The genetic population structure of Buthus occitanus (Scorpiones: Buthidae) across the Strait of Gibraltar: calibrating a molecular clock using nuclear allozyme variation

I assess here the importance of the Strait of Gibraltar as a barrier to gene flow for populations of the scorpion Buthus occitanus . This polytypic buthid scorpion occurs in Europe and in North Africa where it is morphologically more diverse. The phylogenetic relationship between B. occitanus populations across the Strait of Gibraltar is investigated by nuclear allozymes analysis (15 loci scored). Phylogenetic analysis based on estimated gene frequency data results in a tree topology that divides the populations into three clades, i.e. a European, an Atlas (= Morocco samples) and a Tell-Atlas clade (= Tunisian samples). The Tell-Atlas clade grouped with the European clade with a rather high bootstrap support of 70%. Within these clades low levels of genetic variability are observed. Calibrating a molecular clock under the assumption that the European populations are autochthonous and have been isolated from the North African for at least 5.33 Myr reveals a divergence rate of 0.060 genetic distance (D) per Myr estimated between European and Moroccan samples and 0.036D Myr⁻¹ between European and Tunisian samples, respectively.  © 2004 The Linnean Society of London, Biological Journal of the Linnean Society , 2004, 81 , 519–534.     

Genetic structure of the endangered perennial plant Eryngium alpinum (Apiaceae) in an alpine valley

We investigated the genetic structure of Eryngium alpinum (Apiaceae) in an Alpine valley where the plant occurs in patches of various sizes. In a conservation perspective, our goal was to determine whether the valley consists of one or several genetic units. Habitat fragmentation and previous observations of restricted pollen/seed dispersal suggested pronounced genetic structure, but gene dispersal often follows a leptokurtic distribution, which may lead to weak genetic structure. We used nine microsatellite loci and two nested sampling designs (50 × 50 m grid throughout the valley and 2 × 2 m grid in two 50 × 10 m quadrats). Within the overall valley, F -statistics and Bayesian approaches indicated high genetic homogeneity. This result might be explained by: (1) underestimation of long-distance pollen/seed dispersal by in situ experiments and (2) too recent fragmentation events to build up genetic structure. Spatial autocorrelation revealed isolation by distance on the overall valley but this pattern was much more pronounced in the 50 × 10 m quadrats sampled with a 2-m mesh. This was probably associated with limited primary seed dispersal, leading to the spatial clustering of half-sibs around maternal plants. We emphasize the interest of nested sampling designs and of combining several statistical tools.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 93 , 667–677.     

Morphological, mitochondrial DNA and allozyme evolution in representative amphibians and reptiles inhabiting each side of the Strait of Gibraltar

Patterns of differentiation in morphology, mitochondrial DNA and allozymes in amphibians and reptiles inhabiting northern and southern shores of the Strait of Gibraltar are not concordant, suggesting that each taxon was affected differently by events preceding or following the formation of the Strait of Gibraltar. Mitochondrial DNA and allozyme differentiation between Discoglossus jeanneae and Discoglossus scovazzi (Anura, Discoglossidae), Rana perezi and Rana saharica (Anura, Ranidae), and Blanus cinereus and Blanus tingitanus (Squamata, Amphisbaenia, Amphisbaenidae) is substantial, whereas morphological differentiation is moderate in Rana and Blanus , but is substantial in Discoglossus . Differentiation in mitochondrial DNA and morphology between Timon ( Lacerta ) lepidus and Timon ( Lacerta ) tangitanus (Squamata, Lacertoidea, Lacertidae) is considerable, but allozyme differentiation is low. In members of type-I and -II Podarcis vaucheri (Squamata, Lacertoidea, Lacertidae), morphology and mitochondrial DNA are moderately differentiated, but allozyme differentiation is low. Spanish and Moroccan populations of Hyla meridionalis (Anura, Hylidae), Mauremys leprosa (Testudines, Geoemydidae), and Macroprotodon brevis (Squamata, Serpentes, Colubridae) demonstrate little allozyme and mitochondrial DNA differentiation, but whereas morphological differentiation between Mauremys and Macroprotodon populations is moderate, Hyla demonstrate substantial morphological differentiation between continental populations. These data suggest that sex-limited mitochondrial markers are reflective of ancient phylogenetic history, whereas biparentally inherited allozyme markers and morphological characteristics reflect more recent population structure and movement.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 94 , 445–461.     

Phylogeography of five Polytrichum species within Europe

Using allozymes and microsatellites we have analysed the genetic structure among European populations for several Polytrichum species to infer relevant factors, such as historical events or gene flow, that have shaped their genetic structure. As we observed low levels of genetic differentiation among populations, and no decreasing levels of genetic variation with increasing latitude within most of the examined species, no genetic evidence was obtained for a step-wise recolonization of Europe from southern refugia after the latest glacial period for P. commune , P. uliginosum , P. formosum and P. piliferum . The near absence of population substructuring within these species does indicate that extensive spore dispersal is the most important factor determining the genetic structure among European Polytrichum populations. Gene flow levels have apparently been sufficient to prevent genetic differentiation among populations caused by genetic drift, and to wipe out any genetic structure caused by the postglacial recolonization process. On the other hand, increased genetic differentiation of alpine P. formosum populations suggests that mountain ranges might restrict gene flow significantly among Polytrichum populations. In contrast to most examined Polytrichum species, P. juniperinum showed high levels of genetic differentiation and a profound genetic structure. Assuming that gene flow is not more restricted in P. juniperinum , these findings suggest that this species has recolonized Europe after the latest glacial period from two different refugia, one possibly being the British Isles.  © 2003 The Linnean Society of London, Biological Journal of the Linnean Society 2003, 78, 203–213.     

Implications of contrasting patterns of genetic variability in two vespertilionid bats from the Indonesian archipelago

Wallacean island populations of two Vespertilioninae bats, Myotis muricola and Scotophilus kuhlii , which have similar geographical ranges, showed marked contrast in the amount and pattern of genetic variation. Within islands, genetic variation was on average much higher in M. muricola but declined from west to east, whereas all populations of S. kuhlii had uniformly low levels of genetic variation by mammalian standards. S. kuhlii showed little genetic differentiation between islands and estimates of gene flow were substantial whereas island populations of M. muricola differed markedly and there was a strong isolation-by-distance effect associated with the extent of the sea crossing between islands. Furthermore, the lower mean heterozygosity and small genetic distances between eastern island populations of M. muricola is evidence that there has been a bottleneck associated with the colonization of this area. The attenuation of genetic diversity to the east is also seen in some other mammalian species and may indicate limits to dispersal and have implications for species management. The patterns of variability in S. kuhlii may be a consequence of its strong dispersal capacity and close association with human activity, which, together with other factors, suggest a panmictic population.  © 2004 The Linnean Society of London, Biological Journal of the Linnean Society , 2004, 83 , 421–431.     

Conservation genetics of Hebe speciosa (Plantaginaceae) an endangered New Zealand shrub

All extant populations of Hebe speciosa (Plantaginaceae), a threatened endemic New Zealand shrub, were investigated using the amplified fragment length polymorphism technique (AFLP). Genetic diversity indices varied significantly among geographical regions and were positively correlated with population size. Among-population genetic differentiation was high (mean pairwise Φ_ST = 0.47), implying complex historical relationships between disjunct populations and negligible contemporary gene flow. Southern populations exhibited extremely low genetic diversity relative to those found in Northland, suggesting that these populations may be more recent in origin. Patterns of genetic relationship among some populations indicate pre-European Māori dispersal and cultivation. The three northernmost populations were found to contain the majority of the species' remaining genetic diversity and thus, should be a focus for future conservation management. Some southern sites may also be culturally significant as evidence of Māori trade and cultivation of Hebe speciosa .  © 2005 The Linnean Society of London, Botanical Journal of the Linnean Society , 2005, 149 , 229–239.     

Genetic differentiation and gene flow of two sparidae subspecies, Diplodus sargus sargus and Diplodus sargus cadenati in Atlantic and south-west Mediterranean populations

A total of nine enzymes coded by 14 loci were assayed for each of six populations (from the north-eastern Atlantic and the Mediterranean) of two sea bream subspecies ( Diplodus sargus sargus and Diplodus sargus cadenati ). Diagnostic alleles were observed for each subspecies, although there were several common alleles. Estimates of variance in allele frequencies among samples ( F _ST) revealed significant differences ( P  < 0.05) among both subspecies. Genetic divergence was found between Atlantic and Mediterranean samples: values for genetic distances were higher than 0.163. Furthermore, D. sargus cadenati populations displayed a higher mean weight and length than D. sargus sargus populations and significant differences in growth were found among subspecies and populations. These results are discussed in terms of levels of gene flow and its respective relationships with water circulation in the Strait of Gibraltar and geological events.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 89 , 705–717.     

Phylogeographic patterns of two tiger beetle species at both sides of the strait of Gibraltar (Coleoptera: Cicindelini)

The Strait of Gibraltar is one of the major barriers to gene flow between land masses of Europe and Africa at the western end of the Mediterranean. Since the opening of the Strait at the end of the Miocene 5.33 million years ago (Mya) it has exerted a strong influence on the dispersal and colonization of whole biotas, particularly with regard to the retreat of organisms during glaciation peaks and the northward colonization events during the warm periods of Pleistocene. The aim of this study is to elucidate the influence of the Strait of Gibraltar in the gene flow among populations of two tiger beetle species collected in Morocco, Cicindela campestris (L. 1758) and Lophyra flexuosa (Fabricius 1787), with regard to both new and published data from populations of southern Iberia. The phylogeographic analysis showed that Moroccan haplotypes of L. flexuosa belonged to a single coastal mitochondrial clade and that populations at both sides of` the Strait of Gibraltar were genetically well connected. The haplotype network of C. campestris showed that Moroccan populations made up a robust cluster clearly differentiated from those of Iberian and other European populations. These differences are thought to reflect the distinct evolutionary history (dispersal capacity, ecological strategies) of both species, as L. flexuosa shows an almost continuous distribution on the coasts located at both sides of the West Mediterranean, whereas C. campestris shows a patchy distribution and prefers montane habitats in the Western Palaearctic.     

Strong genetic cohesiveness between Italy and North Africa in four butterfly species

The sea acts as an effective dispersal barrier for most terrestrial animal species. Narrow sea straits, therefore, often represent areas where species are able to disperse from one land mass to another. In the Mediterranean Sea, the narrowest connecting points between North Africa and Europe are the Strait of Gibraltar and the Strait of Sicily. In the past, climatic oscillations caused changing sea levels and thus influenced the permeability of these sea straits. We analysed the genetic structure of four butterfly species that all occur on both sides of the Strait of Sicily. In all four species, we observed a lack of genetic differentiation between the populations of North Africa and those of Italy. Species distribution models support the strong cohesiveness in that they show a largely continuous glacial distribution over Italy and North Africa. The data obtained reveal that there was a large exchange of individuals between Italy and the eastern Maghreb during the last ice age. This might not only be the case for the species under investigation in the present study, but also might represent a more general pattern for mobile thermophilic western Palearctic species. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 99 , 818–830.     

Deep biogeographical history and cytonuclear discordance in the black-tailed brush lizard (Urosaurus nigricaudus) of Baja California

Molecular tools help us deduce historical events such as vicariance, dispersal, gene flow and speciation. However, our inferences are inevitably linked to the nature of the characters that we use to infer history. For example, the difference in inheritance patterns of mitochondrial DNA (mtDNA) and nuclear DNA (non-recombining maternal vs. recombining biparental inheritance) may lead us to propose different intraspecific histories. The peninsula of Baja California of north-western Mexico, a region affected by a complex geological history involving temporary seaways, permits evaluation of differences between these character types. We sequenced 1966 bp of mtDNA to reconstruct the genealogical history of Urosaurus nigricaudus (black-tailed brush lizard) from samples spanning the entire peninsula. The genealogy revealed several deep divergences, congruent with temporary vicariance events in the mid-peninsular, Loreto and Cape regions, as well as a major split across the Isthmus of La Paz, possibly resulting from a late Miocene seaway. The results support an emerging picture of the historical biogeography of Baja California, which suggests that key vicariance events are older than commonly perceived. The maternal genealogy of U. nigricaudus sharply contrasts with variation in allozymes that suggests very little differentiation across mitochondrial breaks, consistent with a pattern of ongoing gene flow. We interpret this cytonuclear discordance in relation to the historical biogeography of the region.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 94 , 89–104.     

Seed dispersal and population structure in Vriesea gigantea,a bromeliad from the Brazilian Atlantic Rainforest

Seed dispersal, population structure and the mating system of plant species can have great consequences on the genetic structure of populations. Vriesea gigantea is a bromeliad from southeastern Brazil which is self‐compatible and pollinator dependent for fruit set. Its populations are fertile in terms of the production of flowers, pollen, fruits and seeds. To assess the importance of seed supply for gene flow, colonization and distribution of adult individuals, the seed dispersal and population structure of V. gigantea were studied. Seeds are dispersed over short distances; most seeds land close to the mother plant. This pattern coincides with the reported aggregate distribution of bromeliad seedlings. Population structure results showed high seedling recruitment, because 51.3% developed into adults, although few juveniles reached this stage. This result is different from that for other bromeliad species from different habitat conditions. Seed dispersal and population structural patterns are consistent with previous molecular studies, revealing that V. gigantea populations are genetically structured, with low gene flow and a moderate outcrossing rate. © 2010 The Linnean Society of London, Botanical Journal of the Linnean Society, 2010, 164 , 317–325.     

Phylogenetic relationships and biogeographical history of the genus Rhinoclemmys Fitzinger, 1835 and the monophyly of the turtle family Geoemydidae (Testudines: Testudinoidea)

Rhinoclemmys is an interesting genus of turtles biogeographically and ecologically, being the only genus of the family Geoemydidae that occurs in the New World and inhabiting a wide range of habitats from aquatic to highly terrestrial. Here we present a molecular phylogeny of Rhinoclemmys using both mitochondrial and nuclear genes. Our results strongly support the monophyletic and subfamilial status of Rhinoclemmys within the monophyletic family Geoemydidae. Within Rhinoclemmys , two clades are strongly supported, i.e. R. annulata  +  R. pulcherrima and R. areolata  +  R. punctularia  +  R. diademata  +  R. funerea  +  R. melanosterna , but the positions of R. nasuta and R. rubida are still weakly supported. In terms of the biogeographical history, the results of this study, coupled with palaeontological evidence, corroborate the hypothesis that this group migrated from Asia to the Americas across the Bering Strait during the early Eocene. The radiation of Rhinoclemmys in Central and South America corresponds well with vicariance events, including the emergence of the Sierra Madres of Mexico and the Nuclear Highland, and dispersals across the Panama land bridge. Interestingly, our resulting phylogeny suggests this group invaded South America at least four times and that dispersal of R. nasuta to South America probably took place in the early Miocene before the emergence of the Isthmus of Panama. We finally discuss our phylogenetic results with regard to the monophyly of the family Geoemydidae and in the context of previous morphological analyses. © 2008 The Linnean Society of London, Zoological Journal of the Linnean Society , 2008, 153 , 751–767.     

Phylogeography of a host-specific insect: genetic structure of Ips typographus in Europe does not reflect past fragmentation of its host

The phylogeography of the bark beetle Ips typographus was assessed using five microsatellite markers. Twenty-eight populations were sampled throughout Europe on the host tree Picea abies . I. typographus showed very low levels of genetic diversity, and the study revealed a lack of genetic structure across Europe. No significant barrier to gene flow was found, even though P. abies has a fragmented distribution. A weak but significant effect of isolation by distance was found. These results suggest a high dispersal capacity of I. typographus , which leads to low genetic differentiation between populations. Its high dispersal capacity is likely to have prevented I. typographus from developing important local adaptations to its host, which would have influenced its genetic structure. The nuclear data was compared to previously published mitochondrial data that showed strong differentiation between Central–Northern European populations and Russian–Baltic populations, and a founder effect in Scandinavia, probably reflecting the postglacial history of I. typographus . Discrepancies between nuclear and mitochondrial markers could be due to the maternal inheritance of mitochondrial DNA, and to sex-biased dispersal in I. typographus . The overall low genetic diversity observed on both markers on a large geographical scale is discussed. © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 90 , 239–246.     

Contrasting levels of genetic diversity between the common, self-compatible Liparis kumokiri and rare, self-incompatible Liparis makinoana (Orchidaceae) in South Korea

Levels of allozyme variation and intrapopulation spatial genetic structure of the two terrestrial clonal orchids Liparis kumokiri , a self-compatible relatively common species, and L. makinoana , a self-incompatible rare species, were examined for 17 ( N  = 1875) and four ( N  = 425) populations, respectively, in South Korea. Populations of L. makinoana harboured high levels of genetic variation ( H _e = 0.319) across 15 loci. In contrast, L. kumokiri exhibited a complete lack of allozyme variation ( H _e = 0.000). Considering the lack of genetic variability, it is suggested that current populations of L. kumokiri in South Korea originated from a genetically depauperate ancestral population. For L. makinoana , a significant deficit of heterozygosity (mean F _IS = 0.198) was found in population samples excluding clonal ramets, suggesting that pollen dispersal is localized, generating biparental inbreeding. The significant fine-scale genetic structuring (≤ 2 m) found in a previous study, in addition to the moderate levels of population differentiation ( F _ST = 0.107) and the significant relationship between genetic and geographical distances ( r  = 0.680) found here, suggests a leptokurtic distribution of seed dispersal for L. makinoana . Although populations of L. makinoana harbour high levels of genetic variation, they are affected by a recent genetic bottleneck. This information suggests that genetic drift and limited gene flow could be the main evolutionary forces for speciation of a species-rich genus such as Liparis .  © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 153 , 41–48.     

Genetic and morphological divergence reveals local subdivision of perch (Perca fluviatilis L.)

The level of gene flow is an important factor influencing genetic differentiation between populations. Typically, geographic distance is considered to be the major factor limiting dispersal and should thus only influence the degree of genetic divergence at larger spatial scales. However, recent studies have revealed the possibility for small-scale genetic differentiation, suggesting that the spatial scale considered is pivotal for finding patterns of isolation by distance. To address this question, genetic and morphological differentiation were studied at two spatial scales (range 2–13 km and range 300 m to 2 km) in the perch ( Perca fluviatilis L.) from the east coast archipelago of Sweden, using seven microsatellite loci and geometric morphometrics. We found highly significant genetic differentiation between sampled locations at both scales. At the larger spatial scale, the distance per se was not affecting the level of divergence. At the small scale, however, we found subtle patterns of isolation by distance. In addition, we also found morphological divergence between locations, congruent with a spatial separation at a microgeographic scale, most likely due to phenotypic plasticity. The present study highlights the importance of geographical scale and indicates that there might be a disparity between the dispersal capacity of a species and the actual movement of genes. Thus, how we view the environment and possible barriers to dispersal might have great implications for our ability to fully understand the evolution of genetic differentiation, local adaptation, and, in the end, speciation.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 746–758.     

The colonization of Scottish islands by the common shrew, Sorex araneus (Eulipotyphla: Soricidae)

In this paper we apply molecular methods to study the colonization of islands off the west coast of Scotland by the common shrew ( Sorex araneus L.), and current gene flow. We collected 497 individuals from 13 islands of the Inner Hebrides and Clyde Island groups and six mainland regions. Individuals were genotyped at eight microsatellite loci, and the mitochondrial cytochrome b sequence (1140 base pairs) was obtained for five individuals from each island/mainland region. Based on these molecular data, island colonization apparently proceeded directly from the mainland, except for Islay, for which Jura was the most likely source population. Raasay may also have been colonized by island hopping. Most island populations are genetically very distinct from the mainland populations, suggesting long periods of isolation. Two exceptions to this are the islands of Skye and Seil, which are geographically and genetically close to the mainland, suggesting in each case that there has been long-term gene flow between these islands and the mainland. We consider possible methods of island colonization, including human-mediated movement, swimming, and land and ice bridges.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 94 , 797–808.     

Does provenance matter? Fitness is not determined by genetic distance or the scale of pollen dispersal in Grevillea sphacelata (Proteaceae)

Understanding how the scale of pollen transfer determines the outcome of matings is important evolutionarily and a key issue in restoration ecology. We tested the effects of pollen transfer distance for the self‐incompatible shrub Grevillea sphacelata using (1) open pollination and transfer among (2) near neighbours, (3) neighbouring subpopulations and (4) populations separated by c. 4 km. We used AFLP markers to test for evidence of genetic differentiation within and among populations. Patterns of seed initiation suggest that open pollinated flowers were pollen limited, although in one subpopulation open seed set was greater than that achieved with pollen from near neighbours or other subpopulations. We detected no other effects of pollen source on seed initiation or seed and seedling development. In contrast, our genetic survey revealed significant spatial autocorrelation to 5 m, moderate differentiation of populations separated by up to 4 km and significant isolation by distance > 16 km. Our data suggest that, although dispersal of pollen may typically be localized, gene flow prevents localized adaptation or co‐adaptation and we detected no effects of inbreeding depression. In a restoration context, our results imply that movement of seed between populations separated by 4 km will not have detrimental consequences, despite significant differentiation at neutral genetic markers, and may be beneficial in maintaining genetic diversity and evolutionary potential. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 173 , 290–302.     

Positive effects of flower abundance and synchronous flowering on pollination success,and pollinia dispersal in rewardless Changnienia amoena (Orchidaceae)

Pollination success and pollen dispersal in natural populations depend on the spatial‐temporal variation of flower abundance. For plants that lack rewards for pollinators, pollination success is predicted to be negatively related to flower density and flowering synchrony. We investigated the relationships between pollination success and flower abundance and flowering synchrony, and estimated pollinia dispersal distance in a rewardless species, Changnienia amoena (Orchidaceae). The results obtained in the present study revealed that male pollination success was negatively influenced by population size but was positively affected by population density, whereas female pollination success was independent of both population size and density. Phenotypic analysis suggested that highly synchronous flowering was advantageous through total pollination success, which is in contrast to previous studies. These results indicate that pollination facilitation rather than competition for pollinator visits occurs in this rewardless plant. The median distance of pollinia dispersal was 11.5 m (mean distance = 17.5 m), which is comparable to that of other rewardless plants but longer than for rewarding plants. However, pollen transfer occured mainly within populations; pollen import was a rare event. Restricted gene flow by pollinia and seeds probably explains the previous population genetic reporting a high degree of genetic differentiation between populations. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 99 , 477–488.     

Contrasting temporal dynamics and spatial patterns of population genetic structure correlate with differences in demography and habitat between two closely-related African freshwater snails

The relationship between habitat stability, demography, and population genetic structure was explored by comparing temporal microsatellite variability spanning a decade in two closely-related hermaphroditic freshwater snails from Cameroon, Bulinus forskalii and Bulinus camerunensis . Although both species show similar levels of preferential selfing, microsatellite analysis revealed significantly greater allelic richness and gene diversity in populations of the highly endemic B. camerunensis compared to those of the geographically-widespread B. forskalii . Additionally, B. camerunensis populations showed significantly lower spatial genetic differentiation, higher dispersal rates, and greater temporal stability compared to B. forskalii populations over a similar spatial scale. This suggests that a more stable demography and greater gene flow account for the elevated genetic diversity observed in this geographically-restricted snail. This contrasts sharply with a metapopulation model (which includes extinction/contraction, recolonization/expansion, and passive dispersal) invoked to account for population structuring in B. forskalii . As intermediate hosts for medically important schistosome parasites, these findings have ramifications for determining the scale at which local adaptation may occur in the coevolution of these snails and their parasites.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 90 , 747–760.     

Microsatellite DNA polymorphism confirms reproductive isolation and reveals differences in population genetic structure of cryptic pipistrelle bat species

Previous studies have indicated that the common European pipistrelle bat ( Pipistrellus pipistrellus ) comprises two cryptic species, P. pipistrellus and Pipistrellus pygmaeus , which differ in echolocation call frequency and mitochondrial DNA sequence. However, levels of divergence based on nuclear markers have not been examined, and hence the potential for male-mediated gene flow between the species cannot be discounted. Moreover, little is known about population structure and migration patterns in either species. Here, we describe the use of microsatellites to investigate nuclear DNA differentiation between, and the pattern of population genetic structure within, the two cryptic pipistrelle species. In total, 1300 individuals from 82 maternity colonies were sampled across the British Isles and Continental Europe. We show, using multivariate analyses, that colonies of the same species are generally genetically more similar to each other than to those from the other species regardless of geographical location. Our findings support the hypothesis that the species are reproductively isolated. Significant patterns of genetic isolation by distance were identified in both species, indicating that mating may occur before any long-distance autumnal migration. The presence of a sea channel does not confer higher levels of genetic differentiation among colonies over and above distance alone in either species. Differences in genetic population structure were identified between the species, with P. pipistrellus showing a wider range of levels of genetic differentiation among colonies and a stronger relationship between genetic and geographical distance than P. pygmaeus . Differences in dispersal, mating behaviour, colony size and/or postglacial colonization patterns could contribute to the differences observed.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 90 , 539–550.