Screening Of Streptococcus Suis Virulence-associated Transcription Factors And Proinflammatory Mechanisms Of Proinflammatory Protein HP1717

Streptococcus suis(S.suis),which can cause meningitis,sepsis,pneumonia,endocarditis and arthritis in pigs,is an important infectious disease in pigs.The disease in swine occurs mainly in piglets up to 10 weeks of age.It causes significant economic losses worldwide,which have been estimated in the USA alone at over 300 million dollars per annum.This bacterium can not only infect pigs but also other mammals and birds.In the population,S.suis can be transmitted to humans through close contact with infected pig or pork by-products,and cause meningitis,arthritis,sepsis and Streptococcus toxic shock like syndrome(STSLS).Since it was first reported in 1968,it has become an important zoonotic disease that jeopardizes public health.From 2009 to 2013,the total number of human S.suis cases increased from more than 700 to 1,642,indicating that human S.suis infection is becoming more common.The pathogen is a gram-positive bacterium with a polysaccharide capsule(CPS),and 29 serotypes have been identified based on the CPS antigen.Among them,serotype 2 is the most frequently isolated serotype from diseased pigs in most countries and is most commonly involved in human infections.Two large-scale human infections of S.suis were reported in China,which caused public concern.Unlike in the past,patients developed severe STSLS which was previously thought to be a symptom of Staphylococcus aureus and Streptococcus pyogenes.This suggests that S.suis may have evolved a new virulent strain with more serious harm.In this study,we screened the virulence-related transcription factors from the genome of S.suis and studied them in detail.At the same time,we studied the pro-inflammatory mechanism of a protein in S.suis.1.Research on virulence-related transcription factorsA total of 89 transcription factors were predicted in the S.suis genome,of which 22 were up-regulated and 18 were down-regulated during S.suis infection in mice.To evaluate the roles of these differentially expressed factors in S.suis virulence,deletion mutants were constructed,and 10 mutants were successfully obtained.Among these genes,the deletion of com R,sit R,or sxv R caused significantly decreased virulence in mice,compared to that of the wild-type strain.Moreover,the survival of Δcom R,Δsit R,and Δsxv R mutant strains in blood was significantly reduced both in vitro and in vivo.Further,their pro-inflammatory abilities were also obviously decreased in vivo.The regulatory mechanisms of Com R,Sit R,and Sxv R were then analyzed by whole transcriptome RNA sequencing(RNA-Seq).Results indicated that the absence of com R induced the down-regulation of 17 virulence factors or virulence-related factors,including genes involved in the synthesis of capsules,oxidative stress tolerance,immune evasion,and cell division.Further,three and two virulence factors or virulence-related factors were down-regulated upon deletion of sit R and sxv R,respectively.Thus,this study reports the discovery of three virulence-associated transcriptional regulatory factors in S.suis.These factors could ultimately be targeted to control infection caused by these bacteria.2.Research on pro-inflammatory mechanism of HP1717We found that the extracellular pro-inflammatory protein HP1717 in S.suis induced excessive production of inflammatory cytokines,and further study showed that this pro-inflammatory activity of HP1717 was dependent on Toll-like receptor 2(TLR2)recognition and phosphorylation of both extracellular signal–regulated kinases 1/2(ERK1/2)and nuclear factor kappa-light-chain-enhancer of activated B cells(NF-κB).On the other hand,HP1717 deficiency led to significant changes in the phenotypes of S.suis,including reduced growth,chain length extension,almost complete disappearance of the capsule.Mouse infection experiments showed that compared to those of wild-type bacteria,the virulence and tissue bacterial load of mutant bacteria were significantly reduced.In summary,our study confirms that HP1717 is an important factor related to the virulence of S.suis.HP1717 can induce an excessive inflammatory response and significantly affects the bacterial capsule.This study further expands our understanding of the pathogenesis of S.suis.