我国华南沿海海水养殖鱼类病原菌美人鱼发光杆菌(Photobacterium damselae)分离株的毒力基因和耐药性分析

Analysis of virulence genes and antibiotic resistance of Photobacterium damselae isolated from marine fishes in coastal South China

摘要:

[目的] 为探讨我国华南沿海海水养殖鱼类病原菌美人鱼发光杆菌的非典型毒力基因和耐药性的时空变化，解析影响其毒力和耐药性变化的可能环境因素，为该菌所引起的病害防控提供建议。[方法] 本研究以分离自我国广东和海南沿海患病海水鱼的35株美人鱼发光杆菌为研究对象，利用普通PCR扩增技术，分析5个非典型毒力基因在菌株中的分布情况，并采用纸片扩散法（K-B）分析菌株对15种抗生素的耐药性。[结果] 19株菌含有1–2个被检测的非典型毒力基因，尤其是hlyA和vvh的检出率均高于20%。35株菌多重耐药指数为0.00–0.67，表现出27种耐药谱，多重耐药率（菌株耐抗生素种类>3）达到60.00%，尤其对万古霉素、阿莫西林、麦迪霉素和利福平的耐药率均高于50%，但对庆大霉素、诺氟沙星、环丙沙星、氯霉素和氟苯尼考的耐药率均低于10%。非典型毒力基因含量和耐药性，呈现一定的随年份增加而增强以及海南>广东的时空差异，尤其是耐药性中的谱型丰富度、多重耐药率、某一菌株的最多耐药数量以及多重耐药指数，海南（分别是1.00，69.23%，10，0.32）均大于广东（分别是0.82，54.55%，9，0.25）。[结论] 美人鱼发光菌的非典型毒力基因可能是通过水平基因转移获得，海南与广东区域美人鱼发光菌毒力基因与耐药的差异主要受到温度和抗生素使用的影响。

Abstract:

[Objective] The aim of this research is to study the temporal and spatial changes of atypical virulence genes and drug resistance of the marine fish pathogen Photobacterium damselae in South China, and analyze the potential environmental factors driving the change of virulence and drug resistance in P. damselae, and consequently provide suggestions for disease prevention and control caused by P. damselae. [Methods] Based on PCR and Kirby-Bauer diffusion method, we analyzed the presence of 5 atypical virulence genes and the resistance to 15 tested antibiotics of 35 P. damselae strains isolated from diseased marine fishes in South China coastal area. [Results] The results show that:(1) 19 strains contained 1-2 atypical virulence genes, particularly, the detection rate of hlyA and vvh were higher than 20%; (2) 27 resistance types were observed in these 35 strains, the multi-antibiotic resistance index of was 0.00-0.67 with the multi-antibiotic resistance rate (one strain shows resistance to more than 3 antibiotics) up to 60.00%. Therein, more than 50% strains showed resistance to vancomycin, amoxicillin, midecamycin and rifampin, but less than 10% strains resisted to gentamicin, norfloxacin, ciprofloxacin, chloramphenicol and florfenicol; (3) The atypical virulence gene contents and drug resistance were marginally increased with the year grows, and more/stronger in Hainan than in Guangdong. Further, the richness of resistance types, multi-antibiotic resistance rate, the most resistant number in one strain, and multi-antibiotic resistance index were observed higher/more in Hainan (1.00, 69.23%, 10, 0.32, respectively) than in Guangdong (0.82, 54.55%, 9, 0.25, respectively). [Conclusion] These results indicated that atypical virulence genes probably be transferred into P. damselae by horizontal gene transfer and the variation of virulence and drug resistance of P. damselae isolated form Hainan and Guangdong are mainly affected by temperature and antibiotic pollution.