| 钟小飘,陈政思,李一,等.珍珠龙胆石斑鱼(Epinephelus fuscoguttatus♀×E.lanceolatu♂) 源美人鱼发光杆菌的分离鉴定与致病性分析[J].广东农业科学,2025,(4-5):- |
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珍珠龙胆石斑鱼(Epinephelus fuscoguttatus♀×E.lanceolatu♂) 源美人鱼发光杆菌的分离鉴定与致病性分析 |
| Isolation, identification and pathogenicity analysis of Photobacterium from pearl gentian grouper (Epinephelus fuscoguttatus♀×E.lanceolatu♂) |
| 投稿时间：2024-11-18 修订日期：2025-03-30 |
| DOI： |
| 中文关键词: 石斑鱼 美人鱼发光杆菌 药敏试验 回归感染 致病性 载菌量 毒力基因 |
| 英文关键词: grouper(Epinephelus fuscoguttatus♀×E.lanceolatu♂) photobacterium drug susceptibility test recurrent infection pathogenicity bacterial load virulence gene |
| 基金项目:国家自然科学基金(U20A2065,32002426,32073006)；广东省科技计划项目(2023B0202010016) |
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| 中文摘要: |
| 【目的】近年来水产养殖病害频繁发生，而石斑鱼患美人鱼发光杆菌却鲜有报道。分离鉴定患病珍珠龙胆石斑鱼的病原菌，并对该菌的类型、生理生化特性、药敏特性和致病性展开研究，为其病害防控提供理论基础。【方法】在湛江市某养殖场发现患病石斑鱼，无菌条件下从脾脏中分离得到1株优势菌，对该菌株展开形态学观察、16SrRNA基因序列分析及生理生化特性鉴定，进一步开展药敏试验、回归感染试验，测定脾脏及肝脏载菌量，并检测其毒力基因。【结果】根据分离菌株的形态学观察、16SrRNA序列分析和生理生化鉴定，确定菌株OR539201为美人鱼发光杆菌美人鱼亚种（Photobacterium damselae subsp.damselae）。该菌株在LB平板上呈现圆形菌落，形态圆润、饱满，光滑均匀。在16SrRNA系统发育树中，与美人鱼发光杆菌聚为一支，亲缘关系最近，同为弧菌属。生化鉴定结果显示，氧化酶活性、尿素、赖氨酸等反应呈现阳性，水杨苷、甘露醇等反应呈现阴性。药敏试验结果表明分离菌株对不同药物呈现出不同程度的耐药性，对多粘菌素、丁胺卡那、头孢曲松等中等敏感，对氧氟沙星、环丙沙星、头孢氨苄等13种抗生素敏感。回归感染试验结果表明，该菌对石斑鱼具有致病性，感染后出现体表溃疡、红色增生结节、肝脏肿大、肠道糜烂、脑部红肿等与自然发病相同的症状。菌量测定结果显示，实验组脾脏及肝脏菌量明显高于对照组。毒力基因检测结果显示，该致病菌具有磷脂酶活性相关基因（plpV）。【结论】本研究成功从患病石斑鱼中分离得到一株潜在致病菌，鉴定为美人鱼发光杆菌。该菌株具有明显致病性，能够入侵健康石斑鱼免疫器官，诱发明显病变，引起鱼类死亡。 |
| 英文摘要: |
| Abstract: 【Objective】 In recent years, aquaculture diseases have become increasingly prevalent; however, there is limited literature on luminous Bacillus mermaidis infections in groupers. This study aimed to isolate and identify the pathogenic bacteria from diseased groupers, characterizing their types, physiological and biochemical properties, drug sensitivity profiles, and pathogenicity, thereby providing a theoretical foundation for disease prevention and control. 【Methods】 Diseased Epinephelus specimens were collected from a farm in Zhanjiang City. Under aseptic conditions, a dominant bacterial strain was isolated from the spleen. The strain underwent morphological observation, 16S rRNA gene sequence analysis, and physiological and biochemical identification. Further analyses included drug susceptibility testing and regression infection experiments, with bacterial loads quantified in the spleen and liver，and detect its virulence genes【Results】 Morphological observation, 16S rRNA sequence analysis, and physiological and biochemical identification confirmed that strain OR539201 belonged to Photobacterium damselae subsp. damselae. On LB agar, the strain formed circular colonies with rounded, full, and smooth morphology. Phylogenetic analysis of the 16S rRNA gene revealed a close relationship between this strain and Luminobacter mermaideni, both belonging to the genus Vibrio. Biochemical tests indicated positive reactions for oxidase, urea, glycerol, and lysine, while salicin, aesculin, and mannitol reactions were negative. Drug sensitivity testing revealed varying degrees of resistance to different antibiotics, with moderate sensitivity to polymyxin, butanacana, ceftriaxone, and sensitivity to 13 antibiotics including ofloxacin, ciprofloxacin, and cefalexin. Regression infection experiments demonstrated that the bacteria caused significant pathogenic effects in groupers, resulting in symptoms such as surface ulcers, red proliferative nodules, liver enlargement, intestinal erosion, and brain redness, consistent with natural disease manifestations. Bacterial counts in the spleen and liver of the experimental group were significantly higher than those in the control group, confirming the presence of luminous Bacillus mermaidis,The virulence gene detection results show that the pathogenic bacteria possess the gene (plpV) related to phospholipase activity.【Conclusion】 This study successfully isolated and identified a potential pathogen from diseased groupers, identified as Luminous Bacillus mermaidis. The strain exhibited significant pathogenicity, capable of invading immune organs in healthy groupers, inducing severe lesions, and causing fish mortality. |
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