Figure 1.
Phospholipid (PL) profiles by 2D-TLC.
Phospholipid spots shown in 2D-TLC plates were developed with CuSO4 (100 mg/ml) spray containing 8% phosphoric acid and heated at 180°C. ACP-1 and ACP-2, amino-containing PL-1 and -2; I-LPG, inner CM layer-lysyl-phosphatidylglycerol; O-LPG, outer CM layer lysyl-phosphatidylglycerol; PG, phosphatidylglycerol; CL,cardiolipin; GP-DGDAG, glycerolphospho-diglycodiacylglycerol; PA, phosphatidic acid; Un, unidentified phospholipid. Red arrows indicate the major spots confirmed by LC/ESI-MS/MS analysis. The spot of fluorescamine-labelled O-LPG was detected by UV, indicating flipping of the latter PL to the outer CM.
Table 1.
Cell membrane phospholipid (PL) composition and asymmetry in the E. faecalis and E. faecium clinical strain-pairs.
Table 2.
Polarization index values as indicators of CM fluidity in the enterococcal clinical strain-pairs.
Table 3.
Fatty acid composition of the E. faecalis and E. faecium clinical strain-pairs.
Figure 2.
Transmission electron microscopy of cell envelope of Enterococcus faecium isolates (daptomycin-susceptible, S447 and daptomycin-resistant, R446).
At high magnification, cells in the S447 isolate have multiple septa with easily detectable separation (Panels A and B) and cell envelope of S447 have smooth appearance (panels C, D and E). In contrast, cells from the R446 isolate are in close proximity with each other (panel A). Dividing cells of the R446 isolate tend to have aberrant placement of septa (arrows) and cell envelopes appeared to be altered at forming septa (panels B and D). Localized protrusions of the cell envelope, usually within close proximity to a septal structure, are observed in cells of R446 (panels C, D, and E).
Figure 3.
CM depolarization by DiSC3(5) after daptomycin exposure of daptomycin-susceptible (S447) and daptomycin-resistant (R446) Enterococcus faecium.
Data represent the mean (±SD). * p<0.05; ** p<0.001.
Figure 4.
CM permeabilization by LIVE/DEAD BacLight after daptomycin exposure of daptomycin-susceptible (S447) and daptomycin-resistant (R446) E. faecium.
Data represent mean (±SD); * p<0.05, ** p<0.001.