This review updates the basic biology of moonwort ferns, genus Botrychium, (Ophioglossaceae). It is intended to provide background to readers of this issue and to alleviate the need for repeating this information by subsequent authors. At present, 38 species of Botrychium have been described, with occurrences worldwide in a diverse range of habitats, most commonly in meadows dominated by herbaceous perennial flowering plants that are the alternative host of the mycorrhizal fungi that support belowground growth of Botrychium. The genus produces small aboveground leaves that produce sporangia. Leaves of individual plants may remain dormant belowground in some years. Individual plants may live for decades. Spores germinate belowground and, following colonization by a mycorrhizal fungus, produce belowground bisexual gametophytes that have a high probability of intragametophytic selfing, commonly resulting in populations with low allelic variability. Because of their small size and reduced morphological complexity, Botrychium species have often been referred to as cryptic species. Though difficult to identify, genetic analysis has shown species distinction to be equal to that of non-cryptic species, and has allowed recognition of morphological characteristic of each taxon.

We trace the development of our understanding regarding the mating systems of moonworts (i.e., Botrychium sensu stricto), from the early recognition of inbreeding as the primary mode of sexual reproduction to the discovery of predominantly outcrossing populations in high mountain habitats. We propose an evolutionary trajectory in which both mating systems, in concert with mycorrhizal support, have been able to sustain Botrychium populations through changing environments. We present a depiction of the fern life cycle that portrays the positive role of inbreeding in speciation and evolution of Botrychium and its relevance to ferns and lycophytes in general.

Botrychium farrarii is described as a new species from the Bighorn Mountains of Wyoming, U.S.A. Based on allozyme patterns and chloroplast and nuclear sequence data, it is inferred to be a diploid and the maternal (i.e., chloroplast donor) parent of three allotetraploids, B. furculatum, B. minganense, and B. paradoxum, with which it shares similar morphological traits. Botrychium farrarii is closely related to B. campestre sensu lato, from which it differs primarily by having longer trophophore and sporophore stalks. A key to B. farrarii and morphologically-similar species is presented. At this time, B. farrarii is documented by few individuals from four small, montane forest openings and roadsides in close proximity within a single watershed. Its full geographic range and habitat preferences remain unknown, and we hope this paper will spur discovery of additional populations. Until the species is known to be more abundant, it would be prudent to manage each site to maintain population viability, including avoidance of actions that could adversely impact plant persistence or habitat suitability.

Genetic analyses of the lanceolatum clade in the genus Botrychium supports description of a new diploid species, Botrychium rubellum. It differs genetically, morphologically, and geographically from B. lanceolatum, from which it is segregated, and B. angustisegmentum. Botrychium rubellum is differentiated by the red coloration of the common stalk, upswept basal pinnae, basiscopic pinnules much longer than acroscopic pinnules, spores larger than the spores of the other two species, and in genetic identities reflecting the percent of shared alleles. Botrychium rubellum is currently known to occur in western North America from the southern Rocky Mountains to Alaska, the Yukon Territory, and disjunctively to Greenland.

Recent molecular analyses demonstrate that Botrychium lunaria does not occur in North America except in Greenland, even in a restricted sense, following the description of B. neolunaria. The appropriate name for the remaining continental North American element of the Botrychium lunaria complex is Botrychium onondagense. Because B. neolunaria and B. onondagense are cryptic with respect to each other, we provide a morphological analysis comparing B. onondagense to B. neolunaria, a key, an illustration, and a map based on molecularly identified specimens.

Mitigation translocations of Botrychium (moonworts) were conducted in Colorado and Minnesota due to road and pipeline construction, respectively. Populations in Colorado were monitored for 20 years and for five years in Minnesota following translocation. Botrychium is mycoheterotrophic, so moving the soil containing mycorrhizae surrounding and attached to the plants is key to successful relocation. Two different methods of translocation were used to move plants and soil. Colorado populations declined 99% over 20 years. Minnesota populations declined 64% over five years. The success of these translocations appears to be poor as evidenced by the declining return of aboveground plants. It is possible that populations could return through the spore bank assuming that site conditions remain consistent. Results suggest that translocation is not a viable conservation tool within the typical management planning horizons of natural resource agencies.

Prairie moonwort, Botrychium campestre, was discovered in 1990 in the Bonny Prairie Natural Area of eastern Colorado, the species' southernmost occurrence in the Great Plains. In the years following discovery, the population was monitored by recording the number of aboveground plants. From the initial discovery of three plants in 1990, numbers remained low through 1996, and none were observed from 1997 through 2009. An analysis of soil samples collected in 2007 from the site confirmed belowground viable gemmae (vegetative sporophytic propagules produced by the sporophyte stem). Following two years of above-average precipitation in 2008 and 2009, plus artificial watering in those years, 12 aboveground plants were found in 2010. In 2011, another wet year, 63 aboveground plants were recorded. In the following two drought years, the number returned to zero and has remained low in subsequent years. We address the belowground biology of Botrychium and its mycorrhizal symbiont that make possible the persistence of populations in the absence of aboveground plants, as well as correlation with precipitation and implications for detection and conservation of Botrychium populations.

I review the history of my work on moonwort ferns, genus Botrychium, beginning with my early discovery of biology as a profession and the importance of mentors and networks of professional researchers in the development of my interests in biology. I discuss my initial work on discovery of new species of Botrychium and the utilization of enzyme electrophoresis as an early methodology for establishing species' individual genetic characterization and their relationships among species, emphasizing the importance of codominant nuclear markers for detection of allopolyploid parentages. I review the unusual predominance of allopolyploidy in Botrychium and discuss possible reasons for this relative to breeding systems, and describe a new pathway for overcoming hybrid sterility without polyploidy. I discuss the inadequacy of wind as a universal dispersal mechanism in Botrychium, and suggest avian dispersal as an under-appreciated spore dispersal mechanism. Throughout I note unresolved issues of Botrychium biology and suggest areas of needed research. Lastly, I discuss the unique characteristics of Botrychium that have allowed this lineage to persist through universal ecological disruptions and massive extinctions and compete successfully in modern ecosystems, and the morphological similarities of its descendants to early land plants.