Fig 8.

Fluorescent vancomycin targets positions of active peptidoglycan biogenesis, accumulating within septa in control cells and also in membrane patches and bent regions of sub-MIC daptomycin-treated cells. Cells were stained with the vital membrane stains FM 4-64 (1 μg/ml), DAPI (2 μg/ml), and a fluorescent derivative of vancomycin, as described in Materials and Methods. (A) In cells not treated with daptomycin, Van-BDP (0.5 μg/ml; ∼1× MIC) intensely stains septa and only very faintly stains the sidewalls, as previously shown (9, 17, 54). The septum in the middle of the cell is just beginning to divide and accumulates Van-BDP as a ring of fluorescence, which appears as two dots in the medial focal plan. Van-BDP forms an intense band at completed septa, as previously shown (9, 17, 54). Scale bar, 1 μm. (B) Two examples showing that Van-BDP localizes to the membrane patches and bent regions that are formed in cells grown in a sublethal concentration of daptomycin (2 μg/ml). Scale bar, 1 μm.