| Streptococcus suis 2(SS2)is an important worldwide swine pathogen and zoonotic agent,which account for two recent outbreaks in Jiangsu and Sichuan province in china and pose a great threaten to the pig industry and public health.Despite the growing researches on this pathogen during the last couple of years,however,very little is known about the detailed pathogenic mechanism by which the S.suis evades the host immune system and further results in fatal diseases.Despite the growing public attentions on this pathogen during the last couple of years,however,very little is known about the detailed pathogenic mechanism by which the S.suis evades the host immune system and further results in fatal diseases.Therefore,the primary objective of this study is to explore the potential immune escape mechanism in S.suis,aiming at shedding light on the complicated pathogenesis of S.suis to some extent.In our previous study,we have identified a novel surface-associated protein with unknown function,which exhibits a robust suppression of inflammatory cytokines secretion triggered by S.suis.To investigate the correlation between the function of this protein and the immune evasion mechanism within S.suis,the overall experiments are initiated and devised as the following.In the present study,a membrane protein encoded by SSU05ZY＿1914,which posses the suppression activity towards the S.suis inflammatory attributes,was discovered to be capable of significantly decreasing the cytokines secretions in the host,including the IL-1β,MCP-1,TNF-α and IFN-β.Subsequently,this protein was hence designated as HP1914 in the following assays.In order to provide novel insights into the immune escape mechanism of S.suis by unraveling the underlying function of HP1914,we firstly amplified the CDS sequence from the genomes DNA and then inserted it into the expression vector p ET-28 a.The recombination protein was then over-expressed and purified using Ni-NTA column.To detect its inflammation-suppression efficacy in vitro,the RAW264.7 cell line was employed as the experimental model.The inflammation-suppression ability towards the SS2 train and its lysed supernates in the RAW264.7 cell line model were assessed and measured by the relative quantitative real-time reverse transcription-PCR in ABI system,and the results exhibited its suppression effects on the SS2 pro-inflammation traits.To further explore the contribution of HP1914 in the course of SS2 infection,a deletion mutant was constructed.I ntriguingly,in comparison to the wild strain,the mutant presents a marked increase in the capability of promoting the pro-inflammatory cytokines secretion using the aforementioned RAW264.7 model.This finding,to some extent,also verifies the distinct inflammation-suppression activity in HP1914.To confirm this function in vivo,we further investigated the cytokines level in the blood using the mice intraperitoneal infection model.As expected,the results of elisa experiments indicate that a variety of cytokines in blood have recahed a relative high level during the three hours of post-infection.However,in the six hours post-infection,the viable bacteria load in blood in the mutant group was significant less than that of the wild SC19 group,which undoubtedly hint at the important role of HP1914 in the process of SS2 infection.Moreover,the HP1914 protein was still capable of suppressing the activation of excessive inflmatory response induced by a wealth of inflammatory activators including PGN,LTA,LPS and Pam3CSK4(TLR-2pathway),all of which are already well recognized by the public.Taken togenther,it thus can be postulated that the surfece-associated HP1914,which has the definite suppression activity on inflammatory activation components,can partially facilitate the SS2 to evade the immune system surveillance,thus contributing to the SS2 rapid reproduction in vivo and spread among the host organs. |
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