity and disease progression.It is generally believed that an infected person with CD4+<200cells/μl has immunodeficiency.According to research,virus subtypes and virus helper receptor utilization are closely related to changes in the number of CD4+T lymphocytes.In the early stages of infection,the virus strains use more CCR5 receptors;as HIV replicates in the body,the proportion of strains that use CXCR4 receptors increases.This study analyzes the virus genotypes,the utilization（tropism）of virus co-receptors,and the relationship with the number of CD4+T lymphocytes in newly reported HIV-infected persons in some areas of Guangxi.Based on the results of genotype and phenotype,the characteristic amino acid positions related to the utilization of co-receptor in the V3 segment of the env region were analyzed.Methods:1.Collect the population epidemiological data of some newly reported HIV-infected persons in Qinzhou City,Guangxi from 2014 to 2019,including gender,age,occupation,marital status,infection route,etc.The number of CD4+T lymphocytes was detected,the pol and env gene regions were used to determine the virus genotype,the Geno2pheno tool was used to predict the co-receptor,and the GHOST cell line was used to detect the tropism of some strains.2.Analyze the differences in the number of CD4+T lymphocytes among the main HIV genotypes,subclusters and tropism,and use Logistic regression to analyze the factors related to CD4+<200cells/μl,and P<0.05 as the difference is statistically significant.3.The amino acid site map was used to preliminarily analyze the similarities and differences of the amino acid sites in the V3 segment of the env gene region between the main subtypes and tropisms.Logistic regression analysis was used to explore the amino acid positions related to tropism,and the results obtained were compared with the phenotypic results,so as to further study the relationship between subtypes,amino acid positions and tropism.Results:1.The basic situation of the survey subjects and the distribution of HIV subtypesThis study completed 2010 surveys of newly reported HIV-infected persons in Qinzhou City,Guangxi from 2014 to 2019,and collected whole blood samples,accounting for 33.1%of the total number of new reports in the region.Among the survey respondents,males accounted for 77.1%,aged 49.2±15.1 years old,farmers accounted for 75.7%,elementary school and below accounted for 54.2%,married couples accounted for 57.1%,and heterosexual transmission accounted for 87.9%.The HIV genotype among newly reported infections is determined based on the pol region and the region.CRF01_AE subtype accounted for 55.2%（1109/2010）,CRF08_BC subtype accounted for 29.5%（593/2010）,and CRF07 BC subtype accounted for 8.1%（163/2010）.In the CRF01_AE subtype,the first cluster（Clusterl）accounted for 20.2%（224/1109）,and the second cluster（Cluster2）accounted for 71.7%（795/1109）.In addition,105 cases of unique recombination（multiple infections may also exist）were mainly recombined with CRF01_AE subtype and CRF08 BC or CRF07_BC subtype.With the increase of the year,the proportion of CRFOI AE subtypes among newly reported HIV infections decreased（P=0.001）,and the proportion of CRF08_BC and CRF07_BC subtypes increased accordingly.However,in the CRF01_AE subtype,the proportion of subclusters did not change significantly between years.2.The number of first CD4+T lymphocytes and related factors of the survey subjectsThe number of first CD4+T lymphocytes M（P25,P7s）of the survey subjects was 168（55,333）,and the number of first CD4+T lymphocytes of CRF01_AE subtype was 108（32,272）lower than CRF08_BC’s 220（112,370）and 248（156,380）of CRF07_BC’s.Samples of 317 newly reported infections were selected for restricted antigen affinity experiments.28 new infections accounted for 8.8%,and 289 previous infections accounted for 91.2%.There was no statistically significant difference in the proportion of new infections among subtypes.No difference has been found in the number of CD4+T lymphocytes among the main subtypes of new infections,which may be related to the sample size.In previous infections,the number of CD4+T lymphocytes of CRF01_AE subtype was lower than that of other subtypes.Use the geno2pheno tool to predict co-receptors.From 2014 to 2019,15.9%（319/2010）of newly reported HIV-infected patients with CXCR4 tropism accounted for the virus.There was no statistically significant difference in the proportion of CXCR4 tropism among various years.26.0%（288/1109）of CRF01_AE subtype virus showed CXCR4 tropism,which was higher than 2.5%（15/593）of CRF08_BC,P<0.001.In the CRF01_AE subtype,26.3%（59/224）of the first cluster showed CXCR4 tropism,and 26.5%（211/795）of the second cluster showed CXCR4 tropism.The difference in CXCR4 tropism ratio between the first cluster and the second cluster was not statistically significant.Correlation analysis of the main HIV subtypes,subclusters,tropism and the number of first CD4+T lymphocytes.In the CRF01_AE subtype,there was no significant difference in the number of CD4+T lymphocytes between Clusterl and Cluster2（P=0.374）.In the CRF01_AE subtype,the number of CD4+T lymphocytes in infected individuals with CXCR4 tropism is lower than the number of CD4+T lymphocytes with CCR5 tropism.Logistic regression analysis was performed on the population epidemiology data of 2010 newly reported infected persons and laboratory data such as virus subtype and tropism.The results showed that male,farmer,heterosexual transmission,CRF01_AE subtype,CXCR4 tropism are related to the number of CD4+<200cells/μl.3.Analysis of the amino acid positions of different HIV subtype subclusters and tropism-related strainsFurther analyze the amino acid positions of the V3 segment of the env gene region between different subtypes.CRF01_AE subtype differs from CRF08_BC subtype and CRF07_BC subtype at the 5th.10th.11th.12th,13th.14th,18th,19th,22th,32th and 34th amino acids.No significant difference was observed in the amino acid positions of CRF08_BC subtype and CRF07_BC subtype.The previous research of the research group showed that in the CRF01_AE subtype,the 13th and 32th amino acids of the V3 segment of Cluster4 and Clusters are correlated with tropism,and there are also results showing that the 7th,11th,18th and 32nd positions of the V3 segment of the CRF01_AE subtype.There is a correlation between amino acids and tropism.Further analysis of Logistic regression showed that K10,G11,R11,V12,T12,M12,L12,R13,R18,V19,K32,R32,and Y34 were related to CXCR4 tropism;H13,P13,M18 were related to CCR5 tropism.In this study,the main subtype strains were selected for virus isolation and culture.and 10 virus strains were successfully cultured.The phenotype identified by the GHOST cell line and Geno2pheno predicted tropism,and the agreement between the two results was 90%（9/10）.The phenotypic results showed that among the amino acid positions of the V3 segment of CRF01_AE subtype,the 8th,10th,12th,14th and 18th amino acids were different among different tropisms.Among the amino acid positions in the V3 segment of CRF08_BC subtype,the 25th amino acid differs between different tropisms.Among the amino acid positions of subtype B,there are differences between the 11th and 25th amino acids.The results of phenotype and genotype consistently showed that the amino acids L12,M12 and R18 of the V3 segment of CRF01_AE subtype are related to CXCR4 tropism,and the amino acid R11 of V3 segment of B subtype is related to CXCR4 tropism.Other amino acid positions are not yet in the phenotype.observed.Conclusion:1.From 2014 to 2019,the viral genotypes of 2010 newly reported HIV-infected persons in Qinzhou City.Guangxi were mainly CRF01_AE,followed by CRF08_BC and CRF07_BC subtypes,and there were partially complex recombinant subtypes or multiple infections.The number of CD4+T lymphocytes of CRF01_AE subtype is lower than that of CRF08_BC and CRF07_BC subtypes;but in CRF01_AE subtype,the number of CD4+T lymphocytes of Clusterl and Cluster2 is similar.Regression analysis showed that male,farmer,heterosexual transmission,CRF01_AE subtype,and CXCR4 tropism were related to the number of CD4+<200cells/μl.2.CXCR4 tropism among newly reported infections accounted for a certain proportion.The proportion of CXCR4 tropism in CRF01_AE subtype is higher than the proportion of CXCR4 tropism in CRF08_BC and other subtypes.In the CRF01_AE subtype,the ratio of CXCR4 tropism of Cluster1 and Cluster2 is similar.3.Based on the phenotype and genotype results,the amino acids L12,M12,and R18 of the V3 segment of the env region of the CRF01_AE subtype are related to the CXCR4 tropism,while the amino acid R11 of the V3 segment of the B subtype is related to the CXCR4 tropism.It is suggested that the distribution of amino acid sites related to tropism in CRF01_AE subtype is different from other subtypes.