Teresa M. Brown

U.S. Human Immunodeficiency Virus Type 1 Epidemic: Date of Origin, Population History, and Characterization of Early Strains

ABSTRACT Human immunodeficiency virus (HIV) type 1 subtype B sequences (whole envelope and the p17 region of gag) were obtained from peripheral blood mononuclear cell samples collected in 1981 from seven HIV-infected U.S. individuals and in 1982 from one infected Canadian resident. Phylogenetic and nucleotide distance analyses were performed by using database sequences representing North American strains collected from 1978 to 1995. The estimated phylogeny was starlike, with early strains represented on different lineages. When sequences were grouped by years of collection, nucleotide distance comparisons demonstrated an increase in diversity over time and indicated that contemporary strains are more closely related to early epidemic strains than to each other. Using a recently developed likelihood ratio reduction procedure, the date of origin of the U.S. epidemic was estimated to be 1968 ± 1.4 years. A coalescent approach was also used to estimate the population history of the U.S. subtype B epidemic. Our analyses provide new information that implies an exponential growth rate from the beginning of the U.S. HIV epidemic. The dating results suggest a U.S. introduction date (or date of divergence from the most recent common ancestor) that precedes the date of the earliest known AIDS cases in the late 1970s. Furthermore, the estimated epidemic growth curve shows a period of exponential growth that preceded most of the early documented cases and also indicates a leveling of prevalence rates in the recent past.

The effect of maternal viral load on the risk of perinatal transmission of HIV-1

Objective:To determine the effect of maternal viral load at delivery on the risk of perinatal transmission of HIV-1. Design:A nested case–control study within a prospectively followed cohort of HIV- 1-infected pregnant women and their infants. Setting:The multicenter New York City Perinatal HIV Transmission Collaborative Study. Participants:Fifty-one women who gave birth to HIV-1-infected infants were frequency-matched within CD4+ cell count quintiles with 54 non-transmitting mothers. Main outcome measures:Maternal quantity of HIV-1 viral RNA was assayed in plasma obtained near delivery using the nucleic acid sequence-based amplification assay system. Results:Viral RNA was detected in 73 (70%) out of 105 women and the median viral load was 16 000 RNA copies/ml in transmitters and 6600 in non-transmitters (P < 0.01). When adjusted for maternal CD4+ count near delivery, women with measurable viral load were nearly sixfold more likely to transmit HIV-1 than women with viral load below detection [adjusted odds ratio (AOR), 5.8; 95% confidence interval (CI), 2.2–15.5]. The odds ratio for perinatal transmission of log10 viral load, adjusted for CD4 count was 2.7 (95% CI, 1.5–5.1). When stratified by the stage of HIV-1 disease, the only group with significant association between log10 viral load and transmission were AIDS-free women with CD4+ count > 500 x 106/l (AOR, 9.1; 95% CI, 2.6–31.5). Conclusions:High maternal viral load increases the likelihood of perinatal transmission of HIV-1 in women without AIDS and advanced immunosuppression. HIV-1-infected pregnant women without advanced disease, shown by others to have the lowest risk of perinatal transmission, may benefit the most from efforts to identify and decrease viral load at delivery.

A Novel Recombinant Multisubunit Vaccine against Chlamydia

The administration of an efficacious vaccine is the most effective long-term measure to control the oculogenital infections caused by Chlamydia trachomatis in humans. Chlamydia genome sequencing has identified a number of potential vaccine candidates, and the current challenge is to develop an effective delivery vehicle for induction of a high level of mucosal T and complementary B cell responses. Vibrio cholerae ghosts (VCG) are nontoxic, effective delivery vehicles with potent adjuvant properties, and are capable of inducing both T cell and Ab responses in mucosal tissues. We investigated the hypothesis that rVCG could serve as effective delivery vehicles for single or multiple subunit chlamydial vaccines to induce a high level of protective immunity. rVCG-expressing chlamydial outer membrane proteins were produced by a two-step genetic process, involving cloning of Omp genes in V. cholerae, followed by gene E-mediated lysis of the cells. The immunogenicity and vaccine efficacy of rVCG-expressing single and multiple subunits were compared. Immunologic analysis indicated that i.m. immunization of mice with either vaccine construct induced a strong mucosal and systemic specific Th1 response against the whole chlamydial organism. However, there was an immunogenic advantage associated with the multiple subunit vaccine that induced a higher frequency of Th1 cells and a relatively greater ability to confer protective immunity, compared with the single subunit construct. These results support the operational theory that the ability of a vaccine to confer protective immunity against Chlamydia is a function of the level of Th1 response elicited.

Sensitivity and specificity of a qualitative RNA detection assay to diagnose HIV infection in young infants. Perinatal AIDS Collaborative Transmission Study.

OBJECTIVE To evaluate the sensitivity and specificity of an RNA detection assay for diagnosing perinatal HIV infection. METHODS Plasma and serum specimens taken during the first 3 months of life from HIV-infected and uninfected children enrolled in a cohort study were assayed for HIV RNA using the qualitative nucleic acid sequence-based amplification (NASBA) kit. Sensitivity, specificity, and predictive values were calculated. NASBA results from infected children were compared with DNA PCR results from the same blood samples. Autoantibody patterns of suspected false-positive specimens were compared with those of subsequent specimens from the same child to exclude specimen labelling errors. RESULTS Amongst 131 specimens from 105 HIV-infected children, the sensitivity of the qualitative NASBA assay was 13 out of 34 [38%; 95% confidence interval (CI), 22-56] at < 7 days, 56 out of 58 (97%; 95% CI, 88-100) at 7-41 days, and 37 out of 39 (95%; 95% CI, 83-99) at 42-93 days of life. Of 252 specimens from 206 uninfected children, six tested positive and one tested indeterminate by NASBA. Four of these positive specimens had discordant autoantibody patterns suggesting mislabelling; excluding these, the test specificity was 245 out of 248 (99%; 95% CI, 97-100). Amongst 128 paired specimens from infected children, NASBA results were more often positive than those from DNA PCR (103 versus 92; P=0.01). Amongst infants with specimens drawn in the first week of life, the proportion born after > 4 h of membrane rupture was greater amongst those testing negative (81%) than those testing positive (46%; P=0.05). CONCLUSIONS The qualitative NASBA RNA assay is highly specific and more sensitive than DNA PCR. Qualitative RNA assays may be useful for diagnosing and excluding perinatal HIV infection in children after the first week of life for such purposes as initiating antiretroviral therapy and other treatment, resolving parental uncertainty, determining timing of transmission, and providing endpoints for intervention trials.

Effect of maternal CD4 + cell count, acquired immunodeficiency syndrome, and viral load on disease progression in infants with perinatally acquired human immunodeficiency virus type 1 infection

Among a cohort of 152 infants perinatally infected with human immunodeficiency virus type 1, and their mothers, we correlated infant outcome with material CD4+ lymphocyte count and the presence of maternal acquired immunodeficiency syndrome near delivery. In a subset of 50 mother-infant pairs, we also correlated infant outcome with maternal quantitative viral burden as measured by the nucleic acid sequence based amplification system. We found that low maternal CD4+ cell count and high viral burden were associated with decreased time to category C disease or death in infants infected with human immunodeficiency virus type 1. In a multivariate analysis, high maternal viral load and maternal acquired immunodeficiency syndrome were independently associated with shorter time to category C disease or death in infants with human immunodeficiency virus type 1 infection. High viral load in pregnant women, independent of the presence of advanced maternal disease, appears to increase the risk of rapidly progressive disease in their infected offspring.

Genetic Analysis of Human Immunodeficiency Virus Type 1 Strains in Kenya: A Comparison Using Phylogenetic Analysis and a Combinatorial Melting Assay

We surveyed human immunodeficiency virus (HIV) subtype distribution from peripheral blood mononuclear cells (PBMCs) collected in 1995 from 24 HIV-1-infected Kenyan residents (specimens from predominantly male truck drivers and female sex workers near Mombasa and Nairobi). Processed lysates from the PBMC samples were used for env amplification, directly sequenced, and analyzed by phylogenetic analysis. Envelope amplification products were also used for analysis in a polymerase chain reaction (PCR)-based assay, called the combinatorial melting assay (COMA). Results of the two tests were compared for assignment of subtype for this Kenyan cohort. The COMA, a PCR capture technique with colorimetric signal detection, was used with HIV reference subtype strains as well as regional (East Africa) HIV strains for subtype identification. Performance of the COMA was at 100% concordance (24 of 24) as compared with DNA sequencing analysis. Phylogenetic analysis showed 17 isolates to be subtype A, 3 subtype D, and 4 subtype C viruses. This may represent an increase in subtype C presence in Kenya compared with previously documented reports. The COMA can offer advantages for rapid HIV-1 subtype screening of large populations, with the use of previously identified regional strains to enhance the identification of local strains. When more detailed genetic information is desired, DNA sequencing and analysis may be required.

Early detection of reverse transcriptase activity in plasma of neonates infected with HIV-1: a comparative analysis with RNA-based and DNA-based testing using polymerase chain reaction.

Summary: Plasma viral load from 71 HIV‐1‐infected neonates was measured by using Amp‐RT, an ultrasensitive quantitative reverse transcriptase (RT) assay and by nucleic acid sequence‐based amplification (NASBA), an RNA‐based quantitative assay. Results were then compared with those obtained from detection of proviral DNA in peripheral blood mononuclear cells (PBMCs) by polymerase chain reaction (PCR) using Turnbull analysis. At 5 days of life, 50% of neonates were positive by Amp‐RT, 30% were NASBA positive, and 20% were DNA‐PCR positive. Through the first 12 days of life, Amp‐RT was more sensitive than either NASBA or DNA‐PCR in detecting HIV‐1 infection. Amp‐RT values correlated well with NASBA RNA values, with an overall Pearsons r = 0.63 (95% confidence interval [CI], 0.40‐0.78). In proportional hazards analysis of infants aged 14 to 61 days (N = 31), a one‐log increase in RNA‐based viral load was associated with a > fivefold risk of disease progression when using the U.S. Centers for Disease Control and Prevention (CDC) clinical Category C (CDC‐C) or death as an endpoint (p = .014). Kaplan‐Meier analysis of these data found that RNA viral loads were able to predict disease progression using CDC‐C/death as an endpoint (p = .013). Early quantitative viral load measurements may assist clinicians in diagnosing HIV‐1 infection, stratifying risk of disease progression, and implementing a treatment plan using highly active antiretroviral therapy for infants within the first few weeks of life.

Molecular analysis in support of an investigation of a cluster of HIV-1-infected women.

An investigation of a possible single-source sexual transmission case was conducted in upstate New York in 1997-1998 (MMWR 1999;48:413-416). Of 42 primary female contacts with the putative male index case, 13 tested positive for HIV infection. Blood was available for DNA sequencing (C2V3C3 region of the env gene and the p17-coding region of gag) from 10 of the 13 women, 1 HIV-infected secondary contact, and 2 HIV-infected persons from the community, but not from the index cam. Phylogenetic and distance analyses were performed with the inclusion of reference HIV subtype strains for both the env and gag gene regions, as was the two regions combined. A high degree of relatedness was found among DNA sequences of the 10 primary contacts that excluded reference strains, the secondary contact, and the community HIV control subjects. In conclusion, phylogenetic analysis of HIV strains in an epidemiologic investigation is highly useful in support of cluster identification, even without sampling from the putative index patient.

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

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

Optimal methods for collecting and storing vaginal specimens for prostate-specific antigen testing in research studies☆

BACKGROUND Prostate-specific antigen (PSA) detected in vaginal fluid can be used in studies of HIV/sexually transmitted infection (STI) and pregnancy prevention as an alternative to relying on participant reports of exposure to semen. Optimal methods for collecting and storing specimens for this testing have not been determined. STUDY DESIGN We conducted a controlled, in vitro experiment of 550 specimens spiked with semen to determine the effects of swab type (five types), storage conditions of the swabs (room temperature with or without desiccant or at -80°C without desiccant) and time from collection to testing (seven intervals over the course of 12 months) on the identification of PSA. We performed factorial analysis of variance to identify factors influencing PSA detection. RESULTS Concentrations of PSA detected in the swabs declined with time of storage over the 1-year experiment (p<.01). The 1-mL, rayon-tipped swab stored immediately at -80°C following collection performed best. CONCLUSIONS If immediate testing or freezer storage is not feasible, investigators should use a swab with 1-mL capacity with processing and testing as soon as possible after specimen collection.