The Intestinal Life Cycle of Bacillus subtilis and Close Relatives

Abstract

Bacillus subtilis is considered a soil organism for which endospore formation provides a means to ensure long-term survival in the environment. We have addressed here the question of what happens to a spore when ingested. Spores displaying on their surface a heterologous antigen, tetanus toxin fragment C (TTFC), were shown to generate anti-TTFC responses not to the antigen contained in the primary oral inoculum but to those displayed on spores that had germinated and then resporulated. We then used reverse transcription-PCR to determine expression of vegetative genes and sporulation-specific genes in the mouse gut following oral dosing with spores. Significant levels of germination and sporulation were documented. Using natural isolates of B. subtilis that could form biofilms, we showed that these strains could persist in the mouse gut for significantly longer than the laboratory strain. Moreover, these isolates could grow and sporulate anaerobically and exhibited a novel phenomenon of being able to form spores in almost half the time required for the laboratory isolate. This suggests that spores are not transient passengers of the gastrointestinal tract but have adapted to carry out their entire life cycle within this environment. This is the first report showing an intestinal life cycle of B. subtilis and suggests that other Bacillus species could also be members of the gut microflora.

Bacillus subtilis Spores Germinate in the Chicken Gastrointestinal Tract

“…This principle finding agrees with studies conducted with a mouse model (6,25). However, in those studies, vegetative cells were detected only in the small intestines of mice dosed with spores.…”

“…Between 0.13% and 1.23% of the original spore dose was estimated to have been retained in the GI tract of each chick (Table 2). This finding is comparable with that from work conducted in mice (6,25) and supports the notion that B. subtilis is a transient member of the chick GI microflora, which does not colonize the GI tract. The remainder of the original spore dose may have been shed in feces or germinated in the GI environment and subsequently died due to unfavorable conditions.…”

“…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).…”

Bioavailability of the anti-clostridial bacteriocin thuricin CD in gastrointestinal tract

“…Previous studies have shown that some Bacillus species could germinate and grow in the GIT of humans and mice (Hoa et al, 2001;Tam et al, 2006;Hong et al, 2009a, b). Spores have the advantage over vegetative cells of being resistant to the low pH of the stomach, ensuring the delivery of high numbers to the GIT.…”