The soil saprophyte Bacillus cereus forms biofilms at solid-liquid interfaces. The composition of the extracellular polymeric matrix is not known, but biofilms of other bacteria are encased in polysaccharides, protein, and also extracellular DNA (eDNA). A Tn917 screen for strains impaired in biofilm formation at a solid-liquid interface yielded several mutants. Three mutants deficient in the purine biosynthesis genes purA, purC, and purL were biofilm impaired, but they grew planktonically like the wild type in Luria-Bertani broth. Biofilm populations had higher purA, purC, and purL transcript ratios than planktonic cultures, as measured by real-time PCR. Laser scanning confocal microscopy (LSCM) of BacLight-stained samples indicated that there were nucleic acids in the cell-associated matrix. This eDNA could be mobilized off the biofilm into an agarose gel matrix through electrophoresis, and it was a substrate for DNase. Glass surfaces exposed to exponentially growing populations acquired a DNA-containing conditioning film, as indicated by LSCM. Planktonic exponential-phase cells released DNA into an agarose gel matrix through electrophoresis, while stationary-phase populations did not do this. DNase treatment of planktonic exponential-phase populations rendered cells more susceptible than control populations to the DNA-interacting antibiotic actinomycin D. Exponential-phase purA cells did not contain detectable eDNA, nor did they convey a DNA-containing conditioning film to the glass surface. These results indicate that exponential-phase cells of B. cereus ATCC 14579 are decorated with eDNA and that biofilm formation requires DNA as part of the extracellular polymeric matrix.

Bacillus is commonly isolated from soils, with organisms of Bacillus cereus sensu lato being prevalent. Knowledge of the ecology of B. cereus and other Bacillus species in soil is far from complete. While the older literature favors a model of growth on soil-associated organic matter, the current paradigm is that B. cereus sensu lato germinates and grows in association with animals or plants, resulting in either symbiotic or pathogenic interactions. An in terra approach to study soil-associated bacteria is described, using filtersterilized soil-extracted soluble organic matter (SESOM) and artificial soil microcosms (ASM) saturated with SESOM. B. cereus ATCC 14579 displayed a life cycle, with the ability to germinate, grow, and subsequently sporulate in both the liquid SESOM extract and in ASM inserted into wells in agar medium. Cells grew in liquid SESOM without separating, forming multicellular structures that coalesced to form clumps and encasing the ensuing spores in an extracellular matrix. Bacillus was able to translocate from the point of inoculation through soil microcosms as shown by the emergence of outgrowths on the surrounding agar surface. Microscopic inspection revealed bundles of parallel chains inside the soil. The motility inhibitor L-ethionine failed to suppress outgrowth, ruling out translocation by a flagellar-mediated mechanism such as swimming or swarming. Bacillus subtilis subsp. subtilis Marburg and four Bacillus isolates taken at random from soils also displayed a life cycle in SESOM and ASM and were all able to translocate through ASM, even in presence of L-ethionine. These data indicate that B. cereus is a saprophytic bacterium that is able to grow in soil and furthermore that it is adapted to translocate by employing a multicellular mode of growth.

Antibiotics such as chlortetracycline (CTC) have been used to promote growth of pigs for decades, but concerns over increased antibiotic-resistant infections in humans have prompted the development of alternative strategies. Developing alternatives to antibiotic growth promoters (AGPs) could be informed by information on the mechanisms of growth promotion, notably, how AGPs affect the microbial populations of the gastrointestinal tract. Pigs from three sows were aseptically delivered by cesarean section. Six piglets were distributed to each of two foster mothers until weaning, when piglets were fed a diet with or without 50 mg/kg CTC for 2 weeks. The ileal bacterial microbiota was characterized by using a cultivation-independent approach based on DNA extraction, PCR amplification, cloning, and sequencing of the 16S rRNA gene pool. The ileal and mucosal communities of these growing pigs were dominated by Lactobacillus bacteria, various members of the family Clostridiaceae, and members of the poorly known genus Turicibacter. Overall, CTC treatment resulted in three shifts: a decrease in Lactobacillus johnsonii, an increase in L. amylovorus, and a decrease in Turicibacter phylotypes. The composition of the microbiota varied considerably between individual pigs, as revealed by shared operational taxonomic units (OTUs) and similarity (SONS) analysis ( YC values). While the observed variation between untreated pigs obscured the possible effect of CTC, ͐-LIBSHUFF and SONS analyses of pooled libraries indicated a significant shift due to CTC in both the lumen and the mucosa, with some OTUs unique to either treated or control ileum. DOTUR analysis revealed little overlap between control and treated communities at the 3% difference level, indicating unique ileal communities in the presence of CTC.

BackgroundBacterial biofilms are predominant in natural ecosystems and constitute a public health threat because of their outstanding resistance to antibacterial treatments and especially to antibiotics. To date, several systems have been developed to grow bacterial biofilms in order to study their phenotypes and the physiology of sessile cells. Although relevant, such systems permit analysis of various aspects of the biofilm state but often after several hours of bacterial growth.ResultsHere we describe a simple and easy-to-use system for growing P. aeruginosa biofilm based on the medium adsorption onto glass wool fibers. This approach which promotes bacterial contact onto the support, makes it possible to obtain in a few minutes a large population of sessile bacteria. Using this growth system, we demonstrated the feasibility of exploring the early stages of biofilm formation by separating by electrophoresis proteins extracted directly from immobilized cells. Moreover, the involvement of protein synthesis in P. aeruginosa attachment is demonstrated.ConclusionsOur system provides sufficient sessile biomass to perform biochemical and proteomic analyses from the early incubation period, thus paving the way for the molecular analysis of the early stages of colonization that were inaccessible to date.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-014-0253-z) contains supplementary material, which is available to authorized users.