Middlebrook 合成培地での抗酸菌薬剤感受性試験(第 4 報)：
Nontuberculous Mycobacteria を試験対象とする
微量液体希釈法，BrothMIC NTM の開発評価

山　根　誠　久*1　翁　長　小百合*2　斉　藤　　　宏*3　豊　嶋　俊　光*4
霜　島　正　浩*5　河　原　　　伸*6　高　嶋　哲　也*7　山　下　知　成*8
　　　　　　　　　　　　　　　　　　　　　　
Multicenter Evaluation of a Newly Developed Microdilution Test, BrothMIC NTM
to Determine Minimum Inhibitory Concentrations of Antimicrobial Agents
for Nontuberculous Mycobacteria

Nobuhisa YAMANE, MD*1, Sayuri ONAGA*2, Hiroshi SAITOH*3,
Shunkoh TOYOSHIMA*4, Masahiro SHIMOJIMA*5, Shin KAWAHARA, MD*6,
Tetsuya TAKASHIMA, MD*7 and Tomonari YAMASHITA*8

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 0.1ml 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℃ with clarithromycin at pH7.4, and with the remaining agents at pH6.6. 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 log₂ 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.
[Rinsho Byori 50 : 381〜391, 2002]

*1Department of Laboratory Medicine, Faculty of Medicine, University of the Ryukyus, Nakagami-gun, Okinawa-pref. 903-0215

【Key Words】antimycobacterial susceptibility(抗酸菌薬剤感受性試験)，nontuberculous mycobacteria(非結核性抗酸菌)，broth microdilution test(微量液体希釈法)，minimum inhibitory concentration(最小発育阻止濃度)

受付2001年10月31日・受理2001年12月13日
*1琉球大学医学部臨床検査医学講座，*2同　附属病院検査部(〒903-0215 沖縄県中頭郡西原町字上原207)
*3国立都城病院研究検査科(〒885-0014 都城市祝吉町5033-1)
*4由利組合総合病院中央検査部(〒015-0051 本荘市川口字家後38)
*5 �Mビー・エム・エル(〒350-1101 川越市的場1361-1)
*6国立療養所南岡山病院(〒701-0304 岡山県都窪郡早島町早島4066)
*7大阪府立羽曵野病院(〒583-0872 羽曵野市はびきの3-7-1)
*8 �Mファルコバイオシステムズ(〒613-0036 京都府久世郡久御山町大字田井小字西荒見17-1)

E-mail :nobie@med.u-tyukyu.ac.jp