Starvation-survival in Escherichia coli
The population dynamics of carbon-starved E. coli K12 cultures was investigated. It was found that less cell lysis occurred when cells were previously grown in low glucose concentrations. Exponential-phase cells grown previously in 0.05% (w/v) glucose had survival rates comparable with their stationary-phase counterparts, suggesting that the rate of growth is more important in determining the outcome of starvation than the phase of batch culture growth. Long-termstarved cells (18-24 months) showed very little protein, DNA and RNA synthesis. Methionine was shown to alter the de novo synthesis protein profiles of longterm- starved cells and growth was seen to occur in the presence of methionine. This suggests that radio-labelling of proteins with 35S-methionine in these cells should be interpreted with care as the cells have been subjected to a nutrient upshift. Radio-labelling of proteins with 3H-leucine did not have the same effect. The ATP content of cells during prolonged incubation was shown to decrease in the first 48 hours incubation, increase until 5-7 days incubation then decrease after 7-8 days. After 13 days a slow, steady increase occurred. The ATP content of cells incubated for 16 days was higher than that of 48 hour-incubated cells. The physiology of long-term-starved cells was investigated with respect to their permeability to routine bacteriological stains ( e.g. DAPI, saffranin, Geimsa) and it was found that very few of these dyes were able to penetrate the cells, indicating that a decrease in cell permeability may be an important factor in survival as is seen in endospores of Bacillus species and swarmer cells of Rhodomicrobium vannielii and Caulobacter crescentus. Resistance of long-term starved cells to heat and biocide challenge was increased in comparison with exponential- and short-term (48 hour) stationary-phase cells and the resistance to biocides was shown to be retained through subsequent generations. Examination of the nucleoids of long-term-starved cells revealed that a more condensed form was present in cultures incubated for over 14 days, suggesting that dehydration of the DNA had occurred, similar to the situation found in endospores of Bacillus species and suggestive of dormancy. Analysis of outer-membrane proteins and lipopolysaccharide of long-term-starved cells showed that alterations occurred to the surface of the cells and it was demonstrated that hydrophobicity changes occurred. Hydrophobicity reached a maximum after 48 hours incubation then subsequently declined between days 2 and 3 which corresponded with an increase in cell numbers. Cell surface hydrophobicity was shown to be a potential method for separating heterogeneous, carbon-starved populations into homogeneous subpopulations. The data suggest that E. coli produces a dormant survival cell type which is morphologically and physiologically distinct from the parent cell.