Motivations to participate in a Phase I/II HIV vaccine trial: A descriptive study from Dar es Salaam, Tanzania.

BACKGROUND: The search for an efficacious HIV vaccine is a global priority. To date only one HIV vaccine trial (RV144) has shown modest efficacy in a phase III trial. With existing different HIV-1 subtypes and frequent mutations, multiple trials are needed from different geographical sites particularly in sub-Saharan Africa where most HIV infections occur. Thus, motivations to participate in HIV vaccine trials among Tanzanians need to be assessed. This paper describes the motives of Police Officers who showed great interest to volunteer in HIVIS-03 in Dar es Salaam, Tanzania. METHODS: A descriptive cross-sectional study was conducted among Police Officers who showed interest to participate in the HIVIS-03, a phase I/II HIV vaccine trial in Dar es Salaam. Prior to detailed training sessions about HIV vaccine trials, the potential participants narrated their individual motives to participate in the trial on a piece of paper. Descriptive analysis using content approach and frequency distributions were performed. RESULTS: Of the 265 respondents, 242 (91.3%) provided their socio-demographic characteristics as well as reasons that would make them take part in the proposed trial. Majority, (39.7%), cited altruism as the main motive. Women were more likely to volunteer due to altruism compared to men (P < 0.01). Researchers' explanations about HIV/AIDS vaccine studies motivated 15.3%. More men (19.6%) than women (1.7%) were motivated to volunteer due to researchers' explanations (P < 0.001). Also, compared to other groups, those unmarried and educated up to secondary level of education were motivated to volunteer due to researchers' explanation (P < 0.05). Other reasons were: desire to become a role model (18.6%); to get knowledge for educating others (14.0%); to cooperate with researchers in developing an HIV vaccine (9.5%); to get protection against HIV infection (7.0%), and severity of the disease within families (6.2%). These results were supported by testimonies from both men and women. CONCLUSIONS: Participation in an HIV vaccine trial in a Tanzanian context is likely to be influenced by altruism and comprehensive education about the trial. Gender differences, marital status and education level need to be considered to enhance participation in future HIV vaccine trials.

Enhancing adherence to antiretroviral therapy at the HIV clinic in resource constrained countries; the Tanzanian experience.

OBJECTIVE: To evaluate various strategies aimed at improving adherence to antiretroviral therapy (ART). METHODS: Patients initiated on ART at Muhimbili National Hospital HIV clinic were randomly assigned to either regular adherence counseling, regular counseling plus a calendar, or regular counseling and a treatment assistant. Patients were seen monthly; during these meetings self-reported adherence to treatment was recorded. Disease progression was monitored clinically and immunologically. RESULTS: Of the 621 patients randomized, 312 received regular counseling only, 242 regular counseling and calendars, while 67 had treatment assistants in addition to regular counseling. The mean (SD) follow-up time was 14.5 (4.6) months. During follow-up 20 (3.2%) patients died, and 102 (16.4%) were lost to follow-up; this was similar in all groups. In 94.8% of all visits, patients reported to have adhered to treatment. In only 39 (0.7%) visits did patients report a < or = 95% adherence. There were no differences in adherence (P = 0.573) or differences in CD4 count and weight changes over time in the interventions. CONCLUSIONS: Good adherence to ART is possible in resource constrained countries. Persistent adherence counseling in clinic settings by itself may be effective in improving adherence to ART.

AIDS: an international perspective.

The acquired immunodeficiency syndrome (AIDS) and infection with the human immunodeficiency virus type 1 (HIV-1) constitute a worldwide public health problem. Whereas in Europe and in most of the Americas transmission of HIV-1 has occurred predominantly among homosexual men and intravenous drug abusers, in Africa a distinct epidemiologic pattern has emerged that indicates that HIV-1 infection is mainly heterosexually acquired. Heterosexual transmission appears to be increasing in some parts of Latin America and the Caribbean, and possibly in the United States. In addition to HIV-1, at least one other human retrovirus, namely HIV-2, has been implicated as a cause of AIDS in Africa and Europe. Factors that influence heterosexual transmission of HIV-1 include genital ulcerations, early or late stages of HIV-1 infection in the index case, and possibly oral contraception and immune activation. The rate of perinatal transmission is enhanced when the mother's illness is more advanced. AIDS and HIV-1 infection may have a significant impact not only on public health, but also on the demography and socioeconomic conditions of some developing countries. Programs for the prevention and control of AIDS should be an immediate priority in all countries.

Risky sexual practices among youth attending a sexually transmitted infection clinic in Dar es Salaam, Tanzania.

BACKGROUND: Youth have been reported to be at a higher risk of acquiring STIs with significant adverse health and social consequences. Knowledge on the prevailing risky practices is an essential tool to guide preventive strategies. METHODS: Youth aged between 18 and 25 years attending an STI clinic were recruited. Social, sexual and demographic characteristics were elicited using a structured standard questionnaire. Blood samples were tested for syphilis and HIV infections. Urethral, high vaginal and cervical swabs were screened for common STI agents. RESULTS: A total of 304 youth were studied with mean age of 21.5 and 20.3 years for males and females respectively. 63.5% of youth were seeking STI care. The mean age of coitache was 16.4 and 16.2 years for males and females respectively. The first sexual partner was significantly older in females compared to male youth (23.0 vs 16.8 years) (p < 0.01). 93.2% of male youth reported more than one sexual lifetime partner compared to 63.0% of the females. Only 50% of males compared to 43% of females had ever used a condom and fewer than 8.3% of female youth used other contraceptive methods. 27.1% of pregnancies were unplanned and 60% of abortions were induced. 42.0% of female youth had received gifts/money for sexual favours. The HIV prevalence was 15.3% and 7.5% for females and males respectively. The prevalence of other STIs was relatively low. Among male youth, use of alcohol or illicit drugs was associated with increased risk of HIV infection. However, the age of sexual initiation, number of sexual partners or the age of the first sexual partner were not associated with increased risk of being HIV infected. CONCLUSION: Most female youth seen at the STI clinic had their first sexual intercourse with older males. Youth were engaging in high risk unprotected sexual practices which were predisposing them to STIs and unplanned pregnancies. There is a great need to establish more youth-friendly reproductive health clinics, encourage consistent and correct use of condoms, delay in sexual debut and avoid older sexual partners in females.

Burden of diseases in poor resource countries: meeting the challenges of combating HIV/AIDS, tuberculosis and malaria.

Poverty, ill health and ignorance are closely interlinked and each is a determinant of the other. HIV/AIDS, malaria and tuberculosis are by far the commonest causes of ill-health and death in the poorest countries of the world which happen to be in the tropics and temperate countries in Africa, Asia and South America. Morbidity and mortality from these three diseases have a major socio-economic impact on individuals, communities and nations, due to the vicious cycle of poverty, ill health and ignorance. In Tanzania morbidity due to HIV/AIDS, tuberculosis and malaria leads to irrecoverable losses in productivity, inadequately trained workforce due to absence from training by the sick, heavy health care budgets to treat these otherwise preventable diseases, less competitive economy, higher labour force turnovers and unstable national budgets. If not controlled continuing rise in incidence of HIV/AIDS, malaria and TB may threaten the survival of small enterprises and ability to attract foreign investments leading to a rise in unemployment. Thus, investments in the improvement of health including HIV/AIDS, malaria and TB if done well will bring substantial benefits for the national economy including an increase in productivity. In this paper a review of the impact of HIV/AIDS, TB and malaria in Tanzania is done with an attempt to propose how research can contribute to improved efforts towards more effective prevention and control efforts. The need for multidisplinary research efforts in addressing the three disease conditions is proposed.

Laboratory diagnosis of HIV-1 and HIV-2 in Africa.

PIP: It is well accepted that HIV is the cause of AIDS and that the virus is distributed widely throughout the world. Being able to diagnose infection with HIV through laboratory tests has done much to facilitate the early recognition of the severity and extent of the AIDS pandemic. Many laboratory techniques exist to detect infection with HIV-1 and HIV-2. In recent years, however, African countries have found it difficult to implement such diagnostic tests because the tests have been either ill-suited or too expensive for the developing country setting. This paper describes many of the HIV laboratory diagnostic techniques currently used in both diagnostic and research settings. The review of techniques is, however, neither all-inclusive nor globally applicable, but intended to be simply a view of available techniques from the African perspective. The opening general section on the detection of HIV-1 and HIV-2 is followed by discussion of screening tests, rapid tests, and confirmatory tests to detect HIV antibodies. Techniques to detect virus include viral isolation, the detection of viral antigen, and PCR. HIV testing algorithms are discussed. The authors stress in closing the importance of the effective laboratory diagnosis of HIV in the prevention and control of HIV/AIDS. Laboratory personnel must be trained, cost-effective laboratory techniques made available for the African setting, and test systems chosen which are best adapted to the prevailing epidemiologic, socioeconomic, and cultural contexts. These latter systems often will differ from the types of diagnostic tests and testing algorithms used in more developed countries.^ieng

Evaluation of commercial enzyme immunoassays for anti-HIV-1 using East African sera.

Sera from 622 blood donors collected in 1986 and 1987 in Tanzania were screened for antibodies to HIV-1 by seven different commercial enzyme-linked immunosorbent assay (ELISA) kits. All ELISA-positive sera were tested by Western blot analysis and many of them also by radioimmunoprecipitation assay (RIPA). Sixty-seven sera were confirmed positive. Eight sera, which were repeatedly positive on only one or two of the ELISA kits and showed weak, doubtful reactions on Western blot and RIPA, were considered indeterminate and were not included in the calculations of sensitivity and specificity of the various ELISA kits. The sensitivity of the ELISAs was as follows: Organon Vironostika: low cut-off 74.6%; Organon Vironostika: high cut-off 62.7%; Du Pont: 85.1%; Pasteur: 78.7%; Abbott: 80.8%; Abbott recombinant: 94.0%; Wellcozyme: 82.1%; Wellcozyme monoclonal: 98.5%. The specificity was as follows: Organon Vironostika: low and high cut-off 100%; Du Pont: 94.7%; Pasteur: 99.3%; Abbott: 100%; Abbott recombinant 98.7%; Wellcozyme: 99.8%; Wellcozyme monoclonal: 98.5%. In conclusion, the two new generation kits tested, Wellcozyme monoclonal and Abbott recombinant, had the highest sensitivity whereas the sensitivity of the first-generation tests was unexpectedly low.

Prevalence of HIV-1 infection in the Kagera region of Tanzania: a population-based study.

A population-based survey was carried out in the Kagera region of the United Republic of Tanzania in 1987 to determine the magnitude of HIV-1 infection and to study associated risk factors. The region was divided into one urban and three rural zones. A multistage cluster sampling technique was adopted. Antibodies to HIV-1 were determined by enzyme-linked immunosorbent assay and confirmed by Western blot analysis. A total of 2,475 adults (aged 15-54 years) and 1,961 children (aged 0-14 years) was studied. The overall prevalence of HIV-1 infection among adults was 9.6%, with a higher prevalence in the urban zone (24.2%) than in the three rural zones (10.0, 4.5 and 0.4%, respectively). The corresponding figures for children were 1.3% overall: 3.9% in the urban area and for the rural areas 1.2, 0.8 and 0.0%, respectively. The age-specific seroprevalence for adults was highest in the age group 25-34 years. The age-standardized sex-specific prevalence was higher among women than men in the urban zone, while it was the same in the rural zones. Change of sexual partners among adults was associated with an increased risk of HIV-1 seropositivity. Travelling outside the region but within the country was also found to be associated with increased risk of HIV-1 infection but only in the rural population.

High HIV seroprevalence and increased HIV-associated mortality among hospitalized patients with deep bacterial infections in Dar es Salaam, Tanzania.

OBJECTIVES: To correlate deep bacterial infections with HIV infection and evaluate the influence of HIV on clinical picture and outcome in patients with meningitis, pneumonia or pyomyositis. DESIGN: Case-control comparison of HIV seroprevalence between patients and an age- and sex-matched control group in a prospective cross-sectional study of hospitalized patients. PARTICIPANTS: One hundred and sixty-five patients admitted to hospital with either purulent meningitis, pneumonia or pyomyositis and 165 age- and sex-matched controls from orthopaedic/trauma wards. SETTING: University Hospital, Dar es Salaam, Tanzania. OUTCOME MEASURES: Differences in HIV seroprevalence and mortality. RESULTS: Of 78 patients with purulent meningitis, 19 (24%) were HIV-seropositive, compared with 13 (17%) in the control group (P = 0.345). Of 36 patients with meningitis seen before a meningococcal epidemic affected Dar es Salaam, there was a statistically significant association with HIV infection (P = 0.013). Ten out of 19 (53%) HIV-seropositives died, compared with nine out of 59 (15%) seronegatives (P = 0.028). Of patients with pneumococcal meningitis, five out of six (83%) seropositives died, compared with two out of 12 (17%) seronegatives (P = 0.013). Fifteen out of 45 (33%) patients with pneumonia were HIV-seropositive, compared with four (9%) in the control group (P = 0.001). There was no difference in mortality between seropositive and seronegative patients with pneumonia. HIV seroprevalence was 62% among 42 patients with pyomyositis and 12% among 42 controls (P less than 0.0001). Eighteen out of 25 (72%) seropositive patients with pyomyositis fulfilled the World Health Organization (WHO) clinical case definition for AIDS. Response to recommended antibiotic treatment was satisfactory among patients with pneumonia and pyomyositis. CONCLUSIONS: These results show a strong association between pyomyositis, pneumonia and HIV infection. They also indicate an increased mortality associated with HIV infection in patients with pyogenic meningitis, especially pneumococcal meningitis. Pyomyositis should be considered an indicator of stage III HIV disease in the proposed WHO clinical staging system.

PIP: This study sought to correlate deep bacterial infection with HIV infection and evaluate the influence of HIV on clinical practice and outcome in patients with meningitis, pneumonia, or pyomyositis. At University Hospital, Dar es Salaam, Tanzania, 165 patients were admitted to the hospital with purulent meningitis, pneumonia, or pyomyositis and were evaluated in a prospective, cross-sectional study along with 165 age- and sex-matched controls from orthopedic/trauma wards to determine HIV seroprevalence. Of the 78 patients with purulent meningitis, 19 (24%) were HIV-seropositive, as compared with 13 (17%) in the control group (p=0.345). Of 36 patients with meningitis seen before a meningococcal epidemic affected Dar es Salaam, there was a statistically significant association with HIV infection (p=0.013). 10 of 19 (53%) HIV-seropositives died, compared with 9 of 59 (15%) seronegatives (p=0.028). Of patients with pneumococcal meningitis, 5 of 6 (83%) seropositives died, compared with 2 of 12 (17%) seronegatives (p=0.013). 15 of 45 (33%) patients with pneumonia were HIV-seropositive compared with 4 (9%) in the control group (p=0.001). There was no difference in mortality between seropositive and seronegative patients with pneumonia. HIV seroprevalence was 62% among 42 patients with pyomyositis and 12% among 42 controls (p0.0001). 18 of 25 (72%) seropositive patients with pyomyositis fulfilled the WHO clinical case definition of AIDS. Response to recommended antibiotic treatment was satisfactory among patients with pneumonia and pyomyositis. These results show a strong association between pyomyositis, pneumonia, and HIV infection. They also indicate an increased mortality associated with HIV infection in those patients with pyogenic meningitis, especially pneumococcal meningitis. Pyomyositis should be considered an indicator of stage III HIV disease in the proposed WHO clinical staging system.

Prevalence of HIV infection in healthy subjects and groups of patients in Tanzania.

During 1986 sera from 2508 individuals representing various groups of healthy subjects and patients in Dar es Salaam (the capital city of Tanzania), Bukoba (the capital of Kagera region in the northwest corner of Tanzania), Arusha (in the northeast of Tanzania) and Mbeya (in the southwest of Tanzania) were screened for antibodies to HIV by enzyme-linked immunosorbent assay (ELISA). All ELISA-positive sera were also tested by Western blot analysis. In Dar es Salaam HIV antibodies were demonstrated in 3.6% of 192 pregnant women, 5.2% of 784 blood donors, 29.0% of 224 barmaids, 8.0% of 50 male bar workers, 9.25% of 400 male and 12.2% of 90 female patients attending a clinic for sexually transmitted diseases (STDs), 85.7% of 35 patients with herpes zoster and in 97.6% of 84 patients clinically suspected of AIDS. Among the barmaids the seropositivity rate was higher in younger women (45%) than in middle-aged women (11%). Only three (4.6%) out of 65 HIV-seropositive barmaids had HIV-related symptoms. The prevalence of HIV seropositivity among healthy low-risk subjects was highest in Bukoba, namely 16% of 100 pregnant women and 13.9% of 36 blood donors, while in Arusha only one (0.7%) of the 144 pregnant women and none of 41 bar workers, none of 42 blood donors and none of 61 patients with STD were positive. In Mbeya, 3.4% of 118 pregnant women and 11.8% of 34 men with STD were seropositive. Thus the prevalence of HIV infection differs considerably in various population groups and in various parts of Tanzania.

Selection of primers of optimal sensitivity for the detection of HIV-1 from Africa and Europe by polymerase chain reaction.

In order to facilitate the detection of integrated HIV-1 proviral DNA from African as well as European patients, four new primer pairs for use in the polymerase chain reaction (PCR), localised in the gag, pol, vif and env genes of HIV-1, were constructed. The primer pairs were compared to all accessible HIV-1 sequences from African and European isolates and to some of the earlier published and most commonly used primer pairs. HIV-1 DNA was detected in blood drawn from 13 out of 13 individuals infected in Africa, in three out of three Tanzanian HIV-1 isolates and in three out of three asymptomatic Swedes infected in Europe. The new selection of primer pairs can be used as an alternative to enhance the detection of HIV-1 of different origins.

PIP: In order to facilitate the detection of integrated HIV-1 proviral DNA from African as well as European patients, 4 new primer pairs for use in the polymerase chain reaction (PCR), localized in the gag, pool, vif, and env genes of HIV-1, were constructed. The primer pairs were compared to all accessible HIV-1 sequences from African and European isolates and to some of the earlier published and most commonly used primer pairs. HIV-1 DNA was detected in blood drawn from 13 infected individuals in Africa, in 3 Tanzanian HIV-1 isolates, and in the 3 asymptomatic Swedes infected in Europe. The new selection of primer parts can be used as an alternative to enhance the detection of HIV-1 of different origins.

High proportion of unrelated HIV-1 intersubtype recombinants in the Mbeya region of southwest Tanzania.

BACKGROUND: In Mbeya, a rural region of southwest Tanzania, HIV-1 subtypes A, C and D have been co-circulating since the early 1990s. OBJECTIVE: To define to what extent the co-existence of subtypes has led to recombinant HIV-1 strains and whether there is evidence for epidemic spread of any circulating recombinant form. METHODS: Nine HIV-1-seropositive young adults from Mbeya Town with no evident high-risk behaviour contributed peripheral blood mononuclear cells for this study. Nine virtually full-length-genome-sequences were amplified from this DNA and phylogenetically analysed. RESULTS: Out of the nine samples, two were subtype A (22%), two were subtype C (22%) and five were recombinants (56%): four A/C recombinants and one C/D recombinant. None of the recombinants were related to each other; all of them had different mosaic structures. Most of the genome in the recombinants was subtype C. CONCLUSION: A high proportion of unrelated intersubtype recombinants, none of them apparently spreading in the population, may be present in southwest Tanzania.

The prevalence and incidence of HIV-1 infection and syphilis in a cohort of police officers in Dar es Salaam, Tanzania: a potential population for HIV vaccine trials.

OBJECTIVES: To assess the suitability of a cohort of police officers in Dar es Salaam for HIV vaccine trials by determining the prevalence and incidence of HIV-1 infection, active syphilis and their associated factors. DESIGN AND SETTING: An open cohort study of police officers in Dar es Salaam, Tanzania. METHODS: Recruitment of police officers began in 1994. A standardized questionnaire was completed at enrolment and subsequent visits. HIV antibodies were determined using two consecutive enzyme-linked immunosorbent assays. Samples repeatedly discordant on the two tests were tested by a Western blot assay. Treponema pallidum antibodies were first determined by Venereal Disease Research Laboratory (VDRL) test and reactive sera were confirmed by Treponema pallidum hemagglutination test. RESULTS: At the end of 1996 a total of 2850 police officers had been recruited of whom 2733 (96%) consented to be tested for HIV. The overall HIV-1 seroprevalence at recruitment was 13.8% (378 of 2733). Females had a significantly higher HIV-1 seroprevalence, 18.0% (55 of 306), as compared to males, 13.3% (323 of 2427), P< 0.05. From a total of 2215 married police officers, 585 (26.4%) responded to a question on extramarital sex within the previous 3 months of whom 36.2% (212 of 585) admitted to have had at least one extramarital sexual intercourse. Condoms were not used during these encounters by 178 of 212 (84.0%). As of 31st December 1998, among the 1524 males observed for 2553 person-years (PYAR), 50 had seroconverted and among 200 females observed for 357 PYAR, eight had seroconverted. The overall crude HIV-1 incidence was thus 19.9/1000 PYAR; 19.6 and 22.4/1000 PYAR for males and females, respectively. The overall prevalence and incidence of active syphilis were 3.1% (88 of 2850) and 8.6/1000 PYAR (26 of 3149), respectively. Males had a higher prevalence of active syphilis, 84 of 2525 (3.3%) than females, five of 325 (1.5%), P = 0.09. CONCLUSIONS: There was high risk sexual practice including low condom use in this cohort of police officers. The incidence and prevalence of HIV infection were high. Police officers in Dar es Salaam are therefore a potential population group for HIV vaccine evaluation.

Alternative confirmatory strategies in HIV-1 antibody testing.

Alternatives to confirmation of human immunodeficiency virus (HIV)-1 seropositivity by Western blot analysis were evaluated retrospectively using combinations of six anti-HIV-1 screening assays, including four enzyme-linked immunosorbent assays (ELISA) and two simple tests (a rapid dot immunoassay and an agglutination assay), according to an algorithm where sera are first screened by one assay and those repeatedly reactive on this assay are tested repeatedly by a second assay. Two panels of sera collected in Dar es Salaam, Tanzania, were used. Panel 1 was composed of 1,465 consecutive blood donor sera of which 99 (6.8%) were confirmed HIV-1 antibody positive, and panel 2 was composed of sera from 396 consecutively admitted patients at two medical wards of which 116 (29.3%) were confirmed HIV-1 antibody positive. Sera reactive on any of the six screening assays were also tested by a confirmatory Western blot assay. The sensitivity of the assays at the initial valid testing were as follows: Abbott 99.5%, Behring 99.5%, Organon 97.7%, Wellcozyme 100%, HIV CHEK-1 95.8%, and Serodia 95.8%. After repeat testing of sera that initially gave false-negative results all assays showed 100% sensitivity except HIV CHEK-1 (98.6%). The specificities after repeat testing were between 99.6 and 99.9% for all assays except for the Behring ELISA (98.1%). Several combinations of screening assays were found to give the same diagnostic accuracy as the screening assay followed by Western blot analysis. We conclude that an alternative confirmatory strategy can be fully satisfactory for some testing purposes.

Prevalence of HIV-1 infection and symptomatology of AIDS in severely malnourished children in Dar Es Salaam, Tanzania.

In Dar es Salaam, Tanzania, 200 children with severe malnutrition and controls matched for age, sex, and area of residence were screened for serological evidence of infection with the human immunodeficiency virus type 1 (HIV-1) over 5 months in 1988. The prevalence of HIV-1 antibodies in the malnourished group was 25.5% (51 of 200) compared with 1.5% (three of 200) in the controls. The seroprevalence rate was equally high in malnourished children above the age of 18 months (26 of 102; 25.5%), as in those below this age (25 of 98; 25.5%). The prevalence rate was higher in children with marasmus (38.2%) as compared to children with marasmic-kwashiorkor (12.3%) or kwashiorkor (12.2%). The prevalence of clinical features known to be associated with AIDS was higher in the HIV seropositive malnourished children as compared to the seronegative children. The modified World Health Organization clinical case definition of AIDS in children was also evaluated and found to have a low sensitivity and positive predictive value (62.8 and 57.1%, respectively) but a fairly high specificity (83.9%). It is recommended to routinely rule out HIV infection in malnourished children, especially those with marasmus.

A cross-sectional study of a program for HIV infection control among public house workers.

We report results of a cross-sectional study of a program for human immunodeficiency virus (HIV) infection control among public house workers in Dar es Salaam. Forty percent of the 605 workers sampled had been part of this program, which included behavioral counseling and provision of condoms, for 1 year. The remaining 60% were new recruits. Program participation was associated with both enhanced condom use (p less than 0.001) and behavioral modification (p less than 0.001). Females, and specifically barmaids, were more likely to be condom users but were less likely to have changed their behavior in other respects. Seropositivity to both HIV and Treponema pallidum tended to be higher among females, especially the barmaids. Since barmaids and waitresses in public houses in Dar es Salaam often engage in prostitution, it is felt that to effect a reduction of numbers of their sexual partners, there is a need to address the social and economic factors underlying high-risk sexual behavior.

Lymphocyte subset enumeration in HIV seronegative and HIV-1 seropositive adults in Dar es Salaam, Tanzania: determination of reference values in males and females and comparison of two flow cytometric methods.

The level of CD4(+) T-lymphocytes represents a useful marker with which to monitor the progression of HIV infection. Sex and geographical differences in the reference values of lymphocyte subsets have been reported. We have compared two flow cytometric methods (MultiSET and SimulSET) for the quantification of lymphocyte subsets using whole blood from 92 HIV seropositive and 241 seronegative adults, and determined the reference values of lymphocyte subsets in HIV seronegative Tanzanian subjects. In seronegative Tanzanian subjects, the percentages of CD3(+) and CD4(+) T-lymphocytes and the CD4(+):CD8(+) T-lymphocyte ratios were lower while the percentage of natural killer cells was higher compared to the levels of the corresponding parameters reported for Europeans. Seronegative Tanzanian females had significantly higher levels of CD3(+) and CD4(+) T-lymphocytes and CD4(+):CD8(+) T-lymphocyte ratios compared to seronegative males. The correlation coefficients of CD3(+), CD4(+) and CD8(+) T lymphocyte counts and percentages obtained by the two flow cytometric methods were high. The median values of the number of CD4(+) T-lymphocytes obtained by the two methods were not significantly different. In conclusion, determination of the reference values of lymphocyte subsets in HIV seronegative Tanzanian adults showed significant sex differences and differences in percentage values compared to those reported in certain other geographical areas. There was acceptable agreement in the levels of CD4(+) T-lymphocyte values obtained by the two flow cytometric methods.

Evaluation of the FACScount, TRAx CD4 and Dynabeads methods for CD4 lymphocyte determination.

A study to evaluate the performance of the FACScount, TRAx CD4 and Dynabeads methods for the determination of CD4+ T lymphocyte subset levels was conducted in Tanzania as part of a World Health Organization (WHO) collaborative multicenter field evaluation of alternative methodologies for the enumeration of CD4+ T lymphocytes. The objective was to compare the performance of these alternative methods in a developing country setting, against that of flow cytometry as the reference standard. T lymphocyte subset levels were determined in 91 HIV seronegative and 98 HIV-1 seropositive adults using the three alternative methods. CD4+ and CD8+ T lymphocyte counts were determined by all methods except for TRAx CD4 enzyme linked immunosorbent assay (ELISA) which measures CD4+ T lymphocyte levels only. Linear regression analysis was done to correlate the counts obtained by the alternative methods to those obtained by flow cytometry. The overall correlation coefficients of FACScount and Dynabeads CD4+ and CD8+ T lymphocyte counts with those of flow cytometry were high (r > 0.9). A lower correlation (r = 0.631) was obtained when TRAx CD4+ ELISA counts were compared to those of the reference method. These results show that two of these alternative methodologies are suitable for the determination of CD4+ and CD8+ T lymphocyte counts with the use of African blood samples. Since the methods are simpler and cheaper than flow cytometry, they provide an alternative option for the enumeration of T lymphocyte subsets in laboratories with limited facilities.

Affordable CD4+ T cell counts by flow cytometry. II. The use of fixed whole blood in resource-poor settings.

We tested the feasibility and precision of affordable CD4+ T cell counting in resource-poor settings using a recently standardised fixative, TransFix in whole blood (WB) by flow cytometry (FCM). The precision of the assays was established under optimal conditions for single-platform FCM such as the volumetric CytoronAbsolute and the bead-based FACSCan. Fresh WB samples from HIV-seropositive and seronegative patients were tested in Tanzania and South Africa, fixed and sent to the UK for reanalysis 7 days later. Correlation, bias and limits of agreements were analysed by linear regression and the Bland-Altman test. Absolute CD4+ T cell counts remained stable for at least 10 days when TransFix was added to WB in 1:10 dilution at 20-25 degrees C, and for 7 days when added in 1:10 or 1:5 dilution to samples stored to mimic 'tropical' conditions at 37 degrees C. Higher temperatures such as 42 degrees C were tolerated for only short periods since the recovery had decreased to 63% by day 3. The reproducibility of lymphocyte subset analysis remained unchanged by TransFix with coefficient of variations <6% for all T cell subsets. Absolute CD4+ T cell counts and CD4+ T cell % values on fixed samples in the UK showed a high correlation with the results using fresh samples in Tanzania (r=0.993 and 0.969, respectively) and with the samples handled in Johannesburg (r=0.991 and 0.981) with minimal bias. Primary CD4 gating using only a single CD4 antibody also remained accurate in TransFixed samples (r=0.999). Thus, TransFix permits optimal fixation and transport of WB samples in the developing world for FCM to local regional laboratories and for quality assurance in international centres. When used together with inexpensive primary CD4 gating, TransFix will allow reliable and affordable CD4+ T cell counting by FCM in resource-poor settings.

vpu and env sequence variability of HIV-1 isolates from Tanzania.

PIP: The nucleotide sequences of an approximately 1400-base pair (bp) region spanning the vpu and env (V1-V3) genes of 9 HIV-1 isolates originating from Tanzania were determined. Peripheral blood lymphocyte (PBL) specimens were obtained in 1988 from urban patients with clinical signs of AIDS attending the Muhimbili Medical Center, Dar es Salaam, Tanzania. 9 samples (TZ005, TZ012, TZ016, TZ017, TZ023, TZ030, TZ053, TZ064, and TZ112) were randomly chosen for virus isolation by cocultivation with HIV-negative donor PBLs. Viral DNA sequences between the positions 5543 and 6956 were amplified by polymerase chain reaction (PCR), using 2 sets of primer pairs, subcloned into a Bluescript vector, and sequenced on both strands. Sequence analysis revealed vpu and env open reading frames (ORFs) for all clones, except 2 that had a missense mutation in vpu (TZ016) or env (TZ017). The vpu sequences showed a high degree of homology among all isolates, with TZ005, TZ016, and TZ030 having identical sequences. Phylogenetic tree analysis indicated that most of the isolates fell into the D subtype. The analysis of the deduced protein sequences of the V3 loop, which contains the principal neutralizing domain (PND), revealed an amino acid pattern closely related, but not identical, to known African HIV isolates. The GSGQ motif was found in 4 isolates (TZ005, TZ030, TZ053, and TZ080), and the GPGQ motif was found in 2 cases (TZ016 and TZ017). The V3 variability of the HIV isolates was greater than previously reported for Tanzanian viruses. Although AIDS viruses are believed to have originated from Africa, little is known about the sequence variability of African HIV-1 isolates, compared to the information available on Euro-American viruses. The variability of East African HIV-1 isolates are consistent with the view that these are rapidly changing viruses for which further variants are likely to be discovered.^ieng