Arbuscular mycorrhizal fungi (AMF) are a group of soil microorganisms that establish symbioses with the vast majority of land plants. To date, generation of AMF coding information has been limited to model genera that grow well axenically; Rhizoglomus and Gigaspora. Meanwhile, data on the functional gene repertoire of most AMF families is non-existent. Here, we provide primary large-scale transcriptome data from eight poorly studied AMF species (Acaulospora morrowiae, Diversispora versiforme, Scutellospora calospora, Racocetra castanea, Paraglomus brasilianum, Ambispora leptoticha, Claroideoglomus claroideum and Funneliformis mosseae) using ultra-low input ribonucleic acid (RNA)-seq approaches. Our analyses reveals that quiescent spores of many AMF species harbour a diverse functional diversity and solidify known evolutionary relationships within the group. Our findings demonstrate that RNA-seq data obtained from low-input RNA are reliable in comparison to conventional RNA-seq experiments. Thus, our methodology can potentially be used to deepen our understanding of fungal microbial function and phylogeny using minute amounts of RNA material.

Ribosomal nuclear genes are routinely utilized in the molecular identification of fungi. The variation in the multiple copies of these genes within each Glomeromycota strain and species reduces their usefulness for molecular characterization of arbuscular mycorrhizal fungi. To explore the potential of molecular tools for the identification of Glomus species, a multi-gene analysis approach was undertaken. Three protein-encoding genes were tested, namely elogation factor 1-α (765 bp), V-H+-ATPase VHA5 (1468 bp), and F0F1-ATPase β-subunit (621 bp). The latter is newly reported for the Glomeromycetes. Eleven species, including the type-specimen of Glomus irregulare Blaszk., Wubet, Renker & Buscot, a reference strain of G. intraradices N.C. Schenck & G.S. Sm. (DAOM 225240), and five strains of Glomus sp. formerly identified as G. intraradices, were analysed. These genes did not show polymorphisms within strains, and results indicated a close relationship between molecular identification and morphological characterization. Species with closely related spore morphological features, G. aggregatum N.C. Schenck & G.S. Sm., G. diaphanum Morton & Walker, G. irregulare , and Glomus sp. DAOM 197198, showed more than 99% nucleotide similarity, while the morphologically distinct species, G. cerebriforme McGee, G. clarum T.H. Nicolson & N.C. Schenck, G. claroideum N.C. Schenck & G.S. Sm., G. custos C. Cano & Dalpé, G. mosseae (T.H. Nicolson & Gerd.) Gerd. & Trappe, and G. proliferum Dalpé & Declerck, showed less than 97% similarity for at least one gene. A 100% molecular similarity for all three genes was found between G. irregulare and Glomus sp. DAOM 197198, confirming the new identity of the model arbuscular mycorrhizal fungus. Similarity thresholds for identification by DNA sequencing are discussed.

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The use of microwave irradiation as a source of energy to clear and stain intra-radical arbuscular mycorrhizal fungi propagules has been tested on a variety of indigenous and cultivated herbaceous plants. The aim of the study was to evaluate the efficiency of microwave irradiation on root softening, fungi tissue staining, and preservation of DNA integrity for subsequent molecular analyses. The proposed methodology has been adapted from the standard procedures used to detect and quantify mycorrhizal root colonization levels. Using a domestic microwave oven, tissue clearing and staining required together between 30 s and 1.5 min of microwave treatment to be completed, depending the diameter size of the roots. The well-performing chemical stains tested were acid fuchsin, trypan blue, and aniline blue. The acid fuchsin clearing and staining processes, as performed, were also demonstrated to preserve DNA integrity for further molecular analyses. Irradiation by microwaves has been used with success in our laboratory within the frame of several studies. It offers considerable time saving over traditional method, reducing processing times from several hours to a few minutes while decreasing considerably the amount of chemicals and energy required to perform analyses.