Phylogenetic relationships among Spanish Barbus species (Pisces,Cyprinidae) shown by RAPD markers

Random amplified polymorphic DNA (RAPD) markers were used to estimate the population structure and phylogenetic relationships among the eight species of the genus Barbus that inhabit the Iberian Peninsula. Ten random oligodecamers were used to amplify DNA from 232 fish sampled from 15 populations. A total of 270 markers were detected that revealed low levels of genetic variability. The conclusions of cluster analysis indicate two main branches and three well-differentiated groups: north-eastern, Mediterranean and Atlantic. This clustering mainly reflects the evolutionary history of the genus, which is closely related to the paleogeography of the Iberian Peninsula. The contribution and application of these results to the conservation of the species, to their taxonomic status and to the process of colonization of the Iberian Peninsula by the genus Barbus are discussed.     

Haplotype diversity and phylogenetic relationships among the Iberian barbels (Barbus,Cyprinidae) reveal two evolutionary lineages

The phylogenetic relationships and haplotype diversity of all Iberian barbels were examined by analyzing the complete mitochondrial cytochrome b gene sequence (1141 bp) of 72 specimens from 59 Iberian localities. Phylogenetic findings demonstrated a clear distinction between two mitochondrial lineages and confirmed the existence of two previously considered subgenera: Barbus and Luciobarbus: The first subgenus, Barbus, is represented on the Iberian Peninsula by Barbus haasi and Barbus meridionalis. The second subgenus, Luciobarbus, includes the remaining endemic Iberian species: Barbus comizo, Barbus bocagei, Barbus microcephalus, Barbus sclateri, Barbus guiraonis, and Barbus graellsii. Mean haplotype divergence between these subgenera was 10.40%, providing evidence of a clear subdivision within the Iberian barbels. Our results conflict with those reported in a recent study, based on 307 cytochrome b base pairs, that failed to identify any division within the genus Barbus in the Iberian Peninsula. The inclusion of nine further species belonging to this genus (used as outgroups) allowed us to establish a closer relationship of the Iberian species of the subgenus Barbus with other European taxa than with the Iberian Luciobarbus, which was found to cluster with North African, Caucasian, and Greek species. At the population level, no biogeographic structure was shown by specimens of each species (only 5.98% of the variation was attributable to differences among populations of each species). Given the discrete amount of divergence found among the Luciobarbus species, the formation of current hydrographic basins during the Plio-Pleistocene seems to have played a major role in their isolation and evolution.     

Biogeographic hypothesis explaining the diversity of the genus Amphimallon Berthold, 1827, in the Mediterranean Basin (Coleoptera,Scarabaeidae, Melolonthinae,Rhizotrogini)

In this paper, the analysis of the current distribution of species, or species-groups, of the Western–Palaearctic genus Amphimallon Berthold, 1827, based on a phylogenetic hypothesis recently provided and relevant paleogeographical events that occurred in the Mediterranean Basin from Late Paleogene (35 Myr BP) to the present, supports a biogeographical scenario to explain species diversity within this genus. The center of origin of this genus is hypothesized to be in the Iberian Peninsula. Microplate fragments drifting from the Iberian Peninsula, successive landmass suture events between the Eastern and Western Mediterranean Basins, separation of Tethys and Paratethys Oceans, re-establishment of marine connections, uplift of the Alps, and drying out of the Mediterranean are hypothesized as the principal events allowing dispersal of populations throughout Mediterranean Basin and biogeographical vicariances within the genus Amphimallon.     

Phylogeny and evolution of the genus Barbus in the Iberian Peninsula as revealed by allozyme electrophoresis

Variation of 14 enzyme systems encoded by 31 presumptive loci in different barbel species was studied using starch gel electrophoresis. Eighteen samples representing 11 Barbus tetraploid taxa were taken, including 10 populations from the Iberian Peninsula, six from other parts of western and southern Europe, one from northern Africa and one diploid species as outgroup from Asia Minor. The genetic analysis reassessed of the taxonomic status of the Iberian barbels into two distinct species groups. The first group included B. bocagei, B. comiza, B. graellsii, B. gulraonis, B. microcephalus , and B. sclateri , that aligned with B. callensis from northern Africa and with B. apoensis from Asia Minor; the other group included B. haasi and B. merldlonahs that was related to the European species, B. barbus, B. plebejus and B. peloponnesius . These groups are probably not monophyletic. It is suggested that the isolation of the Iberian Peninsula from Europe since the Oligocene-Miocene may explain the genetic affinities of the Iberian barbels with those of North African rather than with the European group.     

Rapid radiation of the Mediterranean Luciobarbus species (Cyprinidae) after the Messinian salinity crisis of the Mediterranean Sea, inferred from mitochondrial phylogenetic analysis

Phylogenetic relationships of 64 freshwater Barbus s.s. species distributed in basins around the Mediterranean Sea were assessed using cytochrome b sequences. Our results are in concordance with previous morphological and genetic studies, which proposed that these species belong to two major lineages (or subgenera): Barbus and Luciobarbus . We were particularly interested in phylogenetic relationships among species of the Luciobarbus lineage that are primarily found in the southern Mediterranean region from the Iberian Peninsula to the Middle East. In the Luciobarbus lineage, species that were previously attributed to the Capoeta genus were clustered. In this study, we observed short internodes between monophyletic groups having a geographical agreement around the Mediterranean. However, groups from the opposite sides of the Mediterranean Sea (Iberian Peninsula– Capoeta , north-western Africa–Middle East) seem to be phylogenetically close. We therefore infer that rapid radiation of Luciobarbus species in the Late Miocene better fits our data rather than gradual founder events in the southern Mediterranean. We propose that the biogeographical event along an east–west route, responsible for the present distribution of Luciobarbus species, was the 'Lago Mare' phase of the Mediterranean Sea that provided a rapid dispersal route over extensive distances. This provides new insights into the speciation pattern of this group, and may be of general use in the study of freshwater species in these regions.  © 2003 The Linnean Society of London, Biological Journal of the Linnean Society , 2003, 80 , 207–222.     

Karyosystematic study of Potentilla L. subgen. Potentilla (Rosaceae) in the Iberian Peninsula

Chromosome numbers in 80 populations belonging to 18 species of Potentilla L. subgen. Potentilla from the Iberian Peninsula and two of P. maura, a North African endemic taxon, have been counted. The basic number of chromosomes is always x = 7 and these chromosomes are small (between 1 and 2 μm). For three species, the number of chromosomes is reported for the first time and, for another six, this number has been established in Iberian representatives. Moreover, new ploidy levels have been obtained for P. hispanica and P. crantzii with regard to their entire distribution area, and in P. cinerea and P. neumanniana for the Iberian Peninsula. Some taxonomic, phylogenetic and phytogeographic comments are made for several species or groups of species from the West Mediterranean region. In 13 species only one ploidy level has been found, but six species have several ploidy levels. Seven ploidy levels occur in the investigated taxa. The frequency of each ploidy level represented within Iberian Potentilla is analysed and the data are compared with those available for taxa from the rest of the distribution area of the genus.     

The genus Mantisalca (Asteraceae) in southwest Europe

The diversity of the genus Mantisalca in southwest Europe is the subject of much controversy. The present taxonomic revision of the genus on the Iberian Peninsula allows four species to be recognized, whose morphological variability is described and various synonyms are typified. A new species is described, M. cabezudoi Ruiz de Clavijo and Devesa, endemic to southeast Spain. The name Microlonchus spinulosus Rouy is reinstated for a species endemic to the central and east Iberian Peninsula, for which a new combination is made: Mantisalca spinulosa (Rouy) Ruiz de Clavijo and Devesa. A diagnostic key to the taxa is given.     

Molecular identification (RAPD) of the eight species of the genus Barbus (Cyprinidae) in the Iberian Peninsula

Random amplified polymorphic DNA (RAPD) analysis has been used to identify the eight Barbus species of the Iberian Peninsula, whose morphological differentiation is difficult. Ten random primers were employed to generate RAPD markers. Different RAPD profiles were observed for the different species. Four species-specific markers were found in B. bocagei , seven in B. comizo , five in B. graellsii , three in B. guiraonis , eight in B. haasi , 13 in B. meridionalis , four in B. microcephalus and four in B. sclateri. Evidence is presented that RAPD markers constitute useful tools for accurate taxonomic identification of Spanish barbels which is one of the first prerequisites in effective conservation programmes.     

Complex patterns of environmental niche evolution in Iberian columbines (genus Aquilegia,Ranunculaceae)

Aims This study explores the patterns of niche differentiation in a group of seven closely related columbines (genus Aquilegia, Ranunculaceae) from the Iberian Peninsula. Populations of these columbines are subject to complex patterns of divergent selection across environments, which partly explain the taxonomic structure of the group. This suggests the hypothesis that niche divergence must have occurred along the process of diversification of the group.Methods We used MaxEnt to build environmental niche models of seven subspecies belonging to the three species of Aquilegia present in the Iberian Peninsula. From these models, we compared the environmental niches through two different approaches: ENMtools and multivariate methods.Important findings MaxEnt distributions conformed closely to the actual distribution of the study taxa. ENMtools methods failed to uncover any clear patterns of niche differentiation or conservatism in Iberian columbines. Multivariate analyses indicate the existence of differentiation along altitudinal gradients and along a gradient of climatic conditions determined by the summer precipitation and temperatures. However, climatic conditions related to winter temperature and precipitation, as well as soil properties, were equally likely to show conservatism or divergence. The complex patterns of niche evolution we found suggest that Iberian Columbines have not been significantly constrained by forces of niche conservatism, so they could respond adaptively to the fast and profound climate changes in the Iberian Peninsula through the glacial cycles of the Pleistocene.     

Recent radiation of Iberian barbel fish (Teleostei, Cyprinidae) inferred from cytochrome b genes

Nucleotide sequence fragments of the mitochondrial DNA gene encoding cytochrome b were examined in 26 individuals belonging to the seven species of Barbus endemic to the Iberian Peninsula: Barbus haasi, B. bocagei, B. graellsii, B. sclateri, B. comiza, B. guiraonis, and B. microcephalus. Six of the seven currently recognized species can be distinguished on the basis of their cytochrome b nucleotide sequences. Sequence divergence estimates for Spanish species of Barbus (0-6.5%) are, in general, low in comparison to those reported for other fish species, and hybrid individuals were found. All of these observations suggest a recent radiation. The inferred phylogenetic tree has two main clades, one including B. graellsii, B. guiraonis, and B. microcephalus, and the other the remaining species groups.     

Epidermal micromorphology of the genus Festuca L. (Poaceae) in the Iberian Peninsula

Micromorphological features of the leaf-blade, lemma, and palea were examined under scanning electron microscopy (SEM) for 64 species of the genus Festuca L. occurring in the Iberian Peninsula. The objective of the study was to survey these characters for the first time in the whole of the genus Festuca in this region, to assess their taxonomic value, and to compare the variation with recent DNA sequence-based phylogenies. The principal features include the frequency, shape, and distribution of silica bodies, short cells, crown cells, and prickles, and the morphology of long cells. The abaxial lemma surface shows the most important taxonomical and phylogenetic characters. The variation observed was consistent with the subgenera and sections identified in molecular studies.     

Profiling invasive fish species: the importance of phylogeny and human use

Understanding the ecological differences between native and invasive species is of considerable scientific and practical interest. We examined such differences between native and invasive inland fish species from the Iberian Peninsula in order to analyse the importance of phylogenetic correction and variability (in addition to central tendency). We collected 26 quantitative and qualitative variables on the ecology, life‐history traits and human use of the 69 inland fish species of the Iberian Peninsula, including native, invasive and migratory species. The taxonomic distribution of invasive fish species deviated significantly from world freshwater richness and in contrast to native species, invasive fish belongs to only five taxonomic orders but to a wide spectrum of families not native to the Iberian Peninsula. Because the life‐history traits were highly dependent on taxonomy, the results, with or without applying phylogenetic methods, differed and after accounting for phylogeny, invasive species displayed higher and wider latitude in general and a different reproductive season mainly among salmonids and cyprinids. Human use was also significantly different between native and invasive fish species and produced more variability in life‐history traits of invasive species and uneven taxonomic distribution because of the high diversity of species introduced. We show that accounting for taxonomy and studying variability in addition to central tendency is important in the comparison of life‐history traits between native and invasive species.     

Determinant variables of Iberian Peninsula Aphodiinae diversity (Coleoptera, Scarabaeoidea, Aphodiidae)

Aim The aims of this paper are to examine diversity–variability patterns for species of Aphodiinae (Coleoptera, Scarabaeoidea, Aphodiidae) on the Iberian Peninsula, and to determine the factors that influence their geographic distribution. Location Iberian Peninsula (Spain and Portugal). Methods Data from 30 studies and their bibliographies on species of Iberian Peninsula Aphodiinae were compiled. The reliability of the inventories was evaluated using parametric species richness estimators. In addition, a further 11 variables related to rarity, geographic distribution, or phylogenetic diversity were considered. Diversity variables were analysed using principal components analysis to reduce the number of dependent variables. Subsequently, the effect of differences in locality size among the 30 studies was eliminated by calculating and retaining the residuals of the curvilinear relationship of each diversity variable with the area. Generalized linear models were used to examine the relationships between diversity and 17 environmental variables. The diversity variables and their residuals were also subject to trend surface analysis in order to identify the relevance of spatially structured variables that had not been considered. The contribution of explanatory variables was determined through hierarchical variance analysis. Results Principal components analysis of biodiversity variables revealed that most of the variability could be explained using three biodiversity indexes: BI1, correlated positively with species richness, widely distributed species, frequent species, abundant species, species occurring in North Africa, Europe and the Iberian Peninsula, and phylogenetic diversity; BI2, correlated positively with numbers of infrequent and African–Iberian species; and, BI3, correlated positively with numbers of endemic, non‐abundant, European, and Iberian‐restricted species. A latitudinal disjunction emerged in BI1, with maximum scores at the north‐western and southern corners, while maximum BI2 scores were found throughout the south, and maximum BI3 scores in the north‐west. For BI1, it was climate that had the greatest influence, followed by lithology, and livestock presence. Geographic variables were the most significant for BI2, followed by climate and livestock presence. Finally, for BI3, climate variables were the most important, while geography, lithology and livestock presence had some relevance. Main conclusions The relevance of geographic variables indicates that other unaccounted‐for factors that are spatially structured could possibly explain additional variation in Aphodiinae diversity. These factors may be historic in nature, relating to the species groups, namely the Ibero‐European and the Mediterranean or Afro‐Iberian. The northern pattern could reflect the fact that the Iberian Peninsula acted as a colonization route and as a refuge during the glacial/interglacial cycles, while the southern pattern could be a consequence of the connection between the Iberian Peninsula and North Africa during the Messinian crisis, and/or a historic relationship in common, related to human activity.     

Biometrical analysis reveals major differences between the two subspecies of the European rabbit

The climatic oscillations that have occurred in the last few million years have strongly affected species distribution ranges. Highly divergent genetic lineages arose, some of which correspond to recognized subspecies that currently occupy small geographical areas. Understanding the implications of the genetic differences between these subspecies is crucial for proper conservation of Evolutionarily Significant Units. We use the two European rabbit subspecies, Oryctolagus cuniculus cuniculus and O. c. algirus, in the Iberian Peninsula as a model to investigate the repercussions at the biometric level of their largely recognized genetic differentiation. To accomplish this we analysed the ear and hind foot length, and the body mass of 999 adult rabbits from 27 locations across the distribution range of both subspecies in their native range, the Iberian Peninsula. Our results show biometric differences between the two subspecies, also explained by geographical location and sex, O. c. algirus being lighter and having shorter ear and hind foot lengths. We examine these findings under an evolutionary framework, and discuss their implications for current conservation efforts. Future research should focus on the ecological implications of these biometric differences, namely potential different habitat use and anti‐predatory strategies in the species' native range.     

Morphology, biometry and taxonomic position of two peritrich ciliate Protozoa found in activated sludge

The morphology of two sessile peritrich species taken from a plant for the processing of activated sludge is described. One of these species, having a non-contractile, branched peduncle, belongs to the genus, Heteropolaria, and the other having a single, contractile stalk, is a member of the genus Vorticella. A statistical and biometric study is made of their morphological characteristics, and the taxonomic position of both ciliates is discussed.     

Comparative phylogeography of endemic cyprinids in the south-west Iberian Peninsula: evidence for a new ichthyogeographic area

The comparative phylogeography and evolutionary history of three native cyprinid genera, Squalius (subfamily Leuciscinae), Chondrostoma (subfamily Leuciscinae) and Barbus (subgenus Luciobarbus ; subfamily Cyprininae), were examined focusing mainly in the South-Western region of the Iberian Peninsula, where recently described endemic species are present with considerably restricted distribution areas. In order to accomplish that the variation at the complete cytochrome b gene (1140 bp) was analysed for specimens from the South-Western region, and also for representatives of the three genera from all over the Iberian Peninsula. Data indicate different evolutionary histories, with distinct time periods of colonization between the two cyprinid subfamilies in the Iberian Peninsula. Four new Iberian ichthyogeographic areas are accordingly proposed based on congruent phylogeographic and geological evidences: the South-Western, the South-Eastern, the Atlantic and the Mediterranean. Evidence was provided for the older isolation of the South-Western area in the Miocene during the Endorheic Drainages phase, designating a clearly defined and distinct ichthyogeographic area. A new molecular clock calibration is proposed for the subgenus Luciobarbus .     

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.     

Phylogenetic relationships of Iberian cyprinids: systematic and biogeographical implications.

The phylogenetic relationships among all Iberian endemic cyprinids were inferred using the complete nucleotide sequence of the cytochrome b gene. The inferred molecular phylogeny included representatives from Central European, Asian and North African species, and is highly congruent with previous phylogenies based on osteological characters. Iberian cyprinids were grouped into only five, very speciose lineages (with the exception of the monotypic Anaecypris): Barbus, Luciobarbus, Chondrostoma, Leuciscus and Anaecypris. The existence of such a relatively small number of Iberian cyprinid lineages can be explained by the historical isolation of the Iberian Peninsula. North African and Asian barbels are the sister group of Iberian Luciobarbus, supporting a south-eastern route of colonization of the Iberian Peninsula for this subgenus. With leuciscins, Anaecypris hispanica was considered a relict species as it could not be related to any other Iberian cyprinid. The phylogenetic relationships among the main lineages of Iberian cyprinids based on cytochrome b sequence data supported the traditional division of the Cyprinidae into two subfamilies: Cyprininae and Leuciscinae.     

Understanding the biogeography of a group of earthworms in the Mediterranean basin--the phylogenetic puzzle of Hormogastridae (Clitellata: Oligochaeta)

Traditional earthworm taxonomy is hindered due to their anatomical simplicity and the plasticity of the characteristics often used for diagnosing species. Making phylogenetic inferences based on these characters is more than difficult. In this study we use molecular tools to unravel the phylogeny of the clitellate family Hormogastridae. The family includes species of large to mid-sized earthworms distributed almost exclusively in the western Mediterranean region where they play an important ecological role. We analyzed individuals from 46 locations spanning the Iberian Peninsula to Corsica and Sardinia, representing the four described genera in the family and 20 species. Molecular markers include mitochondrial regions of the cytochrome c oxidase subunit I gene (COI), 16S rRNA and tRNAs for Leu, Ala, and Ser, two nuclear ribosomal genes (nearly complete 18S rRNA and a fragment of 28S rRNA) and two nuclear protein-encoding genes (histones H3 and H4). Analyses of the data using different approaches corroborates monophyly of Hormogastridae, but the genus Hormogaster is paraphyletic and Hormogaster pretiosa appears polyphyletic, stressing the need for taxonomic revisionary work in the family. The genus Vignysa could represent an early offshoot in the family, although the relationships with other genera are uncertain. The genus Hemigastrodrilus is related to the Hormogaster elisae complex and both are found in the Atlantic drainage of the Iberian Peninsula and France. From a biogeographic perspective Corsica and Sardinia include members of two separate hormogastrid lineages. The species located in Corsica and Northern Sardinia are related to Vignysa, whereas Hormogaster pretiosa pretiosa, from Southern Sardinia, is closely related to the Hormogaster species from the NE Iberian Peninsula. A molecular dating of the tree using the separation of the Sardinian microplate as a calibration point (at 33 MY) and assuming a model of vicariance indicates that the diversification of Hormogastridae may be ancient, ranging from 97 to 67 Ma.     

Phylogenetic relationships among the Ibero-African cobitids (Cobitis, cobitidae) based on cytochrome b sequence data

We estimated the phylogenetic relationships of all Ibero-African spined loaches of the genus Cobitis using the complete mitochondrial cytochrome b gene (1140bp). We analysed 93 individuals of seven cobitid species found in all the principal drainages of the Iberian Peninsula and North Africa. A molecular phylogeny was used to revise current systematics of the Ibero-African Cobitis species and to infer a biogeographical model for Cobitis in the Western Mediterranean area during the Cenozoic period. Phylogenetic analysis provided support for the monophyly of two mtDNA clades: Clade A or Italian Clade with the Italian species (C. bilineata, C. zanandreai), and Clade B or the Ibero-African Clade. The Ibero-African Clade included all species endemic for the Iberian Peninsula (C. vettonica, C. calderoni, and C. paludica) and North Africa (C. maroccana). The species C. paludica does not constitute a natural group, and could be divided into at least four monophyletic mtDNA lineages with moderate to high bootstrap values and posterior probability support. Phylogenetic relationships of the Ibero-African species were not resolved satisfactorily, but in all analyses C. calderoni from Northern Iberian Peninsula was basal. We have re-calibrated a molecular clock for the genus Cobitis (0.68% per million year by pairwise) using populations inhabiting both sides of the Gibraltar Strait. Application of this Cobitis mtDNA clock provides evidence that the Messinian salinity crisis played a primary role in the diversification of some Ibero-African cobitid species. The basal polytomies of the Ibero-African Clade might suggest that all mtDNA lineages diversified rapidly.