Bacillus Probiotics: Spore Germination in the Gastrointestinal Tract

Abstract

Spores of Bacillus species are being used commercially as probiotics and competitive exclusion agents. Unlike the more commonly used Lactobacillus-type probiotics, spores are dormant life forms. To address how spore probiotics might function we have investigated whether spores can germinate in the gastrointestinal tract by using a murine model. Using a genetically engineered chimeric gene, ftsH-lacZ, which is strongly expressed only in vegetative cells, we have developed a sensitive competitive reverse transcription-PCR assay which has enabled detection of as few as 10 2 vegetative bacteria in the mouse gut. Using this method we have administered doses of ftsH-lacZ spores to groups of mice and shown that spores can germinate in significant numbers in the jejunum and ileum. The levels of detection we obtained suggest that spores may colonize the small intestine, albeit briefly.Bacterial spores are dormant life forms which can exist in a desiccated and dehydrated state indefinitely. The process of spore formation has been extensively studied as a simple model for understanding cellular differentiation and is one of the primary reasons for the interest in spores and spore formation (7). Intriguingly though, spores of Bacillus subtilis are being used as probiotics and competitive exclusion (CE) agents for both human and animal consumption (18). For humans they are available either as over-the-counter prophylactics for mild gastrointestinal disorders such as diarrhea or as health foods or nutritional supplements. In some countries though (e.g., Vietnam), bacterial spores are being used for oral bacteriotherapy of gastrointestinal disorders often under clinical supervision. In the agricultural industry spores are also receiving increasing attention as potential alternatives to antibiotics as growth promoters. The use of probiotics and/or CE agents seems likely to increase as public awareness of their potential benefits increases.While spores are being sold as probiotics, an important question is that of how spores act to enhance the normal microbial flora of the gastrointestinal tract. This question must be addressed, because the majority of probiotics currently available are bacteria which are non-spore formers: i.e., they are given as vegetative cells (usually as lyophilized preparations). The best-known examples of these probiotic bacteria are the lactobacilli and bifidobacteria (2,8,9). If probiotic bacteria are to be taken seriously then we would assume that they would share a common mechanism for enhancing the normal well-being of the gut microflora or for CE of potential pathogens. If there is no common mechanism shared between conventional Lactobacillus-type bacteria and the spore probiotics, then the question must be asked as to whether there is any credibility to some of the claims made about the positive benefits of probiotic bacteria (2, 12).Probiotics and CE agents are thought to enhance the gut microflora by preventing the colonization of the gastrointestinal tract by pathogenic bacteria. There are three...

Identification and characterization of Bacillus subtilis from grass carp (Ctenopharynodon idellus) for use as probiotic additives in aquatic feed

“…For lacking of mucus production, Caco-2 cell monolayer is usually used as an ideal model to investigate bacterial adhesion to intestinal epithelium [59]. In this study, the results demonstrated that both vegetative cells and spores of the isolated B. subtilis could adhere to and interact with Caco-2 cells, suggesting that the isolated B. subtilis strains could colonize the intestinal tract, and the colonized spores might germinate, proliferate and re-sporulate within intestinal tract [60,61].…”

Bacillus subtilis Spores Germinate in the Chicken Gastrointestinal Tract

“…it was shown that vegetative cells of Bacillus subtilis could be detected in the small intestines of mice after they had received an oral dose of spores (6,25). More recently, spore germination has been observed in the GI tracts of pigs, using flow cytometry analysis (17).…”

“…Other researchers have reported similar problems (14,15), the issues being (i) inefficient and nonsynchronous spore germination in the absence of a heat treatment/activation step, which leads to an underestimation of total viable units, and (ii) growth of background contaminants which obscure colonies of B. subtilis on agar plates. Ultimately, our decision to use RT-PCR was based on successful work by researchers elsewhere (6,25).…”

“…To address the fate of orally administered spores, 1-day-old specific pathogen-free White Leghorn chicks were each dosed by oral gavage with 1 ϫ 10 9 spores of B. subtilis SC2362 (6,8,9) suspended in 0.1 ml of sterile distilled water. This strain of B. subtilis harbors a unique rrnO-lacZ fusion gene and a chloramphenicol acetyltransferase (cat) gene in the chromosome.…”

Crosstalk between the microbiome and cancer cells by quorum sensing peptides

“…Moreover, bacterial quorum sensing molecules are likely to play a role in bacterial colonization of mucosa, thus requiring quorum sensingmediated biofilm formation [14]. Finally, Casula and Cutting showed the germination of Bacillus subtilis spores in the murine gastrointestinal tract, thereby probably requiring signaling peptides for quorum sensing pathway activation [15]. Although thus not yet investigated, it is very likely that also quorum sensing peptides are found in the human intestinal tract.…”