Pediococcus pentosaceus IM96 Exerts Protective Effects against Enterohemorrhagic Escherichia coli O157:H7 Infection In Vivo

Abstract

Enterohemorrhagic Escherichia coli (EHEC) is a notorious and prevalent foodborne pathogen which can cause serious intestinal diseases. The antagonistic activity of probiotics against EHEC is promising, but most of the studies concerning this subject have been carried out in vitro. Specifically, the interaction between Pediococcus pentosaceus and EHEC O157:H7 in vivo has not been reported yet. In this study, we investigated the protective effect of P. pentosaceus IM96 on EHEC O157:H7-infected female mice in vivo. The results demonstrated that P. pentosaceus IM96 reduced the level of pro-inflammatory factors and increased the level of anti-inflammatory factors of EHEC O157:H7-infected mice. Furthermore, P. pentosaceus IM96 alleviated intestinal mucosal damage and increased the level of MUC-2, tight junction (TJ) proteins, and short chain fatty acids (SCFAs). The intestinal microbial community structure and the diversity and richness of the microbiota were also changed by P. pentosaceus IM96 treatment. In summary, P. pentosaceus IM96 exerted protective effects against EHEC O157:H7 via alleviating intestinal inflammation, strengthening the intestinal barrier function, and regulating intestinal microbiota, suggesting that P. pentosaceus IM96 might serve as a potential microbial agent to prevent and treat intestinal diseases caused by EHEC O157:H7 infection in the future.

Keywords: EHEC O157:H7; Pediococcus pentosaceus IM96; inflammation; intestinal barrier; intestinal microbiota; probiotics.

Figure 1

P. pentosaceus IM96 relieved the clinical characteristics of EHEC O157:H7-infected mice. (A) Survival rate of the mice; (B) body weight change of the mice; (C) ratio of villus height to crypt depth in jejunum of the mice; (D) representative image of H&E staining in jejunum and ileum of the mice (scale bar, 100 μm). All of the data are expressed as the mean ± SD (n = 8), *** p < 0.001.

Figure 2

P. pentosaceus IM96 relieved EHEC O157:H7-induced inflammation in jejunum of the mice. (A) IL-1β; (B) IL-6; (C) TNF-α; and (D) IL-10. All of the data are expressed as the mean ± SD (n = 8), *** p < 0.001.

Figure 3

P. pentosaceus IM96 enhanced the intestinal epithelial barrier function in jejunum of the mice by upregulating MUC-2 expression. (A) The concentration of MUC-2; (B) the number of goblet cells per microscopic field (scale bar, 100 μm); (C) representative images of immunohistochemical stainings of MUC-2; and (D) representative images of PAS stainings (scale bar, 100 μm). All of the data are expressed as the mean ± SD (n = 8), * p < 0.05, ** p < 0.01, *** p < 0.001.

Figure 4

P. pentosaceus IM96 enhanced the intestinal epithelial barrier function in jejunum of the mice by increasing TJ proteins expression. (A) The concentration of Occludin and (B) ZO-1; (C) representative images of immunohistochemical stainings of Occludin and (D) ZO-1 (scale bar, 100 μm). All of the data are expressed as the mean ± SD (n = 8), * p < 0.05, ** p < 0.01, *** p < 0.001.

Figure 5

P. pentosaceus IM96 return the concentrations of SCFAs to normal levels. (A) Acetic acid; (B) propionic acid; (C) butyric acid. All of the data are expressed as the mean ± SD (n = 6), * p < 0.05, ** p < 0.01.

Figure 6

P. pentosaceus IM96 regulated the bacterial community of the cecum microbiota. (A) NMDS; (B) relative abundance richness at phyla level; (C) heatmap of the top 20 families; (D) LEfSe on family level; and (E) LDA in EHEC group and IM96 group.

Figure 7

Correlation analysis of environmental factors. (A) RDA of intestinal microbiota distribution and environmental factors. The arrows indicate the environmental factors, and the length of the arrow line indicates the degree of correlation between the environmental factor and the sample distribution. The longer the line, the greater the correlation. The angle between the arrow line and the sort axis and the angle between the arrow line indicates the correlation. The smaller the angle, the higher the correlation; (B) the significance of environmental factors was analyzed by envfit function. r² is determinant coefficient of environmental factors on intestinal microbiota distribution; (C) correlation heatmap, * p < 0.05, ** p < 0.01, *** p < 0.001.