Hidden Allies: Decoding the Core Endohyphal Bacteriome of Aspergillus fumigatus

Bacterial-fungal interactions that influence the behaviour of one or both organisms are common in nature. Well-studied systems include endosymbiotic relationships that range from transient to long-term associations. Diverse endohyphal bacteria associate with fungal hosts, emphasising the need to better comprehend the fungal bacteriome. We evaluated the hypothesis that Aspergillus fumigatus harbours an endohyphal community of bacteria that influence the host phenotype. We analysed whether 38 A. fumigatus strains show stable association with diverse endohyphal bacteria; all derived from single-conidium cultures that were subjected to antibiotic and heat treatments. The fungal bacteriome, inferred through analysis of bacterial diversity within the fungal strains (short- and long- read sequencing methods), revealed the presence of core endohyphal bacterial genera. Microscopic analysis further confirmed the presence of endohyphal bacteria. The fungal strains exhibited high genetic diversity and phenotypic heterogeneity in drug susceptibility and in vivo virulence. No correlations were observed between genomic or functional traits and bacteriome diversity, but the abundance of some bacterial genera correlated with fungal virulence or posaconazole susceptibility. The observed endobacteriome may play functional roles, for example, nitrogen fixation. Our study emphasises the existence of complex interactions between fungi and endohyphal bacteria, possibly impacting the phenotype of the fungal host, including virulence.