用 N-乙酰-L-半胱氨酸-氢氧化钠进行化学去污可提高从培养乳中回收活的分枝杆菌副结核亚种的效果。
Chemical decontamination with N-acetyl-L-cysteine-sodium hydroxide improves recovery of viable Mycobacterium avium subsp. paratuberculosis organisms from cultured milk.
机构信息
Roy J. Carver Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, Iowa, USA.
出版信息
J Clin Microbiol. 2013 Jul;51(7):2139-46. doi: 10.1128/JCM.00508-13. Epub 2013 May 1.
Mycobacterium avium subsp. paratuberculosis is shed into the milk and feces of cows with advanced Johne's disease, allowing the transmission of M. avium subsp. paratuberculosis between animals. The objective of this study was to formulate an optimized protocol for the isolation of M. avium subsp. paratuberculosis in milk. The parameters investigated included chemical decontamination with N-acetyl-l-cysteine-sodium hydroxide (NALC-NaOH), alone and in combination with antibiotics (vancomycin, amphotericin B, and nalidixic acid), and the efficacy of solid (Herrold's egg yolk medium [HEY]) and liquid (Bactec 12B and para-JEM) culture media. For each experiment, raw milk samples from a known noninfected cow were inoculated with 10(2) to 10(8) CFU/ml of live M. avium subsp. paratuberculosis organisms. The results indicate that an increased length of exposure to NALC-NaOH from 5 to 30 min and an increased concentration of NaOH from 0.5 to 2.0% did not affect the viability of M. avium subsp. paratuberculosis. Additional treatment of milk samples with the antibiotics following NALC-NaOH treatment decreased the recovery of viable M. avium subsp. paratuberculosis cells more than treatment with NALC-NaOH alone. The Bactec 12B medium was the superior medium of the three evaluated for the isolation of M. avium subsp. paratuberculosis from milk, as it achieved the lowest threshold of detection. The optimal conditions for NALC-NaOH decontamination were determined to be exposure to 1.50% NaOH for 15 min followed by culture in Bactec 12B medium. This study demonstrates that chemical decontamination with NALC-NaOH resulted in a greater recovery of viable M. avium subsp. paratuberculosis cells from milk than from samples treated with hexadecylpyridinium chloride (HPC). Therefore, it is important to optimize milk decontamination protocols to ensure that low concentrations of M. avium subsp. paratuberculosis can be detected.
分支杆菌副结核亚种会随患有晚期副结核病的奶牛的奶和粪便排出，从而在动物之间传播分支杆菌副结核亚种。本研究的目的是制定一种优化的牛奶中分支杆菌副结核亚种分离方案。研究中考察的参数包括使用 N-乙酰-L-半胱氨酸-氢氧化钠（NALC-NaOH）进行化学去污染，单独使用和与抗生素（万古霉素、两性霉素 B 和萘啶酸）联合使用，以及固体（赫罗尔德鸡蛋蛋黄培养基 [HEY]）和液体（Bactec 12B 和 para-JEM）培养基的效果。对于每个实验，均将已知未感染奶牛的生奶样品接种 10(2) 至 10(8) CFU/ml 的活分支杆菌副结核亚种生物体。结果表明，NALC-NaOH 的暴露时间从 5 分钟增加到 30 分钟，NaOH 的浓度从 0.5%增加到 2.0%，不会影响分支杆菌副结核亚种的生存能力。NALC-NaOH 处理后再用抗生素处理牛奶样品，会比单独用 NALC-NaOH 处理降低活分支杆菌副结核亚种细胞的回收率。在三种评估的培养基中，Bactec 12B 培养基是从牛奶中分离分支杆菌副结核亚种的最佳培养基，因为它达到了最低的检测阈值。NALC-NaOH 去污染的最佳条件被确定为在 1.50%的 NaOH 中暴露 15 分钟，然后在 Bactec 12B 培养基中培养。本研究表明，与使用十六烷基吡啶氯（HPC）处理的样品相比，NALC-NaOH 化学去污染从牛奶中回收活分支杆菌副结核亚种细胞的效果更好。因此，优化奶去污染方案以确保能够检测到低浓度的分支杆菌副结核亚种非常重要。
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