HIV-1 Sequences in the Establishment of Chronic Virus Producers: a Thesis

FoxP3 determines the development of CD4+CD25+ regulatory T (Treg) cells and represses interleukin-2 (IL-2) expression in Treg cells. However, human immunodeficiency virus type 1 (HIV-1) infects and replicates efficiently in FoxP3+ Treg cells. We report that, while inhibiting IL-2 gene expression, FoxP3 enhances gene expression from HIV-1 long terminal repeat (LTR). This FoxP3 activity requires both the N- and C-terminal domains and is inactivated by human IPEX (immunodysregulation, polyendocrinopathy, enteropathy, X-linked syndrome) mutations. FoxP3 enhances HIV-1 LTR via its specific NFkappaB binding sequences in an NFkappaB-dependent fashion in T cells but not in HEK293 cells. FoxP3 decreases level of histone acetylation at the interleukin-2 locus but not at the HIV-1 LTR. Although NFkappaB nuclear translocation is not altered, FoxP3 enhances NFkappaB-p65 binding to HIV-1 LTR. These data suggest that FoxP3 modulates gene expression in a promoter sequence-dependent fashion by modulating chromatin structure and NFkappaB activity. HIV-1 LTR has evolved to both highjack the T-cell activation pathway for expression and to resist FoxP3-mediated suppression of T-cell activation.

Depletion of CD4(+) T cells is the hallmark of HIV infection and AIDS progression. In addition to the direct killing of the viral-infected cells, HIV infection also leads to increased apoptosis of predominantly uninfected bystander cells. This is mediated in part through the HIV-1 Tat protein, which is secreted by the infected cells and taken up by uninfected cells. Using an affinity-purification approach, a specific and direct interaction of Tat with tubulin and polymerized microtubules has been detected. This interaction does not affect the secretion and uptake of Tat, but is critical for Tat to induce apoptosis. Tat binds tubulin/microtubules through a four-amino-acid subdomain of its conserved core region, leading to the alteration of microtubule dynamics and activation of a mitochondria-dependent apoptotic pathway. Bim, a pro-apoptotic Bcl-2 relative and a transducer of death signals initiated by perturbation of microtubule dynamics, facilitates the Tat-induced apoptosis. Our findings reveal a strategy by which Tat induces apoptosis by targeting the microtubule network. Thus HIV-1 Tat joins a growing list of pathogen-derived proteins that target the cytoskeleton of host cells.

Transmitted HIV Resistance Among Pregnant Young Women Infected with HIV-1 in Brazil

Human cytomegalovirus (HCMV) is commonly found in the brains of patients with AIDS and in some cases can be detected in the same cells as can human immunodeficiency virus type 1 (HIV-1). In this study, we analyzed the patterns of replication of HIV-1 and HCMV in singly infected cells and the effects of dual infection in human brain-derived cell lines of three different origins: neuroblastoma cell lines SK-N-MC and SY5Y; astrocytoma/glioblastoma cell lines U373-MG and Hs 683; and undifferentiated glioblastoma cell lines A172 and T98G. To bypass the restriction at the adsorption/penetration step in these CD4-negative cells, we used HIV-1 (amphotropic retrovirus) pseudotypes. These HIV-1 pseudotypes infected the majority of the cells in the cultures and expressed high levels of HIV-1 gene products in all except the SY5Y cells. The cell lines differed in the ability to support HCMV infection, but coinfection with HIV-1 had no effect on HCMV replication. The A172 cells were completely nonpermissive for HCMV gene expression, while HCMV replication in the singly infected T98G and SK-N-MC cell lines was restricted at the level of some early gene products. This resulted in complete and partial inhibition, respectively, of viral DNA synthesis. Dual infection of the A172, T98G, and SK-N-MC cells had no effect on HIV-1 replication. The other three cell lines, U373-MG, Hs 683, and SY5Y, were fully permissive for HCMV replication. In the U373-MG and Hs 683 cells, HCMV markedly inhibited the synthesis of HIV-1 gene products. In contrast, a transient stimulation of HIV-1 production followed by a repression was observed in the dually infected SY5Y cells. We conclude from these results that under conditions in which both HIV-1 and HCMV can undergo fully permissive infection, HCMV can repress HIV-1 gene expression. In cells in which HCMV replication is limited but HIV-1 replicates well, there is no effect on HIV-1 gene expression. However, activation of HIV-1, at least transiently, may occur in cells in which HIV-1 gene expression is limited. These studies suggest that a threshold level of some HIV-1 gene product(s) may obscure activation or promote repression of HIV replication by HCMV.

(See the major article by Wertheim et al on pages 304–13) Phylogenetics, the study of relatedness among homologous genetic sequences, is an integral part of virology research. Phylogenetic methods are used to reconstruct ancestral relationships among a set of viral sequences and to provide a framework within which hypotheses about virus evolution can be tested. By accounting for the nonindependence or shared ancestry of sampled sequences, the phylogenetic context makes it possible to distinguish between founder effects and natural selection as explanations for the spread of virus variants [1]. The recognition that circulating virus lineages coalesced at some time in the past makes it possible to use the dates and locations of viral sequences to reconstruct the spatiotemporal dynamics of an epidemic [2]. Viruses such as human immunodeficiency virus type 1 (HIV-1) are measurably evolving and, thus, generate epidemics in which there is a correspondence between the transmission network and phylogenetic branching [3]. Viruses that are phylogenetically clustered are likely to be connected by shorter transmission chains than viruses that are phylogenetically more distant from one another. Factors such as geography, the method of virus transmission, and the risk factors of infected persons also influence the parameters of a phylogenetic tree [3]. When combined with epidemiological data, phylogenetic cluster analyses can inform public health interventions by determining which epidemiological factors are associated with increased virus transmission [4, 5]. The parameters responsible for virus evolution and spread and the extent to which virus phenotypic characteristics correlate with shared ancestry can be estimated from large sequence datasets by subsampling up to several hundred sequences at a time and integrating or averaging the results obtained from the resulting trees [6]. Repeated subsampling makes large phylogenetic analyses tractable and yields parameter estimates that are independent of any single phylogeny. The HIV-1 reverse transcriptase (RT) gene has been sequenced more often than any other virus gene because RT sequences are often used in clinical settings to help guide the use of antiretroviral therapy. In this issue of The Journal of Infectious Diseases, Wertheim and colleagues leveraged the vast amount of publicly available HIV-1 RT sequence data to analyze the relatedness of previously published HIV-1 RT sequences from >80 000 individuals worldwide [7]. To analyze this large number of sequences, the authors dispensed with a phylogenetic approach and instead clustered sequences solely by their genetic distances without considering their ancestral history. Viruses were represented as nodes and were connected to one another if the genetic divergence of their sequences was no greater than 1%. Wertheim and colleagues found that 13 300 sequences, approximately one-sixth of the sequences in the dataset, were connected to at least 1 other sequence and that the mean number of connections per sequence was 3.8. The 1% threshold for connecting sequences from different individuals was sensitive enough to identify many of the inferred transmission clusters reported in the previously published studies from which the sequences were obtained. Most of these clusters were known from smaller regional cohorts of HIV-1–infected individuals. The authors' global analysis made it uniquely possible for them to identify >200 connections among sequences in different countries. As in previous studies of transmission clusters, Wertheim et al found that the degree of connectivity of viruses from different individuals was heterogeneous with some nodes connected to many more nodes than other nodes. Such heterogeneous connectivity patterns characterize most biological and social networks [8]. Consistent with the decreased replication fitness of viruses with drug resistance mutations, sequences containing drug resistance mutations were less likely to be part of a cluster than sequences without drug resistance mutations. The article by Wertheim and colleagues warrants commentary to address the use of a network (rather than a tree) to study HIV-1 transmission dynamics and to review the epidemiological conclusions that can be drawn by analyzing populations of HIV-1 sequences from different individuals either in isolation or in combination with temporal, geographic, epidemiological, and clinical data. A network is derived from a matrix of pairwise distances between each pair of nodes or sequences in a dataset. For large numbers of sequences, a network can be created much more rapidly than a tree because no attempt is made to take into account the patterns of shared ancestry of the individual sequences. One limitation of creating a network directly from a set of sequences, however, is that many more connections may be inferred than could have possibly existed in the real transmission network, a problem that may not occur if a phylogenetic analysis had been performed first. For example, if multiple infections happen in a short time span, several people may be infected with very similar viruses. The viral sequences from these people would all be connected by the method of Wertheim and colleagues, leading to many more edges in the thus constructed network than exist in reality. Indeed, with the exception of a study published by several authors of the study reviewed here [9], previous HIV-1 network analyses used a preliminary phylogenetic analysis either to define the connections between sequences or to characterize the internal structure of a set of connected sequences [4, 5, 10]. Both phylogenetic and network analysis can be used to estimate the cluster size distribution in a population of virus sequences. As in the study by Wertheim et al, the distribution of cluster sizes in populations of HIV-1 sequences is usually found to be heterogeneous, consistent with different rates of transmission for different subpopulations or individuals [4, 5, 10]. This heterogeneity suggests that interventions to prevent transmission should be targeted toward those with the highest risk of transmitting the virus to others. Not surprisingly, studies in which sequence data are combined with behavioral risk factors have shown that sequences from individuals with the same risk factors are likely to cluster with one another [10, 11]. Both phylogenetic and network analyses of HIV-1 sequences combined with information about the duration of infection have shown that sequences from recently infected individuals are more likely to cluster with sequences from other recently infected individuals [4, 12]. This clustering is consistent with the high HIV-1 transmissibility known to occur during acute infection [13]. In agreement with these studies, the authors' previously cited study [9] combined sequences and clinical data from 3700 individuals in 5 US clinics and found that sequences that were part of a cluster (defined as having a genetic distance no more than 1.5% from 1 or more other sequences) were more likely to be from younger antiretroviral-naive individuals with high plasma HIV-1 RNA levels than were sequences that were not part of a cluster. These and other studies support interventions directed toward reducing the risks of secondary transmission events by individuals with acute infection [14]. Although the complete knowledge of a specific transmission network is informative, such perfect knowledge is not required to understand the general processes responsible for the spread of a virus [15]. Neither phylogenetic nor nonphylogenetic network analyses can prove that HIV-1 transmission occurred directly between 2 individuals [16]. Although 2 individuals may carry HIV-1 strains with very similar sequences, these sequences will not necessarily be unique; very similar sequences could be found in viruses from other persons within the same transmission network. The notion that sequences alone can identify specific transmission events between individuals is a misconception that has the potential to jeopardize the continued public benefit that results from the open publication of pathogen genomes [17]. Hypotheses about direct transmission, therefore, make sense only when sequence analysis is combined with contact tracing. Few studies, however, contain both contact data and HIV-1 sequence data, and those that do have been too small to provide insight into the population-level factors responsible for HIV-1 spread. By identifying the large number of connections between individuals in different countries, Wertheim and colleagues demonstrate the novel insights that can be gained by analyzing publicly available sequence data from a variety of published studies. A more complete understanding of the relative importance of different global transmission routes will require further studies and new methods that can incorporate information on the intensity of sampling in different regions, different risk groups, and different years. Nonetheless, the data presented here demonstrate that even a virus such as HIV-1, which requires intimate contact for transmission, can form small-world networks linking virus variants from geographically remote regions [18].

Sterile alpha motif and HD domain-containing protein 1 (SAMHD1) restricts human immunodeficiency virus type 1 (HIV-1) replication in nondividing cells by degrading intracellular deoxynucleoside triphosphates (dNTPs). SAMHD1 is highly expressed in resting CD4+ T cells, which are important for the HIV-1 reservoir and viral latency; however, whether SAMHD1 affects HIV-1 latency is unknown. Recombinant SAMHD1 binds HIV-1 DNA or RNA fragments in vitro, but the function of this binding remains unclear. Here we investigate the effect of SAMHD1 on HIV-1 gene expression and reactivation of viral latency. We found that endogenous SAMHD1 impaired HIV-1 long terminal repeat (LTR) activity in monocytic THP-1 cells and HIV-1 reactivation in latently infected primary CD4+ T cells. Overexpression of wild-type (WT) SAMHD1 suppressed HIV-1 LTR-driven gene expression at a transcriptional level. Tat coexpression abrogated SAMHD1-mediated suppression of HIV-1 LTR-driven luciferase expression. SAMHD1 overexpression also suppressed the LTR activity of human T-cell leukemia virus type 1 (HTLV-1), but not that of murine leukemia virus (MLV), suggesting specific suppression of retroviral LTR-driven gene expression. WT SAMHD1 bound to proviral DNA and impaired reactivation of HIV-1 gene expression in latently infected J-Lat cells. In contrast, a nonphosphorylated mutant (T592A) and a dNTP triphosphohydrolase (dNTPase) inactive mutant (H206D R207N [HD/RN]) of SAMHD1 failed to efficiently suppress HIV-1 LTR-driven gene expression and reactivation of latent virus. Purified recombinant WT SAMHD1, but not the T592A and HD/RN mutants, bound to fragments of the HIV-1 LTR in vitro These findings suggest that SAMHD1-mediated suppression of HIV-1 LTR-driven gene expression potentially regulates viral latency in CD4+ T cells.IMPORTANCE A critical barrier to developing a cure for HIV-1 infection is the long-lived viral reservoir that exists in resting CD4+ T cells, the main targets of HIV-1. The viral reservoir is maintained through a variety of mechanisms, including regulation of the HIV-1 LTR promoter. The host protein SAMHD1 restricts HIV-1 replication in nondividing cells, but its role in HIV-1 latency remains unknown. Here we report a new function of SAMHD1 in regulating HIV-1 latency. We found that SAMHD1 suppressed HIV-1 LTR promoter-driven gene expression and reactivation of viral latency in cell lines and primary CD4+ T cells. Furthermore, SAMHD1 bound to the HIV-1 LTR in vitro and in a latently infected CD4+ T-cell line, suggesting that the binding may negatively modulate reactivation of HIV-1 latency. Our findings indicate a novel role for SAMHD1 in regulating HIV-1 latency, which enhances our understanding of the mechanisms regulating proviral gene expression in CD4+ T cells.

Efficient expression of the human immunodeficiency virus type 1 (HIV-1) structural gene products Gag, Pol, and Env involves the regulation by viral Rev and Rev-responsive elements (RRE). Removal of multiple inhibitory sequences (INS) in the coding regions of these structural genes or modification of the codon usage patterns of HIV-1 genes to those used by highly expressed human genes has been found to significantly increase HIV-1 structural protein expression in the absence of Rev and RRE. In this study, we show that efficient and stable expression of the HIV-1 structural gene products Gag and Env could be achieved by transfection with a noncytopathic Sindbis virus expression vector by using HIV-1 sequences from primary isolates without any sequence modification. Stable expression of these Gag and Env proteins was observed for more than 12 months. The fact that the Sindbis virus expression vector replicates its RNA only in the cytoplasm of the transfected cells and the fact that the lack of expression of HIV-1 Gag by the DNA vector containing unmodified HIV-1 gag sequences was associated with a lack of detectable cytoplasmic gag RNA suggest that a major blockage in the expression of HIV-1 structural proteins in the absence of Rev/RRE is caused by inefficient accumulation of mRNA in the cytoplasm. Efficient long-term expression of structural proteins of diverse HIV-1 strains by the noncytopathic Sindbis virus expression system may be a useful tool for functional study of HIV-1 gene products and vaccine research.

A critical aspect of AIDS pathogenesis that remains unclear is the mechanism by which human immunodeficiency virus type 1 (HIV-1) induces death in CD4(+) T lymphocytes. A better understanding of the death process occurring in infected cells may provide valuable insight into the viral component responsible for cytopathicity. This would aid the design of preventive treatments against the rapid decline of CD4(+) T cells that results in AIDS. Previously, apoptotic cell death has been reported in HIV-1 infections in cultured T cells, and it has been suggested that this could affect both infected and uninfected cells. To evaluate the mechanism of this effect, we have studied HIV-1-induced cell death extensively by infecting several T-cell lines and assessing the level of apoptosis by using various biochemical and flow cytometric assays. Contrary to the prevailing view that apoptosis plays a prominent role in HIV-1-mediated T-cell death, we found that Jurkat and H9 cells dying from HIV-1 infection fail to exhibit the collective hallmarks of apoptosis. Among the parameters investigated, Annexin V display, caspase activity and cleavage of caspase substrates, TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) signal, and APO2.7 display were detected at low to negligible levels. Neither peptide caspase inhibitors nor the antiapoptotic proteins Bcl-x(L) or v-FLIP could prevent cell death in HIV-1-infected cultures. Furthermore, Jurkat cell lines deficient in RIP, caspase-8, or FADD were as susceptible as wild-type Jurkat cells to HIV-1 cytopathicity. These results suggest that the primary mode of cytopathicity by laboratory-adapted molecular clones of HIV-1 in cultured cell lines is not via apoptosis. Rather, cell death occurs most likely via a necrotic or lytic form of death independent of caspase activation in directly infected cells.

Background: N 6-methyladenosine (m 6 A) modifications of human immunodeficiency virus type 1 (HIV-1) and cellular RNA contribute to viral immune evasion and regulation of host and viral gene expression. We reported elevated RNA m 6 A levels in peripheral blood mononuclear cells (PBMCs) from HIV-1 viremic individuals compared to those on antiretroviral therapy (ART). RNA m 6 A dysregulation has been implicated in many types of cancer. However, the role of m 6 A modifications in HIV-1-associated cancers remains to be investigated. In this study, we aim to address this important question using clinical samples. Methods: We quantified RNA m 6 A levels in PBMCs from 43 de-identified people living with HIV-1 (PLWH), comparing those with cancer (n=15) to those without cancer (n=28). We used enzyme-linked immunosorbent assay (ELISA) to measure RNA m 6 A levels in PBMCs. Using an array of reverse transcription quantitative polymerase chain reaction (RT-qPCR), we performed quantitative transcriptomic analysis of 84 IFN-I-responsive genes in PBMCs. Furthermore, we performed linear regression analyses of cellular RNA m 6 A levels with HIV-1 RNA copies and CD4 + T cell counts in peripheral blood. Results: We found that m 6 A levels of PBMCs were 2.8-fold higher in the cancer group and correlated with expression of m 6 A regulatory genes. Higher m 6 A levels were also associated with increased HIV-1 RNA copies and reduced CD4 + T cell counts. HIV-1 viral load in the cancer group was higher than the non-cancer group. Transcriptomic analysis of 84 IFN-I-responsive genes revealed upregulation of many pro-inflammatory and interferon-stimulated genes in PLWH with cancer. Conclusions: Our findings suggest that HIV-1 infection and cancer microenvironment-mediated m 6 A reprogramming may contribute to chronic immune activation and malignancy in PLWH. Our results also highlight a post-transcriptional mechanism linking HIV-1 persistence to cancer risk.

CD8(+) T lymphocytes can inhibit human immunodeficiency virus type 1 (HIV-1) replication by secreting a soluble factor(s) known as CD8(+) T-lymphocyte antiviral factor (CAF). One site of CAF action is inhibition of HIV-1 RNA transcription, particularly at the step of long terminal repeat (LTR)-driven gene expression. The inhibitory effect of CAF on HIV-1 LTR activation is mediated through STAT1 activation. A recent study reports that alpha-defensins 1 to 3 account for CAF activity against HIV-1. Here, we address whether alpha-defensins, particularly alpha-defensin-1, contribute to CAF-mediated inhibition of HIV-1 transcription. Both recombinant alpha-defensin-1 and CAF derived from herpesvirus saimiri (HVS)-transformed CD8(+) cells inhibited HIV-1 infection and gene expression. For both factors, the inhibition of HIV-1 infection did not occur at the level of viral entry. Pretreatment of cells with alpha-defensin-1 followed by a washing out prior to infection blocked infection by HIV-1, indicating that direct inactivation of virions was not required for its inhibitory effect. In contrast to CAF, alpha-defensin-1 did not inhibit phorbol myristate acetate- or Tat-mediated HIV-1 LTR activation in a transient transfection system, nor did it activate STAT1 tyrosine phosphorylation. Furthermore, alpha-defensins 1 to 3 were below the level of detection in a panel of HVS-transformed CD8(+) cells with potent HIV-1 inhibitory activity and a neutralizing antibody against alpha-defensins 1 to 3 did not reverse the inhibitory effect of CAF on HIV-1 gene expression in infected cells and on HIV-1 LTR activation in transfected cells. Taken together, our results suggest that alpha-defensin-1 inhibits HIV-1 infection following viral entry but that alpha-defensins 1 to 3 are not responsible for the HIV-1 transcriptional inhibition by CAF.

Recent studies have demonstrated that genomes of poliovirus with deletions in the P1 (capsid) region contain the necessary viral information for RNA replication. To test the effects of the substitution of foreign genes on RNA replication and protein expression, chimeric human immunodeficiency virus type 1 (HIV-1)-poliovirus genomes were constructed in which regions of the gag, pol, or env gene of HIV-1 were substituted for regions of the P1 gene in the infectious cDNA clone of type 1 Mahoney poliovirus. The HIV-1 genes were inserted between nucleotides 1174 and 2956 of the poliovirus cDNA so that the translational reading frame was maintained between the HIV-1 genes and the remaining poliovirus genes. The chimeric genomes were positioned downstream from a T7 RNA polymerase promoter and transcribed in vitro by using T7 RNA polymerase, and the RNA was transfected into HeLa cells. A Northern (RNA blot) analysis of the RNA from transfected cells demonstrated the appropriate-size RNA, corresponding to the full-length chimeric genomes, which increased over time. Immunoprecipitation with antibodies specific for poliovirus RNA polymerase or sera from AIDS patients demonstrated the expression of the poliovirus RNA polymerase and HIV-1 proteins as fusions with the poliovirus P1 protein. The expression of the HIV-1-poliovirus P1 fusion protein was dependent upon an intact RNA polymerase gene, indicating that RNA replication was required for efficient expression. A pulse-chase analysis of the protein expression from the chimeric genomes demonstrated the initial rapid proteolytic processing of the polyprotein from the chimeric genomes to give HIV-1-poliovirus P1 fusion protein in transfected cells; the HIV-1 gag-P1 and HIV-1 pol-P1 fusion proteins exhibited a greater intracellular stability than the HIV-1 env-P1 fusion protein. Finally, superinfection with wild-type poliovirus of HeLa cells which had been transfected with the chimeric genomes did not significantly affect the expression of chimeric fusion protein. The results are discussed in the context of poliovirus RNA replication and demonstrate the feasibility of using poliovirus genomes (minireplicons) as novel vectors for expression of foreign proteins.

Apoptosis contributes to the loss of CD4 cells during human immunodeficiency virus type 1 (HIV-1) infection. Although the product of the env gene, gp160/gp120, is known to play a role in cell death mediated by HIV-1, the role of other HIV-1 genes in the process is unclear. We found that HIV-1 lacking the env gene (HIVDeltaenv) still induced apoptosis in T-cell lines and primary CD4 T cells. The ability to induce apoptosis was attributable to Tat, a viral regulatory protein. Tat induction of apoptosis was separate from the transactivation function of Tat, required expression of the second exon of Tat, and was associated with the increased expression and activity of caspase-8 (casp-8), a signaling molecule in apoptotic pathways. Moreover, induction of apoptosis could be prevented by treating cells with an inhibitor of casp-8. In addition, we show that HIV-1Deltaenv infection and Tat expression increased the sensitivity of cells to Fas-mediated apoptosis, an apoptotic pathway that signals via casp-8. The up-regulation of casp-8 by HIV-1 Tat expression may contribute to the increased apoptosis and sensitivity to apoptotic signals observed in the cells of HIV-1-infected persons.

An effective vaccine against human immunodeficiency virus type 1 (HIV-1) will have to provide protection against a vast array of different HIV-1 strains. Current methods to measure HIV-1-specific binding antibodies following immunization typically focus on determining the magnitude of antibody responses, but the epitope diversity of antibody responses has remained largely unexplored. Here we describe the development of a global HIV-1 peptide microarray that contains 6564 peptides from across the HIV-1 proteome and covers the majority of HIV-1 sequences in the Los Alamos National Laboratory global HIV-1 sequence database. Using this microarray, we quantified the magnitude, breadth, and depth of IgG binding to linear HIV-1 sequences in HIV-1-infected humans and HIV-1-vaccinated humans, rhesus monkeys and guinea pigs. The microarray measured potentially important differences in antibody epitope diversity, particularly regarding the depth of epitope variants recognized at each binding site. Our data suggest that the global HIV-1 peptide microarray may be a useful tool for both preclinical and clinical HIV-1 research.

This article describes a case of human immunodeficiency virus type 1 (HIV-1) infection transmission caused by a bloody knife fight in a robbery. The victim was a 69-year-old man who was not infected with HIV-1, and his wife was HIV-antibody negative. A robber, a 42-year-old man, was HIV antibody-positive since December 2005 and had not taken antiretroviral therapy. The BED IgG Capture incidence EIA (BED-CEIA assay) data showed that the specimens from the victim were compatible with a recent seroconversion. Phylogenetic analysis of fragments of pol, encompassing protease and a portion of reverse transcriptase, and of env genes isolated from the victim, the robber, and a local population samples of HIV-1 positive individuals showed that the victim's HIV-1 sequences were most closely related to and nested within a lineage comprised of the robber's HIV-1 sequences. We provide HIV-1 seroconversion data and phylogenetic analysis as evidence that the HIV-1 transmission likely occurred from contact during the robbery.

The role of thiamine in HIV infection