奶牛乳房炎病原微生物20联核酸检测试剂盒(PCR-荧光探针法)的临床应用评估  

Clinical Application Evaluation of a 20-plex Nucleic Acid Detection Kit(PCR-Fluorescent Probe Method)for Bovine Mastitis Pathogenic Microorganisms

作　　者：胡秀花 王子华 黄鹏成 刘园园 毛君杰 王桂琴 马翀[1] 王少林[1,4] HU Xiuhua;WANG Zihua;HUANG Pengcheng;LIU Yuanyuan;MAO Junjie;WANG Guiqin;MA Chong;WANG Shaolin(College of Veterinary Medicine,China Agricultural University,Beijing 100193,China;Hunan Shengwei Animal Husbandry Biotechnology Development Co.,Ltd.,Changsha 410205,China;College of Animal Science and Technology,Ningxia University,Yinchuan 750021,China;Hainan Food Safety Monitoring and Testing Technology Innovation Center,Sanya Research Institute,China Agricultural University,Sanya 572025,China)

机构地区：[1]中国农业大学动物医学院,北京海淀100193 [2]湖南圣维动牧生物科技发展有限责任公司,湖南长沙410205 [3]宁夏大学动物科技学院,宁夏银川750021 [4]中国农业大学三亚研究院海南省食品安全监测及检测技术创新中心,海南三亚572025

出　　处：《中国兽医杂志》2024年第11期61-69,共9页Chinese Journal of Veterinary Medicine

基　　金：宁夏回族自治区重点研发计划项目(2021BBF02036);国家现代农业产业技术体系资助项目(CARS36)。

摘　　要：为了评估商品化的奶牛乳房炎病原微生物20联核酸检测试剂盒(PCR-荧光探针法)(后文简称为“检测试剂盒”)在临床应用中的适用性,本试验采用传统细菌分离培养、检测试剂盒、16S rRNA基因扩增子测序和药物敏感性试验对宁夏回族自治区9个牧场155份奶牛乳房炎奶样分别进行菌株鉴定和耐药性检测,并将检测试剂盒与其他3种检测结果进行比较和分析。结果显示,检测试剂盒与传统细菌分离培养在病原菌检出情况上基本保持一致,均主要检出了大肠杆菌、芽孢杆菌属和凝固酶阴性葡萄球菌(CNS)等;检测试剂盒与16S rRNA基因扩增子测序结果在菌属组成上一致性较高,均主要检出了假单胞菌属、链球菌属和葡萄球菌属等;药物敏感性试验结果显示,检出率较高的66株病原菌分离株对β-内酰胺类抗菌药物的耐药率普遍偏高;奶样中β-内酰胺酶耐药基因blaZ的检测试剂盒检出结果与同一奶样中分离菌株的药物敏感性试验结果一致率达58%(7/12)。综上所述,该检测试剂盒具有较高的准确度和病原覆盖率,可为牧场奶牛乳房炎主要病原的快速诊断和耐药基因(blaZ)监测提供有效的技术支持,从而降低抗菌药物的使用量,以减少牧场经济损失。To assess the clinical applicability of a commercially available 20-plex nucleic acid detection kit(PCR-fluorescent probe method)(hereinafter referred to as the"detection kit")for bovine mastitis pathogenic microorganisms,this study employed traditional bacterial isolation and culture,the detection kit,16S rRNA gene amplicon sequencing,and drug sensitivity test to identify strains and assess drug resistance in 155 mastitic milk samples from 9 farms in the Ningxia Hui Autonomous Region.The results of the detection kit were compared and analyzed against the other three methods.The findings indicated that the detection kit largely agreed with traditional bacterial isolation and culture in terms of pathogen detection,with both methods primarily identifying Escherichia coli,Bacillus spp.,and coagulase-negative staphylococci(CNS).The detection kit also showed a high degree of consistency with 16S rRNA gene amplicon sequencing in terms of genus composition,primarily detecting Pseudomonas,Streptococcus,and Staphylococcus spp.Drug sensitivity test revealed that the 66 most frequently isolated strains had generally high resistance rates toβ-lactam antibiotics.The detection results of theβ-lactamase resistance gene blaZ using the detection kit had a 58%(7/12)consistency rate with the antimicrobial susceptibility test results of the isolated strains from the same milk samples.In conclusion,the detection kit demonstrates high accuracy and broad pathogen coverage,offering effective technical support for the rapid diagnosis of major bovine mastitis pathogens and the monitoring of resistance genes(blaZ).This could contribute to reducing the use of antimicrobial agents and minimizing economic losses on farms.

关 键 词：奶牛乳房炎病原微生物20联核酸检测试剂盒(PCR-荧光探针法) 细菌分离培养 16S rRNA基因扩增子测序 药物敏感性试验 

分 类 号：S852.61[农业科学—基础兽医学]