Intraspecific phylogeography of Percina evides (Percidae: Etheostomatinae): an additional test of the Central Highlands pre-Pleistocene vicariance hypothesis

North America exhibits the most diverse freshwater fish fauna among temperate regions of the world. Species diversity is concentrated in the Central Highlands, drained by the Mississippi, Gulf Slope and Atlantic Slope river systems. Previous investigations of Central Highlands biogeography have led to conflicting hypotheses involving dispersal and vicariance to explain the diversity and distribution of the freshwater fish fauna. In this investigation predictions of the Central Highlands pre-Pleistocene vicariance hypothesis are tested with a phylogeographic analysis of the percid species Percina evides, which is widely distributed in several disjunct areas of the Central Highlands. Phylogenetic analysis of complete gene sequences of mitochondrially encoded cytochrome b recover three phylogroups, with very low levels of sequence polymorphism within groups. The two western phylogroups are monophyletic with respect to the eastern phylogroup. The recovery of two monophyletic lineages with an eastern and western distribution in the disjunct highland areas is a pattern expected from vicariance, but is not predicted by the Central Highlands pre-Pleistocene vicariance hypothesis. The recovery of very limited mitochondrial DNA polymorphism and lack of phylogeographic structuring across the entire range of the eastern clade, very shallow polymorphism between the disjunct Missouri River and upper Mississippi River populations, and lack of sequence polymorphism in the upper Mississippi River populations, support a hypothesis of dispersal during or following the Pleistocene. The present distribution of P. evides is best explained by both vicariant and dispersal events.     

Phylogenetic relationships,subdivision, and biogeography of the cyprinid tribe Labeonini (sensu ) (Teleostei: Cypriniformes), with comments on the implications of lips and associated structures in the labeonin classification

The Labeonini (sensu Rainboth, 1991) is a tribe of the subfamily Cyprininae, the largest subfamily of Cypriniformes. With around 400 species in 34 genera, this tribe is widely distributed in the freshwaters of tropical Africa and Asia. Most species are adapted to fast-flowing streams and rivers, and exhibit unique morphological modifications associated with their lips and other structures around the mouth. The monophyly of this tribe has been tested and generally accepted in previous morphological and molecular studies. The major objectives of this study were to reconstruct the phylogenetic relationships within the tribe Labeonini, test its monophyly and explore the taxonomic subdivisions, intrarelationships and biogeography of the group. The value of the morphological characters associated with the lips and other associated structures in the taxonomic classification of labeonins was also discussed. Nucleotide sequences (3867 bp) of four unlinked nuclear loci were obtained from 51 species in 18 Labeonini genera from throughout the range of the tribe. Maximum parsimony, partitioned maximum likelihood and partitioned Bayesian analyses were used for phylogenetic inference from combined and separate gene data sets. Based on our results, the monophyly of Labeonini was well supported. Two major clades could be recovered within the tribe. Three subclades could further be recognized from the first clade. These clades/subclades are not consistent with groupings of any of previous workers using either morphological or molecular characters for phylogenetic inference. Only five currently recognized genera in this analysis are monophyletic. The similarity between some lips and associated structures (e.g. suctorial discs) of labeonins may due to convergence or parallelism instead of common ancestry. Labeonins of Southeast Asia, India and China are closely related to each other; the multiple clades of African taxa do not form a single monophyletic group, indicating multiple, independent dispersal events of labeonins into Africa from Asia.     

Phylogeny of the gudgeons (Teleostei: Cyprinidae: Gobioninae)

The members of the cyprinid subfamily Gobioninae, commonly called gudgeons, form one of the most well-established assemblages in the family Cyprinidae. The subfamily is a species-rich group of fishes, these fishes display diverse life histories, appearances, and behavior. The phylogenetic relationships of Gobioninae are examined using sequence data from four loci: cytochrome b, cytochrome c oxidase I, opsin, and recombination activating gene 1. This investigation produced a data matrix of 4114 bp for 162 taxa that was analyzed using parsimony, maximum likelihood, and Bayesian inference methods. The phylogenies our analyses recovered corroborate recent studies on the group. The subfamily Gobioninae is monophyletic and composed of three major lineages. We find evidence for a Hemibarbus-Squalidus group, and the tribes Gobionini and Sarcocheilichthyini, with the Hemibarbus-Squalidus group sister to a clade of Gobionini-Sarcocheilichthyini. The Hemibarbus-Squalidus group includes those two genera; the tribe Sarcocheilichthyini includes Coreius, Coreoleuciscus, Gnathopogon, Gobiocypris, Ladislavia, Paracanthobrama, Pseudorasbora, Pseudopungtungia, Pungtungia, Rhinogobio, and Sarcocheilichthys; the tribe Gobionini includes Abbottina, Biwia, Gobio, Gobiobotia, Huigobio, Microphysogobio, Platysmacheilus, Pseudogobio, Romanogobio, Saurogobio, and Xenophysogobio. The monotypic Acanthogobio is placed into the synonymy of Gobio. We tentatively assign Belligobio to the Hemibarbus-Squalidus group and Mesogobio to Gobionini; Paraleucogobio and Parasqualidus remain incertae sedis. Based on the topologies presented, the evolution of swim bladder specializations, a distinctive feature among cyprinids, has occurred more than once within the subfamily.     

Comparison of evolutionary rates in the mitochondrial DNA cytochrome b gene and control region and their implications for phylogeny of the Cobitoidea (Teleostei: Cypriniformes)

It is widely accepted that mitochondrial DNA (mtDNA) control region evolves faster than protein encoding genes with few exceptions. In the present study, we sequenced the mitochondrial cytochrome b gene (cyt b) and control region (CR) and compared their rates in 93 specimens representing 67 species of loaches and some related taxa in the Cobitoidea (Order Cypriniformes). The results showed that sequence divergences of the CR were broadly higher than those of the cyt b (about 1.83 times). However, in considering only closely related species, CR sequence evolution was slower than that of cyt b gene (ratio of CR/cyt b is 0.78), a pattern that is found to be very common in Cypriniformes. Combined data of the cyt b and CR were used to estimate the phylogenetic relationship of the Cobitoidea by maximum parsimony, neighbor-joining, and Bayesian methods. With Cyprinus carpio and Danio rerio as outgroups, three analyses identified the same four lineages representing four subfamilies of loaches, with Botiinae on the basal-most clade. The phylogenetic relationship of the Cobitoidea was ((Catostomidae+Gyrinocheilidae)+(Botiinae+(Balitorinae+(Cobitinae+Nemacheilinae)))), which indicated that Sawada's Cobitidae (including Cobitinae and Botiinae) was not monophyletic. Our molecular phylogenetic analyses are in very close agreement with the phylogenetic results based on the morphological data proposed by Nalbant and Bianco, wherein these four subfamilies were elevated to the family level as Botiidae, Balitoridae, Cobitidae, and Nemacheilidae.     

Phylogenetic relationships of the Cobitoidea (Teleostei: Cypriniformes) inferred from mitochondrial and nuclear genes with analyses of gene evolution

The superfamily Cobitoidea of the order Cypriniformes is a diverse group of fishes, inhabiting freshwater ecosystems across Eurasia and North Africa. The phylogenetic relationships of this well-corroborated natural group and diverse clade are critical to not only informing scientific communities of the phylogeny of the order Cypriniformes, the world's largest freshwater fish order, but are key to every area of comparative biology examining the evolution of traits, functional structures, and breeding behaviors to their biogeographic histories, speciation, anagenetic divergence, and divergence time estimates. In the present study, two mitochondrial gene sequences (COI, ND4+5) and four single-copy nuclear gene segments (RH1, RAG1, EGR2B, IRBP) were used to infer the phylogenetic relationships of the Cobitoidea as reconstructed from maximum likelihood (ML) and partitioned Bayesian Analysis (BA). Analyses of the combined mitochondrial/nuclear gene datasets revealed five strongly supported monophyletic Cobitoidea families and their sister-group relationships: Botiidae+(Vaillantellidae+(Cobitidae+(Nemacheilidae+Balitoridae))). These recovered relationships are in agreement with previous systematic studies on the order Cypriniformes and/or those focusing on the superfamily Cobitoidea. Using these relationships, our analyses revealed pattern lineage- or ecological-group-specific evolution of these genes for the Cobitoidea. These observations and results corroborate the hypothesis that these group-specific-ancestral ecological characters have contributed in the diversification and/or adaptations within these groups. Positive selections were detected in RH1 of nemacheilids and in RAG1 of nemacheilids and genus Vaillantella, which indicated that evolution of RH1 (related to eye's optic sense) and RAG1 (related to immunity) genes appeared to be important for the diversification of these groups. The balitorid lineage (those species inhabiting fast-flowing riverine habitats) had, as compared with other cobitoid lineages, significantly different dN/dS, dN and dS values for ND4 and IRBP genes. These significant differences are usually indicative of weaker selection pressure, and lineage-specific evolution on genes along the balitorid lineage. Furthermore, within Cobitoidea, excluding balitorids, species living in subtropics had significantly higher dN/dS values in RAG1 and IRBP genes than those living in temperate and tropical zones. Among tropical cobitoids, genes COI, ND5, EGR2B, IRBP and RH1, had a significantly higher mean dS value than those species in subtropical and temperate groups. These findings suggest that the evolution of these genes could also be ecological-group-specific and may have played an important role in the adaptive evolution and diversification of these groups. Thus, we hypothesize that the genes included in the present study were actively involved in lineage- and/or ecological-group-specific evolutionary processes of the highly diverse Cobitoidea. These two evolutionary patterns, both subject to further testing, are hypothesized as integral in the diversification with this major clade of the world's most diverse group of freshwater fishes.     

Molecular phylogeny of the cyprinid tribe Labeonini (Teleostei: Cypriniformes)

The cyprinid tribe Labeonini (sensuRainboth, 1991) is a large group of freshwater fishes containing around 40 genera and 400 species. They are characterized by an amazing diversity of modifications to their lips and associated structures. In this study, a total of 34 genera and 142 species of putative members of this tribe, which represent most of the generic diversity and more than one third of the species diversity of the group, were sampled and sequenced for four nuclear genes and five mitochondrial genes (totaling 9465bp). Phylogenetic relationships and subdivision of this tribe were investigated and the placement and status of most genera are discussed. Partitioned maximum likelihood analyses were performed based on the nuclear dataset, mitochondrial dataset, combined dataset, and the dataset for each nuclear gene. Inclusion of the genera Paracrossochilus, Barbichthys, Thynnichthys, and Linichthys in the Labeonini was either confirmed or proposed for the first time. None of the genera Labeo, Garra, Bangana, Cirrhinus, and Crossocheilus are monophyletic. Taxonomic revisions of some genera were made: the generic names Gymnostomus Heckel, 1843, Ageneiogarra Garman, 1912 and Gonorhynchus McClelland, 1839 were revalidated; Akrokolioplax Zhang and Kottelat, 2006 becomes a junior synonym of Gonorhynchus; the species Osteochilus nashii was found to be a member of the barbin genus Osteochilichthys. Five historical hypotheses on the classification of the Labeonini were tested and rejected. We proposed to subdivide the tribe, which is strongly supported as monophyletic, into four subtribes: Labeoina, Garraina, Osteochilina, and Semilabeoina. The taxa included in each subtribe were listed and those taxa that need taxonomic revision were discussed.     

Phylogeny, diversity, and species delimitation of the North American Round-Nosed Minnows (Teleostei: Dionda), as inferred from mitochondrial and nuclear DNA sequences

Accurate delimitation of species is a critical first step in protecting biodiversity. Detection of distinct species is especially important for groups of organisms that inhabit sensitive environments subject to recent degradation, such as creeks, springs, and rivers in arid or semi-desert regions. The genus Dionda currently includes six recognized and described species of minnows that live in clear springs and spring-fed creeks of Texas, New Mexico (USA), and northern Mexico, but the boundaries, delimitation, and characterization of species in this genus have not been examined rigorously. The habitats of some of the species in this genus are rapidly deteriorating, and many local populations of Dionda have been extirpated. Considering the increasing concerns over degradation of their habitat, and pending a more detailed morphological revision of the genus, we undertook a molecular survey based on four DNA regions to examine variation over the range of the genus, test species boundaries, and infer phylogenetic relationships within Dionda. Based on analyses of two mitochondrial (cytb and D-loop) and two nuclear (Rag1 and S7) DNA regions from specimens collected throughout the range of Dionda, we identified 12 distinct species in the genus. Formerly synonymized names are available for two of these species, and four other species remain undescribed. We also redefine the known range of six species. The limited distribution of several of the species, coupled with widespread habitat degradation, suggests that many of the species in this genus should be targets for conservation and recovery efforts.     

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Morphologic comparisons of hatchery-reared specimens of Scaphirhynchus albus, Scaphirhynchus platorynchus, and S. albus × S. platorynchus hybrids (Acipenseriformes: Acipenseridae)

Extensive habitat modifications within the Mississippi and Missouri rivers have presumably interfered with the reproductive isolating mechanisms between the endangered pallid sturgeon, Scaphirhynchus albus, and the sympatric shovelnose sturgeon, Scaphirhynchus platorynchus, causing hybridization between these two species. Several character indices were developed to assist fisheries biologists in identifying specimens of S. albus, S. platorynchus, and their putative hybrids. The indices have numerous assumptions, including that pure strains of both parental species are within the sample analyzed and that hybrids are morphologically intermediate relative to their parents. If these indices have produced inaccurate identifications, then previous work on Scaphirhynchus studies in the Mississippi and Missouri rivers are questionable, including status surveys, captive propagation efforts, or the harvesting of tissues for genetic studies. In this study we tested indices by examining progeny of ‘known’ pallid, shovelnose, and hybrid sturgeon propagated, raised, and preserved at hatcheries. These 60 specimens [78–600 mm standard length (SL)] were propagated with breeding stock from the upper Missouri River drainage, where hybridization between these two species presumably does not occur. Existing indices did not correctly identify small (<250 mm SL) or a combination of small and large (>250 mm SL) sizes of S. albus, S. platorynchus, and their hybrids. Indices worked fairly well in identifying large S. platorynchus, but not in differentiating large S. albus from hybrids. We used principal components analysis (PCA) as an alternative approach to character indices. No a priori knowledge of the identity of the specimen is required with this multivariate technique, which avoids potential circular reasoning present in indices. We were able to completely or almost completely separate both sturgeon species and their hybrids by extracting principal components from a correlation matrix of 13 meristic characters in a standard PCA and extracting size‐corrected principal components from a covariance matrix of 51 morphometric variables using a sheared PCA. Additionally, we demonstrated that first generation hybrids were intermediate with respect to their parental species. Multivariate analyses with a reduced character set of six meristic and 12 morphometric variables also led to accurate and reliable specimen identification. Head spines and numerous qualitative characters are also extremely useful in differentiating between Scaphirhynchus species and their hybrids. In addition to all morphometric characters, some meristic characters and the degree of head spine fusion vary significantly with SL of sturgeons. Recording appropriate data from released specimens, including photovouchers, and making this information available is essential for researchers to have any scientific or legal basis for genetic or any other studies involving these sturgeons.