Shigeto E, Taska H
National Hiroshima Hospital, Higashihirosima-shi, Japan.
Kekkaku. 1996 Mar;71(3):245-52.
BACTEC system is a reliable and rapid drug susceptibility test for mycobacteria and is widely accepted in Europe and in the United States of America. In Japan, it is impossible to introduce the BACTEC system in clinical laboratories because of strict regulations for the use of radioactive substances in Japan. To resolve this dilemma, we adopted alpha-antigen (alpha-antigen), a widely distributed secretory protein of mycobacteria, as the index substance replacing the radioactive substance 14C in BACTEC system. Alpha-antigen was detected by reverse passive latex agglutination (LA), using latex sensitized with rabbit anti-alpha-IgG. Susceptibility of 17 M. tuberculosis strains isolated from patients, grown on Ogawa egg medium and then cultured in 7H9 medium, and of 46 M. tuberculosis strains freshly isolated from patients by 7H9 medium of MB check system, was tested for four antituberculosis drugs, isoniazid (INH), rifampicin (RFP), streptomycin (SM) and ethambutol (EB). Ninety four percent of the control cultures were positive for alpha-antigen within 7 days after the inoculation. The MIC values of H37Rv strain in 7H9 medium determined by the method of Heifets were 0.05 micrograms/ml for INH, 0.03 micrograms/ml for RFP, 0.025 micrograms/ml for SM and 1.9 micrograms/ml for EB. In 17 strains from Ogawa egg medium, the results obtained from all but 4 strains for SM, 1 for INH, 1 for RFP and 7 for EB were concurrent with that obtained by the method using 1% Ogawa egg medium. No strains were determined to be resistant to any drug by the alpha-antigen method and be sensitive by the Ogawa medium method. In 46 strains cultured by the MB check system, the results of 42 strains for SM, 35 for INH, 39 for RFP and 36 for EB coincided with those determined by the Microtiter method. Among the strains determined to be resistant by Microtiter method, 1/2 for SM, 10/14 for INH, 4/17 for RFP and 8/10 for EB were determined to be sensitive by alpha-antigen LA method. The disagreement was seen mostly in strains which were determined to be resistant by the method using egg medium, while sensitive by the alpha-antigen LA method. The discrepancy might originate from the difference of critical concentration due to heat inactivation of the drugs and absorption in the egg medium. However, some instability was observed in latex agglutination and its cause should be examined further. This method of utilizing 7H9 medium for culture and alpha-antigen as the index of mycobacterial growth can be an expedient and economical drug susceptibility test because it does not use radioactive substance as in the case of BACTEC system.
BACTEC系统是一种用于分枝杆菌的可靠且快速的药敏试验，在欧洲和美国被广泛接受。在日本，由于日本对放射性物质使用的严格规定，临床实验室无法引入BACTEC系统。为了解决这一困境，我们采用了α-抗原（一种分枝杆菌广泛分布的分泌蛋白）作为替代BACTEC系统中放射性物质14C的指标物质。使用兔抗α-IgG致敏的乳胶通过反向被动乳胶凝集试验（LA）检测α-抗原。对从患者分离出的17株结核分枝杆菌菌株进行药敏试验，这些菌株先在小川鸡蛋培养基上生长，然后在7H9培养基中培养，以及对通过MB检查系统的7H9培养基从患者新鲜分离出的46株结核分枝杆菌菌株，检测四种抗结核药物，即异烟肼（INH）、利福平（RFP）、链霉素（SM）和乙胺丁醇（EB）。94%的对照培养物在接种后7天内α-抗原呈阳性。Heifets法测定H37Rv菌株在7H9培养基中的MIC值为：INH 0.05微克/毫升，RFP 0.03微克/毫升，SM 0.025微克/毫升，EB 1.9微克/毫升。在来自小川鸡蛋培养基的17株菌株中，除4株SM、1株INH、1株RFP和7株EB外，其余所有结果与使用1%小川鸡蛋培养基的方法所得结果一致。没有菌株通过α-抗原法被判定对任何药物耐药而通过小川培养基法被判定敏感。在通过MB检查系统培养的46株菌株中，42株SM、35株INH、39株RFP和36株EB的结果与微量滴定法测定的结果一致。在微量滴定法判定耐药的菌株中，1/2的SM、10/14的INH、4/17的RFP和8/10的EB通过α-抗原LA法被判定敏感。差异主要出现在通过鸡蛋培养基法判定耐药而通过α-抗原LA法判定敏感的菌株中。这种差异可能源于药物热灭活导致的临界浓度差异以及在鸡蛋培养基中的吸附。然而，在乳胶凝集试验中观察到一些不稳定性，其原因应进一步研究。这种利用7H9培养基培养并以α-抗原作为分枝杆菌生长指标的方法可以成为一种方便且经济的药敏试验，因为它不像BACTEC系统那样使用放射性物质。
