Alexa Oniciuc Elena A, Walsh Calum J, Coughlan Laura M, Awad Amal, Simon Cezara A, Ruiz Lorena, Crispie Fiona, Cotter Paul D, Alvarez-Ordóñez Avelino
Department of Food Hygiene and Technology, Universidad de León, León, Spain.
Teagasc Food Research Centre, Fermoy, County Cork, Ireland.
mSystems. 2020 Feb 18;5(1):e00723-19. doi: 10.1128/mSystems.00723-19.
Here, the role of the dairy-processing chain as a reservoir of antimicrobial resistance (AR) determinants and a source of novel biocontrol quorum-sensing inhibitors is assessed through a functional metagenomics approach. A metagenomic library comprising ∼22,000 recombinant clones was built from DNA isolated from raw milk, raw milk cheeses, and cheese-processing environment swab samples. The high-throughput sequencing of 9,216 recombinant clones showed that lactic acid bacteria (LAB) dominated the microbial communities of raw milk cheese, while Gram-negative microorganisms of animal or soil origin dominated the microbiota of raw milk and cheese-processing environments. Although functional screening of the metagenomic library did not recover potential quorum-sensing inhibitors, analysis using an in-house database built specifically for this study identified homologues to several genes encoding proteins with predicted quorum-quenching activity, among which, the QsdH hydrolase was the most abundant. screening of the library identified LAB, and especially , as a relevant reservoir of AR determinants in cheese. Functional screening of the library allowed the isolation of 13 recombinant clones showing an increased resistance toward ampicillin, which in all cases was accompanied by a reduced susceptibility to a wide range of β-lactam antibiotics. This study shows that the dairy-processing environment is a rich reservoir of AR determinants, which vary by sample source, and suggests that combining next-generation sequencing with functional metagenomics can be of use in overcoming the limitations of both approaches. The study shows the potential of functional metagenomics analyses to uncover the diversity of functions in microbial communities prevailing in dairy products and their processing environments, evidencing that lactic acid bacteria (LAB) dominate the cheese microbiota, whereas Gram-negative microorganisms of animal or soil origin dominate the microbiota of milk and cheese-processing environments. The functional and screening of the library allowed the identification of LAB, and especially , as a relevant reservoir of antimicrobial resistance (AR) determinants in cheese. Quorum-quenching (QQ) determinants were not recovered through the execution of wet-lab function-based screenings but were detected through sequencing-based analyses.
在这里，通过功能宏基因组学方法评估了乳制品加工链作为抗菌抗性（AR）决定因素的储存库和新型生物控制群体感应抑制剂来源的作用。从生牛奶、生牛奶奶酪和奶酪加工环境拭子样本中分离的DNA构建了一个包含约22,000个重组克隆的宏基因组文库。对9,216个重组克隆的高通量测序表明，乳酸菌（LAB）在生牛奶奶酪的微生物群落中占主导地位，而动物或土壤来源的革兰氏阴性微生物在生牛奶和奶酪加工环境的微生物群中占主导地位。尽管对宏基因组文库的功能筛选未发现潜在的群体感应抑制剂，但使用专门为此研究构建的内部数据库进行的分析鉴定出了几个编码具有预测群体猝灭活性蛋白质的基因的同源物，其中QsdH水解酶最为丰富。对文库的筛选确定LAB，尤其是，是奶酪中AR决定因素的相关储存库。对文库的功能筛选使得能够分离出13个对氨苄青霉素抗性增加的重组克隆，在所有情况下，这都伴随着对多种β-内酰胺抗生素敏感性的降低。这项研究表明，乳制品加工环境是AR决定因素的丰富储存库，其因样本来源而异，并表明将下一代测序与功能宏基因组学相结合可用于克服这两种方法的局限性。该研究显示了功能宏基因组学分析在揭示乳制品及其加工环境中普遍存在的微生物群落功能多样性方面的潜力，证明乳酸菌（LAB）主导奶酪微生物群，而动物或土壤来源的革兰氏阴性微生物主导牛奶和奶酪加工环境的微生物群。对文库的功能和筛选使得能够确定LAB，尤其是，是奶酪中抗菌抗性（AR）决定因素的相关储存库。通过基于湿实验室功能的筛选未发现群体猝灭（QQ）决定因素，但通过基于测序的分析检测到了它们。
