EFFECTS OF MEDIUM CARBON-TO-NITROGEN RATIO ON BIOFILM FORMATION AND PLASMID STABILITY

Biofilm formation and plasmid segregational instability in biofilm cul tures of Escherichia coli DH5 alpha (pMJR1750) were investigated under different medium carbon-to-nitrogen (CIN) ratios. At CIN ratios of 0. 07 and 1, net accumulation of both biofilm plasmid-bearing and plasmid -free cells continued throughout the entire experiment without attaini ng any apparent steady state. At CIN ratios of 5 and 10, net biofilm c ell accumulation for the two populations reached apparent steady state s after 84 and 72 h, respectively. At CIN ratios of 0.07 and 1, polysa ccharide production increased slowly and reached about 2 mu g alginate equivalent/cm(2) by the end of both experiments. At a C/N ratio of 5, polysaccharide increased significantly after 84 h, reaching about 7 m u g alginate equivalent/cm(2) prior to termination. At a C/N ratio of 10, polysaccharide increased significantly after 72 h and reached 21 m u g alginate equivalent/cm(2) at 108 h. At C/N ratios of 0.07 and 1, p rotein production reached 6.5 and 4 mu g/cm(2), respectively. At C/N r atios of 5 and 10, protein production increased slightly for the first 84 h and reached a maximum at 108 h, at 3 and 2 mu g/cm(2), respectiv ely, then decreased over the last 12 h of the experiment. Ratios of po lysaccharide to protein increased with increasing C/N ratios. At C/N r atios of 0.07 and 1, the ratios between extracellular polysaccharide ( EP) and protein were no more than 2.5 mu g polysaccharide/mu g protein , whereas those at CIN ratios of 5 and 10 increased to about 7 and 12 mu g polysaccharide/mu g protein, respectively. Probabilities, of plas mid loss in the biofilm cultures increased with increasing CIN ratios. At CIN ratios of 0.07, 1, and 5, the probabilities of plasmid loss we re 0.013 +/- 0.011, 0.020 +/- 0.006 and 0.122 +/- 0.021, respectively. At a C/N ratio of 10, the probability of plasmid loss was significant ly higher, reaching 0.388 +/- 0.125. The increase of probability of pl asmid loss at higher CIN ratios results from competition between cell replication and extracellular polysaccharide production. (C) 1994 John Wiley & Sons, Inc.