Contrasting levels of polymorphism in cross-amplified microsatellites in two endangered xerothermophilous, obligatorily myrmecophilous, butterflies of the genus Phengaris (Maculinea) (Lepidoptera: Lycaenidae)

Abstract

Abstract.We analysed the polymorphism of cross-amplified microsatellite loci in two endangered butterflies of the genus Phengaris, which inhabit warm grasslands. Specimens of P. arion and P. 'rebeli' collected in Poland showed contrasting levels of variability in the investigated loci. All six tested microsatellites were highly variable in P. arion, whereas in P. 'rebeli' one locus was monomorphic and the other four loci showed a rather low level of polymorphism. Genetic differentiation was small or moderate among P. arion populations and high among P. 'rebeli' populations. We hypothesise that these differences reflect: (1) the relatively high genetic variability of P. arion compared with other members of the genus, indicated by previous studies on allozymes; (2) the small, in the majority of cases, genetic differentiation among populations of P. arion, which suggests recent gene flow; (3) a decrease in the genetic variability and increased isolation of peripheral populations of P. 'rebeli'. In both species, the occurrence of "null" alleles was suggested for some loci, which should be taken into consideration in future studies of the population structure. Additional analysis performed on the corrected frequency of alleles indicated that "null" alleles constitute only a minor problem when estimating genetic differentiation, using FST and AMOVA, however they strongly influence estimates of heterozygosity and FIS, and inferences about the number of genetic groups and assignment of individuals, especially in the case of ' P. rebeli'.

What keeps ‘living dead’ alive: demography of a small and isolated population of Maculinea (= Phengaris) alcon

“…Small pieces of wings were torn off using tweezers and placed into containers with 95% ethanol. DNA was extracted using a Genomic Mini Kit (A&A Biotechnology) as described in Rutkowski et al (2009). DNA polymorphism was assessed using microsatellite loci.…”

“…DNA polymorphism was assessed using microsatellite loci. We PCR-amplified five microsatellite loci, characterized by Zeisset et al (2005) as Macu5, Macu8, Macu11, Macu15 and Malc169, using primers as described by the authors and reagents and conditions as described by Rutkowski et al (2009). The length of the amplified fragments was estimated using a CEQ8000 Beckman Coulter automated sequencer (Beckman Coulter, Inc., Brea, CA, USA; agency in Poland: Comesa, Warsaw), and subsequently the chromatograms were analysed using Beckman Coulter Fragment Analysis Software in order to obtain into the individual genotypes.…”

Population isolation rather than ecological variation explains the genetic structure of endangered myrmecophilous butterfly Phengaris (=Maculinea) arion

“…Laboratory procedures DNA was extracted from approximate 0.5 cm long fragments of thorax or from the whole leg, as described in Rutkowski et al (2009). Using the PCR reaction we amplified six microsatellite loci, designated by Zeisset et al (2005) as Macu8, Macu9, Macu11, Macu15, Macu16 and Macu17 using primers described by the authors.…”

“…P. alcon and P. nausithous (Zeisset et al 2005), and used in the investigation of the molecular ecology of these species (Anton et al 2007;Nash et al 2008) may also be successfully applied in studies of P. arion. Preliminary data on the genetic structure of the butterfly in Poland also indicated moderate genetic differentiation among populations, but simultaneously suggested that some pairs of populations could be highly differentiated, despite spatial (geographical) closeness (Rutkowski et al 2009).…”

“…The low number of populations analysed made it impossible to characterize a wider pattern or the factors affecting genetic differentiation within the species. Recently Rutkowski et al (2009) showed that microsatellite nuclear markers originally developed for congenera, i.e. P. alcon and P. nausithous (Zeisset et al 2005), and used in the investigation of the molecular ecology of these species (Anton et al 2007;Nash et al 2008) may also be successfully applied in studies of P. arion.…”