rifampin and diacetylfluorescein

Topics: Drug Resistance, Bacterial; Fluoresceins; Humans; Mali; Molecular Diagnostic Techniques; Mycobacterium tuberculosis; Rifampin; Sensitivity and Specificity; Sputum; Tuberculosis, Multidrug-Resistant

Damien Foundation Project, Bangladesh.. To evaluate sputum smear fluorescein diacetate (FDA) vital staining to predict culture-defined failure and rifampicin (RMP) resistance.. A retrospective, operational study.. A total of 1633 episodes of auramine smear-defined late conversion and failure could be evaluated (respectively 640 and 584 on first treatment and 185 and 224 on retreatment). Negative FDA was 95% predictive of negative culture in patients on first treatment, while its positive predictive value was around 95% during retreatment. The predictive value of a positive (not scanty) result for RMP resistance or environmental non-tuberculous mycobacteria (NTM) was at least 90%, except in late converters on first-line treatment; a negative result was over 95% exclusive of the same except in retreatment failures. FDA correctly identified 88-98% of all RMP resistance.. FDA staining increased the proportion of tuberculosis patients put on second-line treatment without receiving the standard first-line retreatment regimen. In our setting, with excellent microscopy, late case presentation and low resistance prevalence, it proved indispensable for efficient culture and referrals of early suspects for rapid drug susceptibility testing (DST). In other settings with low prevalence of NTM and difficult access to accurate and rapid DST, FDA-positive failures might even be considered for immediate start of second-line treatment.

Topics: Antibiotics, Antitubercular; Bangladesh; Drug Resistance, Multiple, Bacterial; Early Diagnosis; Fluoresceins; Fluorescent Dyes; Humans; Microbial Sensitivity Tests; Microscopy, Fluorescence; Mycobacterium tuberculosis; Predictive Value of Tests; Retrospective Studies; Rifampin; Sensitivity and Specificity; Sputum; Treatment Failure; Tuberculosis, Multidrug-Resistant; Tuberculosis, Pulmonary

Susceptibility testing of Mycobacterium tuberculosis is seriously limited by the time required to obtain results. We show that susceptibility testing of clinical isolates of M. tuberculosis can be accomplished rapidly with acceptable accuracy by using flow cytometry. The susceptibilities of 35 clinical isolates of M. tuberculosis to various concentrations of isoniazid, rifampin, and ethambutol were tested by the agar proportion method and by flow cytometry. Agreement between the results from the two methods was 95, 92, and 83% for isoniazid, ethambutol, and rifampin, respectively. Only 11 discrepancies were detected among 155 total tests. The results of flow cytometric susceptibility tests were available within 24 h of inoculation of drug-containing medium, while the proportion method required 3 weeks to complete. The flow cytometric method is also simple to perform.

Topics: Agar; Antibiotics, Antitubercular; Antitubercular Agents; Ethambutol; Flow Cytometry; Fluoresceins; Humans; Isoniazid; Microbial Sensitivity Tests; Mycobacterium tuberculosis; Reproducibility of Results; Rifampin

The resurgence of tuberculosis has caused considerable effort to be focused on the development of rapid methods for determining the susceptibility of Mycobacterium tuberculosis to antimycobacterial agents. We demonstrated that susceptibility testing of M. tuberculosis can be accomplished rapidly by using flow cytometry. Results of tests were available within 24 h after M. tuberculosis organisms were incubated with ethambutol, isoniazid, rifampin, or streptomycin. The method was based on the ability of viable M. tuberculosis organisms to hydrolyze fluorescein diacetate (FDA) and the detection of fluorescent mycobacteria by flow cytometric analysis. The assay system also did not require multiplication of the mycobacteria. In contrast, M. tuberculosis organisms exposed to antimycobacterial agents hydrolyzed significantly less FDA. The use of flow cytometry and FDA staining shows considerable promise as a rapid method for obtaining susceptibility test results.

Topics: Antitubercular Agents; Drug Resistance, Microbial; Ethambutol; Evaluation Studies as Topic; Flow Cytometry; Fluoresceins; Formaldehyde; Humans; Hydrolysis; Isoniazid; Microbial Sensitivity Tests; Mycobacterium tuberculosis; Rifampin; Streptomycin; Time Factors

The susceptibilities of Mycobacterium leprae and M. avium complex (MAC) to the H2-O2-Fe-mediated halogenation system supplemented with antimicrobial agents were evaluated by fluorescein diacetate-ethidium bromide (FDA/EB) staining. In the case of M. leprae, the number of greenstained bacteria (intact cells) was reduced in the presence of the H2O2-Fe-mediated halogenation system supplemented with agents possessing antileprosy activity, such as rifampin, 4,4'-diaminodiphenylsulfone (dapsone), clofazimine, and ofloxacin. In the case of the MAC strain, although viable units of the organisms were reduced by the halogenation system alone, the number of greenstained cells in the FDA/EB stain was not reduced, even when the halogenation system was used in combination with ofloxacin. Because stainability of the cells is related to structural and functional intactness of the membrane, differences between M. leprae and the MAC strain imply possible differences in the rigidity of the cell membrane.

Topics: Animals; Clofazimine; Dapsone; Ethidium; Ferrous Compounds; Fluoresceins; Humans; Hydrogen Peroxide; Iodides; Mice; Mycobacterium avium Complex; Mycobacterium leprae; Ofloxacin; Rifampin; Sodium Iodide; Staining and Labeling