Studies On The The Extra Cellular Products (ECP)and The Flagellin Of The Pathogenic Vibrio Harveyi

The disease induced by Vibrio harveyi is one of the most serious bacterial disease that economically affects marine culture.The disease has made great mortality on aquaculture such as fish,shrimp and shellfish, inducing heavy lose of the marine culture.In this thesis,an attempt had been made to study the characterizations of the extra cellular products (ECP) and flagellin of Vibrio harveyi .The main results are shown as follows:1. A pathogenic strain of Vibrio harveyi named TS-628 was isolated in our laboratory from the farmed Epinephlus awoara. coioides suffering from ulcered syndrome in 2004.The ECP of the bacteria were found to show gelatinase, amylase, lipase, phospholipase, caseinase activities and haemolytic property. ECP samples obtained from the SDS-polyacrylamide gel eletrophoresis (SDS-PAGE) after different incubation periods produced almost the same bands. The molecular weights of the proteins in these ECP mainly ranged from 25.0 kDa to 66.2 kDa.The extracellular proteinase (ECPase) activities of V .harveyi and the other four Vibrio species (V. fischeri,V. parahaemolyticus,V. fluvialis,V. alginolyticus) were compared by SDS-gelatin-polyacrylamide gel electrophoresis (SDS-G-PAGE) staining with coomassie brilliant blue R-250. The results from the SDS-G-PAGE of these ECPases indicated that: (1) For V . harveyi, the activity and the sorts of ECPase under acidic condition (pH4)were higher and more than those under basic condition(pH9); (2) In the same case, ECPase of V . parahaemolyticus had the strongest activity and various styles as well as wide molecular weight among the five Vibrio species, and it seemed the lower proteinase activity and less sorts of the ECPase in V . harveyi.2. Cycles of differential centrifugation and the CsCl density-gradient ultracentrifugation were chosen to isolate and the purificate the flagellin from V. harveyi.The purified flagellin showed the typical filiform with negative stain under electronic microscopy. SDS-PAGE analysis showed that the purified flagellin was composed of 3 flagellin subunits with the estimated molecular weight of 43,43.5 and...