Diverse Bacterial Properties Affect Dispersal on the Fungal Highway

Abstract

Bacterial–fungal interactions are common in microbial communities and often promote bacterial dispersal along fungal hyphae, with the various mechanisms enabling this process collectively referred to as the fungal superhighway. Using a custom microfluidic device, diverse bacterial species were examined for their ability to traverse Aspergillus flavus hyphae in bacterial–fungal co-culture compared to monocultures, and factors influencing their movement were identified. Some non-motile or poorly motile species, including Staphylococcus aureus, showed enhanced dispersal specifically in bacterial–fungal co-culture. Mechanisms varied: for Listeria monocytogenes, flagella were essential and dispersal was impaired in flagellar mutants but enhanced in ∆mogR strains that upregulate flagellar expression. In contrast, Pseudomonas aeruginosa and Ralstonia solanacearum dispersed effectively even when tested with flagellar mutant strains, suggesting the presence of alternative motility mechanisms such as type IV pili- mediated movement. S. aureus exhibited quorum-sensing–dependent, wave-like dispersal, which was abolished in mutants deficient in phenol-soluble modulins (PSMs), amphipathic peptides with surfactant-like properties. It was also demonstrated that fungal hyphal morphology influences bacterial dispersal: altered hyphal branching patterns controlled by the fungal transcription factor ZfpA modulates S. aureus movement, with hyperbranching A. flavus strains promoting greater bacterial accumulation accelerating dispersal. A triculture system involving both L. monocytogenes and S. aureus with A. flavus revealed competitive dynamics, where L. monocytogenes restricted S. aureus dispersal on fungal hyphae, suggesting that interactions between bacteria can impact fungal-mediated transport. Our findings show that bacterial motility on A. flavus arises from diverse, species-specific mechanisms, including the presence of flagella, transcriptional regulation, surfactant production, and physical interactions shaped by fungal hyphal architecture.