Sporulation of Bacillus spp. within biofilms: a potential source of contamination in food processing environments.

Bacillus strains are often isolated from biofilms in the food industries. Previous works have demonstrated that sporulation could occur in biofilms, suggesting that biofilms would be a significant source of food contamination with spores. In this study, we investigated the properties of mono-species and mixed Bacillus biofilms and the ability of Bacillus strains to sporulate inside biofilms. Bacillus strains were able to form mono-species biofilms on stainless steel coupons, with up to 90% spores after a 48 h-incubation. These spores were highly resistant to cleaning but were easily transferred to agar, mimicking the cross-contamination of food, thereby suggesting that biofilms would be of particular concern due to a potential for Bacillus spore food contamination. This hypothesis was strengthened by the fact that Bacillus strains were able to form mixed biofilms with resident strains and that sporulation still occurred easily in these complex structures.

Viability and surface properties of spores subjected to a cleaning-in-place procedure: consequences on their ability to contaminate surfaces of equipment.

This study was designed to evaluate how conditions encountered by spores during cleaning-in-place (CIP) procedures affected their surface properties, their viability and ability to contaminate materials. Spores from five Bacillus cereus strains were treated with NaOH at high temperature. Results revealed that high temperatures (exceeding 60 degrees C) and NaOH concentrations (over 0.5%) were required to significantly decrease spore viability (3-5log decrease). In these conditions, modifications were also clearly observed by microscopy to various surface structures of spores (appendages, exosporium, and especially to the hair-like nap) but also to their coat. Therefore, the ability of culturable spores to adhere decreased for the majority of strains tested. We then demonstrated that spores in suspension in NaOH could adhere to surfaces of a CIP rig and that the contamination level was controlled by flow pattern. Consequently, re-adhesion along the processing line might occur during CIP procedures and this phenomenon must be taken into account when defining cleaning strategies.

The mdoC gene of Escherichia coli encodes a membrane protein that is required for succinylation of osmoregulated periplasmic glucans.

Osmoregulated periplasmic glucans (OPGs) of Escherichia coli are anionic oligosaccharides that accumulate in the periplasmic space in response to low osmolarity of the medium. Their anionic character is provided by the substitution of the glucosidic backbone by phosphoglycerol originating from the membrane phospholipids and by succinyl residues from unknown origin. A phosphoglycerol-transferase-deficient mdoB mutant was subjected to Tn5 transposon mutagenesis, and putative mutant clones were screened for changes in the anionic character of OPGs by thin-layer chromatography. One mutant deficient in succinylation of OPGs was obtained, and the gene inactivated in this mutant was characterized and named mdoC. mdoC, which encodes a membrane-bound protein, is closely linked to the mdoGH operon necessary for the synthesis of the OPG backbone.