一株牛蛙源金黄杆菌的分离鉴定及其致病特性

【背景】美国牛蛙养殖过程中病害问题非常突出,尤其是细菌性病害,其病原种类多、病原菌复杂多样、蔓延速度快、发病死亡率高,一直是牛蛙养殖过程中防控的难点。【目的】确定从患病牛蛙体内分离到的一株细菌NW1203的分类地位和致病性。【方法】无菌操作从牛蛙体内取样划线分离细菌,通过形态观察、生理生化试验、16S rRNA基因序列比对进行种属鉴定,通过人工感染、溶血性试验和病理切片观察分析其致病特性。【结果】经形态和生理生化鉴定及16S rRNA基因序列比对,菌株NW1203为金黄杆菌属细菌,与Chryseobacterium sp. F30的相似性达100%,进化树也显示该菌与金黄杆菌属细菌聚类;溶血性试验表明,菌株NW1203对绵羊、小鼠和牛蛙的血细胞都呈完全溶血;人工感染试验及感染病蛙的组织切片观察显示,菌株NW1203对牛蛙具有较强致病性,可引起牛蛙肝、肾、脾等主要组织严重病变,LD50为4.753×103 CFU/g。【结论】明确了菌株NW1203为牛蛙新病原,为牛蛙疾病的防控提供了理论依据。

Isolation, identification and pathogenic characteristics of Chryseobacterium sp. from Rana catesbiana

Background] Diseases, especially bacterial diseases have been a big challenge in American bullfrog (Rana catesbiana) farming. Due to a high diversity of these pathogens, rapid spread of the disease as well as high mortality rate, the bacterial disease in the bullfrog is difficult to prevention and control. Objective] In order to study the taxonomic status and pathogenicity, one bacterial strain NW1203 was isolated and identified from diseased R. catesbiana. Methods] The strain NW1203 was isolated from R. catesbiana by streaking plate method under aseptic conditions, followed by morphological observation, physiological, biochemical tests and 16S rRNA gene phylogenetic analyses to identify bacterial species. To investigate the pathogenicity of these bacteria, artificial infection followed by hemolytic test and histopathological examination were performed. Results] Based on the characterization of physiological, biochemical examination and 16S rRNA gene sequence alignment, the isolated bacteria NW1203 was identified as Chryseobacterium sp. with 100% sequences similarities to Chryseobacterium sp. F30. Furthermore, phylogenetic trees showed bacteria NW1203 was grouped with Chryseobacterium sp. Hemolytic tests showed the strain NW1203 exhibited strongly hemolytic to the erythrocyte of sheep, mice and bullfrog. Artificial infection test and tissue section observation of infected frogs indicated the strain NW1203 had strongly pathogenic to R. catesbiana with LD50 values of 4.753×103 CFU/g and could cause severe pathological changes in main tissues including liver, kidney and spleen. Conclusion] This study indicates that the bacteria strain NW1203 was a new pathogen causing bacterial disease of R. catesbiana and provides a theoretical basis for the prevention and control of bullfrog diseases.