The seven deadly sins of DNA barcoding Collins, R. A.; Cruickshank, R. H.
Molecular ecology resources,
November 2013, Volume:
13, Issue:
6
Journal Article
Peer reviewed
Despite the broad benefits that DNA barcoding can bring to a diverse range of biological disciplines, a number of shortcomings still exist in terms of the experimental design of studies incorporating ...this approach. One underlying reason for this lies in the confusion that often exists between species discovery and specimen identification, and this is reflected in the way that hypotheses are generated and tested. Although these aims can be associated, they are quite distinct and require different methodological approaches, but their conflation has led to the frequently inappropriate use of commonly used analytical methods such as neighbour‐joining trees, bootstrap resampling and fixed distance thresholds. Furthermore, the misidentification of voucher specimens can also have serious implications for end users of reference libraries such as the Barcode of Life Data Systems, and in this regard we advocate increased diligence in the a priori identification of specimens to be used for this purpose. This commentary provides an assessment of seven deficiencies that we identify as common in the DNA barcoding literature, and outline some potential improvements for its adaptation and adoption towards more reliable and accurate outcomes.
Orthoptera, among the oldest and most numerous insect lineages, is an excellent model for evolutionary studies but has numerous taxonomic problems. To mitigate these issues, the cytochrome c oxidase ...subunit I (COI), standardized with the DNA barcode for Metazoa, is increasingly used for specimen identification and species delimitation. We tested the performance of COI as a DNA barcode in Orthoptera, using two analyses based on intra- and inter-specific distances, barcode gap, and Probability of Correct Identification (PCI); and estimated species richness through Automatic Barcode Gap Discovery (ABGD) and Assemble Species by Automatic Partitioning (ASAP). We filtered all sequences of Orthoptera available in Barcode of Life Data System (BOLD) and used 11605 COI sequences, covering 1132 species, 226 genera, and 18 families. The overall average PCI was 73.86%. For 82.2% of genera, the barcode gap boxplots were classified as "good" or "intermediate", indicating that COI can be effective as a DNA barcode in Orthoptera, although with varying efficiency depending on the need for more information. ABGD and ASAP inferred species richness similar to labels informed by BOLD for the suborders Caelifera and Ensifera. The representation of Orthoptera in the BOLD database and the results of these analyses are discussed.
Telling plant species apart with DNA: from barcodes to genomes Hollingsworth, Peter M.; Li, De-Zhu; van der Bank, Michelle ...
Philosophical transactions of the Royal Society of London. Series B. Biological sciences,
09/2016, Volume:
371, Issue:
1702
Journal Article
Peer reviewed
Open access
Land plants underpin a multitude of ecosystem functions, support human livelihoods and represent a critically important component of terrestrial biodiversity—yet many tens of thousands of species ...await discovery, and plant identification remains a substantial challenge, especially where material is juvenile, fragmented or processed. In this opinion article, we tackle two main topics. Firstly, we provide a short summary of the strengths and limitations of plant DNA barcoding for addressing these issues. Secondly, we discuss options for enhancing current plant barcodes, focusing on increasing discriminatory power via either gene capture of nuclear markers or genome skimming. The former has the advantage of establishing a defined set of target loci maximizing efficiency of sequencing effort, data storage and analysis. The challenge is developing a probe set for large numbers of nuclear markers that works over sufficient phylogenetic breadth. Genome skimming has the advantage of using existing protocols and being backward compatible with existing barcodes; and the depth of sequence coverage can be increased as sequencing costs fall. Its non-targeted nature does, however, present a major informatics challenge for upscaling to large sample sets.
This article is part of the themed issue ‘From DNA barcodes to biomes’.
Armigeres subalbatus, a mosquito species widely found in Thailand and other Asian countries, serves as a vector for filarial parasites, affecting both humans and animals. However, the surveillance of ...this vector is complicated because of its morphological similarity to two other species, Armigeres dohami and Armigeres kesseli. To differentiate these morphologically similar species, our study employed both wing geometric morphometrics (GM) and DNA barcoding, offering a comprehensive approach to accurately identify these closely related Armigeres species in Thailand. Our GM analyses based on shape demonstrated significant accuracy in differentiating Armigeres species. Specifically, the outline-based GM method focusing on the 3rd posterior cell exhibited an accuracy rate of 82.61%, closely followed by the landmark-based GM method with 81.54%. Both these GM techniques effectively distinguished Ar. subalbatus from Ar. dohami and Ar. kesseli. Regarding DNA barcoding, our investigation of pairwise intra- and interspecific divergences revealed a "barcoding gap". Furthermore, the results of species confirmation using both species delimitation methods including the automatic barcode gap discovery method (ABGD) and the Multi-rate Poisson tree process (mPTP) were consistent with those of morphological identification, sequence comparisons with the GenBank and Barcode of Life Data System (BOLD) databases, and the neighbor-joining tree construction. These consistent results emphasize the efficacy of DNA barcoding in the precise identification of Armigeres species.
Because many animal species are undescribed, and because the identification of known species is often difficult, interim taxonomic nomenclature has often been used in biodiversity analysis. By ...assigning individuals to presumptive species, called operational taxonomic units (OTUs), these systems speed investigations into the patterning of biodiversity and enable studies that would otherwise be impossible. Although OTUs have conventionally been separated through their morphological divergence, DNA-based delineations are not only feasible, but have important advantages. OTU designation can be automated, data can be readily archived, and results can be easily compared among investigations. This study exploits these attributes to develop a persistent, species-level taxonomic registry for the animal kingdom based on the analysis of patterns of nucleotide variation in the barcode region of the cytochrome c oxidase I (COI) gene. It begins by examining the correspondence between groups of specimens identified to a species through prior taxonomic work and those inferred from the analysis of COI sequence variation using one new (RESL) and four established (ABGD, CROP, GMYC, jMOTU) algorithms. It subsequently describes the implementation, and structural attributes of the Barcode Index Number (BIN) system. Aside from a pragmatic role in biodiversity assessments, BINs will aid revisionary taxonomy by flagging possible cases of synonymy, and by collating geographical information, descriptive metadata, and images for specimens that are likely to belong to the same species, even if it is undescribed. More than 274,000 BIN web pages are now available, creating a biodiversity resource that is positioned for rapid growth.
Environmental DNA (eDNA) metabarcoding is increasingly used to study the present and past biodiversity. eDNA analyses often rely on amplification of very small quantities or degraded DNA. To avoid ...missing detection of taxa that are actually present (false negatives), multiple extractions and amplifications of the same samples are often performed. However, the level of replication needed for reliable estimates of the presence/absence patterns remains an unaddressed topic. Furthermore, degraded DNA and PCR/sequencing errors might produce false positives. We used simulations and empirical data to evaluate the level of replication required for accurate detection of targeted taxa in different contexts and to assess the performance of methods used to reduce the risk of false detections. Furthermore, we evaluated whether statistical approaches developed to estimate occupancy in the presence of observational errors can successfully estimate true prevalence, detection probability and false‐positive rates. Replications reduced the rate of false negatives; the optimal level of replication was strongly dependent on the detection probability of taxa. Occupancy models successfully estimated true prevalence, detection probability and false‐positive rates, but their performance increased with the number of replicates. At least eight PCR replicates should be performed if detection probability is not high, such as in ancient DNA studies. Multiple DNA extractions from the same sample yielded consistent results; in some cases, collecting multiple samples from the same locality allowed detecting more species. The optimal level of replication for accurate species detection strongly varies among studies and could be explicitly estimated to improve the reliability of results.
Gut microbiota of patients with Parkinson’s disease and healthy volunteers was analyzed by the method of high throughput 16S rRNA sequencing of bacterial genomes. In patients with Parkinson’s ...diseases, changes in the content of 9 genera and 15 species of microorganisms were revealed: reduced content of
Dorea
,
Bacteroides
,
Prevotella
,
Faecalibacterium
,
Bacteroides massiliensis
,
Stoquefichus massiliensis
,
Bacteroides coprocola
,
Blautia glucerasea
,
Dorea longicatena
,
Bacteroides dorei
,
Bacteroides plebeus
,
Prevotella copri
,
Coprococcus eutactus
, and
Ruminococcus callidus
, and increased content of
Christensenella
,
Catabacter
,
Lactobacillus
,
Oscillospira
,
Bifidobacterium
,
Christensenella minuta
,
Catabacter hongkongensis
,
Lactobacillus mucosae
,
Ruminococcus bromii
, and
Papillibacter cinnamivorans
. This microbiological pattern of gut microflora can trigger local inflammation followed by aggregation of α-synuclein and generation of Lewy bodies.
The number of seafood species sold on Western markets is constantly growing and many unconventional species are sold in ethnic food outlets. In this work, 68 ethnic seafood products variously ...processed were collected from the Italian market and a molecular analysis was performed by sequencing a full cytochrome c oxidase (COI) DNA barcode (FDB, ∼655 bp) or a mini COI DNA barcode (MDB, ∼139 bp) using universal primers. Barcodes were then compared with sequences available in BOLD and GenBank. In addition, the label information was assessed according to the European legislation. By using the IDs analysis on BOLD a maximum species identity ≥98% was retrieved for 84% of the sequences. Of these, 67% were unambiguously identified at species level (51.3% of the FDB and 74% of the MDB). Using NCBI BLAST, 74% of the sequences scored a maximum species identity ≥98%, of which 73% were identified at species level (52% of the FDB and 61% of the MDB). Both databases performed better in mollusk identification. Overall, 45 products (66%) were not correctly labeled according to the European requirements. Finally, the comparison between the molecular and the label analysis highlighted that 48.5% of the products presented discrepancies between labeling and molecular identification. In particular, health implications were highlighted for 2 samples labeled as squid but identified as Lagocephalus spp., a poisonous puffer fish species banned from the EU market. The present results confirm DNA barcoding as a reliable tool for protecting consumers' health and economic interests.
•Unconventional seafood species are sold in ethnic food retailers in Western countries.•DNA barcoding is a useful tool for seafood species identification.•Full and mini-DNA barcodes have been used for ethnic seafood identification.•Full and mini-DNA barcodes show high discriminatory ability.•Molecular and labeling analysis highlighted widespread mislabeling.
The analysis of food webs and their dynamics facilitates understanding of the mechanistic processes behind community ecology and ecosystem functions. Having accurate techniques for determining ...dietary ranges and components is critical for this endeavour. While visual analyses and early molecular approaches are highly labour intensive and often lack resolution, recent DNA‐based approaches potentially provide more accurate methods for dietary studies. A suite of approaches have been used based on the identification of consumed species by characterization of DNA present in gut or faecal samples. In one approach, a standardized DNA region (DNA barcode) is PCR amplified, amplicons are sequenced and then compared to a reference database for identification. Initially, this involved sequencing clones from PCR products, and studies were limited in scale because of the costs and effort required. The recent development of next generation sequencing (NGS) has made this approach much more powerful, by allowing the direct characterization of dozens of samples with several thousand sequences per PCR product, and has the potential to reveal many consumed species simultaneously (DNA metabarcoding). Continual improvement of NGS technologies, on‐going decreases in costs and current massive expansion of reference databases make this approach promising. Here we review the power and pitfalls of NGS diet methods. We present the critical factors to take into account when choosing or designing a suitable barcode. Then, we consider both technical and analytical aspects of NGS diet studies. Finally, we discuss the validation of data accuracy including the viability of producing quantitative data.
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