Integrative taxonomy in two free-living nematode species complexes

Integrative taxonomy considers species boundaries from multiple, complementary perspectives, with the main objective being to compare the observed data against the predictions of the methodologies used. In the present study we used three methods for delineating species boundaries within the cosmopolitan nematode species Rhabditis ( Pellioditis ) marina and Halomonhystera disjuncta . First, phylogenetic relationships among molecular sequences from the mitochondrial cytochrome oxidase c subunit 1 gene (COI), and from two nuclear regions, internal transcribed spacer (ITS) and D2D3, were analysed. Subsequently, multivariate morphometric analysis was used to investigate whether concordant molecular lineages were also morphologically distinct. When morphological differences were found, typological taxonomy was performed to identify fixed or non-overlapping characters between lineages. Interbreeding experiments were conducted between the two closest related lineages of R . ( P. ) marina to investigate potential reproductive isolation. This integrative approach confirmed the presence of several species within each nominal species: molecular lineages were concordant across two independent loci (COI and ITS), and were characterized by significant morphological divergence. Most lineages were also detectable in the D2D3 region, but were less resolved. The two lineages investigated in our study did not produce offspring. Our results highlight that classical taxonomy grossly underestimates species diversity within the phylum Nematoda.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 94 , 737–753.     

The evolutionary history of the Chydoridae (Crustacea: Cladocera)

Although much is known about the evolutionary history of the pelagic 'cladocerans', there is little information on benthic families such as the Chydoridae. In this study, we examine the phylogenetic history of 37 chydorid species using sequence variation in two mitochondrial genes, COI and 16S rDNA, and one nuclear gene, 18S rDNA. The four recognized subfamilies of chydorids (Eurycercinae, Saycinae, Aloninae and Chydorinae) were well supported, being separated by large sequence divergences of 14.3–16.4%. By contrast, the existing taxonomic system appears to be less clear at a generic level, since many genera (e.g. Alona , Chydorus , Pleuroxus ) consist of an amalgam of distantly related species. However, among those genera which are monophyletic, levels of divergence are very high, suggesting that they originated somewhere in the mid-Palaeozoic. The factors involved in promoting diversification in this group are discussed.  © 2003 The Linnean Society of London, Biological Journal of the Linnean Society , 2003, 79, 629–643.     

Evolution in the slow lane: molecular rates of evolution in sexual and asexual ostracods (Crustacea: Ostracoda)

Parthenogenetic lineages within non-marine ostracods can occur either in mixed (with sexual and asexual females) or exclusively asexual taxa. The former mode of reproduction is associated with a high intraspecific diversity at all levels (genetic, morphological, ecological) and, at least in the Cypridoidea, with geographical parthenogenesis. Obligate asexuality is restricted to the Darwinuloidea, the strongest candidate for an ancient asexual animal group after the bdelloid rotifers, and is characterized by low diversity. We have compared rates of molecular evolution for the nuclear ITS1 region and the mitochondrial COI gene amongst the three major lineages of non-marine ostracods with sexual, mixed and asexual reproduction. Absolute rates of molecular evolution are low for both regions in the darwinulids. The slow-down of evolution in ITS1 that has been observed for Darwinula stevensoni (Brady & Robertson) apparently does not occur in other darwinulid species. ITS1 evolves more slowly than COI within non-marine ostracod families, including the darwinulids, but not between superfamilies. The ancient asexuals might have a higher relative substitution rate in ITS1, as would be expected from hypotheses that predict the accumulation of mutations in asexuals. However, the speed-up of ITS could also be ancient, for example through the stochastic loss of most lineages within the superfamily after the Permian–Triassic mass extinction. In this case, the difference in rate would have occurred independently from any effects of asexual reproduction.  © 2003 The Linnean Society of London, Biological Journal of the Linnean Society , 2003, 79 , 93–100.     

Phylogeography of Troglophilus (Orthoptera: Troglophilinae) based on Anatolian members of the genus: radiation of an old lineage following the Messinian

Active tectonic history of the Eastern Mediterranean, especially around Aegean area, through the Neogene led to interesting radiation patterns of animal lineages, allowing intriguing biogeographical hypotheses to be tested. Descendants of the ancestral stock in the Miocene Aegean Plate presently occur in the Anatolia, Aegean islands and the Balkan Penninsula. Troglophilus (Orthoptera, Rhaphidophoridae, Troglophilinae) is such a genus represented in these areas with approximately 15 species. The present study addresses the phylogeography of the genus, with a special emphasis on its Anatolian members, aiming to test the biogeographical patterns suggested for this area using mitochondrial [cytochome oxidase I (COI)] and nuclear (ITS1‐5.8S–ITS2) markers. Data matrices obtained from sequences of COI and ITS1‐5.8S–ITS2 were used for phylogenetic analyses using Dolichopoda lycia and Dolichopoda sbordonii as outgroups. All sets of the analyses suggested monophyly of the Anatolian haplotypes, although they are not congruent in revealing their relationships. Anatolian haplotypes constituted three main phylogroups in trees calculated from a matrix of short COI sequences: the ECMA (corresponding to the Eastern part of coastal Mediterranean Anatolia); the CWMA (from the Central and Western part of Mediterranean Anatolia); and NA (from Northern Anatolia). Trees obtained using longer sequences resulted in only two phylogroups, namely ECMA and CWMA + NA. The trees based on the ITS1‐5.8S–ITS2 data matrix supported monophyly of Anatolian phylogroups. BEAST analysis of the COI estimated the time to most recent common ancestor for Dolichopoda and Troglophilus as 10.8 Mya, to that for the Anatolian + Balkan Troglophilus as 7.2 Mya, and to that for the Anatolian Troglophilus as 6.3 Mya. BEAST analysis of ITS1–ITS2 intron regions is largely congruent with that of COI. From these results, several conclusions were drawn. First, the divergence of Dolichopoda and Troglophilus possibly started with the opening of the Mid‐Aegean Trench in the Tortonian. Second, Troglophilus possibly originated from an ancestral stock in the old Aegean Plate. It later diverged as Anatolian and Balkan lineages and, possibly, the Cretan population may be regarded the third lineage. Divergence within the Anatolian lineage is estimated to have occurred through the Pliocene and Pleistocene, although before the last four glacial periods in the late Pleistocene. Additionally, the northern Anatolian Troglophilus appears to originate from the dispersal of an ancestral stock from a mountainous lineage through the Taurus Way. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 108 , 335–348.     

Molecular systematics and patterns of morphological evolution in the Centropagidae (Copepoda: Calanoida) of Argentina

Recent studies have shown the value of complementing standard taxonomy with genetic analyses to reveal cryptic diversity and to aid in the understanding of patterns of evolution. We surveyed variation in the COI mitochondrial gene in members of the three genera of centropagid copepods from the inland waters in Argentina. In general, we found a close association between molecular and morphological systematics in this group. Similar to findings for marine calanoids, genetic distances within Boeckella species were modest (< 4%), while distances among morphospecies were much larger (> 11%). Parabroteas is currently monotypic, although we detected cryptic genetic diversity, with two lineages showing 5.5% divergence. In contrast, Karukinka was not a valid genus, apparently representing an interesting and atavistic offshoot of B. poppei , a result reinforcing the value of considering both morphological and molecular evidence. Moreover, we used combined genetic and morphological information, analysed with maximum likelihood methods, to evaluate the common assumption that evolution tends to proceed via the loss of structures in crustaceans. Although analysis of other taxa and character types is required to evaluate fully the reduction hypothesis, our results suggest that structures may be gained readily as well as lost.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 90 , 279–292.     

Taxonomic status of two South American sympatric squat lobsters, Munida gregaria and Munida subrugosa (Crustacea: Decapoda: Galatheidae), challenged by DNA sequence information

We investigated the taxonomic status of two sympatric morphospecies of squat lobsters from southern South America (Beagle Channel, Strait of Magellan, and Burdwood Bank), Munida gregaria and Munida subrugosa , by DNA sequence analysis of three mitochondrial (mt)DNA gene fragments [416 bp of 16S rDNA(165), 566 bp of cytochrome c oxidase subunit I(COI) and 418 bp of NADH dehydrogenase subunit 1 (ND1)]; and the nuclear rDNA internal transcribed spacer (ITS) 1 (883–952 bp). We obtained a total of 79 sequences from 32 individuals. The 16S sequences of all M. gregaria and M. subrugosa were invariant and identical, whereas COI and ND1 showed 12 and 15 variable sites, respectively. These polymorphisms were shared between morphospecies. Interspecific Tamura–Nei distances for COI and ND1 sequences were 0.0024 and 0.0032, respectively, and were not significantly different from intraspecific distances (Kruskal–Wallis tests: P  = 0.58 and P  = 0.69, for COI and ND1, respectively). Similar to the results obtained from the mtDNA sequences, no relationship was found between the ITS1 maximum parsimony tree topology and the morphologic classification of specimens in M. gregaria and M. subrugosa . We conclude that M. gregaria and M. subrugosa from southern South America may either represent a case of a dimorphic species, or a case of incomplete lineage sorting. The fact that these two morphospecies did not show fixed differences over a total of 1947 bp analysed reinforces the hypothesis of a single dimorphic species.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 94 , 421–434.     

Revision of the New Caledonian endemic genus Gracilipsodes (Trichoptera: Leptoceridae: Grumichellini)

The taxonomy and systematics of the New Caledonian endemic caddisfly genus Gracilipsodes Sykora, 1967 (Trichoptera: Leptoceridae: Grumichellini) are reviewed. Seven new species represented by males are described and illustrated: Gracilipsodes aoupiniensis sp. nov. , Gracilipsodes aureus sp. nov. , Gracilipsodes aurorus sp. nov. , Gracilipsodes grandis sp. nov. , Gracilipsodes koghiensis sp. nov. , Gracilipsodes lanceolatus sp. nov. , and Gracilipsodes robustus sp. nov. Molecular phylogenetic analyses are applied to discern the relationships among the species of the genus and their closest relatives, based on sequence characters from the nuclear gene translation elongation factor-1α (EF-1α) and the three mitochondrial genes cytochrome oxidase I (COI), COII, and ribosomal large subunit (16S). The data are analyzed using parsimony and Bayesian inference, revealing a monophyletic Gracilipsodes with the eastern Australian monotypic genus Triplexa as its closest relative. Gracilipsodes is in turn divided into two major lineages.  © 2008 The Linnean Society of London, Zoological Journal of the Linnean Society, 2008, 153 , 425–452.     

Hidden diversity of Euscorpius (Scorpiones: Euscorpiidae) in Greece revealed by multilocus species‐delimitation approaches

Phylogenetic analysis of the genus Euscorpius (Scorpiones: Euscorpiidae) across the Mediterranean region (86 specimens, 77 localities, four DNA markers: 16S rDNA, COI, COII, and ITS1), focusing on Greek fauna, revealed high variation, deep clade divergences, many cryptic lineages, paraphyly at subgenus level, and sympatry of several new and formerly known lineages. Numerous specimens from mainland and insular Greece, undoubtedly the least studied region of the genus' distribution, have been included. The reconstructed phylogeny covers representative taxa and populations across the entire genus of Euscorpius. The deepest clades detected within Euscorpius correspond (partially) to its current subgeneric division, outlining subgenera Tetratrichobothrius and Alpiscorpius. The rest of the genus falls into several clades, including subgenus Polytrichobothrius and a paraphyletic subgenus Euscorpius s.s. Several cryptic lineages are recovered, especially on the islands. The inadequacy of the morphological characters used in the taxonomy of the genus to delineate species is discussed. Finally, the time frame of differentiation of Euscorpius in the study region is estimated and the distributional patterns of the lineages are contrasted with those of other highly diversified invertebrate genera occurring in the study region. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 110 , 728–748.     

Deep genetic divergences among morphologically similar and parapatric Skistodiaptomus (Copepoda: Calanoida: Diaptomidae) challenge the hypothesis of Pleistocene speciation

We used mitochondrial [cytochrome c oxidase subunit I (CO I ), cytochrome b , and 16S] and nuclear [internal transcribed spacer (ITS) phylogenies of Skistodiaptomus copepods to test hypotheses of Pleistocene divergence and speciation within the genus. Mitochondrial (mt)DNA sequence divergences do not support hypotheses for Pleistocene speciation and instead suggest much more ancient speciation events in the genus. Skistodiaptomus oregonensis and Skistodiaptomus pygmaeus (i.e. two morphologically similar and parapatric species) exhibited uncorrected mtDNA sequence divergences exceeding 20%. Similarly, we identified three divergent clades of Skistodiaptomus pallidus that exhibited mtDNA sequence divergences exceeding 15%, suggesting that even intraspecific divergence within this morphospecies predates the Pleistocene. We found clear evidence of CO I pseudogenes in S. pygmaeus , but their presence did not lead to significant overestimates of sequence divergences for this gene. Substitution saturation and strong purifying selection have most likely led to underestimates of sequence divergences and divergence times among Skistodiaptomus . The widespread phenomenon of morphological stasis among genetically divergent copepod groups indicates that speciation often occurs with little or no morphological change. Instead, morphological evolution may occur idiosyncratically after speciation and create discordant patterns of morphological similarity, shared ancestry and divergence time. Cryptic species complexes are therefore common in copepods, and morphological species concepts underestimate their true species diversity.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 150–165.     

The molecular systematics of Leiopotherapon unicolor (Günther, 1859): testing for cryptic speciation in Australia's most widespread freshwater fish

Leiopotherapon unicolor is the most widespread freshwater fish species in Australia. A comprehensive allozyme and mitochondrial DNA 16S rRNA data set was assembled from 141 specimens of L. unicolor collected Australia-wide in order to test for cryptic speciation in this far-ranging species. Surprisingly, little genetic diversity was observed within L. unicolor and provided no evidence for the existence of cryptic species within this lineage. In contrast, a small sample set of L. aheneus used as the outgroup showed two highly divergent haplotypes strongly suggestive of cryptic speciation. L. unicolor has a number of ecological and life history attributes that may explain the lack of significant genetic divergence over substantial geographical distances. The occurrence of other widespread fish and crustacean species that also display only limited genetic diversity indicate that climate conditions more favourable to dispersal across central and northern Australia than is suggested by the extent of present-day aridity have occurred in the relatively recent geological past.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 87 , 537–552.     

Phylogenetic analyses of mtDNA sequences reveal three cryptic lineages in the widespread neotropical frog Leptodactylus fuscus (Schneider, 1799) (Anura, Leptodactylidae)

Leptodactylus fuscus is a neotropical frog ranging from Panamá to Argentina, to the east of the Andes mountains, and also inhabiting Margarita, Trinidad, and the Tobago islands. We performed phylogenetic analyses of 12S rRNA, 16S rRNA, tRNA-Leu, and ND1 mitochondrial (mt) DNA sequences from specimens collected across the geographic distribution of L. fuscus to examine two alternative hypotheses: (i) L. fuscus is a single, widely distributed species, or (ii) L. fuscus is a species complex. We tested statistically for geographic association and partitioning of genetic variation among mtDNA clades. The mtDNA data supported the hypothesis of several cryptic species within L. fuscus. Unlinked mtDNA and nuclear markers supported independently the distinctness of a 'northern' phylogenetic unit. In addition, the mtDNA data divided the southern populations into two clades that showed no sister relationship to each other, consistent with high differentiation and lack of gene flow among southern populations as suggested by allozyme data. Concordance between mtDNA and allozyme patterns suggests that cryptic speciation has occurred in L. fuscus without morphological or call differentiation. This study illustrates a case in which lineage splitting during the speciation process took place without divergence in reproductive isolation mechanisms (e.g. advertisement call in frogs), contrary to expectations predicted using a biological species framework.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society, 2006, 87 , 325–341. No claim to original US government works.     

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.     

Opening a can of worms: unprecedented sympatric cryptic diversity within British lumbricid earthworms

Earthworms play a major role in many aspects of soil fertility, food web ecology and ecosystem functioning, and hence are frequently the subjects of, for example, ecological and toxicological research. Our aim was to examine the genetic structure of common earthworm species, to identify cryptic lineages or species that may be distinct ecotypes or biotypes (and hence confound current research based upon morphotypes) and to try to explain the massive cryptic diversity that eventually emerged. We demonstrated that species such as Allolobophora chlorotica, Aporrectodea longa, Aporrectodea rosea and Lumbricus rubellus all comprise highly divergent lineages with species-level divergence at the mitochondrial cytochrome oxidase I (COI) gene. In Allo. chlorotica alone, we found 55 haplotypes for COI, with 35 of these being found in pink and 20 in green morph worms. There were no cases of the two colour morphs sharing COI haplotypes. Phylogenetic analyses of mitochondrial COI and 16S genes showed the presence of five highly divergent lineages, suggesting the presence of multiple cryptic species within Allo. chlorotica. There was no clear geographical pattern to lineage distribution and many populations were polymorphic for both mitochondrial DNA lineage and colour morph. Amplified fragment length polymorphism results, based on two primer combinations, were broadly congruent with mitochondrial DNA results with one significant exception. Despite showing over 14% divergence at COI, amplified fragment length polymorphism markers showed that the two green morph lineages may be interbreeding and therefore represent a single taxon. The cryptic diversity revealed by these results has profound consequences for all areas of earthworm research.     

The Straits of Gibraltar: barrier or bridge to Ibero‐Moroccan bat diversity?

Genetic divergence in bat communities was assessed on both sides of the Straits of Gibraltar and cryptic diversity was examined. Screening was carried out using partial sequences of the mitochondrial (mt)DNA cytochrome b gene on 399 individual bats belonging to the 18 species found on both sides of the Straits of Gibraltar. For those bats that showed important genetic discontinuities, molecular markers (ND1 and nuclear RAG2 genes) were added to expand the sampling process. Phylogenetic reconstructions were obtained using maximum parsinomy, genetic distances, maximum likelihood, and Bayesian criteria. As an estimate of bats' flight performance, we measured for each species the wing aspect ratio and wing loading indexes, and correlated them with the maximum pairwise genetic distances obtained between southern Iberian and northern Moroccan populations. Genetic mtDNA distances between populations on both continents exceed 5% in seven out of 18 bat species analysed and unknown lineages were uncovered within the species complexes Myotis nattereri and Myotis mystacinus . We did not find a general pattern in the degree of permeability of the Straits of Gibraltar for bats. Genetic distances were not correlated with the ability to cross the Straits. Our study shows that the cryptic diversity uncovered among bats continues to increase as more regions are studied.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 434–450.     

Molecular variation and speciation in Antitrichia curtipendula s.l. (Leucodontaceae, Bryophyta)

Molecular variation in Antitrichia curtipendula (Hedw.) Brid. s.l. was studied based on the nuclear internal transcribed spacer (ITS) and the chloroplast markers trn L- trn F and rpl 16, and analysed by neighbour joining (for ITS; recombination present), maximum parsimony (chloroplast markers) and TCS (haplotype network). Old World plus E North American populations belong to a different lineage than those of W North America. These are molecularly well differentiated and are treated as A. curtipendula and A. gigantea (Sull. & Lesq.) Kindb. Two distinct groups of Old World haplotypes are separated by one 'missing' haplotype and are interpreted as cryptic species. Tropical African populations share one ITS deletion and form a lineage within one of the cryptic species. Molecular variation within A. gigantea , within each of the two cryptic Old World plus E North American (except tropical African) haplotype groups, and among tropical African populations are of similar magnitude, suggesting that analogous mechanisms and similar time spans explain the found variation. Events related to Pleistocene climatic oscillations are suggested as having caused this differentiation within each group, whereas the African lineage was probably split off before this. Identical tRNA-Gly sequences were found in 33 specimens; new primers were designed for rpl 16 and ITS 1+2.  © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society , 2008, 156 , 341–354.     

A combined genetic-morphometric analysis unravels the complex biogeographical history of Polyommatus icarus and Polyommatus celina common blue butterflies

Widespread species have the potential to reveal large‐scale biogeographical patterns, as well as responses to environmental changes possibly unique to habitat generalists. This study presents a continental‐scale phylogeographical analysis of Polyommatus icarus, one of the most common Palaearctic butterflies, and the morphologically and ecologically similar Polyommatus celina, a recently discovered cryptic species. By combining data from mitochondrial [cytochrome c oxidase subunit I (COI)] and nuclear [internal transcribed spacer (ITS2)] molecular markers with geometric morphometrics, we document a complex phylogeographical history for the two species. Despite morphological similarities, the genetic divergence between these two species is high (more than 5% at COI) and they are not sister species. For the first time, we show that P. celina occurs not only in North Africa but also in Europe, where it inhabits several west Mediterranean islands, as well as large parts of Iberia, where it occurs in parapatry with P. icarus. The two species appear to completely exclude each other on islands, but we provide morphological and molecular evidence that introgression occurred in the Iberian Peninsula. We discovered strongly diverged lineages that seem to represent relict populations produced by past range expansions and contractions: Crete and Iberian isolates for P. icarus, Balearics–Sardinia and Sicily–Lipari for P. celina. This study shows that a combined genetic‐morphometric approach can shed light on cryptic diversity while providing the necessary resolution to reconstruct a fine‐scale phylogeographical history of species at both spatial and temporal levels.     

Cryptic speciation on the high seas; global phylogenetics of the copepod family Eucalanidae

Few genetic data are currently available to assess patterns of population differentiation and speciation in planktonic taxa that inhabit the open ocean. A phylogenetic study of the oceanic copepod family Eucalanidae was undertaken to develop a model zooplankton taxon in which speciation events can be confidently identified. A global survey of 20 described species (526 individuals) sampled from 88 locations worldwide found high levels of cryptic diversity at the species level. Mitochondrial (16S rRNA, CO1) and nuclear (ITS2) DNA sequence data support 12 new genetic lineages as highly distinct from other populations with which they are currently considered conspecific. Out of these 12, at least four are new species. The circumglobal, boundary current species Rhincalanus nasutus was found to be a cryptic species complex, with genetic divergence between populations unrelated to geographic distance. 'Conspecific' populations of seven species exhibited varying levels of genetic differentiation between Atlantic and Pacific basins, suggesting that continental landmasses form barriers to dispersal for a subset of circumglobal species. A molecular phylogeny of the family based on both mitochondrial (16S rRNA) and nuclear (ITS2, 18S rRNA) gene loci supports monophyly of the family Eucalanidae, all four eucalanid genera and the 'pileatus' and 'subtenuis' species groups.     

Genetic cohesion of Eresus walckenaeri (Araneae, Eresidae) in the eastern Mediterranean

The eresid spider genus Eresus is morphologically and ecologically conservative. At least three species occur in Europe. However, deep genetic divergence among geographical samples within two species, E. cinnaberinus and E. sandaliatus , may suggest more cryptic species. In the present study we investigate the genetic cohesion of the third species, Eresus walckenaeri , throughout its eastern Mediterranean distribution range, relative to the E. cinnaberinus–E. sandaliatus species complex. Eresus walckenaeri specimens were monophyletic. Genetic discreteness of E. walckenaeri in a region of sympatry with its sister species in Greece provides evidence for species integrity of E. walckenaeri within the European Eresus species complex. Eresus walckenaeri exhibited high concordance between geographical location and mtDNA genealogy. Two major phylogeographical clades were found in the Greek–Turkish and Syrian–Israel parts of the investigated area, respectively (∼6.5% sequence divergence). Concordance between geography and genetic divergence was further observed between Aegean island samples and their corresponding Greek and Turkish mainland samples, suggesting regional subdivision with gradual but potentially high dispersal propensity. Monophyly and limited regional distribution indicate Mediterranean endemic origin.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 86 , 1–9.     

Geographical diversification of the genus Cicer (Leguminosae: Papilionoideae) inferred from molecular phylogenetic analyses of chloroplast and nuclear DNA sequences

Cicer L. (Leguminosae: Papilionoideae) consists of 42 species of herbaceous or semi-shrubby annuals and perennials distributed throughout the temperate zones of the Northern Hemisphere. The origin and geographical relationships of the genus are poorly understood. We studied the geographical diversification and phylogenetic relationships of Cicer using DNA sequence data sampled from two plastid regions, trnK / matK and trnS - trnG , and two nuclear regions, the internal transcribed spacer (ITS) and external transcribed spacer (ETS) regions of nuclear ribosomal DNA, from 30 species. The results from the phylogenetic analyses of combined nuclear and chloroplast sequence data revealed four well-supported geographical groups: a Middle Eastern group, a West-Central Asian group, an Aegean–Mediterranean group, and an African group. Age estimates for Cicer based on methods that do not assume a molecular clock (for example, penalized likelihood) demonstrate that the genus has a Mediterranean origin with considerable diversification in the Miocene/Pliocene epochs. Geological events, such as mountain orogenesis and environmental changes, are major factors for the dispersal of Cicer species. The early divergence of African species and their geographically distinct region in the genus suggest a broader distribution pattern of the genus in the past than at present.  © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society, 2007, 154 , 175–186.