Anastomosis behavior differs between asymbiotic and symbiotic hyphae of Rhizophagus clarus

The life history of arbuscular mycorrhizal fungi (AMF, Glomeromycota) consists of a short asymbiotic phase when spores germinate and a longer symbiotic phase where hyphae form a network within roots and subsequently in the rhizosphere. Hyphal anastomosis contributes to colony formation, yet this process has been studied mostly in the asymbiotic phase rather than in mycorrhizal plants because of methodological limitations. We sought to compare patterns of anastomosis during each phase of fungal growth by measuring hyphal fusions in genetically identical and different single spore isolates of Rhizophagus clarus from different environments and geographic locations. These isolates were genotyped with two anonymous markers of microsatellite-flanking regions. Anastomosis of hyphae from germinating spores was examined in axenic Petri dishes. A rhizohyphatron consisting of agar-coated glass slides bridging single or paired mycorrhizal sorghum plants allowed evaluation of anastomosis of symbiotic hyphae. Anastomosis of hyphae within a colony, defined here as a mycelium from an individual germinating spore or from mycorrhizal roots of one plant, occurred with similar frequencies (8-38%). However, anastomosis between paired colonies was observed in germinating spores from either genetically identical or different isolates, but it was never detected in symbiotic hyphae. The frequency of anastomosis in asymbiotic hyphae from paired interactions was low, occurring in fewer than 6% of hyphal contacts. These data suggest that anastomosis is relatively unconstrained when interactions occur within a colony but is confined to asymbiotic hyphae when interactions occur between paired colonies. This pattern of behavior suggests that asymbiotic and symbiotic phases of mycelium development by R. clarus may differ in function. Anastomosis in the asymbiotic phase may provide brief opportunities for gene flow between populations of this and possibly other AMF species.