Functional Analysis Of Glycosyl Hydrolase CZGH25 In Streptococcus Suis CZ130302 And Genome Analysis Of WH1609

Streptococcus suis(S.suis)is an important zoonotic pathogen,which can cause meningitis,septicemia,arthritis,toxic shock syndrome and death in pigs or human beings.It seriously endangers the development of swine industry and human health.There are many serotypes of S.suis while the virulence and geographical distribution of S.suis in different serotypes are significantly different.More than 80 virulence factors have been identified in S.suis up to now,but not present in all S.suis and even if the same virulence factors might not play the consistent role in different strains.Studies in genomics and molecular for the newly isolated strains are helpful to understand the molecular characteristics and genetic variation of S.suis,which can also provide a reference for the study of the pathogenic characteristics of S.suis.1 Functional analysis of glycosyl hydrolase CZGH25 in CZ130302In this study,11 secreted proteins of S.suis CZ130302 were identified by MALDI-TOF MS analysis ans CZGH25 was selected for subsequent study.CZGH25 belongs to glycosyl hydrolase 25(GH25)family and is about 45 KDa in size.There are few reports about the glycosyl hydrolase in S.suis.In order to know more about the biological function of CZGH25,the czgh25 deletion strain ?SS-CZGH25 was constructed.CZGH25 was also confirmed to be expressed in CZ130302 and secreted to the extracellular by indirect immunofluorescence assay.The adhesion of ?SS-CZGH25 to HEp-2 cells was was reduced by about 20%compared with wild-type strain.The survival rate of mice infected with ?SS-CZGH25 was 40%,which was significantly higher than that of mice infected with CZ130303,and the colonization ability of ?SS-CZGH25 in blood,brain,kidney,lung decreased by about 10%compared with CZ130302.Far-Western blot showed that CZGH25 could bind to fibrinogen(Fg)specifically.These results indicate that CZGH25 is a virulence factor of CZ130302.The survival rate of mice infected with CZ130302 was 37%after being immunizing with CZGH25 protein which suggested that CZGH25 had certain immunoprotective effect on mice.The autolysis of CZ130302 increased after the deletion of czgh25 gene and the sensitivity to acid and H2O2 increased significantly,suggesting that CZGH25 participate in the resistance of CZ130302 to environmental stress.2 Genome analysis of Streptococcus suis WH1609In 2016,a strain of S.suis type 9,named WH1609,was isolated from clinic in our laboratory.The strain has a strong pathogenicity to mice.Further study on WH1609 helps to elucidate the pathogenic mechanism of S.suis.The LD50 of WH1609 is 3.16 × 102 CFU/mL which was measured with zebrafish as an animal model.The virulence of WH1609 is about 105 times that of S.suis virulent strains.Drug sensitivity test indicated that WH1609 was resistant to aminoglycosides,lincomycin,macrolides antibiotics and sensitive to penicillin,?-lactam,fluoroquinolones antibiotics which was similar to other reports.The whole genome sequencing of WH1609 was carried out,and multiple online tools were used to analyze its genome.There were 3 large gene islands,3 prophages,61 virulence related factors and 11 drug resistance genes were harboured in WH1609,which were speculated to be related to the virulence of bacteria and the horizontal transfer of drug-resistant genes.Genomic comparative analysis revealed that WH1609 has large scale rearrangement,insertion and inversion.The core gene evolution analysis showed that WH1609 locates an independent evolutionary branch and has a closer relationship with S.suis D12.