Basis for Treatment of Tuberculosis among HIV-Infected Patients in Tanzania: The Role of Chest X-Ray and Sputum Culture

Active tuberculosis (TB) is common among HIV-infected persons living in tuberculosis endemic countries, and screening for tuberculosis (TB) is recommended routinely. We sought to determine the role of chest x-ray and sputum culture in the decision to treat for presumptive TB using active case finding in a large cohort of HIV-infected patients. Ambulatory HIV-positive subjects with CD4 counts ≥ 200/mm3 entering a Phase III TB vaccine study in Tanzania were screened for TB with a physical examination, standard interview, CD4 count, chest x-ray (CXR), blood culture for TB, and three sputum samples for acid fast bacillus (AFB) smear and culture

Maintenance of Sensitivity of the T-SPOT.TB Assay after Overnight Storage of Blood Samples, Dar es Salaam, Tanzania

Background. T-SPOT.TB is an interferon gamma release assay for detecting Mycobacterium tuberculosis infection. The requirement to process within 8 hours is constraining, deters use, and leads to invalid results. Addition of T Cell Xtend reagent may allow delayed processing, but has not been extensively field tested. Design. Consecutive AFB smear positive adult tuberculosis patients were prospectively recruited in Dar es Salaam, Tanzania. Patients provided a medical history, 1–3 sputum samples for culture and 1 blood sample which was transported to the laboratory under temperature-controlled conditions. After overnight storage, 25 μL of T Cell Xtend reagent was added per mL of blood, and the sample was tested using T-SPOT.TB. Results. 143 patients were enrolled: 57 patients were excluded because temperature control was not maintained, 19 patients were excluded due to red blood cell contamination, and one did not provide a sputum sample for culture. Among 66 evaluable patients, overall agreement between T-SPOT.TB and culture was 95.4% (95%CI; 87.1–99.0%) with Kappa value 0.548. Sensitivity of T-SPOT.TB when using T Cell Xtend reagent was 96.8% (95%CI; 88.8–99.6%). Conclusions. When T Cell Xtend reagent is added to specimens held overnight at recommended temperatures, T-SPOT.TB is as sensitive as the standard assay in patients with tuberculosis

Maintenance of Sensitivity of the T-SPOT.TB Assay after Overnight Storage of Blood Samples, Dar es Salaam, Tanzania

Background. T-SPOT.TB is an interferon gamma release assay for detecting Mycobacterium tuberculosis infection. The requirement to process within 8 hours is constraining, deters use, and leads to invalid results. Addition of T Cell Xtend reagent may allow delayed processing, but has not been extensively field tested. Design. Consecutive AFB smear positive adult tuberculosis patients were prospectively recruited in Dar es Salaam, Tanzania. Patients provided a medical history, 1–3 sputum samples for culture and 1 blood sample which was transported to the laboratory under temperature-controlled conditions. After overnight storage, 25 μL of T Cell Xtend reagent was added per mL of blood, and the sample was tested using T-SPOT.TB. Results. 143 patients were enrolled: 57 patients were excluded because temperature control was not maintained, 19 patients were excluded due to red blood cell contamination, and one did not provide a sputum sample for culture. Among 66 evaluable patients, overall agreement between T-SPOT.TB and culture was 95.4% (95%CI; 87.1–99.0%) with Kappa value 0.548. Sensitivity of T-SPOT.TB when using T Cell Xtend reagent was 96.8% (95%CI; 88.8–99.6%). Conclusions. When T Cell Xtend reagent is added to specimens held overnight at recommended temperatures, T-SPOT.TB is as sensitive as the standard assay in patients with tuberculosis

Sputum Microscopy for the Diagnosis of HIV-Associated Pulmonary Tuberculosis in Tanzania

Background: In many resource poor settings only sputum microscopy is employed for the diagnosis of HIV-associated pulmonary tuberculosis; sputum culture may not be available. Methods: We determined the diagnostic accuracy of sputum microscopy for active case finding of HIV-associated pulmonary tuberculosis using TB culture as the reference standard. Results: 2216 potential subjects screened for a TB vaccine trial submitted 9454 expectorated sputum specimens: 212 (2.2%) were sputum culture positive for Mycobacterium tuberculosis (MTB), 31 (0.3%) for non-tuberculous mycobacteria, and 79 (0.8%) were contaminated. The overall sensitivity of sputum microscopy was 61.8% (131/212) and specificity 99.7% (9108/9132). Sputum microscopy sensitivity varied from 22.6% in specimens with \u3c 20 colony forming units (CFU)/specimen to 94.2% in patients with \u3e 100 CFU/specimen plus confluent growth. The incremental diagnostic value for sputum microscopy was 92.1%, 1.8% and 7.1% for the first, second and third specimens, respectively. The positive predictive value and negative predictive values for sputum microscopy were 84.5% and 99.1%, respectively. The likelihood ratio (LR) of a positive sputum microscopy was 235.1 (95% CI 155.8 – 354.8), while the LR of a negative test was 0.38 (95CI 0.32 – 0.45). The 212 positive sputum cultures for MTB represented 103 patients; sputum microscopy was positive for 57 (55.3%) of 103 patients. Conclusion: Sputum microscopy on 3 expectorated sputum specimens will only detect 55% of culture positive HIV-infected patients in active screening for pulmonary tuberculosis. Sensitivity is higher in patients with greater numbers of CFUs in the sputum. Culture is required for active case finding of HIV- associated pulmonary tuberculosis

Molecular Epidemiology of HIV-Associated Tuberculosis in Dar es Salaam, Tanzania: Strain Predominance, Clustering, and Polyclonal Disease

Molecular typing of Mycobacterium tuberculosis can be used to elucidate the epidemiology of tuberculosis, including the rates of clustering, the frequency of polyclonal disease, and the distribution of genotypic families. We performed IS6110 typing and spoligotyping on M. tuberculosis strains isolated from HIV-infected subjects at baseline or during follow-up in the DarDar Trial in Tanzania and on selected community isolates. Clustering occurred in 203 (74%) of 275 subjects: 124 (80%) of 155 HIV-infected subjects with baseline isolates, 56 (69%) of 81 HIV-infected subjects with endpoint isolates, and 23 (59%) of 39 community controls. Overall, 113 (41%) subjects had an isolate representing the East Indian “GD” family. The rate of clustering was similar among vaccine and placebo recipients and among subjects with or without cellular immune responses to mycobacterial antigens. Polyclonal disease was detected in 6 (43%) of 14 patients with multiple specimens typed. Most cases of HIV-associated tuberculosis among subjects from this study in Dar es Salaam resulted from recently acquired infection. Polyclonal infection was detected and isolates representing the East Indian GD strain family were the most common

Lymphocyte proliferation to mycobacterial antigens is detectable across a spectrum of HIV-associated tuberculosis

<p>Abstract</p> <p>Background</p> <p>Identifying novel TB diagnostics is a major public health priority. We explored the diagnostic characteristics of antimycobacterial lymphocyte proliferation assays (LPA) in HIV-infected subjects with latent or active TB.</p> <p>Methods</p> <p>HIV-infected subjects with bacille Calmette Guérin (BCG) scars and CD4 counts ≥ 200 cells/mm³entering a TB booster vaccine trial in Tanzania had baseline in vivo and in vitro immune tests performed: tuberculin skin tests (TST), LPA and five day assays of interferon gamma (IFN-γ) release. Assay antigens were early secreted antigenic target 6 (ESAT-6), antigen 85 (Ag85), and <it>Mycobacterium tuberculosis </it>whole cell lysate (WCL). Subjects were screened for active TB at enrollment by history, exam, sputum smear and culture. We compared antimycobacterial immune responses between subjects with and without latent or active TB at enrollment.</p> <p>Results</p> <p>Among 1885 subjects screened, 635 had latent TB and 13 had active TB. Subjects with latent TB were more likely than subjects without TB to have LPA responses to ESAT-6 (13.2% vs. 5.5%, P < 0.0001), Ag85 (18.7% vs. 3.1%, P < 0.0001), and WCL (45.7% vs. 17.1%, P < 0.0001). Subjects with active TB also were more likely than those without active TB to have detectable LPA responses to ESAT-6 (38.5% vs. 8.1%, P = 0.0001), Ag85 (46.2% vs. 8.5%, P < 0.0001), and WCL (61.5% vs. 27.0%, P = 0.0053). In subjects with a positive TST, LPA responses to ESAT-6, Ag85 and WCL were more common during active TB (p < 0.0001 for all tests). In diagnosing active TB, in vivo and in vitro tests of mycobacterial immune responses had sensitivity and specificity as follows: TST 84.6% and 65.5%, ESAT-6 LPA 38.5% and 92.0%, Ag85 LPA 46.2% and 91.5%, and WCL LPA 61.5% and 73.0%. Detectable LPA responses were more common in patients with higher CD4 counts, and higher HIV viral loads.</p> <p>Conclusion</p> <p>Lymphoproliferative responses to mycobacteria are detectable during HIV-associated active TB, and are less sensitive but more specific than TST.</p> <p>Trial registration</p> <p>ClinicalTrials.gov Identifier NCT00052195.</p

Basis for treatment of tuberculosis among HIV-infected patients in Tanzania: the role of chest x-ray and sputum culture

BACKGROUND: Active tuberculosis (TB) is common among HIV-infected persons living in tuberculosis endemic countries, and screening for tuberculosis (TB) is recommended routinely. We sought to determine the role of chest x-ray and sputum culture in the decision to treat for presumptive TB using active case finding in a large cohort of HIV-infected patients. METHODS: Ambulatory HIV-positive subjects with CD4 counts ≥ 200/mm3 entering a Phase III TB vaccine study in Tanzania were screened for TB with a physical examination, standard interview, CD4 count, chest x-ray (CXR), blood culture for TB, and three sputum samples for acid fast bacillus (AFB) smear and culture. RESULTS: Among 1176 subjects 136 (12%) were treated for presumptive TB. These patients were more frequently male than those without treatment (34% vs. 25%, respectively; p = 0.049) and had lower median CD4 counts (319/μL vs. 425/μL, respectively; p < .0001). Among the 136 patients treated for TB, 38 (28%) had microbiologic confirmation, including 13 (10%) who had a normal CXR and no symptoms. There were 58 (43%) treated patients in whom the only positive finding was an abnormal CXR. Blood cultures were negative in all patients. CONCLUSION: Many ambulatory HIV-infected patients with CD4 counts ≥ 200/mm3 are treated for presumptive TB. Our data suggest that optimal detection requires comprehensive evaluation, including CXR and sputum culture on both symptomatic and asymptomatic subjects.National Institutes of Health (A1 45407); Fogarty International Center (D43-TW006807

Polyantigenic Interferon-γ Responses Are Associated with Protection from TB among HIV-Infected Adults with Childhood BCG Immunization

Surrogate immunologic markers for natural and vaccine-mediated protection against tuberculosis (TB) have not been identified. HIV-infected adults with childhood BCG immunization entering the placebo arm of the DarDar TB vaccine trial in Dar es Salaam, Tanzania, were assessed for interferon gamma (IFN-γ) responses to three mycobacterial antigen preparations--secreted Mycobacterium tuberculosis antigens 85 (Ag85), early secretory antigenic target 6 (ESAT-6) and polyantigenic whole cell lysate (WCL). We investigated the association between the number of detectable IFN-γ responses at baseline and the subsequent risk of HIV-associated TB. During a median follow-up of 3.3 years, 92 (9.4%) of 979 placebo recipients developed TB. The incidence of TB was 14% in subjects with no detectable baseline IFN-γ responses vs. 8% in subjects with response to polyantigenic WCL (P = 0.028). Concomitant responses to secreted antigens were associated with further reduction in the incidence of HIV-associated TB. Overall the percentage of subjects with 0, 1, 2 and 3 baseline IFN-γ responses to mycobacterial preparations who developed HIV-associated TB was 14%, 8%, 7% and 4%, respectively (P = 0.004). In a multivariate Cox regression model, the hazard of developing HIV-associated TB was 46% lower with each increment in the number of detectable baseline IFN-γ responses (P<0.001). Among HIV-infected adults who received BCG in childhood and live in a TB-endemic country, polyantigenic IFN-γ responses are associated with decreased risk of subsequent HIV-associated TB. ClinicalTrials.gov NCT0052195

Maintenance of Sensitivity of the T-SPOT.TB Assay after Overnight Storage of Blood Samples, Dar es Salaam, Tanzania

Background. T-SPOT.TB is an interferon gamma release assay for detecting Mycobacterium tuberculosis infection. The requirement to process within 8 hours is constraining, deters use, and leads to invalid results. Addition of T Cell Xtend reagent may allow delayed processing, but has not been extensively field tested. Design. Consecutive AFB smear positive adult tuberculosis patients were prospectively recruited in Dar es Salaam, Tanzania. Patients provided a medical history, 1–3 sputum samples for culture and 1 blood sample which was transported to the laboratory under temperature-controlled conditions. After overnight storage, 25 μL of T Cell Xtend reagent was added per mL of blood, and the sample was tested using T-SPOT.TB. Results. 143 patients were enrolled: 57 patients were excluded because temperature control was not maintained, 19 patients were excluded due to red blood cell contamination, and one did not provide a sputum sample for culture. Among 66 evaluable patients, overall agreement between T-SPOT.TB and culture was 95.4% (95%CI; 87.1–99.0%) with Kappa value 0.548. Sensitivity of T-SPOT.TB when using T Cell Xtend reagent was 96.8% (95%CI; 88.8–99.6%). Conclusions. When T Cell Xtend reagent is added to specimens held overnight at recommended temperatures, T-SPOT.TB is as sensitive as the standard assay in patients with tuberculosis