Morels (Morchella spp.) are iconic edible mushrooms with a long history of human consumption. Some microbial taxa are hypothesized to be important in triggering the formation of morel primordia and development of fruiting bodies, thus, there is interest in the microbial ecology of these fungi. To identify and compare fungal and prokaryotic communities in soils where Morchella sextelata is cultivated in outdoor greenhouses, ITS and 16S rDNA high throughput amplicon sequencing and microbiome analyses were performed. Pedobacter, Pseudomonas, Stenotrophomonas, and Flavobacterium were found to comprise the core microbiome of M. sextelata ascocarps. These bacterial taxa were also abundant in the soil beneath growing fruiting bodies. A total of 29 bacterial taxa were found to be statistically associated to Morchella fruiting bodies. Bacterial community network analysis revealed high modularity with some 16S rDNA operational taxonomic unit clusters living in specialized fungal niches (e.g., pileus, stipe). Other fungi dominating the soil mycobiome beneath morels included Morchella, Phialophora, and Mortierella. This research informs understanding of microbial indicators and potential facilitators of Morchella ecology and fruiting body production.

One of the most crucial steps in high-throughput sequence-based microbiome studies is the taxonomic assignment of sequences belonging to operational taxonomic units (OTUs). Without taxonomic classification, functional and biological information of microbial communities cannot be inferred or interpreted. The internal transcribed spacer (ITS) region of the ribosomal DNA is the conventional marker region for fungal community studies. While bioinformatics pipelines that cluster reads into OTUs have received much attention in the literature, less attention has been given to the taxonomic classification of these sequences, upon which biological inference is dependent. Here we compare how three common fungal OTU taxonomic assignment tools (RDP Classifier, UTAX, and SINTAX) handle ITS fungal sequence data. The classification power, defined as the proportion of assigned OTUs at a given taxonomic rank, varied among the classifiers. Classifiers were generally consistent (assignment of the same taxonomy to a given OTU) across datasets and ranks; a small number of OTUs were assigned unique classifications across programs. We developed CONSTAX (CONSensus TAXonomy), a Python tool that compares taxonomic classifications of the three programs and merges them into an improved consensus taxonomy. This tool also produces summary classification outputs that are useful for downstream analyses. Our results demonstrate that independent taxonomy assignment tools classify unique members of the fungal community, and greater classification power is realized by generating consensus taxonomy of available classifiers with CONSTAX.

Bacterial interactions with animals and plants have been examined for over a century; by contrast, the study of bacterial–fungal interactions has received less attention. Bacteria interact with fungi in diverse ways, and endobacteria that reside inside fungal cells represent the most intimate interaction. The most significant bacterial endosymbionts that have been studied are associated with Mucoromycota and include two main groups: Burkholderia-related and Mycoplasma-related endobacteria (MRE). Examples of Burkholderia-related endobacteria have been reported in the three Mucoromycota subphyla. By contrast, MRE have only been identified in Glomeromycotina and Mucoromycotina. This study aims to understand whether MRE dwell in Mortierellomycotina and, if so, to determine their impact on the fungal host. We carried out a large-scale screening of 394 Mortierellomycotina strains and employed a combination of microscopy, molecular phylogeny, next-generation sequencing and qPCR. We detected MRE in 12 strains. These endosymbionts represent novel bacterial phylotypes and show evidence of recombination. Their presence in Mortierellomycotina demonstrates that MRE occur within fungi across Mucoromycota and they may have lived in their common ancestor. We cured the fungus of its endosymbionts with antibiotics and observed improved biomass production in isogenic lines lacking MRE, demonstrating that these endobacteria impose some fitness costs to their fungal host. Here we provided the first functional insights into the lifestyle of MRE. Our findings indicate that MRE may be antagonistic to their fungal hosts, and adapted to a non-lethal parasitic lifestyle in the mycelium of Mucoromycota. However, context-dependent adaptive benefits to their host at minimal cost cannot not be excluded. Finally, we conclude that Mortierellomycotina represent attractive model organisms for exploring interactions between MRE and fungi.