林麝源肺炎克雷伯菌耐药性及毒力基因分析

doi:10.16431/j.cnki.1671-7236.2025.01.037

摘要/Abstract

摘要： 【目的】对广东地区某规模化林麝养殖场易导致成年林麝肺炎等疾病的肺炎克雷伯菌进行耐药性及毒力基因分析,为有效防控由肺炎克雷伯菌引发的疾病提供参考依据。【方法】采集广东某林麝场成年林麝粪便共81份,采用细菌分离培养、形态学观察、生化分析和PCR扩增16S rRNA和溶血酵素基因的方法分离纯化肺炎克雷伯菌。通过血清型鉴定、药敏试验和PCR扩增耐药基因和毒力基因等方法研究分离菌株特性。【结果】分离菌在麦康凯平板上形成粉红色、光滑、湿润的单菌落;革兰染色镜检显示,菌体呈红色的短杆状,可判断为革兰阴性杆菌。通过生化试验选取与肺炎克雷伯菌生化特性一致的菌株进行PCR扩增。16S rRNA基因PCR扩增结果显示,获得大小约为1 542 bp的条带。共33株分离株与GenBank数据库肺炎克雷伯菌16S rRNA基因核苷酸序列相似性＞98%,且特异性基因khe检测为阳性,即本研究从81份样品中成功分离鉴定出33株肺炎克雷伯菌。血清型检测结果显示,分离株中优势血清型是K57。药敏试验结果显示,分离株对青霉素类、氨基糖苷类、四环素类、磺胺类、氯霉素类、喹诺酮类、单环β-内酰胺类药物均有不同程度的耐药性;对头孢菌素类和碳青霉烯类药物敏感。检测的12种耐药基因中,bla_SHV和oqxA基因检出率均为96.97%;aac(6')-Ⅰb-cr、rmtB、tetA、tetM、sul1、sul2、oqxB和floR基因检出率分别为57.58%、51.52%、78.79%、84.85%、72.73%、66.67%、87.88%和51.52%;未检测出bla_KPC和bla_NDM基因。检测的9种毒力基因中,wabG、uge、mrkD、entB和ureA基因检出率较高,分别为90.91%、96.97%、96.97%、100%和100%;aerobactin和allS基因检出率较低,分别为6.06%和18.18%;未检出ybtA基因。【结论】本研究成功分离鉴定出33株林麝源肺炎克雷伯菌,优势血清型是K57,存在多重耐药情况,耐药基因和毒力基因携带率高。本研究结果可为林麝源肺炎克雷伯菌所致疾病的防控提供数据支持。

关键词: 林麝; 肺炎克雷伯菌; 耐药性; 毒力基因; 抗菌药物

Abstract: 【Objective】 The resistance and virulence genes of Klebsiella pneumoniae,which could easily cause pneumonia and other diseases of adult forest musk deer,were analyzed in a forest musk deer farm in Guangdong,so as to provide reference for effective prevention and control of diseases caused by Klebsiella pneumoniae. 【Method】 In this study,81 fresh feces of forest musk deer were collected from a forest musk deer ranch in Guangdong. Klebsiella pneumoniae was isolated and purified by isolation culture,morphological observation,biochemical analysis,and PCR amplification of 16S rRNA and hemolytic enzyme (khe) genes.The characteristics of isolates were studied by serotype identification,drug susceptibility test,drug resistance and virulence genes detection. 【Result】 The isolates formed pink,smooth and moist single colony on McConkey plate.The strain showed short red rod shape under the microscope after Gram-staining and was Gram-negative.Strains with biochemical characteristics consistent with Klebsiella pneumoniae were selected for PCR amplification.PCR amplification of 16S rRNA gene showed that a band with a size of about 1 542 bp was obtained.A total of 33 isolates showed a nucleotide similarity of over 98% with the 16S rRNA gene of Klebsiella pneumoniae in GenBank database,and the specific gene khe was detected as positive.Therefore,this study successfully isolated and identified 33 strains of Klebsiella pneumoniae from 81 samples.The serotype test results showed that the dominant serotype among the isolates was K57.The drug sensitivity results showed that the isolates exhibited varying degrees of resistance to penicillin,aminoglycosides,tetracyclines,sulfonamides,chloramphenicols,quinolones and monocyclic β-lactams.However,it was sensitive to cephalosporins and carbapenems.Among the 12 drug resistance genes detected,the detection rates of bla_SHV and oqxA genes were both 96.97%.The detection rates of aac(6')-Ⅰb-cr,rmtB,tetA,tetM,sul1,sul2,oqxB and floR genes were 57.58%,51.52%,78.79%,84.85%,72.73%,66.67%,87.88% and 51.52%,respectively.bla_KPC and bla_NDM genes were not detected.Among the 9 virulence genes detected,the detection rates of wabG,uge,mrkD,entB and ureA genes were 90.91%,96.97%,96.97%,100% and 100%,respectively.The detection rates of aerobactin and allS genes were 6.06% and 18.18%,respectively,and ybtA gene was not detected. 【Conclusion】 In this study,33 strains of Klebsiella pneumoniae were isolated and identified from forest musk deer.The dominant serotype was K57,which had multiple drug resistance and a high carrying rate of drug resistance genes and virulence genes.The results of this study could provide data support for the prevention and control of diseases caused by Klebsiella pneumoniae from forest musk deer.

Key words: Moschus berezovskii; Klebsiella pneumoniae; drug resistance; virulence genes; antibacterials

中图分类号:

S852.61

廖慧群, 赵美, 曾国辉, 苏仁伟, 邓衔柏. 林麝源肺炎克雷伯菌耐药性及毒力基因分析[J]. 中国畜牧兽医, 2025, 52(1): 411-421.

LIAO Huiqun, ZHAO Mei, ZENG Guohui, SU Renwei, DENG Xianbo. Analysis of Drug Resistance and Virulence Genes of Klebsiella pneumoniae from Moschus berezovskii[J]. China Animal Husbandry and Veterinary Medicine, 2025, 52(1): 411-421.

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