Effects of genital tract inflammation on human immunodeficiency virus type 1 V3 populations in blood and semen.

We have examined cell-free viral populations in the blood plasma and seminal plasma compartments of men infected with subtype C human immunodeficiency virus type 1 (HIV-1) using the V3-specific heteroduplex tracking assay (V3-HTA). We studied two cohorts of subjects who had visited either a sexually transmitted disease (STD) clinic for genital tract inflammation in the form of urethritis (n = 43) or a dermatology clinic (controls, n = 14) in Malawi. We have previously shown that the presence of urethritis is associated with an eightfold increase in virus load in the seminal plasma compartment (M. S. Cohen et al., Lancet 349:1868-1873, 1997). The purpose of this study was to determine whether genital tract inflammation and its treatment caused genetic instability in cell-free HIV-1 populations. In a cross-sectional analysis at study entry, three-fourths of the STD and control subjects had multiple V3 populations in their blood while 60% of the STD subjects and 79% of the control subjects had multiple V3 populations in their semen. Overall, one-fourth of all of the subjects showed discordance between results with blood and semen specimens when samples were compared for the presence and absence of subpopulations. When differences in the relative levels of abundance of bands were also taken into account, two-fifths of all of the subjects showed discordance between the compartments. Among the subset of subjects in whom multiple virus populations could be detected, half showed discordance between the compartments. There were no differences between STD and control cohorts for these comparisons of the compartments in this cross-sectional analysis at study entry. Longitudinal analysis of the viral populations from two separate clinic visits over 1 to 4 weeks showed that the complexity of each V3 population as measured by Shannon entropy was different in blood and semen at the two time points, indicating that the blood and semen constitute different compartments for HIV-1. The seminal plasma compartment was more dynamic than the blood plasma compartment for the STD subjects who were treated for urethritis, with changes being noted in the presence or absence of V3-HTA bands in the semen of 29% of these subjects but in the blood of only 9% of these subjects. However, the changes were generally small. Overall, our results suggest that 40% of male subjects show discordance between seminal and blood viral populations and that the complexity of each V3 population was different between the two compartments. Both of these results point to the partial independence of the seminal compartment as a viral niche within the body.

Trichomonas vaginalis as a cause of urethritis in Malawian men.

BACKGROUND AND OBJECTIVES: Trichomonas vaginalis is one of the most common sexually transmitted infections. In Malawi, rates of trichomoniasis in women are high. The prevalence of T. vaginalis infection in men is expected to be high but has not previously been documented. GOALS: We sought to determine the prevalence of trichomoniasis in Malawian men with and without urethritis, to evaluate a polymerase chain reaction detection assay for T. vaginalis in urethral swabs and to examine the effect of T. vaginalis infection on excretion of human immunodeficiency virus (HIV) in semen. STUDY DESIGN: Men presenting at the Sexually Transmitted Diseases (STD) and Dermatology Clinics in Malawi were enrolled in a cross-sectional study. We compared a polymerase chain reaction-based test for T. vaginalis detection with wet-mount microscopy and culture of urethral swabs. HIV serology was determined by enzyme-linked immunosorbent assay (ELISA), and HIV-1 RNA concentrations in semen were measured by quantitative nucleic acid sequence-based analysis. RESULTS: T. vaginalis was detected in 51 of 293 men. The estimated prevalence among symptomatic men was 20.8% and among asymptomatic men, 12.2%. Polymerase chain reaction performed with a sensitivity of 0.82 (95% CI: 0.66-0.92) and specificity of 0.95 (95% CI: 0.91-0.97) compared to wet-mount microscopy and culture. There was no difference in the rate of HIV seropositivity in men with and without T. vaginalis infection. However, in men with symptomatic urethritis, the median HIV RNA concentration in seminal plasma from men with T. vaginalis was significantly higher that in seminal plasma from HIV-positive men without trichomonas.

PIP: This study was conducted to determine the prevalence of trichomoniasis in Malawian men, to evaluate a polymerase chain reaction (PCR) detection assay for T. vaginalis in urethral swab samples, and to examine the effect of T. vaginalis infection on HIV excretion in the semen. There were 206 men with symptomatic urethritis in STD clinic and 127 asymptomatic men in the Dermatology Clinic who were enrolled from January to March 1996. Results, according to a wet-mount microscopy and urethral swabs culture combination, showed that, of 293 men, only 38 (13%) men were positive for T. vaginalis. The estimated prevalence among symptomatic and asymptomatic cases was 15.7% and 8.7%, respectively. The PCR yielded a sensitivity of 0.82 (95% CI: 0.66-0.92) and specificity of 0.95 (95% CI: 0.91-0.97); these were compared to the wet-mount microscopy and culture combination. Overall HIV seroprevalence of men was 51%, because gonococcal urethritis was shown to significantly increase seminal HIV RNA levels. The median HIV RNA concentration in seminal plasma from men with symptomatic urethritis plus T. vaginalis infection was significantly higher than in seminal plasma from HIV-positive men with symptomatic urethritis only. Since this study has several important limitations, a randomized clinical trial would be useful for determining whether urethritis cure rates can be significantly improved.

Characterization of V3 sequence heterogeneity in subtype C human immunodeficiency virus type 1 isolates from Malawi: underrepresentation of X4 variants.

We have examined the nature of V3 sequence variability among subtype C human immunodeficiency virus type 1 (HIV-1) sequences from plasma-derived viral RNA present in infected men from Malawi. Sequence variability was assessed by direct sequence analysis of the V3 reverse transcription-PCR products, examination of virus populations by a subtype C V3-specific heteroduplex tracking assay (V3-HTA), and selected sequence analysis of molecular clones derived from the PCR products. Sequence variability in V3 among the subtype C viruses was not associated with the presence of basic amino acid substitutions. This observation is in contrast to that for subtype B HIV-1, where sequence variability is associated with such substitutions, and these substitutions are determinants of altered coreceptor usage. Evolutionary variants in subtype C V3 sequences, as defined by the V3-HTA, were not correlated with the CD4 level in the infected person, while such a correlation was found with subtype B V3 sequences. Viruses were isolated from a subset of the subjects; all isolates used CCR5 and not CXCR4 as a coreceptor, and none was able to grow in MT-2 cells, a hallmark of the syncytium-inducing phenotype that is correlated with CXCR4 usage. The overall sequence variability of the subtype C V3 region was no greater than that of the conserved regions of gp120. This limited sequence variability was also a feature of subtype B V3 sequences that do not carry the basic amino acid substitutions associated with altered coreceptor usage. Our results indicate that altered coreceptor usage is rare in subtype C HIV-1 isolates in sub-Saharan Africa and that sequence variability is not a feature of the V3 region of env in the absence of altered coreceptor usage.

High levels of human immunodeficiency virus type 1 in blood and semen of seropositive men in sub-Saharan Africa.

High levels of human immunodeficiency virus type 1 (HIV-1) replication, as reflected in HIV-1 RNA concentrations in blood and semen, probably contribute to both rapid disease progression and enhanced sexual transmission. Semen and blood were collected from 49 Malawian and 61 US and Swiss (US/Swiss) HIV-1-seropositive men with similar CD4 cell counts and no urethritis or exposure to antiretroviral drugs. Median seminal plasma and blood plasma HIV-1 RNA concentrations were >3-fold (P = .034) and 5-fold (P = .0003) higher, respectively, in the Malawian men. Similar differences were observed in subsets of the Malawian and US/Swiss study groups matched individually for CD4 cell count (P = .035 and P < .002, respectively). These observations may help explain the high rates of HIV-1 sexual transmission and accelerated HIV-1 disease progression in sub-Saharan Africa.

Development of an antimicrobial susceptibility surveillance system for Neisseria gonorrhoeae in Malawi: comparison of methods.

Susceptibility of Neisseria gonorrhoeae to gentamicin, the primary treatment for gonorrhea in Malawi since 1993, was determined by using agar dilution MICs, E-test MICs, disc diffusion, and clinical cure rate. Agar dilution MICs were slightly higher in 1996 than in 1993 isolates, with a concomitant drop in the clinical cure rate. E-test MICs were substantially lower than agar dilution determinations, with only 77.4% within 1 log2 concentration.

PIP: Gentamicin (240 mg single intramuscular dose) was adopted in 1993 as the treatment of choice for gonorrhea in Malawi, following findings of high-level gonococcal resistance to penicillin, tetracycline, and streptomycin. This study sought to assess the clinical efficacy of gentamicin in the treatment of Neisseria gonorrhoeae 2 years after the onset of its widespread use and to evaluate the potential roles of the disc diffusion and E test in the development of a surveillance system for monitoring antimicrobial susceptibility. 206 consecutive men presenting to a sexually transmitted disease clinic in Lilongwe in 1996 and 198 urethritis patients treated at a hospital in Blantyre during 1992-93 were enrolled. The clinical cure rate for gentamicin treatment of gonorrhea was 91.8% in 1996 compared to 95.0% in 1993. The proportion of highly susceptible isolates declined from 17.2% in 1993 to 9.5% in 1996, but the majority of isolates remained in the moderately susceptible category. The E test yielded substantially lower minimum inhibitory concentrations (MICs) than the standard agar dilution assay, with only 77.4% within 1 log2 concentration. Disc diffusion was not helpful in determining the efficacy of gentamicin, since there was little difference in the range of disc diameters at different MICs as determined by agar dilution. Establishment of an MIC that clearly shows resistance to gentamicin was not possible. These findings suggest that the agar dilution assay remains the method of choice for MIC determination, although the less expensive E test may be capable of monitoring antibiotic efficacy under field conditions.

Reduction of concentration of HIV-1 in semen after treatment of urethritis: implications for prevention of sexual transmission of HIV-1. AIDSCAP Malawi Research Group.

BACKGROUND: Transmission of HIV-1 is predominantly by heterosexual contact in sub-Saharan Africa, where sexually transmitted diseases (STDs) are also common. Epidemiological studies suggest that STDs facilitate transmission of HIV-1, but the biological mechanism remains unclear. We investigated the hypothesis that STDs increase the likelihood of transmission of HIV-1 through increased concentration of the virus in semen. METHODS: HIV-1 RNA concentrations were measured in seminal and blood plasma from 135 HIV-1-seropositive men in Malawi; 86 had urethritis and 49 controls did not have urethritis. Men with urethritis received antibiotic treatment according to the guidelines of the Malawian STD Advisory Committee. Samples were analysed at baseline and at week 1 and week 2 after antibiotic therapy in urethritis patients, and at baseline and week 2 in the control group. FINDINGS: HIV-1-seropositive men with urethritis had HIV-1 RNA concentrations in seminal plasma eight times higher than those in seropositive men without urethritis (12.4 vs 1.51 x 10(4) copies/mL, p = 0.035), despite similar CD4 counts and concentrations of blood plasma viral RNA. Gonorrhoea was associated with the greatest concentration of HIV-1 in semen (15.8 x 10(4) copies/mL). After the urethritis patients received antimicrobial therapy directed against STDs, the concentration of HIV-1 RNA in semen decreased significantly (from 12.4 x 10(4) copies/mL to 8.91 x 10(4) copies/mL at 1 week [p = 0.03] and 4.12 x 10(4) copies/mL at 2 weeks [p = 0.0001]). Blood plasma viral RNA concentrations did not change. There was no significant change in seminal plasma HIV-1 RNA concentrations during the 2-week period in the control group (p = 0.421). INTERPRETATION: These results suggest that urethritis increases the infectiousness of men with HIV-1 infection. HIV-1-control programmes, which include detection and treatment of STDs in patients already infected with HIV-1, may help to curb the epidemic. Targeting of gonococcal urethritis may be a particularly effective strategy.