Performance comparison of a pair of Lowenstein-Jensen media supplemented with pyruvate or glycerol, and the combination of both supplements in a single Lowenstein-Jensen medium for the growth support of the Mycobacterium Tuberculosis complex.

OBJECTIVE/BACKGROUND: To evaluate the performance of Lowenstein-Jensen medium (LJ) supplemented with pyruvate and glycerol (LJPG), compared with LJ supplemented with pyruvate (LJP) or glycerol (LJG) for the support of mycobacterial growth. METHOD: This study used 100 Ziehl-Neelsen-confirmed positive mycobacterium growth indicator tube 960 culture samples that were obtained from clinical samples during routine diagnosis. All cultures were inoculated in parallel on LJG/LJP and on LJGP, which were incubated and read weekly for the evidence of growth. The mycobacterial recovery rate, contamination rate, and time to detection were compared. RESULT: The recovery rate for LJG/LJP and for LJPG was 90% (90 samples) and 88% (88 samples), respectively (kappa p-value, 0.9). There was no significant difference in the contamination rate, which was 8% (8 samples) for LJG/LJP and 9% (9 samples) for LJPG. Mycobacterial growth was faster in LJPG (1.6weeks) than in LJG/LJP (2weeks). CONCLUSION: A single LJPG slope was not significantly different, compared with the usual pair of LJG or LJP slopes. This is a promising new culturing approach that could be used in Mycobacterium africanum-endemic in West African countries. It significantly reduces labor time and consumable costs and more quickly detects the M. tuberculosis complex.

Immunogenic Mycobacterium africanum strains associated with ongoing transmission in The Gambia.

In West Africa, Mycobacterium tuberculosis strains co-circulate with M. africanum, and both pathogens cause pulmonary tuberculosis in humans. Given recent findings that M. tuberculosis T-cell epitopes are hyperconserved, we hypothesized that more immunogenic strains have increased capacity to spread within the human host population. We investigated the relationship between the composition of the mycobacterial population in The Gambia, as measured by spoligotype analysis, and the immunogenicity of these strains as measured by purified protein derivative-induced interferon-γ release in ELISPOT assays of peripheral blood mononuclear cells. We found a positive correlation between strains with superior spreading capacity and their relative immunogenicity. Although our observation is true for M. tuberculosis and M. africanum strains, the association was especially pronounced in 1 M. africanum sublineage, characterized by spoligotype shared international type 181, which is responsible for 20% of all tuberculosis cases in the region and therefore poses a major public health threat in The Gambia.

Immunogenicity of antigens from the TbD1 region present in M. africanum and missing from "modern" M. tuberculosis: a cross- sectional study.

BACKGROUND: Currently available tools cannot be used to distinguish between sub-species of the M. tuberculosis complex causing latent tuberculosis (TB) infection. M. africanum causes up to half of TB in West- Africa and its relatively lower progression to disease suggests the presence of a large reservoir of latent infection relative to M. tuberculosis. METHODS: We assessed the immunogenicity of the TbD1 region, present in M. africanum and absent from "modern" M. tuberculosis, in an ELISPOT assay using cells from confirmed M. africanum or M. tuberculosis infected TB patients without HIV infection in the Gambia. RESULTS: Antigens from the TbD1 region induced IFNgamma responses in only 35% patients and did not discriminate between patients infected with M. africanum vs. M. tuberculosis, while PPD induced universally high responses. CONCLUSIONS: Further studies will need to assess other antigens unique to M. africanum that may induce discriminatory immune responses.

Mycobacterium africanum elicits an attenuated T cell response to early secreted antigenic target, 6 kDa, in patients with tuberculosis and their household contacts.

BACKGROUND: Mycobacterium africanum, a member of the M. tuberculosis complex that is infrequently found outside of western Africa, is the cause of up to half of the tuberculosis cases there. METHODS: We genotyped mycobacterial isolates obtained from a study of patients with tuberculosis and their household contacts and compared T cell responses and tuberculin skin test results by infecting genotype. RESULTS: The T cell response to early secreted antigenic target, 6 kDa (ESAT-6), was attenuated in patients with tuberculosis (odds ratio [OR], 0.41 [95% confidence interval {CI}, 0.19-0.89]; P = .024) and household contacts (OR, 0.56 [95% CI, 0.38-0.83]; P = .004) infected with M. africanum, compared with the response in those infected with M. tuberculosis. In these same groups, responses to culture filtrate protein, 10 kDa (CFP-10), were nonsignificantly attenuated (P = .22 and P = .16, respectively), as were tuberculin skin test results (P = .30 and P = .46, respectively). Sequencing of region of difference 1 of M. africanum revealed that Rv3879c is a pseudogene in M. africanum; however, this finding does not provide an obvious mechanism for the attenuated ESAT-6 response. CONCLUSIONS: This is the first evidence, to our knowledge, that strain differences affect interferon- gamma -based T cell responses. Our findings highlight the need to test new diagnostic candidates against different strains of mycobacteria. Integrating additional immunologic and genomic comparisons of M. tuberculosis and M. africanum into further studies may provide fundamental insights into the interactions between humans and mycobacteria.

Large-scale evaluation of enzyme-linked immunospot assay and skin test for diagnosis of Mycobacterium tuberculosis infection against a gradient of exposure in The Gambia.

The purified protein derivative (PPD) skin test for Mycobacterium tuberculosis infection lacks specificity. We assessed 2 more specific M. tuberculosis antigens (ESAT-6 and CFP-10) by enzyme-linked immunospot assay (ELISPOT) compared with PPD by ELISPOT and skin test in The Gambia. Of 735 household contacts of 130 sputum smear-positive tuberculosis cases, 476 (65%) tested positive by PPD ELISPOT, 300 (41%) tested positive by PPD skin test, and 218 (30%) tested positive by ESAT-6/CFP-10 ELISPOT. Only 15 (2%) had positive ESAT-6/CFP-10 results and negative PPD results by ELISPOT. With increasing M. tuberculosis exposure, the percentage of subjects who were PPD skin test positive/ESAT-6/CFP-10 ELISPOT negative increased (P<.001), whereas the percentage of subjects who were PPD skin test negative/PPD ELISPOT positive decreased (P=.011). Eighteen (31%) ESAT-6/CFP-10 ELISPOT-positive subjects in the lowest exposure category had negative PPD skin test results. ESAT-6/CFP-10 ELISPOT probably offers increased specificity in the diagnosis of M. tuberculosis infection in this tropical setting of endemicity, at the cost of some sensitivity.