Abstract
Arbuscular mycorrhizal fungi (AMF) are obligate symbionts of c. 80% of land plants, having enormous ecological and economic impact, as they often improve crop plant nutrition and yield. DNA-based identification with molecular markers is used to analyze AM fungal communities in the field, but reaching species level taxonomic resolution remains challenging. Thus, currently there is no consensus on how to analyze high-throughput sequences and assign them into species. Here, a new sequencing strategy combined with taxonomic affiliations implemented with an evolutionary placement algorithm (EPA) was established. It is based on sequencing a c. 450 bp region of the large subunit (LSU) ribosomal rRNA gene with the MiSeq-Illumina platform. The method is suitable for the discrimination of closely related AMF species and was used to study host-AMF preferences in roots of Pequin pepper, soybean and orange at one location in the arid northeast of Mexico. Twenty AM fungal species from 13 genera were detected. Phylogenetic affiliation of reads to species revealed crop preferential associations. In Pequin pepper roots, several Rhizophagus species represented most of the community, Rhizophagus clarus being the most abundant. The soybean AM fungal community was dominated by Rhizophagus irregularis and Funneliformis mosseae and that of orange by several species of Dominikia, some of them only found in this crop. Unraveling the AMF-plant preferences of important crops by an affordable and robust sequencing method, combined with phylotaxonomic AMF species resolution, is an important tool to obtain taxonomic units that are meaningful in both biological and ecological studies.
Highlights
Arbuscular mycorrhizal fungi (AMF, abbreviation used for ‘arbuscular mycorrhizal fungal’) have received great attention because they form symbiotic relations with the most relevantElectronic supplementary material The online version of this article contains supplementary material, which is available to authorized users.Historically, AMF communities have been characterized by spore morphology studies but this method may be misleading as presence of spores, which are resting stages, does not necessarily represent the active community (Renker et al 2005; Hempel et al 2007) and, e.g., spatiotemporal variations may be mistaken by sporulation dynamics (Pringle and Bever, 2002)
We focused on host-AMF species preferences and the effect of the developmental stage of the crops on the AMF community composition
Using the artificially constructed R. intraradices and R. irregularis dataset, the evolutionary placement algorithm (EPA) approach resulted in the correct affiliation of all 21 sequence variants with a maximum placement likelihood value
Summary
AMF communities have been characterized by spore morphology studies but this method may be misleading as presence of spores, which are resting stages, does not necessarily represent the active community (Renker et al 2005; Hempel et al 2007) and, e.g., spatiotemporal variations may be mistaken by sporulation dynamics (Pringle and Bever, 2002). Species characterization and for molecular ecological field studies (Senés-Guerrero et al 2014). This 1.5 kb barcode comprises a part of the SSU rRNA gene, the complete ITS region (ITS1, 5.8S, ITS2 rRNA gene) and c. High-throughput sequencing of the 1.5 kb AMF barcode fragment has only been attempted with Pacific BioSciences (PacBio) Single-Molecule Real-Time (SMRT) sequencing (Schlaeppi et al 2016). SMRT sequencing is not the most commonly used highthroughput sequencing technology due to its high error rate and limited throughput (Ardui et al 2018)
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