DNA barcoding as a tool for coral reef conservation

DNA Barcoding (DBC) is a method for taxonomic identification of animals that is based entirely on the 5′ portion of the mitochondrial gene, cytochrome oxidase subunit I (COI-5). It can be especially useful for identification of larval forms or incomplete specimens lacking diagnostic morphological characters. DBC can also facilitate the discovery of species and in defining “molecular taxonomic units” in problematic groups. However, DBC is not a panacea for coral reef taxonomy. In two of the most ecologically important groups on coral reefs, the Anthozoa and Porifera, COI-5 sequences have diverged too little to be diagnostic for all species. Other problems for DBC include paraphyly in mitochondrial gene trees and lack of differentiation between hybrids and their maternal ancestors. DBC also depends on the availability of databases of COI-5 sequences, which are still in early stages of development. A global effort to barcode all fish species has demonstrated the importance of large-scale coordination and is yielding promising results. Whether or not COI-5 by itself is sufficient for species assignments has become a contentious question; it is generally advantageous to use sequences from multiple loci.     

昆虫DNA条形码分析中的距离方法

本文介绍了DNA条形码分析中常用的距离方法,包括简单的距离方法、基于阈值的距离方法和基于模糊成员关系的距离方法,并且以松毛虫属7个近缘种的COⅠ数据为例,运用基于阈值的距离方法进行演示分析。     

caos software for use in character-based DNA barcoding

The success of character-based DNA barcoding depends on the efficient identification of diagnostic character states from molecular sequences that have been organized hierarchically (e.g. according to phylogenetic methods). Similarly, the reliability of these identified diagnostic character states must be assessed according to their ability to diagnose new sequences. Here, a set of software tools is presented that implement the previously described Characteristic Attribute Organization System for both diagnostic identification and diagnostic-based classification. The software is publicly available from http://sarkarlab.mbl.edu/CAOS.     

DNA barcoding: a tool for improved taxon identification and detection of species diversity

Recently it was decided that portions of rbcL and matK gene regions are approved and required standard barcode regions for land plants. Ideally, DNA barcoding can provide a fast and reliable way to identify species. Compiling a library of barcodes can be enhanced by the numerous specimens available in botanic gardens, museums and herbaria and in other ex situ conservation collections. Barcoding can strengthen ongoing efforts of botanic gardens and ex situ conservation collections to preserve Earth’s biodiversity. Our study aimed to detect the usability of the universal primers of the standard DNA barcode, to produce standard barcodes for species identification and to detect the discriminatory power of the standard barcode in a set of different groups of plant and fungal taxa. We studied Betula species originating from different parts of the world, and Salix taxa, bryophytes and edible and poisonous fungal species originating from Finland. In Betula and Salix, the standard DNA barcode regions, portions of matK and rbcL, were able to identify species to genus level, but did not show adequate resolution for species discrimination. Thus, supplementary barcode regions are needed for species identification. In Salix, the trnH-psbA spacer was also used, and it proved to have more resolution but, yet, not adequate levels of interspecific divergence for all studied taxa. In a set of bryophyte species, the rbcL gene region was found to possess adequate resolution for species discrimination for most genera studied. In bryophytes, matK failed to amplify properly. In fungi, the combination of ITS1 and ITS2 proved to be effective for species discrimination, although alignment difficulties were encountered. In general, closely related or recently diverged species are the greatest challenge, and the problem is most difficult in plants, both in terms of a suitable combination of barcoding regions and the universality of used primers.     

DNA barcoding and minibarcoding as a powerful tool for feather mite studies

Feather mites (Astigmata: Analgoidea and Pterolichoidea) are among the most abundant and commonly occurring bird ectosymbionts. Basic questions on the ecology and evolution of feather mites remain unanswered because feather mite species identification is often only possible for adult males, and it is laborious even for specialized taxonomists, thus precluding large‐scale identifications. Here, we tested DNA barcoding as a useful molecular tool to identify feather mites from passerine birds. Three hundred and sixty‐one specimens of 72 species of feather mites from 68 species of European passerine birds from Russia and Spain were barcoded. The accuracy of barcoding and minibarcoding was tested. Moreover, threshold choice (a controversial issue in barcoding studies) was also explored in a new way, by calculating through simulations the effect of sampling effort (in species number and species composition) on threshold calculations. We found one 200‐bp minibarcode region that showed the same accuracy as the full‐length barcode (602 bp) and was surrounded by conserved regions potentially useful for group‐specific degenerate primers. Species identification accuracy was perfect (100%) but decreased when singletons or species of the Proctophyllodes pinnatus group were included. In fact, barcoding confirmed previous taxonomic issues within the P. pinnatus group. Following an integrative taxonomy approach, we compared our barcode study with previous taxonomic knowledge on feather mites, discovering three new putative cryptic species and validating three previous morphologically different (but still undescribed) new species.     

DNA barcoding as a complementary tool for conservation and valorisation of forest resources

Since the pre-historic era, humans have been using forests as a food, drugs and handcraft reservoir. Today, the use of botanical raw material to produce pharmaceuticals, herbal remedies, teas, spirits, cosmetics, sweets, dietary supplements, special industrial compounds and crude materials constitute an important global resource in terms of healthcare and economy. In recent years, DNA barcoding has been suggested as a useful molecular technique to complement traditional taxonomic expertise for fast species identification and biodiversity inventories. In this study, in situ application of DNA barcodes was tested on a selected group of forest tree species with the aim of contributing to the identification, conservation and trade control of these valuable plant resources.The “core barcode” for land plants (rbcL, matK, and trnH-psbA) was tested on 68 tree specimens (24 taxa). Universality of the method, ease of data retrieval and correct species assignment using sequence character states, presence of DNA barcoding gaps and GenBank discrimination assessment were evaluated. The markers showed different prospects of reliable applicability. RbcL and trnH-psbA displayed 100% amplification and sequencing success, while matK did not amplify in some plant groups. The majority of species had a single haplotype. The trnH-psbA region showed the highest genetic variability, but in most cases the high intraspecific sequence divergence revealed the absence of a clear DNA barcoding gap. We also faced an important limitation because the taxonomic coverage of the public reference database is incomplete. Overall, species identification success was 66.7%.This work illustrates current limitations in the applicability of DNA barcoding to taxonomic forest surveys. These difficulties urge for an improvement of technical protocols and an increase of the number of sequences and taxa in public databases.     

Comparing the usefulness of distance, monophyly and character-based DNA barcoding methods in species identification: a case study of neogastropoda

Background

DNA barcoding has recently been proposed as a promising tool for the rapid species identification in a wide range of animal taxa. Two broad methods (distance and monophyly-based methods) have been used. One method is based on degree of DNA sequence variation within and between species while another method requires the recovery of species as discrete clades (monophyly) on a phylogenetic tree. Nevertheless, some issues complicate the use of both methods. A recently applied new technique, the character-based DNA barcode method, however, characterizes species through a unique combination of diagnostic characters.

Methodology/Principal Findings

Here we analyzed 108 COI and 102 16S rDNA sequences of 40 species of Neogastropoda from a wide phylogenetic range to assess the performance of distance, monophyly and character-based methods of DNA barcoding. The distance-based method for both COI and 16S rDNA genes performed poorly in terms of species identification. Obvious overlap between intraspecific and interspecific divergences for both genes was found. The “10× rule” threshold resulted in lumping about half of distinct species for both genes. The neighbour-joining phylogenetic tree of COI could distinguish all species studied. However, the 16S rDNA tree could not distinguish some closely related species. In contrast, the character-based barcode method for both genes successfully identified 100% of the neogastropod species included, and performed well in discriminating neogastropod genera.

Conclusions/Significance

This present study demonstrates the effectiveness of the character-based barcoding method for species identification in different taxonomic levels, especially for discriminating the closely related species. While distance and monophyly-based methods commonly use COI as the ideal gene for barcoding, the character-based approach can perform well for species identification using relatively conserved gene markers (e.g., 16S rDNA in this study). Nevertheless, distance and monophyly-based methods, especially the monophyly-based method, can still be used to flag species.     

DNA barcoding is a useful tool for the identification of marine fishes from Japan

In this study, 229 DNA sequences of cytochrome oxidase subunit I gene (COI) from 158 marine fishes of Japan were employed to test the efficacy of species identification by DNA barcoding. The average genetic distance was 60-fold higher between species than within species, as Kimura two parameter (K2P) genetic distances averaged 17.6% among congeners and only 0.3% among conspecifics. There were no overlaps between intraspecific and interspecific K2P distances, and all sequences formed species units in the neighbor-joining dendrogram. Hybridization phenomena in two species (Kyphosus vaigiensis and Pterocaesio digramma) were also detected through searches in Barcode of Life Data Systems (BOLD). DNA barcoding provides a new way for fish identification.     

DNA barcoding of freshwater ichthyoplankton in the Neotropics as a tool for ecological monitoring

Quantifying and classifying ichthyoplankton is one of the most effective ways of monitoring the recruitment process in fishes. However, correctly identifying the fish based on morphological characters is extremely difficult, especially in the early stages of development. We examined ichthyoplankton from tributaries and reservoirs along the middle stretch of the Paranapanema River, one of the areas most impacted by hydroelectric projects in the Neotropics. Matching DNA sequences of the COI gene (628–648 bp) allowed us to identify 99.25% of 536 samples of eggs (293) and larvae (243) subjected to BOLD‐IDS similarity analysis with a species‐level threshold of 1.3%. The results revealed 37 species in 27 genera, 15 families and four orders, some 23.8% of documented fish species in the Paranapanema River. Molecular identification meant that we could include data from egg samples that accounted for about 30% of the species richness observed. The results in this study confirm the efficacy of DNA barcoding in identifying Neotropical ichthyoplankton and show how the data produced provide valuable information for preparing plans for conserving and managing inland waters.     

BCETM: a tool for software integration

The integration of software into special-purpose systems (e.g.for gene sequence analysis) can be a difficult task. We describea general-purpose software integration tool, the BCETM program,that facilitates assembly of VAX-based software into applicationsystems and provides an easy-to-use, intuitive user interface.We describe the use of BCE to integrate a heterogeneous collectionof sequence analysis tools. Many BCE design features are generallyapplicable and can be implemented in other language or hardwareenvironments. Received on May 13, 1987; accepted on October 2, 1987     

DNA barcoding as a heuristic tool for classifying undescribed Nearctic Myrmica ants (Hymenoptera: Formicidae)

The Palearctic species of the ant genus Myrmica are well studied. In contrast, the taxonomy of the Nearctic species is outdated, making identification impossible. We collected Myrmica samples in the Holarctic and investigated their diversity using mtDNA data. We analysed a barcode sequence of the Cytochrome Oxidase I gene for 57 Palearctic and 293 Nearctic Myrmica samples. We used sequences of known Palearctic species to search for Myrmica barcode patterns. All but one Palearctic species groups were recovered. The Nearctic diversity was much higher than known. We retrieved the punctiventris, crassirugis and incompleta groups, and established nine additional tentative species groups. Genetic distance analysis revealed a large overlap of intra- and inter-specific distances in Palearctic species and species groups. We could not find a variation gap to separate Nearctic sequences into species with COI data only. Variation in scape morphology divided two genetic groups further. Scape morphology correlated with most molecular groups, except three specimens. Our results illustrate that barcoding, using only a limited amount of genetic information, cannot serve as a universal proxy for taxonomy and species demarcation. It should be considered a first step in understanding the taxonomic diversity of an unknown group of organisms.     

DNA条形码分析方法研究进展

DNA条形码是一段短的、标准化的DNA序列,DNA条形码技术通过对DNA条形码序列分析实现物种的有效鉴定.随着生物DNA条形码序列的大量测定,DNA条形码分析方法得到迅速发展,推动了其在生物分子鉴定中的应用.2003年以来,DNA条形码技术已广泛应用于动物、植物和真菌等物种的鉴定,并有力地推动了生物分类学、生物多样性和生态学等学科的发展.本文在综述DNA条形码技术的基础上,总结了5类主要的DNA条形码分析方法,即基于遗传距离的分析、基于遗传相似度的分析、基于系统发育树的分析、基于序列特征的分析和基于统计分类法的分析,并进一步展望了DNA条形码技术的发展与应用.     

PROBEmer: A web-based software tool for selecting optimal DNA oligos

PROBEmer (http://probemer.cs.loyola.edu) is a web-based software tool that enables a researcher to select optimal oligos for PCR applications and multiplex detection platforms including oligonucleotide microarrays and bead-based arrays. Given two groups of nucleic-acid sequences, a target group and a non-target group, the software identifies oligo sequences that occur in members of the target group, but not in the non-target group. To help predict potential cross hybridization, PROBEmer computes all near neighbors in the non-target group and displays their alignments. The software has been used to obtain genus-specific prokaryotic probes based on the 16S rRNA gene, gene-specific probes for expression analyses and PCR primers. In this paper, we describe how to use PROBEmer, the computational methods it employs, and experimental results for oligos identified by this software tool.     

Loader Lite: a new software tool for the ABI PRISM 3700 DNA sequencer

Here we describe the development of a novel software tool entitled Loader Lite that generates plate records or sample sheetsfor the ABI PRISMs 3700 DNA sequencer. The major advantage of this program is that it enables the ongoing operation of sequencing instruments without reference to external network(s). The autonomous operation of sequencing instruments is critical if sample throughput is to be maintained during periods of network outage. Loader Lite employs a deliberate strategy of inputting anonymous tray barcodes at run time. After sequencing, the barcodes are reconciled with relevant project details by reference to a database. This software takes advantage of barcode scanning technology by creating plate records directly on the local computer, serving an individual sequencer, immediately before importing and linking. This real-time synthesis of the plate records at the point of loading all but eliminates loading errors. Loader Lite is user-friendly, fully configurable, and permits the running of partial or full 384-well sample trays, using any standard combinations of run modules, dye sets, mobility files, analysis modules, etc. The 96-well format is not supported; however, this capability will appear in subsequent versions that are currently under development. This application is designed as an added value, adjunct program to the regular ABI PRISM 3700 Data Collection software. We have successfully used Loader Lite over the past six months to load approximately 7 million sequencing reactions and believe its utility and functionality will prove to be attractive to the wider sequencing community.     

DNA-BAR: distinguisher selection for DNA barcoding

DNA-BAR is a software package for selecting DNA probes (henceforth referred to as distinguishers) that can be used in genomic-based identification of microorganisms. Given the genomic sequences of the microorganisms, DNA-BAR finds a near-minimum number of distinguishers yielding a distinct hybridization pattern for each microorganism. Selected distinguishers satisfy user specified bounds on length, melting temperature and GC content, as well as redundancy and cross-hybridization constraints.     

The utility of DNA barcoding as a tool to assess the success of ecological restoration using Hemiptera as a biological indicator

DNA barcoding uses a short, standardized DNA fragment to sort individuals into species. This molecular technique has applications in fields including ecology, evolution, conservation, and biogeography. In ecological applications such as species monitoring and habitat restoration, its potential has not been fully realized and implemented. Invertebrates are excellent biological indicators, as changes in species diversity or community assemblage provide important insights into the condition of, or changes in, the environment. This information is particularly useful within the context of restoration ecology. In this study, DNA barcoding is used to assess the potential of Hemiptera as a biological indicator of restoration success for the Buffelsdraai Landfill Site Community Reforestation Project (Durban, South Africa). A total of 393 Hemiptera specimens were collected from sites reforested at distinct phases (plots reforested in 2010, 2012, and 2015) and two reference sites (natural forest and grassland). The Hemiptera species composition and assemblage were assessed by analyzing diversity indices, ordination, unweighted pair‐group average cluster analysis, and phylogenetic analysis. Hemiptera species composition varied significantly across the chronologically different reforested sites, with a higher species richness observed in the older reforested plots. This suggests that Hemiptera diversity can be used to track restoration success, even over the small temporal scale used in this study. This study highlights the utility of DNA barcoding as a taxonomic sorting tool both to monitor ecological restoration and to discover specific taxa within Hemiptera that may be useful biological indicators.     

植物DNA条形码研究展望

<正>物种的鉴定是生物多样性研究的基石之一。在DNA双螺旋结构发现50年之际,加拿大学者提出了通过DNA条形码,即标准化的、较短的DNA序列对物种进行快速、准确鉴定的动议(Hebert et al.,2003)。经过12年的发展,DNA条形码已成为生物多样性研究领域发展最迅速的方向之一。一方面,DNA条形码和分子系统发育的研究使分类学、生态学、进化生物学等传统领域焕发出新的活力,极大