Herman L M, Vaerewijck M J, Moermans R J, Waes G M
Government Dairy Research Station, Agricultural Centre Ghent, Melle, Belgium.
Appl Environ Microbiol. 1997 Aug;63(8):3139-43. doi: 10.1128/aem.63.8.3139-3143.1997.
A PCR method was developed to detect spores of Bacillus sporothermodurans in 1, 10, and 100 ml of raw milk. Two primers were derived from a unique sequence after subtractive hybridization of B. sporothermodurans DNA with DNA of MB 397, a not yet identified spore-forming bacterium isolated from raw milk, closely related to B. sporothermodurans. Specific identification was proven on a large collection of Bacillus strains and on strains from relevant taxa. The detection of B. sporothermodurans in raw milk is based on activation, germination, and outgrowth of the spores, followed by PCR identification. Spores from 10 and 100 ml were concentrated by centrifugation after chemical extraction of the milk components. The total test takes 28 h. The detection limits are 9, 0.4, and 0.22 CFU/ml for 1, 10, and 100 ml, respectively.
开发了一种PCR方法，用于检测1毫升、10毫升和100毫升原料乳中嗜热栖热芽孢杆菌的孢子。从嗜热栖热芽孢杆菌DNA与MB 397（一种从原料乳中分离出的尚未鉴定的产芽孢细菌，与嗜热栖热芽孢杆菌密切相关）的DNA进行消减杂交后的独特序列中获得了两条引物。在大量芽孢杆菌菌株和相关分类群的菌株上证实了特异性鉴定。原料乳中嗜热栖热芽孢杆菌的检测基于孢子的激活、萌发和生长，随后进行PCR鉴定。在对乳成分进行化学提取后，通过离心浓缩10毫升和100毫升中的孢子。整个检测过程需要28小时。1毫升、10毫升和100毫升的检测限分别为9 CFU/毫升、0.4 CFU/毫升和0.22 CFU/毫升。
