title: Defining the pathways of symbiotic Epichloë colonization in grass embryos with confocal microscopy.
abstract: Asexual cool-season grass endophytes of the genus Epichloë (Ascomycota: Clavicipitaceae) are strictly vertically disseminated. The hosts of these mutualistic fungi express no symptoms during the fungal lifecycle that takes place entirely within the plant, while their hosts receive beneficial outcomes. These fungi are distributed in two major locations within the mature seeds of their hosts; namely, within the embryo (including the scutellum, coleoptile, plumule, radicle, and coleorhiza tissues) and between the aleurone and pericarp layers, with the latter hyphae playing no role in transmission of the fungus to the next plant generation. Conflicting evidence remains in the literature on the timing of embryo colonization. In a detailed investigation, utilizing confocal microscopy to observe the distribution of Epichloë coenophiala strain AR601 in tall fescue (Lolium arundinaceum), we tracked endophyte hyphal colonization in the ovary (pre-fertilization) through to the fully mature seed stage. Confocal microscopy images revealed that at the early and mature developmental stages of the embryo sac, before host grass fertilization, there were large quantities of endophyte mycelium present, especially around the antipodal cells, indicating that this endophyte enters the embryo sac before the fertilization stage. After host fertilization, fungal hyphae could be seen in the true embryo and early nonstarchy endosperm. Understanding the mechanisms of transmission to the seed is important for commercial seed producers and end users.
authors.affiliation: a Institute of Agriculture and Environment , Massey University , Private Bag 11222, Palmerston North , New Zealand , 4410.|b Forage Improvement, AgResearch Ltd. , Grasslands Research Centre , Private Bag 11008, Palmerston North , New Zealand , 4410.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: b Forage Improvement, AgResearch Ltd. , Grasslands Research Centre , Private Bag 11008, Palmerston North , New Zealand , 4410.
authors.forename: Stuart D
authors.initials: SD
authors.lastname: Card
authors.affiliation: b Forage Improvement, AgResearch Ltd. , Grasslands Research Centre , Private Bag 11008, Palmerston North , New Zealand , 4410.
authors.forename: Wade J
authors.initials: WJ
authors.lastname: Mace
authors.affiliation: b Forage Improvement, AgResearch Ltd. , Grasslands Research Centre , Private Bag 11008, Palmerston North , New Zealand , 4410.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Christensen
authors.affiliation: a Institute of Agriculture and Environment , Massey University , Private Bag 11222, Palmerston North , New Zealand , 4410.
authors.forename: Craig R
authors.initials: CR
authors.lastname: McGill
authors.affiliation: a Institute of Agriculture and Environment , Massey University , Private Bag 11222, Palmerston North , New Zealand , 4410.
authors.forename: Cory
authors.initials: C
authors.lastname: Matthew
mesh_terms: D060026:Endophytes / Q000166:cytology / Q000254:growth & development*; D055329:Epichloe / Q000166:cytology / Q000254:growth & development*; D031751:Festuca / Q000382:microbiology*; D018613:Microscopy, Confocal; D025282:Mycelium / Q000166:cytology / Q000254:growth & development; D012639:Seeds / Q000382:microbiology; D013559:Symbiosis
keywords: Epichloë coenophiala; Lolium arundinaceum; confocal microscopy; endophyte; fluorescent dye