Methodologies for soil extraction and conservation analysis of ferns and lycophytes with belowground gametophytes

Studies of fern and lycophyte gametophyte biology in natural settings can be challenging, but such analyses are critical to understanding the dispersal, ecology, and conservation of these species. It is especially challenging to study species whose gametophytes and early sporophyte stages develop belowground, as is the case for species of the Ophioglossaceae, Psilotaceae, some species of the Schizeaceae (Actinostachys), and some species of the Lycopodiaceae. To study these taxa, gametophytes and young sporophytes must be extracted from the soil.

In 1989, Mason and Farrar described a methodology for accomplishing the collection of belowground gametophytes and sporophytes using soil centrifugation. Here, we refine this procedure based on subsequent years of experience.

We found that many more sporophytes exist belowground than are represented by aboveground leaves, and that belowground sporophytes can survive indefinitely without production of aboveground leaves. Belowground gametophytes are common in areas where spore-releasing sporophyte leaves are present. Gametophytes are bisexual with male and female gametangia intermixed. Some species of Botrychium also reproduce asexually belowground through production of gemmae.

We conclude that in Botrychium, assessments of population health and structure must include analyses of the belowground plants and their habitat. Conservation management strategies must also include potential changes in the belowground habitat.