肺炎克雷伯菌(K.肺炎)是一种机会性病原体,导致人类和动物严重感染。调查肺炎克雷伯菌的耐药模式和毒力谱,从健康和乳腺炎奶牛以及被褥中收集了887个样本,饲料,粪便,空气,饮用水,喷水,洗涤水,来自湖北五个奶牛场的牛奶杯拭子,中国,2019年和2020年。使用khe的PCR和16SrDNA测序分离和鉴定肺炎克雷伯菌。使用wzi分型和多位点序列分型(MLST)对肺炎克雷伯菌分离株进行基因型表征。使用针对17种抗菌剂的肉汤微量稀释确认了抗菌剂的耐药性,并通过PCR确定了耐药性和毒力基因。肺炎克雷伯菌的患病率为26.94%(239/887),分布在101个wzi等位基因型(199/239,83.26%)和100个序列型(STs)(209/239,87.45%)中,包括5个新的wzi等位基因型和25个新的STs。系统发育分析表明,从牛奶中分离的肺炎克雷伯菌,乳头拭子,饲料,和粪便被分类在同一克隆复合体中。通过与PubMLST数据库进行比较,至少有67种STs有在不同物种和地区传播的风险。有趣的是,已从人类中分离出60种STs。分离株对美罗培南和粘菌素高度敏感,但对氨苄青霉素(100%)耐药,硫化异恶唑(94.56%),头孢菌素(47.28%),链霉素(30.13%),等等。值得注意的是,在这项研究中发现多药耐药(MDR)率为43.93%。通过PCR,鉴定了68个抗菌素耐药性(AMR)基因中的30个;blaTEM的患病率,blaSHV,strA,strB,aadA1和aac(6')-Ib-cr超过50%。发现11个产生CTX-M的肺炎克雷伯菌。FIMH的检出率,mrkD,uge,wabG,entB,IutA,iron,ureA超过85%。这项研究加强了肺炎克雷伯菌在湖北食用动物中的流行病学重要性。环境型肺炎克雷伯菌的出现和传播可能对食品安全和公众健康构成潜在威胁。
Klebsiella pneumoniae (K. pneumoniae) is an opportunistic pathogen, which causes serious infections in humans and animals. To investigate the antimicrobial resistance pattern and virulence profile of K. pneumoniae, a total of 887 samples were collected from both the healthy and mastitis cows and the bedding, feed, feces, air, drinking water, spraying water, washing water, and milk cup swabs from five dairy farms in Hubei, China, during 2019 and 2020. K. pneumoniae was isolated and identified using PCR of the khe and 16S rDNA sequencing. A genotypic characterization was performed for K. pneumoniae isolates using wzi typing and multilocus sequence typing (MLST). Antimicrobial resistances were confirmed using broth microdilution against 17 antimicrobial agents and resistance and virulence genes were determined by PCR. The prevalence of K. pneumoniae was 26.94% (239/887) distributed in 101 wzi allele types (199/239, 83.26%) and 100 sequence types (STs) (209/239, 87.45%), including 5 new wzi allele type and 25 new STs. Phylogenetic analysis showed that K. pneumoniae isolated from milk, nipple swab, feed, and feces is classified in the same clone complex. By comparing with the PubMLST database, at least 67 STs have the risk of spreading in different species and regions. Interestingly, 60 STs have been isolated from humans. The isolates were highly sensitive to meropenem and colistin, but resistant to ampicillin (100%), sulfisoxazole (94.56%), cephalothin (47.28%), streptomycin (30.13%), and so on. Noteworthy, multidrug-resistant (MDR) rate was found to be 43.93% in this study. By PCR, 30 of 68 antimicrobial resistance (AMR) genes were identified; the prevalence rate of blaTEM, blaSHV, strA, strB, aadA1, and aac(6\')-Ib-cr was more than 50%. Eleven CTX-M-producing K. pneumoniae were found. The detection rate of fimH, mrkD, uge, wabG, entB, iutA, iroN, and ureA was over 85%. This study reinforces the epidemiological importance of K. pneumoniae in food-producing animals in Hubei. The emergence and spread of environmental MDR K. pneumoniae may pose a potential threat to food safety and public health.