A narrow hybrid zone between two crayfish species from a Mexican cave

In Northern Chiapas (Mexico), two newly discovered species of Procambarus crayfish inhabit a subterranean stream. These species can be morphologically distinguished only by comparing extreme phenotypes (dark, thick-eyed, surface dwelling-like individuals vs light, elongated, microphtalmic, cave dwelling-like individuals). Individuals with intermediate phenotypes co-occur with those exhibiting extreme phenotypes. Crayfish were assayed electrophoretically and individual patterns at 23 gene loci were obtained. Unusually high levels of heterozygosity in both species and a clear discrimination between the two gene pools were revealed. The relationships between individuals were investigated by means of multivariate analysis on individual multilocus genotype profiles. Results showed the occurrence of individuals genetically intermediate between the two major clusters, which shared allozymic variants with both species. Due to the occurrence of alternative alleles in the two gene pools, we could quantify patterns of introgression, which revealed asymmetric gene flow between the two species. Moreover, differential levels of introgression in subsamples within the surface-like species were found: most introgressed individuals came from the inner section of the cave, where the two species were greatly mixed. These results are also discussed in reference to the morphometric results from a companion paper. A possible evolutionary pathway, leading to the situation in this cave, and possibly in neighbouring cave systems, is outlined. The hypothesis of a past history of allopatric divergence from a common ancestor and a subsequent secondary contact between these two Procambarus species is supported by geological studies. Crayfish sympatry and competitive exclusion are also discussed.     

Evolutionary and biogeographical patterns within Iberian populations of the genus Squalius inferred from molecular data

Origins and biogeography of the chub, genus Squalius (formerly within the genus Leuciscus), in the Iberian Peninsula were inferred from comparison between patterns of geographic distribution and phylogenetic relationships among populations belonging to 14 European Squalius species. The phylogeny recovered was based on the complete sequence of the mitochondrial cytochrome b gene. Squalius species were grouped into three major clades. The basal clade included species distributed across the ancient Paratethys Sea. The second clade included species from Central and East Europe and the northern areas of the Mediterranean basin towards Minor Asia. The third clade included species from the Mediterranean Peninsulas (Iberian, Italy, and Balkans). The Iberian Squalius species do not constitutes a monophyletic group. Our data indicate that the Iberian Peninsula was colonized at least twice by two different monophyletic lineages, a meridional group and a Central Europe group. The amount of species diversity found in the Iberian Peninsula and the phylogenetic relationships among these species, together with their geographic distribution, suggest that the Central Europe lineage colonized the Iberian Peninsula at a latter time. Our data indicate that the northeastern Iberian lineage is phylogenetically close to Greek populations of Squalius cephalus, while the second lineage formed a monophyletic group including Squalius pyrenaicus, Squalius carolitertii, Squalius aradensis, and Squalius torgalensis. The speciation process that generated these species and the geographic structure of their populations, principally in S. pyrenaicus, can be attributed to paleogeographical events like the ancient endorrheism and the development of hydrographic basins.     

Metapopulation pollination in the deceptive orchid Anacamptis pyramidalis

Pollen dispersal between local plant populations within a range of 6 km in a geographically defined metapopulation of the lepidopteran-pollinated deceptive orchid Anacamptis pyramidalis was studied on the island Öland in the Baltic Sea. Local A. pyramidalis populations were examined for pollinators, flowering individuals, and fruit set. Population sizes of pollen vector species were estimated using a mark–release–recapture technique. As pollen vectors, the burnet moth Zygaena minos and the butterfly Aporia crataegi dominated. 205 out of 745 marked lepidopterans were pollinarium carriers. The proportion carriers of the total was considerably higher in Z. minos (50.3%), than in A. crataegi (21.5%) and nymphalidae (8.2%). Furthermore, Z. minos moved much shorter distances than A. crataegi did, while no difference in potential pollen dispersal distances were found between males and females. The number of individual vectors recaptured in another local population of A. pyramidalis was low: A. crataegi (8) and Z. minos (1). The ratio of pollinaria transferred to another local A. pyramidalis population compared to pollinaria remaining within the same local population was 1:41. This study highlights that pollen dispersal distances vary between pollen vector species.     

Ecological, morphological and allozymic differentiation between diploid and tetraploid knapweeds (Centaurea jacea) from a contact zone in the Belgian Ardennes

In the northeastern part of Belgium, the Centaurea jacea complex shows extensive morphological variation and is represented by a diploid (2 n = 22) and a tetraploid (2 n = 44) cytotype. Polysomic inheritance of allozyme markers in the tetraploids, suggesting autopolyploidy, is here demonstrated for the first time. In order to test whether the two cytotypes occupy distinct habitats and possess different gene pools, patterns of allozymic and morphological variation were investigated in relation to ploidy level and site characteristics in 26 populations from the Belgian Ardennes. The two cytotypes showed a parapatric distribution, the diploids occurring at higher elevations (mostly above 500 m) than the tetraploids (mostly below 500 m). Three mixed populations were found near the contact zone of the two cytotypes. Within the mixed populations no triploid plant and no evidence for gene flow between cytotypes were found, despite widely overlapping flowering periods. The two cytotypes can be distinguished on the basis of morphological traits and enzymatic gene pools. The congruence of morphological and allozymic variation with chromosome numbers suggests a secondary contact between the two cytotypes with limited gene flow between them. The origin and persistence of the parapatric distribution are discussed.     

Colonization of Northern Europe by Zygaena filipendulae (Lepidoptera)

Northern and mountainous ice sheets have expanded and contracted many times due to ice ages. Consequently, temperate species have been confined to refugia during the glacial periods wherefrom they have recolonized warming northern habitats between ice ages. In this study, we compare the gene CYP405A2 between different populations of the common burnet moth Zygaena filipendulae from across the Western Palearctic region to illuminate the colonization history of this species. These data show two major clusters of Z. filipendulae populations possibly reflecting two different refugial populations during the last ice age. The two types of Z. filipendulae only co‐occur in Denmark, Sweden, and Scotland indicating that Northern Europe comprise the hybridization zone where individuals from two different refugia met after the last ice age. Bayesian phylogeographic and ecological clustering analyses show that one cluster probably derives from an Alpe Maritime refugium in Southern France with ancestral expansive tendencies to the British Isles in the west, touching Northern Europe up to Denmark and Sweden, and extending throughout Central Europe into the Balkans, the Peleponnes, and South East Europe. The second cluster encompasses East Anatolia as the source area, from where multiple independent dispersal events to Armenia, to the Alborz mountains in north‐western Iran, and to the Zagros mountains in western Iran are suggested. Consequently, the classical theory of refugia for European temperate species in the Iberian, Italian, and Balkan peninsulas does not fit with the data from Z. filipendulae populations, which instead support more Northerly, mountainous refugia.     

Phylogeography and conservation genetics of endangered European Margaritiferidae (Bivalvia: Unionoidea)

Margaritifera margaritifera and M. auricularia are among the most endangered freshwater mussels in the world, and the only species of the genus found in Europe. Our genetic study explores allozymic variability (27 loci) and differentiation at the mitochondrial sequence level (partial COI and 16S rRNA gene sequences). The Spanish M. auricularia population showed genetic parameters of variation that were of the same order as those of other freshwater molluscs (though at the lower end of the range), probably permitting its potential recovery. The difference between this species and M. margaritifera was clearly established (ten diagnostic allozymic loci, Nei = 0.462, and mean nucleotide divergence around 9.4%). The M. margaritifera populations analysed showed a certain degree of population genetic structure (according to allozyme data) that was not, however, related to a geographical cline. Nevertheless, two mitochondrial lineages (albeit very closely related) were identified: a northern lineage extending from Ireland to the Kola Peninsula including the western Atlantic coast, and a second cluster distributed from Ireland to the Iberian Peninsula. The phylogenetic relationships between these two species and other related taxa were established. The putative M. m. durrovensis could be considered an 'ecophenotype'. Palaeobiogeographical scenarios are presented and indicate unexpected 'recent' gene flow between M. margaritifera populations that were theoretically isolated in the early Tertiary.  © 2003 The Linnean Society of London, Biological Journal of the Linnean Society 2003, 78, 235–252.     

A note on chironomids (diptera) of temporary pools in the National Park of Circeo,Central Italy

Preliminary observations on chironomid assemblages in 9 temporary pools of the National Park of Circeo (Central Italy) are reported. A total of 15 genera or species groups (6 Orthocladiinae, 3 Tanypodinae, 1 Tanytarsini and 5 Chironomini) were recorded during March and April, 1986.Psectrotanypus varius, Polypedilum nubeculosum gr.,Chironomus thummi gr. andC. plumosus gr. were the most abundant and frequent taxa in the nine pools. Almost all chironomids collected are eurytopic and widely distributed in Europe, including Italian waters. Only the finding ofGymnometriocnemus is reported in this paper as a new record for Central Italy. Similarity among pools and among taxa (coefficient of Jaccard) shows a major occurrence of aquatic Orthocladiinae in smaller pools and of Chironomini in larger pools. This relationship between chironomid assemblages and pool sizes can be partly related to the duration of the wet phase which affects chironomid species according to their survival strategies.     

Diploid and Tetraploid Cytotypes of Centaurea stoebe (Asteraceae) in Central Europe: Morphological Differentiation and Cytotype Distribution Patterns

The taxomically critical species Centaurea stoebe is represented in Central Europe by a diploid (2n?=?18) and a tetraploid (2n?=?36) cytotype. Their morphological differentiation and taxonomic treatment is still controversial. Karyological (chromosome numbers and flow cytometric measurements) and multivariate morphometric analyses were used here to address cytotype distribution patterns and their morphological differentiation. Material from 38 localities (771 individuals) in Slovakia, Hungary and Austria, including type localities of the names traditionally applied to the different cytotypes, was sampled and evaluated using both morphometric and karyological approaches. The morphological tendency towards cytotype differentiation is evident only at a population level, and is blurred at the level of individual plants. Diploid populations prevail in the area studied, as well as throughout Europe; mixed-cytotype populations were also found. The present data, namely the weak morphological distinction, largely sympatric occurrence of the cytotypes, and the existence of mixed-cytotype populations, favour taxonomic treatment as a single species, without recognition of infraspecific units.     

Morphological and allozymic variation in Hynobius boulengeri and H. stejnegeri (Amphibia: Urodela: Hynobiidae)

We studied morphological and allozymic variation in populations of Japanese salamanders, Hynobius boulengeri and H. stejnegeri. Adult H. boulengeri showed sexual dimorphism, and juveniles differed greatly from adults in many morphological characters. From the results of multivariate analyses of morphological characters, the populations were divided into four groups: (I) H. boulengeri from Honshu, (II) H. boulengeri from Shikoku, (III) H. boulengeri from the Sobo-Katamuki Mountains of Kyushu and H. stejnegeri, and (IV) H. boulengeri from the Amakusa Islands and the Osumi Peninsula. Phenotypic relationships among the four groups were identical to relationships clarified by allozymic analyses, except for group IV, which was included in group III in the allozyme tree. Some morphometric characters were significantly correlated with environmental variables. We consider H. stejnegeri to be a valid species based on its unique color pattern, morphometric characters, and allelic composition, even though it was nested within group III of H. boulengeri by both morphological and allozymic analyses. We propose that group I from Honshu and group II from Shikoku should be treated as H. boulengeri sensu stricto and H. hirosei, respectively. Resolving the taxonomic status of the remaining populations of groups III and IV from Kyushu requires further study.     

Geographical variation in wing polymorphism of the waterstrider Aquarius najas (Heteroptera,Gerridae)

The waterstrider Aquarius najas is wingless in Northern Europe, while winged individuals occur frequently in Central and Southern Europe. To test if the latitudinal difference is genetically controlled, we collected mature individuals from 10 different populations and raised their offspring in ‘common garden’ laboratory conditions. Half of these populations were from southern and the other half from central Finland. Daylength and temperature do influence wing development among other species of waterstriders, and thus we maintained a similar short daylength and warm conditions for all populations. These conditions should be favourable for wing development in general. Among laboratory-bred individuals several winged individuals appeared, and their proportion varied between populations. The relative frequency of winged individuals was highest in the southern populations. Thus, apart from phenotypic plasticity there seems to be some genetic control over the occurrence of wings, and the latitudinal trend coincides with the direction in natural populations over a larger European scale. Overwinter survival in our laboratory conditions was higher among the wingless individuals. The survival cost may explain why the proportion of winged individuals was lower in the northern populations with more extreme overwintering conditions than in the southern ones.     

Pinus mugo (Pinaceae) in the Abruzzi Mountains: high morphological variation in isolated populations

Thirty-three individuals of Pinus mugo from the Abruzzian Apennines (Italy), the most isolated population of the species, were examined biometrically to verify the range of trait variation and compared with six other populations sampled in the north-eastern part of the species range. Ten needles and ten cones from each individual were measured and quantified on the basis of 31 traits. The interpopulation comparison revealed a close affinity of the Abruzzian population of P. mugo to three of the six other populations. The other three north-eastern populations, one from the Tatra Mountains and two from the Chornokhora Mountains, appeared to be different. This suggests that these populations originated from other Pleistocene centres of occurrence. The intrapopulation variation of the population from the Abruzzian Apennines is comparable with those of all six populations of P. mugo from the Sudety and Carpathian Mountains. These findings support the hypothesis that isolated but locally abundant plant populations do not necessarily show a reduction in variability resulting from isolation.  © 2005 The Linnean Society of London, Botanical Journal of the Linnean Society , 2005, 147 , 309–316.     

Morphological clines across a karyotypic zone of house mice in Central Italy

Multivariate morphometric differentiation between chromosomal races of the mouse Mus domesticus in Central Italy was investigated using a population of 2n = 22 “CB” karyotype, three populations of standard 2n = 40 karyotype, five populations of 2n = 22 “CD” karyotype and three populations from the hybrid zone between the latter two karyotypes. Whilst populations of different karyotype generally have significantly different morphometry, canonical analysis does not reveal that the populations ordinate into distinct aggregations based on karyotype, largely because the 2n = 22 “CD” populations are so diverse. Nevertheless, canonical analysis does reveal a significant cline in morphology across the contact zone between the 2n = 40 and 2n = 22 “CD” mice. The nature of this transition, i. e. a cline 1. within the 2n = 40 range, 2. within the hybrid range (but unrelated to chromosome number) and 3. within the 2n = 22 “CD” range, tends to indicate that the morphometric divergence is due to adaptation to the different ecological regimes across which these mice are distributed rather than the phylogenetic divergence of the karyotypic races.     

Morphometric and allozyme variation in central and southern Greek populations ofMicrotus (Terricola) thomasi

The endemic Balkan vole taxonMicrotus (Terricola) thomasi (Barrett-Hamilton, 1903) exhibits great karyological variability in Greece. In this study, populations belonging to two different karyotypic forms (‘atticus’ and ‘thomasi’) are examined both morphometrically and electrophoretically. A total of 140 individuals ofM. (T.) thomasi were collected from 6 localities of south and central Greece. For the morphometric analysis, 27 variables (external body and cranial characters) were examined and evaluated according to multivariate analyses (PCA, MANOVA, CVA and CLUS). For the electrophoretic analysis, 18 putative genetic loci were examined and the allozymic data were treated by the biostatistical package BIOSYS-1. According to the results obtained, all the populations studied show little overall morphometric variability, whereas they are characterized by high electrophoretic variability. The populations studied are not grouped according to the karyotypic form. In almost all the cases, in the two UPGMA-dendrograms plotted according to morphometric and electrophoretic distances (Mahalanobis’ and Nei’s distances, respectively), the populations branching together belong to different karyotypic forms. Conclusively, the morphometric and electrophoretic results of this study revealed that the two karyotypic forms should not be considered separate species or subspecies, as it has been proposed by some authors in the past, and the populations studied can be considered only as different local populations of the rather variable vole speciesM. (T.) thomasi.     

Allozyme analysis of Trichinella isolates from various host species and geographical regions.

Allozyme analysis was carried out on 152 Trichinella isolates from synanthropic and wild animals and from humans; the isolates were collected from 5 continents. The analysis, involving 27 enzymes, revealed the presence of 8 distinct gene pools, termed T1-T8. Four of the genetic groups represent the 4 previously proposed species: Trichinella spiralis sensu stricto (T1), Trichinella nativa (T2), Trichinella nelsoni (T7), and Trichinella pseudospiralis (T4). The other 4, T3, T5, T6, and T8 are distinct from previously described species. The absence of allozymic hybrid patterns among even sympatric groups indicates a lack of gene flow among the groups. Principal component analysis and the unweighted pair group method of analysis were used to assemble allozyme patterns of the 152 isolates into discrete groups and to show their relative relationships. Both analyses indicated the presence of 8 primary clusters that correlated with the gene pools revealed by direct allozyme profile analysis. The absence of evidence of gene flow among the gene pools and the high level of allozymic differentiation between the cluster groups support the concept that the genus Trichinella is composed of several sibling species.     

Morphometric analysis of interspecific and microgeographic variation of crayfish from a Mexican cave

Two undescribed species of Procambarus crayfish (Crustacea, Decapoda) inhabit the subterranean stream running in the recently explored Cueva de Los Camarones, Chiapas, Mexico. Individuals ranging from dark, robust, large-eyed forms with an epigean mien to pale, comparatively slender, microphthalmic ones resembling related troglobites were found, but only the extremes could be distinguished visually. Evolutionary relationships among individuals were investigated morphometrically by means of multivariate analysis'on 14 body dimensions. These analyses revealed a significant discontinuity between the two species, mainly determined by the shape of the rostrum, chelae and telson. The occurrence of a few aberrant phenotypes may indicate the existence of some gene flow between the two species.
Microgeographic variation was demonstrated within the less cavernicolous species. The characters involved were the same as those that discriminate between the two species. These results are compared with allozymic data presented in a parallel paper and the role of genie introgression and environmental induction are discussed.     

Genetic variability and differentiation in Leuciscus (Telestes) souffia. Taxonomic and conservation inferences

Genetic variability of eight populations of Leuciscus (Telestes) souffia from France, Italy and Slovenia, and one of Leuciscus (Squalius) carolitertii from Spain was analysed using allozymic electrophoresis. For L (Telestes) souffia the parameters of variability ranged from P = 1 8.2 to 48.5 % and H_o = 0.023 to 0.092, values included in the global averages found for freshwater fishes. The phylogenetic relationships among these populations lead us to consider the souffia species belonging to the genus Telestes, and to subdivide it into two species: souffia (with at least two subspecies: souffia in France and agassizi in Slovenia), and muticellus. The genetic structure found within the French populations showed a certain differentiation between the Hérault Basin and the Var/Argens populations, recommending the preservation of these entities that could represent different adapted genetic pools. In the same way, the Italian region should be studied in detail, since the populations analysed from Bevera and Staffora Rivers (T. muticellus) showed evidence of a break of genetic flow (including a diagnostic locus between them).     

Distribution and life cycles of cladocerans in temporary pools from Central Italy

SUMMARY 1. A study of twenty-nine temporary pools located in a nature reserve in Central Italy was carried out for two successive years (1982–83 and 1983–84) to determine the distribution and life cycles of the cladocerans they contained.
2. The pools followed a typical autumnal cycle, filling up in early autumn and drying up in late spring. They differed in morphology (size, depth, type of soil) and in the length of the wet phase.
3. Nineteen cladoceran species were found. A few of these are early species, appearing at the beginning of the wet phase, whereas others appear much later in spring and these populations persist for a shorter time.
4. Species distributions did not show any clear correlation with the ponds'morphological features, and the life cycles of individual species were similar in all ponds. Thus populations did not adapt life history patterns to match conditions in particular ponds.     

Postglacial colonization of Europe by the barbastelle bat: agreement between molecular data and past predictive modelling

The barbastelle (Barbastella barbastellus) is a rare forest bat with a wide distribution in Europe. Here, we combine results from the analysis of two mtDNA fragments with species distribution modelling to determine glacial refugia and postglacial colonization routes. We also investigated whether niche conservatism occurs in this species. Glacial refugia were identified in the three southern European peninsulas: Iberia, Italy and the Balkans. These latter two refugia played a major role in the postglacial colonization process, with their populations expanding to England and central Europe, respectively. Palaeo‐distribution models predicted that suitable climatic conditions existed in the inferred refugia during the last glacial maximum (LGM). Nevertheless, the overlap between the current and the LGM distributions was almost inexistent in Italy and in the Balkans, meaning that B. barbastellus populations were forced to shift range between glacial and interglacial periods, a process that probably caused some local extinctions. In contrast, Iberian populations showed a ‘refugia within refugium’ pattern, with two unconnected areas containing stable populations (populations that subsisted during both glacial and interglacial phases). Moreover, the match between LGM models and the refugial areas determined by molecular analysis supported the hypothesis of niche conservatism in B. barbastellus. We argue that geographic patterns of genetic structuring, altogether with the modelling results, indicate the existence of four management units for conservation: Morocco, Iberia, Italy and UK, and Balkans and central Europe. In addition, all countries sampled possessed unique gene pools, thus stressing the need for the conservation of local populations.     

Mitochondrial sequence analysis of Salamandra taxa suggests old splits of major lineages and postglacial recolonizations of central Europe from distinct source populations of Salamandra salamandra

Representatives of the genus Salamandra occur in Europe, Northern Africa and the Near East. Many local variants are known but species and subspecies status of these is still a matter of dispute. We have analysed samples from locations covering the whole expansion range of Salamandra by sequence analysis of mitochondrial D-loop regions. In addition, we have calibrated the rate of divergence of the D-loop on the basis of geologically dated splits of the closely related genus Euproctus. Phylogenetic analysis of the sequences suggests that six major monophyletic groups exist (S. salamandra, S. algira, S. infraimmaculata, S. corsica, S. atra and S. lanzai) which have split between 5 and 13 million years ago (Ma). We find that each of the Salamandra species occupies a distinct geographical area, with the exception of S. salamandra. This species occurs all over Europe from Spain to Greece, suggesting that it was the only species that has recolonized Central Europe after the last glaciation. The occurrence of specific east and west European haplotypes, as well as allozyme alleles in the S. salamandra populations suggests that this recolonization has started from at least two source populations, possibly originating in the Iberian peninsula and the Balkans. Two subpopulations of S. salamandra were found that are genetically very distinct from the other populations. One lives in northern Spain (S. s. bernardezi) and one in southern Italy (S. s. gigliolii). Surprisingly, the mitochondrial lineages of these subpopulations group closer together than the remainder S. salamandra lineages. We suggest that these populations are remnants of a large homogeneous population that had colonized Central Europe in a previous interglacial period, approximately 500 000 years ago. Animals from these populations were apparently not successful in later recolonizations. Still, they have maintained their separate genetic identity in their areas, although they are not separated by geographical barriers from very closely related neighbouring populations.     

Genetic variation of isolated and peripheral populations of Pinus sylvestris (L.) from glacial refugia

Analysis of variation at 10 polymorphic isoenzyme loci in 51 European populations of Pinus sylvestris both from its main continuous range and from isolated, marginal populations out of the continuous range, revealed differences in genetic structure. The highest genetic richness, measured as mean number of alleles per locus, was recorded for populations from the Iberian Peninsula, followed by those from Scotland and the Balkans. Marginal, isolated populations were characterized by much greater interpopulation variation than populations from the continuous range of the species. The highest mean observed heterozygosity was recorded for the Iberian populations. The highest gene flow was observed among populations in the continuous range of the species. The populations could be classified into five groups based on genetic similarities. Populations from the continuous range formed one group, those from Spain two groups, pines from Scotland one group and those from the Balkans one group. One population from Scotland was found to be most distinct from the other Scottish pines and was found to be grouped with the Balkan populations. The occurrence and frequencies of rare alleles in the populations showed a characteristic pattern, suggesting that populations from the Iberian Peninsula probably did not participate in the colonization of Europe by Pinus sylvestris after the last glaciation. In contrast, the migration of populations from the Balkan refugium into Central and Western Europe had a significant effect on the contemporary gene pools of populations of Pinus sylvestris in its continuous range.