[Show abstract][Hide abstract] ABSTRACT: Mutations associated with the use of protease (PR) and reverse transcriptase (RT) inhibitors have been mostly mapped for HIV-1 subtype B. The prevalence of these mutations in drug-naive HIV-1 subtype B-infected individuals is low but occurs at high frequencies in treated individuals. To determine the prevalence of treatment-associated mutations in non-B viruses, we analyzed a 1613-bp pol region of specimens collected from 57 HIV-1-infected treatment-naive individuals from Cameroon. Of the 57 HIV-1 sequences, 43 belonged to CRF02-AG, two to CRF11-cpx, six to subtype A, one to subtype D, and five were unclassifiable. Of the 57 PR sequences, 100% contained at least one codon change giving substitutions at positions 10, 11, 16, 20, 33, 36, 60, 62, 64, 69, 77, and 89. These substitutions gave the following prevalence pattern, 36I/L (100%, 57/57) >89M/I (98%, 56/57)>69K/R (93%, 53/57)>20I/R (89%, 51/57)>16E (16%, 9/57)>64M (12%, 7/57)>10I (11%, 6/57)>11V (5%, 3/57)=62V (5%, 3/57)=77I (5%, 3/57)>233F/V (4%, 2/57)=60E (4%), which differed significantly from subtype B at positions 20, 36, 69, and 89. All but one (98%) of the 57 RT sequences (438 amino acid residues) carried substitutions located at codons 39A (7%), 43E (7%), 122E (7%), 312Q (2%), 333E (2%), 335C/D (89%), 356K (89%), 358K (14%), 365I (2%), 371V (81%), 376S (11%), or 399D (4%); the frequency of these substitutions ranged from <0.5% to 4% in RT of subtype B. The high prevalence of minor mutations associated with protease inhibitors (PI) and reverse transcriptase inhibitors (RTI) represents natural polymorphisms. HIV-1 PR and RT sequences from antiretroviral (ARV)-naive HIV-infected persons in Cameroon are important for monitoring the development of resistance to PIs and RTIs as such mutations could lead to treatment failures in individuals undergoing ARV therapy.
AIDS research and human retroviruses 09/2011; 28(7):675-84. DOI:10.1089/AID.2011.0181 · 2.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: HIV continues to spread globally, mainly through sexual contact. Despite advances in treatment and care, preventing transmission with vaccines or microbicides has proven difficult. A promising strategy to avoid transmission is prophylactic treatment with antiretroviral drugs before exposure to HIV. Clinical trials evaluating the efficacy of daily treatment with the reverse transcriptase inhibitors tenofovir disoproxil fumarate (TDF) or Truvada (TDF plus emtricitabine) are under way. We hypothesized that intermittent prophylactic treatment with long-acting antiviral drugs would be as effective as daily dosing in blocking the earliest stages of viral replication and preventing mucosal transmission. We tested this hypothesis by intermittently giving prophylactic Truvada to macaque monkeys and then exposing them rectally to simian-human immunodeficiency virus (SHIV) once a week for 14 weeks. A simple regimen with an oral dose of Truvada given 1, 3, or 7 days before exposure followed by a second dose 2 hours after exposure was as protective as daily drug administration, possibly because of the long intracellular persistence of the drugs. In addition, a two-dose regimen initiated 2 hours before or after virus exposure was effective, and full protection was obtained by doubling the Truvada concentration in both doses. We saw no protection if the first dose was delayed until 24 hours after exposure, underscoring the importance of blocking initial replication in the mucosa. Our results show that intermittent prophylactic treatment with an antiviral drug can be highly effective in preventing SHIV infection, with a wide window of protection. They strengthen the possibility of developing feasible, cost-effective strategies to prevent HIV transmission in humans.
Science translational medicine 01/2010; 2(14):14ra4. DOI:10.1126/scitranslmed.3000391 · 15.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: New-generation gels that deliver potent antiretroviral drugs against human immunodeficiency virus type 1 have renewed hopes for topical prophylaxis as a prevention strategy. Previous preclinical research with monkey models suggested that high concentrations and drug combinations are needed for high efficacy. We evaluated two long-acting reverse transcriptase inhibitors, tenofovir (TFV) and emtricitabine (FTC), by using a twice-weekly repeat challenge macaque model and showed that a preexposure vaginal application of gel with 1% TFV alone or in combination with 5% FTC fully protected macaques from a total of 20 exposures to simian-human immunodeficiency virus SF162p3. FTC and TFV were detected in plasma 30 min after vaginal application, suggesting rapid absorption. FTC was detected more frequently than TFV and showed higher levels, reflecting the fivefold-higher concentration of this drug than of TFV. Two of 12 repeatedly exposed but protected macaques showed limited T-cell priming, which did not induce resistance to infection when macaques were rechallenged. Thus, single drugs with durable antiviral activity can provide highly effective topical prophylaxis and overcome the need for noncoital use or for drug combinations which are more complex and costly to formulate and approve.
Journal of Virology 09/2009; 83(20):10358-65. DOI:10.1128/JVI.01073-09 · 4.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cross-species transmission of retroviruses is common in Cameroon. To determine risk for simian T-cell lymphotropic virus (STLV) transmission from nonhuman primates to hunters, we examined 170 hunter-collected dried blood spots (DBS) from 12 species for STLV. PCR with generic tax and group-specific long terminal repeat primers showed that 12 (7%) specimens from 4 nonhuman primate species were infected with STLV. Phylogenetic analyses showed broad diversity of STLV, including novel STLV-1 and STLV-3 sequences and a highly divergent STLV-3 subtype found in Cercopithecus mona and C. nictitans monkeys. Screening of peripheral blood mononuclear cell DNA from 63 HTLV-seroreactive, PCR-negative hunters did not identify human infections with this divergent STLV-3. Therefore, hunter-collected DBS can effectively capture STLV diversity at the point where pathogen spillover occurs. Broad screening using this relatively easy collection strategy has potential for large-scale monitoring of retrovirus cross-species transmission among highly exposed human populations.
[Show abstract][Hide abstract] ABSTRACT: Background
Human T-lymphotropic virus type 4 (HTLV-4) is a new deltaretrovirus recently identified in a primate hunter in Cameroon. Limited sequence analysis previously showed that HTLV-4 may be distinct from HTLV-1, HTLV-2, and HTLV-3, and their simian counterparts, STLV-1, STLV-2, and STLV-3, respectively. Analysis of full-length genomes can provide basic information on the evolutionary history and replication and pathogenic potential of new viruses.
We report here the first complete HTLV-4 sequence obtained by PCR-based genome walking using uncultured peripheral blood lymphocyte DNA from an HTLV-4-infected person. The HTLV-4(1863LE) genome is 8791-bp long and is equidistant from HTLV-1, HTLV-2, and HTLV-3 sharing only 62–71% nucleotide identity. HTLV-4 has a prototypic genomic structure with all enzymatic, regulatory, and structural proteins preserved. Like STLV-2, STLV-3, and HTLV-3, HTLV-4 is missing a third 21-bp transcription element found in the long terminal repeats of HTLV-1 and HTLV-2 but instead contains unique c-Myb and pre B-cell leukemic transcription factor binding sites. Like HTLV-2, the PDZ motif important for cellular signal transduction and transformation in HTLV-1 and HTLV-3 is missing in the C-terminus of the HTLV-4 Tax protein. A basic leucine zipper (b-ZIP) region located in the antisense strand of HTLV-1 and believed to play a role in viral replication and oncogenesis, was also found in the complementary strand of HTLV-4. Detailed phylogenetic analysis shows that HTLV-4 is clearly a monophyletic viral group. Dating using a relaxed molecular clock inferred that the most recent common ancestor of HTLV-4 and HTLV-2/STLV-2 occurred 49,800 to 378,000 years ago making this the oldest known PTLV lineage. Interestingly, this period coincides with the emergence of Homo sapiens sapiens during the Middle Pleistocene suggesting that early humans may have been susceptible hosts for the ancestral HTLV-4.
The inferred ancient origin of HTLV-4 coinciding with the appearance of Homo sapiens, the propensity of STLVs to cross-species into humans, the fact that HTLV-1 and -2 spread globally following migrations of ancient populations, all suggest that HTLV-4 may be prevalent. Expanded surveillance and clinical studies are needed to better define the epidemiology and public health importance of HTLV-4 infection.
[Show abstract][Hide abstract] ABSTRACT: Frequent infection with zoonotic simian foamy virus (SFV) has been reported among HIV-negative primate hunters in rural Cameroon. Plasma samples obtained from urban commercial sex workers (CSWs; n = 139), patients with sexually transmitted diseases (n = 41), and blood donors (n = 179) in the Democratic Republic of Congo [formerly known as Zaire] and Cameroon were tested for SFV and HIV-1 infection. One CSW and one blood donor were found to be seropositive for both SFV and HIV-1, thereby documenting what are, to our knowledge, the first reported cases of dual SFV and HIV infection. The findings of the present study suggest opportunities for bloodborne and sexual transmission of SFV and highlight the importance of defining the clinical consequences of dual infections.
The Journal of Infectious Diseases 06/2008; 197(10):1389-93. DOI:10.1086/587493 · 6.00 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Preexposure prophylaxis (PrEP) with antiretroviral drugs constitutes a promising strategy for HIV prevention. Potent PrEP regimens with reverse transcriptase inhibitors can prevent detectable SHIV infection in a repeated low-dose macaque model that resembles human transmission, supporting plans to quickly move this approach into human trials. However, the possibility remains that extremely low levels of virus replication could nonetheless occur during PrEP and seed viral reservoirs in tissues. Therefore, seemingly protected macaques may harbor occult virus that may be initially contained by cytotoxic T cells, but could emerge later. To explore this possibility, we studied whether CD8(+) cells suppress viremia in four rhesus macaques apparently protected by daily or intermittent Truvada (FTC and tenofovir) during 14 low-dose, rectal SHIV(SF162P3) challenges and during a subsequent drug washout period. CD8(+) cells were efficiently ablated with antibodies in these and two additional control macaques that were previously infected but had reached undetectable virus set points. During 4 weeks of follow-up, all four macaques remained free of plasma viremia and provirus in blood lymphocytes. In contrast, plasma viremia resurged to 10(6) to 10(7) copies per milliliter within 2 weeks in both control macaques. Thus, these results indicate that the undetectable viremia in the PrEP-protected macaques was not due to CD8(+) cells that were containing a low-level infection. Rather, the PrEP treatment created conditions in which infection was prevented, eliminated, or controlled by unknown mechanisms. These data provide important information for PrEP usage to prevent HIV transmission, and fully support the continued pursuit of PrEP prevention measures in humans.
AIDS research and human retroviruses 05/2008; 24(4):543-6. DOI:10.1089/aid.2007.0222 · 2.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In the absence of an effective vaccine, HIV continues to spread globally, emphasizing the need for novel strategies to limit its transmission. Pre-exposure prophylaxis (PrEP) with antiretroviral drugs could prove to be an effective intervention strategy if highly efficacious and cost-effective PrEP modalities are identified. We evaluated daily and intermittent PrEP regimens of increasing antiviral activity in a macaque model that closely resembles human transmission.
We used a repeat-exposure macaque model with 14 weekly rectal virus challenges. Three drug treatments were given once daily, each to a different group of six rhesus macaques. Group 1 was treated subcutaneously with a human-equivalent dose of emtricitabine (FTC), group 2 received orally the human-equivalent dosing of both FTC and tenofovir-disoproxil fumarate (TDF), and group 3 received subcutaneously a similar dosing of FTC and a higher dose of tenofovir. A fourth group of six rhesus macaques (group 4) received intermittently a PrEP regimen similar to group 3 only 2 h before and 24 h after each weekly virus challenge. Results were compared to 18 control macaques that did not receive any drug treatment. The risk of infection in macaques treated in groups 1 and 2 was 3.8- and 7.8-fold lower than in untreated macaques (p = 0.02 and p = 0.008, respectively). All six macaques in group 3 were protected. Breakthrough infections had blunted acute viremias; drug resistance was seen in two of six animals. All six animals in group 4 that received intermittent PrEP were protected.
This model suggests that single drugs for daily PrEP can be protective but a combination of antiretroviral drugs may be required to increase the level of protection. Short but potent intermittent PrEP can provide protection comparable to that of daily PrEP in this SHIV/macaque model. These findings support PrEP trials for HIV prevention in humans and identify promising PrEP modalities.
PLoS Medicine 03/2008; 5(2):e28. DOI:10.1371/journal.pmed.0050028 · 14.43 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Persons occupationally exposed to nonhuman primates (NHPs) can be persistently infected with simian foamy virus (SFV). The clinical significance and person-to-person transmissibility of zoonotic SFV infection is unclear. Seven SFV-infected men responded to annual structured interviews and provided whole blood, oral, and urogenital specimens for study. Wives were tested for SFV infection. Proviral DNA was consistently detected by PCR in PBMCs of infected men and inconsistently in oral or urogenital samples. SFV was infrequently cultured from their PBMCs and throat swabs. Despite this and a long period of intimate exposure (median 20 years), wives were SFV negative. Most participants reported nonspecific symptoms and diseases common to aging. However, one of two persons with mild thrombocytopenia had clinically asymptomatic nonprogressive, monoclonal natural killer cell lymphocytosis of unclear relationship to SFV. All participants worked with NHPs before 1988 using mucocutaneous protection inconsistently; 57% described percutaneous injuries involving the infecting NHP species. SFV likely transmits to humans through both percutaneous and mucocutaneous exposures to NHP body fluids. Limited follow-up has not identified SFV-associated illness and secondary transmission among humans.
AIDS Research and Human Retroviruses 12/2007; 23(11):1330-7. DOI:10.1089/aid.2007.0104 · 2.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Within the last few years significant efforts have been made to identify objective reliable diagnostic markers from individuals with chronic fatigue syndrome (CFS). We report the absence of a previously described retroviral marker (HTLV-II gag) in a blinded study of CFS cases. Even with excellent reproducible sensitivities, this marker failed in repeated attempts to distinguish cases from controls. In addition, four other retroviruses (simian T cell leukaemia virus, human spumavirus, bovine leukaemia virus and simian retrovirus) were examined for their presence in these CFS cases and found to be absent. Our findings suggest that these agents, at least as markers, are non-distinguishing for CFS and that other factors may be confounding the resolution of an aetiology to this syndrome.
[Show abstract][Hide abstract] ABSTRACT: We reported previously the absence of systemic infection in a subset of macaques vaccinated with an HIV-1 DNA/MVA vaccine after 18 or more rectal challenges with low (physiologically relevant) doses of SHIV162P3. To further study the apparent protection, we looked for sequestered virus in gut tissues, lymph nodes, spleen, and testes obtained at necropsy using virus co-culture and nested PCR for SIV Gag, Pol and LTR. There was no evidence of sequestered virus in tissues obtained from the four protected macaques. In contrast, at least one tissue from each of 11 infected animals scored positive by one of these sensitive procedures. Activated PBMC from the protected macaques were not inherently resistant to in vitro infection by the challenge virus. These findings demonstrate that some vaccinated macaques appeared to be free from the challenge virus. Therefore, such T cell-based vaccines may provide some protection when challenge virus doses approach physiological equivalencies.
[Show abstract][Hide abstract] ABSTRACT: We examined the efficacy of tenofovir disoproxil fumarate (TDF) in blocking simian human immunodeficiency virus (SHIV) infection
in Chinese rhesus macaques. Once weekly for 14 weeks or until a macaque became infected, 12 male macaques were inoculated
intrarectally with amounts of SHIVSF162P3 (10 median tissue culture infective doses; 3.8×105 virus particles) that were ∼5-fold higher than the human immunodeficiency virus type 1 RNA levels noted in human semen during
an acute infection. Of the 12 macaques, 4 received oral TDF daily, 4 received oral TDF once weekly, and 4 (control animals)
received no TDF. The control animals became infected after receiving a median of 1.5 virus inoculations; macaques receiving
TDF daily (1 macaque remained uninfected after 14 inoculations) and those receiving TDF weekly became infected after a median
duration of 6.0 and 7.0 weeks, respectively. Although infection was delayed in treated macaques, compared with control macaques,
the differences were not statistically significant (P=.315); however, the study was limited by the small numbers of animals
evaluated and the variability in blood levels of TDF that resulted from oral dosing. These data demonstrate that treatment
with oral TDF provided partial protection against SHIV infection but ultimately did not protect all TDF treated animals against
multiple virus challenges
The Journal of Infectious Diseases 11/2006; 194(7):904-11. DOI:10.1086/507306 · 6.00 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Historically, HIV vaccines specifically designed to raise cellular immunity resulted in protection from disease progression but not infection when tested in monkeys challenged with a single high virus exposure. An alternative approach, more analogous to human sexual exposures, is to repetitively challenge immunized monkeys with a much lower dose of virus until systemic infection is documented. Using these conditions to mimic human sexual transmission, we found that a multi-protein DNA/MVA HIV-1 vaccine is indeed capable of protecting rhesus monkeys against systemic infection when repeatedly challenged with a highly heterologous immunodeficiency virus (SHIV). Furthermore, this repetitive challenge approach allowed us to calculate per-exposure probability of infection, an observed vaccine efficacy of 64%, and undertake a systematic analysis for correlates of protection based on exposures needed to achieve infection. Therefore, improved non-human primate models for vaccine efficacy can provide novel insight and perhaps renew expectations for positive outcomes of human HIV clinical trials.
[Show abstract][Hide abstract] ABSTRACT: Human T-lymphotropic virus type 3 (HTLV-3) is a new virus recently identified in two primate hunters in Central Africa. Limited sequence analysis shows that HTLV-3 is distinct from HTLV-1 and HTLV-2 but is genetically similar to simian T-lymphotropic virus type 3 (STLV-3). We report here the first complete HTLV-3 sequence obtained by PCR-based genome walking using uncultured peripheral blood lymphocytes from an HTLV-3-infected person. The HTLV-3(2026ND) genome is 8,917 bp long and is genetically equidistant from HTLV-1 and HTLV-2, sharing about 62% identity. Phylogenetic analysis of all gene regions confirms this relationship and shows that HTLV-3 falls within the diversity of STLV-3, suggesting a primate origin. However, HTLV-3(2026ND) is unique, sharing only 87% to 92% sequence identity with STLV-3. SimPlot and phylogenetic analysis did not reveal any evidence of genetic recombination with either HTLV-1, HTLV-2, or STLV-3. Molecular dating estimates that the ancestor of HTLV-3 is as old as HTLV-1 and HTLV-2, with an inferred divergence time of 36,087 to 54,067 years ago. HTLV-3 has a prototypic genomic structure, with all enzymatic, regulatory, and structural proteins preserved. Like STLV-3, HTLV-3 is missing a third 21-bp transcription element found in the long terminal repeats of HTLV-1 and HTLV-2 but instead contains a unique activator protein-1 transcription factor upstream of the 21-bp repeat elements. A PDZ motif, like that in HTLV-1, which is important for cellular signal transduction and transformation, is present in the C terminus of the HTLV-3 Tax protein. A basic leucine zipper region located in the antisense strand of HTLV-1, believed to play a role in viral replication and oncogenesis, was also found in the complementary strand of HTLV-3. The ancient origin of HTLV-3, the broad distribution of STLV-3 in Africa, and the propensity of STLVs to cross species into humans all suggest that HTLV-3 may be prevalent and support the need for expanded surveillance for this virus.
Journal of Virology 09/2006; 80(15):7427-38. DOI:10.1128/JVI.00690-06 · 4.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Non-human primate models for human immunodeficiency virus (HIV) infection represent a valuable pre-clinical tool to evaluate interventions (e.g., topical microbicides, vaccines, and chemoprophylaxis) designed to prevent transmission or slow disease progression after infection. Standard transmission models use a single-dose exposure with high, non-physiologic levels of virus to approach 100% infection rates of control animals. These single-exposure models do not represent the circumstances of mucosal HIV transmission in humans and may result in misleading data with regard to intervention efficacy. Therefore, we have developed a repetitive mucosal exposure model using doses of virus that better reflects human exposures.
The virus used for these evaluations was simian-human immunodeficiency virus [SHIVSF162P3 (R5-using, subtype B HIV-1 envelope)] and the virus dose used (approximately 10(5)-10(6) viral particle equivalents or approximately 10 tissue culture infectious doses per exposure) approximates viral loads observed in the semen during acute HIV-1 infection. Using the repeated mucosal exposure approach, we have evaluated a candidate vaginal microbicide (cellulose acetate phthalate, CAP) given 15 minutes prior to each weekly virus exposure. Pig-tailed macaques were exposed weekly by vaginal inoculations with and without microbicide until systemic viral RNA was detected.
Groups of naïve control monkeys were infected after an average of three to four exposures for the vaginal route of inoculation. Data from the first application of this monkey model to evaluate the topical microbicide CAP suggested that protection from SHIV infection was possible with three of four CAP-treated monkeys remaining uninfected after 12 exposures (P = 0.015). CAP efficacy was markedly improved from 66% in a previous single-dose virus exposure study to 92% in this repeated exposure system.
Our experience with using repetitive virus exposures to study topical microbicides and the findings to date from this study provides a basis to refine monkey models to more closely resemble human exposure during HIV transmission. This model may be highly relevant to pre-clinical evaluation for a variety of therapeutic interventions which is discussed here.
Journal of Medical Primatology 09/2006; 35(4-5):210-6. DOI:10.1111/j.1600-0684.2006.00169.x · 0.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: HIV-seronegative Cameroonians with exposure to nonhuman primates were tested for simian immunodeficiency virus (SIV) infection. Seroreactivity was correlated with exposure risk (p<0.001). One person had strong humoral and weak cellular immune reactivity to SIVcol peptides. Humans are exposed to and possibly infected with SIV, which has major public health implications.
[Show abstract][Hide abstract] ABSTRACT: Recent HIV infection or divergent HIV or simian immunodeficiency virus (SIV) strains may be responsible for Western blot-indeterminate results on 70 serum samples from Zairian hospital employees that were reactive in an enzyme immunoassay. Using universal polymerase chain reaction HIV-1, HIV-2, and SIV primers, we detected 1 (1.4%) HIV-1 sequence. Except for 1 sample, no molecular evidence for unusual HIV- or SIV-like strains in this sampling was found.