Germination and outgrowth of Bacillus subtilis and Bacillus licheniformis spores in the gastrointestinal tract of pigs

Abstract

Aims: To determine if orally ingested Bacillus spores used as probiotics or direct‐fed microbial feed additives germinate and the vegetative cells grow in the gastrointestinal (GI) tract. Methods and Results: Three independent experiments were done to determine if spores of Bacillus licheniformis and Bacillus subtilis germinate and grow in the GI tract of pigs. After a 2 weeks spore‐feeding period, spores were detected in all segments of the GI tract. The lowest number of spores was found in the stomach, increasing in the small intestine to approx. 55% of the dietary inclusion. When spores were withdrawn from the feed, faecal excretion of spores reflected the dietary inclusion, but decreased gradually to the background level after 1 week. By containing spores in short, sealed pieces of dialysis membrane that were orally administered to the pigs, both the number of spores and vegetative cells could be determined by flow cytometry. Spores accounted for 72% of the total counts after 4–6 h in the stomach and proximal part of the small intestine. After 24 h, spores constituted only 12% of the total counts in the stomach, caecum, and mid‐colon. Less spores and more vegetative cells were detected after 24 h, but total counts increased only 2·14‐fold compared to time zero. Conclusions: The experiments showed that 70–90% of dietary‐supplemented Bacillus spores germinate in the proximal part of the pig GI tract, and that only limited outgrowth of the vegetative cell population occurs. The two Bacillus strains can temporarily remain in the GI system, but will be unable to permanently colonize the GI tract. Significance and Impact of the Study: A substantial population of growing vegetative cells in the GI tract is not a prerequisite for the mode of action of Bacillus feed additives and probiotics.

Probiotic <i>Bacillus</i> species and <i>Saccharomyces boulardii</i> improve performance, gut histology and immunity in broiler chickens

“…Probiotic preparations belonging to single or multispecies of Lactobacillus, Streptococcus, Bacillus, Bifidobacterium, Enterococcus, Aspergillus, Candida, and Saccharomyces had a potential effect on modulation of the intestinal microflora and pathogen inhibition (Kabir, 2009). This could be owing to the production of antimicrobial substances such as bacteriocins and lactic acid, and the adherence and co-aggregation of probiotic bacteria to the mucosa, forming a barrier that prevents colonization by pathogens (Patterson & Burkholder, 2003;Leser et al, 2008). The antimicrobial activity of Bacillus spp.…”

Bacillus subtilis Spores Germinate in the Chicken Gastrointestinal Tract

“…More recently, spore germination has been observed in the GI tracts of pigs, using flow cytometry analysis (17). While all of these findings are important in the context of mammalian hosts, it cannot be assumed that they will translate directly to an avian host, primarily due to differences in GI anatomy and physiology.…”

Selection of Bacillus species for targeted in situ release of prebiotic galacto-rhamnogalacturonan from potato pulp in piglets

“…Under the experimental conditions used, maximal fiber release was observed after 16-20 h. The transit time in the small intestine is normally estimated to be less than 7 hours (Strube et al 2013). Conversely, Leser and co-workers found a substantial amount of vegetative Bacilli in the caecum of pigs 24 h after administration of two strains of B. subtilis and B. licheniformis (Leser et al 2008). While it is not possible to directly extrapolate the kinetics from our simple in vitro system to the conditions in the GI tract in the piglet, our data suggest the possibility that the strains of B. mojavensis, when co-fed with potato pulp as a feed supplement, could release prebiotic fibers in the small intestine.…”

“…Whether Bacillus spores are capable of readily germinating and growing in the GI tract has not been unequivocally proven, but it has been suggested that some of the probiotic effects of Bacillus species might be attributed to the spore itself (Leser et al 2008;LaTorre et al 2014).…”

“…In case of the pathogen Bacillus anthracis it is known that the spore itself can elicit an immunological response (Kang et al 2008), whereas other immunological effects of Bacillus species are attributed to vegetative cells and secreted effectors (Huang et al 2008;Okamoto et al 2012). In pigs, spores of B. subtilis and B. licheniformis can germinate in the GI tract of pigs, but only limited growth of vegetative cells has been observed (Leser et al 2008). This could indicate that the reported probiotic effects are in fact not mediated by a high number of growing cells.…”