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首页> 美国卫生研究院文献>Gut Microbes >How free of germs is germ-free? Detection of bacterial contamination in a germ free mouse unit
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How free of germs is germ-free? Detection of bacterial contamination in a germ free mouse unit

机译：细菌中如何不含细菌？检测无菌鼠标单元中的细菌污染

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Management of germ free animals has changed little since the beginning of the 20th century. The current upswing in their use, however, has led to interest in improved methods of screening and housing. Traditionally, germ free colonies are screened for bacterial colonization by culture and examination of Gram stained fecal samples, but some investigators have reported using PCR-based methods of microbial detection, presumably because of perceived increased sensitivity. The accuracy and detection limit for traditional compared to PCR-based screening assays are not known. The purpose of this study was to determine the limit of detection of bacterial contamination of mouse feces by aerobic and anaerobic culture, Gram stain, and qPCR, and to compare the accuracy of these tests in the context of a working germ free mouse colony. We found that the limit of detection for qPCR (approximately 105 cfu/g of feces) was lower than for Gram stain (approximately 109 cfu/g), but that all 3 assays were of similar accuracy. Bacterial culture was the most sensitive, but the least specific, and qPCR was the least sensitive and most specific. Gram stain but not qPCR detected heat-killed bacteria, indicating that bacteria in autoclaved diet are unlikely to represent a potential confounding factor for PCR screening. We conclude that as a practical matter, bacterial culture and Gram stain are adequate for screening germ free mouse colonies for bacterial contaminants, but that should low numbers of unculturable bacteria be present, they would not be detected with any of the currently available means.
机译：自20世纪初以来，无菌动物的管理几乎没有改变。然而，目前使用它们的兴起引起了对改进的筛选和安置方法的兴趣。传统上，通过培养和检查革兰氏染色的粪便样品来筛选无菌菌落的细菌定植，但是一些研究者已经报道了使用基于PCR的微生物检测方法，这可能是因为感觉到的敏感性增加了。与基于PCR的传统筛查方法相比，传统方法的准确性和检测极限尚不清楚。这项研究的目的是确定通过需氧和厌氧培养，革兰氏染色和qPCR检测小鼠粪便中细菌污染的极限，并在无病菌的小鼠菌落中比较这些检测的准确性。我们发现qPCR的检出限（大约10 5 cfu / g粪便）低于革兰氏染色（大约10 9 cfu / g），但是所有3种测定法的准确性相似。细菌培养是最敏感的，但特异性最低，而qPCR的敏感度和特异性最低。革兰氏染色但qPCR未检测到热灭活细菌，这表明高压灭菌饮食中的细菌不太可能代表PCR筛选的潜在混杂因素。我们得出的结论是，实际上，细菌培养和革兰氏染色足以筛查无菌小鼠菌落中的细菌污染物，但是如果存在少量不可培养的细菌，则无法通过任何当前可用的手段对其进行检测。

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