犊牛腹泻症的病原学检测及鉴定

doi:10.6048/j.issn.1001-4330.2024.06.028

新疆农业科学 ›› 2024, Vol. 61 ›› Issue (6): 1535-1543.DOI: 10.6048/j.issn.1001-4330.2024.06.028

犊牛腹泻症的病原学检测及鉴定

苗书魁(), 王俊伟(), 陆桂丽, 汪萍, 王杰, 魏玉荣, 魏婕, 米晓云, 沙依兰·卡依扎

新疆畜牧科学院兽医研究所(新疆畜牧科学院动物临床医学研究中心)/新疆动物疫病研究重点实验室,乌鲁木齐 830013

收稿日期:2023-11-03 出版日期:2024-06-20 发布日期:2024-08-08
通信作者: 王俊伟(1985-),男,山西人,高级实验师,硕士,研究方向为动物疫病防控,(E-mail)1505115475@qq.com
作者简介:苗书魁(1983- ),男,甘肃人,正高级实验师,硕士,研究方向为动物病毒病原学及免疫学,(E-mail) 903076760@qq.com
基金资助:
新疆维吾尔自治区自然科学基金资助项目(2021D01A60);新疆维吾尔自治区奶产业技术体系资助(XJARS-11)

Etiological detection and identification of calf diarrhea

MIAO Shukui(), WANG Junwei(), LU Guili, WANG Ping, WANG Jie, WEI Yurong, WEI Jie, MI Xiaoyun, Shayilan Kayizha

Xinjiang Key Laboratory of Animal Infectious Diseases/Institute of Veterinary Medicine, Xinjiang Academy of Animal Sciences, Urumqi 830013, China

Received:2023-11-03 Published:2024-06-20 Online:2024-08-08
Correspondence author: WANG Junwei(1985-), male, master, senior experimentalist, research direction: Prevention and control of animal epidemic, (E-mail)E-mail
Supported by:
Project of Natural Science Foundation of Xinjiang Uygur Autonomous Region(2021D01A60);Supported by the earmarked fund for XJARS(XJARS-11)

摘要/Abstract

摘要：

【目的】研究新疆某规模化牛场犊牛腹泻症的主要病原。【方法】采用RT-PCR方法,检测14份腹泻犊牛粪便中的病毒,对4头死亡犊牛肝组织进行细菌分离培养及鉴定、小鼠致病性和药物敏感性试验。将分离的致病菌扩增16S rDNA基因片段并测序,并将其结果在NCBI中用BLAST搜索同源序列,分析遗传进化特性。【结果】该牛场犊牛腹泻的病原为牛冠状病毒(Bovine coronavirus, BCoV)、牛轮状病毒(Bovine rotavirus, BRV)、大肠埃希氏杆菌(Escherichia coli, E.coli)和肺炎克雷伯氏菌(Klebsiella pneumoniae, K.pneumoniae)。14份粪便中,BCoV检出率为64.29%,BRV检出率为50.00%;4份肝组织中, E.coli检出率为100.00%,K.pneumoniae检出率为50.00%。分离的2种致病菌E.coli和K.pneumoniae对阿米卡星敏感,对其它16种抗生素均耐药。【结论】该牛场引起犊牛腹泻的主要病原为BCoV、BRV、E.coli和K.pneumoniae,存在病毒与细菌的混合感染。选择阿米卡星药物防治E.coli和K.pneumoniae。

关键词: 犊牛腹泻; 病原学; 检测; 鉴定

Abstract:

【Objective】 This article aims to identify the main pathogens of calf diarrhea in a large-scale cattle farm in Xinjiang. 【Methods】 Virus detection was performed on 14 diarrhea calf feces using RT-PCR. Besides, bacteria were isolated from the liver tissues of four dead calves, cultured and identified, and mouse pathogenicity and drug sensitivity tests were conducted on these bacteria. Afterwards, the 16 S rDNA gene fragment of the isolated pathogenic bacteria was amplified and sequenced, and the sequencing results were searched for homologous sequences using BLAST in NCBI for genetic evolution analysis. 【Results】 The pathogens of diarrhea in the cattle farm were Bovine coronavirus (BCoV), Bovine rotavirus (BRV), Escherichia coli (E.coli), and Klebsiella pneumoniae (K.pneumoniae). Among the 14 feces, the detection rate of BCoV was 64.29%, and the detection rate of BRV was 50.00%; Among the four liver tissues, the detection rate of E. coli was 100.00%, and the detection rate of K.pneumoniae was 50.00%. The two isolated pathogenic bacteria E.coli and K. pneumoniae were sensitive to amikacin and resistant to all 16 other antibiotics. 【Conclusion】 The main pathogens causing calf diarrhea in this cattle farm are BCoV, BRV, E.coli, and K.pneumoniae, with mixed infection of viruses and bacteria. Amikacin can be used as the preferred drug for the prevention and treatment of E. coli and K. pneumoniae.

Key words: calf diarrhea; etiology; detection; identification

中图分类号:

S858.23

苗书魁, 王俊伟, 陆桂丽, 汪萍, 王杰, 魏玉荣, 魏婕, 米晓云, 沙依兰·卡依扎. 犊牛腹泻症的病原学检测及鉴定[J]. 新疆农业科学, 2024, 61(6): 1535-1543.

MIAO Shukui, WANG Junwei, LU Guili, WANG Ping, WANG Jie, WEI Yurong, WEI Jie, MI Xiaoyun, Shayilan Kayizha. Etiological detection and identification of calf diarrhea[J]. Xinjiang Agricultural Sciences, 2024, 61(6): 1535-1543.

图/表 11

参考文献 31

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引物名称 Name of primers	引物序列(5'→3') Sequence of primers(5'→3')	片段大小 Fragment size (bp)	退火温度 Annealing temperature(℃)
BVDV-F	TACGAGATTTAGAAAAGT	373	50
BVDV-R	TACAGCAGAGATTTTCAG	373	50
BCoV-F	ACAACAGGATGGTACGAT	478	54
BCoV-R	GCCATAATTAACTTCATTCA	478	54
BRV-F	GGCTTTTAAACGAAGTCTTC	1 356	54
BRV-R	GGTCACATCCTCTCACTA	1 356	54
BEV-F	GAGCCCAGTGTTTTCTTTGATCTTT	732	52
BEV-R	AAAGTTGATCAATAAAGTGCTTGC	732	52
BNoV-F	GCTGTCAACTAAGACCAA	610	54
BNoV-R	ATCCTCAATGGCATTCAT	610	54

引物名称 Name of primers	引物序列(5'→3') Sequence of primers(5'→3')	片段大小 Fragment size (bp)	退火温度 Annealing temperature(℃)
BVDV-F	TACGAGATTTAGAAAAGT	373	50
BVDV-R	TACAGCAGAGATTTTCAG	373	50
BCoV-F	ACAACAGGATGGTACGAT	478	54
BCoV-R	GCCATAATTAACTTCATTCA	478	54
BRV-F	GGCTTTTAAACGAAGTCTTC	1 356	54
BRV-R	GGTCACATCCTCTCACTA	1 356	54
BEV-F	GAGCCCAGTGTTTTCTTTGATCTTT	732	52
BEV-R	AAAGTTGATCAATAAAGTGCTTGC	732	52
BNoV-F	GCTGTCAACTAAGACCAA	610	54
BNoV-R	ATCCTCAATGGCATTCAT	610	54

引物名称 Name of primers	引物序列(5'→3') Sequence of primers(5'→3')	片段大小 Fragment size(bp)	退火温度 Annealing temperature(℃)
27F	AGAGTTTGATCCTGGCTCAG	约1 500	55
1492R	GGCTACCTTGTTACGACTT	约1 500	55

引物名称 Name of primers	引物序列(5'→3') Sequence of primers(5'→3')	片段大小 Fragment size(bp)	退火温度 Annealing temperature(℃)
27F	AGAGTTTGATCCTGGCTCAG	约1 500	55
1492R	GGCTACCTTGTTACGACTT	约1 500	55

抗菌药物 Antibiotic	兽用/非兽用 Animal/non animal use	大肠埃希氏杆菌 E.coli	肺炎克雷伯氏菌 K.pneu moniae
四环素 Tetracycline	兽用	R	R
多西环素 Doxycycline	兽用	R	R
氨苄西林 Ampicillin	兽用	R	R
链霉素 Streptomycin	兽用	R	R
庆大霉素 Gentamicin	兽用	R	R
阿米卡星 Amikacin	兽用	S	S
环丙沙星 Ciprofloxacin	兽用	R	R
甲氧苄啶/磺胺甲噁唑 Trimethoprim/ sulfamethoxazole	兽用	R	R
头孢噻吩 Cefoxitin	兽用	R	R
氯霉素 Chloramphenicol	兽用	R	R
红霉素 Erythromycin	兽用	R	R
诺氟沙星 Norfloxacin	兽用	R	R
卡那霉素 Kanamycin	兽用	R	R
米诺环素 Minocycline	兽用	R	R
头孢曲松 Ceftriaxone	非兽用	R	R
妥布霉素 Tobramycin	非兽用	R	R
洛美沙星 Lomefloxacin	非兽用	R	R

犊牛腹泻症的病原学检测及鉴定

Etiological detection and identification of calf diarrhea

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参考文献 31

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细菌 Bacterium	犊牛(肝组织)编号Number of calf (liver tissue)													4份肝组织中细菌检出率 Detection rate of bacteria in 4 liver tissues(%)
	3			11			13			14			合计 Total
	菌落编号Colony number
		1	2	3	4	5	6	7	8	9	10	11			12
E.coli	+	+	+	+	+	+	+	+	-	+	-	-	4	100.00
K.pneumoniae	-	-	-	-	-	-	-	-	+	-	+	+	2	50.00

病原 Pathogens	犊牛编号Calf number															检出率 Detection rate(%)
病原 Pathogens	1	2	3	4	5	6	7	8	9	10	11	12	13	14	合计	检出率 Detection rate(%)
BVDV	-	-	-	-	-	-	-	-	-	-	-	-	-	-	0	0
BCoV	+	-	+	-	+	-	-	+	+	-	+	+	+	+	9	64.29
BRV	-	+	-	+	-	+	-	-	-	+	+	-	+	+	7	50.00
BEV	-	-	-	-	-	-	-	-	-	-	-	-	-	-	0	0
BNoV	-	-	-	-	-	-	-	-	-	-	-	-	-	-	0	0
E.coli			+								+		+	+	4	100.00
K.pneumoniae			-								-		+	+	2	50.00

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