Light restores sporulation in Rhizopus microsporus cured of its endosymbionts, unveiling their role in fitness and virulence

Abstract Rhizopus microsporus is a major cause of mucormycosis, an infection caused by Mucorales that increasingly affects immunocompromised individuals. Certain isolates of R. microsporus harbor bacterial endosymbionts that regulate key fungal functions, particularly asexual and sexual reproduction, but these effects have been explored exclusively in environmental isolates. Although some clinical isolates contain Mycetohabitans endosymbionts, their influence on fungal reproduction remains unknown. This dependence on endosymbionts for asexual spore formation in environmental isolates has established the Rhizopus-Mycetohabitans association as a model for studying fungal-bacterial endosymbiosis, but it has also constrained functional comparative studies across environmental and clinical backgrounds. We show that light exposure partially restores asexual sporulation in endosymbiont-cured environmental strains, enabling the generation of isogenic sporulating lines. Transcriptomic analyses revealed that both light and endobacteria modulate overlapping signal transduction pathways, regulating the expression of conserved genes involved in asexual development in Mucorales and other fungi. Functional assays demonstrated that asexual spores from cured strains are viable; however, the presence of endosymbionts accelerates spore formation, enhances osmotic stress tolerance, and helps maintain cell-wall integrity. Cured strains exhibit altered membrane composition, including reduced ergosterol levels, which may contribute to their resistance to macrophage phagocytosis. Despite these compensatory adaptations, cured strains showed attenuated virulence in a murine mucormycosis model, highlighting the role of bacterial endosymbionts in fungal pathogenicity. The discovery of light-induced sporulation in cured strains provides a valuable experimental framework for future comparative studies requiring asexual spores, offering new opportunities to explore the role of fungal-bacterial endosymbiosis in fungal biology and human disease.