A simple model system for identifying arbuscular mycorrhizal fungal taxa that actively colonize rice (Oryza sativa L.) roots grown in field soil

Abstract

ABSTRACTEven a few centimeters of roots in field soils can be colonized by genetically diverse arbuscular mycorrhizal (AM) fungi. The DNA sequences of AM fungi in roots suggest the fungal identities; however, it is difficult to determine which AM fungal taxa are physiologically active. In this study, we took advantage of the characteristics of rice (Oryza sativa L.) mycorrhizal roots, in which active colonization in roots is easily detected via histochemical staining of fungal succinate dehydrogenase activity (vital staining) and individual active colonization regions (infection units) in roots rarely coalesce. Root segments (< 3 mm) containing an active infection unit were dissected and squashed, large subunit (LSU) ribosomal RNA genes (rDNAs) were amplified using fungal universal primers and the sequences were directly determined by Sanger sequencing. All obtained sequences of colonization regions were of glomeromycotan origin. Phylogenetic analysis revealed that the levels of LSU-rDNA heterogeneity within an active colonization site were different among different clades. The methodology presented in this study offers researchers a novel tool for investigating the DNA information of physiologically active AM fungi in roots, whereas the factors that affect genetic diversity among active colonization remain to be clarified.