Isolation and identification of mycobacteria responsible for tuberculosis of dromedaries in Algeria

peer reviewe

Performance of the Genotype® MTBDRPlus assay in the diagnosis of tuberculosis and drug resistance in Samara, Russian Federation

<p>Abstract</p> <p>Background</p> <p>Russia is a high tuberculosis (TB) burden country with a high prevalence of multidrug resistant tuberculosis (MDRTB). Molecular assays for detection of MDRTB on clinical specimens are not widely available in Russia.</p> <p>Results</p> <p>We performed an evaluation of the GenoType^®MTBDRplus assay (HAIN Lifescience GmbH, Germany) on a total of 168 sputum specimens from individual patients at a public health laboratory in Central Russia, as a model of a middle income site in a region with high levels of drug resistance. Phenotypic drug resistance tests (DST) were performed on cultures derived from the same sputum specimens using the BACTEC 960 liquid media system.</p> <p>Interpretable GenoType^®MTBDRplus results were obtained for 154(91.7%) specimens with readability rates significantly higher in sputum specimens graded 2+ and 3+ compared to 1+ (RR = 1.17 95%CI 1.04–1.32). The sensitivity and specificity of the assay for the detection of rifampicin (RIF) and isoniazid (INH) resistance and MDR was 96.2%, 97.4%, 97.1% and 90.7%, 83.3%, 88.9% respectively. Mutations in codon 531 of the <it>rpoB </it>gene and codon 315 of the <it>katG </it>gene dominated in RIF and INH resistant strains respectively. Disagreements between phenotypical and molecular tests results (12 samples) could be explained by the presence of rare mutations in strains circulating in Russia and simultaneous presence of resistant and sensitive bacilli in sputum specimens (heteroresistance).</p> <p>Conclusion</p> <p>High sensitivity, short turnaround times and the potential for screening large numbers of specimens rapidly, make the GenoType^®MTBDRplus assay suitable as a first-line screening assay for drug resistant TB.</p

High-Resolution Melting Analysis for the Rapid Detection of Fluoroquinolone and Streptomycin Resistance in Mycobacterium tuberculosis

published_or_final_versio

New Insights into Fluoroquinolone Resistance in Mycobacterium tuberculosis: Functional Genetic Analysis of gyrA and gyrB Mutations

Fluoroquinolone antibiotics are among the most potent second-line drugs used for treatment of multidrug-resistant tuberculosis (MDR TB), and resistance to this class of antibiotics is one criterion for defining extensively drug resistant tuberculosis (XDR TB). Fluoroquinolone resistance in Mycobacterium tuberculosis has been associated with modification of the quinolone resistance determining region (QRDR) of gyrA. Recent studies suggest that amino acid substitutions in gyrB may also play a crucial role in resistance, but functional genetic studies of these mutations in M. tuberculosis are lacking. In this study, we examined twenty six mutations in gyrase genes gyrA (seven) and gyrB (nineteen) to determine the clinical relevance and role of these mutations in fluoroquinolone resistance. Transductants or clinical isolates harboring T80A, T80A+A90G, A90G, G247S and A384V gyrA mutations were susceptible to all fluoroquinolones tested. The A74S mutation conferred low-level resistance to moxifloxacin but susceptibility to ciprofloxacin, levofloxacin and ofloxacin, and the A74S+D94G double mutation conferred cross resistance to all the fluoroquinolones tested. Functional genetic analysis and structural modeling of gyrB suggest that M330I, V340L, R485C, D500A, D533A, A543T, A543V and T546M mutations are not sufficient to confer resistance as determined by agar proportion. Only three mutations, N538D, E540V and R485C+T539N, conferred resistance to all four fluoroquinolones in at least one genetic background. The D500H and D500N mutations conferred resistance only to levofloxacin and ofloxacin while N538K and E540D consistently conferred resistance to moxifloxacin only. Transductants and clinical isolates harboring T539N, T539P or N538T+T546M mutations exhibited low-level resistance to moxifloxacin only but not consistently. These findings indicate that certain mutations in gyrB confer fluoroquinolone resistance, but the level and pattern of resistance varies among the different mutations. The results from this study provide support for the inclusion of the QRDR of gyrB in molecular assays used to detect fluoroquinolone resistance in M. tuberculosis

Lateral flow assay for rapid differentiation of Mycobacterium tuberculosis complex and 97 species of mycobacteria other than tuberculosis grown in Löwenstein-Jensen and TK-SLC medium

Background: Mycobacterial antigen MPB64 is a secretory protein specific for Mycobacterium tuberculosis complex. A lateral flow immunochromatographic assay (ICA) is a method used for the rapid differentiation of M. tuberculosis complex. Aim: We aimed to evaluate the performance of ICA in rapid differentiation of M. tuberculosis complex from 97 Mycobacterium species other than tuberculosis (MOTT), which are grown in Lφwenstein-Jensen and TK-selective (SLC) medium. Materials and Methods: The study was performed in our laboratory between January 2009 and January 2010. A total of 394 isolates consisting of reference strains of 34 M. tuberculosis from World Health Organization (WHO) collection, 97 different MOTT bacilli, 7 Mycobacterium bovis BCG substrains and total 256 clinical Mycobacterium isolates were tested by ICA, which is based on anti-MPB64 monoclonal antibodies. All the strains were inoculated onto a TK-SLC (selective) medium and Lφwenstein-Jensen medium. TK-SLC is a new rapid mycobacterial culture medium that indicates mycobacterial growth by colour change. Results: The growth of mycobacterial strains was observed in 10-12 days on TK-SLC medium. ICA test was performed in 15 minutes. All strains belonging to M. tuberculosis complex group were found positive and all MOTT species were found negative on ICA slides. The results were confirmed with nucleic acid amplification by polymerase chain reaction (PCR) using primers specific for M. tuberculosis complex. Conclusion: With the additive effect of growth on TK-SLC medium in 10-12 days, the mycobacterial antigen MPB64 is a very useful and specific tool in rapid differentiation of M. tuberculosis and MOTT grown in culture

Dio-sensimedia: a novel culture medium for rapid detection of extended spectrum beta-lactamases

Background: Resistance to contemporary broad-spectrum beta-lactams, mediated by extended-spectrum beta-lactamases (ESBL), is an increasing problem worldwide. Many of the emerging antimicrobial resistance problems of this decade have been characterized by difficulty in the recognition of resistance in the laboratory, particularly by rapid susceptibility test methods. The plasmid-encoded ESBL represent such a resistance phenomenon that is difficult to recognize. We compared Dio-Sensimedia-ES (DSM-ES; Diomed, Istanbul, Turkey) and Mueller-Hinton (MH) agar in the double-disk synergy test (DDST) as a novel rapid system for detecting ESBL directly from bacterial culture