Evolutionary relationships among hares from North Africa (Lepus sp. or Lepus spp.), cape hares (L. capensis) from South Africa, and brown hares (L. europaeus), as inferred from mtDNA PCR-RFLP and allozyme data

Systematics and taxonomy of hares of the genus Lepus (Lagomorpha) are under contentious debate, and phylogenetic relationships among many taxa are not well understood. Here we study genetic differentiation and evolutionary relationships among North African hares, currently considered subspecies of Lepus capensis , cape hares ( L. capensis ) from the Cape province in South Africa, and brown hares ( L. europeaus ) from Europe and Anatolia, using maternally (mtDNA) and biparentally (allozymes) inherited markers. A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of a c. 1.8 kb long segment of the mitochondrial control region using eight hexanucleotide-recognizing restriction endonucleases yielded 28 haplotypes, and horizontal starch gel electrophoresis of proteins encoded by 25 structural gene loci revealed 52 alleles at 18 polymorphic loci. Diverse phylogenetic analyses (neighbor joining dendrogram, median joining network, multidimensional scaling of pairwise distances, AMOVA, F -statistics, hierarchical F -statistics) of genetic variants revealed marked substructuring of mtDNA into three phylogeographic groups, namely an African, a central European, and an Anatolian, but a somewhat less pronounced overall differentiation of the nuclear genome, despite a relatively high number of population-specific (private) alleles. However, all our results are not incongruent with Petter's (1959: Mammalia 23 , 41; 1961: Z. f. Säugetierkunde 26 , 30; 1972 : Société Des Sciences Naturelles et Physiques du Maroc 52 , 122) hypothesis that North African hares generally belong to L. capensis and that brown hares should be included in this species as well.     

Molecular evidence of conspecificity of South African hares conventionally considered Lepus capensis L., 1758

Conventionally, Lepus capensis is considered to range across large parts of Africa, the Middle East, Central and Far East Asia. However, a recent morphological study restricts cape hares tentatively to a small range in the Western Cape Region of South Africa and groups all other L. capensis-type hares from South Africa into a new species: L. centralis. Here, we studied molecular relationships among L. capensis-type hares from South Africa. Phenotypically and morphologically the individuals matched either the newly described L. capensis or L. centralis. We examined 66 hares for allelic variation at 13 microsatellite loci and for sequence variation of the hypervariable domain 1 of the mitochondrial control region. All tree-generating analyses of the currently obtained sequences and all South African cape hare sequences downloaded from GenBank revealed monophyly when compared to sequences of various other Lepus species. A network analysis indicated close evolutionary relationships between hares of the “L. capensis-phenotype” and the “L. centralis-phenotype” (according to Palacios et al. 2008) from the southwest of the Western Cape, relative to their pronounced evolutionary divergence from all other more central, northern, and north-eastern L. capensis-type hares. F-statistics, a Bayesian admixture STRUCTURE model, as well as a principal coordinate analysis of microsatellite data indicated close genetic relationships among all South African L. capensis-type hares studied presently. A coalescence model-based migration analysis for microsatellite alleles indicated gene flow between most of the considered subspecies of cape hare, including L. capensis capensis and L. capensis centralis, theoretically sufficient to balance stochastic drift effects. Concordantly, AMOVA models revealed only little effects of partitioning microsatellite variation into the two suggested morpho-species “L. capensis” and “L. centralis”. Under an “Interbreeding Species Concept” (e.g. a strict or relaxed Biological Species Concept), the current molecular data demonstrate conspecificity of the two proposed morpho-species “L. capensis” and “L. centralis”. Based on the present molecular data the differentiation of subspecies of cape hares from southern Africa is discussed.     

Phylogenetic analysis of mtCR-1 sequences of Tunisian and Egyptian hares (Lepus sp. or spp., Lagomorpha) with different coat colours

North African hares are currently considered belonging to cape hares (Lepus capensis), except for an isolated occurrence of L. victoriae in NW Algeria. However, the few existing molecular data are not unequivocal. Here, we study sequence variation (415 bp) in the hypervariable domain-1 of the mitochondrial (mt) control region, of hares with different coat colour from north-central Tunisia and NW Egypt, to test Petter's [(1959): Eléments d’une révision des Lièvres africains du sous-genre Lepus. Mammalia 23, 41–67] hypothesis that North African hares belong to L. capensis. Seven Tunisian and one Egyptian haplotypes were revealed from 28 hares and compared phylogenetically to 245 haplotypes of various Lepus species downloaded from GenBank. Neighbour joining (NJ) and principal coordinate (PCO) analyses based on a Tamura-Nei 93 distance matrix, as well as maximum parsimony (MP) analysis concordantly grouped all currently obtained haplotypes together into one monophyletic clade, and revealed relatively close relationships to the clades of African scrub hares (L. saxatilis) and brown hares (L. europaeus). The three distinguished coat colour types of Tunisian hares were paralleled only to a small extent by sequence differentiation. Haplotypes of L. capensis from the nominal Cape province of South Africa, North Africa, and China clustered into different major clades, respectively, with Chinese L. capensis haplotypes forming only a subclade within a major clade that encompassed predominantly “mountain/arctic hare-type sequences” in addition to sequences of several other palaearctic and nearctic species. One further Chinese L. capensis haplotype clustered into the L. comus clade. These results indicated occurrence of introgression and/or shared ancestral polymorphism. Such an evolutionary scenario implies using nucelar markers in addition to mtDNA for phylogenetic inferences in the genus Lepus; nevertheless, mtDNA is still useful for inferring phylogenetic history and biogeography of hares.     

Speciation and paraphyly in western mediterranean hares (Lepus castroviejoi,L. europaeus,L. granatensis,andL. capensis) revealed by mitochondrial DNA phylogeny

Mitochondrial DNA (mtDNA) variation among specimens of the northwestern African hare (Lepus capensis schlumbergeri) and three European hares sampled in Spain (L. castroviejoi andL. granatensis, which are endemic to the Iberian Peninsula, andL. europaeus) was analyzed using seven restriction endonucleases. Fourteen haplotypes were found among the 34 animals examined. Restriction site maps were constructed and the phylogeny of the haplotypes was inferred. mtDNA ofL. capensis was the most divergent, which is consistent with its allopatric African distribution and with an African origin of European hares. We estimated that mtDNA in hares diverges at a rate of 1.5–1.8% per MY assuming that the European and African populations separated 5–6 MYBP. Maximum intraspecies nucleotide divergences were 1.3% inL. capensis, 2.7% inL. castroviejoi, and 2.3% inL. granatensis but 13.0% inL. europaeus. The latter species contained two main mtDNA lineages, one on the branch leading toL. castroviejoi and the other on that leading toL. granatensis. The separation of these two lineages from theL. castroviejoi orL. granatensis lineages appears to be much older than the first paleontological record ofL. europaeus in the Iberian peninsula. This suggests that the apparent polyphyly ofL. europaeus is due not to secondary introgression, but to the retention of ancestral polymorphism inL. europaeus. The results suggest thatL. europaeus either has evolved as a very large population for a long time or has been fractionated. Such a pattern of persistence of very divergent lineages has also been reported in other species of highly mobile terrestrial mammals. As far as mtDNA is concerned,L. europaeus appears to be the common phylogenetic trunk which has diversified during dispersion over the European continent and from whichL. castroviejoi andL. granatensis speciated separately in southwest Europe.     

Differentiation under isolation and genetic structure of Sardinian hares as revealed by craniometric analysis,mitochondrial DNA and microsatellites

Hares (Lepus capensis Linnaeus 1758) were probably introduced into Sardinia in historical times. Previous studies indicated North Africa as the most likely source area but did not exclude the occurrence of hybridization events with continental brown hares (L. europaeus Pallas 1778) perhaps introduced for hunting purposes. We implemented both morphometric and genetic approaches to verify the genetic isolation of the Sardinian population. Specifically, we conducted a multivariate analysis of craniometric data and analysed 461 bp of the mitochondrial control region and 12 autosomal microsatellites in Sardinian hares, using North African cape hares and European brown hares as reference populations. Sardinian hares displayed a peculiar skull shape. In agreement, both nuclear and mitochondrial markers remarked the distinctiveness of this population. Observed and expected heterozygosity were 0.52 and 0.61, while haplotype and nucleotide diversity were 0.822 and 0.0129. Self‐assignment based on Bayesian cluster analysis was high (average membership 0.98), and no evident signs of introgression from continental brown hares were found. Our results support the hypothesis that the Sardinian hares have been introduced from North Africa, remained genetically isolated since the founding event and evolved independently from the source population. This long‐lasting isolation and the consequent genetic drift resulted in a differentiation, perhaps accompanied by an adaptation to local environmental conditions.     

Molecular Approaches Revealing Prehistoric, Historic, or Recent Translocations and Introductions of Hares (Genus Lepus) by Humans

Although only of medium size, and thus of little nutritional value compared to big game such as mammoths and ungulates, hares (Lepus spp.) probably have always been a food source for humans, as documented in archaeological finds. Nowadays, hares, particularly such species as the brown hare (L. europaeus), are among the most important game species in many European countries. For hunting, perhaps religious reasons, and in connection with certain myths, hares have been and are still being intentionally translocated. Ancient translocations by humans can be inferred from the presence of hares on islands that had no mainland connections, at least during the Pleistocene, the major evolutionary period of the genus Lepus. We review some of the literature on anthropogenic translocations of hares. We focus on three examples [the brown hare (L. europaeus), the Corsican hare (L. corsicanus), and the Sardinian hare (L. capensis)], where some molecular data could be used to trace the translocation routes and possible origins of introduced hare populations. Certain molecular marker systems, such as sequences of the hypervariable part I (HV-1) of the mitochondrial control region, show high variability in hare species and are thus promising for tracing both recent and ancient origins of translocated hares. Some other molecular marker systems as well as caveats connected with the use of such marker systems in the genus Lepus are also discussed.     

Factors related to the occurrence of hybrids between brown hares Lepus europaeus and mountain hares L. timidus in Sweden

Hybridization occurs among many species, and may have implications for conservation as well as for evolution. Interspecific gene flow between brown hares Lepus europaeus and mountain hares L. timidus has been documented in Sweden and in continental Europe, and has probably to some extent occurred throughout history in sympatric areas. What local factors or ecological relationships that correlate with or trigger hybridization between these species has however been unclear. We studied spatial distribution of hybrids between brown hares and mountain hares in Sweden in relation to characteristics of the sampled localities (hunting grounds). In a sample of 70 brown hares collected from 39 populations in south‐central Sweden during 2003–2005, 11 (16%) showed introgressed mtDNA from mountain hares. Among the brown hares from their northern range, i.e. in general the most recent establishments, the corresponding figure was 75% (9/12). The frequency of samples with hybrid ancestry increased significantly with latitude, altitude and hilliness, and were higher (p<0.1) in recently established populations and/or where the proportion of arable land was low. Several site‐specific parameters were correlated, e.g. latitude as expected to hilliness, and no parameter explained the occurrence of hybrids exclusively. Instead, the appearance of mountain hare mtDNA among brown hares was associated with a conglomerate of parameters reflecting landscapes atypical for the brown hare, e.g. forest dominated and steep areas where the species quite recently was established. We suggest that these abiotic factors mirror the main aspect influencing hybridization frequency, namely the density or relative frequency of the two species. In atypical brown hare landscapes with recent establishment, mountain hares are probably relatively more common. When one species dominate in numbers, or when both species display low densities, increased frequency of hybridization is expected due to low availability of conspecific partners, a phenomenon referred to as Hubbs’ principle.     

On shortcomings of using mtDNA sequence divergence for the systematics of hares (genus Lepus): An example from cape hares

Despite the well-known fact that evolutionary patterns of single genes or sequences are not necessarily paralleled by organismic evolution, using mitochondrial sequence divergence for inferring phylogenetic relationships among hare species is not uncommon. Earlier studies reported interspecific introgression in the genus Lepus that may slur systematic conclusions drawn exclusively from mtDNA data. We examined pairwise divergence in partial mtHV-1 sequences and nuclear gene pools based on microsatellite and allozyme loci separately in a South and North African population of cape hares, Lepus capensis, and did not find significant correspondence on the individual level within either population. Also, the former population had a significantly higher level of mtHV-1 sequence divergence compared to the latter, but levels of individual nuclear gene pool divergence did not differ significantly between the two populations. Hence, the absence of correspondence between mtHV-1 sequence divergence and nuclear gene pool divergence among individuals within a population might be independent of the population-specific level of differentiation among individuals. Our results complement earlier findings in hares, and strongly recommend to include nuclear gene pool evidence for systematic inferences within this genus, even under the absence of mitochondrial introgression.     

Mitochondrial CR-1 Variation in Sardinian Hares and Its Relationships with Other Old World Hares (Genus Lepus)

Among the European fauna, the Sardinian hare (Lepus sp.) is peculiar in that it differs from all other hares inhabiting the continent. Here, we report on the variation of a 461 bp sequence of hypervariable domain 1 of the mitochondrial control region, examined in 42 hares collected throughout Sardinia and compared to the corresponding sequences of different Lepus taxa. Seventeen novel haplotypes were found in the Sardinian population, resulting in a haplotype diversity of 0.840 and a nucleotide diversity of 0.012. As a result of Bayesian and principal coordinates analyses, Sardinian hares were grouped with North African hares, constituting a monophyletic clade that diverges from all other Old World hares, including Cape hares from South Africa and East Asia. Hence, our data agree that populations inhabiting North Africa and Sardinia form a distinct taxon, which could possibly be included in the L. capensis superspecies. Moreover, two corresponding lineages can be found in Sardinia and Tunisia, providing evidence of a common origin of the two populations and thus supporting the hypothesis that North African hares were introduced into the island in historical times. Our data show that the two lineages differ in their geographic distribution throughout the island and that the wild Sardinian population also shows the signature of a postintroduction demographic expansion.     

Genetic variation in the major histocompatibility complex of the European brown hare (Lepus europaeus) across distinct phylogeographic areas

The major histocompatibility complex is one of the best studied systems in vertebrates providing evidence for the long-term action of selection. Here, we examined the intra- and inter-population genetic diversity of the MHC class II DRB locus in European brown hare (Lepus europaeus) and correlated the results with genetic variability already estimated from the MHC DQA locus and from maternally (mitochondrial DNA (mtDNA)) and biparentally (allozymes, microsatellites) inherited loci. L. europaeus showed remarkable genetic polymorphism in both DQA and DRB1 loci. The Anatolian populations exhibited the highest genetic polymorphism for both loci. Balancing selection has established increased variability in the European populations despite the founder effects after the last glaciation. Different evolutionary rates were traced for DRB1 and DQA loci, as evidenced by the higher number of common DRB1 than DQA alleles and the greater differences between DRB1 alleles with common origin in comparison with DQA alleles. The high number of rare alleles with low frequencies detected implies that frequency-dependent selection drives MHC evolution in the brown hare through the advantage of rare alleles. Both loci were under the influence of positive selection within the peptide-binding region. The functional polymorphism, recorded as amino acid substitutions within the binding pockets, fell also within distinct geographic patterns, yet it was much narrower than the genetic polymorphism. We hypothesize that certain structural and functional characteristics of the binding pockets set limitations to the actual shape of genetic polymorphism in MHC.     

Analysis of MC1R genetic variation in Lepus species in Mediterranean refugia

A study on the inter- and intraspecies variation of MC1R gene was performed in Lepus species inhabiting the Mediterranean basin (L. granatansis, L. europaeus, L. corsicanus, L. castroviejoi and L. mediterraneus) and their neighboring species in Europe (L. timidus) and Africa (L. saxatilis, L. capensis), in order to infer micro- versus macroevolutionary adaptation. Eleven different sequences were isolated that corresponded to five amino acid sequences. Comparison of MC1R nucleotide phylogenetic tree with phylogenies resulting from mtDNA regions of the same species showed absence of congruence between these sets of markers. The Mediterranean area that offered refugia during last glaciation retains more MC1R genotypes compared with populations of North and Central Europe as a consequence of founder effects. L. corsicanus and L. castroviejoi bore identical alleles supportive of their conspecificity, as indicated by other molecular markers. Within L. europaeus, a group of Israeli hares were distinguished by a different MC1R functional allele; additional differences in coat colour and other genetic markers raise doubts about its taxonomic status. Finally, the present data reinforced the idea of bi-directional introgressive hybridization between L. europaeus and L. timidus in Switzerland.     

The distribution of mountain hares Lepus timidus in Europe: a challenge from brown hares L. europaeus?

1. Throughout the most recent glacial period (Weichsel), the mountain hare Lepus timidus had a continuous distribution in the tundra habitat south of the ice‐rim. When the ice retreated, mountain hares colonized deglaciated land, and spread over northern Europe. 2. Since the Weichsel, the mountain hare's distribution in Europe has been gradually reduced and at present comprises Ireland and the Scottish Highlands, high altitudes in the Alps, isolated forests in eastern Poland, most of Fennoscandia and from the Baltic countries eastwards through Russia. Declines during the last century have been observed in Sweden and Russia. 3. This review defines and evaluates causes for this gradual reduction and fragmentation of the mountain hare's distribution, with special focus on interactions with brown hares Lepus europaeus. The relative importance of diseases, predation, cultivation and interactions with other herbivores than brown hares are discussed. 4. A plausible cause of the possible permanent disappearance of mountain hares in Europe appears to be exclusion by interspecific competition and hybridization with, and/or epidemic diseases mediated by, the congeneric brown hare.     

New Genetic Variation in European Hares, Lepus granatensis and L. europaeus

Six genetic polymorphisms for the Iberian hare (Lepus granatensis) and four for the brown hare (L. europaeus) are newly described. The genetic variation of peptidases B (PEPB) and C (PEPC), hemoglobin CHKCHK chain (HBA), hemopexin (HPX), vitamin D binding protein (GC), and properdin factor B (BF) was assessed by conventional electrophoresis and isoelectric focusing in carrier ampholytes and hybrid pH gradients. Six alleles were detected in PEPB, three in PEPC, four in HBA, six in GC, five in HPX, and six in BF. At least one allele was shared between species at all loci except HBA. The allelic overlap between the two species was medium to high in PEPB, GC, and HPX and small in PEPC and BF.     

Diet selection by European hares (Lepus europaeus) in the alpine zone of the Snowy Mountains, Australia

European hares (Lepus europaeus) are grazers and open grassland specialists that are replaced in mountain areas of their natural range in the northern hemisphere by browsing/intermediate feeding mountain hares (Lepus timidus), but in their introduced range in the southern hemisphere, occupy the alpine zone. We used micro-histological identification of plant fragments and germination of seeds in faecal pellets of L. europaeus from the Snowy Mountains, Australia, to determine diet. We asked whether diet shifted and/or diet breadth expanded in response to seasonally reduced food availability, particularly during winter. If so, did the constraints of food availability in the alpine zone lead to the diet mirroring that of L. timidus in its native alpine habitat. The diet of L. europaeus was dominated by grasses, herbs and shrubs. The main diet items in summer were grasses (70 %) and herbs (28 %). Grasses declined in the diet between summer and autumn when herbs increased to co-dominance, with a further change after establishment of the winter snowpack to a greater preponderance of shrubs (43 % compared with a maximum of 3 % in snow-free months). L. europaeus selected a wider range of plants in winter (59 species compared with 39 in summer) and diet was significantly more variable in winter than in autumn or summer (and in autumn than summer). We concluded that the persistence of L. europaeus in alpine areas of the southern hemisphere is testament to their ability to expand their dietary breadth to occupy mountain climatic zones normally occupied by L. timidus.     

Biochemical genetic variability in brown hares (Lepus europaeus) from Greece

Allozyme variability of 91 brown hares (Lepus europaeus) from seven regions in Greece was compared to existing data of Bulgarian populations to test the hypothesis of the occurrence of specific alleles in Greece, likely stemming from an isolated Late Pleistocene refugial population in the southern Balkans. This hypothesis is particularly suggested by some subfossil Late Pleistocene hare remains in Greece and the reported high mtDNA diversity in Greek hares. Allozymic diversity could be higher in Greek hares than in hares from neighboring regions as a result of the accumulation of variants in a long-lasting Pleistocene refugium. Conversely, Greek hares could exhibit reduced genetic diversity because of long-lasting low effective population sizes during the Late Glacial Maximum and a lower chance of postglacial gene flow from other populations into this rather marginal part in the southern Balkans. Horizontal starch gel electrophoresis of proteins from 35~loci revealed three alleles (Es-1 ^–162, Pep-2 ¹¹⁴, Mpi ⁸⁸) at low frequencies, which were not found in Bulgarian or any other brown hare population. In contrast, some alleles from the populations from Bulgaria and other regions of Europe were absent in the Greek samples. Population genetic statistics indicated only a slight tendency of increased gene pool diversity in Greek hares, little substructuring in Greek and Bulgarian populations, respectively, as well as an only slightly lower level of gene flow between the two neighboring regions, as compared to the gene flow within each region. The results conform to the hypothesis of a Late Pleistocene refugial population in the southern Balkans, with some few specific nuclear gene pool characteristics, but little effect on the overall genetic differentiation between Greek and Bulgarian hares.     

Y DNA and mitochondrial lineages in European and Asian populations of the brown hare (Lepus europaeus)

Both the Cytb gene of mtDNA and Y chromosome markers were studied in a relatively large sample of brown hares (L. europaeus) from Europe and Anatolia (Turkey and Israel), together with other seven Lepus species, in order to enable comparative analysis of possible sex-specific gene flow. Furthermore, Y chromosome markers were compared with data from biparentally inherited markers in an attempt to understand whether or not their pattern of distribution was congruent with that of allozymes or whether they rather matched mtDNA phylogenies, with which they share uniparental inheritance. Consistent with the general observation, levels of interspecific genetic variability were very low for the Y chromosome markers compared with mtDNA. Moreover, lack of interspecific variation for the Y-DNA studied within Lepus genus rendered these markers improper for any further phylogenetic analysis. With the highest nucleotide diversity in Anatolia compared with Europe, both marker systems confirmed an unbroken species history in Anatolia, corroborated the hypothesis of continuous gene flow from Anatolia's neighbouring regions, and supported the idea of a quick postglacial colonization followed by expansion of the species in large parts of Europe. Phylogenetic analysis under mtDNA revealed the existence of four different haplogroups with a well defined distribution across Europe and Anatolia. Both genetic systems supported the deep separation of Anatolian and European lineages of L. europaeus. Nevertheless, Anatolian Y-DNA lineages extended across a longer geographic distance in south-eastern Europe than Anatolian mtDNA haplotypes, probably as a result of higher female philopatry that makes mtDNA introgression more difficult in brown hares.     

Morphological evidence of species differentiation within Lepus capensis Linnaeus, 1758 (Leporidae,Lagomorpha) in Cape Province,South Africa

A morphological study was carried out of the hares (Lepus) from Cape Province previously assigned to the subspecies L. capensis capensis, L. c. centralis, and L. c. grantii. The purpose of the study was to characterize the species L. capensis Linnaeus, 1758. In doing so, it was possible to distinguish two populations which we consider different species, as each shows homogeneous cranial, dental and pelage features. One of them, defined as L. capensis, is distributed near Cape Town not far from the coast, between Lambert's Bay and Cape Agulhas. The other species, defined as L. centralis, which includes L. grantii as a synonym, is distributed in central and western Cape Province. L. capensis and L. centralis have a parapatric distribution, with a small area of sympatry in a contact zone in Compagnies Drift area, near Lambert's Bay. With respect to cranial differences between the two species, L. capensis has a stronger maxilla and more robust dental series, while L. centralis has larger tympanic bullae. Among dental characters, L. centralis usually has a deeper groove and more abundant cement than L. capensis in the first upper incisor, and its internal lobe is squared, while in L. capensis is rounded. As for pelage color and pattern, L. capensis shows a more extended white ventral area than L. centralis. Our results are of interest for further research on taxonomic problems regarding Old World hare populations in which L. capensis is concerned.     

The secret of Pianosa island: an Italian native population of European brown hare (<Emphasis Type="Italic">Lepus europaeus meridiei</Emphasis> Hilzheimer, 1906)

Given its relevance as a game species, the brown hare (Lepus europaeus Pallas, 1778) is one of the most managed and translocated mammals in Europe. In Italy, the species shows a genepool consisting of a mix of native and exotic lineages, due to translocations and introductions for hunting purposes. Some authors argued that the introduction of exotic brown hares could have caused the extinction of an endemic subspecies, L. e. meridiei Hilzheimer 1906, once present in central and northern Italy. Here we genetically characterized for the first time the brown hare population living in Pianosa island (part of the Tuscan Archipelago National Park) using 13 STR loci and a fragment of the mtDNA control region. All individuals analyzed share a unique haplotype, the L. europaeus haplotype Leu2, recognized as the ancestral mitochondrial lineage corresponding to the subspecies L. e. meridiei. Furthermore, considering autosomal markers, Pianosa brown hare population and current Italian peninsular population are genetically distinct. The discovery of this ancient population in a protected area, isolated and not affected by recent translocation/restocking events, has a great relevance in conservation and confirms the current presence of the endemic subspecies L. e. meridiei in Italy.     

Genetic diversity within Anatolian brown hares (Lepus europaeus Pallas, 1778) and differentiation among Anatolian and European populations

Genetic variability of Anatolian hares and relationships between Anatolian and European populations were assessed by a multilocus allozyme approach to infer evolutionary relationships between hares from Asia Minor and Europe. Of the 48 loci assayed, 19 (39.6%) were polymorphic with two to four alleles in the Anatolian hares. Among all Anatolian alleles, 14 were so far not found in the compared 717 brown hares from Europe. Overall, genetic diversity was highest in Anatolian hares, intermediate in brown hares from the southern and southeastern Balkans and lowest in central European populations. The rich genetic diversity in Anatolian hares might be a consequence of Anatolias biogeographic position with the chance of multiple gene flow from neighbouring regions, and the likelihood of long-term presence of hares during the last ice age, when large parts of more northern latitudes did not provide suitable habitats.However, among 28 loci used for the comparison between European and Anatolian populations, most common alleles of European brown hares were also common in Anatolian populations and no alternately fixed alleles were found for Anatolian and European populations. This together with only little or moderately varying allele frequencies produced low genetic divergence between Anatolian and European populations. Genetic differentiation among Anatolian populations was also low. Even between the two forms with different coat colour (brownish and yellowish) in Anatolian hares, there was little genetic differentiation. Altogether, all Anatolian hares studied presently are closely related to European brown hare populations, and only some distantly spaced population pairs revealed increased genetic divergence.

Zusammenfassung

Genetische Diversität anatolischer Feldhasen (Lepus europaeus Pallas, 1778) und Differenzierung zwischen anatolischen und europäischen PopulationenZur Beurteilung der phylogenetischen Beziehungen zwischen anatolischen Hasen und europäischen Feldhasenpopulationen wurde die allelische Variabilität anatolischer Hasen mittels horizontaler Stärkegelelektrophorese erfaßt und gemeinsam mit unmittelbar vergleichbaren Daten griechischer, bulgarischer und österreichischer Populationen aus früheren Studien populationsgenetischen Analysen unterzogen. Neunzehn der 48 untersuchten Loci der anatolischen Hasen zeigten allelische Variabilität. Unter den anatolischen Allelen kamen 14 bisher in den europäischen Polulationen nicht vor. Insgesamt zeigten anatolische Hasen die höchste und österreichische Populationen die niedrigste genetische Diversität; die jeweiligen Werte der griechischen und bulgarischen Populationen lagen dazwischen. Dies entspricht unserer Hypothese hoher genetischer Diversität in Anatolien, auf Grund der biogeografischen Position und der klimatischen bzw. Lebensraumbedingungen während des Pleistozäns, die, im Gegensatz zu Mitteleuropa, kontinuierliche Hasenpopulation in Anatolien wahrscheinlich erscheinen lassen. Kontinuierliche Populationen und Genflüsse aus verschiedenen Nachbarregionen könnten bei langfristig relative ungestörten Populationen zur Anreicherung genetischer Varianten in Anatolien geführt haben, während mitteleuropäische Feldhasenpopulationen im Zuge ihrer postglazialen Einwanderung aus Refugial-gebieten an genetischer Vielfalt eingebüßt haben. Allerdings waren die häufigen Allele der anatolischen Hasen ebenfalls häufig bei den europäischen Feldhasen vertreten; somit ergab sich insgesamt nur eine geringe genetische Differenzierung zwischen anatolischen und europäischen Feldhasen. Die zwei in Anatolien gefundenen Fellfärbungstypen (brauner vs. gelber Grundton) zeigten ebenfalls keine besondere genetische Differenzierung.