Multicenter evaluation of a newly developed microdilution test, brothMIC NTM to determine minimum inhibitory concentrations of antimicrobial agents for nontuberculous mycobacteria
Yamane, N.; Onaga, S.; Saitoh, H.; Toyoshima, S.; Shimojima, M.; Kawahara, S.; Takashima, T.; Yamashita, T.
Rinsho Byori. Japanese Journal of Clinical Pathology 50(4): 381-391
2002
ISSN/ISBN: 0047-1860 PMID: 12014018 Document Number: 549351
We developed a new broth microdilution susceptibility test for nontuberculous mycobacteria to determine minimum inhibitory concentrations(MICs). The test method utilized air-dried microplates containing serially diluted antimicrobial agents and the modified Middlebrook 7H9 broth. The nine agents tested were rifampicin, isoniazid, ethambutol, streptomycin, kanamycin, levofloxacin, clarithromycin, ethionamide and amikacin. The test plates were reconstituted by inoculation of 0.1 ml of cell suspensions prepared in distilled water by a 1:100 dilution of the 0.5 McFarland suspension. After inoculation, the isolates resulted in incubation at 36 degrees C with clarithromycin at pH 7.4, and with the remaining agents at pH 6.6. The growth endpoints were visually read after 7-day and 10-day incubations for slowly growing nontuberculous mycobacteria, and after 3-day and 5-day incubations for rapidly growing mycobacteria, respectively. The reproducibility was evaluated with the five ATCC reference strains of nontuberculous mycobacteria. Of the 1,287 repeated tests of the four ATCC slowly growing strains(M. avium, M. intracellulare, M. kansasii and M. gordonae), 1,200(93.2%) of the MICs read after 7-day incubation fell within 3 log 2 dilutions. Also, a strain of rapidly growing mycobacteria, M. fortuitum, was repeatedly tested, and the reproducibility was estimated to be 93.3% after 3-day incubation. A total of 728 clinical isolates of nontuberculous mycobacteria comprising 14 species were tested against nine agents. The MICs against nontuberculous mycobacteria distributed in a wide range, and the activities of rifampicin, levofloxacin, clarithromycin were more potent. These results demonstrate this newly developed test method to be a practical, rapid, quantitative means to determine MICs for nontuberculous mycobacteria in clinical laboratories.