Fig. 3

Microfluidics for bacterial-fungal interaction studies. a The spoke-wheel microfluidic design incorporates ports for separate, spatially-defined inoculations of fungi and bacteria. b Shown are green dye-filled channels (loaded with vacuum-assisted filling) in a glass-bottomed dish. An agar plug fills the center of the chamber introduced through the center culture well and aspirated through the agar exchange port. The agar plug holds the fungal inoculum in place and minimizes dehydration during growth. For scale, the open glass surface (between black arrows) is 30 mm. c Average number of hyphae per primary and secondary radiating channels from a single dish (15 DIV). The inset image shows channel locations that correspond to the graphed data. d, e Bacterial-fungal interactions are conveniently established and imaged with microfluidic systems. Image d and inset e of Pseudomonas fluorescens GM41 navigating the Laccaria bicolor fungal highway within a microfluidic chamber (3 DIV bacteria co-culture). f(i, ii) P. fluorescens GM41 bacterial communities accumulate where fungi contact the glass (i) or PDMS (ii) surface, preferentially forming at the PDMS-hyphae surface d over the glass-hyphae interface (30 DIV co-culture). g In the same device architecture, Pseudomonas fluorescens BBc6 biofilm-like accumulation on ectomycorrhizal fungi (L. bicolor S238N) 16 h after bacterial inoculation. h Vacuum-packed spoke-wheel microfluidics are permissive for even the most sensitive cell cultures, neurons. Here, neurons (DIV 4) were transfected for molecular imaging studies (VAMP2, magenta; PSD-95, green)