Documenting decapod biodiversity in the Caribbean from DNA barcodes generated during field training in taxonomy

DNA barcoding is a useful tool to identify the components of mixed or bulk samples, as well as to determine individuals that lack morphologically diagnostic features. However, the reference database of DNA barcode sequences is particularly sparsely populated for marine invertebrates and for tropical taxa. We used samples collected as part of two field courses, focused on graduate training in taxonomy and systematics, to generate DNA sequences of the barcode fragments of cytochrome c oxidase subunit I (COI) and mitochondrial ribosomal 16S genes for 447 individuals, representing at least 129 morphospecies of decapod crustaceans. COI sequences for 36% (51/140) of the species and 16S sequences for 26% (37/140) of the species were new to GenBank. Automatic Barcode Gap Discovery identified 140 operational taxonomic units (OTUs) which largely coincided with the morphospecies delimitations. Barcode identifications (i.e. matches to identified sequences) were especially useful for OTUs within Synalpheus, a group that is notoriously difficult to identify and rife with cryptic species, a number of which we could not identify to species, based on morphology. Non-concordance between morphospecies and barcode OTUs also occurred in a few cases of suspected cryptic species. As mitochondrial pseudogenes are particularly common in decapods, we investigate the potential for this dataset to include pseudogenes and discuss the utility of these sequences as species identifiers (i.e. barcodes). These results demonstrate that material collected and identified during training activities can provide useful incidental barcode reference samples for under-studied taxa.

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Toward a global reference database of COI barcodes for marine zooplankton

Marine Biology ◽

10.1007/s00227-021-03887-y ◽

2021 ◽

Vol 168 (6) ◽

Author(s):

Ann Bucklin ◽

Katja T. C. A. Peijnenburg ◽

Ksenia N. Kosobokova ◽

Todd D. O’Brien ◽

Leocadio Blanco-Bercial ◽

...

Keyword(s):

Species Diversity ◽

Dna Sequences ◽

Reference Sequence ◽

Global Ocean ◽

Reference Database ◽

Data Repositories ◽

Marine Zooplankton ◽

The North ◽

Coi Sequences ◽

Taxonomic Groups

AbstractCharacterization of species diversity of zooplankton is key to understanding, assessing, and predicting the function and future of pelagic ecosystems throughout the global ocean. The marine zooplankton assemblage, including only metazoans, is highly diverse and taxonomically complex, with an estimated ~28,000 species of 41 major taxonomic groups. This review provides a comprehensive summary of DNA sequences for the barcode region of mitochondrial cytochrome oxidase I (COI) for identified specimens. The foundation of this summary is the MetaZooGene Barcode Atlas and Database (MZGdb), a new open-access data and metadata portal that is linked to NCBI GenBank and BOLD data repositories. The MZGdb provides enhanced quality control and tools for assembling COI reference sequence databases that are specific to selected taxonomic groups and/or ocean regions, with associated metadata (e.g., collection georeferencing, verification of species identification, molecular protocols), and tools for statistical analysis, mapping, and visualization. To date, over 150,000 COI sequences for ~ 5600 described species of marine metazoan plankton (including holo- and meroplankton) are available via the MZGdb portal. This review uses the MZGdb as a resource for summaries of COI barcode data and metadata for important taxonomic groups of marine zooplankton and selected regions, including the North Atlantic, Arctic, North Pacific, and Southern Oceans. The MZGdb is designed to provide a foundation for analysis of species diversity of marine zooplankton based on DNA barcoding and metabarcoding for assessment of marine ecosystems and rapid detection of the impacts of climate change.

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Overcoming limitations to environmental DNA studies: A coastal temperate reference sequence database for multiple chloroplast gene regions generated in a single assay.

10.22541/au.163252330.05592688/v1 ◽

2021 ◽

Author(s):

Nicole Foster ◽

Kor-jent Dijk ◽

Ed Biffin ◽

Jennifer Young ◽

Vicki Thomson ◽

...

Keyword(s):

Dna Sequences ◽

Dna Barcode ◽

Environmental Dna ◽

Reference Sequence ◽

Reference Database ◽

Chloroplast Gene ◽

Coastal Plants ◽

Reference Databases ◽

Targeted Capture ◽

Comprehensive Reference

A proliferation in environmental DNA (eDNA) research has increased the reliance on reference sequence databases to assign unknown DNA sequences to known taxa. Without comprehensive reference databases, DNA extracted from environmental samples cannot be correctly assigned to taxa, limiting the use of this genetic information to identify organisms in unknown sample mixtures. For animals, standard metabarcoding practices involve amplification of the mitochondrial Cytochrome-c oxidase subunit 1 (CO1) region, which is a universally amplifyable region across majority of animal taxa. This region, however, does not work well as a DNA barcode for plants and fungi, and there is no similar universal single barcode locus that has the same species resolution. Therefore, generating reference sequences has been more difficult and several loci have been suggested to be used in parallel to get to species identification. For this reason, we developed a multi-gene targeted capture approach to generate reference DNA sequences for plant taxa across 20 target chloroplast gene regions in a single assay. We successfully compiled a reference database for 93 temperate coastal plants including seagrasses, mangroves, and saltmarshes/samphire’s. We demonstrate the importance of a comprehensive reference database to prevent species going undetected in eDNA studies. We also investigate how using multiple chloroplast gene regions impacts the ability to discriminate between taxa.

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Database establishment for the secondary fungal DNA barcodetranslational elongation factor 1α(TEF1α)

Genome ◽

10.1139/gen-2018-0083 ◽

2019 ◽

Vol 62 (3) ◽

pp. 160-169 ◽

Cited By ~ 8

Author(s):

Wieland Meyer ◽

Laszlo Irinyi ◽

Minh Thuy Vi Hoang ◽

Vincent Robert ◽

Dea Garcia-Hermoso ◽

...

Keyword(s):

Dna Sequences ◽

Fungal Infections ◽

Dna Barcode ◽

Elongation Factor ◽

Its Region ◽

Reference Sequence ◽

Diagnostic Tools ◽

Reference Database ◽

Elongation Factor 1Α ◽

Fungal Dna

With new or emerging fungal infections, human and animal fungal pathogens are a growing threat worldwide. Current diagnostic tools are slow, non-specific at the species and subspecies levels, and require specific morphological expertise to accurately identify pathogens from pure cultures. DNA barcodes are easily amplified, universal, short species-specific DNA sequences, which enable rapid identification by comparison with a well-curated reference sequence collection. The primary fungal DNA barcode, ITS region, was introduced in 2012 and is now routinely used in diagnostic laboratories. However, the ITS region only accurately identifies around 75% of all medically relevant fungal species, which has prompted the development of a secondary barcode to increase the resolution power and suitability of DNA barcoding for fungal disease diagnostics. The translational elongation factor 1α (TEF1α) was selected in 2015 as a secondary fungal DNA barcode, but it has not been implemented into practice, due to the absence of a reference database. Here, we have established a quality-controlled reference database for the secondary barcode that together with the ISHAM-ITS database, forms the ISHAM barcode database, available online at http://its.mycologylab.org/ . We encourage the mycology community for active contributions.

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Molecular characterization of Urocleidoides cuiabai and U. malabaricusi (Monogenea: Dactylogyridae) from the trahira fish Hoplias aff. malabaricus in the Paraná River, Brazil

Journal of Helminthology ◽

10.1017/s0022149x15000966 ◽

2015 ◽

Vol 90 (6) ◽

pp. 693-697 ◽

Cited By ~ 3

Author(s):

L.S. Gasques ◽

R.J. Graça ◽

S.M.A.P. Prioli ◽

R.M. Takemoto ◽

A.J. Prioli

Keyword(s):

Cryptic Species ◽

Molecular Characterization ◽

Dna Sequences ◽

Mitochondrial Gene ◽

Genetic Data ◽

Flood Plain ◽

Fish Parasites ◽

Coi Gene ◽

Coi Sequences

AbstractUrocleidoides ectoparasites are mainly found on fish of the neotropical regions. Although molecular research on monogeneans is available, no genetic data exist characterizing species in the Urocleidoides genus. Some DNA sequences have been efficacious in systematic studies and in the reconstruction of phylogenies of fish parasites. Relevant roles have been given to the sequence of the mitochondrial gene of cytochrome c oxidase I (COI). This study characterized COI sequences of the parasites Urocleidoides malabaricusi and U. cuiabai in trahira fish Hoplias aff. malabaricus of the flood plain of the Upper River Paraná, Brazil. The two species under analysis were distinguished by sequencing and analysing a 420-bp fragment of the COI gene, which suggested the existence of the cryptic species U. malabaricusi.

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DNA barcode data accurately identify higher taxa

10.7287/peerj.preprints.1633v1 ◽

2016 ◽

Author(s):

Jonathan A Coddington ◽

Ingi Agnarsson ◽

Ren-Chung Cheng ◽

Klemen Čandek ◽

Amy Driskell ◽

...

Keyword(s):

Dna Sequences ◽

Dna Barcode ◽

Accurate Method ◽

Reference Database ◽

Accurate Identification ◽

Sequencing Errors ◽

Sequence Identity ◽

Percent Sequence Identity ◽

Near Term ◽

Accuracy Of Results

The use of unique DNA sequences as a method for taxonomic identification is no longer fundamentally controversial, even though debate continues on the best markers, methods, and technology to use. Although both existing databanks such as GenBank and BOLD, as well as reference taxonomies, are imperfect, in best case scenarios “barcodes” (whether single or multiple, organelle or nuclear, loci) clearly are an increasingly fast and inexpensive method of identification, especially as compared to manual identification of unknowns by increasingly rare expert taxonomists. Because most species on Earth are undescribed, a complete reference database at the species level is impractical in the near term. The question therefore arises whether unidentified species can, using DNA barcodes, be accurately assigned to more inclusive groups such as genera and families—taxonomic ranks of putatively monophyletic groups for which the global inventory is more complete and stable. We used a carefully chosen test library of CO1 sequences from 49 families, 313 genera, and 816 species of spiders to assess the accuracy of genus and family-level identifications. We used BLAST queries of each sequence against the entire library and got the top ten hits resulting in 8160 hits. The percent sequence identity was reported from these hits (PIdent, range 75-100%). Accurate identification (PIdent above which errors totaled less than 5%) occurred for genera at PIdent values > 95 and families at PIdent values ≥ 91, suggesting these as heuristic thresholds for generic and familial identifications in spiders. Accuracy of identification increases with numbers of species/genus and genera/family in the library; above five genera per family and fifteen species per genus all identifications were correct. We propose that using percent sequence identity between conventional barcode sequences may be a feasible and reasonably accurate method to identify animals to family/genus. However, the quality of the underlying database impacts accuracy of results; many outliers in our dataset could be attributed to taxonomic and/or sequencing errors in BOLD and GenBank. It seems that an accurate and complete reference library of families and genera of life could provide accurate higher level taxonomic identifications cheaply and accessibly, within years rather than decades.

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Integrative taxonomy ofMetrichiaRoss (Trichoptera: Hydroptilidae: Ochrotrichiinae) microcaddisflies from Brazil: descriptions of twenty new species

PeerJ ◽

10.7717/peerj.2009 ◽

2016 ◽

Vol 4 ◽

pp. e2009 ◽

Cited By ~ 6

Author(s):

Allan P.M. Santos ◽

Daniela M. Takiya ◽

Jorge L. Nessimian

Keyword(s):

New Species ◽

Dna Sequences ◽

Mitochondrial Gene ◽

Dna Barcode ◽

First Record ◽

Integrative Taxonomy ◽

Morphological Data ◽

Mato Grosso ◽

Intraspecific Divergence ◽

Coi Sequences

Metrichiais assigned to the Ochrotrichiinae, a group of almost exclusively Neotropical microcaddisflies.Metrichiacomprises over 100 described species and, despite its diversity, only one species has been described from Brazil so far. In this paper, we provide descriptions for 20 new species from 8 Brazilian states:M. acuminatasp. nov.,M. azulsp. nov.,M. bonitasp. nov.,M. bracuisp. nov.,M. caracasp. nov.,M. circuliformesp. nov.,M. curtasp. nov.,M. farofasp. nov.,M. forcepssp. nov.,M. formosinhasp. nov.,M. goianasp. nov.,M. itabaianasp. nov.,M. longissimasp. nov.,M. peludasp. nov.,M. rafaelisp. nov.,M. simplessp. nov.,M. talhadasp. nov.,M. teresp. nov.,M. ubajarasp. nov., andM. vulgarissp. nov.DNA barcode sequences (577 bp of the mitochondrial gene COI) were generated for 13 of the new species and two previously known species ofMetrichiaresulting in 64 sequences. In addition, COI sequences were obtained for other genera of Ochrotrichiinae (Angrisanoia,Nothotrichia,Ochrotrichia,Ragatrichia, andRhyacopsyche). DNA sequences and morphological data were integrated to evaluate species delimitations. K2P pairwise distances were calculated to generate a neighbor-joining tree. COI sequences also were submitted to ABGD and GMYC methods to assess ‘potential species’ delimitation. Analyses showed a conspicuous barcoding gap amongMetrichiasequences (highest intraspecific divergence: 4.8%; lowest interspecific divergence: 12.6%). Molecular analyses also allowed the association of larvae and adults ofMetrichia bonitasp. nov.from Mato Grosso do Sul, representing the first record of microcaddisfly larvae occurring in calcareous tufa (or travertine). ABGD results agreed with the morphological delimitation ofMetrichiaspecies, while GMYC estimated a slightly higher number of species, suggesting the division of two morphological species, each one into two potential species. Because this could be due to unbalanced sampling and the lack of morphological diagnostic characters, we have maintained these two species as undivided.

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Molecular evidence for sweeping discontinuity between peracarid (Crustacea) fauna of Macaronesian islands and nearby continental coasts: over fifty candidate endemic species

10.1101/2021.06.22.449383 ◽

2021 ◽

Author(s):

Pedro E Vieira ◽

Andrea Desiderato ◽

Carla L Azevedo ◽

Patricia Esquete ◽

Filipe O Costa ◽

...

Keyword(s):

Dna Sequences ◽

Marine Invertebrates ◽

Genetic Distances ◽

Oceanic Islands ◽

Marine Conservation ◽

Molecular Evidence ◽

Network Analyses ◽

Operational Taxonomic Units ◽

La Palma ◽

Macaronesian Islands

Oceanic islands are recognized evolutionary hotspots for terrestrial organisms, but little is known about their impact on marine organisms' evolution and biogeography. The volcanic archipelagos of Macaronesia occupy a vast and complex region which is particularly suitable to investigate marine island biogeography. In this study, we used mitochondrial DNA sequences to investigate the genetic diferentiation between the populations from Webbnesia (i.e. Madeira, Selvagens and Canaries) and adjacent coasts, of 23 intertidal peracarid species. All species had unexpectedly high intraspecific genetic distances, reaching more than 20% in some cases. Between 79 and 95 Molecular Operational Taxonomic Units (MOTUs) were found in these species. Webbnesia populations displayed an impressive genetic diversity and high endemicity, with 83% of the MOTUs being private to these islands, particularly La Palma and Madeira. Network analyses suggested higher similarity between Webbnesia and Azores than with adjacent continental coasts. These results reveal an unanticipated and sweeping biogeographic discontinuity of peracaridean fauna between Webbnesia and the Iberian Peninsula, raising suspicion about the possible occurrence of identical patterns in other groups of marine invertebrates in the region. We emphasize the unique genetic heritage hosted by these islands, underlining the need to consider the fine scale endemicity in marine conservation efforts.

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Alignment-free classification of COI DNA barcode data with the Python package Alfie

Metabarcoding and Metagenomics ◽

10.3897/mbmg.4.55815 ◽

2020 ◽

Vol 4 ◽

Cited By ~ 1

Author(s):

Cameron M. Nugent ◽

Sarah J. Adamowicz

Keyword(s):

Neural Network ◽

Dna Sequences ◽

Dna Barcode ◽

Neural Network Classifier ◽

Alignment Free ◽

Target Sequences ◽

Coi Sequences ◽

Free Classification ◽

Python Package

Characterization of biodiversity from environmental DNA samples and bulk metabarcoding data is hampered by off-target sequences that can confound conclusions about a taxonomic group of interest. Existing methods for isolation of target sequences rely on alignment to existing reference barcodes, but this can bias results against novel genetic variants. Effectively parsing targeted DNA barcode data from off-target noise improves the quality of biodiversity estimates and biological conclusions by limiting subsequent analyses to a relevant subset of available data. Here, we present Alfie, a Python package for the alignment-free classification of cytochrome c oxidase subunit I (COI) DNA barcode sequences to taxonomic kingdoms. The package determines k-mer frequencies of DNA sequences, and the frequencies serve as input for a neural network classifier that was trained and tested using ~58,000 publicly available COI sequences. The classifier was designed and optimized through a series of tests that allowed for the optimal set of DNA k-mer features and optimal machine learning algorithm to be selected. The neural network classifier rapidly assigns COI sequences of varying lengths to kingdoms with greater than 99% accuracy and is shown to generalize effectively and make accurate predictions about data from previously unseen taxonomic classes. The package contains an application programming interface that allows the Alfie package’s functionality to be extended to different DNA sequence classification tasks to suit a user’s need, including classification of different genes and barcodes, and classification to different taxonomic levels. Alfie is free and publicly available through GitHub (https://github.com/CNuge/alfie) and the Python package index (https://pypi.org/project/alfie/).

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DNA-barcoding of sympatric species of ectoparasitic gastropods of the genusCerithiopsis(Mollusca: Gastropoda: Cerithiopsidae) from Croatia

Journal of the Marine Biological Association of the United Kingdom ◽

10.1017/s0025315412000926 ◽

2012 ◽

Vol 93 (4) ◽

pp. 1059-1065 ◽

Cited By ~ 7

Author(s):

M.V. Modica ◽

P. Mariottini ◽

J. Prkić ◽

M. Oliverio

Keyword(s):

Cryptic Species ◽

Dna Barcoding ◽

Dna Barcode ◽

Genetic Data ◽

Distinct Species ◽

Sympatric Species ◽

Species Level ◽

Colour Pattern ◽

Coi Sequences

The ectoparasitic gastropod genusCerithiopsisForbes & Hanley, 1850 was nominally based onMurex tubercularisMontagu, 1803. We have used the DNA barcode COI sequences to assay sympatric samples of morphotypes recently described as distinct species of theCerithiopsis tubercularis-complex. Our results demonstrated that, in the Croatian waters, the gastropods usually calledC. tubercularisin fact comprise a complex of cryptic species, which can be reliably diagnosed only by examining the soft parts. In the present study we have demonstrated that the colour pattern of the head-foot is diagnostic at the species level in this complex and, coupled with genetic data, may provide a sounding base for a revision of the cerithiopsids of the European coasts.

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DNA Barcoding of Mullets (Family Mugilidae) from Pakistan Reveals Surprisingly High Number of Unknown Candidate Species

Diversity ◽

10.3390/d13060232 ◽

2021 ◽

Vol 13 (6) ◽

pp. 232

Author(s):

Ariba Hasan ◽

Pirzada Jamal Ahmed Siddiqui ◽

Shabir Ali Amir ◽

Jean-Dominique Durand

Keyword(s):

Species Diversity ◽

Cryptic Species ◽

Dna Barcoding ◽

Dna Barcode ◽

Morphological Examination ◽

Operational Taxonomic Units ◽

Phylogenetic Studies ◽

Management Measures ◽

Morphological Criteria ◽

Candidate Species

The mullets are a widespread group of ecologically and economically important fishes of disputed taxonomy due to their uniform external morphology. Barcoding and phylogenetic studies from various locations around the world largely highlighted the species diversity underestimation using morphological criteria used to establish the taxonomy of the family. Here, we investigated the mullet species diversity from Pakistan, a biogeographic area where nearly no mullet species were genetically characterized. Morphological examination of 40 mullets reveals 6 known species (Planiliza macrolepis, P. klunzingeri, P. subviridis, Crenimugil seheli, Ellochelon vaigiensis, and Mugil cephalus). Using a references DNA barcode library, the DNA barcode-based species identification flagged eight molecular operational taxonomic units (MOTUs) belonging to five genera (Crenimugil, Ellochelon, Mugil, Osteomugil, and Planiliza). Among these MOTUs, only one was already present in Barcode of Life Data system, all other representing new Barcode Index Numbers (BIN). These results emphasize the importance of the recognition of cryptic species and the necessity to re-evaluate the overall diversity by the genetic characterization of different species of this family. DNA barcoding is an effective tool to reveal cryptic species that need to be considered in conservation and management measures of fisheries in Pakistan.

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