Spatial genetic variation, phylogeography and barcoding of the peritrichous ciliate Carchesium polypinum

Most recent studies of geographic distribution of microbial eukaryotes have focused on marine rather than freshwater protists. Here, we used the freshwater peritrich ciliate Carchesium polypinum to quantify the degree of genetic diversity of four closely related and previously described lineages and to determine whether patterns of genetic differentiation showed geographic partitioning. Using an expanded dataset of 100 isolates and employing the mitochondrial marker cytochrome oxidase c subunit I (cox-1), we enriched the 6 previously identified clades of Carchesium polypinum. We found a large degree of geographic overlap among the different clades (e.g. to the level of range of sampling), but also a spatially restricted clade (e.g. to the level of one river basin). Furthermore, we present evidence of a clear geographic separation in one of the lineages with Canadian and North Carolinian isolates grouping in two distinct clusters.     

Genetic introgression, incomplete lineage sorting and faulty taxonomy create multiple cases of polyphyly in a montane clade of tyrant-flycatchers (Elaenia, Tyrannidae)

Species-level paraphyly and polyphyly are pervasive phenomena in modern phylogenetic research and can be due to a number of factors. We explore a complicated pattern of nuclear and mitochondrial polyphyly in montane Neotropical Elaenia flycatchers. Using a combination of phylogenetic and population genetic methods, we demonstrate that no single factor is sufficient to account for this pattern of polyphyly, and that it is likely based on an interplay of three different factors: (i) faulty taxonomy which has led to the recognition of two polyphyletic species that are better classified as four biological species; (ii) a late Pleistocene hybridization event that resulted in two morphologically and ecologically distinct species sharing extremely similar mitochondrial DNA but distinct nuclear DNA profiles; and (iii) incomplete lineage sorting in a nuclear marker that results in a polyphyletic placement of species that are otherwise well-differentiated in mitochondrial DNA, morphology and ecology. Additionally, we demonstrate that the two clades of montane Elaenia exhibit a reverse pattern of mitochondrial and nuclear diversity, with high mitochondrial and low nuclear genetic diversity in one clade and vice versa in the other clade. A possible cause for this pattern is differences in population histories, with large panmictic population structures being conducive to the retention of ancient nuclear polymorphisms in Elaenia albiceps chilensis .     

Decoupling of molecular and morphological evolution in deep lineages of a meiobenthic harpacticoid copepod

Molecular and biochemical genetic analyses have revealed that many marine invertebrate taxa, including some well-studied and presumably cosmopolitan species, are actually complexes of sibling species. When morphological differences are slight and estimated divergence times are old, data suggest either unusually high rates of sequence evolution or long-term morphological stasis. Here, five gene regions (mitochondrial cytochrome oxidase subunit I and large-subunit ribosomal 16S rDNA and nuclear ITS1, 5.8S rDNA, and ITS2) were analyzed in four geographic samples of the meiobenthic harpacticoid copepod Cletocamptus deitersi. Molecular sequences revealed four extremely differentiated molecular lineages with unalignable nuclear intergenic spacers and mitochondrial uncorrected divergences reaching 25% (cytochrome oxidase) and 36% (16S rDNA). These levels of divergence are greater than those reported previously for congeneric species in diverse invertebrate taxa, including crustaceans. The nominally intraspecific divergence matches or exceeds the corresponding divergence from a known congener (Cletocamptus helobius). A molecular clock applied to the cytochrome oxidase subunit I data suggests that these lineages split in the Miocene, consistent with the fossil record of a North American Cletocamptus from the same period. Morphological differences among the major lineages are subtle but congruent with the patterns of genetic differentiation. Our conclusion, based on concordant patterns of variation in two mitochondrial and three nuclear gene regions, as well as morphological observations, is that C. deitersi in North America is composed of at least four separate species by the genealogical concordance, phylogenetic, and morphological-species criteria. Alternative explanations for the deep phylogenetic nodes and apparent morphological stasis, including high rates of sequence evolution, balancing selection, and genetic signatures of historical events, are considered unlikely.     

Molecular Phylogeny of the Butterfly Genus Polytremis (Hesperiidae,Hesperiinae, Baorini) in China

Background

The genus Polytremis, restricted to the continental part of the southeastern Palaearctic and northern Oriental regions, is one of the largest and most diverse lineages of the tribe Baorini. Previous studies on the genus were focused mainly on morphological classification and identification of new species. Due to the lack of effective and homologous traits of morphology, there were many challenges in the traditional classification. In this report, we reconstruct the phylogeny to provide a solid framework for these studies and to test the traditional limits and relationships of taxa.

Methodology and Principal Findings

We sequenced a mitochondrial and three nuclear gene fragments, coupled with an evaluation of traditional morphological characters, to determine the phylogenetic relationships for a total of 15 species representing all major species groups of the Polytremis genus in China, and to elucidate their taxonomic status.

Conclusions and Significance

Analysis of mitochrondial and nuclear DNA showed considerable congruent phylogenetic signal in topology at the inter-species level. We found strong support for the monophyly of Polytremis and some clades were recognized with morphological data. Thus, the COI sequence in our study could be used as a DNA barcode to identify almost all members of the genus. However, incongruences of phylogenetic analyses occurred: in contrast to the phylogenetic trees of mitochondrial COI, it was not possible for nuclear rDNA to discriminate P. gotama from P. caerulescens, suggesting a possible recent separation of these two species. Additionally, P. theca was the only species with a greater intra-specific genetic distance compared to some inter-specific genetic distances in this study and some problems associated with the cryptic diversity of the species are discussed. The results of this study will helpful to reveal the causes of the high degree of diversity of butterflies, and possibly other groups of insects in China.     

Genetic diversity and evolutionary history of the Schizothorax species complex in the Lancang River (upper Mekong)

The genus Schizothorax (Cyprinidae), one of the most diverse genera of ichthyofauna of the Qinghai‐Tibetan Plateau (QTP), is a good candidate for investigating patterns of genetic variation and evolutionary mechanisms. In this study, sequences from the mitochondrial control region, the cytochrome b gene, and two nuclear genes were used to re‐examine the genetic diversity and investigate the evolutionary history of the Schizothorax species complex inhabiting the Lancang River. Three maternal clades were detected in the Schizothorax species complex, but frequent nuclear allele sharing also occurred among the three maternal clades. A discrepancy between topologies of mitochondrial and nuclear loci might result from introgression or/and incomplete lineage sorting. The divergence of the clades of the Schizothorax species complex was closely related to the Late Pliocene and Early Pleistocene orogenesis of the QTP and Southwest Mountains of China. Demographic analyses indicated that the species complex subsequently persisted in situ with stable populations during Pleistocene glacial cycling, which suggested that Pleistocene climate changes did not exert a remarkable influence on the species complex. Our study provides a comprehensive analysis of the genetic diversity and evolutionary history of the Schizothorax species complex in the Lancang River.     

Population genetic structure of Carchesium polypinum (Ciliophora: Peritrichia) in four Chinese lakes inferred from ISSR fingerprinting: high diversity but low differentiation

Although the peritrichous ciliate Carchesium polypinum is common in freshwater, its population genetic structure is largely unknown. We used inter-simple sequence repeat (ISSR) fingerprinting to analyze the genetic structure of 48 different isolates of the species from four lakes in Wuhan, central China. Using eight polymorphic primers, 81 discernible DNA fragments were detected, among which 76 (93.83%) were polymorphic, indicating high genetic diversity at the isolate level. Further, Nei's gene diversity (h) and Shannon's Information index (I) between the different isolates both revealed a remarkable genetic diversity, higher than previously indicated by their morphology. At the same time, substantial gene flow was found. So the main factors responsible for the high level of diversity within populations are probably due to conjugation (sexual reproduction) and wide distribution of swarmers. Analysis of molecular variance (AMOVA) showed that there was low genetic differentiation among the four populations probably due to common ancestry and flooding events. The cluster analysis and principal component analysis (PCA) suggested that genotypes isolated from the same lake displayed a higher genetic similarity than those from different lakes. Both analyses separated C. polypinum isolates into subgroups according to the geographical locations. However, there is only a weak positive correlation between the genetic distance and geographical distance, suggesting a minor effect of geographical distance on the distribution of genetic diversity between populations of C. polypinum at the local level. In conclusion, our studies clearly demonstrated that a single morphospecies may harbor high levels of genetic diversity, and that the degree of resolution offered by morphology as a marker for measuring distribution patterns of genetically distinct entities is too low.     

A molecular phylogenetic investigation of zoothamnium (ciliophora, peritrichia, sessilida)

The gene coding for 18S small subunit ribosomal RNA (ssu rRNA) was sequenced in seven free-living, marine species of the sessiline peritrich genus Zoothamnium. These were Zoothamnium niveum, Zoothamnium alternans, Zoothamnium pelagicum, and four unidentified species. The ssu rRNA gene also was sequenced in Vorticella convallaria, Vorticella microstoma, and in an unidentified, freshwater species of Vorticella. Phylogenetic trees were constructed using these new sequences to test a previously published phylogenetic association between Zoothamnium arbuscula, currently in the family Zoothamniidae, and peritrichs in the family Vorticellidae. Trees constructed by means of neighbor-joining, maximum parsimony, maximum likelihood, and Bayesian inference methods all had similar topologies. The seven new sequences of Zoothamnium species grouped into three well-supported clades, each of which contained a diversity of morphological types. The three clades formed a poorly supported, larger clade that was deeply divergent from Z. arbuscula, which remained more closely associated with vorticellid peritrichs. It is apparent that Zoothamnium is a richly diverse genus and that a much more intensive investigation, involving both morphological and molecular data and a wider selection of species, will be necessary to resolve its phylogeny. A greater amount of molecular diversity than is predicted by morphological data exists within all major clades of sessiline peritrichs that have been included in molecular phylogenies, indicating that characteristics of stalk and peristomial structure traditionally used to differentiate taxa at the generic level and above may not be uniformly reliable.     

The phylogeny of acorn weevils (genus Curculio) from mitochondrial and nuclear DNA sequences: the problem of incomplete data

We considered the contribution of two mitochondrial and two nuclear data sets for the phylogenetic reconstruction of 22 species of seed beetles in the genus Curculio (Coleoptera: Cuculionidae). A phylogenetic tree from representatives found on various hosts was inferred from a combined data set of mitochondrial DNA cytochrome oxidase subunit I, mitochondrial cytochrome b, nuclear elongation factor 1alpha, and nuclear phosphoglycerate mutase, used for the first time as a molecular marker. Separate parsimony analyses of each data set showed that individual gene trees were mainly congruent and often complementary in the support of clades but the analysis was complicated by failure of PCR amplification of nuclear genes for many taxa and hence missing data entries. When the four gene partitions were combined in a simultaneous analysis despite the missing data, this increased the resolution and taxonomic coverage compared to the individual source trees. Alternative approaches of combining the information via supertree methodology produced a comparatively less resolved tree, and hence seem inferior to combining data matrices even in cases where numerous taxa are missing. The molecular data suggest a classification of the European species into two species groups that are in accordance with morphological characteristics but the data do no support any of the previously recognised American species groups.     

The hidden diversity of Coleodactylus amazonicus (Sphaerodactylinae,Gekkota) revealed by molecular data

Coleodactylus amazonicus, a small leaf-litter diurnal gecko widely distributed in Amazon Basin, has been considered a single species with no significant morphological differences between populations along its range. A recent molecular study, however, detected large genetic differences between populations of central Amazonia and those in the easternmost part of the Amazon Basin, suggesting the presence of taxonomically unrecognised diversity. In this study, DNA sequences of three mitochondrial (16S, cytb, and ND4) and two nuclear genes (RAG-1, c-mos) were used to investigate whether the species currently identified as C.amazonicus contains morphologically cryptic species lineages. The present phylogenetic analysis reveals further genetic subdivision including at least five potential species lineages, restricted to northeastern (lineage A), southeastern (lineage B), central-northern (lineage E) and central-southern (lineages C and D) parts of Amazon Basin. All clades are characterized by exclusive groups of alleles for both nuclear genes and highly divergent mitochondrial haplotype clades, with corrected pairwise net sequence divergence between sister lineages ranging from 9.1% to 20.7% for the entire mtDNA dataset. Results of this study suggest that the real diversity of “C. amazonicus” has been underestimated due to its apparent cryptic diversification.     

Molecular patterns of differentiation in canyon treefrogs (Hyla arenicolor): evidence for introgressive hybridization with the Arizona treefrog (H. wrightorum) and correlations with advertisement call differences

Detection of genetic and behavioural diversity within morphologically similar species has led to the discovery of cryptic species complexes. We tested the hypothesis that US populations of the canyon treefrog (Hyla arenicolor) may consist of cryptic species by examining mate‐attraction signals among three divergent clades defined by mtDNA. Using a multi‐locus approach, we re‐analysed phylogenetic relationships among the three clades and a closely related, but morphologically and behaviourally dissimilar species, the Arizona treefrog (H. wrightorum). We found evidence for introgression of H. wrightorum’s mitochondrial genome into H. arenicolor. Additionally, the two‐clade topology based on nuclear data is more congruent with patterns of call variation than the three‐clade topology from the mitochondrial dataset. The magnitude of the call divergence is probably insufficient to promote isolation of the nuclear DNA‐defined clades should they become sympatric, but further divergence in call properties significant in species identification could promote speciation in the future.     

Phenotypic plasticity of six unusual cercariae in nassariid gastropods and their relationships to the Acanthocolpidae and Brachycladiidae (Digenea)

Species-level relationships of a complex of six putative acanthocolpid cercariae with ventral keels from nassariid gastropods from Capricornia, Queensland (Cercaria capricornia I–VI) were explored using mitochondrial and nuclear DNA sequence data. Analysis of the large subunit of nuclear ribosomal DNA (rDNA), the internal transcribed spacer 2 rDNA and subunit 3 of the nicotinamide-adenine-dinucleotide dehydrogenase mitochondrial gene indicates that the six distinct morphotypes previously identified relate to only three discrete genotypes. Consequently, the six cercarial morphotypes that were inferred to represent six species and distinguished previously based on morphology and behaviour, are considered here to be comprised of three species (i.e. Cercaria capricornia I and II are considered the same species, C. capricornia types III to V are considered to belong to a second species and C. capricornia VI is considered to comprise the third species in this complex). This recognition of reduced diversity following molecular analysis runs contrary to the more frequently reported uncovering of cryptic diversity, especially when larval trematodes are examined. Phylogenetic analysis of these morphologically unusual cercariae supports their inclusion in the Acanthocolpidae. Bayesian inference analysis identifies three distinct clades: (a) Stephanostomum + Monostephanostomum; (b) the Brachycladiidae and Pleorchis; and (c) Tormopsolus and the present cercariae. Genus-level identification of these cercariae remains uncertain given the current lack of available comparative genetic data on nucleic acid databases.     

Multigenic phylogeographic divergence in the paleoendemic southern Appalachian opilionid Fumontana deprehendor Shear (Opiliones, Laniatores, Triaenonychidae)

The paleoendemic opilionid Fumontana deprehendor is restricted to a small area of mid-elevation forested habitats in the southern Blue Ridge province of the Appalachian Mountains. In a recent study we reported on the discovery of 22 new montane populations of this monotypic genus, specimens from which exhibit remarkably little morphological divergence despite their separation by intervening lowlands and large riverine barriers. Here, we further explore spatial and temporal patterns of divergence in this taxon using DNA sequence data from a portion of the mitochondrial cytochrome c oxidase subunit I gene ( approximately 1000 bp) and full-length sequences of both nuclear ribosomal internal transcribed spacer regions, including the intervening 5.8S rRNA region ( approximately 700 bp total). Bayesian phylogenetic analyses of these independent data sets reveal congruent genealogical patterns, with all data partitioning and combination strategies consistently recovering five allopatric, geographically cohesive genetic clades. These clades show an almost complete lack of internal genetic divergence, with most individuals sharing a clade-specific, regionally widespread haplotype. The geographic distribution of these clades corresponds to patterns seen in other upland taxa of the region, possibly indicating coincident vicariance. Because of a lack of quantifiable morphological divergence and relatively modest levels of genetic divergence, we conservatively refer to the geographically cohesive genetic clades as "phylogeographic units", although these may actually represent cryptic species. Conservation implications and the prospect for future comparative arachnid phylogeography in the southern Appalachians are discussed in light of the results presented here.     

The history and geography of diversification within the butterfly genus Lycaeides in North America

The Lycaeides butterfly species complex in North America consists of two nominal, morphologically defined species. These butterflies are ecologically diverse and appear to be distributed as a geographically complex mosaic of locally differentiated populations that may be undergoing adaptive radiation. We asked whether patterns of molecular genetic variation within the species complex are congruent with currently recognized morphological species and whether the distribution of molecular variation is consistent with the hypothesis that Pleistocene climate changes contributed to the process of differentiation within the genus. Variation in the form of the genitalia from 726 males from 59 populations clearly distinguishes both species with only six populations containing morphologically intermediate or ambiguous individuals. However, partitioning of molecular variance in a 236 bp section of the mitochondrial AT-rich region from 628 individuals (57 populations) surveyed using single strand conformation polymorphism analysis (SSCP) indicates that only 26% of the total genetic variation is distributed along nominal species boundaries as defined by morphology. Instead, three phylogeographical groups were detected, represented by three major haplotype clades, which account for 90% of the total genetic variance. Pleistocene glaciations appear to have fostered divergence during glacial maxima, while post-glacial range expansions created opportunities for gene exchange and reticulation along suture zones between geographical groups. Data presented here allow us to make inferences about the history of the species complex. However, evidence of ancestral polymorphism and reticulation limit our ability to define species boundaries based on mitochondrial DNA sequence variation.     

Molecular differentiation of the microgastrine species commonly found in paddy fields from Southeast Asia,with additional data on their phylogeny （Hymenoptera：Braconidae）

Partial DNA sequences of three genes, that is, mitochondrial large ribosomal subunit (16S), nuclear large ribosomal subunit (28S D2) and mitochondrial NADH1 dehydrogenase (NADH1) gene, were sequenced from different microgas trine species(Braconidae: Microgastrinae) collected fresh from paddy fields. The DNA sequences were used to determine the extent of sequence variation among species in order to evaluate the specific status of each species. Cladistic analysis was also used to infer a phylogenetic relationship among these species. The results showed that sequence divergence among species of the same genus Cotesia was much lower than those among different genera, such as Cotesia, Exoryza and Apanteles; the sequence similarity of 16S rDNA and NADH 1 genes between Cotesia sp. and C. chilonis was higher than that between C. sp. and C. ruficrus.Phylogenetic analyses suggested that four species of Cotesia were always grouped in the same clade regardless of using different analysis methods; Cotesia sp. and C. chilonis are more closely related to each other than to C. ruficrus, different from previous morphological results. Additionally, sequence analyses indicated that NADH1 gene has more parsimony informative characters than 28S rDNA D2 and 16S rDNA at the species-level analysis,indicating that NADH1 gene might be a useful marker for species-level analysis.     

A phylogenetic perspective on habitat shifts and diversity in the North American Enallagma damselflies

Community ecologists are increasingly aware that the regional history of taxon diversification can have an important influence on community structure. Likewise, systematists recognize that ecological context can have an important influence on the processes of speciation and extinction that create patterns of descent. We present a phylogenetic analysis of 33 species of a North American radiation of damselflies (Zygoptera: Coenagrionidae: Enallagma Selys), which have been well studied ecologically, to elucidate the evolutionary mechanisms that have contributed to differences in diversity between larval habitats (lakes with and without fish predators). Analysis of molecular variation in 842 bp of the mitochondrial cytochrome oxidase I and II subunit and of the intervening Leu-tRNA and 37 morphological characters resulted in three well-defined clades that are only partially congruent with previous phylogenetic hypotheses. Molecular and morphological data partitions were significantly incongruent (p < .01). Lack of haplotype monophyly within species and small amounts of sequence divergence (< 1%) between related species in three of the four clades suggest that recent, and parallel, speciation has been an important source of community diversity. Reconstruction of habitat preference over the phylogeny suggests that the greater species diversity in fish-containing lake habitats reflects the recency of shifts into the fishless lake habit, although a difference in speciation or extinction rates between the two habitats is difficult to exclude as an additional mechanism.     

Unrecognized diversity of a scale worm,Polyeunoa laevis (Annelida: Polynoidae), that feeds on soft coral

A goal of taxonomy is to employ a method of classification based on phylogeny that captures the morphological and genetic diversity of organismal lineages. However, morphological and genetic diversity may not always be concordant, leading to challenges in systematics. The scale worm Polyeunoa laevis has been hypothesized to represent a species complex based on morphology, although there is little knowledge of its genetic diversity. Commonly found in Antarctic waters and usually associated with gorgonian corals (especially Thouarella), this taxon is also reported from the south-west Atlantic, Magellanic and sub-Antarctic regions. We employ an integrative taxonomic approach to examine the traditional morphological characters used for scale worm identification in combination with COI mitochondrial gene data and whole mtDNA genomes. Moreover, we consider P. laevis's association with Thouarella by examining data from the mMutS gene, a soft-coral phylogenetic marker. Analyses for P. laevis recovered 3 clades, two in Antarctic waters and one from the Argentina-Indian Ocean. Interestingly, genetic and morphological results show differences between specimens from South Argentina and the Antarctic region, suggesting that open ocean barriers might have limited gene flow from these regions. Bayesian phylogenetic analyses for Thouarella resulted in at least 12 lineages, although some of the lineages consist of only a single individual. Our results show different evolutionary histories for both species, confirming that association between these scale worms and their hosts is not restrictive. For both taxonomic groups, biodiversity in the Southern Ocean appears to be underestimated.     

Molecular phylogeny and phylogeography of the Cuban cave-fishes of the genus Lucifuga: evidence for cryptic allopatric diversity

Underground environments are increasingly recognized as reservoirs of faunal diversity. Extreme environmental conditions and limited dispersal ability of underground organisms have been acknowledged as important factors promoting divergence between species and conspecific populations. However, in many instances, there is no correlation between genetic divergence and morphological differentiation. Lucifuga Poey is a stygobiotic fish genus that lives in Cuban and Bahamian caves. In Cuba, it offers a unique opportunity to study the influence of habitat fragmentation on the genetic divergence of stygobiotic species and populations. The genus includes four species and one morphological variant that have contrasting geographical distributions. In this study, we first performed a molecular phylogenetic analysis of the Lucifuga Cuban species using mitochondrial and nuclear markers. The mitochondrial phylogeny revealed three deeply divergent clades that were supported by nuclear and morphological characters. Within two of these main clades, we identified five lineages that are candidate cryptic species and a taxonomical synonymy between Lucifuga subterranea and Lucifuga teresinarum. Secondly, phylogeographic analysis using a fragment of the cytochrome b gene was performed for Lucifuga dentata, the most widely distributed species. We found strong geographical organization of the haplotype clades at different geographic scales that can be explained by episodes of dispersal and population expansion followed by population fragmentation and restricted gene flow. At a larger temporal scale, these processes could also explain the diversification and the distribution of the different species.     

Exploring the molecular diversity of terrestrial nemerteans (Hoplonemertea, Monostilifera, Acteonemertidae) in a continental landmass

We report the finding of terrestrial nemerteans of the family Acteonemertidae from the northern part of Spain. The specimens were studied using molecular data from the nuclear ribosomal 18S rRNA and the mitochondrial cytochrome c oxidase subunit I genes. The 18S rRNA data strongly suggest the presence of three species of terrestrial nemerteans in the Iberian Peninsula, an old landmass where terrestrial nemerteans had not previously been reported. These specimens originate from primarily undisturbed forests across a distance of c . 1000 km. Cytochrome c oxidase subunit I data also indicate the presence of multiple lineages of Iberian terrestrial nemerteans. Nonetheless, the pattern obtained from this marker is obscured most probably by deep genetic divergences. This molecular diversity, at least in some of the clades, suggests that the Iberian species are not the result of recent introductions, as proposed for other terrestrial nemerteans found in Europe. Our data also touch upon the question of a single origin of terrestriality in nemerteans, a hypothesis rejected by both data sets. Nevertheless, terrestriality seems to have had a single origin in the family Acteonemertidae.     

High levels of cryptic species diversity uncovered in Amazonian frogs

One of the greatest challenges for biodiversity conservation is the poor understanding of species diversity. Molecular methods have dramatically improved our ability to uncover cryptic species, but the magnitude of cryptic diversity remains unknown, particularly in diverse tropical regions such as the Amazon Basin. Uncovering cryptic diversity in amphibians is particularly pressing because amphibians are going extinct globally at an alarming rate. Here, we use an integrative analysis of two independent Amazonian frog clades, Engystomops toadlets and Hypsiboas treefrogs, to test whether species richness is underestimated and, if so, by how much. We sampled intensively in six countries with a focus in Ecuador (Engystomops: 252 individuals from 36 localities; Hypsiboas: 208 individuals from 65 localities) and combined mitochondrial DNA, nuclear DNA, morphological, and bioacoustic data to detect cryptic species. We found that in both clades, species richness was severely underestimated, with more undescribed species than described species. In Engystomops, the two currently recognized species are actually five to seven species (a 150-250% increase in species richness); in Hypsiboas, two recognized species represent six to nine species (a 200-350% increase). Our results suggest that Amazonian frog biodiversity is much more severely underestimated than previously thought.     

Genetic diversification without obvious genitalic morphological divergence in harvestmen (Opiliones, Laniatores, Sclerobunus robustus) from montane sky islands of western North America

The southern Rocky Mountains and adjacent Intermontane Plateau Highlands region of western North America is a geographically diverse area with an active geologic history. Given the topological complexity and extensive geologic activity, organisms inhabiting this region are expected to show some degree of morphological and genetic divergence, especially populations found on the southern montane 'sky islands' of this region. Here we examine the phylogeographic history and diversification of a montane forest inhabiting harvestmen, Sclerobunus robustus, using a combination of genetic and morphological data. Divergence time estimates indicate that much of the diversification within and between major groups S. robustus predate the Pleistocene glacial cycles. The most widespread subspecies, Sclerobunus robustus robustus, is recovered as six genetically distinct, geographically cohesive mitochondrial phylogroups. Gene tree data for a single nuclear gene reveals congruent, albeit slightly more conservative, patterns of genetic divergence. Despite high levels of genetic divergence throughout their distribution, phylogroups show extreme conservation in somatic and reproductive morphology. This uncoupling of morphological and genetic differentiation may be due to morphological conservatism associated with a conserved microhabitat preference. Based on these data, it is obvious that S. robustus has undergone some level of cryptic diversification.