Toxin-producing endosymbionts shield pathogenic fungus against micropredators

Abstract

The phytopathogenic fungus Rhizopus microsporus harbours a bacterial endosymbiont (Mycetohabitans rhizoxinica) for the production of the toxin rhizoxin, the causative agent of rice seedling blight. This toxinogenic bacterial-fungal alliance is, however, not restricted to the plant disease, but has been detected in numerous environmental isolates from geographically distinct sites covering all five continents. Yet, the ecological role of rhizoxin beyond rice seedling blight has been unknown. Here we show that rhizoxin serves the fungal host in fending off protozoan and metazoan predators. Fluorescence microscopy and co-culture experiments with the fungivorous amoeba Protostelium aurantium revealed that ingestion of R. microsporus spores is toxic to P. aurantium. This amoebicidal effect is caused by the bacterial rhizoxin congener rhizoxin S2, which is also lethal towards the model nematode Caenorhabditis elegans. By combining stereomicroscopy, automated image analyses, and quantification of nematode movement we show that the fungivorous nematode Aphelenchus avenae actively feeds on R. microsporus that is lacking endosymbionts, while worms co-incubated with symbiotic R. microsporus are significantly less lively. This work uncovers an unexpected ecological role of rhizoxin as shield against micropredators. This finding suggests that predators may function an evolutionary driving force to maintain toxin-producing endosymbionts in non-pathogenic fungi.

Endofungal bacteria boost anthelminthic host protection with the biosurfactant symbiosin

“…The toxin is not only the major virulence factor, but additionally plays a role in the fungal protection. 61,62 Notably, also other Burkholderia-derived metabolites, like the toxin rhizonin or the antibiotic icosalide, were originally thought to be fungal metabolites and later proven to be of bacterial origin. 63,64 Although functions of these and other endobacterial metabolites include symbiosis promoting activites, 25,[65][66][67] host reproduction control 68 and pathogenicity causing traits, 32,63,69 research on their role in host protecting strategies against predators was only conducted recently.…”

Alternative in-vivo models of mucormycosis

“…These findings aligned with results from human macrophage assays, where endosymbiont-containing spores protected the fungus from phagocytosis. Similarly, Richter et al demonstrated that co-culturing the amoeba Prostelium aurantium with Rhizopus spores, hosting rhizoxin producing bacteria, resulted in detrimental effects of the toxin onto the amoeba, ultimately providing protection to the fungus (Richter et al, 2022). Whether virulence factors of other mucorales, such as those inducing apoptosis in macrophages as observed by López-Muñoz et al for M. circinelloides, originated from interactions between amoeba and this Mucor species requires verification in an amoeba model (Loṕez-Muñoz et al, 2018).…”

“…In 2022, Richter et al utilized C. elegans and another nematode, Aphelenchus avenae, to investigate the potential nematodocidial effects of the toxin rhizoxin, produced by endosymbiotic bacteria of Rhizopus microsporus (Richter et al, 2022). Their study revealed that the secondary metabolite exhibits toxic effects towards both organisms.…”

Detecting and characterizing new endofungal bacteria in new hosts: Pandoraea sputorum and Mycetohabitans endofungorum in Rhizopus arrhizus

“…Many EFBs are associated with the fungal phylum Mucoromycota, and belong to Betaproteobacteria (Burkholderia-related endobacteria, BREs) and Mollicutes (Mycoplasma-related endobacteria, MREs) (Pawlowska et al, 2018;Okrasińska et al, 2021;Richter et al, 2022;Uehling et al, 2023). The BRE Mycetohabitans rhizoxinica, highly dependent on its host R. microsporus, was protected by transcription activator-like (TAL) effectors, while produced toxins rhizoxin and rhizonin with implications in causing rice seedling blight disease and hepatotoxicity (Partida-Martinez and Hertweck, 2005;Partida-Martinez et al, 2007a;Richter et al, 2023).…”