| Bacterial adhesion is a ubiquitous and very important biological phenomena.The surface biochemical structure of the bacteria can occured specific adhesion with mucus or epithelial cells. Adhesion plays an important role in in the virulence of pathogens. The pathogens only the first adhere to the surface of the host, they can further infect and cause the body pathogenic.Adhesion of pathogens has become an important aspect of pathogen virulence. In recent years, the adhesion mechanisms of pathogenic bacteria have attracted extensive attention.In order to get a better understanding of adhesion of pathogenic Vibrio fluvialis to the gill mucus of Pseudosciaena crocea, the influences of culture condition, bacterial density, incubation time, incubation temperature, pH, cation, carbohydrate and culture stage, antibody treatment on the bacterial adhesion were investigated by using a new detecting method--indirect ELISA, successfully established by modifying various conditions. The results show that the conditions of maximum adhesion of Vibrio fluvialis in the TSB culture conditions:25℃to30℃, pH6, salinity5to15, incubated time150min. Divalent cations and most carbohydrates can promote the adhesion of V.fluvialis on cell. Antibodies and starvation will significantly reduce bacterial adhesion.In this study, on the basis of the results mentioned above, transposon(mini-Tn10) which can random inserted mutagenesis was used to construct the mutants. Mutant library of V.fluvialis1.1608was generated and206mutants were screened and seven mutants stably exhibited defective in adhesion formation. Tail-PCR amplified the flanking sequences of the transposon insertion site in the3mutants. The results show that only the flanking sequence of insertion site in the mutant C99show a high identity (89%) with anaerobic nituic oxide reductase transcription of Vibrio.sp.EJY3in chromosome2from1176961bp to1179932bp. However, flanking sequences of the insertion site in mutant C43and C94can’t find homologous sequences. To further understand norR gene in adhesion and virulence-related functions of V.fluvialis, the growth, chemotaxis, biofilm formation and other biological characteristics of mutant C99were investigated. The results showed that the growth, chemotaxis, and biofilm formation of the mutant C99were significantly lower than the wild strain, which suggested that disruption the gene norR may affect bacterial growth, chemotaxis, and biofilm formation and maybe even affect the virulence of V.fluvialis. |
