The LRR domain of internalin gene regulates Streptococcus agalactiae crossing the intestinal barrier and intestinal immunity in tilapia

Abstract

To investigate the effect of the LRR domain of the internalin gene on its role in the invasion of S. agalactiae into tilapia through the intestine and the impact on host immunity. In this study, the LRR domain deletion mutant strain of TOS01 was firstly constructed by homologous recombination. The results of biological characteristics showed no significant difference in growth rate between TOS01ΔLRR and TOS01 (P > 0.05). However, compared with TOS01, the capsular polysaccharide of TOS01ΔLRR was significantly thickened, the chain was significantly shortened, and the ability of invasion and adhesion to PCK cells was significantly decreased (P < 0.05). In addition, the animal oral challenge test results showed that compared with TOS01, the strain TOS01ΔLRR significantly reduced the tissue bacterial load of experimental animals at 24 h, suggesting a role of the LRR domain in mediating bacterial crossing intestinal barriers. However, the virulence did not change, which was also confirmed by comparative transcriptome analysis of TOS01ΔLRR and TOS01. To further explore the effect of the LRR domain of the internalin gene of S. agalactiae on host intestinal immunity, we used RNA-seq to examine tissue expression patterns in tilapia after stimulation with TOS01ΔLRR and TOS01. RNA-seq results displayed 1255 genes were differentially expressed, with 521 genes up-regulated and 734 genes down-regulated. KEGG enrichment showed that differentially expressed genes were mainly concentrated in cell adhesion, invasion and immune-related aspects, such as lysosome, phagosome, apoptosis, cell adhesion molecules (CAMs), cytokine-cytokine receptor interaction, intestinal immune network for IgA production, and PPAR signaling pathwayetc. Among them, the cytokine-cytokine receptor interaction, cellular senescence, PPAR signaling pathway and NOD-like receptor signaling pathway were significantly up-regulated and the lysosome and apoptosis was significantly down-regulated. This suggested that LRR domain inhibited innate immune-related pathways to evade host immunity during S. agalactiae invasion. In summary, the LRR domain of the internin gene is a critical component for S. agalactiae to invade and adhere to host cells. Its deletion reduces the ability of the strain to cross the intestinal barrier, and the LRR domain of internin suppresses the host's innate immunity to promote S. agalactiae colonization in the gut.