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Objective To study the phylogenetic evolution and genetic variations of gag gene among the prevalent human immunodeficiency virus (HIV )‐1 strains in Guangxi Zhuang Autonomous Region . Methods Plasma samples of 158 HIV‐1 infected patients in Guangxi area were collected during October 2011 to March 2012 .The gag gene fragments of HIV‐1 were amplified by reverse transcription/nested‐polymerase chain reaction and then sequenced .MEGA 5 .03 was utilized to construct phylogenetic tree and to calculate the genetic distances and selection pressures (globle ω) of gag gene and its coding regions . The comparisons between two groups were tested by Student′s t test ,and the comparisons of multiple groups were tested by one‐way ANOVA .Results A total of 140 amplification products of gag gene were obtained from 158 samples .Four subtypes of HIV‐1 were found ,including CRF01_AE (80 ,57 .1% ) , CRF08_BC (46 ,32 .9% ) ,CRF07_BC (10 ,7 .1% ) ,and subtype B (B′) (4 ,2 .9% ) .The genetic distances of gag gene of the above subtypes were 0 .036 ± 0 .001 ,0 .031 ± 0 .002 ,0 .043 ± 0 .003 and 0 .102 ± 0 .006 ,respectively ,with statistical significance (F=220 .62 ,P<0 .01) .The p17 and p24 coding regions suffered negative selection pressure (globleω<1) .Neither the globle ω in p17 region nor that in p24 region had significant differences among different subtypes (F=0 .761 ,P=0 .469 and F=0 .037 ,P=0 .964 , respectively ) . Conclusion CRF01_AE is the major subtypes of HIV‐1 in Guangxi Zhuang Autonomous Region .The coding regions of gag gene are relatively conserved during evolution .Changes of HIV‐1 prevalence ,however ,may affect the genetic variation of gag gene ,which should be continuously monitored .
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To investigate the genomic properties of HIV-1, we collected 3,081 sequences from the HIV Sequence Database. The sequences were categorized according to sampling region, country, year, subtype, gene name, and sequence and were saved in a database constructed for this study. The relative synonymous codon usage (RSCU) values of matrix, capsid, and gp120 and gp41 genes were calculated using correspondence analysis. The synonymous codon usage patterns based on the geographical regions of African countries showed broad distributions; when all the other regions, including Asia, Europe, and the Americas, were taken into account, the Asian countries tended to be divided into two groups. The sequences were clustered into nine non-CRF subtypes. Among these, subtype C showed the most distinct codon usage pattern. To determine why the codon usage patterns in Asian countries were divided into two groups for four target genes, the sequences of the isolates from the Asian countries were analyzed. As a result, the synonymous codon usage patterns among Asian countries were divided into two groups, the southern Asian countries and the other Asian countries, with subtype 01_AE being the most dominant subtype in southern Asia. In summary, the synonymous codon usage patterns among the individual HIV-1 subtypes reflect genetic variations, and this bioinformatics technique may be useful in conjunction with phylogenetic methods for predicting the evolutionary patterns of pandemic viruses.
Assuntos
HIV-1/genética , Regulação Viral da Expressão Gênica/genética , Europa (Continente)/epidemiologia , Códon/genética , Ásia/epidemiologia , América/epidemiologia , África/epidemiologiaRESUMO
Objective To comprehend the current difference of sequence variation of p24 coding region in gag gene and subtype distribution of HIV 1 in Henan province and Shanghai in China. Methods Plasma specimens were collected from 37 HIV 1 patients, of which 25 were from Henan province and 12 were from Shanghai. Most patients in Henan were infected by illegally donating blood while in Shanghai the major transmission routes were blood or blood product transfusion and sexuality. RNAs from plasma specimens were extracted and target gene were amplified by the method of RT PCR and Nested PCR. The sequences of p24 region, 693 nucleotides were determined, then phylogenetic analyses were performed. Results Subtyping showed that 83.8% (31/37) were B subtype. Of 25 Henan specimens, 23(92%)were B subtype and 2 were A subtype. Of 12 Shanghai specimens, 8(66.7%)were B subtype, 1 was A subtype, 2 were CRF01 -AE and 1 was CRF02 -AG. Comparing with the Consensus sequence (Consensus -B, from HIV database), nucleotide variation in B subtype of Henan was 1.6%～4.2%, with an average of 3.2%, while that of Shanghai was 2.0%～3.8%, with an average of 3.4%. No G to A hypermutation was observed in all variations. The median intrasubtype distance for B subtype in Henan and Shanghai was 2.9% and 3.5%, and the intersubtype distance between B subtype and other subtypes was 11.1%～12.5%. The two Consensus sequences of Henan and Shanghai B subtype obtained by CLUSTAL X were aligned to Consensus -B, both the substitution for predicted amino acid were 2.2%(5/231). Of all the variations,they shared the same three mutations: A14P、I91V and E180D. However, the disparity between the two Consensus sequences was not significant (0.9%). Phylogenetic analyses implied that many specimens were B subtype, and all the B subtype specimens from Henan and most of Shanghai B subtype specimens were close to the isolates in Thailand. Conclusion B subtype was the dominant HIV 1 isolate in Henan and Shanghai of China.The HIV 1 B subtype in Henan and Shanghai had coincident homology and shared some identical variations of amino acid.
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Objective To provide experimental evidences for developing a safe and effective re- combinant fowlpox virus which can prevent the infection of HIV-2.Methods A fowlpox virus(FPV) transferring vector was constructed by inserting HIV-2 gag gene to the downstream of a synthetic complex promoter ATI-p7.5?20 of vector pUTA2.Transfection was then carried out,and recombi- nant FPV(rFPV)was screened by 5'-bromo-deoxyuridine(BrdU),genome PCR and western blot detection.Balb/c mice were immunized with rFPV by muscular injection.Anti-HIV-2 antibody, CD4~+ and CD8~+ T-cell count and specific target-killing activity of spleen CTL in immunized mice were analyzed by ELISA,FACS and LDH release assay,respectively.Results A transferring vector pA- gag was constructed and confirmed by amplifying a fragment of 766 bp from the rFPV genome.Mean- while,HIV-2 multi-antibody-specific protein blot(55 000)was detected from the recombinant virus and the HIV-2 specific antibody was detected from the immunized Balb/c mice.HIV-2 specific target- killing activity of spleen CTL was observed in immunized mice.Conclusion A recombinant fowlpox virus expressing HIV-2 structural protein Gag has been obtained,and it can stimulate HIV-2-specific eelluar and humoral immune reactions in mice.