Global Adaptations Resulting from High Population Densities in Escherichia coli Cultures

Abstract

The scope of population density effects was investigated in steady-state continuous cultures of Escherichia coli in the absence of complications caused by transient environmental conditions and growth rates. Four distinct bacterial properties reflecting major regulatory and physiological circuits were analyzed. The metabolome profile of bacteria growing at high density contained major differences from low-density cultures. The 10-foldelevated level of trehalose at higher densities pointed to the increased role of the RpoS sigma factor, which controls trehalose synthesis genes as well as the general stress response. There was an eightfold difference in RpoS levels between bacteria grown at 10 8 and at 10 9 cells/ml. In contrast, the cellular content of the DNA binding protein H-NS, controlling many genes in concert with RpoS, was decreased by high density. Since H-NS and RpoS also influence porin gene expression, the influence of population density on the intricate regulation of outer membrane composition was also investigated. High culture densities were found to strongly repress ompF porin transcription, with a sharp threshold at a density of 4.4 ؋ 10 8 cells/ml, while increasing the proportion of OmpC in the outer membrane. The density-dependent regulation of ompF was maintained in rpoS or hns mutants and so was independent of these regulators. The consistently dramatic changes indicate that actively growing, high-density cultures are at least as differentiated from low-density cultures as are exponential-from stationary-phase bacteria.As originally described for luminescent organisms, population density is an important factor in bacterial physiology (7, 23). Nevertheless, a distinct problem in studying population density-dependent regulation is that it is difficult to unravel the complex effects operating in commonly used batch cultures. There is no simple way with batch cultures to uncouple population density-and growth-phase effects, which affect expression of dozens of genes approaching stationary phase (11). Likewise, it is unclear whether the extracellular molecules found in spent media of Escherichia coli (3, 31, 36, 42) result from growth phase or population density effects. The purpose of this study was to identify the extent of regulation in E. coli affected purely by population density, as is possible in continuous cultures growing at the same growth rate but with predetermined, steady-state bacterial densities. As shown below, at least four global properties of E. coli are strongly influenced by population density in the absence of growth phase effects.The macromolecular composition and metabolic activity of microbes are known to vary considerably with changes in growth conditions (24, 38). The metabolome, or global pattern of all metabolites, is a function of environmental conditions in steady-state cultures (39,40). Population density may well affect the metabolome, if indeed the extracellular metabolites in spent media result from shifts in intracellular metabolism. In this study, we investigate...

An analysis of multifactorial influences on the transcriptional control of ompF and ompC porin expression under nutrient limitation

“…The amount of RpoS was analysed by Western blotting. The procedures for protein extraction, SDS-PAGE separation, electrophoretic transfer and immunodetection of RpoS were as previously described (Liu et al, 2000), and as specified in the legends.…”

“…Immunoblotting using anti-RpoS antibodies was performed as described by Liu et al (2000). Equivalent numbers of bacteria from glucose-and N-limited chemostats were extracted by the SDSboiling method, from chemostats run at the dilution rates shown.…”

Differential biofilm formation and chemical disinfection resistance of Escherichia coli on stainless steel and polystyrene tissue culture plate

“…42 Slow growth and/or induction of an rpo S -mediated stress response could contribute to biocide resistance. 43,44 The physical and/or chemical structure of exopolysaccharides or other aspects of biofilm architecture could also confer resistance by exclusion of biocides from the bacterial cells (Cabecal et al,. 23 Finally, biofilm-grown bacteria might develop a biofilm specific biocide-resistant phenotype.…”

Kinetic behavior of the major multidrug efflux pump AcrB of Escherichia coli

“…Thus it was often necessary to decrease the expression of the wider OmpF porin to decrease V in so that it will be similar to Ve. We thus used cells in early stationary phase (36). With cefazolin, OmpF was further repressed by adding 5 mM sodium salicylate to the medium (37).…”