Fig. 2: Dap action on B. subtilis strains (WT, ΔmreB and ΔugtP strains) and POPG vesicles by the FliptR lipid packing probe.

FliptR signal: lower lifetimes correlate to lower lipid packings and vice versa. a FliptR image on WT cells: in the absence of Dap, all the cells keep the same distribution, the bacterial poles display higher packing levels than the bacterial sides, a bimodal distribution of lipid packing values appears; at sub-MIC, at the poles, lipid packing level rises; at over-MIC, an inter-cell heterogeneity appears, certain cells show high lipid packing disseminated over the full cell (orange arrow) while others maintain their previous packing distribution (blue arrow), the distribution of packing stretches and a little peak at higher levels of packing becomes apparent. At higher magnification (zoom) also heterogeneity at the intra-cell level becomes evident, zones of high (orange arrow) and low (blue arrow) lipid packing disseminate inside the cell, in agreement with previous observations¹⁹. It is observed that the deformations protruding from the cells present higher levels of lipid packing than the zones that do not protrude. b, c For the ΔmreB and ΔugtP mutants, FliptR signals are akin to WT strain. Yet, the inter-cell heterogeneity appears at lower Dap concentration, sub-MIC instead of over-MIC, supporting that ‘healthier’ bacterial cell wall limits the deformations induced by Dap. d Tracking of the progression of the lipid packing on POPG vesicles exposed to over-MIC Dap by FliptR. Dap is injected at the ‘Dap IN’ label and a two-step process takes place. First the vesicles rigidify, then they collapse and the lipid packing is drastically reduced to values below the initial ones. This two-step process could explain the two zones of low and high lipid packing identified in the bacteria cells exposed to Dap.