Thomas C. VanCott

Probability of HIV-1 transmission per coital act in monogamous, heterosexual, HIV-1-discordant couples in Rakai, Uganda

BACKGROUND The probability of HIV-1 transmission per coital act in representative African populations is unknown. We aimed to calculate this probability overall, and to estimate how it is affected by various factors thought to influence infectivity. METHODS 174 monogamous couples, in which one partner was HIV-1 positive, were retrospectively identified from a population cohort in Rakai, Uganda. Frequency of intercourse and reliability of reporting within couples was assessed prospectively. HIV-1 seroconversion was determined in the uninfected partners, and HIV-1 viral load was measured in the infected partners. Adjusted rate ratios of transmission per coital act were estimated by Poisson regression. Probabilities of transmission per act were estimated by log-log binomial regression for quartiles of age and HIV-1 viral load, and for symptoms or diagnoses of sexually transmitted diseases (STDs) in the HIV-1-infected partners. RESULTS The mean frequency of intercourse was 8.9 per month, which declined with age and HIV-1 viral load. Members of couples reported similar frequencies of intercourse. The overall unadjusted probability of HIV-1 transmission per coital act was 0.0011 (95% CI 0.0008-0.0015). Transmission probabilities increased from 0.0001 per act at viral loads of less than 1700 copies/mL to 0.0023 per act at 38 500 copies/mL or more (p=0.002), and were 0.0041 with genital ulceration versus 0.0011 without (p=0.02). Transmission probabilities per act did not differ significantly by HIV-1 subtypes A and D, sex, STDs, or symptoms of discharge or dysuria in the HIV-1-positive partner. INTERPRETATION Higher viral load and genital ulceration are the main determinants of HIV-1 transmission per coital act in this Ugandan population.

Protection of macaques against vaginal transmission of a pathogenic HIV-1/SIV chimeric virus by passive infusion of neutralizing antibodies.

The development of the human immunodeficiency virus-1 (HIV-1)/simian immunodeficiency virus (SIV) chimeric virus macaque model (SHIV) permits the in vivo evaluation of anti-HIV-1 envelope glycoprotein immune responses. Using this model, others, and we have shown that passively infused antibody can protect against an intravenous challenge. However, HIV-1 is most often transmitted across mucosal surfaces and the intravenous challenge model may not accurately predict the role of antibody in protection against mucosal exposure. After controlling the macaque estrous cycle with progesterone, anti-HIV-1 neutralizing monoclonal antibodies 2F5 and 2G12, and HIV immune globulin were tested. Whereas all five control monkeys displayed high plasma viremia and rapid CD4 cell decline, 14 antibody-treated macaques were either completely protected against infection or against pathogenic manifestations of SHIV-infection. Infusion of all three antibodies together provided the greatest amount of protection, but a single monoclonal antibody, with modest virus neutralizing activity, was also protective. Compared with our previous intravenous challenge study with the same virus and antibodies, the data indicated that greater protection was achieved after vaginal challenge. This study demonstrates that antibodies can affect transmission and subsequent disease course after vaginal SHIV-challenge; the data begin to define the type of antibody response that could play a role in protection against mucosal transmission of HIV-1.

ALVAC-SIV-gag-pol-env-Based Vaccination and Macaque Major Histocompatibility Complex Class I (A*01) Delay Simian Immunodeficiency Virus SIVmac-Induced Immunodeficiency

ABSTRACT T-cell-mediated immune effector mechanisms play an important role in the containment of human immunodeficiency virus/simian immunodeficiency virus (HIV/SIV) replication after infection. Both vaccination- and infection-induced T-cell responses are dependent on the host major histocompatibility complex classes I and II (MHC-I and MHC-II) antigens. Here we report that both inherent, host-dependent immune responses to SIVmac251 infection and vaccination-induced immune responses to viral antigens were able to reduce virus replication and/or CD4+ T-cell loss. Both the presence of the MHC-I Mamu-A*01 genotype and vaccination of rhesus macaques with ALVAC-SIV-gag-pol-env (ALVAC-SIV-gpe) contributed to the restriction of SIVmac251 replication during primary infection, preservation of CD4+ T cells, and delayed disease progression following intrarectal challenge exposure of the animals to SIVmac251 (561). ALVAC-SIV-gpe immunization induced cytotoxic T-lymphocyte (CTL) responses cumulatively in 67% of the immunized animals. Following viral challenge, a significant secondary virus-specific CD8+ T-cell response was observed in the vaccinated macaques. In the same immunized macaques, a decrease in virus load during primary infection (P = 0.0078) and protection from CD4 loss during both acute and chronic phases of infection (P = 0.0099 and P = 0.03, respectively) were observed. A trend for enhanced survival of the vaccinated macaques was also observed. Neither boosting the ALVAC-SIV-gpe with gp120 immunizations nor administering the vaccine by the combination of mucosal and systemic immunization routes increased significantly the protective effect of the ALVAC-SIV-gpe vaccine. While assessing the role of MHC-I Mamu-A*01 alone in the restriction of viremia following challenge of nonvaccinated animals with other SIV isolates, we observed that the virus load was not significantly lower in Mamu-A*01-positive macaques following intravenous challenge with either SIVmac251 (561) or SIVSME660. However, a significant delay in CD4+ T-cell loss was observed in Mamu-A*01-positive macaques in each group. Of interest, in the case of intravenous or intrarectal challenge with the chimeric SIV/HIV strains SHIV89.6P or SHIVKU2, respectively, MHC-I Mamu-A*01-positive macaques did not significantly restrict primary viremia. The finding of the protective effect of the Mamu-A*01 molecule parallels the protective effect of the B*5701 HLA allele in HIV-1-infected humans and needs to be accounted for in the evaluation of vaccine efficacy against SIV challenge models.

Epidemiologic and biologic characterization of a cohort of human immunodeficiency virus type 1 highly exposed, persistently seronegative female sex workers in northern Thailand

Characterization of persons highly exposed to human immunodeficiency virus (HIV)-1 who remain uninfected may help define protective immunity. Seventeen HIV-1-seronegative Thai female sex workers (CSWs) with epidemiologic evidence of exposure to HIV-1 were studied for humoral immune responses and phenotypic and genotypic analyses of HLA class I and CCR5 allelic profiles. Infected CSWs and low-risk HIV-1-seronegative Thai women were controls. Highly exposed, persistently seronegative (HEPS) CSWs did not differ from HIV-infected CSWs in HIV risks, condom use, or sexually transmitted diseases. Significant differences were seen in humoral immune responses: gp160-specific IgA responses were detected in cervicovaginal lavage fluids in 6 of 13 HEPS CSWs but 0 of 21 seronegative subjects. All women had wild-type CCR5. HEPS CSWs were more likely to have the HLA-B18 phenotype and genotype than were matched controls (corrected P=.018). Epidemiologic exposure to HIV-1 without apparent infection, an unusual distribution of HLA class I alleles, and HIV-1 gp160-specific IgA responses suggest a biologic basis for this phenomenon.

Broad Immunogenicity of a Multigene, Multiclade HIV-1 DNA Vaccine Boosted with Heterologous HIV-1 Recombinant Modified Vaccinia Virus Ankara

BACKGROUND A human immunodeficiency virus (HIV) vaccine that limits disease and transmission is urgently needed. This clinical trial evaluated the safety and immunogenicity of an HIV vaccine that combines a plasmid-DNA priming vaccine and a modified vaccinia virus Ankara (MVA) boosting vaccine. METHODS Forty healthy volunteers were injected with DNA plasmids containing gp160 of HIV-1 subtypes A, B, and C; rev B; p17/p24 gag A and B, and RTmut B by use of a needle-free injection system. The vaccine was administered intradermally or intramuscularly, with or without recombinant granulocyte macrophage colony-stimulating factor, and boosted with a heterologous MVA containing env, gag, and pol of CRF01A_E. Immune responses were monitored with HIV-specific interferon (IFN)-gamma and interleukin (IL)-2 ELISpot and lymphoproliferative assays (LPAs). RESULTS Vaccine-related adverse events were mild and tolerable. After receipt of the DNA priming vaccine, 11 (30%) of 37 vaccinees had HIV-specific IFN-gamma responses. After receipt of the MVA boosting vaccine, ELISpot assays showed that 34 (92%) of 37 vaccinees had HIV-specific IFN-gamma responses, 32 (86%) to Gag and 24 (65%) to Env. IFN-gamma production was detected in both the CD8(+) T cell compartment (5 of 9 selected vaccinees) and the CD4(+) T cell compartment (9 of 9). ELISpot results showed that 25 (68%) of 37 vaccinees had a positive IL-2 response and 35 (92%) of 38 had a positive LPA response. Of 38 subjects, a total of 37 (97%) were responders. One milligram of HIV-1 DNA administered intradermally was as effective as 4 mg administered intramuscularly in priming for the MVA boosting vaccine. CONCLUSION This HIV-DNA priming-MVA boosting approach is safe and highly immunogenic. TRIALS REGISTRATION International Standard Randomised Controlled Trial number: ISRCTN32604572 .

Drug Resistance Patterns, Genetic Subtypes, Clinical Features, and Risk Factors in Military Personnel with HIV-1 Seroconversion

Genetic variability is a central feature of HIV-1. The high frequency of mutations during HIV-1 replication leads to the development of viral quasi-species in vivo and contributes to genetic heterogeneity among HIV-1 isolates (1, 2). There is a growing appreciation that HIV-1 genetic diversity, including the existence of distinct genetic subtypes and the evolution of drug-resistant genotypes, can greatly affect the diagnosis and treatment of HIV-1 infection (1, 2). On the basis of DNA sequence analysis, HIV-1 has been classified into genetic subtypes, with subtypes A through I making up the major HIV-1 group (group M). The more genetically diverse groups, O and N, have also recently been described (1). For reasons that are not clear, HIV-1 subtypes are variably dispersed throughout the world. Some regions, such as central Africa and eastern Europe, have multiple circulating subtypes, whereas the distribution in other regions is more restricted (1, 3). Differences among HIV-1 subtypes can affect the sensitivity of some diagnostic assays of antibody (4) and plasma HIV-1 RNA (5, 6). Although non-subtype B infection has been reported in the United States, the prevalence in the U.S. population is unknown (7, 8). Widespread use of antiretroviral drugs has led to transmission of drug-resistant HIV-1, but the prevalence of resistant mutations in treatment-naive persons has not been thoroughly studied. Because all U.S. Navy and U.S. Marine Corps personnel are screened for HIV-1 infection at 1- to 3-year intervals, HIV-1 infection is often detected early and the seroconversion period can be estimated (9). This permits an investigation of the epidemiologic correlates and risk behaviors associated with the acquisition of non-subtype B and drug-resistant infections. Thus, we determined the prevalence of non-subtype B infection and genotypes associated with antiretroviral drug resistance in a well-characterized cohort of military personnel with recently acquired HIV-1 infection. Methods Study Design United States Navy or Marine Corps personnel with HIV-1 seroconversion who are assigned to military bases west of the Mississippi River in the United States or in the Pacific region overseas are referred to the Navy Medical Center San Diego for initial and follow-up HIV-1 evaluations. Between February 1997 and February 1998, 99 of 141 personnel referred were eligible on the basis of documented seroconversion within the past 3 years. Ninety-five of 99 patients enrolled and signed a consent form approved by the institutional review board of the Navy Medical Center San Diego. Thirty-two patients were being referred for their first HIV evaluation, and 63 were enrolled during a follow-up visit. Blood was obtained to determine genetic subtype and the presence of antiretroviral drug resistance. All patients completed a self-administered risk factor questionnaire labeled with a unique code number. Surveys were sealed in an envelope and placed in a locked drop box that was emptied weekly by off-site data entry personnel. Clinical research staff extracted clinical and laboratory information from the medical record and sent this information, identified by code number, to the data entry site. Laboratory Analysis Seroconversion was documented by a previous negative result on whole-virus HIV-1 enzyme immunoassay followed by a positive result on enzyme immunoassay and a confirmatory positive result on Western blot assay. Lymphocyte subset analysis was performed by flow cytometry, and plasma HIV-1 RNA (available after July 1996) was measured by quantitative reverse transcription polymerase chain reaction assay (Amplicor HIV-1 Monitor assay, Roche Molecular Systems, Branchburg, New Jersey). Genetic subtyping of HIV-1 was performed by using a two-step algorithm. Sera were screened by using a competitive binding enzyme immunoassay with peptides derived from the third variable loop of the HIV-1 envelope glycoprotein. For samples that were serologically reactive to non-subtype B peptides, DNA was extracted from corresponding peripheral blood mononuclear cells, and sequence analysis was performed over a 640-base pair segment of the HIV-1 envelope gene (10). Testing for viral drug resistance was successfully performed in 31 of the 32 therapy-naive patients. Plasma-derived viral RNA was reverse transcribed into complementary DNA, amplified by polymerase chain reaction, and directly sequenced by using an automated ABI Sequencer (Applied Biosystems, Foster City, California). Consensus DNA sequences for the protease and reverse transcriptase genes from each participant were examined for mutations associated with HIV-1 antiretroviral resistance (11). Statistical Analysis Descriptive analysis of demographics, risk behaviors, and laboratory results were performed. Duration of HIV infection was calculated starting from the time of onset, which we estimated as the midpoint between the last negative and first positive result on HIV enzyme immunoassay. Bivariate analyses using odds ratios and 95% CIs determined by the Fisher exact method were done to compare the treatment-naive patients with drug-resistant infection and those with drug-sensitive infection. Statistical analyses were done by using Epi-Info, version 6.02 (Centers for Disease Control and Prevention, Atlanta, Georgia). Results Patients Characteristics and risk exposures of the cohort are summarized in Table 1. Duration of infection and laboratory results are presented for the entire cohort on initial evaluation and for the treatment-naive cohort at the time of enrollment and antiretroviral testing. Risk behaviors known to be associated with acquisition of HIV-1 were reported during the period of seroconversion in all but two patients. Table 1. Characteristics and Risk Exposures of Military Personnel with HIV-1 Seroconversion Laboratory Analysis The CD4 cell counts, plasma HIV RNA levels, and results of syphilis and hepatitis serologic testing in study patients are summarized in Table 1. Combined serologic and genetic analysis revealed that 7 of 95 patients were infected with HIV-1 genetic subtype E (7.4%); the remaining patients were infected with subtype B. Eight of 31 treatment-naive patients (26% [95% CI, 12% to 46%]) had one or more primary mutations that have been associated with phenotypic drug resistance (11) (Table 2). All reverse transcriptase mutations were associated with resistance to zidovudine, lamivudine, or nevirapine-delavirdine. Of the 4 patients with reverse transcriptase mutations, 2 (patients 1 and 2) had mutations that could confer resistance to both nucleoside and non-nucleoside reverse transcriptase inhibitors. Table 2. Drug-Resistant Mutations in Eight Treatment-Naive Patients Characteristics of Patients with Subtype E Infection and Drug-Resistant Mutations Infection with HIV-1 subtype E was documented in 6 men and 1 woman; all 6 men reported sexual contact during short deployments in Thailand. The female patient reported having sex with a man on active duty in the United States. Compared with subtype B-infected patients, subtype E-infected patients were more likely to be heterosexual (100% and 38%), to have had overseas exposure (86% and 27%), and to report sex with commercial sex workers (86% and 15%). Comparisons between patients with wild-type (n=23) and treatment-resistant (n=8) genotypes are listed in Table 1. Discussion We found a high prevalence of non-B genetic subtypes and antiretroviral drug-resistant mutations among treatment-naive military personnel with recently acquired HIV-1 infection. Seven of 95 patients (7.4%) were infected with HIV-1 subtype E. Eight of 31 treatment-naive patients (26%) had primary drug-resistant mutations; of these 8 patients, 3 had mutations in the reverse transcriptase gene only, 4 had mutations in the protease gene only, and 1 had mutations in both the reverse transcriptase and protease genes. Although our study is limited to a single clinical referral center and the sample is small, the results are generally similar to those of other studies that focused on selected drug-resistant mutations. Among several recent studies, 6% to 13% of treatment-naive patients had mutations for zidovudine resistance (12-14). Although data on the prevalence of mutations in the protease gene have not yet been published, transmission of HIV-1 that is resistant to multiple reverse transcriptase and protease inhibitors has been reported (15). Our data suggest that patients infected with drug-resistant virus were more likely to have acquired HIV in the United States and to report sexual contact with a person who is known to be infected with HIV; they were also less likely to be heterosexual. Prospective studies of the treatment responses of drug-naive patients with resistant genotypes have not been performed, but some data suggest that baseline nucleoside analogue mutations can diminish the potency and duration of viral suppression by commonly used nucleoside analogue combinations (16). In addition, several reports suggest an inherent decrease in susceptibility to reverse transcriptase or protease inhibitors in some HIV-1 genetic subtypes (17). Thus, both the transmission of acquired mutations and the genetic subtype of HIV-1 may have important implications for treatment of HIV-1 disease. Although we (7) and others (8) have previously described the introduction of non-subtype B infections into the United States, the current study is the first use of a large cohort of recently infected persons to describe the prevalence and associated risk factors for non-subtype B infection. Six of the seven patients infected with HIV-1 subtype E reported sexual contact during short deployments to Thailand; in contrast, most cases of subtype B infection were acquired in the United States. The acquisition of sexually transmitted diseases and HIV during overseas travel is not unique to the military; 5% to 20% of travelers report having sex with a new partner while abroad (18).

Neutralizing Antibodies Elicited by Immunization of Monkeys with DNA Plasmids and Recombinant Adenoviral Vectors Expressing Human Immunodeficiency Virus Type 1 Proteins

ABSTRACT Immunization with recombinant serotype 5 adenoviral (rAd5) vectors or a combination of DNA plasmid priming and rAd5 boosting is known to elicit potent immune responses. However, little data exist regarding these immunization strategies and the development of anti-human immunodeficiency virus type 1 (HIV-1) neutralizing antibodies. We used DNA plasmids and rAd5 vectors encoding the HIV-1 89.6P or chimeric HxB2/BaL envelope glycoprotein to immunize macaque monkeys. A single rAd5 immunization elicited anti-Env antibody responses, but there was little boosting with subsequent rAd5 immunizations. In contrast, rAd5 boosting of DNA-primed monkeys resulted in a rapid rise in antibody titers, including the development of anti-HIV-1 neutralizing antibodies. The potency and breadth of neutralization were evaluated by testing plasma against a panel of 14 clade B primary isolates. Moderate levels of plasma neutralizing activity were detected against about one-third of the viruses tested, and immunoglobulin G fractionation demonstrated that virus neutralization was antibody mediated. After a challenge with a chimeric simian-human immunodeficiency virus (SHIV89.6P), an anamnestic neutralizing antibody response was observed, although the breadth of the response was limited to the subset of viruses that were neutralized after the primary immunization. These data are the first detailed description of the anti-HIV-1 neutralizing antibody response in nonhuman primates elicited by DNA and rAd5 immunization. In addition to the well-established ability of DNA priming and rAd5 boosting to elicit potent anti-HIV-1 cellular immune responses, this immunization strategy elicits anti-HIV-1 neutralizing antibodies and therefore can be used to study novel Env immunogens designed to elicit more potent neutralizing antibodies.

Performance of the OraQuick Rapid Antibody Test for Diagnosis of Human Immunodeficiency Virus Type 1 Infection in Patients with Various Levels of Exposure to Highly Active Antiretroviral Therapy

ABSTRACT With oral mucosal transudate and serum samples from 101 human immunodeficiency virus type 1 (HIV-1)-infected subjects and 100 HIV-1-negative volunteers, the OraQuick HIV-1 test demonstrated 100% specificity and 96% sensitivity. Four false-negative subjects, who were characterized by early initiation of effective antiretroviral therapy, demonstrated waning serum anti-gp41 titers and Western blot band intensities.

Multiple introductions of HIV-1 subtype E into the western hemisphere

There are nine recognised genetic subtypes of HIV-1, and the epidemic in Southeast Asia is largely due to subtype E. We have investigated HIV-1 viral subtypes in 11 Uruguayan military personnel, six with infection acquired during a United Nations deployment to Cambodia and five with infection acquired in South America. We found subtype E in five of the six infections acquired in Southeast Asia, and subtype B in all five of the domestically acquired cases. These findings document multiple introductions of HIV-1 subtype E into the western hemisphere and mean that the genetic diversity of the global HIV-1 pandemic must be considered in strategies for epidemic control.

Modulation of Antigen-Specific Humoral Responses in Rhesus Macaques by Using Cytokine cDNAs as DNA Vaccine Adjuvants

ABSTRACT An important limitation of DNA immunization in nonhuman primates is the difficulty in generating high levels of antigen-specific antibody responses; strategies to enhance the level of immune responses to DNA immunization may be important in the further development of this vaccine strategy for humans. We approached this issue by testing the ability of molecular adjuvants to enhance the levels of immune responses generated by multicomponent DNA vaccines in rhesus macaques. Rhesus macaques were coimmunized intramuscularly with expression plasmids bearing genes encoding Th1 (interleukin 2 [IL-2] and gamma interferon)- or Th2 (IL-4)-type cytokines and DNA vaccine constructs encoding human immunodeficiency virus Env and Rev and simian immunodeficiency virus Gag and Pol proteins. We observed that the cytokine gene adjuvants (especially IL-2 and IL-4) significantly enhanced antigen-specific humoral immune responses in the rhesus macaque model. These results support the assumption that antigen-specific responses can be engineered to a higher and presumably more desirable level in rhesus macaques by genetic adjuvants.