HIV type 1 subtypes among bar and hotel workers in Moshi,Tanzania

The HIV-1 prevalence among bar and hotel workers in Tanzania suggests they are a high-risk group for HIV-1 infection. We determined the HIV-1 subtype of 3'-p24/5'-p7 gag and C2-C5 env sequences from 40 individuals representing this population in Moshi. Genetic patterns composed of A(gag)-A(env), C(gag)-C(env), and D(gag)-D(env) were found in 19 (48.0%), 8 (20.0%), and 3 (8.0%) samples, respectively. The remaining 10 samples (25%) had different subtypes in gag and env, indicative of intersubtype recombinants. Among these recombinants, two contained sequences from HIV-1 subsubtype A2, a new genetic variant in Tanzania. Five bar and hotel workers may have been infected with viruses from a common source, based on phylogenetic analysis. The information obtained by surveillance of HIV-1 subtypes in a high-risk population should be useful in the design and evaluation of behavioral, therapeutic, and vaccine trial interventions aimed at reducing HIV-1 transmission.     

Sequence variability of the integrase protein from a diverse collection of HIV type 1 isolates representing several subtypes

HIV-1 recombinants between viruses from different subtypes appear to be surprisingly common in several regions of the world. To detect such intersubtype recombinants that contain mosaic genomes, we have analyzed sequences from the integrase (IN)-coding region of the polymerase (pol) gene from 23 viruses of known envelope (env) subtype from South America and Africa. As defined by env sequences, these viral genomes included nine subtype A, four subtype B, three subtype C, and four subtype D viruses from group M, and three viruses from group O HIV-1. Mosaic genomes were common, with 7 mosaic genomes among the 20 group M isolates analyzed. Two of these isolates had mosaic IN-coding regions that were distinct, but that had recombination breakpoints at the same location, in the highly conserved polypurine track. Mosaic genomes were particularly common in the viruses from Kenya (five of nine), consistent with our previous prediction that there was a high frequency of intersubtype recombinants circulating in this country. The IN amino acid sequence was highly conserved among the several represented subtypes, including group O. Group M IN sequences shared 94% or greater amino acid sequence identity within a subtype and 91% or greater identity between subtypes. The most divergent M and O variant amino acid sequences differed by only 19%, and the known functional domains were conserved among all of the isolates. The high degree of genetic homogeneity among the virus isolates representing several subtypes indicates that a single drug targeted against IN might be effective for all HIV-1 infections.     

Predominance of HIV type 1 subtype G among commercial sex workers from Kinshasa, Democratic Republic of Congo

We have investigated the genetic diversity and potential mosaic genomes of HIV-1 during the early part of the HIV-1 epidemic among commercial sex workers (CSWs) in Kinshasa, Democratic Republic of Congo (formerly Zaire). Serologic analysis revealed that 27 (28.7%) of the 94 specimens were seropositive by both peptide and whole-virus lysate EIAs and that 24 were positive by molecular screening assays, using generic primers that can detect all known groups of HIV-1. Phylogenetic analyses of the gag(p24), C2V3, and gp41 regions of these 24 specimens showed that all were group M; none of them had any evidence of group O, N, or SIVcpz-like sequences. On the basis of env sequence analysis, the 24 group M specimens were classified as subtypes G (37.5%), A (21%), F1 (12.5%), CRF01_AE (8%), D (4%), and H (4%); 3 (12.5%) were unclassifiable (U). Similar analysis of the gag(p24) region revealed that the majority of infections were subtype A; however, one-third of the specimens were subtype G. Parallel analysis of gag(p24) and env regions revealed discordant subtypes in many specimens that may reflect possible dual and/or recombinant viruses. These data suggest a predominance of subtype G (both pure G and recombinant CRF02_AG) during the early part of the epidemic in Kinshasa. Infections with group N or SIVcpz-like viruses were not present among these CSWs in Kinshasa.     

An unusual HIV type 1 env sequence embedded in a mosaic virus from Cameroon: identification of a new env clade. European Network on the study of in utero transmission of HIV-1

An atypical HIV-1 strain (CAM001) was identified in a pregnant Cameroonian woman in 1995. HMA subtyping of the env region was unsuccessful, and sequence analyses were performed. Unique sequence motifs were found at the V3 tip (GAGRALHA and GAGRAWIHA), and phylogenetic studies showed that the env C2-V5 sequence branched within group M but remained distinct from all known HIV-1 subtypes, while p17 gag branched with the subtype F sequences. Four other HIV group M viruses, undetermined by HMA, of African origin were found to cluster with CAM001 in the C2-V5 sequences. With the BLAST method, we found in databases three strains whose V3 sequences also clustered with CAM001. These unusual env sequences from eight HIV-1 strains derived from Cameroon formed a separate cluster in HIV-1 group M, which we designated k.     

武汉市HIV-1相关基因序列分析及亚型分布

目的研究武汉市HIV感染人群中的HIV毒株的亚型分布特点和流行规律。方法采集武汉市60名已被确认为HIV-1感染者的抗凝全血样品,提取前病毒DNA,用巢式聚合酶链反应方法（nested-PCR）扩增病毒膜蛋白env基因的C2-V5区及gag基因的部分区段,对PCR纯化产物直接测序,并应用GCG软件对序列进行分析。结果通过PCR扩增得到60份样品的结果,其中env基因序列45份、gag基因序列52份。依据env和gag区基因序列,与HIV-1各个亚型国际参考株比较,通过系统进化分析,最后确定武汉市样品分属5个亚型,分别为HIV-1B亚型中的泰国B（B’）亚型32份,流行重组型CRF07-BC12份,流行重组型CRF01-AE9份,A亚型1份和C亚型6份。结论武汉市存在多种HIV-1亚型,应加强对HIV-1毒株亚型变异的监测,及时调整防治策略。     

Spreading of HIV-1 subtype G and envB/gagG recombinant strains among injecting drug users in Lisbon,Portugal

We have evaluated the genetic diversity of HIV-1 strains infecting injecting drug users (IDUs) in Lisbon, Portugal. Heteroduplex mobility assay and/or phylogenetic analysis revealed that env (C2V3C3 or gp41) subtype B is present in 63.7% of the 135 viral samples studied, followed by subtypes G (23.7%), A (6.7%), F (5.2%), and D (0.7%). Similar analysis of gag (p24/p7) performed on 91 of the specimens demonstrated that 49.5% of the infections were caused by subtype G viruses; other gag subtypes identified were B (39.5%), F (3.3%), A and D (1.1.% each), and the recombinant circulating form CRF02_AG (5.5%). Discordant env/gag sub-types were detected in 34.1% of the strains and may reflect the presence of dual infections and/or recombinant viruses. The presumptive B/G recombinant form was highly predominant (21 of 31). The genetic pattern of HIV-1 subtype B and G strains is suggestive of multiple introductions and recombination episodes and of a longstanding presence of both subtypes in the country. C2V3C3 amino acid sequences from IDU-derived subtype G viruses presented highly significant signatures, which distinguish the variants from this transmission group. The unusually high prevalence of subtype G sequences (34.1%), independent of the geographic origin of the infected individuals, makes this IDU HIV-1 epidemic unique.     

Distinct sequence patterns characterize the V3 region of HIV type 1 gp120 from subtypes A and C

The known sequences of HIV-1 viruses have been categorized into subtypes based on the phylogenetic partitioning of their env and gag gene sequences. The env gene encodes the protein gp120, which contains five sequence- variable regions (V1 to V5), of which the V3 loop is of central importance to viral infectivity. The V3 loop consensus sequences of HIV-1 subtype A and C viruses are similar, and more similar to one another than the V3 consensus sequences of any other two HIV-1 subtypes. However, using a position-specific statistical comparison, we found that the V3 region of these two subtypes is statistically distinct (p = approximately 0.0). (The p-value calculated to the lowest limit of representation on the computer used to run the calculation. This lowest limit was 10(16). Although theoretically a p-value cannot be equal to 0.0, the p-value for the comparisons in question can be intuitively considered to be extremely small, or approximately 0.0.).     

Genetic and biological properties of HIV type 1 isolates prevalent in villagers of the Cameroon equatorial rain forests and grass fields: further evidence of broad HIV type 1 genetic diversity

To understand the evolution of HIV-1, the genetic and biological characteristics of viruses that infect persons living in regions in which the virus has been evolving for several decades must be studied. Thus, we investigated teh genetic subtypes, coreceptor usage, and syncytium-inducing ability of viruses in 47 HIV-1-infected blood samples from individuals living in rural villages in the equatorial rain forest and grass field regions in Cameroon. Heteroduplex mobility analysis (HMA) of gag (part of p24 and p7) and env (C2V5) or sequence and phylogenetic analysis of gag (part of p24 and p7), pol (protease), and env (C2V5), revealed a broad HIV-1 group M genetic diversity. Subtype analysis revealed genetic evidence of seven subtypes (A, C, D, F, G, H, and J) and three circulating recombinant froms (CRFs) (CRF01_AE, CRF02_AG, and CRF11_cpx). Only 15 (32%) of the 47 samples analyzed revealed a concordant subtype in all three genes (gag, pol, and env), while discordant subtypes and CRFs were identified for the remaining 32 (68%) samples. Two patterns of HIV-1 diversity could be discerned in two provinces. While more concordant subtypes in gag, pol, and env genes were identified in villages of South province (10 of 13, 77%), the HIV-1 diversity in the West province was characterized by intersubtype recombinants (63%). Five new intersubtype recombinants were identified including Agag Jpol Genv, Ggag Upol Aenv, AGgag Jpol Aenv, Agag AGpol Henv, and Cgag AGpol AGenv. All of the 40 viruses tested used the R5 coreceptor, of which four also used the X4 coreceptor. Four viruses were able to induce syncytia in MT-2 cells, however, syncytium induction did not correlate with coreceptor usage. This study further reveals the complexity of HIV-1 infection in rural Cameron and points to the future of the global epidemic, which may be characterized by more genetically diverse viruses.     

Recombinant strains of HIV type 1 in the United Kingdom

Twenty-five recombinant (mosaic) HIV-1 genomes were detected among 151 samples comprising 118 non-B subtype sequences and 33 samples containing subtype B sequences. Seven of the 25 mosaic patterns were similar to characterized circulating recombinant forms (two A/E, four A/G, and one D/F) and one was a MAL-like A/D recombinant. Eighteen of the recombinants had evidence of subtype A sequences in at least one region of their genome. One sample was found to contain a novel recombinant form (pol F, env K). Two samples could not be characterized unambiguously as recombinant forms and a further one appeared to be a complex C/J/D/A genomic form. The majority of the mosaic genomes were recombinants between gag, pol, or env, whereas the C/J/D/A mosaic had cross-over breakpoints within pol. These findings suggest that almost 20% of non-B subtype isolates of HIV-1 circulating in the United Kingdom have mosaic genomes. This shows the diverse origin of HIV-1 strains circulating in the United Kingdom and may have implications for antiretroviral drug resistance.     

Predominance of subtype A and G HIV type 1 in Nigeria, with geographical differences in their distribution

The purpose of this study was to generate data on the relative prevalences of the HIV-1 subtypes circulating in Nigeria. A total of 252 HIV-1-positive samples collected during an epidemiologic survey conducted in April 1996 were genetically characterized by HMA (heteroduplex mobility assay) and/or sequencing. Samples were collected in Lagos, Calabar, Kano, and Maiduguri. Overall, the predominant env subtypes were A (61.3%) and G (37.5%). Subtype A is more prevalent in the south (p < 0.001), about 70% in Lagos and Calabar, whereas a quarter of the samples was classified as subtype G in these states. In contrast, subtype G is predominant in the north ( < 0.001), representing 58% of the samples in Kano. In the northeastern region, Maiduguri, almost similar proportions of subtype A and G were seen, 49 and 47.4%, respectively. A total of 37 samples was also sequenced in the p24 region from the gag gene; 13 (35%) had discordant subtype designations between env and gag. The majority of the gag (12 of 17) and env (14 of 22) subtype A sequences clustered with the A/G-IBNG strain. Within subtype G, three different subclusters were seen among the envelope sequences. These different subclusters are observed among samples obtained from asymptomatic individuals and AIDS patients from the four Nigerian states studied. In conclusion, we observed a limited number of HIV-1 subtypes circulating in Nigeria, with subtypes A and G being the major env subtypes responsible for the HIV-1 epidemic. Nevertheless, the high rate of recombinant viruses (A/G) and the different A/G recombinant structures indicate a complex pattern of HIV-1 viruses circulating in this country.     

Genetic and biologic characterization of HIV type 1 subtype C isolates from south Brazil

The molecular and biological properties of HIV-1 subtype C strains from South Brazil were investigated. We sequenced gag and env fragments of viruses from 22 HIV-1-infected individuals from Porto Alegre City, which has the highest frequency of subtype C in the country. The sequences were then compared with other subtype B, C, and F strains isolated in Brazil and other countries using phylogenetic methods. Amino acid signatures were identified and correlated with phenotypic characteristics. We identified six strains with subtype C (27.3%), eight subtype B (36.4%), one subtype F (4.5%), six C/B recombinants (27.3%), and one B/F recombinant (4.5%). The Brazilian subtype C sequences formed a unique phylogenetic group and presented 6 and 18 specific amino acid signatures in gag and env, respectively. Three distinct patterns of C/B recombinants presented characteristic Brazilian amino acid substitutions. Subtype C viruses were predominantly R5 and non-syncytium-inducing, while C/B recombinants were R5/X4 and syncytium-inducing viruses. These findings suggest that subtype C viruses circulating in Brazil are the result of a unique introduction into the country. Recombination events between subtypes B and C have been occurring frequently for more than 10 years in South Brazil. Biological characterization confirms the hypothesis that subtype C is distinct from the others in the evolution of coreceptor utilization.     

Subtypes A,C, G,and recombinant HIV type 1 are circulating in Bangladesh

We analyzed the genetic diversity of HIV-1 circulating in Bangladesh by direct sequencing and subsequent phylogenetic analysis of the V3 region of the env gene and p17 fragment of the gag gene from nine unrelated patients. The sequences from one sample grouped into subtype A, five samples grouped into subtype C, and one grouped into subtype G. In addition, two patients appeared to be infected with different recombinant viruses consisting of subtype A and unclassifiable viral sequences. Epidemiological analysis revealed heterosexual transmission in the majority of cases. Furthermore, most subjects had a history of traveling, either to India or to the Arabian Peninsula. This study shows that several HIV-1 subtypes are circulating in Bangladesh, and we conclude that there must have been several introductions of HIV-1 into the Bangladeshi population.     

Evidence of a novel B/C recombinant exhibiting unique breakpoints of near full-length HIV type 1 genome from Northeastern India

Despite the predominance of the HIV-1 clade C in India, the presence of other subtypes and recombinants has been reported. Here we report the identification of a novel HIV-1 B/C recombinant isolated from Northeast India and characterized near full length genome of the recombinant virus. Bootscan analysis of the nearly full-length genome showed a unique mosaic structure consisting of a subtype B backbone with three subtype C genome insertions. Breakpoint analyses revealed insertion of fragments belonging to subtype C at positions 1853-2223 in gag and 3025-3759 and 3998-5073 in pol. Phylogenetic analysis revealed that the segments of subtype B clustered with sequences of subtype B viruses reported from Thailand whereas segments of subtype C clustered with sequences of subtype C viruses reported from India. We report the mosaic structure that is distinct to HIV-1 B/C recombinant viruses reported to date.     

A new human immunodeficiency virus type 1 circulating recombinant form from Tanzania

It is becoming increasingly important to identify and to study human immunodeficiency virus type 1 (HIV-1) circulating recombinant forms (CRFs) with evidence of epidemic spread, since mosaic strains arise frequently, especially in populations where multiple subtypes cocirculate. We describe the almost complete nucleotide sequence of 3 subtype C and D recombinant viruses, selected from a pool of 13 D(gag)-D/C/D(env) perinatally infected infants from Dar es Salaam, Tanzania. All three genomes had cross-over points with approximately the same genomic localization. The subtype C-like sequences were located within pol, vif, vpr, vpu, the first exons of rev and tat, V3, and the U3-R regions of the LTR. Phylogenetic analyses of the full-length genomic sequences from these viruses showed the formation of a distinct subcluster on the HIV-1 subtype D branch. The pattern of recombination of genomes belonging to this new CRF, named CRF10_CD, might have resulted from independent recombination events occurring at high frequency or from a single source that originated earlier in this population. Future surveys will be needed to determine the potential of this CRF for epidemic spread.     

Identification of all HIV type 1 group M subtypes in Senegal, a country with low and stable seroprevalence

A total of 343 HIV-1-positive samples obtained between June 1996 and March 1999 was genetically characterized in the envelope region by HMA and/or sequencing. The env subtype distribution was as follows: 290 (84.6%) A, 22 (6.5%) B, 16 (4.7%) C, 8 (2.5%) D, 1 (0.03%) E, 1 (0.03%) F1, 4 (1.2%) G, and 1 (0.03%) H. For 77 samples the p24 region from the gag gene was also sequenced, and for 9 (11.6%) the subtypes between env and gag were different. Phylogenetic tree analysis showed the predominance of AG-IBNG-like viruses among gag and env subtype A sequences. HMA is relatively simple and requires less sophisticated technical facilities compared with sequencing, and in Senegal 323 (94.2%) of the 343 samples could be identified by this technique. However, in the actual configuration of the assay, discrimination between the recombinant AG-IBNG-like recombinant viruses, which are predominant in Senegal, and the nonrecombinant subtype A viruses is not possible.     

Identification and characterization of HIV type 1 subtypes present in the Kingdom of Saudi Arabia: high level of genetic diversity found

Saudi Arabia has a very low prevalence of HIV infections and nothing is known about HIV strains present in the population. Here specimens were collected from 62 HIV-1-infected patients at the King Faisal Specialist Hospital and Research Center, Jeddah, Saudi Arabia. Viral sequences were PCR amplified using primers for HIV-1 group M in gag p24, pol integrase, and env gp41 and genetic subtype was determined by phylogenetic analysis. HIV-1 viral sequences were amplified from 56 of the 62 specimens. Based on phylogenetic analysis of viral sequences, subtype C was the most common subtype present and accounted for 39.3% of the infections followed by subtype G (25%), subtype B (17.9%), subtype D (3.6%), and subtypes A and CRF02_AG (1.8% each). In addition, for six specimens subtype classifications were discordant between gag, pol, and/or env; these intersubtype recombinant viruses account for 10.7% of the infections and consisted of recombinants of subtypes A/CRF01, A/CRF02, A/G, B/G, and D/CRF02. The high HIV-1 strain diversity suggests that there have been multiple introductions of HIV-1 into Saudi Arabia from several sources. Within the study population, there were five husband/wife pairs. For each pair, the viral sequences obtained were closely related to each other showing that heterosexual transmission occurred.     

Subtype B and subtype C HIV type 1 recombinants in the northeastern state of Manipur, India

The predominant HIV-1 strain circulating in India is subtype C. However, subtype A and B strains of HIV-1 have also been reported in India. In 1999, the first A/C recombinant strain was reported from Pune in India. Intravenous drug users (IVDUs) from the northeastern region of India have a high HIV-1 seroprevalence. Studies carried out in intravenous drug users in the northeastern region of India have shown that HIV-1 subtype C is the predominant strain infecting IVDUs. Fourteen blood samples were collected from HIV-1-infected individuals from the northeastern region of India and screened by env and gag heteroduplex mobility assays (HMA). Where the env and gag HMA results from a sample yielded different subtypes, sequencing of env and gag PCR products was carried out to confirm the presence of HIV-1 recombinants. Of the 14 samples subtyped, nine samples belonged HIV-1 subtype C (gag C/env C), one to HIV-1 subtype B (gag B/env B), and the remaining were B/C recombinants (gag C/env B). This is the first report of HIV-1 B/C recombinants from India.     

Emergence of new forms of human immunodeficiency virus type 1 intersubtype recombinants in central Myanmar

We have previously shown that HIV-1 env subtypes B' (a Thai-B cluster within subtype B) and E (CRF01_AE) are distributed in Yangon, the capital city of Myanmar. However, HIV strains from the rest of country have not yet been genetically characterized. In the present study, we determined env (C2/V3) and gag (p17) subtypes of 25 specimens from central Myanmar (Mandalay). Phylogenetic analyses identified 5 subtype C (20%), in addition to 10 CRF01_AE (40%) and 4 subtype B' (16%). Interestingly, the remaining six specimens (24%) showed discordance between gag and env subtypes; three gag subtype B'/env subtype C, one gag subtype B'/env subtype E, one gag subtype C/env subtype B', and one gag subtype C/env subtype E. These discordant specimens were found frequently among injecting drug users (4 of 12, 33%) and female commercial sex workers (2 of 8, 25%) engaging in high-risk behaviors. The recombinant nature of these HIV-1 strains was verified in three specimens, indicating the presence of new forms of HIV-1 intersubtype C/B' and C/B'/E recombinants with different recombination breakpoints. The data suggest that multiple subtypes of B', C, and CRF01_AE are cocirculating in central Myanmar, leading to the evolution of new forms of intersubtype recombinants among the risk populations exhibiting one of the highest HIV infection rates in the region.     

Inter- and intra-genic intersubtype HIV-1 recombination in rural and semi-urban Uganda.

OBJECTIVE: To investigate the number and variety of viruses with discrepant subtypes between env and gag and within gag in two cohorts in Uganda. METHODS: Sequences were generated from PCR products amplified directly (without cloning) from patient blood and compared in the v3/v4 region of env and the p17 and p24 regions of gag to reference subtype strains by phylogenetic analysis. Gag sequences with a discrepant subtype between p17 and p24 were analysed further to indicate approximate sites of recombination. RESULTS: Envelope subtypes D and A were predominant, but subtypes B, C and G were also found. From analysis of three short regions of the HIV genome we found 15 different combinations of subtype assortment, including 11 different recombinant permutations. Approximately 30% of viruses (29/104) in this part of Uganda appear to be recombinants between the env and gag genes and 10% (11/104) are recombinant within the gag gene. There was no clear pattern of crossover points within the gag gene. There seems to be no evidence of new circulating recombinant forms. CONCLUSION: Both inter-genic and intra-genic inter-subtype recombination appear to be a relatively common occurrence in this geographical region where two subtypes of virus co-circulate. These results have implications for cross-clade vaccine design.