Altruism, Scepticism, and Collective Decision-Making in Foreign-Born U.S. Residents in a Tuberculosis Vaccine Trial

Background: The current vaccine against tuberculosis, BCG, is effective when given in most TB-endemic countries at birth but has diminished efficacy against pulmonary TB after 15–20 years. As a result, new booster vaccines for adolescents and adults are being developed to realize the World Health Organization target of global elimination of TB by 2035. Multiple TB candidates thus are in active clinical development. Methods: One of these, DAR-901, is advancing in human clinical trials. These clinical trials are conducted in BCG immunized adults with and without HIV infection in order to assess safety and efficacy among the people most in need of a new vaccine. A Phase I dose escalation trial of DAR-901 in BCG-immunized adults with or without HIV infection was conducted between 2014 and 2016. This offered an unusual opportunity to qualitatively examine why foreign-born adults living in the United States – a poorly studied population – decide to participate, or not, in clinical trials. Results: We conducted a qualitative study of individuals who were recruited to participate in this Phase I vaccine trial, interviewing those who agreed and declined to participate. We found diverse motivations for participation or refusal; varied understandings of tuberculosis and vaccines; and complex views about how ‘informed consent’ can be at odds with cultural understandings of power, authority, and medical decision-making. These dynamics included: knowledge (direct or indirect) of tuberculosis, a desire to be altruistic and simultaneous hopes for personal gain as well as concerns over what remuneration for participation could mean, the importance of personal relationships with care providers in shaping volunteerism, concerns over privacy, and evidence of how culture and history shape medical decision-making. Conclusions: This US-based trial, aimed at addressing a crucible global health issue, raises productive questions about the interface between altruism and scepticism regarding clinical research participation

Clinical and Epidemiological Correlates of Genotypes within the Mycobacterium avium Complex Defined by Restriction and Sequence Analysis of hsp65

Species identification of isolates of the Mycobacterium avium complex (MAC) remains a difficult task. Although M. avium and Mycobacterium intracellulare can be identified with expensive, commercially available probes, many MAC isolates remain unresolved, including those representing Mycobacterium lentiflavum as well as other potentially undefined species. PCR restriction analysis (PRA) of the hsp65 gene has been proposed as a rapid and inexpensive approach. We applied PRA to 278 MAC isolates, including 126 from blood of human immunodeficiency virus (HIV)-infected patients, 59 from sputum of HIV-negative patients with chronic obstructive pulmonary disease, 88 from environmental sources, and 5 pulmonary isolates from a different study. A total of 15 different PRA patterns were observed. For 27 representative isolates, a 441-bp fragment of the hsp65 gene was sequenced; based on 54 polymorphic sites, 18 different alleles were defined, including 12 alleles not previously reported. Species and phylogenetic relationships were more accurately defined by sequencing than by PRA or commercial probe. The distribution of PRA types and, by implication, phylogenetic lineages among blood isolates was significantly different from that for pulmonary and environmental isolates, suggesting that particular lineages have appreciably greater virulence and invasive potential

Polyclonal Mycobacterium Avium Infections in Patients with AIDS: Variations in Antimicrobial Susceptibilities of Different Strains of M. Avium Isolated from the Same Patient.

Broth microdilution MICs were determined for pairs of strains isolated from five AIDS patients with polyclonal Mycobacterium avium infection. Four (80%) of the five patients were infected simultaneously with strains having different antimicrobial susceptibility patterns. These findings have implications for the interpretation of susceptibility data in M. avium prophylaxis and treatment trials

Polyclonal Infections Due to Mycobacterium Avium Complex in Patients with AIDS Detected by Pulsed-field Gel Electrophoresis of Sequential Clinical Isolates.

Invasive infection with organisms of the Mycobacterium avium complex (MAC) is common among patients with advanced human immunodeficiency virus infection. In previous studies, we analyzed multiple individual colonies of MAC isolated from specimens obtained at the same time and observed that 14 to 20% of patients are simultaneously infected with more than one strain. In this study, we examined sequential isolates from 12 patients with AIDS who had two or more MAC isolates available from clinical specimens collected more than 1 week apart; the intervals between the first and last specimens ranged from 8 to 192 (median, 46) days. For each isolate, restriction digests of genomic DNA were analyzed by pulsed-field gel electrophoresis; DNA was prepared by using a protocol, described here in detail, which had been optimized for conditions of bacterial growth and lysis. The pulsed-field gel electrophoresis analysis identified four patients (33%) infected with two different MAC strains. Both M. avium and M. intracellulare were cultured from blood specimens from two patients. In each of the four patients, the second strain was identified from a culture taken within 14 days of the initial study isolate, and in three of these patients, the first strain was detected again in a subsequent culture. These observations suggest that the presence of two different strains among isolates from sequential cultures may reflect ongoing polyclonal infection. We conclude that polyclonal infection with MAC is common among patients with AIDS. The identification of such infections may be critical in the development of effective treatments

Isolation of Mycobacterium avium from Potable Water in Homes and Institutions of Patients with HIV Infection in Finland and the United States

Symptomatic disease by nontuberculous mycobacteria has been linked to potable water from institutional and domestic potable water systems. Potable water samples were collected from homes and institutions of patients with AIDS. Colonization of potable water with nontuberculous mycobacteria was demonstrated in 230 (15%) of 1489 samples collected from domestic and institutional water systems of patients with HIV infection in the United States and Finland. Mycobacterium avium was the most common species and colonization was favored at temperatures of 40-50 degrees C in recirculating hot water systems. Such systems are a plausible source of human infection and disease.Peer reviewe

Test Characteristics of Urinary Lipoarabinomannan and Predictors of Mortality among Hospitalized HIV-Infected Tuberculosis Suspects in Tanzania.

Tuberculosis is the most common cause of death among patients with HIV infection living in tuberculosis endemic countries, but many cases are not diagnosed pre-mortem. We assessed the test characteristics of urinary lipoarabinomannan (LAM) and predictors of mortality among HIV-associated tuberculosis suspects in Tanzania. We prospectively enrolled hospitalized HIV-infected patients in Dar es Salaam, with ≥2 weeks of cough or fever, or weight loss. Subjects gave 2 mLs of urine to test for LAM using a commercially available ELISA, ≥2 sputum specimens for concentrated AFB smear and solid media culture, and 40 mLs of blood for culture. Among 212 evaluable subjects, 143 (68%) were female; mean age was 36 years; and the median CD4 count 86 cells/mm(3). 69 subjects (33%) had culture confirmation of tuberculosis and 65 (31%) were LAM positive. For 69 cases of sputum or blood culture-confirmed tuberculosis, LAM sensitivity was 65% and specificity 86% compared to 36% and 98% for sputum smear. LAM test characteristics were not different in patients with bacteremia but showed higher sensitivity and lower specificity with decreasing CD4 cell count. Two month mortality was 64 (53%) of 121 with outcomes available. In multivariate analysis there was significant association of mortality with absence of anti-retroviral therapy (p = 0.004) and a trend toward association with a positive urine LAM (p = 0.16). Among culture-negative patients mortality was 9 (75%) of 12 in LAM positive patients and 27 (38%) of 71 in LAM negative patients (p = 0.02). Urine LAM is more sensitive than sputum smear and has utility for the rapid diagnosis of culture-confirmed tuberculosis in this high-risk population. Mortality data raise the possibility that urine LAM may also be a marker for culture-negative 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

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

CD4+ T cell cytokine responses to the DAR-901 booster vaccine in BCG-primed adults:A randomized, placebo-controlled trial

<div><p>Background</p><p>DAR-901 is an inactivated whole cell tuberculosis booster vaccine, prepared using a new scalable, broth-grown method from the master cell bank of SRL172, a vaccine previously shown to prevent tuberculosis. This study examined whether DAR-901 (a) induces CD4+ T cell cytokine profiles previously proposed as correlates of protection and (b) has a specific vaccine-induced immunological signature compared to BCG or placebo.</p><p>Methods</p><p>We analysed CD4+ T cell cytokine immune responses from 10 DAR-901 recipients, 9 BCG recipients and 9 placebo recipients from the Phase I DAR-901 MDES trial. In that study, HIV-negative, IGRA-negative participants with prior BCG immunization were randomized (double-blind) to receive three intradermal injections of DAR-901 or saline placebo or two injections of saline placebo followed by an intradermal injection of BCG. Antigen-specific functional and phenotypic CD4+ T cell responses along with effector phenotype of responder cells were measured by intracellular cytokine staining.</p><p>Results</p><p>DAR-901 recipients exhibited increased DAR-901 antigen-specific polyfunctional or bifunctional T cell responses compared to baseline. Vaccine specific CD4+ IFNγ, IL2, TNFα and any cytokine responses peaked at 7 days post-dose 3. Th1 responses predominated, with most responder cells exhibiting a polyfunctional effector memory phenotype. BCG induced greater CD4+ T cell responses than placebo while the more modest DAR-901 responses did not differ from placebo. Neither DAR-901 nor BCG induced substantial or sustained Th17 /Th22 cytokine responses.</p><p>Conclusion</p><p>DAR-901, a TB booster vaccine grown from the master cell bank of SRL 172 which was shown to prevent TB, induced low magnitude polyfunctional effector memory CD4+ T cell responses. DAR-901 responses were lower than those induced by BCG, a vaccine that has been shown ineffective as a booster to prevent tuberculosis disease. These results suggest that induction of higher levels of CD4+ cytokine stimulation may not be a critical or pre-requisite characteristic for candidate TB vaccine boosters.</p><p>Trial registration</p><p>ClinicalTrials.gov <a href="https://clinicaltrials.gov/ct2/show/NCT02063555" target="_blank">NCT02063555</a>.</p></div