摘要
出身背景
肠杆菌是一组革兰氏阴性细菌，经常表现出延长的抗微生物耐药性（AMR），并参与耐药性基因向同一环境中存在的其他细菌物种的传播。由于碳青霉烯类抗生素对人类健康的影响以及新抗生素的缺乏，世界卫生组织（WHO）将碳青霉素耐药和ESBL产生列为关键因素。肠杆菌普遍存在，在一种健康方法下应对人类和动物的AMR时，必须检查环境在AMR生物体或抗微生物耐药性基因（ARGs）传播中的作用。动物粪便被认为是AMR细菌进入环境的重要来源，耐药基因可以在环境中积累。
方法
为了更好地了解第三代头孢菌素和氟喹诺酮耐药性基因在环境中分离株之间的传播，我们将全基因组测序（WGS）应用于从爱尔兰农场废水中分离的肠杆菌（79株大肠杆菌、1株阴沟肠杆菌、1例肺炎克雷伯菌和1株吉列尼柠檬酸杆菌） = 72）和之后（n = 10） 综合人工湿地（ICW）处理。使用MagNA Pure 96系统（Roche Diagnostics，Rotkreuz，Switzerland）提取DNA，然后在MiSeq平台（Illumina，Eindhoven，Netherlands）上使用v3化学作为300循环配对末端运行进行WGS。鉴定了AMR基因和点突变，并将其与表型结果进行比较，以更好地了解耐药性和耐药性传播的机制。
后果
在分离株中发现了多种头孢菌素和氟喹诺酮耐药性基因（移动遗传元件（MGE）和染色体突变），这些基因主要解释了表型AMR模式。在82个分离株中共鉴定出31种质粒复制子类型，其中一个子集（n = 24），在大肠杆菌分离株中鉴定。五个质粒复制子局限于阴沟肠杆菌分离株，两个局限于肺炎克雷伯菌分离株。鉴定了与应激、生存、调节、铁摄取分泌系统、侵袭、粘附和毒素产生等功能相关的毒力基因。
结论
我们的研究表明，抗微生物微生物（ARO）即使在废水处理后也可以持续存在，并可能将临床相关的AMR传播到环境中，最终对人类或动物健康构成风险。
Abstract 
Background
The Enterobacterales are a group of Gram-negative bacteria frequently exhibiting extended antimicrobial resistance (AMR) and involved in the transmission of resistance genes to other bacterial species present in the same environment. Due to their impact on human health and the paucity of new antibiotics, the World Health Organization (WHO) categorized carbapenem resistant and ESBL-producing as critical. Enterobacterales are ubiquitous and the role of the environment in the transmission of AMR organisms or antimicrobial resistance genes (ARGs) must be examined in tackling AMR in both humans and animals under the one health approach. Animal manure is recognized as an important source of AMR bacteria entering the environment, in which resistant genes can accumulate.
Methods
To gain a better understanding of the dissemination of third generation cephalosporin and fluoroquinolone resistance genes between isolates in the environment, we applied whole genome sequencing (WGS) to Enterobacterales (79 E. coli, 1 Enterobacter cloacae, 1 Klebsiella pneumoniae, and 1 Citrobacter gillenii) isolated from farm effluents in Ireland before (n = 72) and after (n = 10) treatment by integrated constructed wetlands (ICWs). DNA was extracted using the MagNA Pure 96 system (Roche Diagnostics, Rotkreuz, Switzerland) followed by WGS on a MiSeq platform (Illumina, Eindhoven, Netherlands) using v3 chemistry as 300-cycle paired-end runs. AMR genes and point mutations were identified and compared to the phenotypic results for better understanding of the mechanisms of resistance and resistance transmission.
Results
A wide variety of cephalosporin and fluoroquinolone resistance genes (mobile genetic elements (MGEs) and chromosomal mutations) were identified among isolates that mostly explained the phenotypic AMR patterns. A total of 31 plasmid replicon types were identified among the 82 isolates, with a subset of them (n = 24), identified in E. coli isolates. Five plasmid replicons were confined to the Enterobacter cloacae isolate and two were confined to the Klebsiella pneumoniae isolate. Virulence genes associated with functions including stress, survival, regulation, iron uptake secretion systems, invasion, adherence and toxin production were identified.
Conclusion
Our study showed that antimicrobial resistant organisms (AROs) can persist even following wastewater treatment and could transmit AMR of clinical relevance to the environment and ultimately pose a risk to human or animal health.

https://europepmc.org/article/pmc/pmc10073600