The Development And Application Of Rapid Diagnostic Kits For Detecting Pathogenic Bacteria In The Cultivated Eels

Eels (Anguilla spp.), are one of the most popular and valuable economic farmed fish. Eel aquaculture production in China ranks first in the world. European eel (Anguilla anguilla) and Japanese eel (A. japonica) are the main cultivated eels. Now American eel (A. rostrata) has also been cultivated in some areas. In intensive eel cultivation, the diseases occured are inevitable, and bacterial diseases are the main problem and often lead to great economic loss. The different bacteria are very different in the sensitiveness to the medicine. Therefore, to prevent or cure the bacterial diseases it is very important to develop diagnostic kits for rapid and accurate diagnose the species of pathogenic bacterium. In this study, we developed rapid diagnostic kits, including the 10 new made pathogens immune serums and 23 others made before in our laboratory. They have been used to detect pathogenic bacteria of eel on-site at different times and size of eels in different regions of Fujian Province and Guangdong Province. The results are as follows:1.Ten different pathogenic bacteria strains were used to immune the New Zealand rabbits to produce immune serums. Titer of each immune serum was detected using the method of slide agglutination. The results showed the titres of eight serums were above 1024, the titres of other two were 512 and 256, respectively.2. The cross-reaction was assayed by the method of slide agglutination between ten kinds of immune serums and 29 known strains of bacteria, including ten strains of bacteria used for producing the above immune serums. The results show that there are seven serums with no cross-agglutinate while there are three serums with cross-agglutinate. The highest agglutinating titer ratio between cross-strains and the strains for producing serums was up to 1/2, and the lowest was 1/8.3. The three cross-reactive serums were purified with HiTrap Protein-A HP column through the AKTApurifier-100 purification system. After the purification, no obvious agglutination is observed among 29 tested bacterial strains when tested by slide agglutination; the maximum recovery of sample is 30%, the minimum recovery of 7.5%; and the highest multiples of purification is 85-fold, the lowest multiples of 3.1.4.The rapid diagnostic kits have been applied to investigate different pathogenic bacteria in two species of cultivated eels from four regions of Fujian Province (Eastern, Southern, Western, Northern) and Taishan in Guangdong Province during different period of time (from April to June and from August to October). The results are as follows: 1) There are totally 12 different pathogenic bacteria detected in Anguilla anguilla from Fujian Province during the period of time. Although there are nine strains and ten strains of pathogens detected from April to June and from August to October respectively, there are only six pathogens detected at both the period of time. The results indicated that eels could be infected by different pathogenic bacteria during different period of time.2) From April to June, there are nine pathogenic bacteria detected in European eel from three regions of Fujian, in which only one pathogen detected simultaneously. And from August to October, a total of nine pathogens were detected in European eel from all of the four regions of Fujian, in which two pathogens were detected simultaneously. The results show different pathogenic bacteria could be found in different regions at the same time. 3) There are five pathogens detected at Taishan in Guangdong province from August to October, in which only three pathogenic bacteria are the same as detected in Japanese eel fromFujian province. The detection result show it was no significant difference of pathogen infection in Japanese eel at two areas. In addition, there were 12 strains of pathogenic bacteria detected from European eel, and 13 strains detected in Japanese eel, among which only three pathogens could be detected in both the specices of eel at the same time. These results show that pathogenic bacteria are significant different, because of each different species of eels.