Genotypic analysis of the protease and reverse transcriptase of HIV type 1 subtype C isolates from antiretroviral drug-naive adults in Malawi.

The protease (PR) and reverse transcriptase (RT) regions of HIV-1 isolates from 21 antiretroviral (ARV)-naive Malawian adults were sequenced and analyzed to determine the prevalence of drug resistance-associated mutations in this population. Phylogenetic analysis confirmed that all isolates grouped with HIV-1 subtype C, the predominant subtype in Malawi. No major mutations associated with resistance to PR inhibitors (PIs), nucleoside RT inhibitors (NRTIs), or nonnucleoside RT inhibitors (NNRTIs) were found. In contrast, accessory mutations were found in the protease region at positions 10, 20, 36, 63, 77, and 93, and in the RT region at positions 118, 211, and 214. Further studies will be needed to determine the clinical impact of these polymorphisms on viral susceptibility to existing antiretroviral drugs.

Improving sexually transmitted disease management in the private sector: the Jamaica experience.

OBJECTIVES: To improve the quality of sexually transmitted disease (STD) case management in Jamaica by providing comprehensive continuing medical education to private practitioners who manage 60% of all STDs on the island. METHODS: Six half-day STD seminars were presented at 3-4-month intervals and repeated in three separate geographic locations. All Jamaican practitioners received invitations. The subjects were as follows: urethritis, genital ulcer disease, HIV infection, vaginal discharge syndrome, STDs in children and adolescents, and a review seminar. The program effectiveness was evaluated with a written, self-reported pre-test and a telephone post-test that measured changes in clinical management. RESULTS: Six hundred and twenty eight practitioners attended at least one seminar. Comparing pre- versus post-test scores, there were practitioner improvement trends in all four of the general STD management categories: counseling/education (69.8-73.3%; P > 0.05); diagnostics/screening (57.2-71.0%; P= 0.042); treatment (68.3-74.5%; P> 0.05); and knowledge (66.4-83.2%; P= 0.002). Obtaining syphilis serologies during pregnancy rose from 38.3 to 83.8% (P= 0.001), and providing effective treatment for gonorrhea rose from 57.8 to 81.1 % (P= 0.002), but correct responses on treatment for mucopurulent cervicitis at the post-test was a low 32.4%. CONCLUSION: The introduction of continuing medical education for improved STD care targeting private physicians in Jamaica was successful based on high attendance rates and self-reported STD management practices. However, efforts should continue to address the weaknesses found in STD management and counseling and to reach the providers who did not participate. In the global effort to reduce HIV transmission by improving STD care services, continuing education programs that target the private sector can be successful and should be included as a standard activity to improve care and provide a public/private link to STD/HIV control.

PIP: The Jamaican Ministry of Health has estimated that over 60% of all sexually transmitted diseases (STDs) are managed within the private sector, where 800 (66%) of the country's 1200 registered physicians practice. To improve the quality of STD case management provided by these practitioners, the Medical Association of Jamaica organized a series of 6 half-day seminars repeated at 3-4 month intervals in three geographic locations between December 1993 and July 1995. Topics addressed included urethritis, genital ulcer disease, HIV/AIDS, vaginal discharge, pelvic inflammatory disease, and STDs in children and adolescents. A total of 628 private practitioners attended at least one seminar and almost half the physicians attended two or more. Comparisons of scores on a written pretest completed before the seminar and those from a post-test conducted by telephone after the seminar revealed significant improvements in all four general STD management categories: counseling/education, diagnostics/screening, treatment, and knowledge. The proportion of practitioners who obtained syphilis serologies during pregnancy rose from 38.3% to 83.8% and those providing effective treatment for gonorrhea increased from 57.8% to 81.1%. Overall, 96% of practitioners were providing some level of risk-reduction counseling at the time of the post-test and 74% were prescribing correct treatment regimens. Ongoing education and motivation by the national STD control program or the Medical Association are recommended to improve STD case management even further.

Plasma HIV burden in Malawian children co-infected with rotavirus.

Fifty-eight HIV-infected children with acute rotavirus diarrhea were tested for plasma HIV RNA. There was no difference between acute and convalescent mean viral loads, and little change in CD4 cell counts. Compared with the 16 children who died within 4 weeks, 31 survivors had slightly lower viral loads at presentation and significantly higher CD4 cell counts. Low CD4 cell counts, but not HIV-1-RNA concentrations, were predictive of Death. Local, enteric rotavirus infection did not appear to affect blood HIV viral load or CD4 cell counts in this small group of children.

The effect of Plasmodium falciparum malaria on HIV-1 RNA blood plasma concentration.

OBJECTIVES: This study was undertaken to determine the relative effect of malaria infection on HIV concentration in blood plasma, and prospectively to monitor viral concentrations after antimalarial therapy. DESIGN: A prospective, double cohort study was designed to compare the blood HIV-1 RNA concentrations of HIV-positive individuals with and without acute malaria illness. Subjects were followed for 4 weeks after successful malaria therapy, or for 4 weeks from enrollment (controls). METHODS: Malawian adults with symptomatic Plasmodium falciparum parasitemia (malaria group) and asymptomatic, aparasitemic blood donors (control group) were tested for HIV-1 antibodies to identify appropriate study groups. The malaria group received antimalarial chemotherapy only and were followed with sequential blood films. In both groups, blood plasma HIV-1 RNA viral concentrations were determined at enrollment and again at 1, 2 and 4 weeks. RESULTS: Forty-seven malaria patients and 42 blood donors were enrolled. At enrollment blood plasma HIV-1 RNA concentrations were approximately sevenfold higher in patients with malaria than in blood donors (medians 15.1 x 10(4) and 2.24 x 10(4) copies/ml, respectively, P = 0.0001). No significant changes in median HIV-1 concentrations occurred in the 21 blood donors followed to week 4 (P = 0.68). In the 27 subjects successfully treated for malaria who were followed to week 4, a reduction in plasma HIV-1 RNA was observed from a median of 19.1 x 10(4) RNA copies/ml at enrollment, to 12.0 x 10(4) copies/ml at week 4, (P = 0.02). Plasma HIV-1 concentrations remained higher in malaria patients than controls (median 12.0 x 10(4) compared with 4.17 x 10(4) copies/ml, P = 0.086). CONCLUSIONS: HIV-1 blood viral burden is higher in patients with P. falciparum malaria than in controls and this viral burden can, in some patients, be partly reduced with antimalarial therapy.

Selective screening for chlamydial infection: which criteria to use?

BACKGROUND: Screening sexually active women for Chlamydia trachomatis is necessary to detect asymptomatic infections. Selective screening is a common strategy because universal screening is too costly in many settings. In order to guide local programs in the choice of selective screening criteria, we examined the performance of previously proposed screening criteria for C. trachomatis. METHODS: A clinic-based, cross-sectional study was conducted in public family planning and sexually transmitted disease (STD) clinics in ten counties in North Carolina. Women (n = 4471 in family planning and n = 2201 in STD clinics) undergoing pelvic examination were enrolled consecutively. Nine sets of screening criteria, including age alone, were compared using sensitivity, specificity, number of tests required and receiver-operator characteristic (ROC) analysis. All women underwent testing with ligase chain reaction assay of cervical specimens to identify C trachomatis infection. RESULTS: The prevalence of C. trachomatis was 7.8% and 11.0% in family planning and STD clinics, respectively. The sensitivities of published criteria ranged from 0.50 to 0.97. Specificities ranged from 0.05 to 0.66. In family planning clinics, the best performing criteria would detect 84% of infections while screening 51% of women. In STD clinics, the same criteria would detect 83% of infections but require testing 67% of women. Testing women aged < or =22 would detect 77% of infections in family planning and 74% of infections in STD clinics, while testing 51% and 48% of the women, respectively. CONCLUSIONS: When site-specific criteria cannot be developed, age alone is an acceptable strategy for selective screening for chlamydial infection.

Genital ulcer disease. Management in the HIV era.

What are the main causes of genital ulcer disease in the United States? Why is clinical diagnosis so difficult? What role will molecular diagnostics have in the future? The authors answer these questions and provide a functional approach to successful management of three common types of sexually transmitted genital ulcers.

In-hospital mortality rates and HIV: a medical ward review, Lilongwe, Malawi.

In order to determine inpatient hospital mortality rates, causes of mortality and characteristics of inpatients at Kamuzu Central Hospital (KCH) in Lilongwe, Malawi, we conducted a prospective observational study of all patients admitted to KCH medical ward from 20 September 2008 to April 2, 2009. All admission diagnoses, HIV status and antiretroviral therapy (ART) use were recorded. Patients' vital status was determined at discharge. A descriptive analysis and two logistic regression models were used for the analysis. Of the 1895 enrolled patients, the overall hospital mortality rate was 14.6%, substantially higher among known HIV-infected patients (24.2% versus 10.8%, P = 0.0009) and men (17.1% versus 12%, P = 0.033). Patients with multiple diagnoses had significantly higher mortality (odds ratio [OR] 2.33; 95% confidence interval [CI] 1.47, 3.71). Most patients (62.3%) had unknown HIV status at admission. Among HIV-infected patients, ART use did not reduce hospital mortality or alter the spectrum of diseases. The majority of diagnoses were infectious (63.4%). The high inpatient mortality rate, especially among HIV-infected patients combined with the limited spectrum of diagnoses, emphasizes the need for improved inpatient management and diagnostic services. Expansion of HIV testing is warranted. Despite the rollout of ART, there remains a significant need for treatment of HIV-infected individuals.

Can HIV-1 infections in Africa provide insights into acquired immunity to malaria?

PIP: This article examines the feasibility of a Ugandan study by James Whitworth and colleagues on the effect of HIV-1 infection and advancing immunosuppression on falciparum parasitemia and clinical malaria. The researchers observed the interaction between malaria and HIV-1 infection in adults with acquired immunity to malaria. Data were collected at the routine quarterly visits and also during interim visits prompted by symptoms. Both clinical malaria and parasitemia were rare. Results showed that HIV-1-positive individuals were more likely to be parasitemic during routine visits than were HIV-1-negative participants and the risk of finding clinical malaria was significantly higher among HIV-1-positive than among HIV-1-negative individuals, whether the visits were routine or interim. These data provide a fascinating insight into the nature of acquired immunity to malaria. Whitworth and colleagues have shown that HIV-1 infection is associated with increased prevalence and intensity of Plasmodium falciparum infection in adults with acquired immunity to malaria. However, this study should include measures of factors influencing HIV transmission and the development of AIDS, as well as variables associated with malaria transmission and response to treatment.^ieng

Management of vaginal discharge in women treated at a Jamaican sexually transmitted disease clinic: use of diagnostic algorithms versus laboratory testing.

The management of cervical infections is difficult in developing countries because laboratory facilities for diagnosing these infections are seldom available; therefore, syndrome-based management has been recommended by the World Health Organization (WHO). However, such alternative approaches need to be evaluated in real field settings. We used algorithms (flowcharts) for syndromic management of abnormal vaginal discharge to treat 752 women who presented at a Jamaican sexually transmitted disease (STD) clinic. Laboratory testing revealed cervical infection (gonococcal and/or chlamydial) in 34% of these women; trichomoniasis was documented for 25%; and at least one STD was documented for 54% of the women. Use of a clinical algorithm for diagnosing cervical infection was 73% sensitive (95% CI, 67-78) and 55% specific (95% CI, 49-62) when compared with laboratory testing. The risk-assessment-inclusive flowchart developed by WHO was 84% sensitive (95% CI, 80-89) and 40% specific (95% CI, 34-46) for diagnosing cervical infection. Positive predictive values for diagnosing cervical infection with use of the algorithms ranged from 42% to 43%, and negative predictive values ranged from 78% to 81%. The sensitivity of the algorithms for diagnosing trichomoniasis ranged from 85% to 88%. To treat as many infected women as possible, the most sensitive algorithm was selected for routine use in Jamaican STD clinics.

STD/HIV control in Malawi and the search for affordable and effective urethritis therapy: a first field evaluation.

OBJECTIVES: To evaluate gonococcal (GU) and nongonococcal urethritis (NGU), chlamydia antigen, and serostatus for syphilis and human immunodeficiency virus (HIV) among males attending a Malawian STD clinic with complaints of urethral discharge and/or dysuria. To collect demographic and behavioural data and to determine the effectiveness of five treatments for urethritis. METHODS: Urethritis was diagnosed using microscopy and culture for Neisseria gonorrhoeae. Sera were screened with rapid plasma reagin (RPR) and if reactive, with microhaemagglutination for Treponema pallidum (MHA-TP). HIV antibodies and chlamydia antigen were detected using enzyme immunoassay. Patients were randomised for treatment, cure was assessed 8-10 days later. RESULTS: At enrolment, GU was diagnosed in 415 (80.3%) and NGU in 59 (11.2%) of 517 males. Chlamydia antigen was found in 26 (5.2%) of 497 specimens tested. Syphilis seropositivity rate (RPR and MHA-TP reactive) was 10.7%. Overall HIV seroprevalence was 44.2%; 71.7% of men with reactive syphilis serology were HIV(+) compared with 40.9% of syphilis seronegatives (OR: 3.6, p < 0.001). Trimethoprim 320 mg/sulphamethoxazole 1600 mg by mouth for 2 days (TMPSMX), or the combination of amoxicillin 3 gm, probenicid 1 gm, and clavulanate 125 mg by mouth once (APC), failed to cure gonorrhoea effectively. Amoxicillin 3 gm, probenicid 1 gm, and clavulanate 125 mg, by mouth once with doxycycline 100 mg BID for 7 days (APC-D), gentamicin 240 mg IM once (GENT), ciprofloxacin 250 mg by mouth once (CIPRO) cured 92.9% to 95% of gonorrhoea. APC-D treatment did not generate less NGU at follow-up. HIV serostatus did not affect cure of urethritis. CONCLUSION: All patients presenting with urethritis should be treated syndromically using a simple algorithm and screened for syphilis seroreactivity for appropriate treatment and counselling.

Sexually transmitted diseases and human immunodeficiency virus control in Malawi: a field study of genital ulcer disease.

Men with genital ulcer disease (GUD) attending a clinic in Malawi were evaluated and treated with one of five drug regimens. Haemophilus ducreyi was isolated from 204 (26.2%) of 778 patients. Of 677 men, 198 (29.2%) had treponemes detected in ulcer material by direct immunofluorescence or had rapid plasma reagin reactivity of > or = 1:8. Human immunodeficiency virus type 1 (HIV-1) seroprevalence was 58.9% overall and 75.8% among patients reporting a history of GUD (P < .001). By logistic regression analysis, HIV-1 seropositivity was shown to impair ulcer healing (P = .003). Treatment failure rates for culture-proven chancroid were 19% for trimethoprim-sulfamethoxazole, 12.9% and 7.4%, respectively, for low- and high-dose erythromycin regimens, and 8.3% and 0, respectively, for low- and high-dose ciprofloxacin regimens. Herpes antigen was detected by EIA in 6 (23.1%) of 26 nonhealing ulcers. In Malawi, GUD should be managed as a syndrome to assure treatment of both syphilis and chancroid.

Features of urethritis in a cohort of male soldiers.

Of 400 cases of urethritis in male soldiers enrolled in a behavioral intervention project, the etiology of 69% was defined at study enrollment, as well as the etiology of 72% of 25 repeated episodes involving 21 men during the first 78 days of active follow-up (5% of the cohort). Chlamydia trachomatis (36%), Neisseria gonorrhoeae (34%), and Ureaplasma urealyticum (19%) were the most common causes of infection identified at enrollment and during subsequent visits (44%, 28%, and 12%, respectively). By univariate analysis, patients with repeated infection ("repeaters") were significantly more likely to report a history of sexually transmitted disease (STD; relative risk [RR], 3) and sex with sex workers (RR, 4) than were nonrepeaters. By multivariate analysis, only STD history was significant (RR, 2.8). Characteristics of repeaters in this cohort suggest that specific patterns may be used to establish screening "profiles" of potential repeaters, by which such individuals might be targeted for aggressive intervention at the time of the initial diagnosis.

Trends of gonorrhea and chlamydial infection during 1985-1996 among active-duty soldiers at a United States Army installation.

High rates of sexually transmitted diseases (STDs) have been reported in military populations. However, it remains uncertain whether the incidence of STDs is higher among military personnel than in the civilian population. The annual incidence of gonorrhea and chlamydia from 1985 through 1996 at Fort Bragg, North Carolina, was determined by use of a clinic database and demographic information for the entire installation. A direct standardization for age, sex, and race/ethnicity was performed, and the adjusted annual rates among active duty soldiers were compared with rates among men and women in North Carolina and the United States. Results showed that the adjusted incidence of gonorrhea and chlamydia among Fort Bragg soldiers remained higher overall than comparable state and national rates during the period of analyses. The 1996 adjusted chlamydia rates for male and female active duty soldiers were 3-fold to 6-fold higher than rates for males and females in North Carolina and in the United States as a whole. STDs continue to lead to significant morbidity in this representative military population.

Validation of the WHO diagnostic algorithm and development of an alternative scoring system for the management of women presenting with vaginal discharge in Malawi.

OBJECTIVE: To evaluate the performance of the WHO algorithm for the detection of cervical infection in women presenting with vaginal discharge and modify the risk assessment score for optimum effectiveness in Malawi. METHODS: 550 consecutive women presenting with non-ulcerative genitourinary complaints were interviewed and examined. Cervical infection was defined as presence of Neisseria gonorrhoeae on culture and/or Chlamydia trachomatis by EIA. Other laboratory investigations included wet mount microscopy, serology for syphilis and HIV, LED testing of cervical and vaginal secretions, and pH testing of vaginal fluid. Sensitivity, specificity, and positive predictive values (PPV) of different algorithms were determined in the analysis. RESULTS: Cervical infection was identified in 19.5% of women (17.1% gonorrhoea, 3.7% chlamydial infection). The sensitivity/specificity/PPV of the WHO risk assessment were 43%/73%/28%, respectively by history and 62%/61%/27% with the addition of speculum examination. Using Malawi results to modify the risk assessment improved the performance to 61%/68%/31% respectively by history alone, which increased to 73%/64%/33% with bimanual examination and 72%/56%/29% with speculum examination. CONCLUSION: The sensitivity of the WHO risk assessment is low for the detection of cervical infection in Malawi. Although the Malawi risk assessment performed somewhat better on history alone, this study identified external and bimanual examination variables that improved the diagnostic performance of the algorithm in settings where speculum examination is not possible. Although the PPVs of the algorithms are low, country specific risk assessments can provide a framework for management until simple, affordable diagnostic tests for the definitive diagnosis of cervical infection are available.

Association of CD4 cell depletion and elevated blood and seminal plasma human immunodeficiency virus type 1 (HIV-1) RNA concentrations with genital ulcer disease in HIV-1-infected men in Malawi.

CD4 cell counts and blood plasma and seminal plasma human immunodeficiency virus type 1 (HIV-1) concentrations were compared in HIV-1 RNA-seropositive men with urethritis and with or without genital ulcer disease (GUD). GUD was associated with lower CD4 cell counts (median, 258 vs. 348/microL) and increased blood plasma HIV-1 RNA (median, 240 x 10[3] vs. 79.4 x 10[3] copies/mL). Men with nongonococcal urethritis and GUD shed significantly greater quantities of HIV-1 in semen (median, 195 x 10[3] vs. 4.0 x 10[3] copies/mL) than men with nongonococcal urethritis without GUD. These levels decreased approximately 4-fold following antibiotic therapy. The results indicate an association between GUD and increased blood HIV-1 RNA levels. Increased HIV-1 in semen was demonstrated in some men with GUD; such an increase could lead to increased transmission, thus complicating interpretation of the role of the genital ulcer itself in the infectiousness of HIV. Reasons for increased HIV RNA in semen in men with GUD remain to be determined.

Clinical utility of measuring white blood cells on vaginal wet mount and endocervical gram stain for the prediction of chlamydial and gonococcal infections.

BACKGROUND: White blood cells on endocervical Gram stain and vaginal wet mount are frequently used to predict chlamydial and gonococcal infections. Previous studies provide conflicting evidence for the clinical utility of these tests. GOAL: To evaluate the clinical utility of measuring white blood cells on vaginal wet mount and endocervical Gram stain for the prediction of chlamydial infection and gonorrhea. STUDY DESIGN: Women undergoing pelvic examinations at 10 county health department family planning and sexually transmitted disease clinics were tested for chlamydial infection by ligase chain reaction assay (n = 4550) and for gonorrhea by culture (n = 4402). Vaginal wet mount and endocervical Gram stains were performed in county laboratories at the time of examination. RESULTS: The prevalences of chlamydial infection and gonorrhea were 8.8% and 3.2%, respectively. For detection of chlamydial or gonococcal infection, the likelihood ratio was 2.85 (95% CI, 2.10-3.87) for > 30 white blood cells on vaginal wet mount and 2.91 (95% CI, 2.07-4.09) for > 30 white blood cells on endocervical Gram stain. Similar results were seen for individual diagnoses either of chlamydial infection or of gonorrhea. CONCLUSION: Vaginal wet mount and endocervical Gram stain white blood cells are useful for the presumptive diagnosis of chlamydial infection or gonorrhea only in settings with a relatively high prevalence of infection or when other predictors can increase the likelihood of infection.

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.

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.

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.