Linda M. Swartz, Steven J. Brunsfeld
American Fern Journal 92 (4), 249-269, (1 October 2002) https://doi.org/10.1640/0002-8444(2002)092[0249:TMAGDO]2.0.CO;2
Two species of Botrychium subgenus Botrychium (moonworts, Ophioglossaceae), Botrychium minganense Victorin and B. crenulatum W. H. Wagner, can sometimes be confused in the field, even by experts, because of their reduced morphology. Botrychium minganense can imitate B. crenulatum, which is more rare. They are afforded different degrees of protection on Federal lands, making the distinctness of these species a question of management, conservation, and systematic interest. The purpose of this study was to compare a morphometric analysis of these two species with an analysis of DNA markers from the same individuals, and to assess their distinctness under each method. Collections were made in Washington, Oregon, Idaho, and Montana from seven populations of B. crenulatum and 18 populations of B. minganense. Each plant was measured, emphasizing characters cited by authors in the original species descriptions. Canonical variate analysis performed on SAS separated the samples into two species groups with 32% overlap. RAPD genetic markers revealed more genetic variation than has previously been documented in moonworts. UPGMA cluster analysis of the similarity of RAPD profiles showed well-defined B. minganense and B. crenulatum clusters, but no distinct clusters within B. minganense that could be correlated with its morphological variability. Small samples of the moonwort species B. lunaria and B. simplex included for comparison also formed distinct clusters. Botrychium crenulatum had seven unique RAPD bands, and identification of B. crenulatum could be confirmed or ruled out with markers from one or two RAPD primers. Both B. crenulatum and B. lunaria have been suggested as possible diploid parents of tetraploid B. minganense. All RAPD markers absent in B. crenulatum but present in B. minganense were also present or polymorphic in B. lunaria, supporting B. lunaria as a possible parent. One very small population of B. minganense showed a monomorphic RAPD profile, consistent with inbreeding, but all other populations had multiple genotypes. Some plants of B. minganense clustered most closely with plants from populations up to 400 km away, suggesting that variation may be introduced into populations by occasional colonization by spores from distant sources.