Heterogeneity of Escherichia coli population by respiratory activity and membrane potential of cells during growth and long-term starvation

Abstract

Assessment of physiological states of individual bacterial cells can be useful in the monitoring of the biotechnological processes. Physiological heterogeneity of Escherichia coli population by respiration activity and membrane potential during growth and starvation in batch cultures was evaluated using 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) and bis-(1,3-dibutylbarbituric acid) trimethine oxo (DiBAC₄)(3) fluorescent probes in combination with flow cytometry. The shares of CTC-reducing cells (CTC(+)-cells) and cells with positively charged outside cytoplasmic membrane which were not stained by DiBAC₄(3) (ΔΨ(+)-cells), were 90% and 95% in the exponential phase of batch culture, respectively. After short-term starvation for 10 h, the shares of CTC(+)-cells and ΔΨ(+)-cells in the samples taken from the exponential phase dropped to 78% and 72%, respectively. After long-term starvation for 40 days, the share of CTC(+)-cells dropped to 5%, whereas the share of ΔΨ(+)-cells was about 50%. The conclusions from this research are as follows: (a) the physiological heterogeneity of bacterial population increased after starvation; (b) the cell respiratory activity is more sensitive to starvation than the cell membrane potential; (c) a probe for the cell membrane potential DiBAC₄(3) is more suitable than a probe for the cell respiratory activity CTC in the detection of viable indicator bacteria in environment.