Microsatellites reveal substantial among-population genetic differentiation and strong inbreeding in the relict fern Dryopteris aemula

A previous study detected no allozyme diversity in Iberian populations of the buckler-fern Dryopteris aemula. The use of a more sensitive marker, such as microsatellites, was thus needed to reveal the genetic diversity, breeding system and spatial genetic structure of this species in natural populations. Eight microsatellite loci for D. aemula were developed and their cross-amplification with other ferns was tested. Five polymorphic loci were used to characterize the amount and distribution of genetic diversity of D. aemula in three populations from the Iberian Peninsula and one population from the Azores. Most microsatellite markers developed were transferable to taxa close to D. aemula. Overall genetic variation was low (H-T = 0 center dot 447), but was higher in the Azorean population than in the Iberian populations of this species. Among-population genetic differentiation was high (F-ST = 0 center dot 520). All loci strongly departed from Hardy-Weinberg equilibrium. In the population where genetic structure was studied, no spatial autocorrelation was found in any distance class. The higher genetic diversity observed in the Azorean population studied suggested a possible refugium in this region from which mainland Europe has been recolonized after the Pleistocene glaciations. High among-population genetic differentiation indicated restricted gene flow (i.e. lack of spore exchange) across the highly fragmented area occupied by D. aemula. The deviations from Hardy-Weinberg equilibrium reflected strong inbreeding in D. aemula, a trait rarely observed in homosporous ferns. The absence of spatial genetic structure indicated effective spore dispersal over short distances. Additionally, the cross-amplification of some D. aemula microsatellites makes them suitable for use in other Dryopteris taxa.