An Engineered Approach to Fungal Growth in Rhizosphere

Microbial communities play a critical role in shaping the rhizosphere, physically and chemically modifying their local environment through a dynamic exchange with growing plants and other community members. In this manner, they can dramatically impact plant health, and moderate the cycling of compounds that impact climate. The complex interactions among bacteria, fungi, and plants, the main components of the soil, remain largely unidentified. In this study, we describe the development and use of an engineered microhabitat to visualize and quantify the impact of chemical signals, physical confinement, and bacterial-fungal interactions (BFIs) on fungal growth in a microfluidic channels and microfluidic soil-analog devices. Generally, the microfluidics were fabricated using photolithography and conventional soft lithography methods and assembled to glass slides or nano-porous membranes to facilitate culture in environments varied levels of hydration. Single hypha were directed to grow through channels with well-defined pore widths and well-controlled nutrient supplies. The response of fungi to chemical gradients was recorded using time-lapse microscopy and hyphal elongation rates were determined via image analysis. After the fungal response to specific chemical gradients was recorded, hyphal elongation rates were measured in response to the presence of different microbial isolates. Elongation rates, bacterial growth and co-localization were visualized and quantified. Our results demonstrate a microfluidics approach to the systematic and quantitative examination of complex signal exchange and physical organization during multi-kingdom interactions in structured environments comparable to the plant microbiome.