Collaborative research to respond to the HIV epidemic: a case of Uganda (Makerere University)-Case Western Reserve University Research Collaboration 1988-2021.

Background: Collaborative research between institutions may not yield results to transform communities. Many research collaborations come to the end of their life time without achieving their originally set goals and with a dearth of community transformation to show for it. Objective: To delineate and highlight the achievements of the Uganda (Makerere University)-Case Western Reserve University Research Collaboration. Methods: We retrospectively compiled and reviewed the data on research, training and policy impact achievements of the Uganda (Makerere University)-Case Western Reserve University Research Collaboration over a period of 30 years of its existence. Results: Over the last 35 years, the Uganda (Makerere University)-Case Western Reserve University Research Collaboration trained a total of 104 Ugandans with Masters, PhDs and other varied graduate training programs. More than 70 large tuberculosis/TB+HIV studies were conducted with more than 360 manuscripts published including landmark local and global TB/HIV policy impact publications. Conclusion: The Uganda (Makerere University)-Case Western Reserve University Research Collaboration has in the past 35 years built the capacity of Ugandan and international students through conducting landmark research, training and mentoring and contributed to TB HIV management policy changes in Uganda.

Efavirenz Pharmacokinetics and Human Immunodeficiency Virus Type 1 (HIV-1) Viral Suppression Among Patients Receiving Tuberculosis Treatment Containing Daily High-Dose Rifapentine.

BACKGROUND: A 4-month regimen containing rifapentine and moxifloxacin has noninferior efficacy compared to the standard 6-month regimen for drug-sensitive tuberculosis. We evaluated the effect of regimens containing daily, high-dose rifapentine on efavirenz pharmacokinetics and viral suppression in patients with human immunodeficiency virus (HIV)-associated tuberculosis (TB). METHODS: In the context of a Phase 3 randomized controlled trial, HIV-positive individuals already virally suppressed on efavirenz--containing antiretroviral therapy (ART) (EFV1), or newly initiating efavirenz (EFV2) received TB treatment containing rifapentine (1200 mg), isoniazid, pyrazinamide, and either ethambutol or moxifloxacin. Mid-interval efavirenz concentrations were measured (a) during ART and TB cotreatment (Weeks 4, 8, 12, and 17, different by EFV group) and (b) when ART was taken alone (pre- or post-TB treatment, Weeks 0 and 22). Apparent oral clearance (CL/F) was estimated and compared. Target mid-interval efavirenz concentrations were > 1 mg/L. Co-treatment was considered acceptable if > 80% of participants had mid-interval efavirenz concentrations meeting this target. RESULTS: EFV1 and EFV2 included 70 and 41 evaluable participants, respectively. The geometric mean ratio comparing efavirenz CL/F with vs without TB drugs was 0.79 (90% confidence interval [CI] .72-.85) in EFV1 and 0.84 [90% CI .69-.97] in EFV2. The percent of participants with mid-interval efavirenz concentrations > 1mg/L in EFV1 at Weeks 0, 4, 8, and 17 was 96%, 96%, 88%, and 89%, respectively. In EFV2, at approximately 4 and 8 weeks post efavirenz initiation, the value was 98%. CONCLUSIONS: TB treatment containing high-dose daily rifapentine modestly decreased (rather than increased) efavirenz clearance and therapeutic targets were met supporting the use of efavirenz with these regimens, without dose adjustment. CLINICAL TRIALS REGISTRATION: NCT02410772.

Collaborative research to respond to the HIV epidemic: a case of Uganda (Makerere University)-Case Western Reserve University Research Collaboration 1988-2021

Background: Collaborative research between institutions may not yield results to transform communities. Many research collaborations come to the end of their life time without achieving their originally set goals and with a dearth of community transformation to show for it. Objective: To delineate and highlight the achievements of the Uganda (Makerere University)-Case Western Reserve University Research Collaboration Methods: We retrospectively compiled and reviewed the data on research, training and policy impact achievements of the Uganda (Makerere University)-Case Western Reserve University Research Collaboration over a period of 30 years of its existence. Results: Over the last 35 years, the Uganda (Makerere University)-Case Western Reserve University Research Collaboration trained a total of 104 Ugandans with Masters, PhDs and other varied graduate training programs. More than 70 large tuberculosis/TB+HIV studies were conducted with more than 360 manuscripts published including landmark local and global TB/HIV policy impact publications. Conclusion: The Uganda (Makerere University)-Case Western Reserve University Research Collaboration has in the past 35 years built the capacity of Ugandan and international students through conducting landmark research, training and mentoring and contributed to TB HIV management policy changes in Uganda.

Obstructive lung disease and quality of life after cure of multi-drug-resistant tuberculosis in Uganda: a cross-sectional study.

BACKGROUND: Pulmonary multi-drug-resistant tuberculosis (MDR TB) alters lung architecture and involves lengthy treatment duration, high pill burden, drug adverse effects, travel restrictions, and stigma. Literature about pulmonary function and health-related quality of life (QoL) of patients treated for MDR TB is limited. This study sought to determine the prevalence of chronic obstructive pulmonary disease (COPD) and QoL of patients who were treated for pulmonary MDR TB. METHODS: Participants who completed 18 months of pulmonary MDR TB treatment and considered cured were eligible to be evaluated in a cross-sectional study. We performed post-bronchodilator spirometry to measure forced expiratory volume in 1 s (FEV1) and forced vital capacity (FVC). COPD was defined as FEV1/FVC < 0.7; health-related QoL was assessed using the Medical Outcomes Survey for HIV (MOS-HIV) and St. George's Respiratory Questionnaire (SGRQ). Linear and logistic regression models were used to assess associations with COPD, health-related QoL, and other characteristics of the cohort. RESULTS: A total of 95 participants were enrolled. Median age of the cohort was 39 years (interquartile range (IQR), 29-45), and 55 (58%) were HIV-positive. COPD prevalence was 23% (22/95). Median SGRQ score was normal at 7.8 (IQR, 3.1-14.8). Median mental and physical health summary scores were significantly impaired, at 58.6 (IQR, 52.0-61.5) and 52.9 (IQR, 47.8-57.9), respectively, on a scale of 0 to 100 where 100 represents excellent physical or mental health. In this sample, 19% (18/95) of participants were in the lowest relative socioeconomic position (SEP) while 34% (32/95) were in the highest relative SEP. Belonging in the lowest SEP group was the strongest predictor of COPD. CONCLUSION: Individuals who have completed MDR TB treatment have a high prevalence of COPD and low mental and physical health summary scores. Our study highlights the need for pulmonary rehabilitation programs in patients with a low socioeconomic position (SEP) after MDR TB treatment.

Predictors of recurrent TB in sputum smear and culture positive adults: a prospective cohort study.

OBJECTIVE: To explore simple inexpensive non-culture based predictors of recurrent pulmonary tuberculosis (PTB). SETTING AND STUDY POPULATION: HIV-infected and uninfected adults with the first episode of smear positive, culture-confirmed pulmonary tuberculosis in a high tuberculosis burden country. DESIGN: A nested prospective cohort study of participants with pulmonary tuberculosis (PTB) presenting to a hospital out-patient clinic. RESULTS: A total of 630 TB culture confirmed participants were followed up for eighteen months of which 57 (9%) developed recurrent recurrent TB. On univariate analysis,4.7% low grade(1+) pre-treatment sputum smear participants developed recurrent tuberculosis Vs 8.8% with high grade(3+) smears (OR=0.31,95%CI: 0.10-0.93, p=0.037).On multivariate analysis: participants with extensive fibro-cavitation had a high risk of recurrent TB Vs minimal end of treatment fibro-cavitation (18%Vs12%, OR=2.3,95%CI:1.09-4.68, p=0.03). Weight gain with HIV infection was assosciated with a high risk of recurrent TB Vs weight gain with no HIV infection(18%Vs 6%, OR=6.8,95%CI:165-27.83, p=0.008) where as weight gain with a low pre-treatment high bacillary burden was assosciated with a low risk of recurrent TB Vs weight gain with a high pre-treatmentbacillary burden(6.5%Vs7.9%, OR=0.2,95%CI:0.05-0.79, p=0.02). CONCLUSION: Extensive end of treatment pulmonary fibro-cavitation, high pre-treatment bacillary burden with no weight gain and HIV infection could be reliable predictors of recurrent tuberculosis.

Polypharmacy among HIV positive older adults on anti-retroviral therapy attending an urban clinic in Uganda.

BACKGROUND: Polypharmacy has not been investigated in patients living with HIV in developing countries. The aims of this study were to determine the prevalence of polypharmacy, the factors associated with polypharmacy and whether polypharmacy was associated with adverse effects among older adults on anti-retroviral therapy (ART). METHODS: Cross-sectional study in older adults aged 50 and over on ART attending an outpatient HIV/AIDS care centre in Uganda. Demographic and clinical data collected on number and type of medications plus supplements, possible medication related side-effects, comorbidity, frailty, cognitive impairment, current CD4 count and viral load. RESULTS: Of 411 participants, 63 (15.3, 95% C.I. 11.9, 18.8) had polypharmacy (≥ 4 non- HIV medications). In multivariate analyses, polypharmacy was associated with one or more hospitalisations in the last year (Prevalence Ratio PR = 1.8, 95% C.I. 1.1, 3.1, p = 0.02), prescription by an internist (PR = 3.6, 95% C.I. 1.3, 10.5, p = 0.02) and frailty index scores of 5 to 6 (PR = 10.6, 95% C.I. 1.4, 78, p = 0.02), and 7 or more (PR = 17.4, 95% C.I. 2.4, 126.5, p = 0.005). Polypharmacy was not associated with frequency and severity of possible medication related side effects and falls. CONCLUSION: Polypharmacy is common among older HIV infected patients in sub-Saharan Africa. It's more prevalent among frail people, who have been in hospital in the last year and who have been seen by an internist. We found no evidence that polypharmacy results in any harm but this is worth exploring further.

Identification of Host Proteins Predictive of Early Stage Mycobacterium tuberculosis Infection.

The objective of this study was to identify blood-based protein biomarkers of early stage Mycobacterium tuberculosis (Mtb) infection. We utilized plasma and serum specimens from TB patients and their contacts (age≥12) enrolled in a household contact study in Uganda. In the discovery phase cross-sectional samples from 104 HIV-uninfected persons classified as either active TB, latent Mtb infection (LTBI), tuberculin skin test (TST) converters, or persistent TST-negative were analyzed. Two hundred eighty-nine statistically significant (false discovery rate corrected p<0.05) differentially expressed proteins were identified across all comparisons. Proteins associated with cellular immunity and lipid metabolism were induced early after Mtb infection. One hundred and fifty-nine proteins were selected for a targeted mass spectrometry assay. A set of longitudinal samples from 52 TST-negative subjects who converted to TST-positive or remained TST-negative were analyzed, and multivariate logistic regression was used to identify unique protein panels able to predict TST conversion with cross-validated AUC>0.85. Panel performance was confirmed with an independent validation set of longitudinal samples from 16 subjects. These candidate protein biomarkers may allow for the identification of recently Mtb infected individuals at highest risk for developing active TB and most likely to benefit from preventive therapy.

A single-nucleotide-polymorphism real-time PCR assay for genotyping of Mycobacterium tuberculosis complex in peri-urban Kampala.

BACKGROUND: Accurate and high-throughput genotyping of Mycobacterium tuberculosis complex (MTBC) may be important for understanding the epidemiology and pathogenesis of tuberculosis (TB). In this study, we report the development of a LightCycler® real-time PCR single-nucleotide-polymorphism (LRPS) assay for the rapid determination of MTBC lineages/sublineages in minimally processed sputum samples from TB patients. METHOD: Genotyping analysis of 70 MTBC strains was performed using the Long Sequence Polymorphism-PCR (LSP-PCR) technique and the LRPS assay in parallel. For targeted sequencing, 9 MTBC isolates (three isolates per MTBC lineage) were analyzed for lineage-specific single nucleotide polymorphisms (SNPs) in the following three genes to verify LRPS results: Rv004c for MTB Uganda family, Rv2962 for MTB lineage 4, and Rv0129c for MTB lineage 3. The MTBC lineages present in 300 smear-positive sputum samples were then determined by the validated LRPS method without prior culturing. RESULTS: The LSP-PCR and LRPS assays produced consistent genotyping data for all 70 MTBC strains; however, the LSP-PCR assay was 10-fold less sensitive than the LRPS method and required higher DNA concentrations to successfully characterize the MTBC lineage of certain samples. Targeted sequencing of genes containing lineage-specific SNPs was 100 % concordant with the genotyping results and provided further validation of the LRPS assay. Of the 300 sputum samples analyzed, 58 % contained MTBC from the MTBC-Uganda family, 27 % from the MTBC lineage 4 (excluding MTBC Uganda family), 13 % from the MTBC lineage 3, and the remaining 2 % were of indeterminate lineage. CONCLUSION: The LRPS assay is a sensitive, high-throughput technique with potential application to routine genotyping of MTBC in sputum samples from TB patients.

Distribution and transmission of Mycobacterium tuberculosis complex lineages among children in peri-urban Kampala, Uganda.

BACKGROUND: To gain insight into the transmission of tuberculosis (TB) in peri-urban Kampala-Uganda, we performed a household contact study using children as a surrogate for recent transmission of Mycobacterium tuberculosis (MTB). Using this approach, we sought to understand M. tuberculosis complex (MTBC) lineage diversity, distribution and how these relate to TB transmission to exposed children. METHOD: MTBC isolates from children aged ≤ 15 years, collected from 2002 to 2010 in a household-contact study, were analyzed using a LightCycler RT-PCR SNP genotyping assay (LRPS). The resultant genotypic data was used to determine associations between MTBC lineage and the children's clinical and epidemiological characteristics. RESULTS AND DISCUSSION: Of the 761 children surveyed, 9% (69/761) had culture-positive TB an estimate in the range of global childhood TB; of these 71% (49/69) were infected with an MTBC strain of the "Uganda family", 17% (12/69) infected with MTBC lineage 4 strains other than MTBC Uganda family and 12% (8/69) infected with MTBC lineage 3, thereby disproportionately causing TB in the study area. Overall the data showed no correlation between the MTBC lineages studied and transmission (OR = 0.304; P-value = 0.251; CI: 95%; 0.039-2.326) using children a proxy for TB transmission. CONCLUSIONS: Our findings indicate that MTBC Uganda family strains are the main cause of TB in children in peri-urban Kampala. Furthermore, MTBC lineages did not differ in their transmissibility to children.

Motivations and concerns about adolescent tuberculosis vaccine trial participation in rural Uganda: a qualitative study.

INTRODUCTION: Research is being carried out to develop and test new potentially more effective tuberculosis vaccines. Among the vaccines being developed are those that target adolescents. This study explored the stakeholders' perceptions about adolescent participation in a hypothetical tuberculosis vaccine trial in Ugandan adolescents. METHODS: Focus group discussions with adolescents, parents of infants and adolescents, and key informant interviews with community leaders and traditional healers were conducted. RESULTS: The majority of the respondents expressed potential willingness to allow their children participate in a tuberculosis vaccine trial. Main motivations for potential participation would be being able to learn about health-related issues. Hesitations included the notion that trial participation would distract the youths from their studies, fear of possible side effects of an investigational product, and potential for being sexually exploited by researchers. In addition, bad experiences from participation in previous research and doubts about the importance of research were mentioned. Suggested ways to motivate participation included: improved clarity on study purpose, risks, benefits and better scheduling of study procedures to minimize disruption to participants' academic schedules. CONCLUSION: Findings from this study suggest that the community is open to potential participation of adolescents in a tuberculosis vaccine trial. However, there is a need to communicate more effectively with the community about the purpose of the trial and its effects, including safety data, in a low-literacy, readily understood format. This raises a challenge to researchers, who cannot know all the potential effects of a trial product before it is tested.

Wasting among Uganda men with pulmonary tuberculosis is associated with linear regain in lean tissue mass during and after treatment in contrast to women with wasting who regain fat tissue mass: prospective cohort study.

BACKGROUND: Nutritional changes during and after tuberculosis treatment have not been well described. We therefore determined the effect of wasting on rate of mean change in lean tissue and fat mass as measured by bioelectrical impedance analysis (BIA), and mean change in body mass index (BMI) during and after tuberculosis treatment. METHODS: In a prospective cohort study of 717 adult patients, BMI and height-normalized indices of lean tissue (LMI) and fat mass (FMI) as measured by BIA were assessed at baseline, 3, 12, and 24 months. RESULTS: Men with wasting at baseline regained LMI at a greater rate than FMI (4.55 kg/m2 (95% confidence interval (CI): 1.26, 7.83 versus 3.16 (95% CI: 0.80, 5.52)) per month, respectively during initial tuberculosis therapy. In contrast, women with wasting regained FMI at greater rate than LMI (3.55 kg/m2 (95% CI: 0.40, 6.70) versus 2.07 (95% CI: -0.74, 4.88)), respectively. Men with wasting regained BMI at a rate of 6.45 kg/m2 (95% CI: 3.02, 9.87) in the first three months whereas women, had a rate of 3.30 kg/m2 (95% CI: -0.11, 6.72). There were minimal changes in body composition after month 3 and during months 12 to 24. CONCLUSION: Wasted tuberculosis patients regain weight with treatment but the type of gain differs by gender and patients may remain underweight after the initial phase of treatment.

Long-term dominance of Mycobacterium tuberculosis Uganda family in peri-urban Kampala-Uganda is not associated with cavitary disease.

BACKGROUND: Previous studies have shown that Mycobacterium tuberculosis (MTB) Uganda family, a sub-lineage of the MTB Lineage 4, is the main cause of tuberculosis (TB) in Uganda. Using a well characterized patient population, this study sought to determine whether there are clinical and patient characteristics associated with the success of the MTB Uganda family in Kampala. METHODS: A total of 1,746 MTB clinical isolates collected from 1992-2009 in a household contact study were genotyped. Genotyping was performed using Single Nucleotide Polymorphic (SNP) markers specific for the MTB Uganda family, other Lineage 4 strains, and Lineage 3, respectively. Out of 1,746 isolates, 1,213 were from patients with detailed clinical data. These data were used to seek associations between MTB lineage/sub-lineage and patient phenotypes. RESULTS: Three MTB lineages were found to dominate the MTB population in Kampala during the last two decades. Overall, MTB Uganda accounted for 63% (1,092/1,746) of all cases, followed by other Lineage 4 strains accounting for 22% (394/1,746), and Lineage 3 for 11% (187/1,746) of cases, respectively. Seventy-three (4 %) strains remained unclassified. Our longitudinal data showed that MTB Uganda family occurred at the highest frequency during the whole study period, followed by other Lineage 4 strains and Lineage 3. To explore whether the long-term success of MTB Uganda family was due to increased virulence, we used cavitary disease as a proxy, as this form of TB is the most transmissible. Multivariate analysis revealed that even though cavitary disease was associated with known risk factors such as smoking (adjusted odds ratio (aOR) 4.8, 95% confidence interval (CI) 3.33-6.84) and low income (aOR 2.1, 95% CI 1.47-3.01), no association was found between MTB lineage and cavitary TB. CONCLUSION: The MTB Uganda family has been dominating in Kampala for the last 18 years, but this long-term success is not due to increased virulence as defined by cavitary disease.

Low nutrient intake among adult women and patients with severe tuberculosis disease in Uganda: a cross-sectional study.

BACKGROUND: Information regarding dietary nutrient intake during tuberculosis disease is lacking. We established the relationship between disease severity or wasting during pulmonary tuberculosis and nutrient intake. METHODS: In a cross-sectional study of 131 adults with or without pulmonary tuberculosis were screened for human immune-deficiency virus (HIV), wasting, disease severity using 13 item validated clinical TBscore, and 24-hour dietary intake recall. RESULTS: Of the 131 participants, 61 were males and 70 females. Overall men and women had similar age. In average 24-hour nutrient intake, the following nutrients: energy, protein, total fat, carbohydrate, calcium, vitamin A, and folate were low among patients with severe tuberculosis disease. Patients with moderate-to-severe clinical TBscore had lower average energy intake than patients with mild TBscores (6.11 vs. 9.27 MJ, respectively) (p<0.05). The average 24-hour nutrient intakes between wasted and non-wasted tuberculosis patients were comparable. Nutrient intake among men was higher when compared to women regardless of wasting and severity of tuberculosis. Among those with wasting, men had higher average energy intake than women (8.87 vs. 5.81 MJ, respectively) (p<0.05). Among patients with mild disease, men had higher average energy intake than women with mild disease (12.83 vs. 7.49 kcal, respectively) (p<0.001). CONCLUSIONS: Findings suggest that severity of pulmonary tuberculosis and female gender had reduced nutrient intake. Early tuberculosis diagnosis and nutritional support may be important in management of tuberculosis patients.

Lean tissue mass wasting is associated with increased risk of mortality among women with pulmonary tuberculosis in urban Uganda.

OBJECTIVES: We assessed the impact of wasting on survival in patients with tuberculosis by using a precise height-normalized lean tissue mass index (LMI) estimated by bioelectrical impedance analysis and body mass index (BMI). METHODS: In a retrospective cohort study, 747 adult pulmonary patients with tuberculosis who were screened for HIV and nutritional status were followed for survival. RESULTS: Of 747 patients, 310 had baseline wasting by BMI (kg/m(2)) and 103 by LMI (kg/m(2)). Total deaths were 105. Among men with reduced BMI, risk of death was 70% greater (hazard ratio [HR] 1.7, 95% confidence interval [95% CI] 1.03-2.81) than in men with normal BMI. Survival did not differ by LMI among men (HR 1.1; 95% CI 0.5-2.9). In women, both the BMI and LMI were associated with survival. Among women with reduced BMI, risk of death was 80% greater (HR 1.8; 95% CI 0.9-3.5) than in women with normal BMI; risk of death was 5-fold greater (HR 5.0; 95% CI 1.6-15.9) for women with low LMI compared with women with normal LMI. CONCLUSIONS: Wasting assessed by reduced BMI is associated with an increased risk for death among both men and women whereas reduced LMI is among women with tuberculosis.

Innate and adaptive immune responses during acute M. tuberculosis infection in adult household contacts in Kampala, Uganda.

Contacts of active pulmonary tuberculosis (TB) patients are at risk for Mycobacterium tuberculosis (MTB) infection. Because most infections are controlled, studies during MTB infection provide insight into protective immunity. We compared immune responses of adult household contacts that did and did not convert the tuberculin skin test (TST). Innate and adaptive immune responses were measured by whole blood assay. Responses of TST converters (TSTC) were compared with persistently TST negative contacts (PTST-) and contacts who were TST+ at baseline (TST+). TLR-2, TLR-4, and IFN-γR responses to IFN-γ did not differ between the groups, nor did γδ T cell responses. T cell responses to MTB antigens differed markedly among TSTC, PTST-, and TST+ contacts. Thus, no differences in innate responses were found among the three household contact groups. However, adaptive T cell responses to MTB antigens did differ before and during MTB infection among PTST-, TSTC, and TST+ contacts.

Detection of multiple strains of Mycobacterium tuberculosis using MIRU-VNTR in patients with pulmonary tuberculosis in Kampala, Uganda.

BACKGROUND: Many studies using DNA fingerprinting to differentiate Mycobacterium tuberculosis (MTB) strains reveal single strains in cultures, suggesting that most disease is caused by infection with a single strain. However, recent studies using molecular epidemiological tools that amplify multiple targets have demonstrated simultaneous infection with multiple strains of MTB. We aimed to determine the prevalence of MTB multiple strain infections in Kampala, and the impact of these infections on clinical presentation of tuberculosis (TB) and response to treatment. METHODS: A total of 113 consecutive smear and culture positive patients who previously enrolled in a house-hold contact study were included in this study. To determine whether infection with multiple MTB strains has a clinical impact on the initial presentation of patients, retrospective patient data (baseline clinical, radiological and drug susceptibility profiles) was obtained. To determine presence of infections with multiple MTB strains, MIRU-VNTR (Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem Repeats) -PCR was performed on genomic DNA extracted from MTB cultures of smear positive sputum samples at baseline, second and fifth months. RESULTS: Of 113 patients, eight (7.1%) had infection with multiple MTB strains, coupled with a high rate of HIV infection (37.5% versus 12.6%, p = 0.049). The remaining patients (105) were infected with single MTB strains. The proportions of patients with MTB smear positive cultures after two and five months of treatment were similar. There was no difference between the two groups for other variables. CONCLUSION: Infection with multiple MTB strains occurs among patients with first episode of pulmonary tuberculosis in Kampala, in a setting with high TB incidence. Infection with multiple MTB strains had little impact on the clinical course for individual patients. This is the first MIRU-VNTR-based study from in an East African country.

Mycobacterium tuberculosis microbiologic and clinical treatment outcomes in a randomized trial of immediate versus CD4(+)-initiated antiretroviral therapy in HIV-infected adults with a high CD4(+) cell count.

In a prospective randomized, controlled trial in Uganda comparing the efficacy of antiretroviral therapy during tuberculosis therapy with the efficacy of tuberculosis therapy alone in HIV-infected patients with tuberculosis who have a CD4(+) cell count >350 cells/microL, it was found that antiretroviral therapy did not accelerate microbiologic, radiographic, or clinical responses to tuberculosis therapy: 18% of participants had sputum smears positive for Mycobacterium tuberculosis after 5 months of tuberculosis therapy, despite having had negative culture results. Trial registration. ClinicalTrials.gov identifier: NCT00078247 .

Immunogenicity of novel DosR regulon-encoded candidate antigens of Mycobacterium tuberculosis in three high-burden populations in Africa.

Increasing knowledge about DosR regulon-encoded proteins has led us to produce novel Mycobacterium tuberculosis antigens for immunogenicity testing in human populations in three countries in Africa to which tuberculosis (TB) is endemic. A total of 131 tuberculin skin test-positive and/or ESAT-6/CFP10-positive, human immunodeficiency virus-negative adult household contacts of active pulmonary TB cases from South Africa (n = 56), The Gambia (n = 26), and Uganda (n = 49) were tested for gamma interferon responses to 7 classical and 51 DosR regulon-encoded M. tuberculosis recombinant protein antigens. ESAT-6/CFP10 fusion protein evoked responses in >75% of study participants in all three countries. Of the DosR regulon-encoded antigens tested, Rv1733c was the most commonly recognized by participants from both South Africa and Uganda and the third most commonly recognized antigen in The Gambia. The four most frequently recognized DosR regulon-encoded antigens in Uganda (Rv1733c, Rv0081, Rv1735c, and Rv1737c) included the three most immunogenic antigens in South Africa. In contrast, Rv3131 induced the highest percentage of responders in Gambian contacts (38%), compared to only 3.4% of Ugandan contacts and no South African contacts. Appreciable percentages of TB contacts with a high likelihood of latent M. tuberculosis infection responded to several novel DosR regulon-encoded M. tuberculosis proteins. In addition to significant similarities in antigen recognition profiles between the three African population groups, there were also disparities, which may stem from genetic differences between both pathogen and host populations. Our findings have implications for the selection of potential TB vaccine candidates and for determining biosignatures of latent M. tuberculosis infection, active TB disease, and protective immunity.

Genomic survey of the non-cultivatable opportunistic human pathogen, Enterocytozoon bieneusi.

Enterocytozoon bieneusi is the most common microsporidian associated with human disease, particularly in the immunocompromised population. In the setting of HIV infection, it is associated with diarrhea and wasting syndrome. Like all microsporidia, E. bieneusi is an obligate, intracellular parasite, but unlike others, it is in direct contact with the host cell cytoplasm. Studies of E. bieneusi have been greatly limited due to the absence of genomic data and lack of a robust cultivation system. Here, we present the first large-scale genomic dataset for E. bieneusi. Approximately 3.86 Mb of unique sequence was generated by paired end Sanger sequencing, representing about 64% of the estimated 6 Mb genome. A total of 3,804 genes were identified in E. bieneusi, of which 1,702 encode proteins with assigned functions. Of these, 653 are homologs of Encephalitozoon cuniculi proteins. Only one E. bieneusi protein with assigned function had no E. cuniculi homolog. The shared proteins were, in general, evenly distributed among the functional categories, with the exception of a dearth of genes encoding proteins associated with pathways for fatty acid and core carbon metabolism. Short intergenic regions, high gene density, and shortened protein-coding sequences were observed in the E. bieneusi genome, all traits consistent with genomic compaction. Our findings suggest that E. bieneusi is a likely model for extreme genome reduction and host dependence.

Evaluation of a low-cost method, the Guava EasyCD4 assay, to enumerate CD4-positive lymphocyte counts in HIV-infected patients in the United States and Uganda.

OBJECTIVE: To evaluate the EasyCD4 assay, a less expensive method to enumerate CD4+ lymphocytes, in resource-limited settings. DESIGN: Cross-sectional study conducted in the United States and Uganda. METHODS: We compared CD4+ cell counts obtained on replicate samples from HIV-infected patients by the EasyCD4 assay, a microcapillary flow-based system, and by standard flow cytometry or FACSCount with linear regression and the Bland-Altman method. RESULTS: Two hundred eighteen samples were analyzed (77 in the United States and 141 in Uganda). In the United States, mean +/- SD CD4 was 697 +/- 438 cells/microL by standard flow cytometry and 688 +/- 451 cells/microL by EasyCD4. In Uganda, the mean +/- SD CD4 was 335 +/- 331 cells/microL by FACSCount and 340 +/- 327 cells/microL by EasyCD4. The 2 methods were highly correlated (US cohort, r2 = 0.97, slope = 1.0, intercept = -18; Ugandan cohort, r2 = 0.92; slope = 0.95; intercept = 23). The mean differences in CD4 cell counts were 9.0 and -4.6 cells/microL for the US and Ugandan cohorts, respectively, and they were not significant in either cohort. In the Ugandan cohort, sensitivity and specificity of the EasyCD4 for CD4 below 200 cells/microL were 90% and 98%, respectively. Positive predictive value was 96%; negative predictive value was 93%. CONCLUSIONS: Our results suggest that EasyCD4 may be used with high positive and negative predictive value in resource-limited settings.