草鱼肠道约氏不动杆菌的分离鉴定及药敏特性研究

研究草鱼肠道不动杆菌,为鱼类肠道细菌的分离鉴定与微生态机制研究积累资料。从草鱼肠道中分离纯化获得1株细菌,编号为SW-2,采用细菌形态观察、生理生化特性分析、16S rDNA克隆测序及系统发育树构建等进行研究。结果表明SW-2菌株为革兰氏阴性短杆菌,对葡萄糖、甘露糖、木糖、枸椽酸盐利用等均为阴性;PCR方法扩增SW-2菌株16S rDNA并亚克隆至pMD18-T载体,测序获得序列长度为1 503 bp,与约氏不动杆菌(Acinetobacter johnsonii)序列相似性最高为99.7%;进一步构建系统发育树显示与A.johnsonii亲缘关系最近,自然聚类为一支,从而确定为约氏不动杆菌(A.johnsonii);药敏试验结果显示SW-2菌株对庆大霉素、羧苄青霉素、阿莫西林、氨苄西林、卡那霉素等药物敏感。人工回归感染草鱼7 d后未出现发病死亡现象,但是否为条件致病菌有待进一步研究。

Isolation, Identification and Antibiotic Susceptibility of Acinetobacter johnsonii from the Intestine of Grass Carp

Acinetobacter bacteria are opportunistic pathogens, widely distributed in water and soil, and can infect humans, animals and fish. Reports in the literature have described the isolation of several Acinetobacter species from sturgeon, catfish, crucian carp and grass carp including A. junnii, A. bauman and A. xiamenensis. However, no study has been reported on the isolation and identification of Acinetobacter johnsonii in fish. In this study, a bacterial strain isolated from the intestine of grass carp and labeled SW-2 was identified through morphological examination, physiological and biochemical analysis, and phylogenetic tree analysis based on 16S rDNA sequencing. Simultaneously, an artificial infection study was carried out on healthy grass carp by injecting different concentrations of a suspension of the SW-2 strain. An antibiotic susceptibility test was carried out using 21 antibiotics. The SW-2 strain is gram-negative, rod shaped, milky white, moist with a smooth surface and regular edges. It cannot produce acid from glucose, mannose or xylose and the test for citrate utilization was negative. The 16S rDNA gene sequence of the SW-2 strain was amplified and sub-cloned with the pMD18-T vector. The length of the 16S rDNA sequence was 1,503 bp, 99.7% homologous to the 16S rDNA sequences of A. johnsonii. Phylogenetic tree analysis shows that the SW-2 strain clustered into one group with A. johnsonii. Combined with the morphological characteristics and physiological and biochemical characteristics of the colony, the SW-2 strain was conclusively identified as A. johnsonii. Antibiotic sensitivity results showed that the isolated strain is sensitive to gentamicin, penicillin, amoxicillin, ampicillin and kanamycin. There was no mortality in grass carp within 7 days of artificial infection, but further study should be carried out to determine if it is an opportunistic pathogen in fish. This study provides a scientific reference on the antibiotic sensitivity and intestinal microbial ecology of Acinetobacter spp. in fish.