Abstract. Currently it remains difficult to obtain robust microsatellite markers for Lepidoptera. In an attempt to overcome the problems associated with developing microsatellite markers for this insect order we combined (i) biotin-enrichment protocol, (ii) next generation pyrosequencing (through 454 GS-FLX Titanium technology) and (iii) the use of individuals collected from eight geographically distant European populations representing three subspecies of Euphydryas aurinia. Out of 96 stringently designed primer pairs, 12 polymorphic microsatellite loci amplified without obvious evidence of null alleles in eight individuals from different subspecies. Between five and seven of these loci showed full within population applicability and three revealed to be robust and transferable between populations and sub-species, providing a first step towards the development of a valuable and robust tool for studying conservation issues and evolution in E. aurinia populations. Nevertheless, as in most studies dealing with Lepidoptera microsatellites, null alleles were detected in most of the developed markers. Our results emphasize the need for further research in order to better understand the complex evolution and organization of Lepidopteran genomes. 261* These authors contributed equally to this work.

High mountain ecosystems are extreme habitats for all organisms and therefore demand specific adaptations. In this context, we studied the ecology of the butterfly Euphydryas aurinia debilis in the High Tauern (Austria) and compared the obtained data against the ecology of the species in lower elevation habitats. We performed mark-release-recapture studies over the entire flight periods (end of June to end of July) in 2007 and 2008 to analyse the fundamental ecological parameters of a population. The demography of males and females was similar in both years, and no indication of typical protandry was detected. We observed a generally low dispersal of the individuals in both years, but males dispersed significantly more than females in 2008; this finding of low vagility was supported by allozyme analyses. Furthermore, butterflies survived periods of several days of continuously closed snow cover without any indication of increased mortality rates. In these three traits, this alpine population of E. aurinia apparently has ecological and physiological adaptations to the extreme requirements of high-altitude habitats and strongly deviates from the lower elevation populations.

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Ice ages within Europe forced many species to retreat to refugia, of which three major biogeographic basic types can be distinguished: "Mediterranean", "Continental" and "Alpine / Arctic" species. However, this classification often fails to explain the complex phylogeography of European species with a wide range of latitudinal and altitudinal distribution. Hence, we tested for the possibility that all three mentioned faunal elements are represented within one species. Our data was obtained by scoring 1,307 Euphydryas aurinia individuals (46 European locations) for 17 allozyme loci, and sequencing a subset of 492 individuals (21 sites) for a 626 base pairs COI fragment. Genetic diversity indices, F statistics, hierarchical analyses of molecular variance, individual-based clustering, and networks were used to explore the phylogeographic patterns. The COI fragment represented 18 haplotypes showing a strong geographic structure. All but one allozyme loci analysed were polymorphic with a mean F ST of 0.20, supporting a pronounced among population structure. Interpretation of both genetic marker systems, using several analytical tools, calls for the recognition of twelve genetic groups. These analyses consistently distinguished different groups in Iberia (2), Italy, Provence, Alps (3), Slovenia, Carpathian Basin, the lowlands of West and Central Europe as well as Estonia, often with considerable additional substructures. The genetic data strongly support the hypothesis that E. aurinia survived the last glaciation in Mediterranean, extra-Mediterranean and perialpine refugia. It is thus a rare example of a model organism that combines attributes of faunal elements from all three of these sources. The observed differences between allozymes and mtDNA most likely result from recent introgression of mtDNA into nuclear allozyme groups. Our results indicate discrepancies with the morphologically-based subspecies models, underlining the need to revise the current taxonomy.

In human-altered landscapes, species with specific habitat requirements tend to persist as metapopulations, forming colonies restricted to patches of suitable habitats, displaying mutually independent within-patch dynamics and interconnected by inter-colony movements of individuals. Despite intuitive appeal and both empirical and analytical evidence, metapopulations of only relatively few butterfly systems had been both monitored for multiple years to quantify metapopulation dynamics, and assayed from the point of view of population genetics. We used allozyme analysis to study the genetic make-up of a metapopulation of a declining and EU-protected butterfly, Euphydryas aurinia, inhabiting humid grasslands in western Czech Republic, and reanalysed previously published demography and dispersal data to interpret the patterns. For 497 colony x year visits to the 97 colonies known at that time, we found annual extinction and colonisation probabilities roughly equal to 4%. The genetic diversity within colonies was intermediate or high for all assessed parameters of population genetic diversity and hence higher than expected for such a habitat specialist species. All the standard genetic diversity measures were positively correlated to adult counts and colony areas, but the correlations were weak and rarely significant, probably due to the rapid within-colony population dynamics. Only very weak correlations applied to larval nests numbers. We conclude that the entirety of colonies forms a well-connected system for their majority. Especially in its core parts, we assume a metapopulation structure with a dynamic equilibrium between local extinction and recolonization. It is vital to conserve in particular these structures of large and interconnected colonies.Implications for insect conservation: Conservation measures should focus on considering more in depth the habitat requirements of E. aurinia for management plans and on stabilisation strategies for colonies, especially of peripheral ones, e.g. by habitat restoration.