Kirii Y, Danbara H, Komase K, Arita H, Yoshikawa M
Department of Bacteriology, University of Tokyo, Japan.
J Clin Microbiol. 1987 Oct;25(10):1962-5. doi: 10.1128/jcm.25.10.1962-1965.1987.
By using biotinylated enterotoxin DNA probes, a method to detect enterotoxigenic Escherichia coli by colony hybridization was developed. The treatment of colonies on nitrocellulose membrane filters with proteinase K and Triton X-100 was essential for obtaining the specific hybridization. A total of 200 E. coli strains isolated from travelers with diarrhea were tested for colony hybridization by using a probe encoding heat-labile toxin (LT) type h. All strains (86 of 86) that produced LT, but none of the non-LT producers, hybridized with 32P-labeled and biotinylated LT type h probes. A total of 36 strains chosen randomly from the 200 isolates were tested for colony hybridization by using heat-stable enterotoxin (ST) probes. All but two strains that hybridized with the 32P-labeled ST type Ia probe also hybridized with the biotinylated ST type Ia probe. All strains that hybridized with the 32P-labeled ST type Ib probe also hybridized with the biotinylated ST type Ib probe. Thus, almost all E. coli strains tested were judged to be the same by colony hybridization with biotinylated or 32P-labeled enterotoxin probes. These results demonstrate that the biotinylated enterotoxin probes are useful in the diagnosis of enterotoxigenic E. coli strains by colony hybridization.
通过使用生物素化的肠毒素DNA探针，开发了一种通过菌落杂交检测产肠毒素大肠杆菌的方法。用蛋白酶K和 Triton X - 100处理硝酸纤维素膜滤器上的菌落对于获得特异性杂交至关重要。使用编码h型不耐热毒素（LT）的探针，对从腹泻旅行者中分离出的200株大肠杆菌进行菌落杂交检测。所有产生LT的菌株（86株中的86株），但非LT产生菌均未与32P标记和生物素化的h型LT探针杂交。从200株分离物中随机选择36株菌株，使用耐热肠毒素（ST）探针进行菌落杂交检测。除两株外，所有与32P标记的Ia型ST探针杂交的菌株也与生物素化的Ia型ST探针杂交。所有与32P标记的Ib型ST探针杂交的菌株也与生物素化的Ib型ST探针杂交。因此，通过与生物素化或32P标记的肠毒素探针进行菌落杂交，几乎所有测试的大肠杆菌菌株都被判定为相同。这些结果表明，生物素化的肠毒素探针可用于通过菌落杂交诊断产肠毒素大肠杆菌菌株。
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