Figure 5
CLSEM for analyses of adhesion to, and invasion of polarized MDCK cells by Salmonella enterica. Polarized Lifeact-eGFP MDCK cells (green) were infected with STM WT (red). LCI was performed by SDCM and monolayers were fixed 17 min post infection (p.i.). (A) Time series showing the accumulation of F-actin into membrane ruffle in a frame interval of 25.43 s. The corresponding image sequence recorded over 17 min is shown in Movie 2. (B) Overlay of SDCM and SEM images for correlation of appropriate structures. (C) Representative morphological changes of the host cell membrane during adhesion and invasion of STM at individual ageing stages during point of fixation. (i) Early contact of STM to apical surface of MDCK. F-actin accumulation not yet visible. (ii) Triangular-shaped membrane ruffle (MR) already visible in SDCM. (iii) STM in SEM visible below a very thin carbon film. (iv) STM in the centre of MR. STM, which did not adhere tightly in SDCM, are no more visible in SEM. (v) MR retraction and reduced F-actin accumulation. Three STM were detected in SDCM, two STM were visible in the centre of MR in SEM modality, where later the Salmonella-containing vacuole (SCV) is formed. ROI is indicated by dotted circle. (vi) Complete internalization of STM into host cell in SDCM modality. Absence of MR and F-actin almost reorganized into brush border. Not all microvilli were fully recovered at this time point. Time stamp, min:sec. Scale bars, 1 µm.
