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从南京市某集贸市场所售6种淡水鱼体内分离细菌,根据细菌染色、培养特性及16SrRNA基因扩增等特点进行嗜水气单胞菌的鉴定,对鉴定阳性的菌株测定其溶血、溶蛋白特性,采用PCR技术调查毒力因子的基因分布情况,以斑马鱼作为实验动物研究了不同毒力基因型代表菌株的致病性。试验共分离得到39株嗜水气单胞菌,检出率为65.0%(39/60);检测气溶素(aer)、细胞毒性肠毒素(act)、细胞兴奋性肠毒素(alt)、丝氨酸蛋白酶(ahp)和温敏胞外蛋白酶(eprCAI)等5种胞外毒力因子的基因分布情况表明,以携带3种毒力基因的菌株比例最高。随机选取3个代表菌株进行斑马鱼的致病性试验,结果发现携带全部毒力因子的E7菌株,具有很强的溶血与溶蛋白特性,对斑马鱼的LD50为3.20×106 CFU/mL,显示出较强的致病性。表明临床健康鱼体内嗜水气单胞菌的检出率很高,且携带的毒力因子类型多样,具有潜在的致病性。
Bacteria were isolated from six kinds of freshwater fish sold in a mart in Nanjing, and identified according to the characteristics of bacterial staining, culture and amplification of 16S rRNA gene. The positive strains were tested for hemolysis and dissolution To investigate the gene distribution of virulence factors by PCR, zebrafish was used as experimental animal to study the pathogenicity of representative strains of different virulence genotypes. A total of 39 strains of Aeromonas hydrophila were isolated from the test, the detection rate was 65.0% (39/60). The levels of aer, cytotoxic enterotoxin, The distribution of five extracellular virulence factors, serine protease (ahp) and eprCAI, showed that the highest proportion of strains carrying the three virulence genes was found. Three representative strains were randomly selected for pathogenicity test of zebrafish. The results showed that the strain E7 carrying all the virulence factors had strong hemolysis and hemolysin properties, and the LD50 of zebrafish was 3.20 × 106 CFU / mL, indicating that A strong pathogenicity. It is indicated that the detection rate of Aeromonas hydrophila in clinical healthy fish is very high, and the types of virulence factors carried are diverse and have potential pathogenicity.