Fig. 4

SspA-S47D ∆sspB imposes delay on spore revival. (a, b) Spores of PY79 (wild type, WT), AR186 (∆sspB), AR179 (∆sspA), AR195 (∆sspA ∆sspB), AR187 (sspA-S47A ∆sspB), and AR188 (sspA-S47D ∆sspB) were incubated at 37 °C in S7-defined medium (a) or LB (b) supplemented with L-Ala (10 mM) and optical density (OD₆₀₀) was measured at the indicated time points. Data are presented as a fraction of the initial OD₆₀₀ of the phase-bright spores. Decreasing OD₆₀₀ signifies spore germination while increasing OD₆₀₀ indicates spore outgrowth. The data points are averages of results obtained from three independent biological repeats. Error bars designate SD. c Spores of AR200 (sspA-gfp _A206K ∆sspB) and AR206 (sspA-S47D-gfp _A206K ∆sspB) were incubated in S7-defined medium supplemented with L-Ala (10 mM) and monitored by time lapse microscopy. Shown are phase-contrast (upper) and fluorescent (lower) images taken at the indicated time points. Arrows signify the dominating revival stages of each population at a given time point (germination, red; ripening, orange; elongation, green). Shown is a representative experiment out of three independent biological repeats. d Quantification analysis of the level of the GFP signal measured from the spores in (c) at the indicated time points, calculated as percent of the initial intensity at t = 0. For each time point, the calculated net average fluorescence intensity from at least three different fields (spores n >300) was averaged, and the intensity of wild type spores (PY79), lacking gfp, was subtracted. Error bars designate SD