Evaluating the Effectiveness of Lactobacillus zeae against Enterotoxigenic Escherichia coli F4 Infection in an In Vitro Porcine Intestinal Epithelial Cell Model

Lactobacillus zeae (LB1 strain) has been shown to prevent the death of Caenorhabditis elegans caused by enterotoxigenic Escherichia coli (ETEC) infection by downregulating the gene expression of ETEC toxins and mediating antimicrobial peptides/defense molecules under the p38 mitogen-activated protein kinase (MAPK) and insulin/insulin-like growth factor (DAF/IGF) signaling pathways in the nematode. However, the protective effects of L. zeae LB1 at cellular and molecular levels have not yet been investigated in pigs. In this study, porcine intestinal cells (IPEC-J2) were used to investigate the potential of L. zeae LB1 in modulating intestinal barrier and innate immune functions and protecting against intestinal injuries and inflammatory reactions induced by ETEC F4 infection. The results showed that the pretreatment of IPEC-J2 cells with L. zeae LB1 significantly alleviated the ETEC F4-induced elevation of pro-inflammatory cytokines and damage in the cell monolayer ( P < 0.05). Whereas L. zeae LB1 administration had no effect ( P > 0.05) on the ETEC F4 adhesion and the mRNA abundance of antimicrobial defensins (pBD2 and pBD3), it reduced the mRNA abundance of Toll-like receptor 4 and 5 (TLR4 and -5, respectively) and virulence-related factors of ETEC F4 ( P < 0.05). Moreover, a DNA marker-based test targeting the mucin 4 gene that encodes the F4 fimbria receptor indicated that the IPEC-J2 cell line was from a pig that was identified as being resistant to developing ETEC-F4 diarrhea. These results suggested that probiotic L. zeae LB1 effectively protected the intestinal cells from ETEC F4 infection by inhibiting inflammation and maintaining barrier integrity via downregulating TLR4, TLR5, and ETEC F4 virulence-related factors expression instead of preventing ETEC F4 adhesion. Our data provide further evidence of the mechanisms at the cellular and molecular levels that probiotic L. zeae LB1 could improve gut health in pigs.