Slower growth of E. coli leads to longer survival in carbon starvation (2020)
Figure 2

Figure 2: Quantification of maintenance rate and recycling yield of E. coli.

(A) In carbon starvation death rate γ, is determined by ratio of maintenance rate β to the recycling yield α (Schink et al., 2019).

(B) Death rates of wild-type and GlpK22 mutants plotted versus growth rates. Wild type is grown in glycerol minimal medium in batch cultures (black circle) and in 'chemostat' continuous cultures with growth rates coded in colors, see legend on the center right. GlpK22 mutants (white circle) are grown in glycerol minimal medium in batch culture. Data shown as mean ± (standard deviation) SD. Two replicates per condition. The black line shows the fit of Fig. 1C.

(C) Graphical synopsis of the assay to measure recycling yield: At different times during starvation, a sample is (1) extracted from a starved culture, (2) sterilized with UV-light (3) inoculated with 1% of the original starved culture and (4) growth is measured using plate counting.

(D) Example experiment of the assay sketched in panel C. Cells previously grown at 0.3 h − 1 in continuous culture are starved (green circles). After 2 days of starvation, a sample is (1) extracted and (2) UV-sterilized, see dashed line, followed by (3) inoculation of 1% of starved and (4) growth of the culture. The difference between the maximum viability after growth, and the initial viability at point (3) is the absolute yield.

(E) The absolute yield measured at three different time points for each of the growth conditions shown in panel A is plotted against the viability of the starvation culture at the extraction time. The recycling yield is extracted as the slope of the linear fits.

(F) Recycling yield from panel E plotted versus growth rate. Data shown as mean ± SD. Two replicates per condition.

(G) Graphical synopsis of the assay to measure the maintenance rate. At one point during starvation, a sample (1) extracted from a starvation culture, and split into several tubes. (2) Different concentrations of glycerol are added to each tube, small enough to support survival, but not growth and (3) viability is measured.

(H) Example experiment from a starved culture previously grown at a rate of 0.3 h − 1 with a viability of $\frac{{1.21 \bullet 10}^{8}\ \text{CFU}}{\text{ml}}$ after 5 days of starvation. After (1) extraction and (2) addition of 40 µM of glycerol the decay of viability is delayed (black circles) compared to a control without glycerol (green symbols). Per viable cell, the glycerol addition in this experiment is 0.33 fmol CFU − 1. After an initial period of survival, the culture with added glycerol (black) dies at the same rate as the control (green). The 'lag time' fitted to this data is 1 day.

(I) Lag time of the experiments of panel B for different glycerol concentrations. Lag increases linearly with glycerol concentration and maintenance rate is extracted as the inverse of the slope of the linear fits.

(J) Maintenance rate values extracted from panel I plotted versus growth rate. Data shown as mean ± SD. Two replicates per condition.

(K) Consistency check of the equation in panel A. Fold changes of maintenance rate, FC(β), divided by the fold changes of the recycling yield, FC(α), plotted versus the fold changes of death rate, FC(γ). All fold changes are relative to wild-type in batch, growing at 0.7 h − 1. The dashed line shows the unity line FC(γ)= FC(β)/FC(α).

Data information: Symbol color is identical in all panels and depicted in the legend on the right. Table EV2 shows growth rate, yield and maintenance rate for all experiments. Solid black lines show linear fits (on log transformed data, if necessary), using weighted least square fits. Slopes are reported with standard error and take uncertainty in data into account when necessary (see Methods).