Identification For Common Antigens Of Three Vibrio Species By Immunoproteomics

In recent years, with the rapid development of aquaculture, diseases of farmed fish occur more frequently, which has caused tremendous economic losses for aquaculture industry. Bacterial diseases caused by Vibrio species are known to be the main reason for the mortality and Vibrio harveyi, Vibrio alginolyticus and Vibrio parahaemolyticus are proved to be the most important pathogens of marine fish in South China according to the formerly research on the epidemiology of Vibrio diseases.In this paper, the immuno-reaction proteins of V. harveyi, V. alginolyticus and V. parahaemolyticus recoginzed by rabbit anti-V. harveyi antibody have been identified by immunoproteomics method. Furthermore the common antigens of these three Vibrio species have been screened and the protective function of one of the common antigens has been studied.Immuno-reaction Proteins of V. harveyi: whole cell extract protein of V. harveyi through two-dimensional electrophoresis, Western Blot to get 2-DE of whole cell protein of V. harveyi map and 2-DE Western blotting of whole cell protein of V. harveyi map. Through Image Master 2D Platinum to match to get 45 protein sopts; through mass spectrometry identification we get 40 proteins. Although four pairs of protein spots in the gel get different locations, they are identified as the same protein; one protein spot can not be found the corresponding protein in the database; in addition, H17 and H19 are two subunits of the same protein.Immuno-reaction Proteins of V. alginolyticus: whole cell extract protein of V. alginolyticus through two-dimensional electrophoresis, Western Blot has got 2-DE of whole cell protein of V. alginolyticus map and 2-DE Western blotting of whole cell protein of V. alginolyticus map. Through Image Master 2D Platinum to match we get 40 sopts, through mass spectrometry identification we get 36 proteins. Although two pairs of protein spots in the gel have different locations, they are identified as the same protein; one protein spot can not be found the corresponding protein in the database; a protein spots has not been identified out; in addition, A24 and A29 are two subunits of the same protein.Immuno-reaction Proteins of V. parahaemolyticus: whole cell extract protein of V. parahaemolyticus through two-dimensional electrophoresis, Western Blot has got 2-DE of whole cell protein of V. parahaemolyticus map and 2-DE Western blotting of whole cell protein of V. parahaemolyticus map. Through Image Master 2D Platinum to match we get 58 sopts, Checking 17 spots out of them through mass spectrometry identification, we get 14 kinds of proteins. Although two pairs of protein spots in the gel have get the different locations, they are identified as the same protein; one protein spot can not be found the corresponding protein in the database.Common antigens of the three Vibrio species.: By ImageMaster 2D Platinum analyzes of three Vibrio sp. whole cell protein 2-DE maps, we found 10 protein spots. Each of the three Vibrio sp. of these 10 protein spots were identified separately by MS, we got five kinds of proteins. They are common antigens of the three Vibrio sp.: dihydrolipoamide dehydrogenase, succinate dehydrogenase flavoprotein subunit, elongation factor Ts, glyceraldehyde-3-phosphate dehydrogenase, triosephosphate isomerase.Relative percentage survival (RPS): Firstly, Dihydrolipoamide dehydrogenase was injected intraperitoneally into each E. coioides Cuvier; then three Vibrio sp. was injected intraperitoneally into each E. coioides Cuvier after 4 weeks. Cumulative mortality was recorded over two weeks. The different RPSs were 70% (V. harveyi), 60% (V. alginolyticus), and 70% (V. parahaemolyticus).