The Salmonella pathogenicity islands(SPIs) play crucial roles in the progression of Salmonella infection. In this study, we constructed an improved λ Red homologous recombination system to prepare single and triple d...The Salmonella pathogenicity islands(SPIs) play crucial roles in the progression of Salmonella infection. In this study, we constructed an improved λ Red homologous recombination system to prepare single and triple deletion mutants of 3 prominent SPIs(SPI-1, 2, and 3), aiming at the impact of deletion on morphology, carbon source metabolism, adhesion and invasion capacity, in vivo colonization, and immune efficacy in chicks. Our examination revealed that the surface of the single deletion mutants(SM6ΔSPI1, ΔSPI2, and ΔSPI3) exhibited a more rugged texture and appeared to be enveloped in a layer of transparent colloid, whereas the morphology of the triple deletion mutant(SM6ΔSPI1&2&3) remained unaltered when compared to the parent strain. The carbon metabolic spectrum of the SPI mutants underwent profound alterations, with a notable and statistically significant modification observed in 30 out of 95 carbon sources, primarily carbohydrates(17 out of 30). Furthermore, the adhesion capacity of the 4 mutants to Caco-2 cells was significantly reduced when compared to that of the parent strain. Moreover,the invasion capacity of mutants SM6ΔSPI1 and SM6ΔSPI1&2&3 exhibited a substantial decrease, while it was enhanced to varying degrees for SM6ΔSPI3 and SM6ΔSPI2. Importantly, none of the 4 mutants induced any clinical symptoms in the chicks. However, they did transiently colonize the spleen and liver. Notably, the SM6ΔSPI1&2&3mutant was rapidly cleared from both the spleen and liver within 8 days post-infection and no notable pathological changes were observed in the organs. Additionally, when challenged, the mutants immunized groups displayed a significant increase in antibody levels and alterations in the CD3+CD4+ and CD3+CD8+ subpopulations, and the levels of IL-4 and IFN-γ cytokines in the SM6ΔSPI1&2&3 immunized chicken serum surpassed those of other groups.In summary, the successful construction of the 4 SPI mutants lays the groundwork for further exploration into the pathogenic(including metabolic) mechanisms of SPIs and the development of safe and effective live attenuated Salmonella vaccines or carriers.展开更多
Salmonella enterica serovar Typhimurium,the causative agent of gastroenteritis,is one of the most successful intracellular pathogens.Although certain host factors for Salmonella infection have been unveiled,the factor...Salmonella enterica serovar Typhimurium,the causative agent of gastroenteritis,is one of the most successful intracellular pathogens.Although certain host factors for Salmonella infection have been unveiled,the factors mediating Salmonella entry,particularly the invasion process,remain obscure.Here,we have unearthed β2 integrin,a crucial member of the integrin family,as an important host factor facilitating Salmonella invasion.It is demonstrated that overexpression of β2 integrin promotes Salmonella invasion,while the knockdown of β2 integrin significantly diminishes the extent of invasion.Moreover,Salmonella exhibits specific binding affinity towards β2 integrin,and the block of β2 integrin on cell surface substantially reduces the infection of cells in vitro.The ectodomain soluble protein of β2 integrin neutralized Salmonella infection both in cells(in vitro)and in mice(in vivo).Additionally,Salmonella protein YrbD directly interacts with β2 integrin to facilitate its invasion.To our knowledge,this study showed for the first time that the protein YrbD mediates Salmonella adhesion and internalization into host cells by interacting with β2 integrin.These findings not only broaden our understanding of the mechanisms underlying Salmonella entry,but also identify a prospective target for therapeutic control.展开更多
基金supported by the National KeyR&DProgramof China(2022YFF0710500)the National Natural Science Foundation of China(32172853 and 32373013)the Central Public-interest Scientific Institution Basal Research Fund,China(1610302022001).
文摘The Salmonella pathogenicity islands(SPIs) play crucial roles in the progression of Salmonella infection. In this study, we constructed an improved λ Red homologous recombination system to prepare single and triple deletion mutants of 3 prominent SPIs(SPI-1, 2, and 3), aiming at the impact of deletion on morphology, carbon source metabolism, adhesion and invasion capacity, in vivo colonization, and immune efficacy in chicks. Our examination revealed that the surface of the single deletion mutants(SM6ΔSPI1, ΔSPI2, and ΔSPI3) exhibited a more rugged texture and appeared to be enveloped in a layer of transparent colloid, whereas the morphology of the triple deletion mutant(SM6ΔSPI1&2&3) remained unaltered when compared to the parent strain. The carbon metabolic spectrum of the SPI mutants underwent profound alterations, with a notable and statistically significant modification observed in 30 out of 95 carbon sources, primarily carbohydrates(17 out of 30). Furthermore, the adhesion capacity of the 4 mutants to Caco-2 cells was significantly reduced when compared to that of the parent strain. Moreover,the invasion capacity of mutants SM6ΔSPI1 and SM6ΔSPI1&2&3 exhibited a substantial decrease, while it was enhanced to varying degrees for SM6ΔSPI3 and SM6ΔSPI2. Importantly, none of the 4 mutants induced any clinical symptoms in the chicks. However, they did transiently colonize the spleen and liver. Notably, the SM6ΔSPI1&2&3mutant was rapidly cleared from both the spleen and liver within 8 days post-infection and no notable pathological changes were observed in the organs. Additionally, when challenged, the mutants immunized groups displayed a significant increase in antibody levels and alterations in the CD3+CD4+ and CD3+CD8+ subpopulations, and the levels of IL-4 and IFN-γ cytokines in the SM6ΔSPI1&2&3 immunized chicken serum surpassed those of other groups.In summary, the successful construction of the 4 SPI mutants lays the groundwork for further exploration into the pathogenic(including metabolic) mechanisms of SPIs and the development of safe and effective live attenuated Salmonella vaccines or carriers.
基金supported by the National Key R&D Program of China(2022YFF0710500)the the National Natural Science Foundation,China(31802192,32172853 and 32373013)+2 种基金the Natural Science Foundation of Heilongjiang Province of China(C2018070)China Postdoctoral Science Foundation(2017M620076)the Central Public-interest Scientific Institution Basal Research Fund,China(1610302022001)。
文摘Salmonella enterica serovar Typhimurium,the causative agent of gastroenteritis,is one of the most successful intracellular pathogens.Although certain host factors for Salmonella infection have been unveiled,the factors mediating Salmonella entry,particularly the invasion process,remain obscure.Here,we have unearthed β2 integrin,a crucial member of the integrin family,as an important host factor facilitating Salmonella invasion.It is demonstrated that overexpression of β2 integrin promotes Salmonella invasion,while the knockdown of β2 integrin significantly diminishes the extent of invasion.Moreover,Salmonella exhibits specific binding affinity towards β2 integrin,and the block of β2 integrin on cell surface substantially reduces the infection of cells in vitro.The ectodomain soluble protein of β2 integrin neutralized Salmonella infection both in cells(in vitro)and in mice(in vivo).Additionally,Salmonella protein YrbD directly interacts with β2 integrin to facilitate its invasion.To our knowledge,this study showed for the first time that the protein YrbD mediates Salmonella adhesion and internalization into host cells by interacting with β2 integrin.These findings not only broaden our understanding of the mechanisms underlying Salmonella entry,but also identify a prospective target for therapeutic control.
