Study On The Mechanism Of Pgm Gene Of Streptococcus Equi Ssp.zooepidemicus Affecting Pathogenicity

Streptococcus equi ssp.zooepidemicus（SEZ）is an important pathogen,which mainly causes swine streptococcus disease in China.In recent years,there have been reports of SEZ-related cases worldwide,but there are still no effective measures to prevent and eradicate the invasion of this pathogen.Studying the pathogenesis of SEZ is helpful to the prevention and treatment of SEZ disease.Previous studies have shown that caspase-1 can regulate the airframe’s elimination mechanism of bacteria.Using this as an entry point to study the host’s clearance mechanism of SEZ can become a therapeutic target for SEZ disease.In this study,the C55138 strain of Streptococcus equi ssp.zooepidemicus was used as the research object to study the pathogenic function of its pgm gene and provide a theoretical basis for in-depth understanding of the pathogenic mechanism of SEZ.The main findings are as follows:（1）Analyzing the expression characteristics of SEZ pgm gene during in vivo and in vitro induction by q PCR,it was found that the expression of pgm gene in vivo was down-regulated,which may be related to the virulence of SEZ.In order to study the function of the pgm gene,SEZ C55138 was used as the parent strain,and the pgm gene deletion mutant strain?pgm was successfully obtained by homologous recombination technology.The capsule morphology of the?pgm strain was compared with the C55138 strain and the?pgm strain by scanning electron microscopy.Membrane secretion defects,analysis of the growth curve rate of the two strains showed that the growth and reproduction of?pgm strain and C55138strain were not significantly different.（2）By measuring the virulence of the mutant strain to mice and the deposition test of complement C3 on the surface of the bacteria,the mutant strain’s virulence to mice was significantly weakened,and the deposition of complement C3 on the surface of the mutant strain was significantly higher than that of the parent strain.It can promote the phagocytosis mediated by the body’s complement system.（3）Establish a mouse infection model,and the results of serum cytokine detection by ELISA showed that the IL-18 level of the?pgm infection group was significantly lower than that of the C55138 infection group;and it was confirmed by Western Blot that the mutant strain?pgm activated the expression of caspase-1protein after infecting mice.（4）A caspase-1^-/-mouse infection model was established,and the changes of serum cytokines were detected by ELISA.The results showed that the level of IL-18in the?pgm infection group was not significantly different from that in the C55138infection group,further verifying that mice infected with the mutant strain?pgm depend on the activation of caspase-1 to cause a decrease in IL-18 secretion.（5）The results of the pathogenicity test of caspase-1^+/+and caspase-1^-/-mice showed that whether the parent strain or the mutant strain infected caspase-1^-/-mice,the mouse mortality rate reached 100%.When infected with caspase-1^+/+mice,the mortality of the?pgm infection group was significantly lower than that of the C55138 infection group.It shows that the mutant strain?pgm weaken the pathogenicity relies on caspase-1.The above test results show that:caspase-1 mediates the secretion of IL-18 by the mutant strain?pgm,thereby reducing the pathogenicity of the mutant strain?pgm.This study provides a research basis for revealing the pathogenesis of SEZ,and can be used as a target molecule for the treatment of SEZ-related diseases.