Abstract
Compared to free (free-living) cells, biofilm cells show increased resistance and stability to high-pressure fermentation conditions, although the reasons underlying these phenomena remain unclear. Here, we investigated biofilm formation with immobilized Saccharomyces cerevisiae cells grown on fiber surfaces during the process of ethanol fermentation. The development of biofilm colonies was visualized by fluorescent labeling and confocal microscopy. RNA from yeast cells at three different biofilm development periods was extracted and sequenced by high-throughput sequencing. We quantitated gene expression differences between biofilm cells and free cells and found that 2098, 1556, and 927 genes were significantly differentially expressed, respectively. We also validated the expression of previously reported genes and identified novel genes and pathways under the control of this system. Statistical analysis revealed that biofilm genes show significant gene expression changes principally in the initial period of biofilm formation compared to later periods. Carbohydrate metabolism, amino acid metabolism, signal transduction, and oxidoreductase activity were needed for biofilm formation. In contrast to previous findings, we observed some differential expression performances of FLO family genes, indicating that cell aggregation in our immobilized fermentation system was possibly independent of flocculation. Cyclic AMP-protein kinase A and mitogen-activated protein kinase pathways regulated signal transduction pathways during yeast biofilm formation. We found that carbohydrate metabolism, especially glycolysis/gluconeogenesis, played a key role in the development of S. cerevisiae biofilms. This work provides an important dataset for future studies aimed at gaining insight into the regulatory mechanisms of immobilized cells in biofilms, as well as for optimizing bioprocessing applications with S. cerevisiae.
Highlights
Biofilms, as microbial communities, are dynamic environments wherein constituent cells propagate attached to biotic or abiotic surfaces (O’Toole et al, 2000; Hoyer, 2001; Reynolds and Fink, 2001)
When comparing gene expression levels observed in the three phases with those observed in free cells, we found that most gene expression differences occurred during the attachment period
Analysis of signaling pathways mediated by the differentially expressed genes identified in this study showed that biofilms are regulated by the MAPK and cAMP-protein kinase A (PKA) pathways
Summary
As microbial communities, are dynamic environments wherein constituent cells propagate attached to biotic or abiotic surfaces (O’Toole et al, 2000; Hoyer, 2001; Reynolds and Fink, 2001). The high antibiotic resistance of biofilms in the pathogenesis of some chronic human infections is widely accepted. Several in vitro model systems have been developed to mimic biofilm growth occurring on infected medical devices. These models have provided the foundation for the investigating biofilm composition, architecture, and mechanisms of drug resistance (Baillie and Douglas, 2000; Chandra et al, 2001; Ramage et al, 2002). The structure and shear strength of microbial biofilms have been determined by confocal laser-scanning microscopy and www.frontiersin.org
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