Pavel Hulva

Multilocus detection of wolf x dog hybridization in Italy, and guidelines for marker selection

Hybridization and introgression can impact the evolution of natural populations. Several wild canid species hybridize in nature, sometimes originating new taxa. However, hybridization with free-ranging dogs is threatening the genetic integrity of grey wolf populations (Canis lupus), or even the survival of endangered species (e.g., the Ethiopian wolf C. simensis). Efficient molecular tools to assess hybridization rates are essential in wolf conservation strategies. We evaluated the power of biparental and uniparental markers (39 autosomal and 4 Y-linked microsatellites, a melanistic deletion at the β-defensin CBD103 gene, the hypervariable domain of the mtDNA control-region) to identify the multilocus admixture patterns in wolf x dog hybrids. We used empirical data from 2 hybrid groups with different histories: 30 presumptive natural hybrids from Italy and 73 Czechoslovakian wolfdogs of known hybrid origin, as well as simulated data. We assessed the efficiency of various marker combinations and reference samples in admixture analyses using 69 dogs of different breeds and 99 wolves from Italy, Balkans and Carpathian Mountains. Results confirmed the occurrence of hybrids in Italy, some of them showing anomalous phenotypic traits and exogenous mtDNA or Y-chromosome introgression. Hybridization was mostly attributable to village dogs and not strictly patrilineal. The melanistic β-defensin deletion was found only in Italian dogs and in putative hybrids. The 24 most divergent microsatellites (largest wolf-dog FST values) were equally or more informative than the entire panel of 39 loci. A smaller panel of 12 microsatellites increased risks to identify false admixed individuals. The frequency of F1 and F2 was lower than backcrosses or introgressed individuals, suggesting hybridization already occurred some generations in the past, during early phases of wolf expansion from their historical core areas. Empirical and simulated data indicated the identification of the past generation backcrosses is always uncertain, and a larger number of ancestry-informative markers is needed.

Mechanisms of radiation in a bat group from the genus Pipistrellus inferred by phylogeography, demography and population genetics

Here, we present a study of the Pipistrellus pipistrellus species complex, a highly diversified bat group with a radiation centre in the Mediterranean biodiversity hotspot. The study sample comprised 583 animals from 118 localities representatively covering the bats’ range in the western Palearctic. We used fast‐evolving markers (the mitochondrial D‐loop sequence and 11 nuclear microsatellites) to describe the phylogeography, demography and population structure of this model taxon and address details of its diversification. The overall pattern within this group includes a mosaic of phylogenetically basal, often morphologically distant, relatively small and mostly allopatric demes in the Mediterranean Basin, as well as two sympatric sibling species in the large continental part of the range. The southern populations exhibit constant size, whereas northern populations show a demographic trend of growth associated with range expansion during the Pleistocene climate oscillations. There is evidence of isolation by distance and female philopatry in P. pipistrellus sensu stricto. Although the northern populations are reproductively isolated, we detected introgression events among several Mediterranean lineages. This pattern implies incomplete establishment of reproductive isolating mechanisms in these populations as well as the existence of a past reinforcement stage in the continental siblings. The occurrence of reticulations in the radiation centre among morphologically and ecologically derived relict demes suggests that adaptive unequal gene exchange within hybridizing populations could play a role in speciation and adaptive radiation within this group.

Phylogeography and postglacial recolonization of Europe by Rhinolophus hipposideros: evidence from multiple genetic markers

The demographic history of Rhinolophus hipposideros (lesser horseshoe bat) was reconstructed across its European, North African and Middle‐Eastern distribution prior to, during and following the most recent glaciations by generating and analysing a multimarker data set. This data set consisted of an X‐linked nuclear intron (Bgn; 543 bp), mitochondrial DNA (cytb‐tRNA‐control region; 1630 bp) and eight variable microsatellite loci for up to 373 individuals from 86 localities. Using this data set of diverse markers, it was possible to determine the species’ demography at three temporal stages. Nuclear intron data revealed early colonization into Europe from the east, which pre‐dates the Quaternary glaciations. The mtDNA data supported multiple glacial refugia across the Mediterranean, the largest of which were found in the Ibero‐Maghreb region and an eastern location (Anatolia/Middle East)–that were used by R. hipposideros during the most recent glacial cycles. Finally, microsatellites provided the most recent information on these species’ movements since the Last Glacial Maximum and suggested that lineages that had diverged into glacial refugia, such as in the Ibero‐Maghreb region, have remained isolated. These findings should be used to inform future conservation management strategies for R. hipposideros and show the power of using a multimarker data set for phylogeographic studies.

Molecules, morphometrics and new fossils provide an integrated view of the evolutionary history of Rhinopomatidae (Mammalia: Chiroptera)

BackgroundThe Rhinopomatidae, traditionally considered to be one of the most ancient chiropteran clades, remains one of the least known groups of Rhinolophoidea. No relevant fossil record is available for this family. Whereas there have been extensive radiations in related families Rhinolophidae and Hipposideridae, there are only a few species in the Rhinopomatidae and their phylogenetic relationship and status are not fully understood.ResultsHere we present (a) a phylogenetic analysis based on a partial cytochrome b sequence, (b) new fossils from the Upper Miocene site Elaiochoria 2 (Chalkidiki, Greece), which represents the first appearance datum of the family based on the fossil record, and (c) discussion of the phylogeographic patterns in both molecular and morphological traits. We found deep divergences in the Rhinopoma hardwickii lineage, suggesting that the allopatric populations in (i) Iran and (ii) North Africa and the Middle East should have separate species status. The latter species (R. cystops) exhibits a shallow pattern of isolation by distance (separating the Middle East and the African populations) that contrasts with the pattern of geographic variation in the morphometrical traits. A deep genetic gap was also found in Rhinopoma muscatellum (Iran vs. Yemen). We found only minute genetic distance between R. microphyllum from the Levant and India, which fails to support the sub/species distinctness of the Indian form (R. microphyllum kinneari).ConclusionThe mtDNA survey provided phylogenetic tree of the family Rhinopomatidae for the first time and revealed an unexpected diversification of the group both within R. hardwickii and R. muscatellum morphospecies. The paleobiogeographic scenario compiled in respect to molecular clock data suggests that the family originated in the region south of the Eocene Western Tethyan seaway or in India, and extended its range during the Early Miocene. The fossil record suggests a Miocene spread into the Mediterranean region, followed by a post-Miocene retreat. Morphological analysis compared with genetic data indicates considerable phenotypic plasticity in this group.

Systematic status of African populations of Pipistrellus pipistrellus complex (Chiroptera: Vespertilionidae), with a description of a new species from Cyrenaica, Libya

The distribution of pipistrelles of the Pipistrellus pipistrellus complex (= P. pipistrellus s.l.) reaches only marginally the African continent. These bats are known only from a narrow belt of the Mediterranean zone in Maghreb and from NE Libya. We analysed museum specimens of African populations of P. pipistrellus s.l. using both morphologic and genetic techniques and compared them with Eurasian specimens of the complex. The African representatives of P. pipistrellus complex include two morphologically, genetically and geographically distinct populations. One of them inhabits the Mediterranean part of Cyrenaica, Libya. Belonging to the P. pygmaeus genetic lineage, these bats are represented by larger and more rusty coloured individuals with large massive rostrum and canines. In morphologic traits, this population differs significantly from all Western Palaearctic populations of the P. pipistrellus complex. These bats differ by about 6–7% in genetic distance from P. pygmaeus s. str. Within the P. pygmaeus lineage Libyan bats seem to be unique in their echolocation calls: the maximum energy of terminal frequencies was recorded at about 45 kHz. We consider the Libyan pipistrelles to represent a separate species, Pipistrellus hanaki sp. nov. Another distinct African pipistrelle population inhabits the Mediterranean parts of NW African countries, Morocco, Algeria and Tunisia. Individuals from the latter population are small and somewhat darker members of the P. pipistrellus genetic lineage, with relatively short and narrow mesial part of rostrum. Although both morphological and genetic differences between this population and Eurasian P. pipistrellus s. str. were found (genetic distance about 3–5%), they are probably not sufficient for the separation of this form at the specific level. However, the differences from European samples show rather not a cline character and therefore potential subspecific level of NW African P. pipistrellus has to be taken into consideration.

When the Body Hides the Ancestry: Phylogeny of Morphologically Modified Epizoic Earwigs Based on Molecular Evidence

Here, we present a study regarding the phylogenetic positions of two enigmatic earwig lineages whose unique phenotypic traits evolved in connection with ectoparasitic relationships with mammals. Extant earwigs (Dermaptera) have traditionally been divided into three suborders: the Hemimerina, Arixeniina, and Forficulina. While the Forficulina are typical, well-known, free-living earwigs, the Hemimerina and Arixeniina are unusual epizoic groups living on molossid bats (Arixeniina) or murid rodents (Hemimerina). The monophyly of both epizoic lineages is well established, but their relationship to the remainder of the Dermaptera is controversial because of their extremely modified morphology with paedomorphic features. We present phylogenetic analyses that include molecular data (18S and 28S ribosomal DNA and histone-3) for both Arixeniina and Hemimerina for the first time. This data set enabled us to apply a rigorous cladistics approach and to test competing hypotheses that were previously scattered in the literature. Our results demonstrate that Arixeniidae and Hemimeridae belong in the dermapteran suborder Neodermaptera, infraorder Epidermaptera, and superfamily Forficuloidea. The results support the sister group relationships of Arixeniidae+Chelisochidae and Hemimeridae+Forficulidae. This study demonstrates the potential for rapid and substantial macroevolutionary changes at the morphological level as related to adaptive evolution, in this case linked to the utilization of a novel trophic niche based on an epizoic life strategy. Our results also indicate that the evolutionary consequences of the transition to an ectoparazitic mode of living, which is extremely rare in earwigs, have biased previous morphology-based hypotheses regarding the phylogeny of this insect group.

Alcathoe Bat (Myotis alcathoe) in the Czech Republic: Distributional Status, Roosting and Feeding Ecology

Between 2001 and 2008, we recorded Myotis alcathoe at nine sites within three distant areas in the Czech Republic. The species identification was confirmed with cyt b sequences and four distinct haplotypes were identified. All the localities exhibit surprisingly uniform habitat characteristics: (1) old full-grown oak-hornbeam forests, with (2) numerous large trees in advanced stages of decay are present, and (3) a very small to large water bodies and/or patches of riparian vegetation surrounded by the forest. Using radiotracking techniques, we discovered 27 day roosts of M. alcathoe, located mostly in big oak, birch and lime trees inside extensive forest stands. All roosts were fissures or small cavities in a tree trunk and in branches in the canopies, some 16m above the ground. Bats preferred trees that were higher, had higher canopy and canopy basement and had larger diameter at breast height than other available trees. Roost trees were surrounded by lower trees with lower canopy basements than available trees. Roost trees were in a poorer condition than other available trees. Roosts were occupied by up to 83 individuals in July but usually single individuals were found in the roosts in September. In contrast to syntopic M. mystacinus and M. brandtii, M. alcathoe has never been found in an anthropogenic roost (except for a fissure in concrete electricity pole). Preliminary analysis of the diet showed that nematoceran flies were the most important prey item along with spiders, caddis flies, small moths and neuropterans. In the observed ecological characteristics, M. alcathoe markedly differs from other European species of the genus Myotis. Its restricted habitat requirements are perhaps responsible for an islet-like pattern of its distribution and suggest an essential conservation value of the habitats of its occurrence.

Craseonycteris thonglongyai (Chiroptera: Craseonycteridae) is a Rhinolophoid: Molecular Evidence from Cytochrome b

Craseonycteris thonglongyai (Chiroptera: Craseonycteridae), an enigmatic taxon which shares morphological traits with both Rhinopomatidae and Emballonuridae was for the first time investigated with the aid of molecular phylogenetic techniques. Three methods of phylogenetic inference, parsimony, maximum-likelihood, and Bayesian phylogenetics were used. Based on 402 bp of DNA sequence from the mitochondrial cytochrome b gene, placement of Craseonycteridae within the superfamily Rhinolophoidea was demonstrated. Our results also suggest close proximity of Craseonycteridae to Hipposideridae rather than to Rhinopomatidae, close relationships between Megadermatidae and Rhinolophidae, sister group position of Pteropodidae to Rhinolophoidea, and closer affiliation of Nycteridae with the infraorder Yangochiroptera. Spectral analysis was in agreement with all these outcomes except for closer relationships of Craseonycteris with Rhinopomatidae.

Syntopic Occurrence in Turkey Supports Separate Species Status for Miniopterus schreibersii schreibersii and M. schreibersii pallidus (Mammalia: Chiroptera)

Miniopterus schreibersii is a complex, polytypic species group with a wide distribution ranging from Northern Africa, Southern Europe to Asia, the Solomon Islands and Northern Australia. Two subspecies previously recognized in Turkey, M. s. schreibersii and M. s. pallidus, differ significantly in nuclear and mitochondrial DNA, and in morphology. Until now, the distribution records of M. s. schreibersii and M. s. pallidus showed that they were allopatric and hence even though there was clear morphological and genetic differentiation between the two taxa, whether they represented separate species or subspecies was still not determined with certainty. Here we present the first data on syntopic occurrence of both forms in three caves in south-eastern Turkey. We show that the three caves host individuals of both taxa by using mitochondrial DNA, nuclear DNA and morphometric analyses. These findings provide the final line of evidence to date, for designation of M. s. schreibersii and M. s. pallidus as two separate species, M. schreibersii and M. pallidus. This will raise the number of species in the Eurobats agreement area from 45 to 46.

Hidden diversity in Senegalese bats and associated findings in the systematics of the family Vespertilionidae

IntroductionThe Vespertilionidae is the largest family of bats, characterized by high occurrence of morphologically convergent groups, which impedes the study of their evolutionary history. The situation is even more complicated in the tropics, where certain regions remain under-sampled.ResultsTwo hundred and thirteen vespertilionid bats from Senegal (West Africa) were studied with the use of non-differentially stained karyotypes and multi-locus sequence data analysed with maximum likelihood and Bayesian methods. These bats were identified as 10 different taxa, five of which were distinctive from their nominate species (Pipistrellus hesperidus, Nycticeinops schlieffenii, Scotoecus hirundo, Neoromicia nana and N. somalica), based on both karyotypes and molecular data. These five cryptic taxa are unrelated, suggesting that these West African populations have long been isolated from other African regions. Additionally, we phylogenetically analysed 166 vespertilionid taxa from localities worldwide using GenBank data (some 80% of the genera of the family) and 14 representatives of closely related groups, together with our Senegalese specimens. The systematic position of several taxa differed from previous studies and the tribes Pipistrellini and Vespertilionini were redefined. The African Pipistrellus rueppellii was basal to the Pipistrellus/Nyctalus clade and the Oriental species Glischropus tylopus was basal to the East Asian pipistrelles within the tribe Pipistrellini. The African genus Neoromicia was confirmed to be diphyletic. Based on GenBank data, Eptesicus was polyphyletic, with the Asian E. nasutus and E. dimissus both supported as phylogenetically distinct from the Eptesicus clade. The subfamily Scotophilinae was confirmed as one of the basal branches of Vespertilionidae.ConclusionsNew taxa and new systematic arrangements show that there is still much to resolve in the vespertilionids and that West Africa is a biogeographic hotspot with more diversity to be discovered.