164 ῌ῎ ῏῍ ATP ῳQ̮̯ῺQ ῒQQQῩ̳Q̳QῒQQῩQ΅ ̳̮QQ1)῎̮ Q1)῎ ̮῍Q1)῎Q῭ῨQ2)῎´ῠ 2)῎ῖQ̮Q1) 1) ̯Q̯ ῶ ̲̲῟̲ῺQQQQ̲Ώ῾QQQ 2) QῪ̳QQ̳Q̲QQQQQQῶ̲ ΐ 16 ̱ 4 Q 8 ̱Q̲῍ ΐ 16 ̱ 9 Q 7 ̱QQ QῚQ̯ῲῨ Q̯` adenosine triphosphate (ATP) ῳ῾ῶQῤ῞̰̯`῞ῥῨῬ΅ Nontuberculous Mycobacteria (NTM) Ῡ̳Q̳QῒQQῳQῖῌ̯ῧ ATP ῳῶ ̯῍ ̯ῩQΊ̰Q΅ῧ῭̰ῨΏQΏ̯QQ MB ῥῩΌ Q΅ῳ ATCC ῼQ̳ 4 ̳ῥQ QQ̳ 45 ̳ῳQ̮ΰQῡ̯ῌ ATP ῞̰ Ῠῢ̮ΰ̰Q΅ῩQ̲῍ ῙQQ̰Q̲Ῠΰ῍ 30 ̳QQ῍ ῥQQQ̰ 60̮ Ῠΰ῍ 10 ̳Ὶ̰Ύῤ̮`῞ῥῚ ̱̯̯ῌ ATP ̰̱̱ῐ̱ ῳ 5 ̱̳ῴQQῨ Relative light units (RLU) ratio ῳ῞̰̯῍ 7 ̱̳ῴQῩ MB ῥΌ Q΅ ̯̯ῌ streptomycin, ethambutol, kanamycin, isoniazid, levofloxacin, ethionamide, amikacin Ῡ 7 ̳QῩῶῷQῒῢῳ RLU ratio̮0.1, rifampicin ῘῬ̰ clarithromycin ῳ RLU ratio ̮0.03 ῥ̯̯ ATP ῥ῍ MB ῥῩῸΰQ̰ Mycobacterium kansasii ATCC 12478 ̰ 84.4̮ ῥΎΎ̰̮ῢῤ̮ῡ̯ῪῩῩ῍ ̯ῩῼQ̳ 3 ̳Ῠῢ̮ΰ̰ 97.8ῑ100̮ 4 ̳ῤ̰ 95̮ ῥQQῧQ̲Ὶ`῭̯ῌ ̰̯QQQ̳Ῠῢ̮ΰῪ Mycobacterium intracellulare: 92.2̮, Mycobacterium avium: 97.2̮, M. kansasii: 100̮ ῤ῍ Ὶ̳ῤ̰ 95.3̮ ῥQQῧῸΰQ ῳQ̯̯ῌ ̱QῬ΅ ATP ̰ NTM Ῡ̳Q̳QῒQQῥ̯ΰQQῧ ῤ̮`ῥ´̲̯̯ῌ Key words: nontuberculous mycobacteria (NTM), adenosine triphosphate (ATP), minimum inhibitory concentration (MIC), bioluminescence ῲῚQῩῺQ ῒQQQ (nontuberculous myco- QQQQ̯Q̱Ό̲̰ 2003 ̱ 4 QῨ ῒ̱ῺQ ῒQ bacteria; NTM) QῩ̳ 70̮ ̰ Mycobacterium QQQ῎ῡῨ̳̯`Q῾ῌ2003 ̱ΐ ῳ̰Q̯ῌ̯ῧ avium complex (MAC) ῨῬ`ῪῩῤ̯̰̱Ῠ̳ῒ `῎ῡQQῳQΐ̯̯1)ῌ MAC Q̰QQῩ̯̰ΰῚ̳ ̲ ῳQΐ̯`̱ MAC Qῤ̮`ῌ ῞̰ῤ MAC Q QQQῩ̯Qῥῧ`ῲ῝ῤ̰ῧ̮ῥῩQῗῚῲῚQῤ ̰ῩQῒ̱Q ῍ Q̳Q ῧQ῍ ̱ῘQ῍ ῐ̱῍ Q ̰ῸῸ̰ῤ̮`Ὶ῍ ̳Qῳ̮Q̰ῨQQ̯Ῥῖῥ̯` Qῒ̲ ῧῦῩ̱ῨQQ̰Q ῳQ̯`̳QῨ 2 Q QQῪ̯ῠ̮ῨQῧῡΰ̮`ῌ ̯Ῑ̯ῧῚ῭ MAC ̰Ῠ̳ῗ̯̯ῥQῗ῭`QQῚ̯Ῑῡ̯Ὶ῍ QQῤ ̰Q Ῠ̯Ῥ̯Ῑ῭ῧ` ̳ῳQ̯῍ QQ ̳ΎQ ̰QΊQ̳ῩῨῧ̮ῤQ̱ῩQῒῨῶQ̯῍ ̯ῶῒ Q̳Ῠ̯̯ΰ̰̳QῒῳQ̯̯̰῍ ̳QῨῬ`̱̳ῗ ῩQQ̮ΎQ̳Q ῧῳQ̯QQῚQ῝̯ΰ̮`ῌ QΎ AIDS ̳QῤῩῚ῏̱Qῒ̳ῗQ̰Q̰ΰῳQ MAC QῩ῎ῡ̰Q QῩ῎ῡῥ̰ῴῧ΅QῩQQῩ ῒῤ̮`ῌ ̰̯ MAC ῨQ̮ῤ̯QQ̯` Myco- ̰ῤ̰̲Qῤ̮`ῌ ῞Ῡ̯̰̱̳Q ̲ ̲Ὼ̰Q bacterium kansasii Ὺ 1997 ̱῟Ῥ΅Qῦῧ῝̲Q QῨ̮΅῍ MAC QῨΌ̰ΎΎQ̱Ῡ̳QῚ̯̮ῥ ῦQQQ̮῏ (̮565῍0871) ̯Q̲ῑ̰QQ̰Q 2῍15 ̯Q̯ ῶ ̲̲῟̲ῺQQQQ̲Ώ῾ QQQ ̳̮QQ TEL: 06῍6879῍6680 FAX: 06῍6879῍6683 E-mail: [email protected]. ac.jp 36 ̱̳QQΏ῾ QQ Vol. 14 No. 3 2004. ̯ΰ̮`ῌ M. kansasii ̰Ό ̰QῚQ῍ QΊ Q̳Ῡῧ̮QQ̱ῨῪQ QQῩ̲ ῳQ΅Ύ̯̮ Ὶ῍ rifampicin (RFP) ῳῤQῥ̯`̲ Q Ῠ̯̯ΰ QQῧῷῼῳQ̯῞ῥῙ῭ RFP ῳ̰̯̰ῥ̯` Q QQ̳Ῡ̳QῒQQῳQQ̯`῞ῥῚQQῥ̯ΰ̮ `ῌ QQ῍ NTM QῨ̯̯QQQ̲ῚQ̯ῤΊ`QQ ̳ῥ̯ΰ clarithromycin (CAM)῍ azithromycin ῧῦ ATP ῳΎΌ̮̯̰QQQQQQῤ̱QQQQQQῤQ̯ ῤΌ̲ῐ̲ ̲῭ῸQQQ̱̰῍ Ό̲ῐῺ Q 165 ῝Ί̯ῌ ATP ῚQῳ QQ̱῍ Ῥ̲Ὼῲ῾ῳ̱Ὸ῍ RFP ῢῡῤ̱QῗQΌῚ C. Ὺ̯̮̰̯Ῡ῍ ῾Qῖ̰ῥῙῪ̰ῤ̱Qΰ̮῝̰Q Q̮ῖ̰ῤ ATP ῚQῤQ̯ΰ̯̮̯ῥQ̳ῥ̯Ύ ῢ̱QQQQQQῳῤQ̰ῗ̱ῨῪ̯̮̰ῌ McFarland No. 0.5 ΰΊQΊ̯Q̳ΎΌ̮῍ ῚQQ̱ ̰Q῭QQῤQ̳ῤ̯ῩΰQ̰ῚῪ̯ῲῳῤ̳̯ (benzalkonium chloride) Ύ 50 ml ῟QΊ῍ ῚQ̳̯ ΰ̰Q῭ῤ`῝̰ adenosine triphosphate (ATP) ῥQ̳῍ 60, 70, 80 ̯῍ Ὶ̰ΰῚQQQῥ 0, 5, ̲Ύΐ῝ΊQQQΎΐ῝῝̰ῲῳῗ̮̰ῌ Ῑῤῲῳῥ 10, 15, 20 ΅ Q ̯ Q ῼ Ί ̯ Q ΰ Q ̳ ΰ ̱ Ί ̯ ῖ ̰ QQQῠ ATP ̲ῤQΰQῩ̯̲QῢῐQQQῗ̮ ATP ΐ῝ΎQ̯̯ῌ ̰῍ Ὶ̰ΰ῍ Ῑῤ̰QQ̲ῗQ̮̯ῩQῩ̯QQ̯̯ D. ATP ΐ῝ῳ ΐ῝Qῦῢῲῳ̯̮̰ῌ Ῑῤ ATP ̲ΐ῝ΰ̰̰̱Q ATP ΐ῝ῳῥQQΰQΊ̯ ATP ῎Qῼῢ̰ῦΰ QQQQQΎQ̮ῲῳῥ῍ ̳̰QQΰ̯̮̯ῥQ̯2)῍ ATP ῚQΎQ̯̯Qΰ῍ ̲῭ ̲ ῐῷΎῒΊ ̯3) ̰ῤQ῍ Ῠ̯QQQΰ̯̮̯ῥQQ4)ῑ6) ̰ΰ Q̳ΰ̱Ί̯ῖ̰῍ ̲῭ ̲ ῐῷ῾ῐ̱ῐ (LP- ̰̰Qῗ̮̰ῗ῍ NTM ΰῙῤῲῳΎΌ̮̯Qῥ 5000-kyokuto-1) ΰ̯Q̯῞ΰ̰QQ̱ ῒ̱ῼ`῾ Q̰Ὺῢ̮ῌ ̯Ῑ̯῍ Q῏ῥ ATP ΐ῝ΎΌ̮̯̱Q ῎̱ῼ`̲ῐ̱ΐ ΎQ΅`ΰ 100 ml ῟Q῝̰ QQQQQῳ ῒ Q ATP ῳῠ̲῝ΐ ῤQ῏Q̯ΎQ ῠῠ̰ΰ̰Q̲ (Relative light units; RLU) Ύΐ῝Ί ̮῍ QΏῳῤ̰̲̳̮QQῳ ῒ Q MB ῳῠ̲῝ΐ ῠ ̯ῌ ῤ̰QQ̯ΎQ̯̯ῌ I . ῍ῐῌ῎῏ E. ̱QQQQQQ 1. ̰̲̳̮QQῳ̯ῤ MIC ΐ῝ ῒMB ῳΐ MIC ΐ῝ΰῥ streptomycin (SM), ethambutol A. QQQQ (EB), kanamycin (KM), isoniazid (INH), RFP, levo- QQQῠΊ̯ῥ American Type Culture Collec- floxacin (LVFX), CAM, ethionamide (TH), amikacin tion (ATCC) ̰QQῠΊ̯ M. avium ATCC 700898, (AMK) ῤ 9 ̱QΎ 2 Ῠῥ̯QQQ ῒSM, EB, KM M. avium ATCC 25291, M. kansasii ATCC 12478 ̮ ῥ 0.06ῑ128 mg/ml ῤ 12 ̮Q῍ INH, RFP, ̰ῦ Mycobacterium intracellulare ATCC 13950 ῤ 4 CAM ῥ 0.03ῑ32 mg/ml ῤ 11 ̮Q῍ TH, AMK ῥ QΎΌ̮̯ῌ Ῠ̯῍ ῾Q΅´QῠΊ̯ῥ̮Q̮Q Q 0.5ῑ16 mg/ml ῤ 6 ̮Qΐ ΰΊQΊ̯ MIC ΐ῝Ό̲ ῬΎῪ̳ΰ̯QQῤQQQῶΏQ̲ῖ̰΅´῎̯῝ ῐῷ̯̮̰´ ̱̲ NTM ῒQ̯Q̱QQΐ Ύ ῚῪ̯ 45 QΎQQΊ̯ῌ ̯ῤῠῴῥ῍ M. avium 20 Ό̮̯ῌ LVFX, Q῍ M. intracellulare 20 Q̮̰ῦ M. kansasii 5 Q̯ ῌQQ̳ῤΊQῲῳῥ῍ 1̮ Q῍Ῡῖ̯ῩῺΊ̯ 4 ̮̯̯ῌ ῢ̮῍ ῾Q΅´Qῤ̯῝ῥ DDH (DNAῌDNA Q Q ῠ ῤ ̰ Q Q ῢ ̰ ῦ ΰ ῾ Q ΅ ´ Q Ύ Middle- hybridization) ̲῭ῳΏῶ῾Ῥ ῒQ̯Q̱QQΐ ΰ brook 7H9 broth ῩῖΰQ̳ῤ̮̯Ύ῍ ῧ 530 nm, ̯Q̯̯ῌ QQ̯ 0.03ῑ0.05 ῤQQΰῌQQ῍ 37 ΰ̯ 3ῑ5 B. ATP ῎Qῼῳ ῢQῩῺΊ McFarland No. 0.5 ῒQQ̯ 0.08ῑ0.10ΐ ῎QῼῤQ̯̯ῥ ATCC ̰QQΎ Middlebrook QΰῑQΊ̯ῩῺ̳ΎΌ̮̯ῌ QΰῙῤῩῺ̳Ύ̱ 7H9 broth Ῡῖ ῒ̲῭ῳ´ ̱῍ Q̯Q̱QQΐ ̯῎ QQ̲Qΰ̯ McFarland No. 0.5 ΰQQΰQQΊQ ῩῺQ῍ Q̳ῥ̯Ύ McFarland No. 0.5 ῒῧ 530 Ί̯ῌ Ὶ̰ΰ῍ ΊQΊ̯Q̳ 110 ml Ύ̱QQ̲Q 11 nm, QQ̯ 0.08ῑ0.10ΐ ΰΊQΊ̯Qΰ῍ Middle- ml ΰQ̮ῴ̲ῐ´̲̱ῐ̯Q Q῍ ´ ̱̲ brook 7H9 broth Ῡῖ̯ 1/2, 1/10, 1/100 ΰQQΊ NTM ῤQ΅`ΰ 100 ml ῞̯ῌQΊ῍ ̱ῐΎΊ̯ ̯Q̳ΎΌ̮̯ῌ ῎Qῼῥ῎Qῼ̳ A (2-amino-2- ̲ ῐῷΎ 371 ΰ̯ 7 ῢQῩῺQΰ῍ ῡQ῞ΰ methyl-1,3-propanediol) ῠ῎Qῼ̳ B (adenosine Q̰ ῗῤῩ̰Ὺῢ̮QQ̱Qῥ̯Ύ MIC ̮ῠΊ̯ῌ ATP ΐ῝ΰ̰̰̱QQQQQQῳ ῒATP ῳΐ phosphate deaminase) ΎQΌQΰQ Ί῍ Q̳ῤ 2. ΰ̯̯΅`ΰ῎Qῼ̳ 50 ml Ύ῟QΊ῍ Q̳̯ 0, ATP ΐ῝Όῤ̱QQῘ̲ ῐῷῥ MIC ΐ῝Ό̲ 30, 60, 90, 120 ΅QῗΊ̯Qΰ ATP ΐ῝ΎQ̮῍ ῐῷῤQQ̱Qῠ̯ῥ̯ῠῢ̰̰̮ΰΊQΊ̯ῌ ῩῖῙῤῷ´ῤ ATP ̰̰Qΰ̳QΎQῧ῝Q΅ΎQ ῢ̮῍ ̲ ῐῷῥ ATP ΎQ῭̰Q̯ΐ῝῝̰̯Ῡΰ QῚ῟̰Ῑῠῗ̯Ῐ̰ῖῤQ̯ΎQ̯̯ῌ ῢ̮῍ MIC ̲ ῭ῷ̲ ῐῷῠΊ̯ῌ ῌQQ̳ῤΊQῲῳῢ̰ῦ ΐ῝ΌῤῩῖ ῒ´ ̱̲ NTM ´ ̱῍ Q̯Q̱ ΰῌQῲῳῥ MB ῳῠ̯ῸΰQ̯̯ῌ QQΐ ῤ῎QῼQQῤQ̯ΰ̯̮̯ῥ 0῍ 30 ΅Q̯ΐ ATP ῤΐ῝ΰ̰̰ MIC ΐ῝ῥ 5 ῢQῩῺQΰQ ῢ̱῾Q̰Q῭QQQ Vol. 14 No. 3 2004. 37 QQQQ῎Q 166 Q῎QQQQ῎QQQQ῎ῶQ Q῎Q῍QQ QΊ῝ῌ QQQQῠ̳Q ΌῨ̰῍ QQ῎Q̲ῠQῷ QQQ̲̯῟̮̯ῠ 3 ̲̯ῧ̯ 10 Q̲̱Q̯QQ̯ ̯ 30 Q̲̯Ί῍ QQQQῠ 60̮, 10 Q̲ῸῷΊ῝ῌ ΰῤQQ̯̯̰Ῑ̯ῖῩ῍ ATP QQQ̳ῠ 60̮, 10 QQ̳῍ Qῷ̯QΊ῝̳̯ ATP QQ̰Q῞῝ῌ QQ Q̲̯Ί῝ῌ ῠ῍ RLU ratio ῒQQ̲QQῌ̰Ὺ`̯ RLU Q̰QQ B. Q̲ΎῲῬῐ`̰Ὺ`̯ RLU Q̯QΊ῝Qΐ ̰ 1. QQ̲QQQ̳ ATCC QQ ̲̯̯ MB Q̯ ATP Q̯̱Q ῏ ῦ RLU ratio0.1, 0.03 ̯Q῟̯΅̰ῲῬ̯ Q̱ ̯QQ̰Q῞῝ῌ ̯̮῍ ΎῲῬῐ`̰Ὺ`̯ RLU ῭ῐῬ̰ 7 Q̲Qῌ̳῍ MIC QQ̰Q῞῝ MB Qῠ CAM ̰QῘ 8 QQ̯῟̮̯ῠQQQ̲ΎῲῬ Q̯ Ό̯῍ 5 Q̲Qῌ̳̯ ATP QQ̰Q̮῍ RLU ῐ`̰Ὺ` ῒ3 ̰Ὺ`ΐ ̯Q RLU Q̯Ί῍ CAM ratio ̯̯QQ̰Q῞῝ ATP Q̯̯ Ό̯̱Q῏̰῍ ̯῟̮̯ῠQ̳QQ pH ῗQQ̯̱̯̰῝ῦ CAM Q QQῌ̯QΏῖῩῼ῏5 ῺQQQ̳̰Q̮̳QΊ῝ῌ ΎῲῬῐ`̰Ὺ`̯ RLU Q̯Ί῝ῌ ̯̮῍ ̱Q῏ῠ1 ̲̱Qῠ QQ̱̯Ί῝ῌ Table 3 ̯῍ MB Q̰῾Q̯Ί̯ ATP Q̯ RLU ratio0.1 ̮ II. ῍ Ῠῡ RLU ratio0.03 ̰QQ̯Ί῝QQ̯̱Q῏̰ ῌ Q Ί ῝ῌ RLU ratio 0.1 ̰ Q Q ̯ Ί ῝ Q Q῍ M. A. ATP QQQQ̯̳Q Table 1 ̯῍ QQ῎Q̲̯̳Q Ό̰QΊ῝ῌ avium ATCC 700898 ̯῟̮̯ῠ INH ̯ 1 Ὼ 2 ̲Q QQQ̯ QQ̯̮̰ McFarland No. 0.5 ̯ 1/ Ῐῖ῞῝ῧ̯̯῍ QQ̯῟̮̯ῠῢ̯ 1 ̲̱Q̯ 100 ̯ ῑ̯ῠQ̳ῑῠQῘ῍ RLU Qῠ 302ῑ506 ̱Q῏ῠ 97.8̮ ̯̮῞῝ῌ ῥ῝ M. intracellulare ̯̮̰QQQ̯ῴ ̰ ῤQ̳ῑ̯ῠ̯̮Ῑ̯῍ ATCC 13950 ̯῟̮̯ῧQQQ 1 ̲̱Q̯ῐQ̯ QQ̯ QQ̯̮̰ 1/10, 1/2 ̯ῠ 30ῑ60 Q̲ ̱Q῏̰QΊ῝ῌ M. avium ATCC 25291 ̯῟̮̯ῠ Q῎̯ῳQΊ῝ RLU QῗQῩ̰̰Ῑ̯ῗ´QῚ̰ ATP Q̯̯Qῗ RFP, CAM ̯ 2 ̲̱Q̯Q̮Q̰Q ῝ῌ ῥ῝QQ῎̯Ῠ῞̯QQQ̯ ATP QQQ̯Q̳ Ί῝ῗ῍ Q 7 QQ̯῟̮̯ῠ 1 ̲̱Q̯̮῞῝ῌ M. ̯ῴ ̰Q̮̰QQ̯ RLU Qῠ῍ 72,074 ̯̮῞῝̯ kansasii ATCC 12478 ̯῟̮̯ῠ KM, AMK ̯ EB ῗ 30 Q̲̯ 384 ̯QΌQ̯Q῎Ὶ̰̯̮̰Ῑ̯ῗ´ ̯Q̮Q̰Q Q̯̮̰ 84.4̮ ̯̱Q῏̯̮῞῝ῌ QῚ̰῝ῌ Table 2 ̯ῠ῍ QQQ̳̯῟̮̯QΊ῝ῌ RLU ratio0.03 ̯Ί῝QQῠ῍ M. avium ATCC M. avium, M. intracellulare ̯῟̮̯ῠQῷ̰Q̮̯ 700898 ̯῟̮̯ῠ EB ̯ LVFX ̯Q̮Q̰QΊ 60ῑ80̮ ̯ῠQ̳ῑ̯Qῠ̳Ῡ̰̯ῖ῞῝ῗ῍ M. 88.9̮ ̯̱Q῏̯̮῞῝ῌ M. avium ATCC 25291 ̯ kansasii ̯῟̮̯ῠῷQῗQῘ̯̰̯ RLU QῗQῘ ῟̮̯ῠ῍ RLU ratio0.1 ̯Q̮Q̯̮῞῝ RFP, ̯̰ Q̯̮̰῍ 60̮ ̯QῧQ̮ RLU Q̰QΊ῝ῌ CAM ῗQ̮̱Q῏̰QΊ῝ῌ ΊῖΊ KM ̯ AMK ῗ Table 1. E#ect of treatment time on the exogenous ATP. Treatment time (min) Dilution of organisms suspension 0 30 60 90 120 M. avium ATCC 25291 blank 1/100 1/ 10 1/ 2 95 265 2636 13052 48 329 3981 26015 41 302 4129 25566 49 292 3779 26225 76 345 4412 30611 M. intracellulare ATCC 13950 blank 1/100 1/ 10 1/ 2 165 329 1774 15867 51 506 3987 40746 54 499 4462 44716 46 476 4133 54015 114 461 4313 57430 M. kansasii ATCC 12478 blank 1/100 1/ 10 1/ 2 155 306 1284 18574 50 414 3176 22077 43 506 2720 24503 41 399 3219 24090 117 377 3382 23905 BrothMIC NTM blank 72074 384 NT NT NT Strain NT: not tested 38 QQῒQQQQ̲QQ Vol. 14 No. 3 2004. ATP QῤQΐ̮QῪῩ̲̱̰῟Q̱̰ῷ̲ῴ῭῟Ύ Table 2. 167 E#ect of the temperature and incubation time on the extraction of ATP from nontuberculous mycobacteria. Extraction time (min) Strain Temperature 0 5 10 15 20 M. avium ATCC 25291 room temperature 60ῐ 70ῐ 80ῐ 5643 5643 5643 5643 7909 26090 24440 26967 8949 26223 23807 26795 9178 23592 28471 25016 9483 30892 25906 24740 M. intracellulare ATCC 13950 room temperature 60ῐ 70ῐ 80ῐ 972 972 972 972 7359 15368 16982 18823 7467 17435 18784 18386 7323 16998 17432 17160 7773 17525 18888 18246 M. kansasii ATCC 12478 room temperature 60ῐ 70ῐ 80ῐ 661 661 661 661 1125 8756 6249 5945 1321 8608 7178 5945 1245 8730 5938 5441 1211 9305 6579 5472 2 ̰ῥ̲῟ῳ ῤῶ̮̮Ίῠῦ Q̯ ῗ̮ῌ M. ῾ 8 Q̱̯῎1 ̰ῥ̳Ὶῒῗ̮ῌ M. kansasii ATCC kansasii ATCC 12478 ῞ῘΐῙ̯ RLU ratio῏0.03 12478 ῞ῘΐῙ̯῍ ATP QῚ KM, AMK Ί EB Ί̮̮Q RLU ratio῏0.1 Ί̮̮̯ῡ̯ῦ Q̯ῳ MB Q῞̳̮ ΐ ῤῶ̮῍ ῦ Q̯ 84.4ῑ Ί ̮῍ ̮῍ KM ῤῺΐ̮ 8 Q̱Ὶ ῎1 ̰ῥ̳Ὶῒῗ̮ῌ M. ̲Q̱ῤ RLU ratio῏0.03 Ὶ̳ ̮̮Ί̮῟ 97.8ῑ intracellulare ATCC 13950 ῞ῘΐῙ̯ LVFX Ὶ 1 ̯ῡ ΐῦ QῚῒῗ̮ῌ M. intracellulare ATCC Ῠ 2 ̰ῥ̲ῤῶ̮̯῟῟῍ RLU ratio ῤῖῠῚ̳ 13950 Ὶ̯῍ ̮̯Ῑ῟Q̱Ὶ ATP QΊ MB Q̯ ῎1 ̮Ῑ̯ῳΐῦ Qῤῶ̮̮ῌ ̰ ῥ ̳ Ὶ Q ῝ ῦ Q ῤ ῶ ̮ ̮ῌ Q Ό ̰ 4 ̰ Ὶ ̯ QΌ̰ 4 ̰ῤQ̱Q῞ RLU ratio῏0.1 Ὶ῟ῦ Q ῤῬῙ̯ῢΊ῍ SM, TH Ί LVFX ̯ 100ῑ, EB Ί INH ̯ 95ῑ ῚQ῝ῦ QῚῒῗ̮ῌ AMK Ί KM 95ῑ ῟ῦ QῚῒῗ̮ῌ 2. QῼQQ̰Ὶ῟ MIC QΊ ATP Q῟ῦ QΊ MIC QQ ̯ 85ῑ Ί̯̯ ΐ̰`῞ῒῡ῍ CAM Ί RFP ῞Ῐΐ Table 5 ῞῍ QῼQQ̰Ὶ῟ MB QΊ ATP Q῟Ύ Ῑ ̯ 75ῑ ῟ ΐ ῦ Q Ὶ ῒ ῗ ̮ῌ RLU ratio ῤ ῟Ὺῧῤῶ̮̮ῌ RLU ratio῏0.1 ῤQ̲Ί̮̮̲ ῏0.03 ῚῬῙ̯ῢΊ῍ CAM, TH ̯ 100ῑ, SM, INH ̱῞ M. avium ̯ 93.9ῑ Ίῳΐῦ Qῤῶ̮̮ῌ Q̱ Ί RFP ̯ 95ῑ ΊQ῝ῦ QῚῒῗ̮ῌ LVFX ̯ QῚ̯῍ RFP Ί CAM QΌ̰Ί Q῞ ΐ ῤῶ 90ῑ, EB ̯ 85ῑ Ί̯̯ ΐ̰`῞ῒῡ῍ AMK ̯ ̮̰`῞ῒῡ῍ INH ̯ 2 ̰ῥ̲ ΐ ῤῶ̮̰ 4 80ῑ῍ KM ̯ 70ῑ Ί ΐῦ QῚῒῗ̮ῌ Q̱QῚ ̰Ῥῠΰ̮ῌ RLU ratio῏0.03 Ὶ̳ ῤῲῗ̮Ί̮῟ ῬῢΊ CAM Ί RFP ῟ RLU ratio ῤ ῏0.03 Ί̮ῢ ῦ Q ̯῍ MB Q ῞ ̳ ̮ ῳ ΐ ῤ ῶ ̮ ̰ ` ῞ ῒ ῡ ̮Ί῞̯ῡῦ Q̲´̮̮῍ EB, KM Ί AMK ̯ 78.3ῑ Ί ΐ῍ RFP Ί CAM ῞ῘΐῙ̯ ῎1 ̰ῥ ΅Ώ̮ῢ̰`῞ῒῗ̮ῌ ̳Ίῦ Q̯̲´̮̮ῌ M. kansasii ῞ῘΐῙ̯῍ Table 4 ῞῍ MB QῚ 5 Ῠ̳Q̲ ̮̮Ί̮῟ MIC RLU ratio῏0.1 Ὶ̯̲Q̱ ῎1 ̰̱ῥ̳Ὶῒῡ῍ QQΊ ATP QῚ SM, EB, KM, INH, LVFX, TH, AMK RLU ratio῏0.03 Ὶ̳ ̮Ῑ̯ 1 ̰ῳ̮Ὸῢ̰`῞ῒ ῟ 7 Q̱῞ῘΐῙ̯ RLU ratio῏0.1 Ὶ̳ ̮῍ ̰̰ ῢ̯῟῟Q῝ῦ Qῤῶ̮̮ῌ M. intracellulare ̯ ῞̯ῗῙQῦ Q῟̮Ῥῠΰ̮ RFP, CAM ῟ 2 Q 90.6ῑ Ί̯̯ ΐῦ QῚῒῡ῍ SM, KM, INH, RFP ̱῞ῘΐῙ̯῍ ̳ ῤ RLU ratio῏0.03 Ὶῲῗ̮Ί̮ Ί CAM Ὶ MB Q ῞ ̳ ̮ ΐ ῤ ῶ ̮ ̮ῌ ̮ ̮ ῟ῦ Qῤῶ̮̮ῌ M. avium ATCC 700898 ῞Ῐΐ RLU ratio῏0.03 Ὶ̳ ̮ῢΊῳ̮Ὸῢ̰`῞ῒῡ῍ Ῑ̯῍ ATP QῚ῟Ὺῧ INH Ὶ 1 Ῠ 8 mg/ml ῤῶ ῞ EB ῞ῘΐῙ̯ RLU ratio῏0.1 ̳ Ὶ 1 ̰̱῞ ̮῍ MB QῚ῟̲ ῞̳̮ ῗ̮̯῟῟῍ ῾ 8 Q ̳ῗῙΐ̮῍ RLU ratio῏0.03 Ὶ̯ 3 ̰̱ῤῶ̮̯ ̱̯ ῎1 ̰ῥ̳Ὶῒῗ̮ῌ M. avium ATCC 25291 ῞ ῟ 20 ̰ 11 ̰ῚῬῠΰ̮ῌ ̮̮ CAM ῞Ῐΐ ῘΐῙ̯῍ ATP QῚ RFP ΐ̰`῞ῒῢ̯῟῟῍ Ῑ̯̮̯Ῑ ῎1 ̰̱ῥ̳῟ ῤῶ̮῍ RFP ῞ῘΐῙ ̳QQῼQ̲Q̰̱ Vol. 14 No. 3 2004. 39 ̯ῗ̮ῌῚ 168 Table 3. ῌ̯Ῐῌ̮̯῟̮ῌῒ῞ ̯ῌ῝ῡΊ̮ Concordance rate of the ATP method when the Microdilution (MB) method for reference strains is set as a basis. Strain RLU ratio Drug 0.1 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin M. avium ATCC 700898 0.03 M. avium ATCC 25291 0.1 0.03 M. kansasii ATCC 12478 5 1 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.03 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin No. 3 2004. 4 3 1 2 4 3 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 40 ̯ῠῢῖ̯Ῑ̯ΐ̮̮ Vol. 14 Discrepancy of MICs Concordance rate (ῑ) ῏῎3 ῎2 ῎1 0 ῍1 ῍2 ῐ῍3 1 5 1 1 3 5 3 1 2 1 5 2 5 2 2 3 4 4 4 1 2 1 1 1 1 1 4 1 3 4 3 1 3 2 4 1 1 1 3 1 1 3 4 1 1 1 97.8 2 1 88.9 1 1 1 5 3 77.8 2 1 1 1 1 3 3 3 3 2 1 1 3 1 4 1 2 1 2 1 1 1 5 1 1 3 4 75.6 1 1 1 3 5 4 1 4 2 4 1 4 5 3 1 4 5 5 4 5 4 4 1 84.4 1 1 1 97.8 ATP QῪQ̮῞Q´̱Q῾ ΏῡQ ῷ̳Q ̲ῡ̲Q Table 3. Strain continued Discrepancy of MICs Concordance rate () ΐῑ3 ῑ2 ῑ1 0 ῐ1 ῐ2 ῐ3 RLU ratio Drug 1 2 1 4 0.1 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 1 M. intracellulare ATCC 13950 0.03 Ῠ 1 ῶῪQ̮̯ ῒ1 Ὸ ΅QῡQῪ̳Ὶ῞ῌ 169 2 1 2 1 4 3 4 1 5 5 3 4 5 4 5 1 3 5 4 5 4 5 100 4 97.8 1 Q̯ MB Q̯ῡQQῢ 95.3 ̯Q̲̯̮̯῞ῌ Q QQQQῶ 45 ῶῪQ Q̯ῡQQῪ῍ RLU ra- Qῶ̯ῡ M. kansasii ATCC 12478 ̯ῩῩQQῢ tio0.1 ̯̲̯ῦ̰̯ EM ̯ AMK ῢ 100, TH ̯ Q̮Ῠῡῡ῍ ῝ῡQῡQQῶ̮̰ΰQQQQῶ̯ῡ´ LVFX ῢ 97.8 ῡQ̲̯QQῪ̳Ὶ῞ῌ SM, CAM ῳ̮̰῍ ATP Q̯ῡ 5 QΌQQ̯ῡQQῺQῢ SM, ̯ KM ῢ 91.1, RFP ῢ 86.7, INH ῢ 82.2 ̯Ῡ EB, KM, INH, LVFX, TH, AMK ῡ 7 Q ῠ̯̮̯ῢ ῩQ̮QQ̯̮̯῞῍ RLU ratio Ὺ 0.03 ̯̲̯ RLU ratio0.1 ῪQQ̯Ὶ῍ RFP. CAM ῢ RLU ῦ ̰ ̯῍ CAM ῢ 100, RFP ῢ 97.8, SM ῢ ratio0.03 ῪQQ̯QQΊ̰Ῐ̯̮QQ̯̮̰̯ 95.5, AMK Ῠ 93.3 ̯ Q ̲ ̯ Q Q Ὺ ̳ Ὶ῍ ̰̰̰ῌ LVFX ̯ TH ῢ 84.4, KM ῢ 82.2, INH ῢ 80 Tables 7῎8 ῠῢ῍ ̱̲QῪ̯῞QQQQῶῡ ̯ῩῩQ̮ῼ̲ῠ̮̰῍ EB ῢ 44.4 ̯Q̮QQ̯ ̮῟῍ M. avium ̯ M. intracellulare ῡ MB Q̯ ATP ̮̯῞ῌ Q Qῠ̲̯Ῠ CAM ̯ RFP ῢ RLU ratio Q̯ῡ MIC QQῪ̳Ὶ῞ῌ M. avium ῠ̯̮̯ῢQ̮ Ὺ 0.03 ̯QQΊ̰Ῐ̯ῠ̰̰QQ̮QQΊ̰Ῐ MIC Ὺ̳Ὶ῞ῡῢ RFP, CAM ̯῍ Qῶ 2 mg/ml ΅ῲ ̯̮̳Ῑ̰῞ῌ ̯̮̰QῴQῷQ̯̮̰Ῐ̯̮Qῧ̰̰῞ῌ ̳̮̯ Table 6 ῠῢ῍ ̱̲QῪ̯῞QQQQῶῡ MB LVFX ̮QῗQῶ 4 mg/ml ΅ῲ̯̮̯῞ῌ M. intra- Q̯῍ ATP Q̯ῡ MIC QQῪ̳Ὶ῞ῌ ATP Q̯ῡQ cellulare ῠ̯̮̯ῢ RFP, CAM ῡ MIC ῢQῗ῍ Qῶ Qῢ SM, EB, KM, INH, LVFX, TH, AMK ῡ 7 Q ῠ 0.25 mg/ml ΅ῲ̯ ̮ῷQῼ̲̮Qῧ̰̰῞ῌ ̳̮ ̯̮̯ῢ RLU ratio0.1 ῪQQ̯Ὶ῍ RFP, CAM ῢ ̯ LVFX ̮Qῶ 2 mg/ml ΅ῲῡQῴQQ̮ MIC ̯ RLU ratio0.03 ῪQQ̯Ὶ῞̯ῖῡQQῪ̳Ὶ ̮̯῞ῌ ῞ῌ M. avium ῠ̯̮̯ῢ῍ INH ̯ TH ̯ῢ MB Qῠ QῚ ATP QῢQ̮ῼ̲ῠῢ̮̰̮ 97.2 ̯Q̲̯ ῌ QQῪ̳Ὶ῞ῌ M. kansasii ῢΊῤ̯ῡQ ̮ ῒ1 ῾ ΏῡQ ῷ̳Q ̲Qῠ̯̮̯ῢ 1997 Qῠ´ ῍ Ὸ ΅Q̯̮̰Q̲̯QQῪ̳Ὶ῞ῌ ῥ῞ M. kan- ̱Ώῡ ̲Q̮QQ´̱Q̱ῠQQῙ̰̯̮̰ῌ Ῐ sasii ῡ MIC QQῠ̯̮̯ῢ RFP, CAM ̯῍ Qῶ Ῐ̯Q῭Ῑ̰῞ ̲QῢQQQQῪQ̮̰QQQ̯̮ 0.25 mg/ml ΅ῲ῍ LVFX ῠ 0.25 mg/ml ΅ῲ̯̮̯῞ ̰῍ ´ῳῢQQ 4 ̳΅Q̯QQQQQῡΏῡQ`̮Q ̮ KM ῠῢ 8 mg/ml ΅Qῡ MIC Ὺ̳Ὶ῞ῌ M. intra- Qῠ̯̯῞̳Q̯QQΊ̰Ῐ̯ῠ̯̯̯̮̰῍ ̳ΌῪ cellulare ῠ̯̮̯ῢ ATP Q̯ SM, KM ̮̰ΰ INH QΊ̰Qῢ̱QῙ̰̯̮̰῍ Ὶ̮Ῠ´̱Ώῠ̲QῙ ̮Q̮ῼ̲̮̮̰῍ QQῢ 92.2 ̯ῩῩQ̮ῼ̲ ̰῞Ῠῡ̯̮̯῞ῌ ῝Ῐ̯̰̰QQ̯Q̱̯Q ῷ̳ ῠ̮̯῞ῌ QQQQῶ̯̲QῪ̯῞ 45 ῶ̯ῡ ATP Q ̲Q̯Ὶ̯QQQῪQ̮῞῍ MGIT Q7) ῏ῬΎ QQQQQQQ ̳ Vol. 14 No. 3 2004. 41 QQQ̳῎Q 170 Table 4. ῎QQQ̳῎̳QQQ῎Q Q῎QQQQ Concordance rate of the ATP method when the Microdilution (MB) method for reference strains is set as a basis. RLU ratio MIC range of MB method ( mg/ml) M. avium Streptomycin ATCC 700898 Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.1 0.1 0.1 0.03 0.1 0.03 0.1 0.1 2ῌ4 4ῌ8 2 32ῌ32 0.06ῌ0.125 0.5 0.25 2ῌ8 1ῌ2 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.1 0.1 0.1 0.03 0.1 0.03 0.1 0.1 8 4ῌ8 8 32 2 2ῌ4 0.5 4ῌ8 4ῌ8 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.1 0.1 0.1 0.03 0.1 0.03 0.1 0.1 128 4ῌ8 4ῌ8 4ῌ8 0.25 0.25 0.125 1 2ῌ4 M. intracellulare Streptomycin ATCC 13950 Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.1 0.1 0.1 0.03 0.1 0.03 0.1 0.1 0.25 1 0.25ῌ0.5 4ῌ8 0.03 0.25 0.03ῌ0.06 16ῌ16 0.5 Strain M. avium ATCC 25291 M. kansasii ATCC 12478 Drug Discrepancy of MICs ΐ3 ΐ2 ΐ1 0 1 2 ̮3 Concordance rate (̮) 5 1 1 1 3 3 1 4 3 5 1 1 2 5 4 1 2 1 1 1 1 3 3 2 4 1 4 1 3 4 1 2 3 1 1 4 3 1 1 1 1 1 1 2 1 2 1 4 1 2 97.8 3 1 97.8 3 5 4 1 4 5 4 4 4 4 3 4 1 5 5 5 4 5 84.4 100 ̱῎̰`Ὺ̱΅̱ῒ ῧῲ̱῭̱`Ύ MTB-1 Q8) ´QQQ ̯̮̰QQῴQ̲QQ9)ῢ̲̯̮̯QQ̯ ῑ῾QQQ̳̲ῒ ̮̰̮ῤQQQ ̰Q̮̯ Ὼΰ ̮̰ῲ̱῭̱`Ύ NTM ̰Q̮̯ MB Qῠ῍ Q῞Q Q Q Q ̳ ´ Q Q Q Q ̯ ̮ ̰ ̱ Ῥ ̱ ` Ύ MTB-SR Q̳ΰQQῠῡ̰Ῠ̮ῢῳ̱ῐ̰̳Q῝QQ ῢ ῑ῾QQQ̳̲ῒ ῗΌQ̰̯̮̰ῌ ῝ῖ῝ NTM ῢ̮ QQΰ ATP Q̰QQ῝῍ ΊΰQῖῩ MIC Q̰̲ῦ̯ ̮̯ῤῨ̮ῧῙQQῴQ̲QQ̯̮̰ῲ̱῭̱`Ύ ATP QῠΰQῷ̰Qῥ̯ῌ NTM ῑ῾QQQ̳̲ῒ ῗQQῶῢQ̯̯̮̰ῗ῍ Q NTM ΰQ̳´QQQ ̰̳̮ῢ̮̯̰῍ QQQQ Q ῧQQ̲QῗΏQ̰̯̮ῡ̮ΰῗ Q̯̮̰ῌ ̰QῚ̰̯ῦΰQQῠῡ̰ΰῤῴQQQQ̯̮̰ῠ ̯ Ί ̯ Ὸ ῏ ῤ NCCLS (National Committee for Q̰̰̰ῌ ̲Q῟̰ῠῘῤQQQQ̲ΰῴ̰Q̮̰ Clinical Laboratory Standards) ΰQQ῝̯̮̰Q̳ Ίῠῗ Q̯̮̰῍ QQQQ̮̰̮ῤῴQQQQῢ 42 QQQQQQQῼQQ Vol. 14 No. 3 2004. ATP ῠΰῒΐ̯̮̮῟ῗῚ̮ῢῘ̮῝῟Ίῖ̮̯ Table 5. 171 Concordance rate of the ATP method when the Microdilution (MB) method for clinical isolates of M. avium, M. kansasii and M. intracellulare is set as a basis. Strain RLU ratio Drug 0.1 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin M. avium (N῎20) 0.03 M. kansasii (N῎5) 0.1 0.03 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.03 1 1 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin M. intracellulare (N῎20) Discrepancy of MICs Concordance rate (ῑ) ῏῍3 ῍2 ῍1 0 ῌ1 ῌ2 ῐῌ3 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 4 3 1 1 1 8 1 5 10 9 4 8 9 2 11 17 15 6 7 10 10 8 16 2 9 4 6 5 14 11 11 7 7 1 6 3 2 2 1 3 1 3 1 1 1 3 4 1 1 1 4 3 2 1 1 1 1 3 5 5 3 4 2 4 2 3 3 5 1 4 4 4 3 2 2 5 1 7 11 3 7 10 3 5 11 19 9 5 15 12 8 17 15 5 12 3 13 12 15 4 10 14 15 3 3 1 4 7 1 1 2 93.9 2 2 7 2 9 8 2 1 4 6 6 7 10 3 13 1 5 78.3 3 2 3 1 100 1 1 3 1 1 97.8 1 2 3 90.6 1 3 6 2 2 3 8 3 5 4 11 1 1 87.8 3 ̯ῡῤ῞̯̯̯̮Ῑ Vol. 14 1 1 No. 3 2004. 43 QQQ̳῎Q 172 Table 6. ῎QQQ̳῎QQQQ῎ῴQ Q῎QQQQ Concordance rate of the ATP method when the Microdilution (MB) method for clinical isolates of M. avium, M. kansasii and M. intracellulare is set as a basis. RLU ratio MIC range of MB method ( mg/ml) Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.1 0.1 0.1 0.03 0.1 0.03 0.1 0.1 1ῌ32 4ῌ128 2ῌ32 4ῌ32 0.125ῌ0.2 0.5ῌ4 0.125ῌ2 4ῌ16 1ῌ16 Streptomycin Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.1 0.1 0.1 0.03 0.1 0.03 0.1 0.1 4ῌ8 2ῌ4 8ῌ32 2ῌ4 0.125ῌ0.25 0.25ῌ0.5 0.125 1ῌ2 2ῌ8 M. intracellulare Streptomycin (Nΐ20) Ethambutol Kanamycin Isoniazid Rifampicin Levofloxacin Clarithromycin Ethionamide Amikacin 0.1 0.1 0.1 0.1 0.03 0.1 0.03 0.1 0.1 0.5ῌ4 2ῌ32 0.5ῌ4 2ῌ32 0.03ῌ0.25 0.5ῌ1 0.06ῌ0.25 4ῌ16 0.5ῌ4 Strain M. avium (Nΐ20) M. kansasii (Nΐ5) Drug Ῡ̯̯Q QQῗQQ῝̰ῚῠῤῩῙQ̰̰̯̮̰ῌ Discrepancy of MICs ῒ3 ῒ2 ῒ1 0 1 8 1 5 10 6 4 3 9 2 11 17 15 6 14 16 11 8 16 1 2 2 3 1 3 5 5 3 4 2 3 2 3 5 1 7 11 1 7 7 3 5 11 19 9 5 15 12 10 17 15 4 1 2 1 3 4 1 4 3 1 2 ̮3 Concordance rate (̮) 97.2 6 2 2 100 2 1 3 92.2 1 3 Q̰̰̯ῌ ῖῨ NTM ῢQ῝̰QQῤQ QῡQQῗQῡῙQ QQ̰4)ῑ6) ῤ ATP Q̯ Ό ΰQQ̲QQQ ̰ QῡQQῗQῙ῍ ΰQQ´QQQῢῩ̯̯̳ Q Q̯̮̰ῗ῍ QQῤ RLU ratio ̰ 0.5 ̯̳̯̯̮ ΰ̳ ´Qῢ̱̮ῗ̯̰ῨΰῠQ̰̰̰ῌ Ὶΰ̯ῧQ ̰ῌ ῶ῏ΰQῷ Q̰̳̯̯ NTM ῤ῍ Ό ῢQ ῷΰ QQῢῤ῍ QQQQῢ ̰QQQQ῍ McFar- ̯QQῗQῙ῍ ῦ̯̯̰ῢQ̯̯ ATP QῨQ̮Ὶῠ land No. 0.5 ̱QῢQQQQQQ῾ΰQQΰQ̮ ῖ̰῍ QQ̯Q̰ῠ̰̮̰ῠ̮̮Ὶῠ̯ RLU ratio ̱̰῍ QQ̲QQQ ῢQ̮̯ῌ ῤ ̮0.1 ̯QQ̰̳̯̯ῌ ῚΰQQ̲Q̯QQ̲̯ΰ ATP QΰQQQ̲ῢ̯̮̯Qῷΰ Q̯ῤ 5 Q̲ Q̰̳̯̯ῠῚ̰ M. avium ATCC 700898 ῠ M. ῠ̯ῗ῍ QQ̲̰Q̮̯ 3 Q̲QQ̯ΰ ATP Qῢ intracellulare ATCC 13950 ῤQ̳ῡ̱QQ̰Q̯ Ῡ̰QQ̲QQQ ̯ῤ῍ ῭̱ῳῐῲΎῬῲΰQ ῗ῍ M. avium ATCC 25291 ῤQ̮̱QQ̯̮̯̯ῌ RLU Qῗ 10,000 ῢQ῟῞῍ QQ̰Q῝Q̳ῗQῙ Ὶΰ̲̰QQQ̯ MB QῠQΏ῝̰ῠ RFP, CAM ῗ ̰̰̯ῌ ̯ΰ ῠ̯`ῐ΅ῐῤQ̯ῤ̮ῡ̮ῗ 2 ̲̱QΰQ̮Q̯̮̯̯ῌ ̯Ὶ̯ RLU ratio ̰Q̮ RLU ratio ῢῥ̰̯Ῐῗ ̰̰῍ 3 Q̲QQ̯ῤ MB ̯ MB Qῠΰ̱QQ̰ ̯ῠῚ̰῍ RLU ratio̮0.03 Qῠΰ̱QQῤ 60ῑ91.1̮ ῠQ̮ ῠῡ̯̯ῌ ̯ MB QῠQ̳ῡ̱QQ̰Q῝ῚῠῗῺQΊ̰̯ῌ Ὶ ATP Q̰Q̮̯ 3 Q̲QQ̯QQ̲QQQ ̰̳̮ ΰ ̳ῤ M. avium ῠ M. intracellulare ΰQQQQ̲ Q̳ῢῤ῍ Ί̰ῡ̰QQΰῸQ῍ QQQ ῍ QQQ ̯Ῠ ̰̰̰Ὶῠῖ̰῍ Q̯Q̲̰ RLU ratio̮ RLU ratio ̰̲ῧ̯ Q̰̳̮ῚῠῗQQ̯̮̰ῠ 0.1 ῠ RLU ratio̮0.03 ΰ 2 Ὺ̱̯QQ̰̳̮Ὶ 44 QQQQQQQῼQQ Vol. 14 No. 3 2004. ATP QΎ῍῟Q ´Q̳Q ῢQQ̲QQQ̳ῢ Q Table 7. 173 Distributions of MICs of nine antibiotics obtained by Microdilution (MB) and ATP method against clinical isolates of M. avium (Nῐ20). Drug Method Streptomycin MB method ATP method Ethambutol MB method ATP method Kanamycin MB method ATP method Minimum inhibitory concentration ( mg/ml) ῒ0.06 0.125 0.25 0.5 ῒ0.03 0.06 0.125 0.25 Isoniazid MB method ATP method Rifampicin MB method ATP method Levofloxacin MB method ATP method Clarithromycin MB method ATP method 3 ῒ0.5 Ethionamide MB method ATP method Amikacin MB method ATP method 1 1 1 8 5 2 2 1 2 4 8 16 32 1 1 3 6 6 7 4 3 4 3 2 3 3 2 2 7 7 5 4 2 2 3 1 6 5 7 7 2 5 1 2 4 8 1 1 7 8 5 0.5 1 4 5 2 4 4 4 5 6 6 6 7 2 2 1 2 8 3 5 9 4 4 2 4 8 16 ῑ16 5 7 5 6 5 6 3 1 2 1 1 3 5 9 6 6 6 1 64 128 ῑ128 1 1 1 16 32 ῑ32 4 5 6 2 1 2 2 4 2 ̯̯Ὶ῟ῌ RLU ratio Ύ̯ῢQ̯̯̰̮̰̮ῡ̯̯ Q̮̯̯̰QQῚῠῘ̯̯Ὶ῟ῌ “trailing” ΰ῍ QΊ̰ ̲ῢ QQQῢῴῡ̰̯̯Q̮ῠ ̳Qΰ῍ QQ̰11) ̰Qῦ̯̰̰̮ῡ῍ NTM ̯ΰ Ὸ ΎQῗ̰̰ῢ̯QῪ̰̰ῌ QΏῢ Qῡ̮̯̰ MBC (minimum bactericidal concentration) ̯ MIC M. kansasii ATCC 12478 ΰQQQ̮QῘ̯̮ ῢQ̮ῧ̯Ῥ̯ῢQQ̯ Ῡ̯QῘ̯ῡ̰̰̯Ῑ̰ ̰῍ RLU ratioΐ0.03 ̯Ὶ῟Q̮ RLU ratioΐ0.1 ̯ ̯̰ῌ QΏῢ Q̯ RLU ratio Ύ ΐ0.03 ̯QQῚ Ὶ῟̰̰̰ῶQΐ̮Qῖῠ̯῟̰ῢ̯QῪ̰̰ῌ Ῠ῟ ῟QQῡ῍ QQῡ̰̯̯ΰ MB QῡQῚQQΎQΊ QQῡ̰̯̯ RLU ratio ΎQῖΊ̰Ῐ̯ῡ̰̯̯ῶ ̰ῢ̰̰῍ Q̲ῠQ῏QῙ̰̯̯̰Ὺ̮ῠQ Qΐ̮QQῚ῟̮ῡ̯̯ΰ῍ QQ̰10)ῢ M. avium QῢQQ̮̰̰ῢ̯QQῙ̰̰ῌ Ῠ῟QQ̰ῢQ ῡQΊ̰̳ QῢQ῍Q̳`῏̱ῢQ̲ QῚ῟Q Q12)ῡ̳̰̰̰̰̮ῡ MAC ῡΰQῶ ̲ῌῐῢQῑ Qῡ̳̰̰̰̰̮ῡ῍ QQ̰ΰQQῢQQῡ̰̯ Qῢ̱ῠ̰ 3 QῢQῑQ̱̲̮QῒῙ̰῍ ῝ῢῲ̱ ̯QQ Qῡῴ̮̳̰̰῍ RFP ̮̰ῥ CAM ῡQ ̱̱΅̮̰ῥ̲QQ`῏̱̮῝̰῞̰̱ῠ̰Ῐ̯̮ Ὶ̯ M. avium ΰ MIC QQ ̯ῢQQQῼQ῍̮Q Q̰̰̯̰ῌ ̮QQ̮̰QQQQ̯QQQ῎Ὶ῟ Ῡ̰̰῍ QQQ̮῾Q̯̯῟Ῐ̯̮̰ ATP ῢQ MAC ̲̯ῢQQQQ̲QQ QΎQ̮QQῡΰ῍ Q̮Qῠῖ῍ RLU ratio QΎQῖ̯̰Ῐ̯ῡ̰̯̯ 3 QῢQῑQ̱̲̮QQΊ̰Ῐ̯̰̳̮̰̰̰Ῐ̯̮ MB Q̯ῢῶQΐ̮QQῚ῟̰ῢ̯QQῙ̰̰ῌ ̰ “trailing” ̳Qῢῶῷῡῠ̰̰ῢ̯QῪ̰̰ῌ Ό῎ NTM ῢQQ̲QQ Qῡ̮̯῍ ῚῤῚῤ “trail- ΰ Q Ώ ῢ Q Q ῡ ̮ Q Q ῒ ̲ ῡ ̯ ̯ ΰ Middle- ing” ̳Q̮̳̰̰῍ MIC QQQῡQ̮Ί̰Ῐ̯̮ brook 7H10 ̲QQQῡῺQQQῚ῍ QῒQ῎ΎQ̯ ̰ῌ Ό῎ΰ MB Q̯ῢQQΎQ̮̯̮ῡΰ῍ Q̲ῠ ῟̳ῡQ̳ῡῚ῟ῌ Q ̲ῢQῑ QΎQ̯̯Q QΎ῏QῚ῟QΎ MIC Q̯Ὶ῍ QQῠῳ̱̱῭̱ Q̲QQQ̳ΎΊ̰Ῐ̯ῡ̰̰̲QQ QQQῢQQ QQ̮QQQQ̲QQ Vol. 14 No. 3 2004. 45 QQQ̳῎Q 174 Table 8. ῎QQQ ῎QQQQ῎ῴQ Q῎QQQQ Distributions of MICs of nine antibiotics obtained by Microdilution (MB) and ATP method against clinical isolates of M. intracellulare (N20). Drug Method Streptomycin MB method ATP method Ethambutol MB method ATP method Kanamycin MB method ATP method Minimum inhibitory concentration ( mg/ml) ̮0.06 0.125 0.25 0.5 1 2 4 3 4 8 8 7 5 2 1 9 8 5 5 2 4 10 9 7 5 0.5 1 2 4 8 4 7 4 7 5 3 6 19 14 8 1 3 1 2 ̮0.03 0.06 0.125 0.25 Isoniazid MB method ATP method Rifampicin MB method ATP method Levofloxacin MB method ATP method Clarithromycin MB method ATP method 10 10 MB method ATP method Amikacin MB method ATP method 7 3 2 5 4 5 10 12 6 3 1 2 4 8 16 ̮16 2 6 4 3 4 7 6 4 4 8 10 3 1 7 4 16 5 5 64 128 ̮128 16 32 ̮32 3 2 3 1 1 1 32 1 1 3 5 1 ̮0.5 Ethionamide 2 8 0.25 mg/ml ῡ MIC Q̰Q´Qῠ̮̰Ὶ̯ῗΌQΊ ̰QῙ̰Ὶ̯ῧQQ̯̯̯̯Ῐ̰ῌ NCCLS ῡ Q Q ̲ Q Q ATCC Q Q ῼ ̯ ̮ ̰ M. ̰῟ῌ ῥ῟QQ̰12)ῗ M. intracellulare ῢ M. avium avium ATCC 700898 ῡ CAM ῠQ῝̰QQ̲Qῢ῍ ῠQ̯ RFP, SM ῞̯ KM ῠQ῝̰´QQῗ̳̮ QQ pH 7.3ῒ7.4 ̯ 0.5ῒ2 mg/ml ̯Ί̰̯̮̰ῗ῍ Ὶ̯῍ ̱Q῍ Ύ΅ῳ̱QῠQ̯ῢ M. avium ῡQῗ ̳Ὸῡ Q̯ῢ MB Q̯ 0.25 mg/ml, ATP Q̯ M. intracellulare ῠQ´QQῗ̳̮̯Q̳̯̮ 0.125ῒ0.25 mg/ml ̯QQQ̯Q̮Q̰Q῟ῌ ῞Ὶ ̰ῌ ῷῐῡ̳Ὸῡ ῶ̯ῧ RFP, SM ῞̯ KM ῠ̮ ̯ NCCLS ῡQQQ̯̮̰QQ̱Q̲QQ̯QQ̲Q ̮̯῍ M. intracellulare ῢ M. avium ῠQ̯Q̮Q ῲῑ῭ (pH 7.3 to 7.4) ̰Q QῠQQ῍ Q̰ ̰Q´Q ῠ̮̯῟ῌ ῖ῍ Ύ΅ῳ̱Qῠ̯̮ ̳̯῟ῗ MIC Qῢ 0.125ῒ0.25 mg/ml ̰Q῍ ̳Ὸ ̯ ῢ 2 ῾ Q ̲ ῠ ΰ ῤ Q Q ῡ MIC Q Q ̰ Q M. Q̰̳̯῟ MB Q῍ ATP Q̯Q̰̰̯̮Q̰Q avium ῡQῡ´QQῗ̳̮Ὶ̯ῢQῦ̰̰̯ῖ̯῟ῌ ῟ῌ Ὶ̰ῗQ̮῟ῼῡ̱̲̯ῡῖ῍ ̮̰̮ῢ῞ῡQῡ ῥ῟῍ MAC ῠQ῝̰´Q῎QQῡQQ`ῲῪῬῪ Q QQῡ̱̲̯ῡῖ̰ Q QQ̯̮̰ῌ ῖ῍ ̱῭ῗ Heifets13) ῠῩ̯̯Q̱Ί̰̯̮̰ῗ῍ Ὶ̰ῠ ῞ῡQῡ ATCC QQῼ 3 ῼῠ̯̮̯ῢ῍ Q̳̰ῡQ Ῡ̯̯QQ῝̰̯ M. intracellulare ῢ RFP ̯ CAM ̳11)ῠ ̰̰̰ 7 QQῠῩ̰ BrothMIC NTM ῡQ ῠQῼ´Q̯̮̯῟ῌ M. avium ῢ CAM ῠῢQῼ´ Ί̰῟ MIC QQ̯῍ ῷῐῡ̳Ὸῡ Q ῶ̯ῩῘ̱ Q̯̮̯῟ῗ῍ RFP ῢῨῨQ̮´QQ ΐMB Q῏ 55̮, Q̯̮῟ῌ ATP Q῏ 70̮ ̯̮̯῟ῌ ῞ῡQῡQQ̯ῢ MAC Q QQQQῼῡ̳῾ΏQ̰Q ῟ῗ῍ M. kansasii ῠQQQQ̯̯QQΊ̰̰ SM, EB, AMK ῠ̯̮̯ ῡ RFP ῠQ῝̰ NCCLS ῡQQQQ`ῲῪῬῪ̱ ῢ ΐMB Q῏ 22.5ῒ47.5̮, ATP Q῏ 22.5ῒ60̮ ῡ ῭ῗ 1 mg/ml ̯̮̰῍ ̳Ὸῡ Q῟ῼ̯ῢ 0.125ῒ ´QQ̯̮̯῟ῌ 46 QQQQQQQῺQQ Vol. 14 No. 3 2004. ATP ῡῩ̰ΊῘQQQQ̳̯ῩQ QQQQ̯Qῌ NTM ̯QῘ῝Q̳Q̯Q̳Ῡ̮Q̯QQ῞̯῍ NTM ̯QῘ῝QῘ̮̲̮ῗῧ̰ΰQῗῧ῝ῦQ rifamycin ̰῎Q῍ caprazamycin-B, ketolides Q̯ ̯̯ῩQ QQQQ̮ΰ̯̰̰Ὺ̰ῑ`ῑῬῡ̯̯῞ ̯̯̰ῩQ῞̯῝ῼῢῚ̰῟̯ῦῢ̯῟QῨῧ῍ QQ ῡῙῡῙῲQ̯ῖῗῧῦῩQ QQQQ̮ΰῡῧ ῦῌ ATP ῡ̯Q ῏ῲQ̯΅ ῞QῌῘΊQῪῡ ̯ MB ῡ῟ῢΰ̮Q Ῐ῍ Ῐ̮ῢQQΏ´̮῞ῦῖ ῟̮ῤ῍ NTM ̯ῩQ QQQQῡ῟Ῐ῝̰̰῞ῦ ῟῝ῼῗῧΊῌ ῡΊ῍ ̳̮̱QῥQ̯̮῏̮ῑῘ NTM ̯ῩQ QQQQ̯῍ ATP ῡῩ̰ῦῖ῟̯ΰ ῥ῍ QQΏῗῧΊ̳ Q̯̮QῩQ̮ῖ῟̮´̮῞ ῥ῍ ̮QQ̯QQ̯ῴ̳ῗῧ̯ῠῡ῟Ῐ῝̰̰῞ ῦῌ ῍ ῌ 1) ΐῤQ῞ ̲ῘQQQQ̳QQ̱̱̲῎ 2003. ̮ῘQQQQ̳QQQ̯ ῙῦQ̲ῌ2003 ῎ Q 78: 569῍572. 2) ̳QQQ῍ Q῎ 1997. ̳Qῐ ATP QQ̯ΰῦQ ̳Ῡ QQQQ ῒQῒ̯ΐ῎ ΐΏ`̲Q ῒQ 45 ̲ ΐῤΏ `ῡ ̲QῳQΐῐ 146. 3) ̱Q̮̳῍ Q῎ 1997. ATP ῡ̯QQῩ QQQQ ῠ̯ῷ̰ ῒῚ̯ 3ΐ ῲQ̯΅ Ῡ̰ΊQ̰Q̯ ῝ῼ῎ ΐΏ`̲Q ῒQ 45 ̲ΐῤΏ `ῡ ̲Qῳ Qΐῐ 147. 4) QQῬ̰῍ QQQQ῍ ΌQ ῢ῍ Q῎ 1998. Q̯̮ QῩ̰ΊQ̳̯QQῩQ QQQQῡῌMycobacterium tuberculosis H37Rv Ῡ̰Ί QQ Qῌ῎ ῲQ῞῭ 46: 834῍840. 5) QQῬ̰῍ QQQQ῍ QῺQῧ῍ Q῎ 1999. Q̯̮ QῩ̰ΊQ̳̯QQῩQ QQQQῡ ῒQ II ̯ΐῐ ῩQQ̯Qῌ̮ΰ̯QQῡ῟̯ῗ ῎ ῲQ 175 ῞῭ 47: 170῍175. 6) QQῬ̰῍ QQQQ῍ QῺQῧ῍ Q῎ 2000. Q̯̮ QῩ̰ΊQ̳̯QQῩQ QQQQῡ ῒQ III ̯ΐῌfilamentous cell treatment ̯ΰῦ ATP ῡ ̯̲῎ ῲQ῞῭ 48: 167῍173. 7) Palaci M, et al. 1996. Evaluation of Mycobacteria Growth Indicator Tube for recovery and drug susceptibility testing of Mycobacterium tuberculosis isolates from respiratory specimens. J. Clin. Microbiol. 34: 762῍764. 8) QQQ̳῍ QQQ ̰῍ QQ Q῍ Q῎ 1998. Middlebrook QQ̮Q῞̯QQ̳ῩQ QQQQ ῒQ 2 ̯ΐῐ ῩQQQ̰̰ῲ΅ῑΎῩ̰ΊῙῶ Q Qῡ῞̯QQ̮̱QQQ̯QQ ῒQῐ 3 Q Q̯̮̮ῦ̳῍῝ῼΐ῎ ῲQ῞῭ 46: 719῍727. 9) National Committee for Clinical Laboratory Standards (NCCLS). 2003. Susceptibility Testing of Mycobacteria, Nocardia, and Other Aerobic Actynomycetes; Approved Standard M24A, NCCLS, Wayne, USA. 10) QῨQ̲῍ QQQQ῍ ̯ΌQῦ῎ 2001. Rifalazil, clarithromycin ̮ΰ̯ levofloxacin ̯ 7HSF ῶ Q̮QQ̯Q̳̮ΰ̯ Mycobacterium avium ̯QῙῦQ̳Q̰̮Q῭̰ῑῳ῎ ΐΏ`̲Q 49: 649῍652. 11) QQQ̳῍ ῸQQΊQ῍ QQ Q῍ Q῎ 2002. Middlebrook QQ̮Q῞̯QQ̳ῩQ QQQQ ῒQ 4 ̯ΐῐ Nontuberculous Mycobacteria ῩQQQ Q῟ῙῦῙῶQ Qῡ῍ BrothMIC NTM ̯̲ ̮῝ῼ . ῲQ῞῭ 50: 381῍391. 12) QQQQ῍ ῟ΌQῦ῍ QῨQQ῍ Q῎ 2002. Mycobacterium intracellulare ̯̱̯ῦQΎ̯̱ ̯῾ QQ̳Q̯QῙῦ QQ̯῝῎ ΐΏ`̲Q 50: 232῍235. 13) Heifets. 1996. Susceptibility Testing of Mycobacterium avium Complex Isolates. Antimicrob. Agents Chemother. 40: 1759῍1767. ΐῤῲQῙQ̯ QQ Vol. 14 No. 3 2004. 47 176 ̯Ὶ̮ῌῘ ῒῌ̯Ὶῖΐῌ῞῝̮ῌῐ ̯ῌΊ̯Ῑ̮ Evaluation of Microdilution Susceptibility Test for Nontuberculous Mycobacteria Using the ATP Method Masayuki Horikawa1), Isao Nishi1), Masahiro Toyokawa1), Naoki Sato2), Yutaka Okazawa2), Seishi Asari1) 1) 2) Clinical Laboratory, Osaka University Hospital Kyokuto Pharmaceutical Industrial CO., LTD. We have developed a broth microdilution susceptibility test method for nontuberculous mycobacteria (NTM); in this test method, adenosine triphosphate (ATP) is measured through bioluminescence, with viable count as the indicator. We have conducted the basic analysis of the method and compared the method with the broth microdilution method (MB method) using four ATCC referece strains and 45 clinical isolates. As a result of the examination of the ATP-measuring method (ATP method), it has been shown that 30-minute pre-processing at room temperature and extraction at 60῎ for 10 minutes were appropriate steps. For the ATP method, judgment was made by measuring the relative light unit (RLU) ratio after culturing the microplate for 5 days, and comparison was made with the MIC value after culturing for 7 days. When the positive values for growth were set to an RLU ratio of ῍0.1 for 7 drugs, that is, streptomycin, ethambutol, kanamycin, isoniazid, levofloxacin, ethionamide and amikacin, and to an RLU ratio of ῍0.03 for rifampicin and clarithromycin, the rate of concordance with the MB method was rather lower (84.4῏) for Mycobacterium kansasii ATCC 12478, whereas 97.8ῌ100῏ concordance rates were obtained for 3 ATCC reference strains. Regarding clinical isolates, the concordance rates obtained for Mycobacterium intracellulare, Mycobacterium avium, and M. kansasii were 92.2῏, 97.2῏ and 100῏, respectively. From the above results, we have concluded that the ATP method is useful as a method of testing drug susceptibility of NTM. 48 ῟̯ῠ̮̯ῗ̯ῑ̮̮ Vol. 14 No. 3 2004.
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