Microfluidic monitoring of the growth of individual hyphae in confined environments

Soil fungi have the ability to form large mycelial networks. They rely on the resources available in the soil to produce biomass and are able to degrade complex biomolecules. Some of them can even degrade recalcitrant organic pollutants and are considered as promising candidates for soil bioremediation strategies. However, the success of this approach depends on the ability of fungi to colonize the soil matrix, where they encounter spatial and temporal variations of confinement, humidity and nutrient concentration. In this paper, we present a study of fungal growth at the scale of single hyphae in a microfluidic device, allowing fine control of nutrient and water supply. Time-lapse microscopy allowed simultaneous monitoring of the growth of dozens of hyphae of Talaromyces helicus, a soil isolate, and of the model fungus Neurospora crassa through parallel microchannels. The distributions of growth velocity obtained for each strain were compared with measurements obtained in macroscopic solid culture. For the two strains used in the study, confinement caused the growth velocity to drop in comparison with unconfined experiments. In addition, N. crassa was also limited in its growth by the nutrient supply, while the microfluidic culture conditions seemed better suited for T. helicus. Qualitative observations of fungi growing in microfluidic chambers without lateral confinement also revealed that side walls influence the branching behaviour of hyphae. This study is one of the first to consider the confinement degree within soil microporosities as a key factor of fungal growth, and to address its effect, along with physicochemical parameters, on soil colonization, notably for bioremediation purposes.