Signatures of Mollicutes-related endobacteria in publicly available Mucoromycota genomes

Abstract

Mucoromycota fungi and their Mollicutes-related endobacteria (MRE) are an ideal system for studying bacterial-fungal interactions and evolution due to the long-term and intimate nature of their interactions. However, methods for detecting MRE face specific challenges due to the poor representation of MRE in sequencing databases coupled with the high sequence divergence of their genomes, making traditional similarity searches unreliable. This has precluded estimations on the diversity of MRE associated with Mucoromycota. To determine the prevalence of previously undetected MRE in fungal genome sequences, we scanned 389 Mucoromycota genome assemblies available from the National Center for Biotechnology Information for the presence of MRE sequences using publicly available tools to map contigs from fungal assemblies to publicly available MRE genomes. We demonstrate a higher diversity of MRE genomes than previously described in Mucoromycota and a lack of cophylogeny between MRE and the majority of their fungal hosts. This supports the late invasion hypothesis regarding MRE acquisition across most of the examined fungal families. In contrast with other Mucoromycota lineages, MRE from the Gigasporaceae displayed some degree of cophylogeny with their hosts, which may indicate that horizontal transmission is restricted between members of this family or that transmission is strictly vertical. These results underscore the need for a refined process to capture sequencing data from potential fungal endosymbionts to discern their evolution and transmission. Screens of fungal genomes for MRE can help improve the quality of fungal genome assemblies while identifying new MRE lineages to further test hypotheses on their origin and evolution.IMPORTANCEMollicutes-related endobacteria (MRE) are obligate intracellular bacteria found within Mucoromycota fungi. Despite their frequent detection, MRE roles in host functioning are still unknown. Comparative genomic investigations can improve our understanding of the impact of MRE on their fungal hosts by identifying similarities and differences in MRE genome evolution. However, MRE genomes have only been assembled from a small fraction of Mucoromycota hosts. Here, we demonstrate that MRE can be present yet undetected in publicly available Mucoromycota genome assemblies. We use these newfound sequences to assess the broader diversity of MRE and their phylogenetic relationships with respect to their hosts. We demonstrate that publicly available tools can be used to extract novel MRE sequences from assembled fungal genomes leading to insights on MRE evolution. This work contributes to a greater understanding of the fungal microbiome, which is crucial to improving knowledge on the dynamics and impacts of fungi in microbial ecosystems.

Keywords: Mollicutes; bacterial–fungal interactions; bioinformatics; endobacteria; genomics; intermicrobial; mucormycota.

Fig 1

Phylogenetics provides further support for the late invasion hypothesis due to broad lack of cophylogeny between MRE and fungal hosts. (A) Maximum likelihood phylogeny of MRE extracted from fungal assemblies and previously published MRE genome assemblies. (B) Maximum likelihood phylogeny of fungal hosts of MRE created using 22 shared BUSCO genes. (Table S3). ** represents novel MRE assemblies identified from fungal genome assemblies. * represents MRE assemblies binned from publicly available Endogonales MRE meta-assembly. Lines connect MRE taxa to their fungal host species. Branch labels indicate bootstrap values (1,000 replicates). Bootstrap values > 50 are shown.

Fig 2

Heatmap showing pairwise AAI between MRE taxa. AAI is below 70% between most MRE taxa, but MRE from Gigasporaceae hosts generally display higher AAI. Black rectangle indicates MRE from Gigasporaceae hosts.