羊源溶血性曼氏杆菌的分离、鉴定以及毒力基因和耐药性检测

doi:10.12160/j.issn.1672-5190.2025.01.002

摘要/Abstract

摘要： [目的]对河南省开封市羊源溶血性曼氏杆菌进行分离、鉴定,分析其生物学特性,了解其耐药性和毒力基因的携带情况。[方法]从开封市8家养羊场采集患呼吸道疾病的羊只鼻咽拭子530份,使用胰蛋白胨大豆琼脂培养基（TSA）分离溶血性曼氏杆菌;通过生化反应、形态学观察和特异性基因LKT的检测对分离菌株进行鉴定;利用多重PCR对分离菌株进行分型;采用PCR检测分离菌株的毒力基因;采用K-B纸片扩散法检测分离菌株对12种抗菌药物的敏感性;选用BALB/C小鼠开展分离菌株的致病性试验。[结果]①在530份样品中共鉴定出52株溶血性曼氏杆菌,检出率为9.81%。②对分离株溶血性曼氏杆菌进行血清分型,有17株属于血清1型,占比32.69%;28株属于血清2型,占比53.85%;7株属于其他血清型,占比13.46%。③对溶血性曼氏杆菌携带11种毒力基因检测分析,全部都有检出,其中gcp、lktC、nanH阳性检出率较高,分别为73.08%、67.31%、65.38%,其他检出的毒力基因阳性率介于17.31%~51.92%。④所有菌株对头孢噻肟、头孢氨苄、头孢曲松比较敏感,敏感率分别为86.54%、78.85%、84.62%;其余药物敏感率介于17.31~51.92%,其中,红霉素、庆大霉素、氨苄西林敏感率低于20%。⑤小鼠致病性试验表明,血清1型和血清2型的溶血性曼氏杆菌在36 h内的致死率都为100%。[结论]开封地区羊鼻咽拭子中分离鉴定出的羊源溶血性曼氏杆菌主要具有2种血清型,携带多种毒力基因,具有较强的致病性。研究结果为开封市羊源溶血性曼氏杆菌病的合理用药和科学防治提供数据参考。

关键词: 羊, 溶血性曼氏杆菌, 分离, 鉴定, 毒力基因, 耐药性

Abstract: [Objective] To isolate and identify Mannheimia haemolytica from sheep in Kaifeng City, Henan Province, analyze its biological characteristics, and understand its drug resistance and the presence of virulence genes. [Methods] A total of 530 nasopharyngeal swabs were collected from sheep with respiratory diseases in 8 sheep farms in Kaifeng City. Mannheimia haemolytica was isolated using Tryptic Soy Agar （TSA）. The isolated strains were identified through biochemical reactions, morphological observations, and detection of the specific gene LKT. Multiplex PCR was used for serotyping of the isolates. PCR was employed to detect virulence genes in the isolates. The sensitivity of the isolates to 12 antimicrobial agents was tested using the Kirby-Bauer （K-B） disk diffusion method. Pathogenicity tests of isolates were conducted using BALB/c mice. [Results] ①A total of 52 strains of Mannheimia haemolytica were identified from the 530 samples, with a detection rate of 9.81%. ②Serotyping of the isolated Mannheimia haemolytica revealed that 17 strains belonged to serotype 1, accounting for 32.69%; 28 strains belonged to serotype 2, accounting for 53.85%; and 7 strains belonged to other serotypes, accounting for 13.46%. ③Detection of 11 virulence genes in Mannheimia haemolytica showed that all genes were detected. The detection rates of gcp, lktC, and nanH were relatively high, at 73.08%, 67.31%, and 65.38%, respectively, while the detection rates of the remaining virulence genes ranged from 17.31% to 51.92%. ④The isolated strains were highly sensitive to cefotaxime, cephalexin, and ceftriaxone, with sensitivity rates of 86.54%, 78.85%, and 84.62%, respectively. The sensitivity rates to the remaining drugs ranged from 11.54% to 46.15%, among which the sensitivity rates to erythromycin, gentamicin, and ampicillin were lower than 20%. ⑤Pathogenicity tests in mice showed that the lethality rates of Mannheimia haemolytica of serotype 1 and serotype 2 were both 100% within 36 hours. [Conclusion] The Mannheimia haemolytica isolated and identified from sheep nasopharyngeal swabs in the Kaifeng region primarily exhibited two serotypes, carried multiple virulence genes, and demonstrated high pathogenicity. The research results provide data references for the rational use of drugs and scientific prevention and control of Mannheimia haemolytica infections in sheep in Kaifeng City.

Key words: sheep, Mannheimia haemolytica, isolation, identification, virulence genes, drug resistance

中图分类号:

S852.612

刘亚辉, 杜娟. 羊源溶血性曼氏杆菌的分离、鉴定以及毒力基因和耐药性检测[J]. 畜牧与饲料科学, 2025, 46(1): 10-17.

LIU Yahui, DU Juan. Isolation, Identification, Detection of Virulence Genes and Drug Resistance of Mannheimia haemolytica from Sheep[J]. Animal Husbandry and Feed Science, 2025, 46(1): 10-17.

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