美国牛蛙蝌蚪约氏不动杆菌的分离、鉴定及耐药性分析  

Isolation,identification and drug resistance analysis of Acinetobacter johnsonii from American bullfrog(Rana catesbeiana)tadpole

作　　者：丁戈野 李纯 徐杨 陈晓颖 张秋实 胡亮 邓新兰 刘小燕[1] 冯兴浪 李军华 DING Geye;LI Chun;XU Yang;CHEN Xiaoying;ZHANG Qiushi;HU Liang;DENG Xinlan;LIU Xiaoyan;FENG Xinglang;LI Junhua(College of Fisheries,Hunan Agricultural University,Changsha 41028,China)

机构地区：[1]湖南农业大学水产学院,湖南长沙410128

出　　处：《水产学报》2024年第5期129-138,共10页Journal of Fisheries of China

基　　金：国家自然科学基金(32173021);湖南省科技专项(2022sfq30,20231F18);湖南省教育厅重点项目(23A0196);湖南省大学生创新创业训练计划项目(S202310537085)。

摘　　要：为探明2023年3月初引起中国重庆市渝北区某牛蛙养殖场暴发疾病的原因,本实验从患病美国牛蛙蝌蚪肝脏和肠道中分离优势菌,采用形态学观察、生理生化实验、16S rDNA和rpoB基因序列测定、系统发育学分析进行鉴定。为确定该菌株的致病机理,对该菌株进行了人工感染实验、组织病理学观察、毒力基因检测、生长特性研究及药物敏感性实验。结果显示,在患病濒死美国牛蛙蝌蚪的肝脏和肠道中分离得到的其中一株优势菌命名为LT202303,经鉴定为约氏不动杆菌。该菌株携带OmpA、Omp34和OmpTsx等3种毒力基因,具有较强的耐盐性和在广泛的pH范围内生存的能力,对美国牛蛙蝌蚪的半数致死浓度为6.8×10^(6)CFU/mL,发病临床症状为肝脏红肿并伴有大量白色结节,肠道透明发黄。组织病理学观察发现该菌株可引起美国牛蛙蝌蚪肝脏和肠道的明显炎症反应和病灶性坏死。药敏实验结果显示,分离菌LT202303是一种产β-内酰胺酶的多重耐药细菌。本研究表明,约氏不动杆菌对牛蛙蝌蚪具有高致病力,可引起肝脏和肠道等多器官组织的病理损伤最终引发机体的损伤甚至死亡。本研究首次报道约氏不动杆菌感染引起的美国牛蛙蝌蚪传染病的暴发,为该疾病的诊断和防控提供理论参考。American bullfrogs(Rana catesbeiana)are a globally distributed invasive anuran species.They are extensively farmed in aquaculture facilities,particularly in countries like China,where they are highly valued as a protein-rich delicacy.In early March 2023,an outbreak of a serious infectious disease occurred in a bullfrog breeding farm in Yubei District,Chongqing,China.To elucidate the cause of the disease and find treatment options,this study isolated predominant bacteria from the liver and intestine tissues of diseased bullfrog tadpole.Morphological observation,physiological and biochemical experiments,16S rDNA and rpoB sequencing,and phylogenetic analysis were employed to identify the bacteria.To ascertain the pathogenic mechanism of these dominant bacteria,artificial infection experiments,histopathological observations,virulence gene detection,growth characteristic studies,and drug sensitivity testing were conducted.The results showed that a predominant strain isolated from the liver and intestine of the diseased bullfrog tadpoles,LT202303,was identified as A.johnsonii.It harbored a trio of virulence genes,OmpA,Omp34,and OmpTsx,exhibited strong salt tolerance and the ability to survive within a wide pH range,and had a half-lethal concentration of 6.8×10^(6)CFU/mL.Clinical symptoms included liver redness and swelling with large white nodules,and a transparent yellowing of the intestine.Histopathological observations indicated significant inflammation and focal necrosis in bullfrog tadpole liver and intestine.Drug sensitivity testing revealed that the isolated strain LT202303 is aβ-lactamase-producing multidrug-resistant bacterium.In summary,A.johnsonii has high pathogenicity to R.catesbeiana tadpoles,causing pathological damage to multiple organ tissues including the liver and intestine,ultimately resulting in diseases or even death of bullfrog tadpoles.This study reports,for the first time,an outbreak of R.catesbeiana tadpole infectious disease caused by A.johnsonii infection and provides theoretical reference

关 键 词：约氏不动杆菌 美国牛蛙 蝌蚪 分离鉴定 耐药性 

分 类 号：S947.2[农业科学—水产养殖]