Biological Function Of Pili-like Protein Ssu2101 In Streptococcus Suis Serotype 2

Streptococcus suis serotype 2 (S. suis 2) is an important zoonotic pathogen associated with a wide range of diseases in pigs, including septicaemia, meningitis, pneumonia, endocarditis, arthritis and even sudden death. Occasionally, S. suis 2 can also be transmitted to human beings through wound contamination,posing the serious threat to the pig-breeding industry and the correlation jobholders. Furthermore, two major emerging infectious disease outbreaks of S. suis 2 in China (one in Jiangsu Province, 1998, and the other in Sichuan Province, 2005), raised considerable international concerns among the public health professionals. A key feature of these two Chinese outbreaks is the prevalence of streptococcal toxic shock syndrome (STSS) manifesting itself as acute high fever, multiple organ failures, short course of disease and high lethality. Therefore, control of S. suis 2 infection is becoming an urgent problem, because the pathogenic mechanisms of S. suis 2 are still unclear, coupled with the emergence of multiple drug-resistant strains. For prevention and control of S. suis 2 there are no effective drugs and therapentic measures, thus speeding up the study of pathogenesis and vaccine candidate is needed urgently.In the last few years, pili were described in highly prevalent gram-positive human pathogens, including Streptococcus pyogenes, Streptococcus agalactiae and Streptococcus pneumoniae. These structures are considered as virulence factors playing key roles in the adhesion and invasion process and in pathogensis. Moreover, pilus subunits are effective immunoprotecitive agents and excellent vaccine candidates for the prevention of infections by a number of pathogens. Bioinformatics analysis of the whole genome of S. suis 2 Chinese high virulent strain 05ZYH33 found two gene clusters similar to pilus-associated gene clusters of some Gram-positive bacteria in genetic organization phenomena, designated srtBCD and srtF clusters, which srtBCD cluster exixts only in the virulent strain. In this study, recombinant BP (backbone protein) of SSU2101 was prokaryotic expressed and purified. Western blot suggested that this protein can react strongly with convalescent-phase sera from pigs infected by S. suis 2, which indicated that S. suis 2 could express BP during the infection process and elicit specific antibodies. Animal infection experiments demonstrated that recombinant BP protected mice from lethal challenge with S. suis 2 infection, which suggested that recombinant BP might be a candidate component of S. suis 2 protein vaccine.Based on the principle of homologous recombination, we constructed gene knock-out mutant of the pilus backbone gene in S. suis 2 virulent strain 05ZYH33. PCR analysis, Southern hybridization, RT-PCR and Flow cytometry analysis showed that the target gene was replaced by SpcR cassette; Analysis of biological characteristics showed that there were no differences in mycelial morphology, hemolytic activity and dyeing properties between the mutant and the wild type strain 05ZYH33; Cell adherence assays suggested that BP might be dispensable during the bacterial adhesion to host cells. Virulence assays with murine model confirmed that the mutant was significantly less virulent than the wild type strain. The results suggested that BP played an important role in the pathogenesis, which laid the foundation for the study of the mechanisms of pilus assembly and biological function in the S. suis 2.