Fig. 3

N. meningitidis interaction with the iBEC-LMC direct co-culture model. Models were infected from the iBEC side (MOI = 10, unless specified otherwise) on day 2 of co-culture for the times indicated. a Enumeration of adherent and intracellular cfu/monolayer on iBEC layers with or without LMC co-culture after infection with N. meningitidis at an MOI of 100, determined by gentamicin protection assays. b TEM of iBEC-LMC co-culture models infected with N. meningitidis for 6 h (I), 24 h (II), and 30 h (III, IV). Scale bars represent 1 µm, unless labeled otherwise. c Structured illumination microscopy showing iBEC or LMC layers from iBEC-LMC co-culture, stained for f-actin (phalloidin-546, magenta) and DNA (DAPI, blue) after 24 h of infection with a GFP expressing N. meningitidis strain. Images presented as maximum intensity projection in Z and including maximum intensity projections in X and Y (orthoslices) as indicated by the crosshairs. Scale bars represent 7 µm. d N. meningitidis transmigration rates determined by enumeration of cfu in the basolateral compartment after 1 h of incubation in fresh basolateral media following the indicated infection time points. e TEER of infected and uninfected iBEC monoculture and iBEC-LMC co-culture over a time-course of 32 h post-infection. Data presented as mean ± SD from three independent experiments and iBEC differentiations performed in duplicate (a, n = 6) or triplicate (d, e; n = 9). *p < 0.05, **p < 0.01, ***p < 0.001; Student’s t test; direct co-culture (blue) vs monoculture (grey) (d), infected vs uninfected control (e)