Human immunodeficiency virus type 1 env sequences from Calcutta in eastern India: identification of features that distinguish subtype C sequences in India from other subtype C sequences

India is experiencing a rapid spread of human immunodeficiency virus type 1 (HIV-1), primarily through heterosexual transmission of subtype C viruses. To delineate the molecular features of HIV-1 circulating in India, we sequenced the V3-V4 region of viral env from 21 individuals attending an HIV clinic in Calcutta, the most populous city in the eastern part of the country, and analyzed these and the other Indian sequences in the HIV database. Twenty individuals were infected with viruses having a subtype C env, and one had viruses with a subtype A env. Analyses of 192 subtype C sequences that included one sequence for each subject from this study and from the HIV database revealed that almost all sequences from India, along with a small number from other countries, form a phylogenetically distinct lineage within subtype C, which we designate C(IN). Overall, C(IN) lineage sequences were more closely related to each other (level of diversity, 10.2%) than to subtype C sequences from Botswana, Burundi, South Africa, Tanzania, and Zimbabwe (range, 15.3 to 20.7%). Of the three positions identified as signature amino acid substitution sites for C(IN) sequences (K340E, K350A, and G429E), 56% of the C(IN) sequences contained all three amino acids while 87% of the sequences contained at least two of these substitutions. Among the non-C(IN) sequences, all three amino acids were present in 2%, while 22% contained two or more of these amino acids. These results suggest that much of the current Indian epidemic is descended from a single introduction into the country. Identification of conserved signature amino acid positions could assist epidemiologic tracking and has implications for the development of a vaccine against subtype C HIV-1 in India.     

Origin and dynamics of HIV-1 subtype C infection in India

Objective

To investigate the geographical origin and evolution dynamics of HIV-1 subtype C infection in India.

Design

Ninety HIV-1 subtype C env gp120 subtype C sequences from India were compared with 312 env gp120 reference subtype C sequences from 27 different countries obtained from Los Alamos HIV database. All the HIV-1 subtype C env gp120 sequences from India were used for the geographical origin analysis and 61 subtype C env gp120 sequences with known sampling year (from 1991 to 2008) were employed to determine the origin of HIV infection in India.

Methods

Phylogenetic analysis of HIV-1 env sequences was used to investigate the geographical origin and tMRCA of Indian HIV-1 subtype C. Evolutionary parameters including origin date and demographic growth patterns of Indian subtype C were estimated using a Bayesian coalescent-based approach under relaxed molecular clock models.

Findings

The majority of the analyzed Indian and South African HIV-1 subtype C sequences formed a single monophyletic cluster. The most recent common ancestor date was calculated to be 1975.56 (95% HPD, 1968.78–1981.52). Reconstruction of the effective population size revealed three phases of epidemic growth: an initial slow growth, followed by exponential growth, and then a plateau phase approaching present time. Stabilization of the epidemic growth phase correlated with the foundation of National AIDS Control Organization in India.

Interpretation

Indian subtype C originated from a single South African lineage in the middle of 1970s. The current study emphasizes not only the utility of HIV-1 sequence data for epidemiological studies but more notably highlights the effectiveness of community or government intervention strategies in controlling the trend of the epidemic.     

Molecular cloning and phylogenetic analysis of human immunodeficiency virus type 1 subtype C: a set of 23 full-length clones from Botswana

To better understand the virological aspect of the expanding AIDS epidemic in southern Africa, a set of 23 near-full-length clones of human immunodeficiency virus type 1 (HIV-1) representing eight AIDS patients from Botswana were sequenced and analyzed phylogenetically. All study viruses from Botswana belonged to HIV-1 subtype C. The interpatient diversity of the clones from Botswana was higher than among full-length isolates of subtype B or among a set of full-length HIV-1 genomes of subtype C from India (mean value of 9. 1% versus 6.5 and 4.3%, respectively; P < 0.0001 for both comparisons). Similar results were observed in all genes across the entire viral genome. We suggest that the high level of HIV-1 diversity might be a typical feature of the subtype C epidemic in southern Africa. The reason or reasons for this diversity are unclear, but may include an altered replication efficiency of HIV-1 subtype C and/or the multiple introduction of different subtype C viruses.     

Phenotypic and genotypic characteristics of human immunodeficiency virus type 1 from patients with AIDS in northern Thailand.

Primary human immunodeficiency virus type 1 (HIV-1) isolates were obtained from 22 patients with AIDS from northern Thailand, where HIV-1 is transmitted primarily through the heterosexual route. Viral sequences were determined for the 22 patients with AIDS, and all were subtype E HIV-1 on the basis of sequence analysis of a region from the envelope protein gp120. Syncytium-inducing (SI) viruses were detected for 16 of 22 patients with AIDS by using MT-2 cells. Characteristics of amino acid sequences in V3 which have not been reported previously for subtype B SI HIV-1 were associated with the subtype E HIV-1 SI phenotype. The SI viruses from our study population contain predominantly a GPGR or GPGH motif at the tip of the V3 loop, in contrast to the previously described subtype E HIV-1 from Thailand which contained predominantly GPGQ. All the SI viruses lost a potential N-linked glycosylation site in V3 which is highly conserved among previously described subtype E HIV-1 isolates from asymptomatic patients from Thailand. HIV-1 envelope sequences including V3 from some patients with AIDS were significantly more divergent than viruses from asymptomatic patients in Thailand characterized 2 years ago or earlier. These results suggest that emergence of subtype E SI HIV-1 variants is associated with the development of AIDS, as it is for subtype B HIV-1. The divergence of subtype E HIV-1 in patients with AIDS as the disease progresses, and the divergence of subtype E HIV-1 in the infected population as the epidemic continues in Thailand, may have important implications for vaccine development.     

Full-Length Human Immunodeficiency Virus Type 1 Genomes from Subtype C-Infected Seroconverters in India, with Evidence of Intersubtype Recombination

The development of an effective human immunodeficiency virus type 1 (HIV-1) vaccine is likely to depend on knowledge of circulating variants of genes other than the commonly sequenced gag and env genes. In addition, full-genome data are particularly limited for HIV-1 subtype C, currently the most commonly transmitted subtype in India and worldwide. Likewise, little is known about sequence variation of HIV-1 in India, the country facing the largest burden of HIV worldwide. Therefore, the objective of this study was to clone and characterize the complete genome of HIV-1 from seroconverters infected with subtype C variants in India. Cocultured HIV-1 isolates were obtained from six seroincident individuals from Pune, India, and virtually full-length HIV-1 genomes were amplified, cloned, and sequenced from each. Sequence analysis revealed that five of the six genomes were of subtype C, while one was a mosaic of subtypes A and C, with multiple breakpoints in env, nef, and the 3′ long terminal repeat as determined by both maximal χ² analysis and phylogenetic bootstrapping. Sequences were compared for preservation of known cytotoxic T lymphocyte (CTL) epitopes. Compared with those of the HIV-1_LAI sequence, 38% of well-defined CTL epitopes were identical. The proportion of nonconservative substitutions for Env, at 61%, was higher (P < 0.001) than those for Gag (24%), Pol (18%), and Nef (32%). Therefore, characterized CTL epitopes demonstrated substantial differences from subtype B laboratory strains, which were most pronounced in Env. Because these clones were obtained from Indian seroconverters, they are likely to facilitate vaccine-related efforts in India by providing potential antigens for vaccine candidates as well as for assays of vaccine responsiveness.     

Fighting a virus with a virus: a dynamic model for HIV-1 therapy

A mathematical model examined a potential therapy for controlling viral infections using genetically modified viruses. The control of the infection is an indirect effect of the selective elimination by an engineered virus of infected cells that are the source of the pathogens. Therefore, this engineered virus could greatly compensate for a dysfunctional immune system compromised by AIDS. In vitro studies using engineered viruses have been shown to decrease the HIV-1 load about 1000-fold. However, the efficacy of this potential treatment for reducing the viral load in AIDS patients is unknown. The present model studied the interactions among the HIV-1 virus, its main host cell (activated CD4+ T cells), and a therapeutic engineered virus in an in vivo context; and it examined the conditions for controlling the pathogen. This model predicted a significant drop in the HIV-1 load, but the treatment does not eradicate HIV. A basic estimation using a currently engineered virus indicated an HIV-1 load reduction of 92% and a recovery of host cells to 17% of their normal level. Greater success (98% HIV reduction, 44% host cells recovery) is expected as more competent engineered viruses are designed. These results suggest that therapy using viruses could be an alternative to extend the survival of AIDS patients.     

Human immunodeficiency virus type-1 subtypes of infected patients in Espírito Santo, Brazil

Genetic variability of human immunodeficiency virus type-1(HIV-1) is a potential threat for both diagnosis and treatment of HIV/AIDS, as well as the development of effective vaccines. Up to now, HIV subtypes circulating among HIV-positive patients in the state of Espírito Santo were not known. In the present study, blood samples from 100 therapy-na?ve HIV-1 infected patients were collected and the HIV subtype was determined through the Heteroduplex Mobility Assay (HMA). Ninety-seven out of 100 studied samples were subtyped by HMA, 73 samples (75.2%) were from subtype B, 9 (9.3%) from subtype F, 3 (3.1%) from subtype C, 6 (6.2%) Benv/Fgag, and another 6 (6.2%) Fenv/Bgag, what suggests that recombinant viruses were present in the studied samples. Twenty-eight percent of the subtype B samples were represented by the Brazilian B" subtype, which were identified by RFLP with Fok I. Data presented here demonstrate that the epidemiological characteristics of the HIV epidemic in the state of Espírito Santo are similar to those from the other Southeastern states and helped to better understand the genetic polymorphism of HIV in Brazil.     

Identification of human immunodeficiency virus type 1 subtype C Gag-, Tat-, Rev-, and Nef-specific elispot-based cytotoxic T-lymphocyte responses for AIDS vaccine design

The most severe human immunodeficiency virus type 1 (HIV-1) epidemic is occurring in southern Africa. It is caused by HIV-1 subtype C (HIV-1C). In this study we present the identification and analysis of cumulative cytotoxic T-lymphocyte (CTL) responses in the southern African country of Botswana. CTLs were shown to be an important component of the immune response to control HIV-1 infection. The definition of optimal and dominant epitopes across the HIV-1C genome that are targeted by CTL is critical for vaccine design. The characteristics of the predominant virus that causes the HIV-1 epidemic in a certain geographic area and also the genetic background of the population, through the distribution of common HLA class I alleles, might impact dominant CTL responses in the vaccinee and in the general population. The enzyme-linked immunospot (Elispot) gamma interferon assay has recently been shown to be a reliable tool to map optimal CTL epitopes, correlating well with other methods, such as intracellular staining, tetramer staining, and the classical chromium release assay. Using Elispot with overlapping synthetic peptides across Gag, Tat, Rev, and Nef, we analyzed HIV-1C-specific CTL responses of HIV-1-infected blood donors. Profiles of cumulative Elispot-based CTL responses combined with diversity and sequence consensus data provide an additional characterization of immunodominant regions across the HIV-1C genome. Results of the study suggest that the construction of a poly-epitope subtype-specific HIV-1 vaccine that includes multiple copies of immunodominant CTL epitopes across the viral genome, derived from predominant HIV-1 viruses, might be a logical approach to the design of a vaccine against AIDS.     

Genetic analysis of Indian HIV-1 nef: subtyping, variability and implications

HIV-1 subtype C is predominant in India and globally. In the present study, we analyze HIV-1 subtype C regulatory protein Nef sequences from five recent Indian seroconverters and five long-term survivors (LTSs) for variability at crucial functional domains. Sequence analysis suggested the possibility of using regulatory gene sequences for viral subtyping and evolutionary studies apart from structural genes. In the phylogenetic tree, Indian nef sequences segregated away from other reported subtype C sequences, forming an Indian subclade within subtype C. Our studies also suggested no evidence for the association of truncated Nef with slow progression of disease, as all LTSs had intact Nef. We could identify some variations in juxtapositions to crucial functional domains, especially in seroconverter sequences, when comparing them with others. In phylogenetic analysis, specifically for the base-pair regions 411-428 and 478-525, our seroconverter sequences segregated away from those reported earlier in the literature, indicating specific evolutionary changes in these conserved regions of nef in currently circulating viruses. But the dN/dS ratio for our samples was less than one on comparing them with reported subtype C and representative sequences of different clades, strongly emphasizing the necessity of sequence conservation at different disease stages and even across clades. HLA-I binding epitope predictions for common Indian HLAs indicated that specific mutations in seroconverter Nef may alter the intensity of epitope binding, which may alter the outcome of the immune response. Hence these data would be useful in designing Nef epitopes to be included in multi-epitope HIV-1 vaccine for the Indian population and would also be of immense help in HIV-1 evolutionary studies.     

The development of HIV-1 subtype C vaccines for Southern Africa

One in nine people in South Africa are estimated to be HIV-1 infected, with the majority of these infections being due to HIV-1 subtype C. Until recently, most HIV-1 candidate vaccines were not based on subtype C genes. In response to this epidemic, therefore, the South African AIDS Vaccine Initiative (SAAVI) was established to facilitate the development and testing of candidate HIV-1 subtype C vaccines. The first HIV-1 subtype C candidate vaccine is due to be, tested at the end of 2002, and is based on Venezuelan encephalitis virus replicons expressing Gag protein. The next candidate vaccines to be tested will be DNA and modified vaccinia Ankara vaccines expressing subtype C genes.     

Identification of CD4-Binding Site Dependent Plasma Neutralizing Antibodies in an HIV-1 Infected Indian Individual

Dissecting antibody specificities in the plasma of HIV-1 infected individuals that develop broadly neutralizing antibodies (bNAbs) is likely to provide useful information for refining target epitopes for vaccine design. Several studies have reported CD4-binding site (CD4bs) antibodies as neutralization determinants in the plasma of subtype B-infected individuals; however there is little information on the prevalence of CD4bs specificities in HIV-infected individuals in India. Here, we report on the presence of CD4bs antibodies and their contribution to virus neutralization in the plasma from a cohort of HIV-1 infected Indian individuals. Plasma from 11 of the 140 HIV-1 infected individuals (7.9%) studied here exhibited cross-neutralization activity against a panel of subtype B and C viruses. Analyses of these 11 plasma samples for the presence of CD4bs antibodies using two CD4bs-selective probes (antigenically resurfaced HXB2gp120 core protein RSC3 and hyperglycosylated JRFLgp120 mutant ΔN2mCHO) revealed that five (AIIMS 617, 619, 627, 642, 660) contained RSC3-reactive plasma antibodies and only one (AIIMS 660) contained ΔN2mCHO-reactive antibodies. Plasma antibody depletion and competition experiments confirmed that the neutralizing activity in the AIIMS 660 plasma was dependent on CD4bs antibodies. To the best of our knowledge, this is the first study to report specifically on the presence of CD4bs antibodies in the plasma of a cohort of HIV-1 infected Indian donors. The identification of CD4bs dependent neutralizing antibodies in an HIV-1 infected Indian donor is a salient finding of this study and is supportive of ongoing efforts to induce similar antibodies by immunization.     

AIDS as a zoonosis? Confusion over the origin of the virus and the origin of the epidemics

Based on findings demonstrating the simian ancestry of HIV, AIDS has been reported to be a zoonosis. However, this theory has never been proved and must seriously be questioned. Several arguments show that HIV-AIDS is not a zoonosis. (i) If AIDS were a zoonosis, there must be evidence of AIDS being directly acquired from an animal species, as is rabies, a disease that is directly acquired from animals. (ii) Despite long-term and frequent human exposure to SIV-infected monkeys in Africa, only 11 cross-species transmission events are known, and only four of these have resulted in significant human-to-human transmission, generating HIV-1 groups M and O and HIV-2 groups A and B. The closest relatives of SIVcpz (HIV-1 group N) and of SIVsm (HIV-2 groups C-H) are extremely rare, with only six HIV-1 group N-infected patients and only single individuals known to be infected by HIV-2 groups C-H. SIV, while capable of cross-species transmission, is thus poorly adapted for disease and epidemic spread. If AIDS were a zoonosis that is capable of significant human-to-human spread, there would be a plethora of founder subtypes and groups. (iii) Human exposure to SIV is thousands of years old, but AIDS emerged only in the 20th century. If AIDS were a zoonosis that spread into the human population, it would have spread to the West during slave trade. (iv) Experimental transmission of SIVs to different species of monkeys is often well controlled by the new host, showing that the virus and not the disease is transmitted. Therefore, we conclude that cross-species transmission of SIV does not in itself constitute the basis for a zoonosis. Transmission per se is not the major requirement for the generation of the AIDS epidemic. All HIVs do derive from simian species, but AIDS does not qualify as a zoonosis and this explanation cannot in itself account for the origin of AIDS epidemic. It is important to distinguish AIDS from true zoonoses (e.g. rabies) because research is needed to understand the processes by which animal viruses cause sustained human-to-human transmission, epidemics and even pandemics. Much is known about emerging viruses, but almost nothing is known about emerging viral diseases.     

A Comprehensive Mapping of HIV-1 Genotypes in Various Risk Groups and Regions across China Based on a Nationwide Molecular Epidemiologic Survey

Background

China is experiencing a dynamic HIV/AIDS epidemic. While serology based surveillance systems have reported the spread of HIV/AIDS, detailed tracking of its transmission in populations and regions is not possible without mapping it at the molecular level. We therefore conducted a nationwide molecular epidemiology survey across the country.

Methods

HIV-1 genotypes were determined from 1,408 HIV-positive persons newly diagnosed in 2006. The prevalence of each genotype was estimated by weighting the genotype’s prevalence from each province- and risk-specific subpopulation with the number of reported cases in the corresponding subgroups in that year.

Results

CRF07_BC (35.5%), CRF01_AE (27.6%), CRF08_BC (20.1%), and subtype B'' (9.6%) were the four main HIV-1 strains in China. CRF07_BC and CRF08_BC were the primary drivers of infection among injecting drug users in northeastern and southeastern China, respectively, and subtype B'' remained dominant among former plasma donors in central China. In contrast, all four strains occurred in significant proportions among heterosexuals nationwide, pointing to an expansion of the HIV-1 epidemic from high-risk populations into the general population. CRF01_AE also replaced subtype B as the principal driver of infection among men-who-have-sex-with-men.

Conclusions

Our study provides the first comprehensive baseline data on the diversity and characteristics of HIV/AIDS epidemic in China, reflecting unique region- and risk group-specific transmission dynamics. The results provide information critical for designing effective prevention measures against HIV transmission.     

Infectious Simian/Human Immunodeficiency Virus with Human Immunodeficiency Virus Type 1 Subtype C from an African Isolate: Rhesus Macaque Model

Human immunodeficiency virus type 1 (HIV-1) subtype C is responsible for more than 56% of all infections in the HIV and AIDS pandemic. It is the predominant subtype in the rapidly expanding epidemic in southern Africa. To develop a relevant model that would facilitate studies of transmission, pathogenesis, and vaccine development for this subtype, we generated SHIV(MJ4), a simian/human immunodeficiency virus (SHIV) chimera based on HIV-1 subtype C. SHIV(MJ4) contains the majority of env, the entire second exon of tat, and a partial sequence of the second exon of rev, all derived from a CCR5-tropic, primary isolate envelope clone from southern Africa. SHIV(MJ4) replicated efficiently in human, rhesus, and pig-tailed macaque peripheral blood mononuclear cells (PBMCs) in vitro but not in CEMx174 cells. To assess in vivo infectivity, SHIV(MJ4) was intravenously inoculated into four rhesus macaques (Macaca mulatta). All four animals became infected as determined through virus isolation, PCR analysis, and viral loads of 10(7) to 10(8) copies of viral RNA per ml of plasma during the primary infection phase. We have established a CCR5-tropic SHIV(MJ4)/rhesus macaque model that may be useful in the studies of HIV-1 subtype C immunology and biology and may also facilitate the evaluation of vaccines to control the spread of HIV-1 subtype C in southern Africa and elsewhere.     

Dynamic interplay between viral adaptation and immune recognition during HIV-1 infection

Untreated human immunodeficiency virus (HIV) infections usually lead to death from AIDS, although the rate of the disease progression varies widely among individuals. The cytotoxic T lymphocyte (CTL) response, which is restricted by highly polymorphic MHC class I alleles, plays a central role in controlling HIV replication. It is now recognized that the antiviral efficacy of CTLs at the single cell level is dependent on their antigen specificity and is important in determining the quality of host response to viruses so that the individual will remain asymptomatic. However, because of the extreme mutational plasticity of HIV, HIV-specific CTL responses are continuously and dynamically changing. In order to rationally design an effective vaccine, the questions as to what constitutes an effective antiviral CTL response and what characterizes a potent antigenic peptide to induce such responses are becoming highlighted as needing to be answered.     

Inflammatory Genital Infections Mitigate a Severe Genetic Bottleneck in Heterosexual Transmission of Subtype A and C HIV-1

The HIV-1 epidemic in sub-Saharan Africa is driven largely by heterosexual transmission of non-subtype B viruses, of which subtypes C and A are predominant. Previous studies of subtype B and subtype C transmission pairs have suggested that a single variant from the chronically infected partner can establish infection in their newly infected partner. However, in subtype A infected individuals from a sex worker cohort and subtype B individuals from STD clinics, infection was frequently established by multiple variants. This study examined over 1750 single-genome amplified viral sequences derived from epidemiologically linked subtype C and subtype A transmission pairs very early after infection. In 90% (18/20) of the pairs, HIV-1 infection is initiated by a single viral variant that is derived from the quasispecies of the transmitting partner. In addition, the virus initiating infection in individuals who were infected by someone other than their spouse was characterized to determine if genital infections mitigated the severe genetic bottleneck observed in a majority of epidemiologically linked heterosexual HIV-1 transmission events. In nearly 50% (3/7) of individuals infected by someone other than their spouse, multiple genetic variants from a single individual established infection. A statistically significant association was observed between infection by multiple genetic variants and an inflammatory genital infection in the newly infected individual. Thus, in the vast majority of HIV-1 transmission events in cohabiting heterosexual couples, a single genetic variant establishes infection. Nevertheless, this severe genetic bottleneck can be mitigated by the presence of inflammatory genital infections in the at risk partner, suggesting that this restriction on genetic diversity is imposed in large part by the mucosal barrier.     

High-resolution molecular epidemiology and evolutionary history of HIV-1 subtypes in Albania

Background

HIV-1 epidemic in Western Europe is largely due to subtype B. Little is known about the HIV-1 in Eastern Europe, but a few studies have shown that non-B subtypes are quite common. In Albania, where a recent study estimated a ten-fold increase of AIDS incidence during the last six years, subtype A and B account for 90% of the know infections.

Methodology/Principal Findings

We investigated the demographic history of HIV-1 subtype A and B in Albania by using a statistical framework based on coalescent theory and phylogeography. High-resolution phylogenetic and molecular clock analysis showed a limited introduction to the Balkan country of subtype A during the late 1980s followed by an epidemic outburst in the early 1990s. In contrast, subtype B was apparently introduced multiple times between the mid-1970s and mid-1980s. Both subtypes are growing exponentially, although the HIV-1A epidemic displays a faster growth rate, and a significantly higher basic reproductive number R₀. HIV-1A gene flow occurs primarily from the capital Tirane, in the center of the country, to the periphery, while HIV-1B flow is characterized by a balanced exchange between center and periphery. Finally, we calculated that the actual number of infections in Albania is at least two orders of magnitude higher than previously thought.

Conclusions/Significance

Our analysis demonstrates the power of recently developed computational tools to investigate molecular epidemiology of pathogens, and emphasize the complex factors involved in the establishment of HIV-1 epidemics. We suggest that a significant correlation exists between HIV-1 exponential spread and the socio-political changes occurred during the Balkan wars. The fast growth of a relatively new non-B epidemic in the Balkans may have significant consequences for the evolution of HIV-1 epidemiology in neighboring countries in Eastern and Western Europe.     

The replicative fitness of primary human immunodeficiency virus type 1 (HIV-1) group M, HIV-1 group O, and HIV-2 isolates

The main (M) group of human immunodeficiency virus type 1 (HIV-1) is responsible for the global AIDS epidemic while HIV-1 group O (outlier) and HIV type 2 are endemic only in west and central Africa. The failure of HIV-2 and especially HIV-1 group O to spread following the initial zoonotic jumps is not well understood. This study was designed to examine the relative replicative capacities between these human lentiviruses. A pairwise competition experiment was performed with peripheral blood mononuclear cells with eight HIV-2 isolates, 6 group O viruses, and 15 group M viruses of subtype A (2 viruses), B (5 viruses), C (4 viruses), D (2 viruses) and CRF01_AE (2 viruses). HIV-1 group M isolates of any subtype were typically 100-fold-more fit than group O or HIV-2 strains when competed in peripheral blood mononuclear cells from various humans. This order in replicative fitness was also observed when virus pairs were added to human dendritic cells and then cocultured with primary, quiescent T cells, which is the model for HIV-1 transmission. These results suggest that reduced replicative and transmission fitness may be contributing to the low prevalence and limited geographical spread of HIV-2 and group O HIV-1 in the human population.