The Difference In Expression Levels Of Edwardsiella Tarda LuxS Gene In Different Growth Periods

Edwardsiella tarda is major pathogenic bacteria of aquaculture, after infection, many fish will appear systemic sepsis, the skin ulceration, internal organ necrosis, the whole body severe bleeding; the quorum sensing system(Quorum, sensing, QS) is an important signaling molecule for communication, it is the general language of communication between bacteria,The synthesis mechanism of Autoinducer-2 is only a QS system between gram positive bacteria and gram negative bacteria at the same time; AI-2’s synthetase Lux S is the key of AI-2 synthesis, as well as the inherent component of Activated Methyl Cycle, Lux S/AI-2 system mediated by QS in addition to participate in the induction group, also with the bacterial bioluminescence, plankton, and the phenomenon of plasmid transfer in biofilm formation,infection process and host cross communication and so on.The full-length of AI-2/Lux S of Edwardsiella tarda was cloned by PCR based on the sequence on NCBI, then characteristics and conservative structure of this protein-coding gene were analyzed using web database and bioinformatics tools. The lux S gene was obtained by PCR, its length was 516 bp, and the sequence was highly conversation in Edwardsiella tarda.it encoding a protein of 172 amino acids, with predicted molecular weight of about 19.00 k Da, PI is 5.993; the gene sequence analysis results showed that, lux S gene sequence of E.t CD and E.t 080813 the homology was 99.8%, and Edwardsiella ictaluri and Edwardsiella piscicida homology in more than 95%. NCBI amino acid sequence analysis showed that, the homology between E.t CD strain and E.t 080813 strain were 100%,and Edwardsiella ictaluri and Edwardsiella piscicida homology homology is above 94.8%.This study also conducted a prokaryotic express Lux S proteins, rabbit anti Lux S polyclonal antibody was prepared after purified, then the polyclonal antibody was used in the Western blot technology to detect the Lux S protein from different strains of Edwardsiella tarda, the results show that, the lux S gene was ubiquitous.The anti-rabbits serum was prepared after this protein was purified through prokaryotic expression. The expression level of lux S gene was analyzed during difference growth period using Western blot and the distribution of lux S gene in Edwardsiella tarda was further studiedby this technique. Results of Western blot analysis showed that Lux S expression level was the lowest in the lag phase and began increasing when entered index phase. It reached the peak in the late index phase and decreased in decline phase. To explore whether the specific Lux S is AI-2 dependent, we used the method of antibody neutralization to analyze the effect of antirabbits serum on the growth of Edwardsiella tarda. Moreover, antibody neutralization results showed that, it can elongate the growth plateau phase, but it has no significant effect on bacterial growth.The key gene of lux S was highly conserved, and Lux S/AI-2 was widely distributed among Edwardsiella tarda. The expression level of lux S gene was different during every growth period, expression of Lux S protein reached the highest level in the late index phase.