用点酶法检测鱼类致病性嗜水气单胞菌HEC毒素

DOT IMMUNOBINDING ASSAY (DIA) FOR THE DETECTION OF HEC TOXIN OF AEROMONAS HYDROPHILA

本试验建立的检测嗜水气单胞菌HEC毒素的点酶法,可在嗜水气单胞菌培养上清和临床病料中检出HEC毒素,可检出的最低水平为95ng/ml。用该法检测大肠杆菌、沙门氏菌、鳗弧菌、荧光假单胞菌、温和气单胞菌等的培养上清及患草鱼出血病的病鱼病料,均为阴性反应,仅与豚鼠气单胞菌、杀鲑气单胞菌新亚种及吕克氏耶尔森氏菌有轻度的交叉反应。对72株病鱼分离菌及33份病鱼组织的检测结果表明,点酶法与溶血试验的符合率分别为80.7%和90.9%。以上试验重复3次,结果完全一致。综上可见,用点酶法检测嗜水气单胞菌HEC毒素,无须作复杂的细菌鉴定,并能直接用于病料,是一个快速、敏感、特异、有效的方法,显示出广阔的应用前景。

Aeromonas hydrophila is a pathogen of fulminating infection occurred among freshwater fish in China since 1989. The bacterium can produce a toxin with MW 52. 5KD. This heat-labile protein possesses hemolytic activity、 enterotoxicity and cytotoxicity, and was proposed to name 'hec' toxin. The production of hec toxin is very closely related to its pathogenecity. We have developed a dot immunobinding assay (DIA) for Aeromonas hydrophila septicemia in fish. Assays were done on bacterial isolates and viscera samples from moribund carp and bream collected in Jiangsu province. The DIA was compared with hemolysis test (ML) which was commonly used to detect hec toxin. The agreement between the DIA and HL in the bacterial isolates(n = 72)and viscera samples(n=33)were 80 . 7% and 90. 9% respectively. The detectable level of hec toxin by DIA was 95ng/ml. It is concluded that the DIA may be useful for rapid and effective detection of hec toxin tan large samples.