Journal
PLOS ONE
Volume 13, Issue 7, Pages -Publisher
PUBLIC LIBRARY SCIENCE
DOI: 10.1371/journal.pone.0200011
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Funding
- Spanish Ministry of Economy, Industry and Competitiveness (INIA) [RTA2014-00045-C03-02]
- Regional Government of Castile and Leon (Operational Programme for Youth Employment, European Social Fund and Youth Employment Initiative)
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Salmonella is a major food-borne pathogen able to persist in food processing environments because of its ability to form biofilms. A Salmonella enterica serotype Agona isolate from poultry (S24) was grown at 37 degrees C in biofilms for up to 144 hours (H144) in attachment to polystyrene surfaces. Biofilm structures were examined at different stages in their development (H3, H24, H48, H72, H96 and H144) using confocal laser scanning microscopy (CLSM) in conjunction with fluorescent dyes for live cells (SYTO 9), dead cells (propidium iodide), proteins (fluorescein isothiocyanate isomer I), lipids (DiD'oil), alpha-polysaccharides (concanavalin A, tetramethylrhodamine conjugate), and beta-polysaccharides (calcofluor white M2R). Strain S24 developed a robust biofilm at H72 (biovolume of 166,852.5 +/- 13,681.8 mu m(3) in the observation field of 16,078.2 mu m(2)). The largest biovolume of live cells was also detected at H72 (128,110.3 +/- 4,969.1 mu m(3)), decreasing thereafter, which was probably owing to the detachment of cells prior to a new phase of colonization. The percentage of dead cells with regard to total cells in the biofilms increased throughout the incubation, ranging from 2.3 +/- 1.1% (H24) to 44.2 +/- 11.0% (H144). Proteins showed the greatest biovolume among the extracellular components within the biofilms, with values ranging from 1,295.1 +/- 1,294.9 mu m(3)(H3) to 19,186.2 +/- 8,536.0 mu m(3) (H96). Maximum biovolume values of 15,171.9 +/- 660.7 mu m(3) (H48), 7,055.3 +/- 4,415.2 mu m(3) (H144), and 2,548.6 +/- 1,597.5 mu m(3) (H72) were observed for beta-polysaccharides, alpha-polysaccharides and lipids, respectively. A strong (P < 0.01) positive correlation was found between the total biovolume of biofilm and the biovolume of live cells, proteins and beta-polysaccharides, which may serve as useful markers of biofilm formation. The present work provides new insights into the formation of S. Agona biofilms. Our findings may contribute to the designing of reliable strategies for preventing and removing these bacterial communities.
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