The Complete Mitochondrial Genomes of Two Octopods Cistopus chinensis and Cistopus taiwanicus: Revealing the Phylogenetic Position of the Genus Cistopus within the Order Octopoda

In the present study, we determined the complete mitochondrial DNA (mtDNA) sequences of two species of Cistopus, namely C. chinensis and C. taiwanicus, and conducted a comparative mt genome analysis across the class Cephalopoda. The mtDNA length of C. chinensis and C. taiwanicus are 15706 and 15793 nucleotides with an AT content of 76.21% and 76.5%, respectively. The sequence identity of mtDNA between C. chinensis and C. taiwanicus was 88%, suggesting a close relationship. Compared with C. taiwanicus and other octopods, C. chinensis encoded two additional tRNA genes, showing a novel gene arrangement. In addition, an unusual 23 poly (A) signal structure is found in the ATP8 coding region of C. chinensis. The entire genome and each protein coding gene of the two Cistopus species displayed notable levels of AT and GC skews. Based on sliding window analysis among Octopodiformes, ND1 and DN5 were considered to be more reliable molecular beacons. Phylogenetic analyses based on the 13 protein-coding genes revealed that C. chinensis and C. taiwanicus form a monophyletic group with high statistical support, consistent with previous studies based on morphological characteristics. Our results also indicated that the phylogenetic position of the genus Cistopus is closer to Octopus than to Amphioctopus and Callistoctopus. The complete mtDNA sequence of C. chinensis and C. taiwanicus represent the first whole mt genomes in the genus Cistopus. These novel mtDNA data will be important in refining the phylogenetic relationships within Octopodiformes and enriching the resource of markers for systematic, population genetic and evolutionary biological studies of Cephalopoda.     

Monophyly of clade III nematodes is not supported by phylogenetic analysis of complete mitochondrial genome sequences

Background

The orders Ascaridida, Oxyurida, and Spirurida represent major components of zooparasitic nematode diversity, including many species of veterinary and medical importance. Phylum-wide nematode phylogenetic hypotheses have mainly been based on nuclear rDNA sequences, but more recently complete mitochondrial (mtDNA) gene sequences have provided another source of molecular information to evaluate relationships. Although there is much agreement between nuclear rDNA and mtDNA phylogenies, relationships among certain major clades are different. In this study we report that mtDNA sequences do not support the monophyly of Ascaridida, Oxyurida and Spirurida (clade III) in contrast to results for nuclear rDNA. Results from mtDNA genomes show promise as an additional independently evolving genome for developing phylogenetic hypotheses for nematodes, although substantially increased taxon sampling is needed for enhanced comparative value with nuclear rDNA. Ultimately, topological incongruence (and congruence) between nuclear rDNA and mtDNA phylogenetic hypotheses will need to be tested relative to additional independent loci that provide appropriate levels of resolution.

Results

For this comparative phylogenetic study, we determined the complete mitochondrial genome sequences of three nematode species, Cucullanus robustus (13,972 bp) representing Ascaridida, Wellcomia siamensis (14,128 bp) representing Oxyurida, and Heliconema longissimum (13,610 bp) representing Spirurida. These new sequences were used along with 33 published nematode mitochondrial genomes to investigate phylogenetic relationships among chromadorean orders. Phylogenetic analyses of both nucleotide and amino acid sequence datasets support the hypothesis that Ascaridida is nested within Rhabditida. The position of Oxyurida within Chromadorea varies among analyses; in most analyses this order is sister to the Ascaridida plus Rhabditida clade, with representative Spirurida forming a distinct clade, however, in one case Oxyurida is sister to Spirurida. Ascaridida, Oxyurida, and Spirurida (the sampled clade III taxa) do not form a monophyletic group based on complete mitochondrial DNA sequences. Tree topology tests revealed that constraining clade III taxa to be monophyletic, given the mtDNA datasets analyzed, was a significantly worse result.

Conclusion

The phylogenetic hypotheses from comparative analysis of the complete mitochondrial genome data (analysis of nucleotide and amino acid datasets, and nucleotide data excluding 3^rd positions) indicates that nematodes representing Ascaridida, Oxyurida and Spirurida do not share an exclusive most recent common ancestor, in contrast to published results based on nuclear ribosomal DNA. Overall, mtDNA genome data provides reliable support for nematode relationships that often corroborates findings based on nuclear rDNA. It is anticipated that additional taxonomic sampling will provide a wealth of information on mitochondrial genome evolution and sequence data for developing phylogenetic hypotheses for the phylum Nematoda.

     

A reconsideration of Gallicolumba (Aves: Columbidae) relationships using fresh source material reveals pseudogenes,chimeras, and a novel phylogenetic hypothesis

Phylogenetic analyses of nuclear and mitochondrial DNA sequences were used to test a recent phylogenetic hypothesis for the avian genus Gallicolumba that relied heavily on preserved museum study skins for DNA samples. Our comparisons show that several published gene sequences represent pseudogenes or chimeras of multiple taxa, and these sequences had an adverse influence on phylogeny estimation. A new phylogenetic hypothesis strongly supports the monophyly of Philippine bleeding-hearts, the monophyly of Gallicolumba sensu stricto, and places Leucosarcia as part of an Australasian radiation rather than sister to Alopecoenas. These findings clarify Gallicolumba biogeography and also highlight the need for greater caution in the use of suboptimal source material in molecular systematic studies.     

Y DNA and mitochondrial lineages in European and Asian populations of the brown hare (Lepus europaeus)

Both the Cytb gene of mtDNA and Y chromosome markers were studied in a relatively large sample of brown hares (L. europaeus) from Europe and Anatolia (Turkey and Israel), together with other seven Lepus species, in order to enable comparative analysis of possible sex-specific gene flow. Furthermore, Y chromosome markers were compared with data from biparentally inherited markers in an attempt to understand whether or not their pattern of distribution was congruent with that of allozymes or whether they rather matched mtDNA phylogenies, with which they share uniparental inheritance. Consistent with the general observation, levels of interspecific genetic variability were very low for the Y chromosome markers compared with mtDNA. Moreover, lack of interspecific variation for the Y-DNA studied within Lepus genus rendered these markers improper for any further phylogenetic analysis. With the highest nucleotide diversity in Anatolia compared with Europe, both marker systems confirmed an unbroken species history in Anatolia, corroborated the hypothesis of continuous gene flow from Anatolia's neighbouring regions, and supported the idea of a quick postglacial colonization followed by expansion of the species in large parts of Europe. Phylogenetic analysis under mtDNA revealed the existence of four different haplogroups with a well defined distribution across Europe and Anatolia. Both genetic systems supported the deep separation of Anatolian and European lineages of L. europaeus. Nevertheless, Anatolian Y-DNA lineages extended across a longer geographic distance in south-eastern Europe than Anatolian mtDNA haplotypes, probably as a result of higher female philopatry that makes mtDNA introgression more difficult in brown hares.     

Hybridization and skua phylogeny

The close similarity in mitochondrial DNA (mtDNA) between Pomarine and (North Atlantic) Great skuas, Stercorarius pomarinus and Stercorarius skua, has several possible explanations. Two of the main alternatives are stochastic lineage sorting of mtDNA and introgressive hybridization between S. pomarinus and S. skua. Here, a new probabilistic approach to testing these alternatives using different phylogenies based on external morphology, mtDNA and ectoparasites leads to rejection of the lineage sorting and corroboration of the hybridization hypothesis. If great skuas are a monophyletic group, as indicated by the present cladistic analysis based on morphology and by other evidence, the available molecular genetic data are as expected if mtDNA has been transferred from S. pomarinus to S. skua by introgressive hybridization. Further evidence from several unlinked regions of nuclear DNA is crucial for critical testing of the alternative hypotheses on skua evolution.     

Molecular phylogeny of Eurasian badgers (Meles) around the distribution boundaries,revealed by analyses of mitochondrial DNA and Y-chromosomal genes

Based on previous molecular and morphological analyses, Eurasian badgers are currently classified into Meles meles distributed in Europe, M. canescens in the Caucasus and Middle East, M. leucurus in continental Asia and M. anakuma in Japan. The precise locations of their distribution boundaries are still unclear. Therefore, in the present study, we clarified the phylogenetic relationships in the genus Meles around its distribution boundaries, based on mitochondrial DNA (mtDNA) and Y-chromosomal genes. From 71 badgers examined, 29 mtDNA haplotypes were identified, including new 26 types. Multiple haplotypes of SRY and CAN-SINEs were identified from 23 males, including two new haplotypes of SRY. The mtDNA phylogeny showed that the Ural Mountains were not a current distribution boundary between M. meles and M. leucurus. In addition, our results supported the hypothesis that the Bosphorus Strait is a geographical barrier between M. meles and M. canescens. The badgers from Bulgaria north of the Bosphorus Strait shared haplotypes with M. meles. On the other hand, badgers from Far Eastern Russia distributed in the eastern peripheral region of the Asian Continent had haplotypes of M. leucurus. The badgers from Norway and Finland shared haplotypes with M. meles.     

Effects of Asymmetric Nuclear Introgression,Introgressive Mitochondrial Sweep,and Purifying Selection on Phylogenetic Reconstruction and Divergence Estimates in the Pacific Clade of Locustella Warblers

When isolated but reproductively compatible populations expand geographically and meet, simulations predict asymmetric introgression of neutral loci from a local to invading taxon. Genetic introgression may affect phylogenetic reconstruction by obscuring topology and divergence estimates. We combined phylogenetic analysis of sequences from one mtDNA and 12 nuDNA loci with analysis of gene flow among 5 species of Pacific Locustella warblers to test for presence of genetic introgression and its effects on tree topology and divergence estimates. Our data showed that nuDNA introgression was substantial and asymmetrical among all members of superspecies groups whereas mtDNA showed no introgression except a single species pair where the invader''s mtDNA was swept by mtDNA of the local species. This introgressive sweep of mtDNA had the opposite direction of the nuDNA introgression and resulted in the paraphyly of the local species'' mtDNA haplotypes with respect to those of the invader. Тhe multilocus nuDNA species tree resolved all inter- and intraspecific relationships despite substantial introgression. However, the node ages on the species tree may be underestimated as suggested by the differences in node age estimates based on non-introgressing mtDNA and introgressing nuDNA. In turn, the introgressive sweep and strong purifying selection appear to elongate internal branches in the mtDNA gene tree.     

The first complete mitochondrial genome from Bostrychus genus (Bostrychus sinensis) and partitioned Bayesian analysis of Eleotridae fish phylogeny

To understand the phylogenetic position of Bostrychus sinensis in Eleotridae and the phylogenetic relationships of the family, we determined the nucleotide sequence of the mitochondrial (mt) genome of Bostrychus sinensis. It is the first complete mitochondrial genome sequence of Bostrychus genus. The entire mtDNA sequence was 16508 bp in length with a standard set of 13 protein-coding genes, 22 transfer RNA genes (tRNAs), two ribosomal RNA genes (rRNAs) and a noncoding control region. The mitochondrial genome of B. sinensis had common features with those of other bony fishes with respect to gene arrangement, base composition, and tRNA structures. Phylogenetic hypotheses within Eleotridae fish have been controversial at the genus level. We used the mitochondrial cytochrome b (cytb) gene sequence to examine phylogenetic relationships of Eleotridae by using partitioned Bayesian method. When the specific models and parameter estimates were presumed for partitioning the total data, the harmonic mean –lnL was improved. The phylogenetic analysis supported the monophyly of Hypseleotris and Gobiomorphs. In addition, the Bostrychus were most closely related to Ophiocara, and the Philypnodon is also the sister to Microphlypnus, based on the current datasets. Further, extensive taxonomic sampling and more molecular information are needed to confirm the phylogenetic relationships in Eleotridae.     

The giant Cafeteria roenbergensis virus that infects a widespread marine phagocytic protist is a new member of the fourth domain of Life

Background

A recent work has provided strong arguments in favor of a fourth domain of Life composed of nucleo-cytoplasmic large DNA viruses (NCLDVs). This hypothesis was supported by phylogenetic and phyletic analyses based on a common set of proteins conserved in Eukarya, Archaea, Bacteria, and viruses, and implicated in the functions of information storage and processing. Recently, the genome of a new NCLDV, Cafeteria roenbergensis virus (CroV), was released. The present work aimed to determine if CroV supports the fourth domain of Life hypothesis.

Methods

A consensus phylogenetic tree of NCLDVs including CroV was generated from a concatenated alignment of four universal proteins of NCLDVs. Some features of the gene complement of CroV and its distribution along the genome were further analyzed. Phylogenetic and phyletic analyses were performed using the previously identified common set of informational genes present in Eukarya, Archaea, Bacteria, and NCLDVs, including CroV.

Findings

Phylogenetic reconstructions indicated that CroV is clearly related to the Mimiviridae family. The comparison between the gene repertoires of CroV and Mimivirus showed similarities regarding the gene contents and genome organization. In addition, the phyletic clustering based on the comparison of informational gene repertoire between Eukarya, Archaea, Bacteria, and NCLDVs unambiguously classified CroV with other NCLDVs and clearly included it in a fourth domain of Life. Taken together, these data suggest that Mimiviridae, including CroV, may have inherited a common gene content probably acquired from a common Mimiviridae ancestor.

Conclusions

This further analysis of the gene repertoire of CroV consolidated the fourth domain of Life hypothesis and contributed to outline a functional pan-genome for giant viruses infecting phagocytic protistan grazers.     

The Complete Mitochondrial Genome of Aix galericulata and Tadorna ferruginea: Bearings on Their Phylogenetic Position in the Anseriformes

Aix galericulata and Tadorna ferruginea are two Anatidae species representing different taxonomic groups of Anseriformes. We used a PCR-based method to determine the complete mtDNAs of both species, and estimated phylogenetic trees based on the complete mtDNA alignment of these and 14 other Anseriforme species, to clarify Anseriform phylogenetics. Phylogenetic trees were also estimated using a multiple sequence alignment of three mitochondrial genes (Cyt b, ND2, and COI) from 68 typical species in GenBank, to further clarify the phylogenetic relationships of several groups among the Anseriformes. The new mtDNAs are circular molecules, 16,651 bp (Aix galericulata) and 16,639 bp (Tadorna ferruginea) in length, containing the 37 typical genes, with an identical gene order and arrangement as those of other Anseriformes. Comparing the protein-coding genes among the mtDNAs of 16 Anseriforme species, ATG is generally the start codon, TAA is the most frequent stop codon, one of three, TAA, TAG, and T-, commonly observed. All tRNAs could be folded into canonical cloverleaf secondary structures except for tRNASer (AGY) and tRNALeu (CUN), which are missing the "DHU" arm.Phylogenetic relationships demonstrate that Aix galericula and Tadorna ferruginea are in the same group, the Tadorninae lineage, based on our analyses of complete mtDNAs and combined gene data. Molecular phylogenetic analysis suggests the 68 species of Anseriform birds be divided into three families: Anhimidae, Anatidae, and Anseranatidae. The results suggest Anatidae birds be divided into five subfamilies: Anatinae, Tadorninae, Anserinae, Oxyurinae, and Dendrocygninae. Oxyurinae and Dendrocygninae should not belong to Anserinae, but rather represent independent subfamilies. The Anatinae includes species from the tribes Mergini, Somaterini, Anatini, and Aythyini. The Anserinae includes species from the tribes Anserini and Cygnini.     

Phylogenetic relationships of phrynosomatid lizards based on nuclear and mitochondrial data,and a revised phylogeny for Sceloporus

Phrynosomatid lizards are among the most common and diverse groups of reptiles in western North America, Mexico, and Central America. Phrynosomatidae includes 136 species in 10 genera. Phrynosomatids are used as model systems in many research programs in evolution and ecology, and much of this research has been undertaken in a comparative phylogenetic framework. However, relationships among many phrynosomatid genera are poorly supported and in conflict between recent studies. Further, previous studies based on mitochondrial DNA sequences suggested that the most species-rich genus (Sceloporus) is possibly paraphyletic with respect to as many as four other genera (Petrosaurus, Sator, Urosaurus, and Uta). Here, we collect new sequence data from five nuclear genes and combine them with published data from one additional nuclear gene and five mitochondrial gene regions. We compare trees from nuclear and mitochondrial data from 37 phrynosomatid taxa, including a “species tree” (from BEST) for the nuclear data. We also present a phylogeny for 122 phrynosomatid species based on maximum likelihood analysis of the combined data, which provides a strongly-supported hypothesis for relationships among most phrynosomatid genera and includes most phrynosomatid species. Our results strongly support the monophyly of Sceloporus (including Sator) and many of the relationships within it. We present a new classification for phrynosomatid lizards and the genus Sceloporus, and offer a new tree with branch lengths for use in comparative studies.     

Evidence of a secondary interspecific mitochondrial DNA introgression in the pond loach Misgurnus dabryanus (Teleostei: Cobitidae) population introduced in Japan

The pond loach Misgurnus dabryanus is a freshwater fish with a distribution range spanning the eastern part of the Asian continent, the Korean Peninsula, and Taiwan. The pond loach was transplanted to the Japanese archipelago through the co-inclusion with dojo loach (Misgurnus anguillicaudatus species complex) populations, which were imported live from China for food materials, and it is currently distributed widely across Japan. A previous mitochondrial DNA (mtDNA) analysis revealed that a pond loach population in Ehime Prefecture (Shikoku Island, Japan) included two highly diverged mtDNA groups (Groups I and II). To examine the origin of these two distinct forms of mtDNA within the Japanese pond loach population, we performed phylogenetic analyses using sequences based on the mtDNA of cytochrome oxidase b (cyt b) and the nuclear DNA recombination activating gene 1 (RAG-1). We also conducted a random amplified polymorphic DNA (RAPD) analysis to examine the establishment of reproductive isolation between sympatric pond loaches with two different mtDNA groups. Our mtDNA phylogenetic results indicated that the two diverged pond loach mtDNA sequences showed polyphyletic relationships among Misgurnus species and its related genus Cobitis. In contrast, there were no clear divergence in nuclear DNA among the pond loaches irrespective of their mtDNA groups, and they all formed monomorphic clades in the phylogenetic relationships among the species. The discrepancy between the mtDNA and nuclear DNA genes support that the existence of two diverged forms of DNA within the pond loach population could be attributed to past mtDNA introgressions from other species rather than convergent evolution. Previous mtDNA phylogenetic studies among Cobitidae revealed that the dojo loach also consisted of two genetically diverged polyphyletic clades: an original Misgurnus mtDNA and an introgressed mtDNA from Cobitis species. In our mtDNA result, the Group II haplotype of the pond loach was included in the mtDNA from the introgressed dojo loach. This suggested that the Group II haplotype was derived from introgressed dojo loach mtDNA. The close relationships between the introgressed dojo loach and the pond loach mtDNA indicated that this secondary introgression had recently occurred via hybridization in a recent artificial aquaculture or transportation process. Common RAG-1 alleles and RAPD bands were shared between the sympatric pond loaches with original and introgressed mtDNAs. This indicates that the introgressed mtDNA haplotype is included as one of the polymorphic genotypes within the pond loach populations, and does not represent existence of different cryptic species.     

Phylogeny of Rhigonematomorpha based on the complete mitochondrial genome of Rhigonema thysanophora (Nematoda: Chromadorea)

We determined the complete mitochondrial genome sequence of Rhigonema thysanophora, the first representative of Rhigonematomorpha, and used this sequence along with 57 other nematode species for phylogenetic analyses. The R. thysanophora mtDNA is 15 015 bp and identical to all other chromadorean nematode mtDNAs published to date in that it contains 36 genes (lacking atp8) encoded in the same direction. Phylogenetic analyses of nucleotide and amino acid sequence data for the 12 protein‐coding genes recovered Rhigonematomorpha as the sister group to the heterakoid species, Ascaridia columbae (Ascaridomorpha). The organization of R. thysanophora mtDNA resembles the most common pattern for the Rhabditomorpha+Ascaridomorpha+Diplogasteromorpha clade in gene order, but with some substantial gene rearrangements. This similarity in gene order is in agreement with the sequence‐based analyses that indicate a close relationship between Rhigonematomorpha and Rhabditomorpha+Ascaridomorpha+Diplogasteromorpha. These results are consistent with certain analyses of nuclear SSU rDNA for R. thysanophora and some earlier classification systems that asserted phylogenetic affinity between Rhigonematomorpha and Ascaridomorpha, but inconsistent with morphology‐based phylogenetic hypotheses that suggested a close (taxonomic) relationship between rhigonematomorphs and oxyuridomorphs (pinworms). These observations must be tempered by noting that few rhigonematomorph species have been sequenced and included in phylogenetic analyses, and preliminary studies based on SSU rDNA suggest the group is not monophyletic. Additional mitochondrial genome sequences of rhigonematids are needed to characterize their phylogenetic relationships within Chromadorea, and to increase understanding of mitochondrial genome evolution.     

Sperm ultrastructure of shrimps from the family Penaeidae (Crustacea: Dendrobranchiata) in a phylogenetic context

We describe the sperm ultrastructure of six penaeid species, including at least one member of each tribe (Penaeini, Parapenaeini and Trachypenaeini). Fragments of the vas deferens of the Penaeidae Farfantepenaeus brasiliensis, Farfantepenaeus paulensis, Litopenaeus schmitti, Parapenaeus americanus, Rimapenaeus constrictus and Xiphopenaeus kroyeri were fixed and processed according to the routine for transmission electron microscopy. The morphological results were contextualized in an evolutionary perspective using molecular markers for the phylogenetic reconstruction of this group. A phylogram was proposed by Bayesian inference based on 1007 bp of 33 sequences of the combined genes (16S rDNA and COI mtDNA) from 27 dendrobranchiate specimens. Our findings show that morphological differences in the sperm ultrastructures of members among the tribes of Penaeidae can be used as a baseline to understand their evolutionary relationships. Individuals from the Penaeini tribe show plesiomorphic characteristics in the sperm ultrastructure compared to the Trachypenaeini tribe from which they were derived, such as shrimp from family Sicyoniidae. The morphological complexity of the sperm of the different penaeid members corroborated with the genetic phylogeny, which showed different clades for each tribe and the close relationship with Sicyoniidae. The sperm features of the selected species studied here reflected their evolutionary history. These features confirm the previous phylogenetic hypothesis and question the monophyly of Penaeidae, which should be verified in the future with a more complete set of representative members of each tribe.     

The complete mitochondrial genome of Parasesarma pictum (Brachyura: Grapsoidea: Sesarmidae) and comparison with other Brachyuran crabs

Mitochondrial DNA (mtDNA) is an extrachromosomal genome which can provide important information for evolution and phylogenetic analysis. In this study, we assembled a complete mitogenome of a crab Parasesarma pictum (Brachyura: Grapsoidea: Sesarmidae) from next generation sequencing reads at the first time. P. pictum is a mudflat crab, belonging to the Sesarmidae family (subfamily Sesarminae), which is perched on East Asia. The 15,716 bp mitogenome covers 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs), 2 ribosomal RNA genes (rRNAs), and one control region (CR). The control region spanns 420 bp. The genome composition was highly A+T biased 75.60% and showed negative AT-skew (−0.03) and negative GC-skew (−0.47). Compared with the ancestor of Brachyura, the gene order of Sesarmidae has several differences and the gene order of P. pictum is typical for mitogenomes of Sesarmidae. Phylogenetic tree based on nucleotide sequences of mitochondrial 13 PCGs using BI and ML determined that P. pictum has a sister group relationship with Parasesarma tripectinis and belongs to Sesarmidae.     

Morphological and molecular differentiation of Parastrigea (Trematoda: Strigeidae) from Mexico,with the description of a new species

Parastrigea plataleae n. sp. (Digenea: Strigeidae) is described from the intestine of the roseate spoonbill Platalea ajaja (Threskiornithidae) from four localities on the Pacific coast of Mexico. The new species is mainly distinguished from the other 18 described species of Parastrigea based on the ratio of its hindbody length to forebody length. A principal component analysis (PCA) of 16 morphometric traits for 15 specimens of P. plataleae n. sp., five of Parastrigea cincta and 11 of Parastrigea diovadena previously recorded in Mexico, clearly shows three clusters, which correspond to the three species. DNA sequences of the internal transcribed spacers (ITSs) of ribosomal DNA and the mitochondrial gene cytochrome c oxidase subunit I (cox 1) were used to corroborate this morphological distinction. The genetic divergence estimated among P. plataleae n. sp., P. cincta and P. diovadena ranged from 0.5 to 1.48% for ITSs and from 9.31 to 11.47% for cox 1. Maximum parsimony (MP) and maximum likelihood (ML) analyses were performed on the combined datasets (ITSs + cox 1) and on each dataset alone. All of the phylogenetic analyses indicated that the specimens from the roseate spoonbill represent a clade with strong bootstrap support. The morphological evidence and the genetic divergence in combination with the reciprocal monophyly in all of the phylogenetic trees support the hypothesis that the digeneans found in the intestines of roseate spoonbills represent a new species.     

Genetic diversity of Garra rufa Heckel, 1843 (Teleostei: Cyprinidae) in Anatolia

In the present study, mitochondrial DNA polymerase chain reaction-restriction-fragment length polymorphism (PCR-RFLP) and nuclear DNA inter-simple sequence repeat (ISSR) assays were used to assess the phylogenetic and phylogeographic relationships among Garra rufa samples from Anatolia. The complete mtDNA NADH 3/4 dehydrogenase (ND-3/4) gene amplified by PCR was digested with eight restriction enzymes. These enzymes produced 20 composite haplotypes for G. rufa populations. All the mtDNA haplotypes detected were highly diverged from each other and each lineage had a unique genetic profile. The evaluation of mtDNA PCR-RFLP data coupled with geological history of Anatolia indicated a deep genetic divergence among the mtDNA haplotypes of G. rufa populations from drainages of the Mediterranean and Persian Gulf, suggesting an early isolation of Tigris-Euphrates with Orontes river and other rivers draining into the Mediterranean Sea. In general, data from both mtDNA and nDNA were congruent.     

Combining multiple autosomal introns for studying shallow phylogeny and taxonomy of Laurasiatherian mammals: Application to the tribe Bovini (Cetartiodactyla,Bovidae)

Mitochondrial sequences are widely used for species identification and for studying phylogenetic relationships among closely related species or populations of the same species. However, many studies of mammals have shown that the maternal history of the mitochondrial genome can be discordant with the true evolutionary history of the taxa. In such cases, the analyses of multiple nuclear genes can be more powerful for deciphering interspecific relationships.Here, we designed primers for amplifying 13 new exon-primed intron-crossing (EPIC) autosomal loci for studying shallow phylogeny and taxonomy of Laurasiatherian mammals. Three criteria were used for the selection of the markers: gene orthology, a PCR product length between 600 and 1200 nucleotides, and different chromosomal locations in the bovine genome. Positive PCRs were obtained from different species representing the orders Carnivora, Cetartiodactyla, Chiroptera, Perissodactyla and Pholidota.The newly developed markers were analyzed in a phylogenetic study of the tribe Bovini (the group containing domestic and wild cattle, bison, yak, African buffalo, Asian buffalo, and saola) based on 17 taxa and 18 nuclear genes, representing a total alignment of 13,095 nucleotides. The phylogenetic results were compared to those obtained from analyses of the complete mitochondrial genome and Y chromosomal genes. Our analyses support a basal divergence of the saola (Pseudoryx) and a sister-group relationship between yak and bison. These results contrast with recent molecular studies but are in better agreement with morphology. The comparison of pairwise nucleotide distances shows that our nuDNA dataset provides a good signal for identifying taxonomic levels, such as species, genera, subtribes, tribes and subfamilies, whereas the mtDNA genome fails because of mtDNA introgression and higher levels of homoplasy. Accordingly, we conclude that the genus Bison should be regarded as a synonym of Bos, with the European bison relegated to a subspecies rank within Bos bison. We compared our molecular dating estimates to the fossil record in order to propose a biogeographic scenario for the evolution of Bovini during the Neogene.     

Limited Phylogeographic Signal in Sex-Linked and Autosomal Loci Despite Geographically,Ecologically, and Phenotypically Concordant Structure of mtDNA Variation in the Holarctic Avian Genus Eremophila

Phylogeographic studies of Holarctic birds are challenging because they involve vast geographic scale, complex glacial history, extensive phenotypic variation, and heterogeneous taxonomic treatment across countries, all of which require large sample sizes. Knowledge about the quality of phylogeographic information provided by different loci is crucial for study design. We use sequences of one mtDNA gene, one sex-linked intron, and one autosomal intron to elucidate large scale phylogeographic patterns in the Holarctic lark genus Eremophila. The mtDNA ND2 gene identified six geographically, ecologically, and phenotypically concordant clades in the Palearctic that diverged in the Early - Middle Pleistocene and suggested paraphyly of the horned lark (E. alpestris) with respect to the Temminck''s lark (E. bilopha). In the Nearctic, ND2 identified five subclades which diverged in the Late Pleistocene. They overlapped geographically and were not concordant phenotypically or ecologically. Nuclear alleles provided little information on geographic structuring of genetic variation in horned larks beyond supporting the monophyly of Eremophila and paraphyly of the horned lark. Multilocus species trees based on two nuclear or all three loci provided poor support for haplogroups identified by mtDNA. The node ages calculated using mtDNA were consistent with the available paleontological data, whereas individual nuclear loci and multilocus species trees appeared to underestimate node ages. We argue that mtDNA is capable of discovering independent evolutionary units within avian taxa and can provide a reasonable phylogeographic hypothesis when geographic scale, geologic history, and phenotypic variation in the study system are too complex for proposing reasonable a priori hypotheses required for multilocus methods. Finally, we suggest splitting the currently recognized horned lark into five Palearctic and one Nearctic species.