Lalak Anna, Wasyl Dariusz, Zając Magdalena, Skarżyńska Magdalena, Hoszowski Andrzej, Samcik Ilona, Woźniakowski Grzegorz, Szulowski Krzysztof
Department of Microbiology, National Veterinary Research Institute, Puławy, Poland.
Department of Microbiology, National Veterinary Research Institute, Puławy, Poland.
Vet Microbiol. 2016 Oct 15;194:69-73. doi: 10.1016/j.vetmic.2016.01.023. Epub 2016 Feb 2.
Resistance to β-lactams is considered one of the major global problems and recently it became the most frequently studied topic in the area of antimicrobial resistance. The study was focused on phenotypic and genetic characterisation of commensal Escherichia coli (E. coli), including those producing cephalosporinases, isolated from gut flora of healthy slaughter animals. E. coli were cultured simultaneously on MacConkey agar (MCA) and cefotaxime supplemented MCA. The isolates were confirmed with ONPG and indol tube tests as well as PCR targeting uspA gene. Microbroth dilution method was applied for determination of Minimal Inhibitory Concentrations and interpreted according to EUCAST epidemiological cut-off values. Cephalosporin resistance phenotypes were defined by E-tests (BioMerieux) and relevant gene amplicons from selected strains were sequenced. A total of 298 E. coli isolates with cephalosporin resistance (ESC) found in 99 ones, were obtained from 318 cloacal or rectal swabs deriving from broilers, layers, turkeys, pigs and cattle. Both extended spectrum β-lactamase (ESBL) and ampC-cephalosporinase resistance phenotypes were noted in all tested animal species but cattle. At least one of the analysed genes was identified in 90 out of 99 cephalosporin-resistant isolates: bla (n=44), bla (n=38), bla (n=33) and bla (n=12). None of the phenotypes was identified in nine isolates. Sequencing of PCR products showed occurrence of ESBL-genes: bla, bla, bla and ampC-gene bla. They were located on numerous and diverse plasmids and resistance transferability was proved by electroporation of bla and bla located on X1 plasmids. Detection of cephalosporin resistant E. coli confirms the existence of resistance genes reservoir in farm animals and their possible spread (i.e. via IncX1 plasmids) to other bacteria including human and animal pathogens. The identified genetic background indicates on ecological aspects of selection and dissemination of cephalosporin resistance in E. coli isolated from food-producing animals rather than its potential role for public health threats.
对β-内酰胺类抗生素的耐药性被认为是全球主要问题之一，最近它成为抗菌药物耐药性领域研究最频繁的课题。该研究聚焦于共生大肠杆菌（E. coli）的表型和基因特征，包括那些产生头孢菌素酶的菌株，这些菌株分离自健康屠宰动物的肠道菌群。大肠杆菌同时在麦康凯琼脂（MCA）和添加头孢噻肟的MCA上培养。通过ONPG和吲哚管试验以及靶向uspA基因的PCR对分离株进行确认。采用微量肉汤稀释法测定最低抑菌浓度，并根据欧盟CAST流行病学临界值进行解释。通过E试验（生物梅里埃公司）确定头孢菌素耐药表型，并对选定菌株的相关基因扩增子进行测序。从318份来自肉鸡、蛋鸡、火鸡、猪和牛的泄殖腔或直肠拭子中获得了298株对头孢菌素有耐药性的大肠杆菌分离株，其中99株发现有头孢菌素耐药性（ESC）。除牛外，在所有受试动物物种中均发现了超广谱β-内酰胺酶（ESBL）和AmpC-头孢菌素酶耐药表型。在99株对头孢菌素有耐药性的分离株中，90株鉴定出至少一种分析基因：bla（n = 44）、bla（n = 38）、bla（n = 33）和bla（n = 12）。9株分离株未鉴定出任何表型。PCR产物测序显示存在ESBL基因：bla、bla、bla和AmpC基因bla。它们位于众多不同的质粒上，通过电穿孔位于X1质粒上的bla和bla证明了耐药性的可转移性。对头孢菌素耐药大肠杆菌的检测证实了农场动物中存在耐药基因库，以及它们可能（即通过IncX1质粒）传播到包括人类和动物病原体在内的其他细菌。所确定的遗传背景表明了从产食动物分离出的大肠杆菌中头孢菌素耐药性选择和传播的生态方面，而非其对公共卫生威胁的潜在作用。
