Molecular epidemiological characteristics of HIV-1 strains isolated from newly diagnosed MSM subjects (2006-2010) in Beijing, China / 病毒学报

This study aims to analyze the molecular epidemiological characteristics of HIV-1 strains prevailing among men who have sex with men (MSM) in Beijing, China. The pol gene fragments from 250 newly diagnosed HIV-1-infected MSM individuals during 2006-2010 in Beijing were amplified by RT-nested PCR, sequenced, and phylogenetically analyzed. HIV-1 pol gene from 189 individuals were amplified and analyzed; 81 (42. 9%), 3 (1. 6%), 2 (1.0%), 88 (46. 6%), and 15 (7.9%) individuals were infected with HIV-1 subtypes B, B', C, CRF01_AE, and CRF07_BC, respectively. The subtypes B and CRF01_AE could both be grouped into two clusters, and CRFO7_BC strains shared high homology and were presumed to originate from a common ancestor. The HIV-1 circulating in MSM in Beijing had a lower genetic diversity than in heterosexuals. The HIV-1 epidemic (2006-2010) in MSM in Beijing was actually a rapid spread of HIV-1 CRF01 AE and B, or rather native strains of the two viruses.

Simplified version of prejudice scale of minnesota multiphasic personality inventory and its reliability and validity / 第二军医大学学报

Objective To conduct item analysis on MMPI Prejudice Scale(Pr) and to develop a simplified version of the scale. Methods Pr and Symptom Checklist 90CSCL-90) were used to examine 3, 767 newly recruited soldiers. The difficulty degree and discrimination degree of each item of Pr were analyzed; the correlation between each item score and total score was analyzed; and a simplified version of Pr was developed based on the results of item analysis. The Cronbach's α coefficient values of the original scale and the simplified scale were calculated, and the criterion validity was analyzed using SCL-90 as the validity criterion. For other samples, 16PF, EPQ, and SAS were tested and the simplified scale was also tested at the same time. Results The difficulty degrees of the original scale ranged 0. 04-0. 69, with those of 17 items being less than 0. 20. The discrimination degree ranged 0. 10-0. 72, with those of 12 items being less than 0. 30. The correlation coefficient of the item score and the total score ranged 0. 19-0. 60, with those of 9 items being less than 0. 30. The simplified scale composed of 20 items with better quality. The Cronbach's α coefficient values of the original scale and the short-versioned scale were 0. 801 and 0. 757, respectively. The total scores of both the original scale and the short-versioned scale were significantly correlated with the scores of SCL-90, with the correlation coefficient being above 0. 38. The correlation coefficient between the score of the simplified scale and that of the original scale was 0. 967. The correlation coefficient between the score of the simplified scale and those of 16PF, EPQ, SAS was significant. Conclusion The simplified scale in this study has a higher reliability and validity.

Genotyping and variability of HIV-1 in 26 cases of paid blood donors / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To analyze the genome mutations of HIV-1 gag, pol and env genes from HIV-infected paid blood donors in rural central China.</p><p><b>METHODS</b>DNA was extracted from peripheral blood mononuclear cells, gag (p17-p24), pol (PR-RT), env (C2-V5) genes were amplified by nested polymerase chain reaction (PCR), purified products were sequenced, and sequence data was analyzed by MEGA5.0 soft wares.</p><p><b>RESULTS</b>Twenty-three samples were subtype B, two samples were recombinant of subtype B and subtype C, one sample was recombinant of subtype CRF01_AE and subtype B. PI major resistance mutations were not found in the PR region. M184V, K101E and G190A were detected in the RT region, respectively.</p><p><b>CONCLUSION</b>Subtype B was the major HIV circulating genetic forms in this area. Most strains were sensitive to high active anti-retroviral therapy (HARRT). 91.7% V3 loop tip motifs of X4-tropic strains was GPGR. It showed that GPGR might be associated with accelerate disease progression to AIDS.</p>

Analyses on antigen epitopes and drug resistance mutations of HIV-1 gag and pol genes / 病毒学报

To study the CTL antigen epitopes and drug resistance mutations of HIV-1 gag and pol genes through analyzing gag and pol gene sequences. The HIV-1 gag and pol gene fragments were amplified using nested polymerase chain reaction. A total of 23 PCR sequences, 449 cloned gag sequences and 402 cloned pol sequences were obtained. Sequence analyses showed the 23 samples were subtype B or B'. A total of 4 in 8 CTL antigen epitopes appeared 8 mutations in consensus sequence of subtype B and B'. There were no mutations found in the PCR sequences, whereas a few mutations were found in clone sequences (9.80%) in 5 antigen epitopes in p24 region. Eighteen PIs-related mutations and 24 RTIs-related mutations were found in PCR sequences and clone sequences in pol gene region, in which 17 (94.44%) PIs-related mutations and 15 (62.50%) RTIs-related mutations were found only in the clone sequences, respectively. The results showed that the prevalence of HIV-1 drug resistance strains in this study was at a higher level (17.39%), suggesting that some samples were resistant.to existing antiviral drugs.

Variation of the V3 loop tip motifs and primary drug resistance of HIV-1 strains / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the characteristics of the variation of the V3 loop tip motifs and the drug resistance in the primary treatment patients.</p><p><b>METHODS</b>The partial region of the HIV-1 env and pol gene in 51 samples was amplified by nested polymerase chain reaction (PCR) ,purified products were cloned into the vectors, the obtained were analyzed by MEGA soft wares.</p><p><b>RESULTS</b>The V3 loop tip motifs had four types in our study (GPGR, GPGQ, GPGK, GQGR); the study on the drug resistance in primary treatment patients, showed that there were not major resistance associated with PI, and the resistance were minor mutations in protease gene. In the RT region, there were nine resistance mutants were single NRTIs or NNRTIs.</p><p><b>CONCLUSION</b>The GPGR which was the typical western V3 loop tip motifs attained to 44.44%. This results showed that the percentage of primary drug resistance was still low in our study region, suggesting no need for genotyping detection in blood donor patients before primary therapy.</p>

Study of Gag gene antigen epitypes variation and the quasispecies group characteristics in Henan area HIV-1 strains / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the characteristics of the variation of antigen epitopes and quasispecies group in the HIV-1 viruses in Henan province in China.</p><p><b>METHODS</b>The region of the p17-p24 of the HIV-1 gag gene was amplified by nested polymerse chain reaction (PCR), purified products were cloned into the vector, the obtained were analyzed by MEGA soft wares.</p><p><b>RESULTS</b>B' subtype strains were predominant in Henan province, the mutations in antigen epitypes of the p17 region of the gag gene focus on E62g (55.8%), Y79f (48.9%), T84V (48.9), I44V (44.2%), the p24 region had not found the distinct mutation.</p><p><b>CONCLUSION</b>Both the PCR sequences and clone strains sequences demonstrated that four antigen epitopes mutations in p17 region of the HIV B' subtype gag gene, and the region of p24 was more conservative, which was suitable for development of the epitope vaccien.</p>

Expression and purification of HIV-1 subtype C Gp120, and its antibodies preparation / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To prepare HIV-1 subtype C Gp120 protein and to produce its polyclonal antibodies.</p><p><b>METHODS</b>A C-terminal fragment of gp120 gene was amplified by PCR from a plasmid expressing full-length HIV-1 subtype C gp160 gene. The length of the subtype C gp120 fragment was 612 nt and it encodes 204 amino acid residues. The resulting DNA construct was cloned into a prokaryotic expression vector (pET-30a) and recombinant pET-30a-gp120 was expressed in Escherichia coli BL21 (DE3) as an insoluble protein. The vector also contained a six-histidine (His6) tag at the C-terminus for convenient purification. To produce subtype C Gp120-specific polyclonal antibodies, New-Zealand rabbit was immunized with the purified Gp120 protein. Serum samples were tested by enzyme-linked immunosorbent assays (ELISA) to determine the level of antibodies. And Western blotting was used to further verify whether the polyclonal antibodies could specifically recognize subtype C Gp160 protein expressed in mammalian cells.</p><p><b>RESULTS</b>HIV-1 subtype C Gp120 protein was successfully acquired and the titer of its polyclonal antibodies was 1:204 800. The polyclonal antibodies efficiently recognized Subtype C Gp160 protein expressed in COS-1 cells.</p><p><b>CONCLUSION</b>HIV-1 subtype C Gp120 fusion protein with high purity was obtained and its corresponding polyclonal antibodies with high titer were produced.</p>

Immune potency of recombinant adeno-associated virus combined with recombinant adenovirus vaccine containing HIV-1 gp120 / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the immune effect of recombinant adeno-associated virus (rAAV) combined with recombinant adenovirus (rAdV) vaccine in BALB/c mice.</p><p><b>METHODS</b>The codon-modified HIV-1 gp120 gene was inserted into plasmid of adeno-associated virus and adenovirus vector separately. Then the rAAV and rAdV vaccines were constructed. BALB/c mice were immunized with rAAV and rAdV vaccines in different administration scheme. The IgG antibody was detected by ELISA and CTL response was detected by intracellular cytokine stain assay.</p><p><b>RESULTS</b>Both rAAV and rAdV vaccine could express gp120 gene; the mice primed with rAAV at week 0, 2 and boosted with rAdV at week 5, 14 and 20 elicited the strongest gp120 specific CTL and IgG antibody response.</p><p><b>CONCLUSION</b>The mice primed with rAAV and boosted with rAdV could elicit specific CTL response and IgG antibody.</p>

Construction and immune potency of recombinant adenovirus containing codon-modified HIV-1 gp120 / 中华实验和临床病毒学杂志

<p><b>BACKGROUND</b>To construct replication-deficient recombinant adenovirus expressing wild and codon-modified HIV-1 gp120.</p><p><b>METHODS</b>The viral codons were changed to the codon usage of highly expressed mammal gene, the resulting modified gp120 gene was synthesized. The wild and modified gp120 genes were cloned into shuttle vector pShuttle-CMV respectively, and then the constructed plasmids containing gp120 gene was cotransformed with the backbone vector pADeasy-1 into E.coli BJ5183. Transfection of the recombinant AdEasy plasmid into 293 cells was performed to obtain recombinant adenoviruses. The mice were immunized with the recombinant adenoviruses. Their immunogenicity was evaluated by testing antibody and CTL levels of immunized mice.</p><p><b>RESULTS</b>Two strains of recombinant adenovirus expressing wild and codon-modified HIV-1 gp120 were obtained. The protein expressing level of the recombinant adenoviruses containing modified genes was much higher than that containing wild genes. The mice immunized with recombinant adenoviruses elicited HIV-1 specific antibody and CTL response. The rAd-mod gp120 group was better than the rAd-wt gp120 group.</p><p><b>CONCLUSION</b>Replication-deficient recombinant adenovirus expressing HIV-1 gp120 can elicit HIV-1 specific humoral and cellular response, the codon-modified recombinant virus was more efficient than the native.</p>

Construction and analysis of activity of an HIV-1/bovine immunodeficiency virus chimeric clone cDNA / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>Chimeric human/bovine immunodeficiency virus (HBIV) cDNA was constructed by replacing HIV tat and LTR with bovine immunodeficiency virus (BIV) tat and LTR to study the activity of BIV tat and LTR in the chimerae.</p><p><b>METHODS</b>The target fragments of BIV tat, LTR and HIV gag, pol, env were respectively amplified by using PCR and sequentially inserted into pBluescript SK(+) vector. The chimeric clone was transfected into human MT4 cells. The transcript and gene expression of the HBIV chimeric virus were detected by using RT-PCR and a reverse transcriptase assay, respectively.</p><p><b>RESULTS</b>BIV tat mRNA and HIV gag mRNA were detected. The reverse transcriptase activity of the chimeric virus was analyzed in the fluctuation curve.</p><p><b>CONCLUSIONS</b>In chimeric HBIV cDNA transfected MT?4 cells, BIV tat and HIV gag were transcripted. The reverse transcriptase of the chimeric virus had biological activity. These data suggest that in MT4 cells, BIV LTR had promoter activity and BIV tat had the function of transactivation in the chimeric virus. The study of the chimeric virus with infectivity is in progress.</p>