Genetic characterization of HIV type 1 and type 2 from Bissau, Guinea-Bissau (West Africa)

Previous studies from Guinea-Bissau (West Africa) have demonstrated a unique epidemiology with respect to both HIV-1 and HIV-2 infection. In order to evaluate the prevalence and dynamics of HIV-1 and HIV-2 subtypes in Bissau, the capital city of Guinea-Bissau, a cross-sectional study was set up using serological and molecular techniques. Plasma samples from 103 individuals were screened for HIV-1 and HIV-2 antibodies by ELISA and Western-blot. Seropositive results were confirmed by PCR amplification of proviral sequences in primary peripheral blood mononuclear cells (PBMC) with env and LTR primer sets for HIV-2 and env, LTR and pol primers for HIV-1. A total of 38/103 individuals were HIV-seroreactive (four HIV-1, 15 HIV-2, 19 HIV-1/HIV-2). A total of eight out of 19 dually seropositive specimens showed double PCR amplification of HIV-1 and HIV-2 proviral sequences, accounting for 21% of the infected individuals. In the remaining 11 individuals either HIV-2 or HIV-1 sequences were detected, the majority (n=9) amplifying only HIV-2. These screening data demonstrate a high discrepancy between serology and PCR results for dually seroreactive samples, Western-blot giving an overestimation of double infection. Additionally, HIV-1 strains were subtyped by heteroduplex mobility assay (HMA) on the basis of gp120 sequences. Subtyping of HIV-2 was carried out by DNA sequencing and phylogenetic analysis of env V3 molecular clones. For both HIV-1 and HIV-2 strains circulating in Bissau, our results indicate dominance of subtype A.     

Determination of human immunodeficiency virus type 1 subtypes in Taiwan by vpu gene analysis

The genetic diversity of human immunodeficiency virus (HIV) type 1 (HIV-1) has been characterized mainly by analysis of the env and gag genes. Information on the vpu genes in the HIV sequence database is very limited. In the present study, the nucleotide sequences of the vpu genes were analyzed, and the genetic subtypes determined by analysis of the vpu gene were compared with those previously determined by analysis of the gag and env genes. The vpu genes were amplified by nested PCR of proviral DNA extracted from 363 HIV-1-infected individuals and were sequenced directly by use of the PCR products. HIV-1 subtypes were determined by sequence alignment and phylogenetic analysis with reference strains. The strains in all except one of the samples analyzed could be classified as subtype A, B, C, E, or G. The vpu subtype of one strain could not be determined. Of the strains analyzed, genetic subtypes of 247 (68.0%) were also determined by analysis of the env or gag gene. The genetic subtypes determined by vpu gene analysis were, in general, consistent with those determined by gag and/or env gene analysis except for those for two AG recombinant strains. All the strains that clustered with a Thailand subtype E strain in the vpu phylogenetic analyses were subtype E by env gene analysis and subtype A by gag gene analysis. In summary, our genetic typing revealed that subtype B strains, which constituted 73.8% of all strains analyzed, were most prevalent in Taiwan. While subtype E strains constituted about one-quarter of the viruses, they were prevalent at a higher proportion in the group infected by heterosexual transmission. Genetic analysis of vpu may provide an alternate method for determination of HIV-1 subtypes for most of the strains, excluding those in which intersubtype recombination has occurred.     

Evaluation of commercially available assays for antibodies to HIV-1 in serum obtained from South African patients infected with HIV-1 subtypes B,C, and D

Between 1984 and 1990, virus was routinely isolated and serum collected from patients diagnosed at hospitals in the Western Cape as suffering from AIDS or AIDS-related conditions (ARC). From these, 17 virus strains were selected at random for sequencing and molecular characterisation of the env gene. The strains were previously characterised as belonging to HIV-1 subtypes B, C and D. The purpose of the present study was to evaluate retrospectively the serological diagnosis of HIV-1 in these 17 South African patients. Thirteen anti-HIV screening assays, including 7 rapid/simple test devices (RTDs), 4 enzyme-linked immunosorbent assays (ElAs) and 2 Western immunoblot assays were evaluated. Using commercial ElAs, 16 serum samples were HIV antibody-positive and these results were confirmed by Western immunoblot analysis. Serum from one terminal AIDS patient was found negative with all the serological tests. Some RTDs gave false negative antibody reactions on specimens from patients infected with subtype D strains. To investigate the false negative antibody reactions, the polymerase chain reaction (PCR) was used to amplify, clone and sequence proviral DNA from the immunodominant gp41 region from 7 of the HIV-1 strains. Two patients, both subtype D strains (D214 and D482) with false negative results in the RTDs, showed a significant amino acid substitution, i.e., substitution of a histidine residue for leucine at env position 607. It was concluded that although there were false negative RTD reactions on patients with HIV-1 subtype D strains, the commercial ElAs tested are sensitive and are able to detect patients infected with HIV-1 subtypes B, C and D that are present in South Africa. © 1994 Wiley-Liss, Inc.     

Genetic characteristics of variants of human immunodeficiency virus type 1, causing an epidemic among substance abusers in Commonwealth of Independent States countries]

Gag/env nucleotide sequences of human immunodeficiency virus type 1 (HIV-1) variants detected in drug users in Russia, Ukraine, and Belarus are analyzed. Two HIV-1 subtypes A and B circulate in this risk group. Genetic variability within one subtype is no higher than 3.1 and 3.9% for gag and env genes, respectively, suggesting the same source of infection in populations of drug users infected with the same subtype. Recombinant viruses with gagAenvB genotype, genetically related to parental strains of subtypes A and B, circulate in this risk group. This is the first report about HIV-1 recombinant of the two subtypes, for which both parental strains are known, directly confirming the in vivo recombination between different subtypes.     

Near full-length genomic characterization of a HIV type 1 BC recombinant strain from Manipur, India

Genetic complexity of HIV-1 is brought about by recombination between HIV-1 subtypes which leads to the development of epidemiologically significant founder strains. In the present study, the near full-length genome sequence of an HIV-1 isolate from an injecting drug user of Manipur (India) was determined, which evidenced the presence of a novel HIV-1 BC recombinant strain. Near full-length genome was amplified by polymerase chain reaction using primer walking approach. The recombination break points were detected using bootscan and simplot analyses. This isolate exhibited a mosaic structure consisting of subtype C backbone with subtype B insertions at the upstream of pol gene (3026-3259) and the downstream of env gene which spanned till the nef gene (8183-8961). Phylogenetic relationships determined with neighbor-joining trees, revealed that the subtype C sequences clustered with sequences from Indian subtype C HIV-1 strains, and the subtype B sequences clustered with HIV-1 subtype B strains from Thailand. This finding may create a complex scenario of HIV-1 epidemic among the injecting drug users of Manipur in near future.     

Molecular epidemiology of HIV-1 in Switzerland: evidence for a silent mutation in the C2V3 region distinguishing intravenous drug users from homosexual men. Swiss HIV Cohort Study

OBJECTIVES: To study the molecular epidemiology of HIV-1 strains found in Switzerland and to determine possible genetic linkages among strains sorted by risk group or geographic region. DESIGN: A cross-sectional, clinic-based survey of HIV-1 molecular sequences and linked patient history from Swiss people. METHODS: Specimens were collected from 215 HIV-1-infected people in HIV outpatient clinics of four tertiary referral centers (Lausanne, St. Gallen, Zurich, and Basel) between May and August 1996, mainly from homosexual men, injecting drug users (IDU), and heterosexually infected people. In addition, specimens collected between 1991 and 1995 in the HIV outpatient clinic at University of Geneva were included into this survey. These specimens were collected primarily for an ongoing, prospective cohort (Swiss HIV Cohort Study). Direct C2V3C3 sequences of the env gene were determined from 158 samples of peripheral blood mononuclear cells. Genetic data were analyzed with the available patient history on each specimen. RESULTS: As found in other previous studies in Europe, primarily subtype B viruses were identified, whereas seven (4%) of 158 were non-subtype B: one subtype D, four subtype A, and two subtype E. Five of seven non-B subtypes occurred in immigrants from African or Asian countries and all seven were found exclusively in individuals who had been infected by heterosexual contact. No significant clustering of strains within different study sites or risk groups was found. A silent mutation (LAI env 834) occurred significantly more often in IDU than in homosexual men (p<.001). CONCLUSIONS: Although the lack of significant clustering of strains by risk group or geographic region may result from early introduction of subtype B viruses in Switzerland, the strong association of a silent mutation with IDU suggests that, early in the epidemic, there was a unique founder virus among IDUs. The HIV epidemic in Switzerland is still predominantly caused by subtype B viruses.     

Discrimination of subtype B and non-subtype B strains of human immunodeficiency virus type 1 by serotyping: correlation with genotyping

The ability of a peptide-based serotyping assay to differentiate human immunodeficiency virus (HIV) type 1 (HIV-1) subtype B infections from non-subtype B infections was investigated with 166 anti-HIV-1- and HIV RNA-positive (by PCR) serum or plasma specimens. The specimens were divided genetically into those infected with subtype B and non-subtype B by application of a screening heteroduplex mobility assay (HMA) that used plasmids for subtypes A and B alone. Specimens that were not clearly infected with HIV-1 subtype B by HMA or for which the two methods had discordant results in distinguishing those infected with subtype B from those infected with non-subtype B were then investigated with a full HMA plasmid panel and, for selected specimens, env sequencing. For the 141 genotyped and serotypically reactive specimens, the correlation between genotyping and serotyping (all subtypes) was 69%. Of the 67 specimens that reacted monotypically as serotype B, 64 were shown to be infected with genotype B (positive predictive value, 96%). Of the 82 specimens that contained genotype B nucleic acid, 64 reacted monotypically as serotype B (sensitivity, 78%), and 4 specimens reacted with a single non-subtype B peptide; the viruses in 14 specimens could not be assigned a serotype. Initial screening results had indicated that 12 samples had results discordant between restricted HMA and serotyping. The V3 loop amino acids of the infecting HIV strains from the seven specimens with discordant serology results were analyzed. For five specimens discordance occurred when the amino acid sequence of the infecting virus closely resembled those of more than one consensus peptide antigen or when the observed V3 crown motif of the strain was atypical for the genetic subtype present. For the other two specimens no explanation for the discordance was identified. Five specimens gave unclear or discordant results in the initial HMA screen, but the results were resolved when the full plasmid panel was used. Serotyping, although of limited sensitivity, distinguishes between subtype B and non-subtype B infections with a high degree of specificity. However, it poorly differentiates the major non-subtype B subtypes, particularly subtypes A and C. When HIV-1 subtype B predominates, serological typing and/or subtype-restricted HMA screening usefully distinguishes between subtype B and non-subtype B infections.     

High genetic diversity of HIV-1 strains in Chad,West Central Africa

The genetic diversity of HIV-1 strains in Chad was documented with a total of 107 samples from patients attending the general hospital in N'Djamena, the capital city of Chad. The genetic subtypes were identified in the V3-V5 env and p24 gag regions by sequence and phylogenetic tree analyses. Of the 107 strains, 78 had the same subtype/CRF designation between env and gag. Four subtypes and three CRFs were found to cocirculate: subtype A, 20.5%; subtype D, 18.7%; CRF02_AG, 13.1%; CRF11_cpx, 13.1%; subtype G, 3.7%; CRF01_AE, 2.8%; and subtype F1, 0.9%. The remaining 29 strains (27%) had discordant subtypes or CRF designations between env and gag; in 15 of these 29 strains, a CRF was involved in the recombination event, and 10 were subtype G in gag and subtype A in env, forming a separate subcluster within subtypes G and A. Subtype D strains represent almost 20% of the HIV-1 strains circulating in Chad and form a separate subcluster in gag and env. Nearly full-length genome sequencing for two such strains (99TCD-MN011 and 99TCD-MN012) revealed that they represent nonrecombinant subtype D variants. Compared with neighboring countries, the genetic subtype distribution of HIV-1 strains in Chad is unique for several reasons: lower prevalence of CRF02, high prevalence of CRF11 and subtype D, and absence of CRF06. These data clearly show that subtype distribution is very heterogeneous in Africa, probably the result of different founder effects.     

Segregation of human immunodeficiency virus type 1 subtypes by risk factor in Australia

The aim of this study was to determine which human immunodeficiency virus type 1 (HIV-1) subtypes were circulating in Australia and to correlate the subtypes with risk factors associated with the acquisition of HIV-1 infection. DNA was extracted from peripheral blood mononuclear cells, and HIV-1 env genes were amplified and subtyped using heteroduplex mobility analysis, with selected samples sequenced and phylogenetic analysis performed. The HIV-1 env subtypes were determined for 141 samples, of which 40 were from female patients and 101 were from male patients; 13 samples were from children. Forty-seven patients were infected by homosexual or bisexual contact, 46 were infected through heterosexual contact, 21 were infected from injecting drug use (IDU), 13 were infected by vertical transmission, 8 were infected from nosocomial exposure, and 6 were infected by other modes of transmission, including exposure to blood products, ritualistic practices, and two cases of intrafamilial transmission. Five subtypes were detected; B (n = 104), A (n = 5), C (n = 17), E (CRF01_AE; n = 13), and G (n = 2). Subtype B predominated in HIV-1 acquired homosexually (94% of cases) and by IDU (100%), whereas non-subtype B infections were mostly seen in heterosexually (57%) or vertically (22%) acquired HIV-1 infections and were usually imported from Africa and Asia. Subtype B strains of group M viruses predominate in Australia in HIV-1 transmitted by homosexual or bisexual contact and IDU. However, non-B subtypes have been introduced, mostly acquired via heterosexual contact.     

Presence of multiple HIV subtypes and a high frequency of subtype chimeric viruses in heterosexually infected women

The HIV-1 subtype distribution was determined in 41 HIV-positive women (-8% of all HIV-infected women in Denmark) belonging to different risk groups. HIV p17 gag and env gene subtypes were determined by DNA sequence analysis. Five different HIV subtypes were detected across all patients. Most HIV-1-positive women of Danish origin carried subtype B viruses, and a minority had virus belonging to subtypes A or C. All injecting drug users (IDUs) were infected with HIV subtype B viruses, whereas all non-B subtypes were present in cases linked to heterosexual transmission. In contrast, all women of African origin carried non-B HIV subtypes (subtypes A, C, D, or G) regardless of transmission mode. Of these women, 21% infected with non-B HIV subtypes appeared to be infected by subtype chimeric viruses and 7% were jointly infected with viruses belonging to two different subtypes (A and C). Data demonstrate a preferential representation of non-B HIV subtypes in women infected through heterosexual contact, as well as a high degree of recombination between viruses derived from endemic areas in which several HIV subtypes predominate. Combined with the increased incidence of heterosexual transmission of HIV, the results imply that an increased subtype diversity can be anticipated in newly infected individuals.     

Molecular-epidemiological characteristics of basic foci of the HIV epidemic among substance abusers in Russia]

A serological test has been developed, allowing a rapid and reliable differentiation between envA and envB genotypes in the sera from HIV-1-infected IDUs and their sexual partners. We analyzed 556 specimens from HIV-1-seropositive IDUs and their sexual partners. The sera were collected in five regions of Russia with the greatest number of HIV-1-infected IDUs. High homogeneity of HIV-1 in each of the regions was observed. In Tver, Rostov-on-Don, Nizhny Novgorod, and Krasnodar 100% sera tested belonged to the env subtype A. All the patients from the Kalinigrad region were infected with the env subtype B virus, which is apparently a recombinant.     

淮安市经异性性传播的HIV感染者中HIV-1病毒基因亚型特征分析

目的 了解淮安市经异性性传播HIV感染者中HIV-1病毒基因亚型分布及其流行特征.方法 从60名异性性传播HIV-1感染者血液中提取DNA或RNA,用巢式PCR或RT-PCR方法扩增gag、env基因区片段,测定序列并分析.结果 60份标本中,确定了48份标本的基因亚型,发现有4种HIV-1亚型和重组型.其中CRF01_AE亚型占62.5％,CRF07_BC亚型占22.9％,CRF08_BC亚型占6.3％,B亚型占6.3％.结论淮安市异性性传播感染HIV者中,流行着CRF01_AE、CRF07_BC、CRF08_BC、B亚型这4种亚型病毒株,CRF01_AE为主要流行亚型.     

Description of a "trans-Saharan" strain of human T-lymphotropic virus type 1 in West Africa

The aim of this study was to assess the prevalence and the molecular epidemiology of human T-lymphotropic virus type 1 (HTLV-1) in a group of pregnant women living in Guinea Bissau. We studied 427 consecutive pregnant women attending 10 centers for HIV-1 infection monitoring in Bissau. HTLV-1 infection was found in 2.6% of the patients. Phylogenetic analysis of the long terminal repeat region showed that 10 isolates were of the cosmopolitan subtype (HTLV-1a) and that only 1 was of the widespread Central African subtype (HTLV-1b). All the cosmopolitan isolates belonged to the HTLV-1aD subgroup, which was first described in North Africa and clustered with other Senegal and Guinea isolates to form a significant West African clade. Our data show a high prevalence of HTLV-1 in Guinea Bissau and suggest the existence of a trans-Saharan strain distributed in North and West Africa, which probably crossed the desert in the past as a result of contacts between nomadic and sedentary populations or along trading routes.     

First molecular characterization of HIV-1 Tunisian strains

In order to identify HIV-1 subtypes circulating in Tunisia, blood specimens from 25 HIV-1 infected Tunisian patients were collected. Proviral HIV-1 DNA was genotyped by sequence analysis of the C2-V3 env region. HIV-1 subtypes were determined in 21 DNA sequences: 20 were of subtype B and one was a circulating recombinant form (CRF02 AG). Subtype B largely dominates the epidemiology of HIV-1 infection in Tunisia, suggesting the probable imported origin of HIV-1 infection, but further studies are needed to investigate the potential diversification of HIV-1 isolates in Tunisia.     

我国西南西北地区吸毒人群重组人类免疫缺陷病毒1？…

目的 寻找人类免疫缺陷病毒１型在中国可能的重组。方法从流行２种以上ＨＩＶ－１亚型的地区收集ＨＩＶ感染者的血样。从ＰＭＣｓ中应用套式聚合酶链反应方法,对ＨＩＶ病毒的ｔａｔ和ｅｎｖ基因进行扩增,ＰＣＲ产物直接测序并进行序列分析。结果 对中国Ｂ’亚型和Ｃ亚型浒区域收集的１４个ＨＩＶ－１毒株进行序列分析,对ｅｎｖ基因进行测序后没有发现重组毒株的证据。     

Plasma RNA viral load in human immunodeficiency virus type 2 subtype A and subtype B infections

Human immunodeficiency virus type 2 (HIV-2) is much less pathogenic than HIV-1, and HIV-2 infection is associated with plasma viral loads significantly lower than those found in HIV-1 infection. We have developed a real-time quantitative PCR method for measuring the HIV-2 RNA load that covers the range of genetic diversity of HIV-2 isolates and that detects extremely low viral loads. Samples from 49 patients were studied. Proviral DNA was first detected and quantified. The strains that were detected were then genotyped: 21 patients were infected with HIV-2 subtype A and 15 patients were infected with HIV-2 subtype B; 1 patient was infected with a highly divergent strain. Env PCR failed for the remaining 12 patients, so subtypes could not be determined. For viral RNA quantification, a stock of HIV-2 strain NIHZ, which was counted by electron microscopy, was used as the standard. Several primer sets targeting the highly conserved gag region were evaluated. Various primer combinations failed to amplify subtype B strains. With the final primer pair selected, which detected both subtype A and subtype B strains, the sensitivity of the assay was 100% at a viral load of 250 copies/ml and 66% at a viral load of 125 copies/ml. We found a correlation between the CD4(+)-cell count, the clinical stage, and the plasma HIV-2 RNA level. The median plasma HIV-2 RNA value for the 33 asymptomatic patients was 2.14 log(10), whereas it was 3.1 log(10) for the 16 patients with AIDS (P < 0.01). Proviral DNA was detectable in 18 symptom-free patients with high CD4(+)-cell counts, in whom viral RNA was undetectable.     

HIV-2 subtype circulating in India (south)

The genetic types and subtypes of HIV vary from region to region. This study looked at the relative prevalence of HIV-2 subtypes in south India. Nucleotide sequencing of the HIV-2 envelope V3 region of strains from 11 individuals infected with HIV-2 and one individual who had dual infection with HIV-1 and HIV-2 was performed. Phylogenetic analysis showed that all individuals were infected with HIV-2 subtype A. These strains were from individuals who live in different regions of south India. In all, up to present, 17 strains inclusive of the authors' 12 have been sequenced, and subtype A of HIV-2 is seen in both monoinfections and dual infections with HIV-1 in India.     

Genetic diversity of HIV type 1 in rural eastern Cameroon

To monitor the presence of genotypic HIV-1 variants circulating in eastern Cameroon, blood samples from 57 HIV-1-infected individuals attending 3 local health centers in the bordering rural villages with Central African Republic (CAR) were collected and analyzed phylogenetically. Out of the 40 HIV-1 strains with positive polymerase chain reaction (PCR) profile for both gag and env-C2V3,12 (30.0%) had discordant subtype or CRF designation: 2 subtype B/A (gag/env), 1 B/CRF01, 2 B/CRF02, 1 CRF01/CRF01.A, 2 CRF11/CRF01, 1 CRF13/A, 1 CRF13/CRF01, 1 CRF13/CRF11, and 1 G/U (unclassified). Twenty-eight strains (70.0%) had concordant subtypes or CRF designation between gag and env: 27 subtype A and 1 F2. Out of the remaining 17HIV-1 strains negative for PCR with the env-C2V3 primers used, 10 (58.8%) had discordant subtype or CRF, and 7 (41.2%) had concordant one based on gag/pol/env-gp41 analysis. Altogether, a high proportion (22/57, 38.6%) of the isolates were found to be recombinant strains. In addition, an emergence of new forms of HIV-1 strains, such as subtype B/A (gag/env), B/CRF01 and B/CRF02, was identified. The epidemiologic pattern of HIV-1 in eastern Cameroon, relatively low and high prevalence of CRF02 and CRF11, respectively, was more closely related to those of CAR and Chad than that of other regions of Cameroon, where CRF02 is the most predominant HIV-1 strain. These findings strongly suggest that this part of Cameroon is a potential hotspot of HIV-1 recombination, with a likelihood of an active generation of new forms of HIV-1 variants, though epidemiologic significance of new HIV-1 forms is unknown.     

Two HIV-1 epidemics in Argentina: different genetic subtypes associated with different risk groups

This study determined the risk behaviors and viral subtypes of HIV-1 found in 134 heterosexual HIV-seroprevalent maternity patients, 41 of their sexual partners (men who have sex with women [MSW]), and 95 homosexual men (men who have sex with men [MSM]) from Buenos Aires, Argentina. Peripheral blood mononuclear cells (PBMCs) were purified from blood and used for DNA extraction, amplification, and genotyping by the envelope heteroduplex mobility assay (env HMA).Most of the women had been infected by having sex with an already infected partner (84%), whereas most of the male partners had been infected via drug use (76%). Both the patients and their sexual partners were poorly educated, only 30% having completed secondary school. The MSM study subjects, however, were significantly better educated and had a lower prevalence of injecting drug use.Env HMA subtype F was found in 77% (103 of 134) of the maternity patients, with similar rates in their partners (73%). Most of the remaining samples were env subtype B. All but one of the couples was concordant in subtype. In the MSM risk group, however, only 10% were env HMA subtype F. Ninety percent of the MSM samples were subtype B.There are at least two independent epidemics of HIV-1 infection in Buenos Aires, Argentina. One, in heterosexual men and women, is dominated by env subtype F whereas the other, in homosexual men, is dominated by env subtype B, as determined by env HMA.