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Arbuscular mycorrhizal fungi (AMF) form symbiotic associations with the majority of land plants and deliver a wide range of soil-based ecosystem services. Due to their conspicuous belowground lifestyle in a dark environment surrounded by soil particles, much is still to be learned about the influence of environmental ( i.e. , physical) cues on spore germination, hyphal morphogenesis and anastomosis/hyphal healing mechanisms. To fill existing gaps in AMF knowledge, we developed a new microfluidic platform - the AMF-SporeChip - to visualise the foraging behaviour of germinating Rhizophagus and Gigaspora spores and confront asymbiotic hyphae with physical obstacles. In combination with timelapse microscopy, the fungi could be examined at the cellular level and in real-time. The AMF-SporeChip allowed us to acquire movies with unprecedented visual clarity and therefore identify various exploration strategies of AMF asymbiotic hyphae. We witnessed tip-to-tip and tip-to-side hyphal anastomosis formation. Anastomosis involved directed hyphal growth in a "stop-and-go" manner, yielding visual evidence of pre-anastomosis signalling and decision-making. Remarkably, we also revealed a so-far undescribed reversible cytoplasmic retraction, including the formation of up to 8 septa upon retraction, as part of a highly dynamic space navigation, probably evolved to optimise foraging efficiency. Our findings demonstrated how AMF employ an intricate mechanism of space searching, involving reversible cytoplasmic retraction, branching and directional changes. In turn, the AMF-SporeChip is expected to open many future frontiers for AMF research.

Soil is the habitat of countless organisms and encompasses an enormous variety of dynamic environmental conditions. While it is evident that a thorough understanding of how organisms interact with the soil environment may have substantial ecological and economical impact, current laboratory-based methods depend on reductionist approaches that are incapable of simulating natural diversity. The application of Lab-on-a-Chip or microfluidic technologies to organismal studies is an emerging field, where the unique benefits afforded by system miniaturisation offer new opportunities for the experimentalist. Indeed, precise spatiotemporal control over the microenvironments of soil organisms in combination with high-resolution imaging has the potential to provide an unprecedented view of biological events at the single-organism or single-cell level, which in turn opens up new avenues for environmental and organismal studies. Herein we review some of the most recent and interesting developments in microfluidic technologies for the study of soil organisms and their interactions with the environment. We discuss how so-called "Soil-on-a-Chip" technology has already contributed significantly to the study of bacteria, nematodes, fungi and plants, as well as inter-organismal interactions, by advancing experimental access and environmental control. Most crucially, we highlight where distinct advantages over traditional approaches exist and where novel biological insights will ensue.