Effects Of Bacillus On Immune Responses And Gut Microbial Diversity Of Piglets Infected By Escherichia Coli

Aims:Although the probiotic Bacillus has been used as an promising alternative to protect piglets from post-weaning diarrhea, the mechanism of probiotic in regulating the intestinal health still remains elusive, especially involving with F4receptor-negative weaning piglets. Moderate immune responses and stable gut microbiota community possesses the capacities in defensing the exogenous pathogens-induced infections. The present study aimed at exploring the preventing mechanism of the amalgam of Bacillus subtilis and Bacillus licheniformis in an F4Enterotoxigenic Escherichia coli (F4+ETEC) infection model of F4receptor-negative weaning piglets by comparing the dose effects on gut microbial diversity and immune regulation.Methods:Exact21-day-old weaning F4receptor negative-piglets were randomly divided into four groups and each group is comprised of eight individuals:1) CONT, piglets orally administered with10mL of sterile physiological saline daily during first8days;2) ETEC, pigs orally administered with10mL sterile physiological saline every day for the first week and then orally challenged with10mL F4+ETEC culture (1.0×109CFU/mL) on day8;3) LDBE, pigs orally administered with low dose of Bacillus amalgam dissolved in10mL sterile physiological saline (3.9×107CFU/mL) every day for the first week and then orally challenged with10mL of F4+ETEC culture (1.0R109CFU/mL) on day8; and4) HDBE, pigs orally administered with high dose of Bacillus amalgam dissolved in10mL sterile physiological saline (7.8×10’CFU/mL) every day for the first week and then orally challenged with10mL of F4+ETEC culture (1.0×109CFU/mL) on day8. Piglets were euthanized on day15. Feed intake, body weight and rectal temperature were recorded regularly. The fecal and intestinal contents samples were analysed for microbial diversity by plate counting and high-throughput DNA sequence methods. The jejunal and ileal enteritis were assessed by H&E straining. Blood samples were analysed for routine examination and Serum IL-10and TNF-a concentrations were detected by ELISA.Results:Administration of both low and high dose of Bacillus significantly inhibited the F4+ETEC-induced decrease of daily feed intake. The low dose of Bacillus remarkly attenuated the decline of crude protein and gross energy degestibility, as well as ameliorated the fever and jejunum inflammation induced by F4+ETEC.The surging percentage of banded neutrophils induced by F4+ETEC in12,48,96and144h following challenge were attenuated by both low and high dose of Bacillus. Administration of the high dose of Bacillus remarkly inhibited the elevation of TNF-a concentration induced by F4+ETEC in6h. The IL-10concentrations of piglets administrated with low dose of Bacillus were lower than piglets challenged with F4ETEC in12h. Compared to exposure to F4ETEC, administration of high dose of Bacillus increased the fecal Lactobacillus and Bifidbacteria counts on day9, whereas low dose of Bacillus only multiplied the fecal Lactobacillus number on day12. The fecal Coliformis numbers of piglets administrated with high dose of Bacillus was significantly lower than either of piglets with other treatments. The low and high dose of Bacillus administration lessened the numbers of fecal Enterococci compared to CONT and ETEC treatments on day12. Compared with F4+ETEC challenge, high dose of Bacillus augmented the Lactobacillus numbers in caecum and colon, while low dose of Bacillus boosted the Lactobacillus numbers only in colon. Piglets administrated with low or high dose of Bacillus had higher Bifidbacteria number and lower Coliformis in ileum, caecum and colon than piglets with CONT and ETEC treatments.Conclusions:Both low and high dose of Bacillus ameliorated the decline of growth performance induced by F4+ETEC in F4receptor-negative weaning piglets, but the low dose of Bacillus were more efficient to inhibit the decrease of nutrient digestibility and attenuated F4+ETEC-induced fever and intestinal inflammation. Although high dose of Bacillus was associated with better regulation of microbial balance, it may impose negative effects in immune responses and thus increased the risk of being infected.