摘要

       出于治疗和预防原因，在奶牛场中长期使用抗生素，而且农场环境中抗生素抗性细菌 (ARB) 的流行率较高，这引起了人们对食用生牛奶及其衍生产品的担忧。本研究的目的是通过鸟枪宏基因组分析预测抗生素抗性基因 (ARG) 的存在，该基因主要与革兰氏阴性细菌相关，这些基因主要与来自南蒂罗尔（意大利北部）健康动物的无抗生素残留生牛奶中的革兰氏阴性菌相关，选择为一个模型系统。对重建支架的散弹枪宏基因组数据的评估揭示了假单胞菌属的存在。作为含有 ARG 的生牛奶样品中最丰富的革兰氏阴性菌。此外，ARGs 还与乳酸菌如 Lactococcus sp. 相关。和乳酸菌。与牛奶样品中发现的微生物组相关的 ARG 赋予对氨基糖苷类链球菌素、β-内酰胺酶、大环内酯、四环素、碳青霉烯、头孢菌素、青霉烯、肽、青霉烯、氟喹诺酮、氯霉素和伊法霉素抗生素的抗性。进一步的生物信息学处理包括将所有牛奶样品的所有宏基因组序列从头重新组装，以重建宏基因组组装基因组 (MAG)，并进一步用于研究移动遗传元件 (MGE)。对重建的 MAG 的分析表明，MAG 9 (Pseudomonas sp1.) 包含转移毒力因子所需的 oriT 基因（转移基因的起源）。虽然假单胞菌在生牛奶中很常见，但巴氏杀菌处理降低了它们的存活率。然而，应注意假单胞菌属。由于它们对抗生素的内在抗性以及它们将毒力因子转移给其他细菌的能力。

       Extended use of antibiotics in dairy farming for therapeutic and prophylactic reasons, but also the higher prevalence of antibiotic resistant bacteria (ARB) in the farm environment raised the concern of consuming raw cow's milk and its derived products. The aim of this study was to predict by shotgun metagenomic analyses the presence of antibiotic resistance genes (ARGs) mainly correlated with Gram-negative bacteria in antibiotic residue free raw cow's milk derived exclusively from healthy animal from South Tyrol (Northern Italy), chosen as a model system. Assessment of shotgun metagenomic data of reconstructed scaffolds, revealed the existence of Pseudomonas spp. as the most abundant Gram-negative species in the raw cow's milk samples bearing ARGs. Besides, ARGs also linked to lactic acid bacteria such as Lactococcus sp. and Lactobacillus sp. ARGs correlated to microbiome found in milk samples conferred resistance towards aminoglycoside-streptothricin, beta-lactamase, macrolide, tetracycline, carbapenem, cephalosporin, penam, peptide, penem, fluoroquinolone, chloramphenicol and elfamycin antibiotics. Further bioinformatic processing included de-novo reassembly of all metagenomic sequences from all milk samples in one, to reconstruct metagenome assembled genomes (MAGs), which were further used to investigate mobile genetic elements (MGE). Analyses of the reconstructed MAGs showed that, MAG 9 (Pseudomonas sp1.) contained the oriT gene (origin of transfer gene) needed for transferring virulent factors. Although the presence of Pseudomonas is common in raw cow's milk, pasteurization treatment reduces their survivability. Nevertheless, attention should be paid on Pseudomonas spp. due to their intrinsic resistance to antibiotics and their capability of transferring virulent factors to other bacteria.

https://www.sciencedirect.com/science/article/abs/pii/S0168160521000040