Mycobacterium abscessus multispacer sequence typing.

BACKGROUND: Mycobacterium abscessus group includes antibiotic-resistant, opportunistic mycobacteria that are responsible for sporadic cases and outbreaks of cutaneous, pulmonary and disseminated infections. However, because of their close genetic relationships, accurate discrimination between the various strains of these mycobacteria remains difficult. In this report, we describe the development of a multispacer sequence typing (MST) analysis for the simultaneous identification and typing of M. abscessus mycobacteria. We also compared MST with the reference multilocus sequence analysis (MLSA) typing method. RESULTS: Based on the M. abscessus CIP104536T genome, eight intergenic spacers were selected, PCR amplified and sequenced in 21 M. abscessus isolates and analysed in 48 available M. abscessus genomes. MST and MLSA grouped 37 M. abscessus organisms into 12 and nine types, respectively; four formerly "M. bolletii" organisms and M. abscessus M139 into three and four types, respectively; and 27 formerly "M. massiliense" organisms grouped into nine and five types, respectively. The Hunter-Gaston index was off 0.912 for MST and of 0.903 for MLSA. The MST-derived tree was similar to that based on MLSA and rpoB gene sequencing and yielded three main clusters comprising each the type strain of the respective M. abscessus sub-species. Two isolates exhibited discordant MLSA- and rpoB gene sequence-derived position, one isolate exhibited discordant MST- and rpoB gene sequence-derived position and one isolate exhibited discordant MST- and MLSA-derived position. MST spacer n°2 sequencing alone allowed for the accurate identification of the different isolates at the sub-species level. CONCLUSIONS: MST is a new sequencing-based approach for both identifying and genotyping M. abscessus mycobacteria that clearly differentiates formerly "M. massiliense" organisms from other M. abscessus subsp. bolletii organisms.

Drug-resistant tuberculosis viewed from bacterial and host genomes.

The outcome of infection with Mycobacterium tuberculosis (MTB) is largely influenced by the host-pathogen interaction in which both the human host and the MTB genetic backgrounds play an important role. Whether this interaction also influences the selection and expansion of drug-resistant MTB strains is the primary focus of this review. We first outline the main and recent findings regarding MTB determinants implicated in the development of drug resistance. Second, we examine data regarding human genetic factors that may play a role in TB drug resistance. We highlight interesting openings for TB research and therapy.

Contribution of the P2X7 1513A/C loss-of-function polymorphism to extrapulmonary tuberculosis susceptibility in Tunisian populations.

The P2X7 receptor has been found to be linked to an increased risk for tuberculosis in some populations. In this study, we investigate whether the P2X7 receptor plays a role in increasing susceptibility to tuberculosis in Tunisia. We examined two 1513A/C and -762T/C polymorphisms at the P2X7 receptor in 168 patients with pulmonary TB (pTB), 55 patients with extrapulmonary TB (epTB) and 150 blood donors from Tunisia. Genotyping of 1513A/C and -762T/C polymorphisms was performed in purified genomic DNA using PCR-restriction fragment length polymorphism and allele-specific PCR, respectively. The 1513C, CC and AC loss-of-function allele and genotypes were overrepresented in the epTB group compared with the control group (45% vs. 17%, P=10(-8) ; 24% vs. 4%, P=3 × 10(-7) ; 42% vs. 27%, P=10(-3) , respectively). Additionally, they were associated with 3.83-, 11.86- and 3.15-fold risks of developing this clinical tuberculosis form, respectively. No associations between the -762T/C polymorphism and tuberculosis disease, as well as disease anatomic location were observed. Collectively, our results suggest that the P2X7 1513A/C loss-of-function polymorphism may contribute to susceptibility to epTB in Tunisian populations.

Interferon gamma +874T/A polymorphism is associated with susceptibility to active pulmonary tuberculosis development in Tunisian patients.

Interferon gamma (IFN-γ) is a key cytokine involved mainly in the defense against intracellular pathogens such as Mycobacterium tuberculosis. Given its key role in the control of tuberculosis (TB), in the present article we have investigated a possible association between IFN-γ gene single-nucleotide polymorphism linked to high and low producer phenotypes (IFN-γ [+874T(high) → A(low)]) (rs2430561) and risk development of active TB in Tunisian patients. Genomic DNA samples were obtained from 223 patients with active TB (168 pulmonary and 55 extrapulmonary cases) and 150 healthy blood donors. Genotypes were analyzed using polymerase chain reaction-restriction fragment length polymorphism method. The +874 AA genotype (low IFN-γ producer) was significantly associated with increased risk of developing of active pulmonary TB (odds ratio [OR] = 2.18; 95% confidence intervals [CI], 1.33-3.57; P corrected for the number of genotypes [Pc] = 0.003). By contrast, the AT genotype was found to be significantly associated with resistance to pulmonary TB (OR = 0.46; 95% CI, 0.28-0.74; Pc = 0.0018) and extrapulmonary TB development (OR = 0.46; 95% CI, 0.23-0.91; Pc = 0.045). Collectively, our data showed that the IFN-γ +874T/A polymorphism is a determinant in the resistance or susceptibility to the development of active TB in the studied population.

Polymorphisms in the RANTES gene increase susceptibility to active tuberculosis in Tunisia.

RANTES plays a pivotal role in attracting and activating various leukocyte populations that control Mycobacterium tuberculosis infection. The present study investigated the relationship between the RANTES polymorphisms (-28C/G; rs2280788, and -403G/A; rs2107538) and susceptibility to active tuberculosis (TB) in Tunisian populations. A total of 168 patients with pulmonary TB (pTB), 55 with extrapulmonary TB (epTB), and 150 control subjects were studied. Genotype analyses were carried out using polymerase chain reaction-restriction fragment length polymorphism method. We found that the -28 GG genotype was significantly associated with susceptibility to pTB (odds ratio [OR]=11.19; 95% confidence intervals [CI], 5.14-25; P corrected for the number of genotypes [Pc]=10(-8)) and epTB (OR=11.67; 95% CI, 4.74-29.33; Pc=10(-8)). However, the -28 CC genotype was found to be significantly associated with resistance to pTB (OR=0.08; 95% CI, 0.04-0.16; Pc=10(-8)) and epTB development (OR=0.11; 95% CI, 0.05-0.27; Pc=10(-8)). -403A allele was associated with increased risk development of epTB (OR=2.21; 95% CI, 1.18-4.14; p=0.007). G-G and A-C haplotypes and the AG/GC diplotype were associated with increase susceptibility to pTB (OR=7.88, 95% CI, 5.38-11.55; Pc=3.10(-8); OR=2.32, 95% CI, 1.32-4.11; Pc=3.10(-3); OR=13.26, 95% CI, 6.06-29.89; Pc=3.10(-8); respectively) and epTB (OR=6.64, 95% CI, 4-11.05; Pc=3.10(-8); OR=2.6, 95% CI, 1.26-5.35; Pc=12.10(-3); OR=11.26, 95% CI, 4.44-29.28; Pc=3.10(-8); respectively). Collectively, our findings suggested an association of the RANTES -28C/G and -403G/A functional polymorphisms with susceptibility to Mycobacterium tuberculosis infection in Tunisian populations.

Evaluation of the diagnostic value of measuring IgG, IgM, and IgA antibodies to mycobacterial A60 antigen in active tuberculosis.

The purpose of the present study was to evaluate the clinical usefulness of detection of serum immunoglobulin A (IgA), IgG, and IgM antibodies raised against the mycobacterial A60 antigen for the diagnosis and discrimination of active tuberculosis (TB) from other pulmonary diseases. Three commercially available ELISA kits (IgA, IgG, and IgM) (ANDA Biologicals, Strasbourg, France) were evaluated simultaneously in 246 serum samples from 3 groups of patients: group I, 171 patients with active TB (128 pulmonary TB and 43 extrapulmonary TB); group II, 73 patients with pulmonary non-TB diseases; and group III, 2 leprosies patients. The sensitivities of tests ranged from 31.3% (IgA) to 94% (IgG) in pulmonary TB patients and from 21% (IgA) to 84% (IgG) in extrapulmonary TB patients. The specificities of assays varied from 92% (IgG) to 96% (IgA) in the pulmonary non-TB group. Combination of IgG with IgA and/or IgM does not improve its sensitivity. Clinical use of the A60-based serodiagnostic IgG assay is of great value for the rapid diagnosis and discrimination between active TB and pulmonary non-TB diseases. Moreover, this test could be used to increase diagnostic accuracy, especially for smear-negative TB cases, which are difficult to diagnose.

Rapid detection of Mycobacterium tuberculosis in sputum by Patho-TB kit in comparison with direct microscopy and culture.

The usefulness of a new rapid diagnostic test (Patho-TB) using antibodies specific to mycobacterial antigens was evaluated for the rapid discrimination between pulmonary tuberculosis (TB) and non-TB pulmonary diseases on sputa. One hundred sputa collected from 79 active TB patients and from 21 patients with non-TB pulmonary diseases (asthma and chronic obstructive pulmonary disease) were enrolled into the study and tested for the presence of Mycobacterium tuberculosis by Ziehl-Neelsen smear, Patho-TB kit, and Löwenstein-Jensen culture. The sensitivity, specificity, positive predictive value, and negative predictive value of the Patho-TB test were 95%, 100%, 100%, and 84%, respectively. Patho-TB test is simple, quick, and easy to perform. Its sensitivity, specificity, and positive predictive value are satisfactory. Therefore, it could be used as a screening test in poorly equipped laboratories of TB endemic areas.