Simple technique for distinguishing Yellow‐bellied Flycatchers from Cordilleran and Pacific‐slope flycatchers

Flycatchers of the genus Empidonax are readily misidentified in the field, in the hand, and even in museum collections. We describe a novel plumage feature that can be used to distinguish Yellow‐bellied Flycatchers (E. flaviventris) from the two species that comprise the Western Flycatcher complex, Cordilleran Flycatchers (E. occidentalis) and Pacific‐slope Flycatchers (E. difficilis). The length of the buffy fringing on the anterior edge of each secondary feather, visible on the folded wing, is significantly shorter in Yellow‐bellied Flycatchers than in Western flycatchers, with minimal overlap. A definitive identification can be made using a simple formula that includes measurements of wing chord and the length of the buffy fringing along the outer edge of the first secondary (S1). This method provides definitive in‐hand identification, and the difference in length of the buffy fringing on the secondaries is also a useful field mark for visual identification. Testing our method with 113 museum specimens that had been identified a priori based on locality, we correctly identified 112 specimens. The exception was a specimen from Illinois that had been assumed to be a Yellow‐bellied Flycatcher. However, based on our formula, it was a Western flycatcher and analysis of its mtDNA sequence confirmed this result, proving the utility of our method.     

Fall migration of Empidonax flycatchers in northwestern Colombia

ABSTRACT South of Mexico, little is known about the fall migration patterns of most Neotropical migrants. I studied the migration of Empidonax flycatchers using mist‐net surveys in northwestern Colombia from late September to mid‐October in 4 yr (2003–2005, 2008). Empidonax species were identified using linear measurements and color patterns. About 62% of captured individuals were reliably identified to species, with 86% identified as Willow Flycatchers (Empidonax traillii) and 14% as Alder Flycatchers (E. alnorum). No Acadian Flycatchers (E. virescens) were identified. Most birds captured were adults (84.9%) and, due to overlap in measurements, I was only able to determine the sex of 16.3% of the birds. Most Empidonax flycatchers migrated through northwestern Colombia during September and October, with individuals migrating through my study area over a period of at least 1 mo. Willow Flycatchers tended to migrate earlier than Alder Flycatchers, a pattern consistent with the fall movements of these two species at other locations. No captured flycatchers were molting either remiges or rectrices, and most (89%) had either no or slight traces of subcutaneous fat. No Empidonax flycatchers were recaptured, suggesting that stopover duration at my study site was brief. My results show that many Empidonax flycatchers can be identified as Willow and Alder flycatchers during the nonbreeding period, and such identification will enhance our knowledge of their geographical distribution and improve our understanding of possible patterns of segregation on their wintering grounds.     

Polymorphic microsatellite loci for assigning parentage in least flycatchers (Empidonax minimus)

Least flycatchers (Empidonax minimus) are socially monogamous birds that form tight territorial aggregations on the breeding grounds. We designed five polymorphic microsatellite loci for assigning parentage to offspring within least flycatcher clusters. The number of alleles per locus ranged from 7 to 18. Mean polymorphic information content was 83.8%; the probability of exclusion with known maternal genotype was 0.999. These microsatellites are powerful DNA markers for identifying extra‐pair paternity in this species. Preliminary data also suggest that these loci may be useful for other members of this genus.     

Social stimulation of dawn singing in Dusky Flycatchers: a serendipitous experiment

Hypotheses regarding dawn singing in birds remain largely unsupported by quantitative data, especially for suboscine passerines (suborder Tyranni). During a study of the singing behavior of a suboscine, the Dusky Flycatcher (Empidonax oberholseri), in Alberta, Canada, spring storms in 2002 caused the disappearance and presumed mortality of several territorial males and some of their replacements, creating a serendipitous experiment. Overall, 15 males disappeared and, as a result, the number of territorial males in our study area declined from 13 prior to the storms to six afterward. Only one male maintained the same territory throughout the breeding season. During the period of inclement weather and social instability (20 May to 9 June), males sang at high rates during the predawn period. From 10 to 19 June, following this period of unstable weather and turnover in territorial males, males began dawn singing later in the morning, and sang shorter bouts at lower rates. The proportion of males engaging in dawn singing also decreased between the two periods. In contrast, daytime singing activity of paired males was low during both time periods. Playback of dawn songs to territorial males between 20 June and 21 July caused a resumption of some dawn singing. Singing rates were higher on the day of playback than on the day before playback, and the times when dawn singing was initiated were earlier on each of 2 d after playback than on the day before playback. In addition, the proportion of males engaging in dawn singing increased between the day before and the day after playback. Both the decrease in dawn singing when population density was reduced and its partial restoration by playback suggest that dawn singing in this species functions in male–male interactions and support the social‐dynamics hypothesis as an explanation for the dawn chorus in this species.     

DNA barcoding of Oryx leucoryx using the mitochondrial cytochrome C oxidase gene

The massive destruction and deterioration of the habitat of Oryx leucoryx and illegal hunting have decimated Oryx populations significantly, and now these animals are almost extinct in the wild. Molecular analyses can significantly contribute to captive breeding and reintroduction strategies for the conservation of this endangered animal. A representative 32 identical sequences used for species identification through BOLD and GenBank/NCBI showed maximum homology 96.06% with O. dammah, which is a species of Oryx from Northern Africa, the next closest species 94.33% was O. gazella, the African antelope. DNA barcode sequences of the mitochondrial cytochrome C oxidase (COI) gene were determined for O. leucoryx; identification through BOLD could only recognize the genus correctly, whereas the species could not be identified. This was due to a lack of sequence data for O. leucoryx on BOLD. Similarly, BLAST analysis of the NCBI data base also revealed no COI sequence data for the genus Oryx.     

Identifying gastropod spawn from DNA barcodes: possible but not yet practicable

Identifying life stages of species with complex life histories is problematic as species are often only known and/or described from a single stage. DNA barcoding has been touted as an important tool for linking life-history stages of the same species. To test the current efficacy of DNA barcodes for identifying unknown mollusk life stages, 24 marine gastropod egg capsules were collected off the Philippines in deep water and sequenced for partial fragments of the COI, 16S and 12S mitochondrial genes. Two egg capsules of known shallow-water Mediterranean species were used to calibrate the method. These sequences were compared to those available in GenBank and the Barcode of Life Database (BOLD). Using COI sequences alone, only a single Mediterranean egg capsule was identified to species, and a single Philippine egg capsule was identified tentatively to genus; all other COI sequences recovered matches between 76% and 90% with sequences from BOLD and GenBank. Similarity-based identification using all three markers confirmed the Mediterranean specimens' identifications. A phylogenetic approach was also implemented to confirm similarity-based identifications and provide a higher-taxonomic identification when species-level identifications were not possible. Comparison of available GenBank sequences to the diversity curve of a well-sampled coral reef habitat in New Caledonia highlights the poor taxonomic coverage achieved at present in existing genetic databases, emphasizing the need to develop DNA barcoding projects for megadiverse and often taxonomically challenging groups such as mollusks, to fully realize its potential as an identification and discovery tool.     

Exploring the molecular diversity of Eleotridae (Gobiiformes) using mitochondrial DNA

Species with wide geographic distributions can consist of different lineages or even of different species. The present study evaluated the molecular diversity of the family Eleotridae by analyzing sequences of the COI gene from species collected in different locations in a Neotropical region as well as sequences from BOLD and GenBank. The analysis revealed the distinction and differentiation of a variety of species with high statistical support, including Gobiomorus dormitor (two lineages), Dormitator maculatus (three lineages) and Eleotris fusca (three lineages), and a number of different lineages of Microphilypnus. This indicates the existence of a number of unknown cryptic species that have not been reported previously, increasing the potential diversity of eleotrid species in the Neotropics.     

Mitochondrial DNA sequence-based phylogenetic relationship among flesh flies of the genus <Emphasis Type="Italic">Sarcophaga</Emphasis> (Sarcophagidae: Diptera)

The phylogenetic relationships among flesh flies of the family Sarcophagidae has been based mainly on the morphology of male genitalia. However, the male genitalic character-based relationships are far from satisfactory. Therefore, in the present study mitochondrial DNA has been used as marker to unravel genetic relatedness and to construct phylogeny among five sympatric species of the genus Sarcophaga. Two mitochondrial genes viz., cytochrome oxidase subunit 1 (COI) and NAD dehydrogenase subunit 5 (ND5) were sequenced and genetic distance values were calculated on the basis of sequence differences in both the mitochondrial genes. The data revealed very few genetic difference among the five species for the COI and ND5 gene sequences.     

Complete sequence and gene organization of the mitochondrial genome of scaly-sided merganser (<Emphasis Type="Italic">Mergus squamatus</Emphasis>) and phylogeny of some Anatidae species

The scaly-sided merganser (Mergus squamatus) is an endangered bird species on the IUCN Red List with the estimated global population of less than 2,500 individuals at present. In the present study, we studied the complete mitochondrial genome (mtDNA) and the phylogenetic of M. squamatus by PCR amplification and GenBank data. The genome was 16,595 bp in length and contained 37 genes (13 protein coding genes, two rRNAs, and 22 tRNAs) and a non-coding control region (D-loop). All protein-coding genes of M. squamatus mtDNA start with a typical ATG codon, except ND1, COI, and COII uses GTG as their initial codon. TAA, T- and TAG as the terminate codon occurred very commonly in the sequence. All tRNA genes can be folded into canonical cloverleaf secondary structure except for tRNA^Ser (AGY) and tRNA^Leu (CUN), which lose ‘‘DHU’’ arm. The genome sequences had been deposited in GenBank under accession number HQ833701. Based on the concatenated nucleotide sequences of mtDNA genes (Cyt b and D-loop), we reconstructed phylogenetic trees and discussed the phylogenetic relationships among ten Anatidae species. The results are different from the present classification, and we support Lophodytes cucullatus and Mergullus albellus to be members of the genus Mergus.     

The role of ecologic diversification in sibling speciation of Empidonax flycatchers (Tyrannidae): multigene evidence from mtDNA

Avian genera characterized by sibling species with distinctive habitat preferences present an evolutionary enigma in view of the more commonplace occurrence of syntopic congeners that differ strikingly in colour and pattern. No existing theory has explained the evolutionary background that led to these differences. Here we propose that great phenotypic similarity among some groups of sibling species limits their coexistence and that clues to their radiation can be seen in patterns of geographical occurrence. To illustrate our thesis we focused on the New World flycatcher genus Empidonax, a group of 15 species notorious for their great phenotypic similarity. Using 3069 base pairs of mitochondrial DNA from four genes, we produced a complete molecular phylogeny that identified four clades, three of which represent close relatives. The fourth clade includes only E. virescens, which apparently has no close living relatives. The majority of species, including many distant relatives, are completely (58.1%) or essentially (6.7%) allopatric in breeding distribution and exhibit striking ecological segregation into distinctive climate-vegetation zones. Even where ranges overlap, occupancy of the same habitat by different species is rare. Phylogenetic and distributional patterns in Empidonax suggest a peripatric model of stepwise colonization and then range expansion of small groups of pioneers during glacial periods into initially enlarging, distinctive habitats destined to be widespread during interglacials. Vicariance is not indicated in the absence of barriers of appropriate age and geographical position. Rapoport's rule that northern species have larger ranges than southern species is strongly supported.     

Systematics of Pipizini and taxonomy of European Pipiza Fallén: molecular and morphological evidence (Diptera,Syrphidae)

Vuji?, A., Ståhls, G., A?anski, J., Bartsch, H., Bygebjerg, R. & Stefanovi?, A. (2013). Systematics of Pipizini and taxonomy of European Pipiza Fallén: molecular and morphological evidence (Diptera, Syrphidae). —Zoologica Scripta, 42, 288–305. In the present work the monophyly and molecular phylogenetic relationships of the genera of tribe Pipizini (Syrphidae) were investigated based on mitochondrial cytochrome c oxidase subunit I (COI) and nuclear 28S rDNA sequences, and the relationships among species of genus Pipiza Fallén, 1810 based on mtDNA COI sequences. Molecular phylogenetic analyses of Pipizini supported Pipiza as monophyletic and as sister group to all other Pipizini, and resolved other Pipizini genera as monophyletic lineages except for genus Heringia Rondani, 1856. To recognize the distinctness and maintain the monophyly the genus Heringia was redefined, generic rank was assigned to Neocnemodon Goffe, 1944 stat. n., and the genus Claussenia Vuji? & Ståhls gen. n., type‐species Claussenia hispanica (Strobl, 1909), was described. A revision of the European Pipiza species, including a discussion of taxonomic characters and a morphological redefinition of all included species, is presented. One new species, Pipiza laurusi Vuji? & Ståhls sp. n. was described. The taxa Pipiza carbonaria Meigen, 1822; Pipiza fasciata, Meigen 1822; Pipiza lugubris (Fabricius, 1775), Pipiza noctiluca (Linneaues, 1758), Pipiza notata Meigen, 1822 were redefined. Lectotypes are designated for 17 taxa, and neotypes were designated for seven taxa. Fourteen new synonymies were proposed. Male genitalia were illustrated for all the species, and a key of the 12 European species for males and females was provided. Geometric morphometrics of wing landmarks and extended sampling of mtDNA COI sequences was employed to delimitate taxa of the P. noctiluca and P. lugubris complexes. Despite subtle morphological differences, wing geometric morphometrics variables of wing size and shape showed highly significant differences among species within P. noctiluca and P. lugubris complexes, which were supported by the molecular data.     

Efficacy of using DNA barcoding to identify parasitoid wasps of the melon-cotton aphid (<Emphasis Type="Italic">Aphis gossypii</Emphasis>) in watermelon cropping system

Parasitoid wasps have received a great deal of attention in the biological control of melon-cotton aphid (Aphis gossypii Glover). The species of parasitoids are often difficult to identify because of their small body size and profound diversity. DNA barcoding offers scientists who are not expert taxonomists a powerful tool to render their field studies more accurate. Using DNA barcodes to identify aphid parasitoid wasps in specific cropping systems may provide valuable information for biological control. Here, we report the use of DNA barcoding to confirm the morphological identification of 14 species (belonging to 13 genera of 7 families) of parasitoid wasps from two-year field samples in a watermelon cropping system. We generated DNA sequences from the mitochondrial COI gene and the nuclear D2 region of 28S rDNA to assess the genetic variation within and between parasitoid species. Automatic Barcode Gap Discovery (ABGD) supported the presence of 14 genetically distinct groups in the dataset. Among the COI sequences, we found no overlap between the maximum K2P distance within species (0.49%) and minimum distance between species (6.85%). The 28S sequences also showed greater interspecific distance than intraspecific distance. DNA barcoding confirmed the morphological identification. However, inconsistency and ambiguity of taxonomic information available in the online databases has limited the successful use of DNA barcoding. Only five species matched those in the BOLD and GenBank. Four species did not match the entries in GenBank and five species showed ambiguous results in BOLD due to confusing nomenclature. We suggested that species identification based on DNA barcodes should be performed using both COI and other genes. Nonetheless, we demonstrate the potential of the DNA barcoding approach to confirm field identifications and to provide a foundation for studies aimed at improving the understanding of the biocontrol services provided by parasitoids in the melon ecosystem.     

Combined molecular and morphological phylogenetic analyses of the New Zealand wolf spider genus Anoteropsis (Araneae: Lycosidae)

Datasets from the mitochondrial gene regions NADH dehydrogenase subunit I (ND1) and cytochrome c oxidase subunit I (COI) of the 20 species in the New Zealand wolf spider (Lycosidae) genus Anoteropsis were generated. Sequence data were phylogenetically analysed using parsimony and maximum likelihood analyses. The phylogenies generated from the ND1 and COI sequence data and a previously generated morphological dataset were significantly congruent (p<0.001). Sequence data were combined with morphological data and phylogenetically analysed using parsimony. The ND1 region sequenced included part of tRNA^Leu(CUN), which appears to have an unstable amino-acyl arm and no TψC arm in lycosids. Analyses supported the existence of five species groups within Anoteropsis and the monophyly of species represented by multiple samples. A radiation of Anoteropsis species within the last five million years is inferred from the ND1 and COI likelihood phylograms, habitat and geological data, which also indicates that Anoteropsis arrived in New Zealand some time after it separated from Gondwana.     

Utility of GenBank and the Barcode of Life Data Systems (BOLD) for the identification of forensically important Diptera from Belgium and France

Fly larvae living on dead corpses can be used to estimate post-mortem intervals. The identification of these flies is decisive in forensic casework and can be facilitated by using DNA barcodes provided that a representative and comprehensive reference library of DNA barcodes is available.We constructed a local (Belgium and France) reference library of 85 sequences of the COI DNA barcode fragment (mitochondrial cytochrome c oxidase subunit I gene), from 16 fly species of forensic interest (Calliphoridae, Muscidae, Fanniidae). This library was then used to evaluate the ability of two public libraries (GenBank and the Barcode of Life Data Systems – BOLD) to identify specimens from Belgian and French forensic cases. The public libraries indeed allow a correct identification of most specimens. Yet, some of the identifications remain ambiguous and some forensically important fly species are not, or insufficiently, represented in the reference libraries. Several search options offered by GenBank and BOLD can be used to further improve the identifications obtained from both libraries using DNA barcodes.     

Molecular phylogenetic analysis of all members of the genus Elminia confirms their presence within the Stenostiridae clade

The genus Elminia has had a jumbled taxonomic history, being placed among ‘old world flycatchers’ or ‘monarch flycatchers’, where it was for a long time lumped with Trochocercus. It was recently suggested that it might represent a deep clade in the large sylvioid radiation. Using one mitochondrial protein‐coding gene (ND2, 1041 bp) and one nuclear intron (myoglobin intron 2, 700 bp) DNA sequences, we obtained robust evidence for the phylogenetic placement of Elminia in the new family Stenostiridae, which is strongly supported by a synapomorphic insertion of one base in the nuclear myoglobin intron 2 sequence. Our analyses confirm the monophyly of Elminia and resolve relationships within this genus, but cannot confidently identify its sister‐taxon within the stenostirid clade. Two clades were strongly supported within the genus Elminia: one with the two fairy blue flycatchers and another with the three white‐tailed crested‐flycatchers. Within the first clade, Elminia longicauda appears non‐monophyletic but remains strongly related to E. albicauda. In the second clade, E. albiventris is sister to E. albonotata while the Dusky Crested Flycatcher (E. nigromitrata) appears in a basal position within this clade. According to our molecular dating, several geological events in western Africa and the Albertine Rift area seem to be related to the historical distribution of Elminia. Thus, the differentiation between E. albonotata and E. albiventris could be directly related to the tectonic history of these two regions. According to our molecular dating, at least one intercontinental dispersal event involving Culicicapa took place within the Stenostiridae clade at a time when the Middle East was forested.     

An update on DNA barcoding: low species coverage and numerous unidentified sequences

DNA barcoding was proposed in 2003, the Consortium for the Barcode of Life was established in 2004, and the movement has since attracted more than $80 million funding. Here we investigate how many species of multicellular animals have been barcoded. We compare the numbers in a public database (GenBank as of January 2012) with those in the Barcode of Life Database (BOLD) and find that GenBank contains COI (cytochrome c oxidase subunit 1) sequences for ca. 60 000 species while BOLD reports barcodes for ca. 150 000 species. The discrepancy is likely due to a large amount of unpublished data in BOLD. Overall, the species coverage remains sparse, growth rates are low, and the barcode accumulation curve for Metazoa is linear with only 4788 species having been added in 2011. In addition, the vast majority of species in the public database (73%) were barcoded by projects that are unlikely to be related to the DNA barcoding movement. Particularly surprising was the large number of DNA barcodes in GenBank that were not identified to species (Jan 2012: 74%), with insect barcodes often being identified only to order. Of these several hundred thousand have since been suppressed by NCBI because they did not satisfy the iBOL/GenBank early release agreement. Species coverage is considerably better for target taxa of DNA barcoding campaigns (e.g. birds, fishes, Lepidoptera), although it also falls short of published campaign targets. © The Willi Hennig Society 2012     

Genetic diversity and phylogenetic relationship among the western and the Asian honey bees based on two mitochondrial gene segments (COI and ND5)

The Asian honey bee species i.e., Apis cerana (the eastern honey bee), A. dorsata (the giant honey bee), and the western or European honey bee (A. mellifera) collected from Pakistan were studied using partial sequences from two mitochondrial genes (i) the Cytochrome c oxidase I (COI) and (ii) the mitochondrially encoded NADH dehydrogenase 5 (ND5) and then compared with other honey bees sequences (already submitted from different countries around the globe) obtained after the national center for biotechnology information (NCBI). DNA sequences were analyzed employing molecular evolutionary genetics analysis and Kimura 2-parameter model, neighbor-joining method was applied to investigate phylogenetic relationships, and DNA sequence polymorphism was applied to measure the genetic diversity within the genus Apis. The phylogenetic analyses yielded consistent results. Based on COI gene fragment in two Asian and European honey bee species from Pakistan and from other countries showed considerable genetic diversity levels and deviation among the species. While in contrast the phylogenetic analyses based on ND5 gene fragment in Asian and European honey bee species from Pakistan and other countries showed comparatively higher genetic diversity indices and variations than the COI gene. So, in the genus Apis, the mitochondrial ND5 region has shown the possibility to answer the interactions among species. A further detailed work (by linking the analysis of other genomic and mitochondrial genes) is required for good quality solution to establish the concise genetic diversity and interaction among the Apis species. The objective of this study was to explore the extent of genetic differences and phylogenetic links among the three kinds of honey bee species from Pakistan and comparing them with other bee species around the globe.     

Southwestern Willow Flycatcher Population and Habitat Response to Reservoir Inundation

ABSTRACT One of the largest known populations of the federally endangered southwestern willow flycatcher (Empidonax traillii extimus) occurs at Roosevelt Lake, Arizona, USA. Modifications to Roosevelt Dam, completed in 1996, raised the height of the dam and resulted in a high probability of willow flycatcher habitat inundation within the reservoir's conservation pool. We collected habitat measurements and monitored 922 willow flycatcher nests from 1996 to 2006 to investigate effects of inundation on willow flycatcher habitat and subsequent changes in nest success, productivity, and distribution. Inundation of willow flycatcher habitat at Roosevelt Lake occurred in 2005, changing the location and amount of suitable breeding habitat and significantly altering habitat structure (e.g., thinner vegetation, more canopy gaps) of formally occupied nest sites. The willow flycatcher population at Roosevelt Lake decreased 47% from 209 territories in 2004 to 111 territories in 2006 in response to habitat changes. Willow flycatchers made fewer nesting attempts and nest success rates were significantly lower during inundation (2005 and 2006: 45%) than preinundation (1996–2004: 57%). Combined, these factors negatively affected the population's productivity during inundation. Although inundation caused extensive vegetation die-off, we did observe regeneration of vegetation in some areas at Roosevelt Lake in 2006. The Roosevelt Lake population remains one of the largest willow flycatcher populations in the state and territory numbers remain high enough that the population may not suffer long-term effects if sufficient suitable habitat continues to exist during the cycle of inundation and regeneration. Reservoir managers may be able to develop dam management guidelines that reduce damage to habitat, encourage habitat growth, and mimic the dynamic nature of unaltered riparian habitat. These guidelines can be implemented, as appropriate, at reservoirs throughout the willow flycatcher's range.     

DNA barcodes successfully delimit morphospecies in a superdiverse insect genus

Polypedilum Kieffer (Diptera: Chironomidae), with 520 currently known species worldwide, can be extremely difficult to identify species level based on the morphology. We used 3,670 cytochrome c oxidase subunit I (COI) barcodes to explore the efficiency of the COI barcodes to differentiate between species in a superdiverse aquatic insect genus. The Barcode of Life Data System (BOLD) presented 286 BIN clusters in Polypedilum, representing 163 morphospecies, of which 93 were contributed from our laboratory. Molecular operational taxonomic units (OTUs) ranged from 158 to 345, based on Automatic Barcode Gap Discovery (ABGD), the Barcode Index Number (BIN), Bayesian Poisson tree processes (bPTP), generalized mixed Yule coalescent (GMYC), jMOTU, multi‐rate Poisson tree processes (mPTP), neighbor‐joining (NJ) tree and prethreshold clustering. In comparison, GMYC, bPTP, mPTP and BIN suggested more species than warranted by morphology, while ABGD, jMOTU, NJ, prethreshold clustering and ABGD yielded a conservative number of species when setting higher thresholds. Nine species complexes with deep intraspecific divergences indicated 18 potentially cryptic species, which require further taxonomic research including complete life histories and nuclear genetic data to be resolved. The discrimination of Polypedilum species by DNA barcodes proved to be successful in 94.4% of all studied morphological species.     

DNA Barcoding for Species Assignment: The Case of Mediterranean Marine Fishes

Background

DNA barcoding enhances the prospects for species-level identifications globally using a standardized and authenticated DNA-based approach. Reference libraries comprising validated DNA barcodes (COI) constitute robust datasets for testing query sequences, providing considerable utility to identify marine fish and other organisms. Here we test the feasibility of using DNA barcoding to assign species to tissue samples from fish collected in the central Mediterranean Sea, a major contributor to the European marine ichthyofaunal diversity.

Methodology/Principal Findings

A dataset of 1278 DNA barcodes, representing 218 marine fish species, was used to test the utility of DNA barcodes to assign species from query sequences. We tested query sequences against 1) a reference library of ranked DNA barcodes from the neighbouring North East Atlantic, and 2) the public databases BOLD and GenBank. In the first case, a reference library comprising DNA barcodes with reliability grades for 146 fish species was used as diagnostic dataset to screen 486 query DNA sequences from fish specimens collected in the central basin of the Mediterranean Sea. Of all query sequences suitable for comparisons 98% were unambiguously confirmed through complete match with reference DNA barcodes. In the second case, it was possible to assign species to 83% (BOLD-IDS) and 72% (GenBank) of the sequences from the Mediterranean. Relatively high intraspecific genetic distances were found in 7 species (2.2%–18.74%), most of them of high commercial relevance, suggesting possible cryptic species.

Conclusion/Significance

We emphasize the discriminatory power of COI barcodes and their application to cases requiring species level resolution starting from query sequences. Results highlight the value of public reference libraries of reliability grade-annotated DNA barcodes, to identify species from different geographical origins. The ability to assign species with high precision from DNA samples of disparate quality and origin has major utility in several fields, from fisheries and conservation programs to control of fish products authenticity.