Enterococcus faecium NCIMB 10415 modulates epithelial integrity, heat shock protein, and proinflammatory cytokine response in intestinal cells.

Probiotics have shown positive effects on gastrointestinal diseases; they have barrier-modulating effects and change the inflammatory response towards pathogens in studies in vitro. The aim of this investigation has been to examine the response of intestinal epithelial cells to Enterococcus faecium NCIMB 10415 (E. faecium), a probiotic positively affecting diarrhea incidence in piglets, and two pathogenic Escherichia coli (E. coli) strains, with specific focus on the probiotic modulation of the response to the pathogenic challenge. Porcine (IPEC-J2) and human (Caco-2) intestinal cells were incubated without bacteria (control), with E. faecium, with enteropathogenic (EPEC) or enterotoxigenic E. coli (ETEC) each alone or in combination with E. faecium. The ETEC strain decreased transepithelial resistance (TER) and increased IL-8 mRNA and protein expression in both cell lines compared with control cells, an effect that could be prevented by pre- and coincubation with E. faecium. Similar effects were observed for the increased expression of heat shock protein 70 in Caco-2 cells. When the cells were challenged by the EPEC strain, no such pattern of changes could be observed. The reduced decrease in TER and the reduction of the proinflammatory and stress response of enterocytes following pathogenic challenge indicate the protective effect of the probiotic.

Analysis of in vitro and in vivo effects of probiotics against Campylobacter spp.

Campylobacter (C.) spp. are well recognised as the leading cause of bacterial food-borne diarrheal disease worldwide, with C. jejuni and C. coli as the most important species: C. coli is highly abundant in pigs and pork meat has often been implicated as a source for human infection. Intestinal colonisation of C. coli in pigs plays a role in carcass contamination during slaughter. Different pre-harvest intervention measures are proposed to reduce the C. coli burden in the porcine intestine. Among others, the use of probiotics to prevent or reduce the colonisation of intestinal pathogens is discussed. One aim of this study was to screen a variety of probiotics to evaluate their inhibitory activity against Campylobacter spp. in vitro. Therefore, cell-free culture supernatants of Lactobacillus spp., Bifidobacterium spp., Enterococcus (E.) faecium NCIMB 10415, and Escherichia coli Nissle 1917 were tested against C. jejuni and C. coli by a well-diffusion agar assay. Seven out of eleven Lactobacillus strains showed an inhibitory activity against at least one of the three tested Campylobacter strains. This antagonistic activity against Campylobacter spp. was caused by the production of organic acids that lowered the pH. Application with pH neutralised cell-free culture supernatants abolished this inhibitory effect. Other tested strains with probiotic properties showed no inhibitory activity against any Campylobacter spp. strain. The strain E. faecium NCIMB 10415 was chosen to test its inhibitory activity against C. coli in vivo. Twenty weaned piglets were allocated into two groups, a probiotic group and a control group.The diet of the probiotic group was supplemented with E. faecium NCIMB 10415 (10(9) cfu/kg feed, Cylactin) since weaning, whereas the control group received no probiotic treatment. All piglets were naturally colonised with C. coli. The excretion load of C. coli was monitored for 28 days. The results indicate that dietary supplementation of E. faecium NCIMB 10415 did not significantly affect C. coli excretion levels in pigs. In this study, E. faecium NCIMB 10415 showed no antagonistic activity against C. coli in vitro and in vivo and had no impact on the growth performance of weaned piglets.

Experimental infection of weaned piglets with Campylobacter coli--excretion and translocation in a pig colonisation trial.

Campylobacter (C.) is one of the most common food-borne pathogen causing bacterial enteric infections in humans. Consumption of meat and meat products that have been contaminated with Campylobacter are the major source of infection. Pigs are a natural reservoir of Campylobacter spp. with C. coli as the dominant species. Even though some studies focussed on transmission of C. coli in pig herds and the excretion in faeces, little is known about the colonisation and excretion dynamics of C. coli in a complex gut microbiota present in weaned piglets and the translocation to different tissues. Therefore, an experimental trial was conducted to evaluate the colonisation and translocation ability of the porcine strain C. coli 5981 in weaned pigs. Thus, ten 35 days old piglets were intragastrically inoculated with strain C. coli 5981 (7 × 10(7)CFU/animal) encoding resistances against erythromycin and neomycin. Faecal samples were taken and C. coli levels were enumerated over 28 days. All piglets were naturally colonised with C. coli before experimental inoculation, and excretion levels ranged from 10(4) to 10(7)CFU/g faeces. However, no strain showed resistances against the additional antimicrobials used. Excretion of C. coli 5981 was seen for all piglets seven days after inoculation and highest counts were detectable ten days after inoculation with 10(6)CFU/g faeces. Post-mortem, translocation and subsequent invasion of luminal C. coli was observed for gut tissues of the small intestine and for the gut associated lymphatic tissues, such as jejunal mesenteric lymph nodes and tonsils as well as for spleen and gall bladder. In conclusion, this pig colonisation trial offers the opportunity to study C. coli colonisation in weaned piglets using the porcine strain C. coli 5981 without the need for gnotobiotic or specific pathogen-free animals.