Salmonella Enteritidis activates inflammatory storm via SPI-1 and SPI-2 to promote intracellular proliferation and bacterial virulence

Salmonella Enteritidis is an important intracellular pathogen, which can cause gastroenteritis in humans and animals and threaten life and health. S. Enteritidis proliferates in host macrophages to establish systemic infection. In this study, we evaluated the effects of Salmonella pathogenicity island-1 (SPI-1) and SPI-2 to S. Enteritidis virulence in vitro and in vivo, as well as the host inflammatory pathways affected by SPI-1 and SPI-2. Our results show that S. Enteritidis SPI-1 and SPI-2 contributed to bacterial invasion and proliferation in RAW264.7 macrophages, and induced cytotoxicity and cellular apoptosis of these cells. S. Enteritidis infection induced multiple inflammatory responses, including mitogen-activated protein kinase (ERK-mediated) and Janus kinase-signal transducer and activator of transcript (STAT) (STAT2-mediated) pathways. Both SPI-1 and SPI-2 were necessary to induce robust inflammatory responses and ERK/STAT2 phosphorylation in macrophages. In a mouse infection model, both SPIs, especially SPI-2, resulted in significant production of inflammatory cytokines and various interferon-stimulated genes in the liver and spleen. Activation of the ERK- and STAT2-mediated cytokine storm was largely affected by SPI-2. S. Enteritidis Delta SPI-1-infected mice displayed moderate histopathological damage and drastically reduced bacterial loads in tissues, whereas only slight damage and no bacteria were observed in Delta SPI-2- and Delta SPI-1/SPI-2-infected mice. A survival assay showed that Delta SPI-1 mutant mice maintained a medium level of virulence, while SPI-2 plays a decisive role in bacterial virulence. Collectively, our findings indicate that both SPIs, especially SPI-2, profoundly contributed to S. Enteritidis intracellular localization and virulence by activating multiple inflammatory pathways.

Automated summary

Salmonella Enteritidis is a significant intracellular pathogen that can cause gastroenteritis and pose a threat to life and health. This study evaluated the impact of Salmonella pathogenicity island-1 (SPI-1) and SPI-2 on S. Enteritidis virulence in vitro and in vivo, as well as the host inflammatory pathways affected by these SPIs. The results showed that both SPI-1 and SPI-2 contributed to bacterial invasion and proliferation in macrophages, induced cytotoxicity and cellular apoptosis, and triggered inflammatory responses mediated by ERK and STAT2 pathways. In a mouse infection model, SPI-2 played a crucial role in bacterial virulence by causing significant production of inflammatory cytokines and interferon-stimulated genes. Delta SPI-1 mutant mice showed moderate histopathological damage and reduced bacterial loads, while Delta SPI-2 and Delta SPI-1/SPI-2 mutant mice displayed minimal damage and no bacteria. A survival assay demonstrated that SPI-2 played a decisive role in bacterial virulence. Overall, both SPIs, especially SPI-2, profoundly contributed to S. Enteritidis intracellular localization and virulence by activating multiple inflammatory pathways.