Figure 3.

T7SS mutants display increased membrane permeability upon LA binding. (A) Chemical structure of azide functionalised linoleic acid (azide-LA; N⁶-diazo-N²-((9Z,12Z)-octadeca-9,12-dienoyl)lysine, N₃-LA). Highlighted in green is the azido lysine. (B) S. aureus USA300 WT, ΔessC, and ΔesxC were grown with shaking in TSB to OD₆₀₀ of 1.0. Bacteria were then stained for 15 min with 10 µM azide-LA prior to labelling for 1 h with alkyne Alexa Fluor 488. Mean percentage of fluorescence values relative to WT (100%) are presented; error bars represent SD, n = 5. (C) Micrographs of bacteria grown in TSB and treated as described in (B) and additionally stained with propidium iodide (PI). (D) ImageJ was used to quantitate PI fluorescence of bacterial clusters from 12 different fields per strain. Each box‐and‐whisker plot depicts the minimal and maximal PI intensities, the median is the vertical bar inside the box, which is delimited by the lower and upper quartiles. **Indicates P < 0.01 using one-way ANOVA with Dunnett’s test.