Department of Animal Management and Treatment, Fakous Veterinary Administration, Sharkia Governorate, Fakous, Egypt.
Department of Microbiology, Faculty of Veterinary Medicine, Zagazig University, Zagazig, Egypt.
J Appl Microbiol. 2020 Jun;128(6):1595-1605. doi: 10.1111/jam.14584. Epub 2020 Feb 5.
This study was conducted to early detect the negative culture bacterial pathogens causing subclinical mastitis for the fast diagnosis of the disease and the reduction of some milk-transmitted pathogenic bacteria to human consumers.
A total of 171 positive California mastitis test (CMT) milk samples collected from asymptomatic dairy cows in Sharkia Governorate, Egypt were examined by conventional bacteriological methods. The obtained results revealed that Streptococcus species (77·2%), followed by Staphylococcus species (48·6%) and Escherichia coli (25·7%) were the most predominant bacterial pathogens isolated from positive culture milk samples, whereas Enterobacter and Pseudomonas species were the lowest ones (1·2%, for each). Herein, 13 (7.6%) negative culture milk samples were subjected to propidium monoazide (PMA) conventional PCR assay, followed by DNA sequencing of purified PCR amplicons. Sequence analysis identified seven different types of negative culture bacterial pathogens comprising as following; 4 Enterococcus hirae, 2 Bacillus cereus, 2 Staphylococcus aureus, 1 Bacillus mycoides, 1 Bacillus subtilis, 1 Enterococcus faecium and 1 Escherichia coli.
All the detected negative culture bacterial pathogens by PMA-PCR assay, followed by DNA sequencing were incriminated in causing subclinical mastitis disease and had serious implications on human public health through consumption of milk contaminated with those recovered bacterial pathogens.
The used methods could be useful in the routine detection of negative culture bacterial pathogens present in milk and consequently, it will help in the rapid diagnosis of subclinical mastitis disease and the reduction of many milk-transmitted diseases to human.
本研究旨在早期检测引起亚临床乳腺炎的阴性培养细菌病原体，以便快速诊断该疾病，并减少一些经牛奶传播给人类消费者的致病菌。
从埃及 Sharkia 省无症状奶牛的 171 份阳性加州乳腺炎测试（CMT）奶样中，采用常规细菌学方法进行检测。结果显示，链球菌属（77.2%）、葡萄球菌属（48.6%）和大肠杆菌（25.7%）是从阳性培养奶样中分离出的最主要的细菌病原体，而肠杆菌属和假单胞菌属则是最低的（各占 1.2%）。在此，13 份（7.6%）阴性培养奶样进行了吖啶橙单染（PMA）常规 PCR 检测，随后对纯化的 PCR 扩增子进行 DNA 测序。序列分析鉴定出 7 种不同类型的阴性培养细菌病原体，包括 4 株屎肠球菌、2 株蜡样芽孢杆菌、2 株金黄色葡萄球菌、1 株蕈样芽孢杆菌、1 株枯草芽孢杆菌、1 株屎肠球菌和 1 株大肠杆菌。
PMA-PCR 检测和 DNA 测序鉴定的所有阴性培养细菌病原体都与亚临床乳腺炎有关，通过食用受这些恢复细菌病原体污染的牛奶，对人类公共健康构成严重威胁。
所使用的方法可用于常规检测牛奶中存在的阴性培养细菌病原体，从而有助于快速诊断亚临床乳腺炎，并减少许多经牛奶传播的疾病。
