Abstract：Abstract: Actinobacillus pleuropneumoniae (APP) is a very important respiratory pathogen for swine and causes great economic losses in pig industry worldwide. Biofilm formation contributes to full virulence in many bacterial pathogens. In our previous STM studies we found that the disruption of the hns gene encoding a histone-like nucleoid structuring protein (H-NS) could enhance biofilm formation of APP. To further investigate the effect of H-NS on APP biofilm formation, the 408 bp complete coding sequence of hns gene was amplified by PCR from the genomic DNA of APP serotype 1 strain 4074 and cloned into the prokaryotic expression vector pET-28c. The resultant recombinant plasmid pET-hns was transformed into Escherichia coli BL21 (DE3). A 19KDa of recombinant protein (rH-NS) was obtained by IPTG induction, and purified using the Ni-NTA agarose columns. Different concentrations of rH-NS were added into the culture medium of the hns transposal mutant strain 1-21 and its parental strain 4074, and biofilms were quantified using the microtiter plate biofilm assay. Without rH-NS in the medium, strain 1-21 formed obvious biofims, but strain 4074 did not. Supplemented with 0.1-0.3μM rH-NS in the culture medium, the biomass of biofilms formed by strain 1-21 continuously decreased, whereas that formed by strain 4074 was increased. No obvious changes could be observed when more than 0.4μM of rH-NS was supplemented in both strains. Our data indicated that H-NS negatively regulates biofilm formation of A. pleuropneumoniae in a dose-dependent manner.
