引用本文:	曹馨月,黄嘉怡,杜诗钰,侯天牧,项维,郜卫华,张付贤.一株大口黑鲈源ST-873型肺炎克雷伯菌(Klebsiella pneumoniae)的分离鉴定及生物学特性研究.海洋与湖沼,2023,3(3):836-847.

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*一株大口黑鲈源ST-873型肺炎克雷伯菌(Klebsiella pneumoniae)的分离鉴定及生物学特性研究*
曹馨月¹, 黄嘉怡¹, 杜诗钰¹, 侯天牧¹, 项维¹, 郜卫华¹, 张付贤^1,2,3
1.长江大学动物科学学院湖北荆州 434023;2.长江大学湿地生态与农业利用教育部工程研究中心湖北荆州 434023;3.长江大学涝渍灾害与湿地农业湖北省重点实验室湖北荆州 434023

摘要:

从患腹水病大口黑鲈(Micropterus salmoides)的头肾、脾脏、肝脏组织中分离到一株优势菌, 生长出圆形、边缘整齐的灰白色黏液状菌落; 染色镜检可见短粗、卵圆形、有荚膜的革兰氏阴性杆菌。结合形态、生理生化特征、16S rRNA基因序列分析鉴定为肺炎克雷伯菌(Klebsiella pneumoniae), MLST分析进一步确定其为ST-873型, 与ST-65型聚为一支; 该菌株携带aere、alls、wca和ybt四种毒力基因, 具有溶血活性; 人工感染大口黑鲈, 发现患病鱼呈现腹水等与自然发病类似的症状, 且病鱼内脏的分离菌株与攻毒菌株相同; 经统计, 其LD₅₀为3.4×10⁷CFU/mL, 具有中等生物被膜形成能力。耐药性分析结果显示, 该菌株携带bla-shv、sul2、aadA和tetB四种耐药基因, 对β-内酰胺类、氨基糖苷类、喹诺酮类、黏菌素类药物敏感, 对四环素类、磺胺类、林可酰胺类、利福霉素类耐药; 中药三七、款冬花对分离菌株有明显抑制作用。研究探明了大口黑鲈腹水病的主要病原, 可为鱼源肺炎克雷伯菌的诊断和防治提供科学依据和数据参考。

关键词: 大口黑鲈(Micropterus salmoides) 肺炎克雷伯菌(Klebsiella pneumoniae) ST-873型分离鉴定药敏分析中药

DOI：10.11693/hyhz20220900235

分类号:

基金项目:广东省重点领域研发计划资助项目,2021B0202050002号；湖北省现代农业技术体系项目,HBHZD-ZB-2020-005号；湿地生态与农业利用教育部工程研究中心开放基金项目,KFT202003号

附件

ISOLATION AND CHARACTERIZATION OF KLEBSIELLA PNEUMONIAE TYPE ST-873 FROM MICROPTERUS SALMOIDES

CAO Xin-Yue¹, HUANG Jia-Yi¹, DU Shi-Yu¹, HOU Tian-Mu¹, XIANG Wei¹, GAO Wei-Hua¹, ZHANG Fu-Xian^1,2,3

1.College of Animal Science, Yangtze University, Jingzhou 434023, China;2.Engineering Research Center for Wetland Ecology and Agricultural Utilization, Ministry of Education, Yangtze University, Jingzhou 434023, China;3.Hubei Key Laboratory of Waterlogging Disaster and Agricultural Use of Wetland, Yangtze University, Jingzhou 434023, China

Abstract:

A dominant bacterial strain was isolated from tissues of Micropterus salmoides with ascites disease. It grew into a gray colony with round and neat edges. Short, oval, capsular Gram-negative bacilli were observed under microscope. In terms of morphological, physiological and biochemical characteristics and 16S rRNA gene sequence analysis, the isolated strain was identified as Klebsiella pneumoniae. MLST (multilocus sequence typing) analysis confirmed that it was ST-873 type and merged with ST-65 type. The isolated strain carried four virulence genes, aere, alls, wca, and ybt, and showed hemolytic activity. After artificial infection of M. salmoides, it was found that the infected fish presented ascites and other symptoms similar to the natural diseases, and the isolated strains from the viscera of the infected fish were the same to the challenged strains. The LD₅₀ of the isolated strain was 3.4×10⁷ CFU/mL, showing moderate biofilm formation ability. The results of drug resistance analysis showed that the strain carried four resistance genes, bla-shv, sul2, aadA, and tetB 4, and was sensitive to β-lactam, aminoglycosides, quinolones and colistin, and resistant to tetracycline, sulfa, lincoamide, and rifamycin. In addition, traditional Chinese medicine Panax Notoginseng and Flos Farfarae had obvious inhibitory effect on the isolated strain in vitro. This study found out the main pathogen of ascites disease of M. salmoides, which provided a scientific basis and data reference for the diagnosis and control of K. pneumoniae in fish.

Key words: Micropterus salmoides Klebsiella pneumoniae ST-873 type separation and identification antimicrobial susceptibility test traditional Chinese medicine