Extended Data Fig. 8: The effect of cell wall targeting antibiotics on OMP biogenesis.

(A) Epifluorescence images of FepA in an inducible expression strain (GM07) treated with mecillinam. Shown are images of cells before and after induction with 0.4% arabinose. (B) Co-labelling of PG and OMP biogenesis in mecillinam treated cells. HADA and 0.4% arabinose were added simultaneously for 7 min. Shown are epifluoresence images (left) and pixel by pixel cytofluorogram of a single image (right). (C) Co-labelling of BamA and OMP biogenesis in aztreonam treated cells. Aztreonam was added 30 min before induction with 0.4% arabinose for an additional 7 min. (D) Co-labelling of PG and OMP biogenesis in aztreonam treated cells. HADA and 0.4% arabinose were added simultaneously for 7 min. Shown are epifluoresence images (left) and pixel by pixel cytofluorogram of a single image (right). (E) OMP biogenesis following the removal of aztreonam. Aztreonam was washed from the media at T = 0 min. OMP biogenesis was induced with 0.4% arabinose for 7 min prior to each time point. (F) Co-labelling of PG and OMP biogenesis 45 min after the removal of aztreonam from the media. HADA and 0.4% arabinose were added simultaneously for 7 min. (G) Comparison of the fluorescence intensity profiles of PG and OMP biogenesis across emerging septa as shown in panel F. Shown are profiles of cells at two different stages of septum formation (Group 1/Group 2). Inset displays the width of individual cells at the designated division plane. The chart demonstrates that PG biogenesis precedes the emergence of OMPs at division sites upon aztreonam removal. n_early = 50 cells, n_late = 30 cells. Statistical significance was calculated using two-tailed Student’s unpaired t-test (P = 0.0001). Scale bars, 1 μm.