Characterization of B/C recombinants of near full-length HIV type 1 from northeastern India with mosaics identical to ARE195FL but with a different ancestral origin

We have characterized near full-length genomes of three B/C recombinants of HIV-1 from the northeastern state of India. The recombinant viruses showed a backbone of subtype C virus with a single insertion of the subtype B genome in the envelope region. While all of them were distinct from B/C recombinants CRF_07 and CRF_08 circulating in China and CRF_04BR137 circulating in Brazil, two of them presented with break-points identical to the Argentinean B/C recombinant ARE195FL. However, neighbor-joining analysis followed by phylogenetic clustering showed that gp120 belonging to subtype B of all the recombinants clustered with Thai B sequences, while subtype C gag clustered with an Indian subtype C sequence, suggesting a unique ancestral origin of these recombinants.     

Circulation of novel HIV type 1 A, B/C, and F subtypes in Argentina

The Argentine HIV-1 epidemic is considered to be represented mainly by subtype B and diverse B/F recombinants, with apparent absence of pure subtype F. In this study we describe three novel HIV-1 variants isolated from four infants born in different and distant provinces of Argentina. Partial analysis of different gene fragments spanning 18.5-40.8% of the HIV-1 complete genome revealed two subtype A HIV-1 strains in siblings, a B/C recombinant with a novel mosaic structure, and a putative subtype F. Characteristic patterns of genomic and amino acid sequences of the newly reported subtype F isolate suggest a closer genetic relationship to Argentine B/F recombinants than any other subtype F strain described so far, while the A and B/C subtypes found correspond to unusual genotypes in Argentina. Understanding the origin, diversity, and spread of HIV-1 strains worldwide will be necessary for the development of an effective vaccine approach.     

Sequence analysis of a South American HIV type 1 BC recombinant

We report the characterization of a near full-length sequence of a South American HIV-1 BC intersubtype recombinant, ARE195FL. This isolate contains a C(gag/pol) B(vpr/vpu/gp160(env)) C(gp41(env)/nef) mosaic structure differing from any BC full-length sequence described to date, and is the first full-length sequence obtained from a South American BC strain. Neighbor-joining analysis revealed that subtype C genomic segments of ARE195FL clustered with the South American group of subtype C strains, suggesting a local emergence of the BC recombinant. Despite having dissimilar genomic structures, ARE195FL shares a common recombination breakpoint in vif, which is a "hot spot" for recombinatory events, with the Chinese BC recombinant prototype CRF07_BC. Further full-length sequence analysis of South American HIV-1 C and BC strains is necessary to determine the relevance and spread of emerging HIV-1 BC recombinants in Latin America.     

New HIV type 1 CRF01_AE/B recombinants displaying unique distribution of breakpoints from incident infections among injecting drug users in Thailand

The goals of this study were to identify and characterize recombinant human immunodeficiency virus type 1 (HIV-1) genomes among incident infections in a prospective cohort study of injecting drug users (IDUs) in Bangkok, Thailand. Through cross-sectional, comparative phylogenetic analysis of the protease and env (C2-V4) gene regions, subtype discordance was observed in HIV-1 sequences from 4 of 111 IDUs (3.5%). Near-full-length HIV-1 genome sequences of the four strains revealed that in all four, the gp120 sequences clustered with a CRF01_AE prototype, while the remainder of the genomes displayed distinct mosaic patterns, with multiple breakpoints between HIV-1 CRF01_AE and subtype B-like regions. Two of the four HIV-1 recombinant strains displayed a nearly identical mosaic structure, suggesting the possible emergence and spread of a potentially new circulating recombinant form of HIV-1. Further characterization of these and other recombinant genomes through long-term follow-up will be important in understanding the generation of viral diversity and escape from the hosts immune responses. This information will be especially important for vaccine development.     

Identification of unique B/C recombinant strains of HIV-1 in the southern state of Karnataka, India

We characterized the molecular nature of a large number of primary HIV-1 isolates in the four southern states of India. In addition to confirming a predominance of subtype C infection, for the first time we identified three B/C recombinant viruses in a subset of 115 samples. Unexpectedly, env sequences of two of the three B/C recombinants phylogenetically clustered with subtype B strains of the USA. The determination of the real incidence of the recombinant viruses is of importance.     

HIV type 1 BF recombinant strains exhibit different pol gene mosaic patterns: descriptive analysis from 284 patients under treatment failure

Different complex structures of the pol gene have been identified in 284 HIV-1 B/F recombinant sequences obtained from a group of 587 patients under treatment failure from Argentina. To analyze the mosaic structures of these viral sequences and to determine their phylogenetic relationship, the 284 partial pol gene sequences of BF recombinant viruses were amplified by RT-PCR and sequenced. Intersubtype breakpoints were analyzed by bootscanning. Phylogenetic relationships were determined by means of neighbor-joining trees. The analysis of the sequences showed multiple phylogenetic topologies clustering within intersubtype BF reference sequences. At least three different mosaic patterns were found compared to previously described BF-type viruses with unequal distribution in the studied population. The analysis also showed that HIV-1 BF recombinant viruses with diverse mosaic structures are phylogenetically related in their F segments and in selected B fragments with the F1 subtype and with BF recombinant viruses from Brazil, respectively, suggesting a common recombinant ancestor. No association was observed between the prevalence of each mosaic pattern and the frequency of major drug-resistance mutations in PR and RT.     

Full-length gag sequences of HIV type 1 subtype C recent seroconverters from Pune, India

Although HIV-1 subtype C is the most prevalent subtype worldwide, data on subtype C viruses are rather limited. Very little information is available on the complete HIV-1 subtype C gag sequences from India. We report full-length gag (p55) sequences from six Indian early seroconverters. The samples were collected within few weeks of seroconversion and may represent immunologically naive viruses. The comparison of p55 sequences with other Indian and non-Indian subtype C sequences as well as with nonsubtype C sequences obtained from the Los Alamos database revealed gag as a well-conserved region of the HIV genome (range: 84-95%). The phylogenetic tree indicated that the sequences compared here cluster together within clade C. Two epitopes in the p24 region of the gag gene were subtype C specific while many epitopes in the same region were also present in other clades. The data on HIV-1 subtype C full-length gag sequences would be useful in the design and evaluation of effective subtype C-based HIV vaccines.     

Identification of a novel HIV-1 complex circulating recombinant form (CRF18_cpx) of Central African origin in Cuba

BACKGROUND: Analysis of partial pol and env sequences have indicated a high diversity of HIV-1 genetic forms in Cuba, including two potential novel circulating recombinant forms (CRF): U/H and D/A. OBJECTIVES: To determine whether U/H recombinant viruses from Cuba, detected in 7% of samples, represent a novel HIV-1 CRF, and to identify non-Cuban viruses related to this recombinant form. METHODS: Near full-length genome amplification was carried out by nested polymerase chain reaction in four overlapping DNA segments of two epidemiologically unlinked viruses in uncultured peripheral blood mononuclear cells. The sequences were analysed phylogenetically. Recombinant structures and phylogenetic relationships were analysed by bootscanning and by maximum likelihood. Searches for related viruses in databases were initially based on sequence homology and sharing of signature nucleotides. RESULTS: Both Cuban viruses clustered uniformly in bootscans all along the genome with each other and with a virus from Cameroon, CM53379, indicating that all three represent the same recombinant form. Their genome comprised multiple segments clustering with subtypes A1, F, G, H and K, as well as segments failing to cluster with recognized subtypes. The newly defined CRF, designated CRF18_cpx, was phylogenetically related in partial segments to CRF13_cpx, CRF04_cpx and 36 additional viruses, most of them from Central Africa. One of the viruses from Cameroon, sequenced in the near full-length genome, was a CRF18_cpx/subtype G secondary recombinant. CONCLUSIONS: A novel HIV-1 complex circulating recombinant form (CRF18_cpx) has been identified that is circulating in Cuba and Central Africa.     

Highly divergent HIV type 1 group M sequences evident in Karonga District,Malawi in early 1980s

Six divergent HIV-1 partial env and gag genome sequences have been characterized in five subjects in Malawi, from whom blood spot samples were collected between 1982 and 1989, at the time that the AIDS epidemic there was starting. These sequences could not be classified with any of the recognized subtypes or circulating recombinant forms of HIV-1. They showed no consistent and/or supported associations with other subtypes by either env or gag gene phylogenetic analysis. Their genetic distances from defined subtypes suggest that they may be diverse subsubtype C viruses or, alternatively, that they may have mosaic genomes. Bootscanning analyses are consistent with their being mosaic viruses. These sequences highlight early HIV-1 diversity in a population otherwise dominated by subtype C.     

Characterization of five nearly full-length genomes of early HIV type 1 strains in Ruili city: implications for the genesis of CRF07_BC and CRF08_BC circulating in China

To trace the genesis of HIV-1 CRF07_BC and CRF08_BC, two predominant circulating recombinants among intravenous drug users in China, a retrospective molecular epidemiological investigation (1996-1998) was conducted in Ruili city of Yunnan, where the first AIDS epidemic among IDUs was reported in 1989. Fifty-four HIV-1 env C2V3 sequences were determined and genotyped with 49 subtype B and only 5 subtype C strains. The nearly full-length genome analyses of these five env-based subtype C samples revealed that four of them were actually BC recombinants and only one was pure subtype C. The first identified nonrecombinant HIV-1 subtype C, genetically close to Indian C representatives, provided direct evidence for the hypothesis that subtype C in China was introduced from India. Interestingly, three BC recombinants with subtype B as backbones were identified; one BC recombinant that precisely shared a common subtype B segment (nef region) with CRF07_BC and CRF08_BC was described, which indicated a close evolutionary relationship to these two CRFs. The sequences undoubtedly lead us to a better understanding of the emergence of CRF07_BC and CRF08_BC.     

Identification of a complex env subtype E HIV type 1 virus from the democratic republic of congo, recombinant with A, G, H, J, K, and unknown subtypes

Up to now, all known env subtype E viruses (CRF01-AE) have had the same mosaic structure with subtype A, and no other env subtype E HIV-1 viruses with non-A subtypes in their genomes have been described. In this report we describe the full-length genome sequence of an env subtype E isolate with a recombinant genome different from the prototype CRF01-AE strains. The 97CD-KTB49 strain, obtained from a tuberculosis patient in Kinshasa, has a complex mosaic genome involving subtypes A, E, G, H, J, K, and several unknown fragments. The U sequences formed well-separated clusters together with previously described unknown fragments from CRF04-cpx (subtype I), and from Z321, the oldest intersubtype recombinant isolated in 1976 in the Democratic Republic of Congo. The complex recombinant virus from our study is not an isolated strain; partial sequencing of a second strain, 97CD-KFE45, confirmed the breakpoints observed in the 97CD-KTB49 strain in the regions sequenced. The complexity of these recombinant strains suggests a longstanding presence of subtype E in Central Africa.     

Near full-length clones and reference sequences for subtype C isolates of HIV type 1 from three different continents

Among the major circulating HIV-1 subtypes, subtype C is the most prevalent. To generate full-length subtype C clones and sequences, we selected 13 primary (PBMC-derived) isolates from Zambia, India, Tanzania, South Africa, Brazil, and China, which were identified as subtype C by partial sequence analysis. Near full-length viral genomes were amplified by using a long PCR technique, sequenced in their entirety, and phylogenetically analyzed. Amino acid sequence analysis revealed 10.2, 6.3, and 17.3% diversity in predicted Gag, Pol, and Env protein sequences. Ten of 13 viruses were nonmosaic subtype C genomes, while all three isolates from China represented B/C recombinants. One of them was composed primarily of subtype C sequences with three small subtype B portions in gag, pol, and nef genes. Two others exhibited these same mosaic regions, but contained two additional subtype B portions at the gag/pol overlap and in the accessory gene region, suggesting ongoing B/C recombination in China. All subtype C genomes contained a prematurely truncated second exon of rev, but other previously proposed subtype C signatures, including three potential NF-kappa B-binding sites in the viral promoter-enhancer regions, were found in only a subset of these genomes.     

HIV type 1 strains common in Europe, Africa, and Asia cocirculate in Yemen

To determine the HIV-1 genetic diversity in Yemen, 19 strains collected from men and women were sequenced in pol and six of those were full genome sequenced; all were phylogenetically analyzed. Using the pol sequence data, nine (47.3%) were subtype B, six (31.6%) subtype C, two (10.5%) subtype D, one strain (5.3%) subtype A, and another (5.3%) a unique recombinant form (URF). Concordant phylogenies were also obtained for the six strains full genome sequenced. Most of the strains were from the capitol, Sana'a (n=16). Five of the C strains clustered with African Cs, and one clustered with the Indian C strains. Of the two subtype D strains, one clustered with Ugandan strains and one with Cameroon. The subtype A strain was similar to a Cameroon variant of subtype A and the URF strain was a recombinant between CRF11, CRF13, and subtype B. The HIV epidemic in Yemen is extremely complex, with strains of HIV-1 that have originated in East and West Africa, Europe, and India.     

Identification of new CRF43_02G and CRF25_cpx in Saudi Arabia based on full genome sequence analysis of six HIV type 1 isolates

Recently, we reported a high level of HIV-1 strain diversity in patients at the King Faisal Specialist Hospital and Research Center, Jeddah, Saudi Arabia. Based on phylogenetic analysis of gag p24, pol integrase, and env gp41 sequences, subtypes A, B, C, D, and G, and CRF02_AG, as well as unique recombinant forms were identified. Subtype G accounted for 25% of the infections in the Saudi population and this high prevalence was unexpected. Although subtype G is found in west central Africa, pure subtype G strains are uncommon. To further characterize the subtype G infections in Saudi Arabia, six strains that appeared to be pure subtype G were selected for full genome sequencing. Near full-length genomes were obtained using RT-PCR amplification to generate overlapping fragments from viral RNA extracted from plasma. The six strains are not subtype G throughout their entire genome. Four isolates have a recombinant structure composed of CRF02_AG and subtype G and share three identical breakpoints. This recombinant form defines a new CRF designated CRF43_02G. The remaining two isolates are CRF25_cpx, a circulating recombinant form identified in Cameroon composed of subtypes A and G and unclassified segments. Reanalysis of the previously reported Saudi HIV-1 partial genome sequences revealed additional isolates classified as CRF43_02G and CRF25_cpx and one isolate was reclassified to CRF22_01A. Identification of CRF43_02G in Saudi Arabia could indicate a transmission network within the country. Alternatively, the new CRF could have been introduced from an external source where this CRF is not yet recognized.     

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.     

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.