梅花鹿源A型产气荚膜梭菌的分离鉴定及对小鼠的致病性分析

doi:10.11843/j.issn.0366-6964.2025.08.035

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (8): 3976-3984.doi: 10.11843/j.issn.0366-6964.2025.08.035

梅花鹿源A型产气荚膜梭菌的分离鉴定及对小鼠的致病性分析

曾圣鑫¹(), 宋承琦², 申恺源², 郝国鑫², 王雅琨², 王鑫², 王晓旭³, 刘志杰³, 刘永波², 刘永生², 杨顺利², 付志新²^,*()

1. 四川农业大学动物医学院, 成都 611130
2. 河北科技师范学院动物科技学院, 河北省预防兽医学重点实验室, 秦皇岛 066500
3. 中国农业科学院特产研究所, 长春 130112

收稿日期:2024-11-01 出版日期:2025-08-23 发布日期:2025-08-28
通讯作者: 付志新 E-mail:zengshengxin@126.com;fzxljl@163.com
作者简介:曾圣鑫(2000-)，男，四川宜宾人，硕士生，主要从事产气荚膜梭菌相关研究，E-mail：zengshengxin@126.com
基金资助:
“十四五”国家重点研发项目子课题(2023YFD1800704-02)

Isolation, Identification of a Deer Clostridium perfringens and Pathogenicity Analysis in Mouse

ZENG Shengxin¹(), SONG Chengqi², SHEN Kaiyuan², HAO Guoxin², WANG Yakun², WANG Xin², WANG Xiaoxu³, LIU Zhijie³, LIU Yongbo², LIU Yongsheng², YANG Shunli², FU Zhixin²^,*()

1. College of Veterinary Medicine, Sichuan Agricultural University, Chengdu 611130, China
2. Hebei Key Laboratory of Preventive Veterinary Medicine/College of Animal Science and Technology, Hebei Normal University of Science & Technology, Qinhuangdao 066500, China
3. Institute of Special Animal and Plant Sciences of Chinese Academy of Agricultural Sciences, Changchun 130112, China

Received:2024-11-01 Online:2025-08-23 Published:2025-08-28
Contact: FU Zhixin E-mail:zengshengxin@126.com;fzxljl@163.com

摘要/Abstract

摘要：

产气荚膜梭菌(Clostridium. perfringens)是危害动物养殖业的重要病原，养鹿业深受其害。本研究从临床患鹿肠内容物分离到一株C. perfringens菌株，运用16S rRNA基因序列比对、毒素型PCR及生化等方法进行菌株鉴定，并开展耐药性分析；通过腹腔注射对小鼠进行攻毒，记录其发病、死亡情况和组织病理学变化，建立小鼠感染模型。结果表明，分离株鉴定为A型C. perfringens，符合产气荚膜梭菌生化特征，对阿米卡星、多黏菌素B、链霉素、卡那霉素、克林霉素、复方新诺明几种抗生素表现出耐药特征。菌液攻毒小鼠精神萎靡，被毛凌乱，扎堆不动，剖检可见肝脏、脾脏肿大，肠壁变薄，小肠内有黄色内容物。该菌株半数致死量为7.99×10⁷ CFU·mL^-1。组织切片可见各组织器官充血、出血，炎性细胞浸润，小肠肠绒毛部分断裂脱落，结构不完整。本研究结果将为临床鹿梭菌防治提供指导，为C. perfringens鹿用疫苗研制奠定基础。

关键词: 产气荚膜梭菌, 鹿, 分离鉴定, 小鼠模型

Abstract:

Clostridium perfringens is an important pathogen that endangers the deer breeding industry. In this study, a strain of Clostridium perfringens was isolated from clinical samples obtained from deer using selective identification media. The strain was identified through 16S rRNA sequencing, toxin-type polymerase chain reaction (PCR), and biochemical assays. An analysis of antimicrobial resistance was also performed. To establish a murine infection model, mice were subjected to intraperitoneal injection with the isolated strain, and subsequent observations were made regarding incidence rates, mortality, and histopathological alterations. The findings indicated that the isolated strain was identified as Clostridium perfringens type A, aligning with the biochemical characteristics of Clostridium perfringens. Furthermore, this strain exhibited resistance to the antibiotics amikacin, polymyxin B, streptomycin, kanamycin, clindamycin, and cotrimoxazole. The mice of the experimental group exhibited poor vitality, characterized by unkempt fur and reduced mobility. Examination revealed hepatosplenomegaly, thinning of the intestinal wall, and the presence of yellow material within the small intestine. The median lethal dose was determined to be 7.99×10⁷ CFU·mL^-1. Histopathological analysis indicated hyperemia and hemorrhage across various tissues and organs, infiltration of inflammatory cells, partial villous atrophy and loss in the small intestine, and structural disintegration.

Key words: Clostridium perfringens, deer, isolation, mouse model

中图分类号:

S852.616.3

曾圣鑫, 宋承琦, 申恺源, 郝国鑫, 王雅琨, 王鑫, 王晓旭, 刘志杰, 刘永波, 刘永生, 杨顺利, 付志新. 梅花鹿源A型产气荚膜梭菌的分离鉴定及对小鼠的致病性分析[J]. 畜牧兽医学报, 2025, 56(8): 3976-3984.

ZENG Shengxin, SONG Chengqi, SHEN Kaiyuan, HAO Guoxin, WANG Yakun, WANG Xin, WANG Xiaoxu, LIU Zhijie, LIU Yongbo, LIU Yongsheng, YANG Shunli, FU Zhixin. Isolation, Identification of a Deer Clostridium perfringens and Pathogenicity Analysis in Mouse[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(8): 3976-3984.

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基因 Genes	引物序列(5′→3′) Primer sequences	片段长度/bp Product length
16S rRNA	F-AGAGTTTGATCCTGGCTCAG	1 465^[7]
16S rRNA	R- GGTTACCTTGTTACGACTT	1 465^[7]
cpa	F-GCTAATGTTACTGCCGTTGA	324
cpa	R-CCTCTGATACATCGTGTAAG	324
cpb	F-GCGAATATGCTGAATCATCTA	196
cpb	R-GCAGGAACATTAGTATATCTTC	196
etx	F-GCGGTGATATCCATCTATTC	655
etx	R-CCACTTACTTGTCCTACTAAC	655
itx	F-ACTACTCTCAGACAAGACAG	446
itx	R-CTTTCCTTCTATTACTATACG	446
NetB	F-GCTGGTGCTGGAATAAATGC	384
NetB	R-TCGCCATTGAGTAGTTTCCC	384
cpe	F-GGAGATGGTTGGATATTAGG	233
cpe	R-GGACCAGCAGTTGTAGATA	233

药物种类 Drug type	药物名称 Drugs	缩写 Abbreviations	药物含量/(μg·片^-1) Drug content	结果 Results
氨基糖苷类 Aminoglycoside	阿米卡星 Amikacin	AMK	30	R
	链霉素 Streptomycin	S	10	R
	卡那霉素 Kanamycin	KAN	30	R
	庆大霉素 Gentamicin	GM	120	S
β-内酰胺类 β-lactams	氨苄西林 Ampicillin	AMP	10	S
	头孢氨苄 Cefalexin	CN	30	S
	苯唑西林 Oxacillin	OX	1	R
	头孢唑林 Cefazolin	CZ	30	S
	头孢他啶 Ceftazidime	CAZ	30	S
	青霉素 Penicillin	PEN	10	S
四环素类 Tetracyclines	强力霉素 Doxycycline	DO	30	S
	米诺环素 Minocycline	MI	30	S
	四环素 Tetracycline	TET	30	S
多肽类 Polypeptide	万古霉素 Vancomycin	VAN	30	R
多肽类 Polypeptide	多黏菌素B Polymyxin B	PB	300 IU	S
氯霉素类 Chloram phenicols	氟苯尼考 Florfenicol	FFC	30	S
喹诺酮类 Quinolones	环丙沙星 Ciprofloxacin	CIP	5	I
林可酰胺类 Lincosamides	克林霉素 Clindamycin	CC	2	R
大环内酯类 Macrolides	红霉素 Erythromycin	E	15	R
碳青霉烯类 Carbapenem	亚胺培南 Imipenem	IPM	10	S
磺胺类 Sulfonamides	复方新诺明 Compound sulfamethoxazole	SXT	25	R

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梅花鹿源A型产气荚膜梭菌的分离鉴定及对小鼠的致病性分析

Isolation, Identification of a Deer Clostridium perfringens and Pathogenicity Analysis in Mouse

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