大口黑鲈源豚鼠气单胞菌的分离鉴定及病理性分析  

Isolation,Identification and Pathological Analysis of Aeromonas caviae from Micropterus salmoides

作　　者：罗伟杰 史磊 黄瑜[1] 雷美华 喻大鹏 简纪常[1] 黄浦江 LUO Weijie;SHI Lei;HUANG Yu;LEI Meihua;YU Dapeng;JIAN Jichang;HUANG Pujiang(College of Fisheries,Guangdong Ocean University/Guangdong Key Laboratory of Aquatic Animal Disease Control and Healthy Culture/Key Laboratory of Control for Diseases of Aquatic Economic Animals of Guangdong Higher Education Institutes,Zhanjiang 524088,China;Shenzhen Fisheries Development Research Center,Shenzhen 518067,China;Shenzhen Institute of Guangdong Ocean University,Shenzhen 518120,China)

机构地区：[1]广东海洋大学水产学院/广东省水产动物病害防控与健康养殖重点实验室/广东省水产经济动物病害控制重点实验室,广东湛江524088 [2]深圳市渔业发展研究中心,广东深圳518067 [3]广东海洋大学深圳研究院,广东深圳518120

出　　处：《广东农业科学》2024年第7期66-78,共13页Guangdong Agricultural Sciences

基　　金：国家自然科学基金(32073006);中山市社会公益与基础研究项目(2021B2058)。

摘　　要：【目的】分析深汕特别合作区某鲈鱼养殖基地大口黑鲈(Micropterus salmoides)突发性死亡原因,探究其病原菌生物学特性,为大口黑鲈养殖中的疾病防控与诊治提供理论参考。【方法】从患病大口黑鲈肝脏、脾、肠等病灶组织中分离得到一株优势菌,命名为SZNH-S20230817;开展形态学特征观察,生理生化特性鉴定,人工感染试验,药物敏感性测试,最小抑菌浓度(MIC)和最小杀菌浓度(MBC)测定,16S rRNA序列、耐药基因及毒力基因的鉴定,分析其耐药性和病理。【结果】菌株SZNH-S20230817的序列与豚鼠气单胞菌(Aeromonas caviae)AP019195具有99.38%同源性;同时携带aer、act、lip和fl a 4个毒力基因;生化鉴定结果显示,该菌的吲哚、氧化酶、苯丙氨酸、甘露醇、木糖、精氨酸双水解酶和葡萄糖产酸等结果为阳性,确定菌株SZNH-S20230817为豚鼠气单胞菌。回归感染试验发现菌株SZNH-S20230817具有较强致病性,对大口黑鲈的半致死浓度(LC_(50))为1.39×10^(7)CFU/mL。药敏试验结果显示,该菌对头孢唑林、卡那霉素、四环素、万古霉素、氨苄西林和克林霉素等12种抗生素耐药,同时携带qnrS、catI、emrB、Sul1、rmtB等13种耐药基因。组织病理学分析表明,菌株SZNH-S20230817感染可引起鱼体脾脏细胞肿胀、游离、肠黏膜断裂破损、绒毛脱落、肾脏细胞坏死、炎症细胞浸润、肝脏细胞呈空泡化等。体外抑菌试验结果显示大黄和五倍子对该菌的MIC和MBC较低,其余7种中草药对该菌的抑菌能力较弱。【结论】豚鼠气单胞菌是引起本次大口黑鲈突发性死亡的病原菌,其同时携带aer、act、lip和fl a 4个毒力基因,对大口黑鲈的LC_(50)为1.39×10^(7)CFU/mL,对头孢唑林、卡那霉素、四环素等抗生素耐药。低剂量的大黄和五倍子能够有效抑制该菌生长,具备良好的预防和抑制效果。【Objective】The study was conducted to investigate the causes of sudden death of largemouth bass(Micropterus salmoides)in a bass culture base in Shenzhen-Shanwei Special Cooperation Zone,and reveal the biological characteristics of the Pathogen,with an aim to provide theoretical reference for disease prevention,treatment and prevention in breeding largemouth bass.【Method】A dominant bacterial strain,named SZNH-S20230817,was isolated from the liver,spleen,intestine and lesion tissues of the diseased largemouth bass,and the isolate was subsequently subjected to morphological characteristics observation,physiological and biochemical characteristics identification,artificial infection experiment,drug sensitivity test,MIC and MBC determination,16S rRNA sequencing,identification of resistance genes and virulence genes to analyse its drug resistance and pathology.【Result】The results showed that the extended 16S rRNA sequence of SZNH-S20230817 had a 99.38%homology with Aeromonas caviae AP019195,and also carried four virulence genes,including aer,act,lip and fl a.The results of physiological and biochemical identification showed that,indole,oxidase,phenylalanine,mannitol,xylose,arginine dihydrolysate and glucose acid production of the strain were positive,and the strain SZNH-S20230817 was identified as Aeromonas caviae.Regression infection revealed that the strain SZNH-S20230817 was highly pathogenic,with an LC_(50)of 1.39×10^(7)CFU/mL for largemouth bass.The results of drug sensitivity tests showed that the bacterium was resistant to 12 antibiotics such as cefazolin,kanamycin,tetracycline and vancomycin,ampicillin,and clindamycin,and also carried 13 resistance genes such as qnrS,catI,emrB,Sul1,rmtB and etc.Histopathological analysis showed that infection with the strain SZNH-S20230817 could cause swelling and freeing of splenocytes,fracture and rupture of intestinal mucosa,shedding of villi,necrosis of renal cells,infiltration of inflammatory cells and vacuolisation of hepatic cells in the fish.The results of

关 键 词：大口黑鲈 豚鼠气单胞菌 16S rRNA 药敏试验 毒力基因 组织病理 

分 类 号：S816.3[农业科学—饲料科学]