Matrix is everywhere: extracellular DNA is a link between biofilm and mineralization in Bacillus cereus planktonic lifestyle

Abstract To date, the mechanisms of biomineralization induced by bacterial cells in context of biofilm formation remain unclear. Even less information is available on the biofilm formation in planktonic cells with relation to calcifying microorganisms. In this study, we show the details of the first hours of Bacillus cereus growth in a Ca2+- and urea-rich liquid medium, in which the formation of mineral calcium carbonate precipitates was induced. These precipitates were localized in the extracellular matrix (ECM) formed in planktonic culture at the stage of exponential growth and comprised of extracellular DNA, amyloid proteins and polysaccharides. While accumulation of extracellular polysaccharides and amyloids appeared to be independent of the presence of calcium and urea during the growth, accumulation of eDNA required the presence of both Ca and urea in the medium. Removal of eDNA, which was sensitive to treatment by DNase, did not affect other matrix components, but resulted in disruption of cell network formation and strong inhibition of the mineral formation. Our results indicate that the mechanism of biomineralization by planktonic B. cereus cells appeared to involve a combination of factors including changes in protein expression pattern in response to stress, accumulation of extracellular polysaccharides, protein amyloids and eDNA leading to formation of ECM, and changes in cell metabolism and the medium pH. It was eDNA in extracellular matrix that appeared to provide centers for calcium carbonate precipitation, thus being a necessary factor for triggering the biomineralization. Precipitation of amorphous CaCO3 on ECM was followed by crystal formation via the ACC-vaterite-calcite/aragonite pathway and formation of larger mineral aggregates in complex with extracellular polymeric substances.