内蒙古地区牛呼吸道感染的主要病原菌及支原体分离鉴定及耐药性和毒力基因检测

doi:10.16431/j.cnki.1671-7236.2025.03.034

摘要/Abstract

摘要： 【目的】了解内蒙古地区某规模化牧场牛呼吸道疾病主要致病菌及支原体的感染情况、耐药性及毒力基因携带情况，为该牧场牛呼吸道疾病的防治提供用药依据。【方法】采集死亡牛的肺脏组织进行病原菌分离培养、生化鉴定、16S rRNA基因测序、PCR鉴定、药敏试验、耐药基因及毒力基因检测等。【结果】分离得到在TSA平板上显示圆润、灰白色、透明或半透明菌落，特异性基因及血清型基因扩增符合A型多杀性巴氏杆菌、A6型溶血性曼氏杆菌预期条带的细菌各1株；在PPLO固体培养基出现肉眼可观察的小气泡，镜检呈煎蛋样，特异性基因扩增符合预期条带的牛支原体1株。16S rRNA测序比对结果发现，3种病原体分别与多杀性巴氏杆菌、溶血性曼氏杆菌和牛支原体相似性在99%以上。耐药性及毒力基因检测结果显示，多杀性巴氏杆菌、溶血性曼氏杆菌对氧氟沙星高度敏感，对卡那霉素、阿米卡星、庆大霉素等抗菌药物敏感；多杀性巴氏杆菌携带strA、strB、tetA、tetB、qnrA、bla_ROB耐药基因，携带exbB、exbD、ompA、psl等22种毒力基因；溶血性曼氏杆菌携带strA、strB耐药基因，携带gcp、gs60、lktC、tbpB 4种毒力基因；牛支原体对链霉素、红霉素、克林霉素耐药，对环丙沙星、阿米卡星和卡那霉素中介，对庆大霉素、氧氟沙星、恩诺沙星、四环素敏感，携带strA、strB、mph、msr-E耐药基因，携带VspX、VspY2、VpmA、P81毒力基因。【结论】该牧场病死牛呼吸道疾病为多杀性巴氏杆菌、溶血性曼氏杆菌及牛支原体混合感染所致，且分离菌株携带多种毒力基因，建议临床中治疗牛呼吸道疾病使用氧氟沙星作为主要治疗抗菌药物，不建议使用红霉素、克林霉素等抗菌药物。

关键词: 多杀性巴氏杆菌; 溶血性曼氏杆菌; 牛支原体; 分离鉴定; 耐药性; 毒力基因

Abstract: 【Objective】 This study was aimed to understand the infection,drug resistance and virulence genes of the main causative organisms and Mycoplasma of bovine respiratory diseases in a large-scale ranch in Inner Mongolia,and provide medication basis for the prevention and treatment of bovine respiratory diseases in this ranch.【Method】 The lung tissues of dead cattle were collected for isolation and culture of pathogenic bacteria,biochemical identification,16S rRNA gene sequencing,PCR identification,drug susceptibility test,drug resistance genes and virulence genes detection.【Result】 One strain of Pasteurella multocida type A and one strain of Mannheimia haemolytica type A6 were isolated from the TSA plate,which showed rounded,grayish-white,transparent or translucent colonies,and the amplification of specific and serotypic genes was in accordance with the expected bands.One strain of Mycoplasma bovis was isolated from the solid medium of PPLO,which appeared in the solid medium of PPLO,and showed the appearance of omelette in the microscopic examination,and the amplification of specific genes was in accordance with the expected bands.The 16S rRNA sequencing and aligning results showed that the three pathogens had similarities of over 99% with Pasteurella multocida,Mannheimia haemolytica and Mycoplasma bovis,respectively.The results of drug resistance analysis and virulence gene detection showed that Pasteurella multocida and Mannheimia haemolytica were highly sensitive to oxfloxacin,and sensitive to kanamycin,amikacin,gentamicin and other antimicrobials.Pasteurella multocida carried resistance genes strA,strB,tetA,tetB,qnrA and bla_ROB,and 22 virulence genes such as exbB,exbD,ompA and psl.Mannheimia haemolytica carried resistance genes strA and strB,and four virulence genes such as gcp,gs60,lktC and tbpB.Mycoplasma bovis was resistant to streptomycin,erythromycin,clindamycin,intermediary to ciprofloxacin,amikacin and kanamycin,and sensitive to gentamicin,ofloxacin,enrofloxacin and tetracycline,carrying resistance genes strA,strB,mph and msr-E,and the virulence genes of VspX,VspY2,VpmA and P81.【Conclusion】 The respiratory disease of sick and dead cattle in this ranch was caused by Pasteurella multocida,Mannheimia haemolytica and Mycoplasma bovis,and the isolated strains carried a variety of virulence genes,it was recommended to use oxfloxacin as the main therapeutic antimicrobial drug for the treatment of bovine respiratory disease in the clinic and the use of antimicrobial drugs such as erythromycin and clindamycin were not recommended.

Key words: Pasteurella multocida; Mannheimia haemolytica; Mycoplasma bovis; separation and identification; resistance; virulence genes

中图分类号:

尹凯雯, 毛伟, 曹金山, 孙月, 董海燕, 韩凯凡, 王博, 李培锋, 张志丹, 樊宏亮, 郭宇, 赵红霞. 内蒙古地区牛呼吸道感染的主要病原菌及支原体分离鉴定及耐药性和毒力基因检测[J]. 中国畜牧兽医, 2025, 52(3): 1328-1341.

YIN Kaiwen, MAO Wei, CAO Jinshan, SUN Yue, DONG Haiyan, HAN Kaifan, WANG Bo, LI Peifeng, ZHANG Zhidan, FAN Hongliang, GUO Yu, ZHAO Hongxia. Isolation and Identification of Major Pathogenic Bacteria and Mycoplasma Causing Bovine Respiratory Tract Infection and Detection of Drug Resistance and Virulence Genes[J]. China Animal Husbandry and Veterinary Medicine, 2025, 52(3): 1328-1341.

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