A new method for species identification via protein-coding and non-coding DNA barcodes by combining machine learning with bioinformatic methods.

PubMed

Zhang, Ai-bing; Feng, Jie; Ward, Robert D; Wan, Ping; Gao, Qiang; Wu, Jun; Zhao, Wei-zhong

2012-01-01

Species identification via DNA barcodes is contributing greatly to current bioinventory efforts. The initial, and widely accepted, proposal was to use the protein-coding cytochrome c oxidase subunit I (COI) region as the standard barcode for animals, but recently non-coding internal transcribed spacer (ITS) genes have been proposed as candidate barcodes for both animals and plants. However, achieving a robust alignment for non-coding regions can be problematic. Here we propose two new methods (DV-RBF and FJ-RBF) to address this issue for species assignment by both coding and non-coding sequences that take advantage of the power of machine learning and bioinformatics. We demonstrate the value of the new methods with four empirical datasets, two representing typical protein-coding COI barcode datasets (neotropical bats and marine fish) and two representing non-coding ITS barcodes (rust fungi and brown algae). Using two random sub-sampling approaches, we demonstrate that the new methods significantly outperformed existing Neighbor-joining (NJ) and Maximum likelihood (ML) methods for both coding and non-coding barcodes when there was complete species coverage in the reference dataset. The new methods also out-performed NJ and ML methods for non-coding sequences in circumstances of potentially incomplete species coverage, although then the NJ and ML methods performed slightly better than the new methods for protein-coding barcodes. A 100% success rate of species identification was achieved with the two new methods for 4,122 bat queries and 5,134 fish queries using COI barcodes, with 95% confidence intervals (CI) of 99.75-100%. The new methods also obtained a 96.29% success rate (95%CI: 91.62-98.40%) for 484 rust fungi queries and a 98.50% success rate (95%CI: 96.60-99.37%) for 1094 brown algae queries, both using ITS barcodes.

DNA Barcoding Reveals Cryptic Diversity within Commercially Exploited Indo-Malay Carangidae (Teleosteii: Perciformes)

PubMed Central

Mat Jaafar, Tun Nurul Aimi; Taylor, Martin I.; Mohd Nor, Siti Azizah; de Bruyn, Mark; Carvalho, Gary R.

2012-01-01

Background DNA barcodes, typically focusing on the cytochrome oxidase I gene (COI) in many animals, have been used widely as a species-identification tool. The ability of DNA barcoding to distinguish species from a range of taxa and to reveal cryptic species has been well documented. Despite the wealth of DNA barcode data for fish from many temperate regions, there are relatively few available from the Southeast Asian region. Here, we target the marine fish Family Carangidae, one of the most commercially-important families from the Indo-Malay Archipelago (IMA), to produce an initial reference DNA barcode library. Methodology/Principal Findings Here, a 652 bp region of COI was sequenced for 723 individuals from 36 putative species of Family Carangidae distributed within IMA waters. Within the newly-generated dataset, three described species exhibited conspecific divergences up to ten times greater (4.32–4.82%) than mean estimates (0.24–0.39%), indicating a discrepancy with assigned morphological taxonomic identification, and the existence of cryptic species. Variability of the mitochondrial DNA COI region was compared within and among species to evaluate the COI region's suitability for species identification. The trend in range of mean K2P distances observed was generally in accordance with expectations based on taxonomic hierarchy: 0% to 4.82% between individuals within species, 0% to 16.4% between species within genera, and 8.64% to 25.39% between genera within families. The average Kimura 2-parameter (K2P) distance between individuals, between species within genera, and between genera within family were 0.37%, 10.53% and 16.56%, respectively. All described species formed monophyletic clusters in the Neighbour-joining phylogenetic tree, although three species representing complexes of six potential cryptic species were detected in Indo-Malay Carangidae; Atule mate, Selar crumenophthalmus and Seriolina nigrofasciata. Conclusion/Significance This study confirms that COI is an effective tool for species identification of Carangidae from the IMA. There were moderate levels of cryptic diversity among putative species within the central IMA. However, to explain the hypothesis of species richness in the IMA, it is necessary to sample the whole family across their broad geographic range. Such insights are helpful not only to document mechanisms driving diversification and recruitment in Carangidae, but also to provide a scientific framework for management strategies and conservation of commercially-important fisheries resources. PMID:23209586

[Identification of antler powder components based on DNA barcoding technology].

PubMed

Jia, Jing; Shi, Lin-chun; Xu, Zhi-chao; Xin, Tian-yi; Song, Jing-yuan; Chen Shi, Lin

2015-10-01

In order to authenticate the components of antler powder in the market, DNA barcoding technology coupled with cloning method were used. Cytochrome c oxidase subunit I (COI) sequences were obtained according to the DNA barcoding standard operation procedure (SOP). For antler powder with possible mixed components, the cloning method was used to get each COI sequence. 65 COI sequences were successfully obtained from commercial antler powders via sequencing PCR products. The results indicates that only 38% of these samples were derived from Cervus nippon Temminck or Cervus elaphus Linnaeus which is recorded in the 2010 edition of "Chinese Pharmacopoeia", while 62% of them were derived from other species. Rangifer tarandus Linnaeus was the most frequent species among the adulterants. Further analysis showed that some samples collected from different regions, companies and prices, contained adulterants. Analysis of 36 COI sequences obtained by the cloning method showed that C. elaphus and C. nippon were main components. In addition, some samples were marked clearly as antler powder on the label, however, C. elaphus or R. tarandus were their main components. In summary, DNA barcoding can accurately and efficiently distinguish the exact content in the commercial antler powder, which provides a new technique to ensure clinical safety and improve quality control of Chinese traditional medicine

DNA barcoding Indian freshwater fishes.

PubMed

Lakra, Wazir Singh; Singh, M; Goswami, Mukunda; Gopalakrishnan, A; Lal, K K; Mohindra, V; Sarkar, U K; Punia, P P; Singh, K V; Bhatt, J P; Ayyappan, S

2016-11-01

DNA barcoding is a promising technique for species identification using a short mitochondrial DNA sequence of cytochrome c oxidase I (COI) gene. In the present study, DNA barcodes were generated from 72 species of freshwater fish covering the Orders Cypriniformes, Siluriformes, Perciformes, Synbranchiformes, and Osteoglossiformes representing 50 genera and 19 families. All the samples were collected from diverse sites except the species endemic to a particular location. Species were represented by multiple specimens in the great majority of the barcoded species. A total of 284 COI sequences were generated. After amplification and sequencing of 700 base pair fragment of COI, primers were trimmed which invariably generated a 655 base pair barcode sequence. The average Kimura two-parameter (K2P) distances within-species, genera, families, and orders were 0.40%, 9.60%, 13.10%, and 17.16%, respectively. DNA barcode discriminated congeneric species without any confusion. The study strongly validated the efficiency of COI as an ideal marker for DNA barcoding of Indian freshwater fishes.

FBIS: A regional DNA barcode archival & analysis system for Indian fishes.

PubMed

Nagpure, Naresh Sahebrao; Rashid, Iliyas; Pathak, Ajey Kumar; Singh, Mahender; Singh, Shri Prakash; Sarkar, Uttam Kumar

2012-01-01

DNA barcode is a new tool for taxon recognition and classification of biological organisms based on sequence of a fragment of mitochondrial gene, cytochrome c oxidase I (COI). In view of the growing importance of the fish DNA barcoding for species identification, molecular taxonomy and fish diversity conservation, we developed a Fish Barcode Information System (FBIS) for Indian fishes, which will serve as a regional DNA barcode archival and analysis system. The database presently contains 2334 sequence records of COI gene for 472 aquatic species belonging to 39 orders and 136 families, collected from available published data sources. Additionally, it contains information on phenotype, distribution and IUCN Red List status of fishes. The web version of FBIS was designed using MySQL, Perl and PHP under Linux operating platform to (a) store and manage the acquisition (b) analyze and explore DNA barcode records (c) identify species and estimate genetic divergence. FBIS has also been integrated with appropriate tools for retrieving and viewing information about the database statistics and taxonomy. It is expected that FBIS would be useful as a potent information system in fish molecular taxonomy, phylogeny and genomics. The database is available for free at http://mail.nbfgr.res.in/fbis/

Classification of Sharks in the Egyptian Mediterranean Waters Using Morphological and DNA Barcoding Approaches

PubMed Central

Moftah, Marie; Abdel Aziz, Sayeda H.; Elramah, Sara; Favereaux, Alexandre

2011-01-01

The identification of species constitutes the first basic step in phylogenetic studies, biodiversity monitoring and conservation. DNA barcoding, i.e. the sequencing of a short standardized region of DNA, has been proposed as a new tool for animal species identification. The present study provides an update on the composition of shark in the Egyptian Mediterranean waters off Alexandria, since the latest study to date was performed 30 years ago, DNA barcoding was used in addition to classical taxonomical methodologies. Thus, 51 specimen were DNA barcoded for a 667 bp region of the mitochondrial COI gene. Although DNA barcoding aims at developing species identification systems, some phylogenetic signals were apparent in the data. In the neighbor-joining tree, 8 major clusters were apparent, each of them containing individuals belonging to the same species, and most with 100% bootstrap value. This study is the first to our knowledge to use DNA barcoding of the mitochondrial COI gene in order to confirm the presence of species Squalus acanthias, Oxynotus centrina, Squatina squatina, Scyliorhinus canicula, Scyliorhinus stellaris, Mustelus mustelus, Mustelus punctulatus and Carcharhinus altimus in the Egyptian Mediterranean waters. Finally, our study is the starting point of a new barcoding database concerning shark composition in the Egyptian Mediterranean waters (Barcoding of Egyptian Mediterranean Sharks [BEMS], http://www.boldsystems.org/views/projectlist.php?&#Barcoding%20Fish%20%28FishBOL%29). PMID:22087242

Assessment of snake DNA barcodes based on mitochondrial COI and Cytb genes revealed multiple putative cryptic species in Thailand.

PubMed

Laopichienpong, Nararat; Muangmai, Narongrit; Supikamolseni, Arrjaree; Twilprawat, Panupon; Chanhome, Lawan; Suntrarachun, Sunutcha; Peyachoknagul, Surin; Srikulnath, Kornsorn

2016-12-15

DNA barcodes of mitochondrial cytochrome c oxidase I (COI), cytochrome b (Cytb) genes, and their combined data sets were constructed from 35 snake species in Thailand. No barcoding gap was detected in either of the two genes from the observed intra- and interspecific sequence divergences. Intra- and interspecific sequence divergences of the COI gene differed 14 times, with barcode cut-off scores ranging over 2%-4% for threshold values differentiated among most of the different species; the Cytb gene differed 6 times with cut-off scores ranging over 2%-6%. Thirty-five specific nucleotide mutations were also found at interspecific level in the COI gene, identifying 18 snake species, but no specific nucleotide mutation was observed for Cytb in any single species. This suggests that COI barcoding was a better marker than Cytb. Phylogenetic clustering analysis indicated that most species were represented by monophyletic clusters, suggesting that these snake species could be clearly differentiated using COI barcodes. However, the two-marker combination of both COI and Cytb was more effective, differentiating snake species by over 2%-4%, and reducing species numbers in the overlap value between intra- and interspecific divergences. Three species delimitation algorithms (general mixed Yule-coalescent, automatic barcoding gap detection, and statistical parsimony network analysis) were extensively applied to a wide range of snakes based on both barcodes. This revealed cryptic diversity for eleven snake species in Thailand. In addition, eleven accessions from the database previously grouped under the same species were represented at different species level, suggesting either high genetic diversity, or the misidentification of these sequences in the database as a consequence of cryptic species. Copyright © 2016 Elsevier B.V. All rights reserved.

Spiders (Araneae) of Churchill, Manitoba: DNA barcodes and morphology reveal high species diversity and new Canadian records.

PubMed

Blagoev, Gergin A; Nikolova, Nadya I; Sobel, Crystal N; Hebert, Paul D N; Adamowicz, Sarah J

2013-11-26

Arctic ecosystems, especially those near transition zones, are expected to be strongly impacted by climate change. Because it is positioned on the ecotone between tundra and boreal forest, the Churchill area is a strategic locality for the analysis of shifts in faunal composition. This fact has motivated the effort to develop a comprehensive biodiversity inventory for the Churchill region by coupling DNA barcoding with morphological studies. The present study represents one element of this effort; it focuses on analysis of the spider fauna at Churchill. 198 species were detected among 2704 spiders analyzed, tripling the count for the Churchill region. Estimates of overall diversity suggest that another 10-20 species await detection. Most species displayed little intraspecific sequence variation (maximum <1%) in the barcode region of the cytochrome c oxidase subunit I (COI) gene, but four species showed considerably higher values (maximum = 4.1-6.2%), suggesting cryptic species. All recognized species possessed a distinct haplotype array at COI with nearest-neighbour interspecific distances averaging 8.57%. Three species new to Canada were detected: Robertus lyrifer (Theridiidae), Baryphyma trifrons (Linyphiidae), and Satilatlas monticola (Linyphiidae). The first two species may represent human-mediated introductions linked to the port in Churchill, but the other species represents a range extension from the USA. The first description of the female of S. monticola was also presented. As well, one probable new species of Alopecosa (Lycosidae) was recognized. This study provides the first comprehensive DNA barcode reference library for the spider fauna of any region. Few cryptic species of spiders were detected, a result contrasting with the prevalence of undescribed species in several other terrestrial arthropod groups at Churchill. Because most (97.5%) sequence clusters at COI corresponded with a named taxon, DNA barcoding reliably identifies spiders in the Churchill fauna. The capacity of DNA barcoding to enable the identification of otherwise taxonomically ambiguous specimens (juveniles, females) also represents a major advance for future monitoring efforts on this group.

A DNA Barcode Library for Korean Chironomidae (Insecta: Diptera) and Indexes for Defining Barcode Gap

PubMed Central

Kim, Sungmin; Song, Kyo-Hong; Ree, Han-Il; Kim, Won

2012-01-01

Non-biting midges (Diptera: Chironomidae) are a diverse population that commonly causes respiratory allergies in humans. Chironomid larvae can be used to indicate freshwater pollution, but accurate identification on the basis of morphological characteristics is difficult. In this study, we constructed a mitochondrial cytochrome c oxidase subunit I (COI)-based DNA barcode library for Korean chironomids. This library consists of 211 specimens from 49 species, including adults and unidentified larvae. The interspecies and intraspecies COI sequence variations were analyzed. Sophisticated indexes were developed in order to properly evaluate indistinct barcode gaps that are created by insufficient sampling on both the interspecies and intraspecies levels and by variable mutation rates across taxa. In a variety of insect datasets, these indexes were useful for re-evaluating large barcode datasets and for defining COI barcode gaps. The COI-based DNA barcode library will provide a rapid and reliable tool for the molecular identification of Korean chironomid species. Furthermore, this reverse-taxonomic approach will be improved by the continuous addition of other speceis’ sequences to the library. PMID:22138764

DNA barcoding of Clarias gariepinus, Coptodon zillii and Sarotherodon melanotheron from Southwestern Nigeria

PubMed Central

Falade, Mofolusho O.; Opene, Anthony J.; Benson, Otarigho

2016-01-01

DNA barcoding has been adopted as a gold standard rapid, precise and unifying identification system for animal species and provides a database of genetic sequences that can be used as a tool for universal species identification. In this study, we employed mitochondrial genes 16S rRNA (16S) and cytochrome oxidase subunit I (COI) for the identification of some Nigerian freshwater catfish and Tilapia species. Approximately 655 bp were amplified from the 5′ region of the mitochondrial cytochrome C oxidase subunit I (COI) gene whereas 570 bp were amplified for the 16S rRNA gene. Nucleotide divergences among sequences were estimated based on Kimura 2-parameter distances and the genetic relationships were assessed by constructing phylogenetic trees using the neighbour-joining (NJ) and maximum likelihood (ML) methods. Analyses of consensus barcode sequences for each species, and alignment of individual sequences from within a given species revealed highly consistent barcodes (99% similarity on average), which could be compared with deposited sequences in public databases. The nucleotide distance between species belonging to different genera based on COI ranged from 0.17% between Sarotherodon melanotheron and Coptodon zillii to 0.49% between Clarias gariepinus and C. zillii, indicating that S. melanotheron and C. zillii are closely related. Based on the data obtained, the utility of COI gene was confirmed in accurate identification of three fish species from Southwest Nigeria. PMID:27990256

FBIS: A regional DNA barcode archival & analysis system for Indian fishes

PubMed Central

Nagpure, Naresh Sahebrao; Rashid, Iliyas; Pathak, Ajey Kumar; Singh, Mahender; Singh, Shri Prakash; Sarkar, Uttam Kumar

2012-01-01

DNA barcode is a new tool for taxon recognition and classification of biological organisms based on sequence of a fragment of mitochondrial gene, cytochrome c oxidase I (COI). In view of the growing importance of the fish DNA barcoding for species identification, molecular taxonomy and fish diversity conservation, we developed a Fish Barcode Information System (FBIS) for Indian fishes, which will serve as a regional DNA barcode archival and analysis system. The database presently contains 2334 sequence records of COI gene for 472 aquatic species belonging to 39 orders and 136 families, collected from available published data sources. Additionally, it contains information on phenotype, distribution and IUCN Red List status of fishes. The web version of FBIS was designed using MySQL, Perl and PHP under Linux operating platform to (a) store and manage the acquisition (b) analyze and explore DNA barcode records (c) identify species and estimate genetic divergence. FBIS has also been integrated with appropriate tools for retrieving and viewing information about the database statistics and taxonomy. It is expected that FBIS would be useful as a potent information system in fish molecular taxonomy, phylogeny and genomics. Availability The database is available for free at http://mail.nbfgr.res.in/fbis/ PMID:22715304

Potential of DNA sequences to identify zoanthids (Cnidaria: Zoantharia).

PubMed

Sinniger, Frederic; Reimer, James D; Pawlowski, Jan

2008-12-01

The order Zoantharia is known for its chaotic taxonomy and difficult morphological identification. One method that potentially could help for examining such troublesome taxa is DNA barcoding, which identifies species using standard molecular markers. The mitochondrial cytochrome oxidase subunit I (COI) has been utilized to great success in groups such as birds and insects; however, its applicability in many other groups is controversial. Recently, some studies have suggested that barcoding is not applicable to anthozoans. Here, we examine the use of COI and mitochondrial 16S ribosomal DNA for zoanthid identification. Despite the absence of a clear barcoding gap, our results show that for most of 54 zoanthid samples, both markers could separate samples to the species, or species group, level, particularly when easily accessible ecological or distributional data were included. Additionally, we have used the short V5 region of mt 16S rDNA to identify eight old (13 to 50 years old) museum samples. We discuss advantages and disadvantages of COI and mt 16S rDNA as barcodes for Zoantharia, and recommend that either one or both of these markers be considered for zoanthid identification in the future.

DNA barcoding commercially important aquatic invertebrates of Turkey.

PubMed

Keskin, Emre; Atar, Hasan Hüseyin

2013-08-01

DNA barcoding was used in order to identify aquatic invertebrates sampled from fisheries bycatch and discards. A total of 440 unique cytochrome c oxidase sub unit I (COI) barcodes were generated for 22 species from three important phyla (Arthropoda, Cnidaria, and Mollusca). All the species were sequenced and submitted to GenBank and Barcode of Life Database (BOLD) databases using 654 bp-long fragment of mitochondrial COI gene. Two of them (Pontastacus leptodactylus and Rapana bezoar) were first records of the species for the BOLD database and six of them (Carcinus aestuarii, Loligo vulgaris, Melicertus kerathurus, Nephrops norvegicus, Scyllarides latus, and Scyllarus arctus) were first standard (>648 bp) COI barcode records for the GenBank database. COI barcodes were analyzed for nucleotide composition, nucleotide pair frequencies, and Kimura's two-parameter genetic distance. Mean genetic distance among species was found increasing at higher taxonomic levels. Neighbor-joining trees generated were congruent with morphometric-based taxonomic classification. Findings of this study clearly demonstrate that DNA barcodes could be used as an efficient molecular tool in identification of not only target species from fisheries but also bycatch and discard species, and so it could provide us leverage for a better understanding in monitoring and management of fisheries and biodiversity.

Evaluation of DNA barcoding and identification of new haplomorphs in Canadian deerflies and horseflies.

PubMed

Cywinska, A; Hannan, M A; Kevan, P G; Roughley, R E; Iranpour, M; Hunter, F F

2010-12-01

This paper reports the first tests of the suitability of the standardized mitochondrial cytochrome c oxidase subunit I (COI) barcoding system for the identification of Canadian deerflies and horseflies. Two additional mitochondrial molecular markers were used to determine whether unambiguous species recognition in tabanids can be achieved. Our 332 Canadian tabanid samples yielded 650 sequences from five genera and 42 species. Standard COI barcodes demonstrated a strong A + T bias (mean 68.1%), especially at third codon positions (mean 93.0%). Our preliminary test of this system showed that the standard COI barcode worked well for Canadian Tabanidae: the target DNA can be easily recovered from small amounts of insect tissue and aligned for all tabanid taxa. Each tabanid species possessed distinctive sets of COI haplotypes which discriminated well among species. Average conspecific Kimura two-parameter (K2P) divergence (0.49%) was 12 times lower than the average divergence within species. Both the neighbour-joining and the Bayesian methods produced trees with identical monophyletic species groups. Two species, Chrysops dawsoni Philip and Chrysops montanus Osten Sacken (Diptera: Tabanidae), showed relatively deep intraspecific sequence divergences (∼ 10 times the average) for all three mitochondrial gene regions analysed. We suggest provisional differentiation of Ch. montanus into two haplotypes, namely, Ch. montanus haplomorph 1 and Ch. montanus haplomorph 2, both defined by their molecular sequences and by newly discovered differences in structural features near their ocelli. © 2010 Brock University. Medical and Veterinary Entomology © 2010 The Royal Entomological Society.

DNA Barcode Identification of Freshwater Snails in the Family Bithyniidae from Thailand

PubMed Central

Kulsantiwong, Jutharat; Prasopdee, Sattrachai; Ruangsittichai, Jiraporn; Ruangjirachuporn, Wipaporn; Boonmars, Thidarut; Viyanant, Vithoon; Pierossi, Paola; Hebert, Paul D. N.; Tesana, Smarn

2013-01-01

Freshwater snails in the family Bithyniidae are the first intermediate host for Southeast Asian liver fluke (Opisthorchis viverrini), the causative agent of opisthorchiasis. Unfortunately, the subtle morphological characters that differentiate species in this group are not easily discerned by non-specialists. This is a serious matter because the identification of bithyniid species is a fundamental prerequisite for better understanding of the epidemiology of this disease. Because DNA barcoding, the analysis of sequence diversity in the 5’ region of the mitochondrial COI gene, has shown strong performance in other taxonomic groups, we decided to test its capacity to resolve 10 species/ subspecies of bithyniids from Thailand. Our analysis of 217 specimens indicated that COI sequences delivered species-level identification for 9 of 10 currently recognized species. The mean intraspecific divergence of COI was 2.3% (range 0-9.2 %), whereas sequence divergences between congeneric species averaged 8.7% (range 0-22.2 %). Although our results indicate that DNA barcoding can differentiate species of these medically-important snails, we also detected evidence for the presence of one overlooked species and one possible case of synonymy. PMID:24223896

[Identification of common medicinal snakes in medicated liquor of Guangdong by COI barcode sequence].

PubMed

Liao, Jing; Chao, Zhi; Zhang, Liang

2013-11-01

To identify the common snakes in medicated liquor of Guangdong using COI barcode sequence,and to test the feasibility. The COI barcode sequences of collected medicinal snakes were amplified and sequenced. The sequences combined with the data from GenBank were analyzed for divergence and building a neighbor-joining(NJ) tree with MEGA 5.0. The genetic distance and NJ tree demonstrated that there were 241 variable sites in these species, and the average (A + T) content of 56.2% was higher than the average (G + C) content of 43.7%. The maximum interspecific genetic distance was 0.2568, and the minimum was 0. 1519. In the NJ tree,each species formed a monophyletic clade with bootstrap supports of 100%. DNA barcoding identification method based on the COI sequence is accurate and can be applied to identify the common medicinal snakes.

Spiders (Araneae) of Churchill, Manitoba: DNA barcodes and morphology reveal high species diversity and new Canadian records

PubMed Central

2013-01-01

Background Arctic ecosystems, especially those near transition zones, are expected to be strongly impacted by climate change. Because it is positioned on the ecotone between tundra and boreal forest, the Churchill area is a strategic locality for the analysis of shifts in faunal composition. This fact has motivated the effort to develop a comprehensive biodiversity inventory for the Churchill region by coupling DNA barcoding with morphological studies. The present study represents one element of this effort; it focuses on analysis of the spider fauna at Churchill. Results 198 species were detected among 2704 spiders analyzed, tripling the count for the Churchill region. Estimates of overall diversity suggest that another 10–20 species await detection. Most species displayed little intraspecific sequence variation (maximum <1%) in the barcode region of the cytochrome c oxidase subunit I (COI) gene, but four species showed considerably higher values (maximum = 4.1-6.2%), suggesting cryptic species. All recognized species possessed a distinct haplotype array at COI with nearest-neighbour interspecific distances averaging 8.57%. Three species new to Canada were detected: Robertus lyrifer (Theridiidae), Baryphyma trifrons (Linyphiidae), and Satilatlas monticola (Linyphiidae). The first two species may represent human-mediated introductions linked to the port in Churchill, but the other species represents a range extension from the USA. The first description of the female of S. monticola was also presented. As well, one probable new species of Alopecosa (Lycosidae) was recognized. Conclusions This study provides the first comprehensive DNA barcode reference library for the spider fauna of any region. Few cryptic species of spiders were detected, a result contrasting with the prevalence of undescribed species in several other terrestrial arthropod groups at Churchill. Because most (97.5%) sequence clusters at COI corresponded with a named taxon, DNA barcoding reliably identifies spiders in the Churchill fauna. The capacity of DNA barcoding to enable the identification of otherwise taxonomically ambiguous specimens (juveniles, females) also represents a major advance for future monitoring efforts on this group. PMID:24279427

DNA Barcode Analysis of Thrips (Thysanoptera) Diversity in Pakistan Reveals Cryptic Species Complexes.

PubMed

Iftikhar, Romana; Ashfaq, Muhammad; Rasool, Akhtar; Hebert, Paul D N

2016-01-01

Although thrips are globally important crop pests and vectors of viral disease, species identifications are difficult because of their small size and inconspicuous morphological differences. Sequence variation in the mitochondrial COI-5' (DNA barcode) region has proven effective for the identification of species in many groups of insect pests. We analyzed barcode sequence variation among 471 thrips from various plant hosts in north-central Pakistan. The Barcode Index Number (BIN) system assigned these sequences to 55 BINs, while the Automatic Barcode Gap Discovery detected 56 partitions, a count that coincided with the number of monophyletic lineages recognized by Neighbor-Joining analysis and Bayesian inference. Congeneric species showed an average of 19% sequence divergence (range = 5.6% - 27%) at COI, while intraspecific distances averaged 0.6% (range = 0.0% - 7.6%). BIN analysis suggested that all intraspecific divergence >3.0% actually involved a species complex. In fact, sequences for three major pest species (Haplothrips reuteri, Thrips palmi, Thrips tabaci), and one predatory thrips (Aeolothrips intermedius) showed deep intraspecific divergences, providing evidence that each is a cryptic species complex. The study compiles the first barcode reference library for the thrips of Pakistan, and examines global haplotype diversity in four important pest thrips.

When COI barcodes deceive: complete genomes reveal introgression in hairstreaks

PubMed Central

Shen, Jinhui; Borek, Dominika; Robbins, Robert K.; Opler, Paul A.; Otwinowski, Zbyszek; Grishin, Nick V.

2017-01-01

Two species of hairstreak butterflies from the genus Calycopis are known in the United States: C. cecrops and C. isobeon. Analysis of mitochondrial COI barcodes of Calycopis revealed cecrops-like specimens from the eastern US with atypical barcodes that were 2.6% different from either USA species, but similar to Central American Calycopis species. To address the possibility that the specimens with atypical barcodes represent an undescribed cryptic species, we sequenced complete genomes of 27 Calycopis specimens of four species: C. cecrops, C. isobeon, C. quintana and C. bactra. Some of these specimens were collected up to 60 years ago and preserved dry in museum collections, but nonetheless produced genomes as complete as fresh samples. Phylogenetic trees reconstructed using the whole mitochondrial and nuclear genomes were incongruent. While USA Calycopis with atypical barcodes grouped with Central American species C. quintana by mitochondria, nuclear genome trees placed them within typical USA C. cecrops in agreement with morphology, suggesting mitochondrial introgression. Nuclear genomes also show introgression, especially between C. cecrops and C. isobeon. About 2.3% of each C. cecrops genome has probably (p-value < 0.01, FDR < 0.1) introgressed from C. isobeon and about 3.4% of each C. isobeon genome may have come from C. cecrops. The introgressed regions are enriched in genes encoding transmembrane proteins, mitochondria-targeting proteins and components of the larval cuticle. This study provides the first example of mitochondrial introgression in Lepidoptera supported by complete genome sequencing. Our results caution about relying solely on COI barcodes and mitochondrial DNA for species identification or discovery. PMID:28179510

DNA barcodes for Nearctic Auchenorrhyncha (Insecta: Hemiptera).

PubMed

Foottit, Robert G; Maw, Eric; Hebert, P D N

2014-01-01

Many studies have shown the suitability of sequence variation in the 5' region of the mitochondrial cytochrome c oxidase I (COI) gene as a DNA barcode for the identification of species in a wide range of animal groups. We examined 471 species in 147 genera of Hemiptera: Auchenorrhyncha drawn from specimens in the Canadian National Collection of Insects to assess the effectiveness of DNA barcoding in this group. Analysis of the COI gene revealed less than 2% intra-specific divergence in 93% of the taxa examined, while minimum interspecific distances exceeded 2% in 70% of congeneric species pairs. Although most species are characterized by a distinct sequence cluster, sequences for members of many groups of closely related species either shared sequences or showed close similarity, with 25% of species separated from their nearest neighbor by less than 1%. This study, although preliminary, provides DNA barcodes for about 8% of the species of this hemipteran suborder found in North America north of Mexico. Barcodes can enable the identification of many species of Auchenorrhyncha, but members of some species groups cannot be discriminated. Future use of DNA barcodes in regulatory, pest management, and environmental applications will be possible as the barcode library for Auchenorrhyncha expands to include more species and broader geographic coverage.

DNA Barcodes for Nearctic Auchenorrhyncha (Insecta: Hemiptera)

PubMed Central

Foottit, Robert G.; Maw, Eric; Hebert, P. D. N.

2014-01-01

Background Many studies have shown the suitability of sequence variation in the 5′ region of the mitochondrial cytochrome c oxidase I (COI) gene as a DNA barcode for the identification of species in a wide range of animal groups. We examined 471 species in 147 genera of Hemiptera: Auchenorrhyncha drawn from specimens in the Canadian National Collection of Insects to assess the effectiveness of DNA barcoding in this group. Methodology/Principal Findings Analysis of the COI gene revealed less than 2% intra-specific divergence in 93% of the taxa examined, while minimum interspecific distances exceeded 2% in 70% of congeneric species pairs. Although most species are characterized by a distinct sequence cluster, sequences for members of many groups of closely related species either shared sequences or showed close similarity, with 25% of species separated from their nearest neighbor by less than 1%. Conclusions/Significance This study, although preliminary, provides DNA barcodes for about 8% of the species of this hemipteran suborder found in North America north of Mexico. Barcodes can enable the identification of many species of Auchenorrhyncha, but members of some species groups cannot be discriminated. Future use of DNA barcodes in regulatory, pest management, and environmental applications will be possible as the barcode library for Auchenorrhyncha expands to include more species and broader geographic coverage. PMID:25004106

The Mycetophila ruficollis Meigen (Diptera, Mycetophilidae) group in Europe: elucidating species delimitation with COI and ITS2 sequence data

PubMed Central

Jürgenstein, Siiri; Kurina, Olavi; Põldmaa, Kadri

2015-01-01

Abstract European species of the Mycetophila ruficollis group are compared on the basis of morphology and sequences of mitochondrial cytochrome oxidase subunit one (COI) and the ITS2 region of nuclear ribosomal DNA. The study represents the first evaluation of morphology-based species delimitation of closely related fungus gnat species by applying molecular information. Detailed descriptions and illustrations of the male terminalia are presented along with a key for the identification of all nine European species of the group. Phylogenetic analyses of molecular data generally supported the morphological species discrimination. The barcoding region of COI superseded ITS2 rDNA in resolving species. In the COI barcoding region interspecific differences ranged from 2.9 to 10.6% and the intraspecific distance from 0.08 to 0.8%. Only COI data distinguished between the similar and closely related Mycetophila ichneumonea and Mycetophila uninotata of which the latter was observed to include cryptic species. The host range of some species is suggested to be narrower than previously considered and to depend on the forest type. Presented evidence indicates the importance of analysing sequence data of morphologically very similar mycetophages reared from identified host fungi for elucidating species delimitation as well as their geographic and host ranges. New country records, viz. Estonia for Mycetophila evanida, Georgia for Mycetophila ichneumonea, Mycetophila idonea and Mycetophila ruficollis, and Norway for Mycetophila strobli, widen the known distribution ranges of these species. PMID:26167119

Identifying Canadian Freshwater Fishes through DNA Barcodes

PubMed Central

Hubert, Nicolas; Hanner, Robert; Holm, Erling; Mandrak, Nicholas E.; Taylor, Eric; Burridge, Mary; Watkinson, Douglas; Dumont, Pierre; Curry, Allen; Bentzen, Paul; Zhang, Junbin; April, Julien; Bernatchez, Louis

2008-01-01

Background DNA barcoding aims to provide an efficient method for species-level identifications using an array of species specific molecular tags derived from the 5′ region of the mitochondrial cytochrome c oxidase I (COI) gene. The efficiency of the method hinges on the degree of sequence divergence among species and species-level identifications are relatively straightforward when the average genetic distance among individuals within a species does not exceed the average genetic distance between sister species. Fishes constitute a highly diverse group of vertebrates that exhibit deep phenotypic changes during development. In this context, the identification of fish species is challenging and DNA barcoding provide new perspectives in ecology and systematics of fishes. Here we examined the degree to which DNA barcoding discriminate freshwater fish species from the well-known Canadian fauna, which currently encompasses nearly 200 species, some which are of high economic value like salmons and sturgeons. Methodology/Principal Findings We bi-directionally sequenced the standard 652 bp “barcode” region of COI for 1360 individuals belonging to 190 of the 203 Canadian freshwater fish species (95%). Most species were represented by multiple individuals (7.6 on average), the majority of which were retained as voucher specimens. The average genetic distance was 27 fold higher between species than within species, as K2P distance estimates averaged 8.3% among congeners and only 0.3% among concpecifics. However, shared polymorphism between sister-species was detected in 15 species (8% of the cases). The distribution of K2P distance between individuals and species overlapped and identifications were only possible to species group using DNA barcodes in these cases. Conversely, deep hidden genetic divergence was revealed within two species, suggesting the presence of cryptic species. Conclusions/Significance The present study evidenced that freshwater fish species can be efficiently identified through the use of DNA barcoding, especially the species complex of small-sized species, and that the present COI library can be used for subsequent applications in ecology and systematics. PMID:22423312

Bridging two scholarly islands enriches both: COI DNA barcodes for species identification versus human mitochondrial variation for the study of migrations and pathologies.

PubMed

Thaler, David S; Stoeckle, Mark Y

2016-10-01

DNA barcodes for species identification and the analysis of human mitochondrial variation have developed as independent fields even though both are based on sequences from animal mitochondria. This study finds questions within each field that can be addressed by reference to the other. DNA barcodes are based on a 648-bp segment of the mitochondrially encoded cytochrome oxidase I. From most species, this segment is the only sequence available. It is impossible to know whether it fairly represents overall mitochondrial variation. For modern humans, the entire mitochondrial genome is available from thousands of healthy individuals. SNPs in the human mitochondrial genome are evenly distributed across all protein-encoding regions arguing that COI DNA barcode is representative. Barcode variation among related species is largely based on synonymous codons. Data on human mitochondrial variation support the interpretation that most - possibly all - synonymous substitutions in mitochondria are selectively neutral. DNA barcodes confirm reports of a low variance in modern humans compared to nonhuman primates. In addition, DNA barcodes allow the comparison of modern human variance to many other extant animal species. Birds are a well-curated group in which DNA barcodes are coupled with census and geographic data. Putting modern human variation in the context of intraspecies variation among birds shows humans to be a single breeding population of average variance.

DNA barcoding and the identification of tree frogs (Amphibia: Anura: Rhacophoridae).

PubMed

Dang, Ning-Xin; Sun, Feng-Hui; Lv, Yun-Yun; Zhao, Bo-Han; Wang, Ji-Chao; Murphy, Robert W; Wang, Wen-Zhi; Li, Jia-Tang

2016-07-01

The DNA barcoding gene COI (cytochrome c oxidase subunit I) effectively identifies many species. Herein, we barcoded 172 individuals from 37 species belonging to nine genera in Rhacophoridae to test if the gene serves equally well to identify species of tree frogs. Phenetic neighbor joining and phylogenetic Bayesian inference were used to construct phylogenetic trees, which resolved all nine genera as monophyletic taxa except for Rhacophorus, two new matrilines for Liuixalus, and Polypedates leucomystax species complex. Intraspecific genetic distances ranged from 0.000 to 0.119 and interspecific genetic distances ranged from 0.015 to 0.334. Within Rhacophorus and Kurixalus, the intra- and interspecific genetic distances did not reveal an obvious barcode gap. Notwithstanding, we found that COI sequences unambiguously identified rhacophorid species and helped to discover likely new cryptic species via the synthesis of genealogical relationships and divergence patterns. Our results supported that COI is an effective DNA barcoding marker for Rhacophoridae.

DNA Barcoding the Medusozoa using mtCOI

NASA Astrophysics Data System (ADS)

Ortman, Brian D.; Bucklin, Ann; Pagès, Francesc; Youngbluth, Marsh

2010-12-01

The Medusozoa are a clade within the Cnidaria comprising the classes Hydrozoa, Scyphozoa, and Cubozoa. Identification of medusozoan species is challenging, even for taxonomic experts, due to their fragile forms and complex, morphologically-distinct life history stages. In this study 231 sequences for a portion of the mitochondrial Cytochrome Oxidase I (mtCOI) gene were obtained from 95 species of Medusozoans including; 84 hydrozoans (61 siphonophores, eight anthomedusae, four leptomedusae, seven trachymedusae, and four narcomedusae), 10 scyphozoans (three coronatae, four semaeostomae, two rhizostomae, and one stauromedusae), and one cubozoan. This region of mtCOI has been used as a DNA barcode (i.e., a molecular character for species recognition and discrimination) for a diverse array of taxa, including some Cnidaria. Kimura 2-parameter (K2P) genetic distances between sequence variants within species ranged from 0 to 0.057 (mean 0.013). Within the 13 genera for which multiple species were available, K2P distance between congeneric species ranged from 0.056 to 0.381. A cluster diagram generated by Neighbor Joining (NJ) using K2P distances reliably clustered all barcodes of the same species with ≥99% bootstrap support, ensuring accurate identification of species. Intra- and inter-specific variation of the mtCOI gene for the Medusozoa are appropriate for this gene to be used as a DNA barcode for species-level identification, but not for phylogenetic analysis or taxonomic classification of unknown sequences at higher taxonomic levels. This study provides a set of molecular tools that can be used to address questions of speciation, biodiversity, life-history, and population boundaries in the Medusozoa.

Cytochrome c oxidase subunit I barcoding of the green bee-eater (Merops orientalis).

PubMed

Arif, I A; Khan, H A; Shobrak, M; Williams, J

2011-10-21

DNA barcoding using mitochondrial cytochrome c oxidase subunit I (COI) is regarded as a standard method for species identification. Recent reports have also shown extended applications of COI gene analysis in phylogeny and molecular diversity studies. The bee-eaters are a group of near passerine birds in the family Meropidae. There are 26 species worldwide; five of them are found in Saudi Arabia. Until now, GenBank included a COI barcode for only one species of bee-eater, the European bee-eater (Merops apiaster). We sequenced the 694-bp segment of the COI gene of the green bee-eater M. orientalis and compared the sequences with those of M. apiaster. Pairwise sequence comparison showed 66 variable sites across all the eight sequences from both species, with an interspecific genetic distance of 0.0362. Two and one within-species variable sites were found, with genetic distances of 0.0005 and 0.0003 for M. apiaster and M. orientalis, respectively. This is the first study reporting barcodes for M. orientalis.

Patterns of DNA barcode variation in Canadian marine molluscs.

PubMed

Layton, Kara K S; Martel, André L; Hebert, Paul D N

2014-01-01

Molluscs are the most diverse marine phylum and this high diversity has resulted in considerable taxonomic problems. Because the number of species in Canadian oceans remains uncertain, there is a need to incorporate molecular methods into species identifications. A 648 base pair segment of the cytochrome c oxidase subunit I gene has proven useful for the identification and discovery of species in many animal lineages. While the utility of DNA barcoding in molluscs has been demonstrated in other studies, this is the first effort to construct a DNA barcode registry for marine molluscs across such a large geographic area. This study examines patterns of DNA barcode variation in 227 species of Canadian marine molluscs. Intraspecific sequence divergences ranged from 0-26.4% and a barcode gap existed for most taxa. Eleven cases of relatively deep (>2%) intraspecific divergence were detected, suggesting the possible presence of overlooked species. Structural variation was detected in COI with indels found in 37 species, mostly bivalves. Some indels were present in divergent lineages, primarily in the region of the first external loop, suggesting certain areas are hotspots for change. Lastly, mean GC content varied substantially among orders (24.5%-46.5%), and showed a significant positive correlation with nearest neighbour distances. DNA barcoding is an effective tool for the identification of Canadian marine molluscs and for revealing possible cases of overlooked species. Some species with deep intraspecific divergence showed a biogeographic partition between lineages on the Atlantic, Arctic and Pacific coasts, suggesting the role of Pleistocene glaciations in the subdivision of their populations. Indels were prevalent in the barcode region of the COI gene in bivalves and gastropods. This study highlights the efficacy of DNA barcoding for providing insights into sequence variation across a broad taxonomic group on a large geographic scale.

DNA barcoding for identifying synanthropic flesh flies (Diptera, Sarcophagidae) of Colombia.

PubMed

Buenaventura, Eliana; Valverde-Castro, César; Wolff, Marta; Triana-Chavez, Omar; Gómez-Palacio, Andrés

2018-06-01

The first step for a successful use of any insect as indicator in forensic sciences is providing a precise taxonomic identification at species level. Due to morphology-based identification of Sarcophaginae flies (Diptera, Sarcophagidae) is often difficult and requires strong taxonomic expertise, their use as forensic indicators has been limited. Consequently, molecular-based approaches have been accepted as alternative means of identification. Thus, we aimed testing the efficiency of the barcode region of the mitochondrial cytochrome oxidase subunit I (COI) gene for identification of synanthropic flesh flies of several species of the genera Peckia, Oxysarcodexia, Ravinia, and Tricharaea collected in Colombia. The 645-bp fragment of COI was amplified and aligned (215 parsimoniously informative variable sites). We calculated Kimura two-parameter genetic distances and reconstruct a Neighbor-Joining phylogenetic tree. Our Neighbor-Joining tree recovered all species as monophyletic, and confirmed a new species of the genus Ravinia as also indicated by the interspecific genetic divergences and morphological observations. We obtained a 100% of identification success. Thus, the COI barcodes showed efficiency as an alternative mean of identification of species of flesh flies collected on decaying organic matter in Colombia. Copyright © 2018 Elsevier B.V. All rights reserved.

DNA barcode reference data for the Korean herpetofauna and their applications.

PubMed

Jeong, Tae Jin; Jun, Jumin; Han, Sanghoon; Kim, Hyun Tae; Oh, Kyunghee; Kwak, Myounghai

2013-11-01

Recently, amphibians and reptiles have drawn attention because of declines in species and populations caused mainly by habitat loss, overexploitation and climate change. This study constructed a DNA barcode database for the Korean herpetofauna, including all the recorded amphibians and 68% of the recorded reptiles, to provide a useful, standardized tool for species identification in monitoring and management. A total of 103 individuals from 18 amphibian and 17 reptile species were used to generate barcode sequences using partial sequences of the mitochondrial cytochrome c oxidase subunit I (COI) gene and to compare it with other suggested barcode loci. Comparing 16S rRNA, cytochrome b (Cytb) and COI for amphibians and 12S rRNA, Cytb and COI for reptiles, our results revealed that COI is better than the other markers in terms of a high level of sequence variation without length variation and moderate amplification success. Although the COI marker had no clear barcoding gap because of the high level of intraspecific variation, all of the analysed individuals from the same species clustered together in a neighbour-joining tree. High intraspecific variation suggests the possibility of cryptic species. Finally, using this database, confiscated snakes were identified as Elaphe schrenckii, designated as endangered in Korea and a food contaminant was identified as the lizard Takydromus amurensis. © 2013 John Wiley & Sons Ltd.

Barcoding Neotropical birds: assessing the impact of nonmonophyly in a highly diverse group.

PubMed

Chaves, Bárbara R N; Chaves, Anderson V; Nascimento, Augusto C A; Chevitarese, Juliana; Vasconcelos, Marcelo F; Santos, Fabrício R

2015-07-01

In this study, we verified the power of DNA barcodes to discriminate Neotropical birds using Bayesian tree reconstructions of a total of 7404 COI sequences from 1521 species, including 55 Brazilian species with no previous barcode data. We found that 10.4% of species were nonmonophyletic, most likely due to inaccurate taxonomy, incomplete lineage sorting or hybridization. At least 0.5% of the sequences (2.5% of the sampled species) retrieved from GenBank were associated with database errors (poor-quality sequences, NuMTs, misidentification or unnoticed hybridization). Paraphyletic species (5.8% of the total) can be related to rapid speciation events leading to nonreciprocal monophyly between recently diverged sister species, or to absence of synapomorphies in the small COI region analysed. We also performed two series of genetic distance calculations under the K2P model for intraspecific and interspecific comparisons: the first included all COI sequences, and the second included only monophyletic taxa observed in the Bayesian trees. As expected, the mean and median pairwise distances were smaller for intraspecific than for interspecific comparisons. However, there was no precise 'barcode gap', which was shown to be larger in the monophyletic taxon data set than for the data from all species, as expected. Our results indicated that although database errors may explain some of the difficulties in the species discrimination of Neotropical birds, distance-based barcode assignment may also be compromised because of the high diversity of bird species and more complex speciation events in the Neotropics. © 2014 John Wiley & Sons Ltd.

Limitations of cytochrome oxidase I for the barcoding of Neritidae (Mollusca: Gastropoda) as revealed by Bayesian analysis.

PubMed

Chee, S Y

2015-05-25

The mitochondrial DNA (mtDNA) cytochrome oxidase I (COI) gene has been universally and successfully utilized as a barcoding gene, mainly because it can be amplified easily, applied across a wide range of taxa, and results can be obtained cheaply and quickly. However, in rare cases, the gene can fail to distinguish between species, particularly when exposed to highly sensitive methods of data analysis, such as the Bayesian method, or when taxa have undergone introgressive hybridization, over-splitting, or incomplete lineage sorting. Such cases require the use of alternative markers, and nuclear DNA markers are commonly used. In this study, a dendrogram produced by Bayesian analysis of an mtDNA COI dataset was compared with that of a nuclear DNA ATPS-α dataset, in order to evaluate the efficiency of COI in barcoding Malaysian nerites (Neritidae). In the COI dendrogram, most of the species were in individual clusters, except for two species: Nerita chamaeleon and N. histrio. These two species were placed in the same subcluster, whereas in the ATPS-α dendrogram they were in their own subclusters. Analysis of the ATPS-α gene also placed the two genera of nerites (Nerita and Neritina) in separate clusters, whereas COI gene analysis placed both genera in the same cluster. Therefore, in the case of the Neritidae, the ATPS-α gene is a better barcoding gene than the COI gene.

Developing diagnostic SNP panels for the identification of true fruit flies (Diptera: Tephritidae) within the limits of COI-based species delimitation

PubMed Central

2013-01-01

Background Rapid and reliable identification of quarantine pests is essential for plant inspection services to prevent introduction of invasive species. For insects, this may be a serious problem when dealing with morphologically similar cryptic species complexes and early developmental stages that lack distinctive characters useful for taxonomic identification. DNA based barcoding could solve many of these problems. The standard barcode fragment, an approx. 650 base pairs long sequence of the 5′end of the mitochondrial cytochrome oxidase I (COI), enables differentiation of a very wide range of arthropods. However, problems remain in some taxa, such as Tephritidae, where recent genetic differentiation among some of the described species hinders accurate molecular discrimination. Results In order to explore the full species discrimination potential of COI, we sequenced the barcoding region of the COI gene of a range of economically important Tephritid species and complemented these data with all GenBank and BOLD entries for the systematic group available as of January 2012. We explored the limits of species delimitation of this barcode fragment among 193 putative Tephritid species and established operational taxonomic units (OTUs), between which discrimination is reliably possible. Furthermore, to enable future development of rapid diagnostic assays based on this sequence information, we characterized all single nucleotide polymorphisms (SNPs) and established “near-minimal” sets of SNPs that differentiate among all included OTUs with at least three and four SNPs, respectively. Conclusions We found that although several species cannot be differentiated based on the genetic diversity observed in COI and hence form composite OTUs, 85% of all OTUs correspond to described species. Because our SNP panels are developed based on all currently available sequence information and rely on a minimal pairwise difference of three SNPs, they are highly reliable and hence represent an important resource for developing taxon-specific diagnostic assays. For selected cases, possible explanations that may cause composite OTUs are discussed. PMID:23718854

The campaign to DNA barcode all fishes, FISH-BOL.

PubMed

Ward, R D; Hanner, R; Hebert, P D N

2009-02-01

FISH-BOL, the Fish Barcode of Life campaign, is an international research collaboration that is assembling a standardized reference DNA sequence library for all fishes. Analysis is targeting a 648 base pair region of the mitochondrial cytochrome c oxidase I (COI) gene. More than 5000 species have already been DNA barcoded, with an average of five specimens per species, typically vouchers with authoritative identifications. The barcode sequence from any fish, fillet, fin, egg or larva can be matched against these reference sequences using BOLD; the Barcode of Life Data System (http://www.barcodinglife.org). The benefits of barcoding fishes include facilitating species identification, highlighting cases of range expansion for known species, flagging previously overlooked species and enabling identifications where traditional methods cannot be applied. Results thus far indicate that barcodes separate c. 98 and 93% of already described marine and freshwater fish species, respectively. Several specimens with divergent barcode sequences have been confirmed by integrative taxonomic analysis as new species. Past concerns in relation to the use of fish barcoding for species discrimination are discussed. These include hybridization, recent radiations, regional differentiation in barcode sequences and nuclear copies of the barcode region. However, current results indicate these issues are of little concern for the great majority of specimens.

Towards a global barcode library for Lymantria (Lepidoptera: Lymantriinae) tussock moths of biosecurity concern.

PubMed

deWaard, Jeremy R; Mitchell, Andrew; Keena, Melody A; Gopurenko, David; Boykin, Laura M; Armstrong, Karen F; Pogue, Michael G; Lima, Joao; Floyd, Robin; Hanner, Robert H; Humble, Leland M

2010-12-09

Detecting and controlling the movements of invasive species, such as insect pests, relies upon rapid and accurate species identification in order to initiate containment procedures by the appropriate authorities. Many species in the tussock moth genus Lymantria are significant forestry pests, including the gypsy moth Lymantria dispar L., and consequently have been a focus for the development of molecular diagnostic tools to assist in identifying species and source populations. In this study we expand the taxonomic and geographic coverage of the DNA barcode reference library, and further test the utility of this diagnostic method, both for species/subspecies assignment and for determination of geographic provenance of populations. Cytochrome oxidase I (COI) barcodes were obtained from 518 individuals and 36 species of Lymantria, including sequences assembled and generated from previous studies, vouchered material in public collections, and intercepted specimens obtained from surveillance programs in Canada. A maximum likelihood tree was constructed, revealing high bootstrap support for 90% of species clusters. Bayesian species assignment was also tested, and resulted in correct assignment to species and subspecies in all instances. The performance of barcoding was also compared against the commonly employed NB restriction digest system (also based on COI); while the latter is informative for discriminating gypsy moth subspecies, COI barcode sequences provide greater resolution and generality by encompassing a greater number of haplotypes across all Lymantria species, none shared between species. This study demonstrates the efficacy of DNA barcodes for diagnosing species of Lymantria and reinforces the view that the approach is an under-utilized resource with substantial potential for biosecurity and surveillance. Biomonitoring agencies currently employing the NB restriction digest system would gather more information by transitioning to the use of DNA barcoding, a change which could be made relatively seamlessly as the same gene region underlies both protocols.

DNA barcoding and phylogeny of Calidris and Tringa (Aves: Scolopacidae).

PubMed

Huang, Zuhao; Tu, Feiyun

2017-07-01

The avian genera Calidris and Tringa are the largest of the widespread family of Scolopacidae. The phylogeny of members of the two genera is still a matter of controversial. Mitochondrial cytochrome c oxidase subunit I (COI) can serve as a fast and accurate marker for the identification and phylogeny of animal species. In this study, we analyzed the COI barcodes of thirty-one species of the two genera. All the species had distinct COI sequences. Two hundred and twenty-one variable sites were identified. Kimura two-parameter distances were calculated between barcodes. Neighbor-joining and maximum likelihood methods were used to construct phylogenetic trees. All the species could be discriminated by their distinct clades in the phylogenetic trees. The phylogenetic trees grouped all the species of Calidris and Tringa into different monophyletic clade, respectively. COI data showed a well-supported phylogeny for Calidris and Tringa species.

DNA barcoding of freshwater fishes and the development of a quantitative qPCR assay for the species-specific detection and quantification of fish larvae from plankton samples.

PubMed

Loh, W K W; Bond, P; Ashton, K J; Roberts, D T; Tibbetts, I R

2014-08-01

The barcoding of mitochondrial cytochrome c oxidase subunit 1 (coI) gene was amplified and sequenced from 16 species of freshwater fishes found in Lake Wivenhoe (south-eastern Queensland, Australia) to support monitoring of reservoir fish populations, ecosystem function and water health. In this study, 630-650 bp sequences of the coI barcoding gene from 100 specimens representing 15 genera, 13 families and two subclasses of fishes allowed 14 of the 16 species to be identified and differentiated. The mean ± s.e. Kimura 2 parameter divergence within and between species was 0.52 ± 0.10 and 23.8 ± 2.20% respectively, indicating that barcodes can be used to discriminate most of the fish species accurately. The two terapontids, Amniataba percoides and Leiopotherapon unicolor, however, shared coI DNA sequences and could not be differentiated using this gene. A barcoding database was established and a qPCR assay was developed using coI sequences to identify and quantify proportional abundances of fish species in ichthyoplankton samples from Lake Wivenhoe. These methods provide a viable alternative to the time-consuming process of manually enumerating and identifying ichthyoplankton samples. © 2014 The Fisheries Society of the British Isles.

DNA Barcodes for Forensically Important Fly Species in Brazil.

PubMed

Koroiva, Ricardo; de Souza, Mirian S; Roque, Fabio de Oliveira; Pepinelli, Mateus

2018-04-07

Here, we analyze 248 DNA barcode sequences of 35 fly species of forensic importance in Brazil. DNA barcoding can be effectively used for specimen identification of these species, allowing the unambiguous identification of 31 species, an overall success rate of 88%. Our results show a high rate of success for molecular identification using DNA barcoding sequences and open new perspectives for immature species identification, a subject on which limited forensic investigations exist in Tropical regions. We also address the implications of building a robust forensic DNA barcode database. A geographic bias is recognized for the COI dataset available for forensically important fly species in Brazil, with concentration of sequences from specimens collected mainly in sites located in the Cerrado, Mata Atlântica, and Pampa biomes.

Limited efficiency of universal mini-barcode primers for DNA amplification from desert reptiles, birds and mammals.

PubMed

Arif, I A; Khan, H A; Al Sadoon, M; Shobrak, M

2011-10-31

In recent years, DNA barcoding has emerged as a powerful tool for species identification. We report an extended validation of a universal DNA mini-barcode for amplification of 130-bp COI segments from 23 specimens collected from a desert environment, including 11 reptiles, five mammals and seven birds. Besides the standard double-annealing protocol, we also tested a more stringent single-annealing protocol. The PCR success rate for the amplification of the mini-barcode region was: mammals (4/5), reptiles (5/11) and birds (4/7). These findings demonstrate the limited utility of universal primers for mini-barcoding, at least for these vertebrate taxa that we collected from the Saudi Arabian desert.

The Application of COI Gene for Species Identification of Forensically Important Muscid Flies (Diptera: Muscidae).

PubMed

Ren, Lipin; Chen, Wei; Shang, Yanjie; Meng, Fanming; Zha, Lagabaiyila; Wang, Yong; Guo, Yadong

2018-05-17

Muscid Flies (Diptera: Muscidae) are of great forensic importance due to their wide distribution, ubiquitous and synanthropic nature. They are frequently neglected as they tend to arrive at the corpses later than the flesh flies and blow flies. Moreover, the lack of species-level identification also hinders investigation of medicolegal purposes. To overcome the difficulty of morphological identification, molecular method has gained relevance. Cytochrome c oxidase subunit I (COI) gene has been widely utilized. Nonetheless, to achieve correct identification of an unknown sample, it is important to survey certain muscid taxa from its geographic distribution range. Accordingly, the aim of this study is to contribute more geographically specific. We sequenced the COI gene of 51 muscid specimens of 12 species, and added all correct sequences available in GenBank to yield a total data set of 125 COI sequences from 33 muscid species to evaluate the COI gene as a molecular diagnostic tool. The interspecific distances were extremely high (4.7-19.8%) in either the standard barcoding fragment (658 bp) or the long COI sequence (1,019-1,535 bp), demonstrating that these two genetic markers were nearly identical in the species identification. However, the intraspecific distances of the long COI sequences were significantly higher than the barcoding region for the conspecific species that geographical locations vary greatly. Therefore, genetic diversity presented in this study provides a reference for species identification of muscid flies. Nevertheless, further investigation and data from more muscid species are required to enhance the efficacy of species-level identification using COI gene as a genetic marker.

DNA Barcoding of Marine Metazoa

NASA Astrophysics Data System (ADS)

Bucklin, Ann; Steinke, Dirk; Blanco-Bercial, Leocadio

2011-01-01

More than 230,000 known species representing 31 metazoan phyla populate the world's oceans. Perhaps another 1,000,000 or more species remain to be discovered. There is reason for concern that species extinctions may outpace discovery, especially in diverse and endangered marine habitats such as coral reefs. DNA barcodes (i.e., short DNA sequences for species recognition and discrimination) are useful tools to accelerate species-level analysis of marine biodiversity and to facilitate conservation efforts. This review focuses on the usual barcode region for metazoans: a ˜648 base-pair region of the mitochondrial cytochrome c oxidase subunit I (COI) gene. Barcodes have also been used for population genetic and phylogeographic analysis, identification of prey in gut contents, detection of invasive species, forensics, and seafood safety. More controversially, barcodes have been used to delimit species boundaries, reveal cryptic species, and discover new species. Emerging frontiers are the use of barcodes for rapid and increasingly automated biodiversity assessment by high-throughput sequencing, including environmental barcoding and the use of barcodes to detect species for which formal identification or scientific naming may never be possible.

Deep COI sequencing of standardized benthic samples unveils overlooked diversity of Jordanian coral reefs in the northern Red Sea.

PubMed

Al-Rshaidat, Mamoon M D; Snider, Allison; Rosebraugh, Sydney; Devine, Amanda M; Devine, Thomas D; Plaisance, Laetitia; Knowlton, Nancy; Leray, Matthieu

2016-09-01

High-throughput sequencing (HTS) of DNA barcodes (metabarcoding), particularly when combined with standardized sampling protocols, is one of the most promising approaches for censusing overlooked cryptic invertebrate communities. We present biodiversity estimates based on sequencing of the cytochrome c oxidase subunit 1 (COI) gene for coral reefs of the Gulf of Aqaba, a semi-enclosed system in the northern Red Sea. Samples were obtained from standardized sampling devices (Autonomous Reef Monitoring Structures (ARMS)) deployed for 18 months. DNA barcoding of non-sessile specimens >2 mm revealed 83 OTUs in six phyla, of which only 25% matched a reference sequence in public databases. Metabarcoding of the 2 mm - 500 μm and sessile bulk fractions revealed 1197 OTUs in 15 animal phyla, of which only 4.9% matched reference barcodes. These results highlight the scarcity of COI data for cryptobenthic organisms of the Red Sea. Compared with data obtained using similar methods, our results suggest that Gulf of Aqaba reefs are less diverse than two Pacific coral reefs but much more diverse than an Atlantic oyster reef at a similar latitude. The standardized approaches used here show promise for establishing baseline data on biodiversity, monitoring the impacts of environmental change, and quantifying patterns of diversity at regional and global scales.

Testing the performance of a fragment of the COI gene to identify western Palaearctic stag beetle species (Coleoptera, Lucanidae).

PubMed

Cox, Karen; Thomaes, Arno; Antonini, Gloria; Zilioli, Michele; De Gelas, Koen; Harvey, Deborah; Solano, Emanuela; Audisio, Paolo; McKeown, Niall; Shaw, Paul; Minetti, Robert; Bartolozzi, Luca; Mergeay, Joachim

2013-12-30

Lucanidae) remains challenging, mainly due to the sexual dimorphism and the strong allometry in males. Such conjecture confounds taxonomic based conservation efforts that are urgently needed due to numerous threats to stag beetle biodiversity. Molecular tools could help solve the problem of identification of the different recognized taxa in the "Lucanus cervus complex" and in some related Palaearctic species. We investigated the potential use of a 670 bp region at the 3' end of the mitochondrial cytochrome c oxidase subunit I gene (COI) for barcoding purposes (different from the standard COI barcoding region). Well resolved species and subspecies were L. tetraodon, L. cervusakbesianus, L. c. laticornis, as well as the two eastern Asian outgroup taxa L. formosanus and L. hermani. Conversely, certain taxa could not be distinguished from each other based on K2P-distances and tree topologies: L. c. fabiani / L. (P.) barbarossa, L. c. judaicus / an unknown Lucanus species, L. c. cervus / L. c. turcicus / L. c. pentaphyllus / L. (P.) macrophyllus / L. ibericus. The relative roles of phenotypic plasticity, recurrent hybridisation and incomplete lineage sorting underlying taxonomic and phylogenetic discordances are discussed.

DNA barcoding of Bemisia tabaci complex (Hemiptera: Aleyrodidae) reveals southerly expansion of the dominant whitefly species on cotton in Pakistan.

PubMed

Ashfaq, Muhammad; Hebert, Paul D N; Mirza, M Sajjad; Khan, Arif M; Mansoor, Shahid; Shah, Ghulam S; Zafar, Yusuf

2014-01-01

Although whiteflies (Bemisia tabaci complex) are an important pest of cotton in Pakistan, its taxonomic diversity is poorly understood. As DNA barcoding is an effective tool for resolving species complexes and analyzing species distributions, we used this approach to analyze genetic diversity in the B. tabaci complex and map the distribution of B. tabaci lineages in cotton growing areas of Pakistan. Sequence diversity in the DNA barcode region (mtCOI-5') was examined in 593 whiteflies from Pakistan to determine the number of whitefly species and their distributions in the cotton-growing areas of Punjab and Sindh provinces. These new records were integrated with another 173 barcode sequences for B. tabaci, most from India, to better understand regional whitefly diversity. The Barcode Index Number (BIN) System assigned the 766 sequences to 15 BINs, including nine from Pakistan. Representative specimens of each Pakistan BIN were analyzed for mtCOI-3' to allow their assignment to one of the putative species in the B. tabaci complex recognized on the basis of sequence variation in this gene region. This analysis revealed the presence of Asia II 1, Middle East-Asia Minor 1, Asia 1, Asia II 5, Asia II 7, and a new lineage "Pakistan". The first two taxa were found in both Punjab and Sindh, but Asia 1 was only detected in Sindh, while Asia II 5, Asia II 7 and "Pakistan" were only present in Punjab. The haplotype networks showed that most haplotypes of Asia II 1, a species implicated in transmission of the cotton leaf curl virus, occurred in both India and Pakistan. DNA barcodes successfully discriminated cryptic species in B. tabaci complex. The dominant haplotypes in the B. tabaci complex were shared by India and Pakistan. Asia II 1 was previously restricted to Punjab, but is now the dominant lineage in southern Sindh; its southward spread may have serious implications for cotton plantations in this region.

Molecular barcoding of venomous snakes and species-specific multiplex PCR assay to identify snake groups for which antivenom is available in Thailand.

PubMed

Supikamolseni, A; Ngaoburanawit, N; Sumontha, M; Chanhome, L; Suntrarachun, S; Peyachoknagul, S; Srikulnath, K

2015-10-30

DNA barcodes of mitochondrial COI and Cytb genes were constructed from 54 specimens of 16 species for species identification. Intra- and interspecific sequence divergence of the COI gene (10 times) was greater than that of the Cytb gene (4 times), which suggests that the former gene may be a better marker than the latter for species delimitation in snakes. The COI barcode cut-off scores differed by more than 3% between most species, and the minimum interspecific divergence was greater than the maximum intraspecific divergence. Clustering analysis indicated that most species fell into monophyletic clades. These results suggest that these species could be reliably differentiated using COI DNA barcodes. Moreover, a novel species-specific multiplex PCR assay was developed to distinguish between Naja spp, Ophiophagus hannah, Trimeresurus spp, Hydrophiinae, Daboia siamensis, Bungarus fasciatus, and Calloselasma rhodostoma. Antivenom for these species is produced and kept by the Thai Red Cross for clinical use. Our novel PCR assay could easily be applied to venom and saliva samples and could be used effectively for the rapid and accurate identification of species during forensic work, conservation study, and medical research.

Identification of cephalopod species from the North and Baltic Seas using morphology, COI and 18S rDNA sequences

NASA Astrophysics Data System (ADS)

Gebhardt, Katharina; Knebelsberger, Thomas

2015-09-01

We morphologically analyzed 79 cephalopod specimens from the North and Baltic Seas belonging to 13 separate species. Another 29 specimens showed morphological features of either Alloteuthis mediaor Alloteuthis subulata or were found to be in between. Reliable identification features to distinguish between A. media and A. subulata are currently not available. The analysis of the DNA barcoding region of the COI gene revealed intraspecific distances (uncorrected p) ranging from 0 to 2.13 % (average 0.1 %) and interspecific distances between 3.31 and 22 % (average 15.52 %). All species formed monophyletic clusters in a neighbor-joining analysis and were supported by bootstrap values of ≥99 %. All COI haplotypes belonging to the 29 Alloteuthis specimens were grouped in one cluster. Neither COI nor 18S rDNA sequences helped to distinguish between the different Alloteuthis morphotypes. For species identification purposes, we recommend the use of COI, as it showed higher bootstrap support of species clusters and less amplification and sequencing failure compared to 18S. Our data strongly support the assumption that the genus Alloteuthis is only represented by a single species, at least in the North Sea. It remained unclear whether this species is A. subulata or A. media. All COI sequences including important metadata were uploaded to the Barcode of Life Data Systems and can be used as reference library for the molecular identification of more than 50 % of the cephalopod fauna known from the North and Baltic Seas.

Applications of three DNA barcodes in assorting intertidal red macroalgal flora in Qingdao, China

NASA Astrophysics Data System (ADS)

Zhao, Xiaobo; Pang, Shaojun; Shan, Tifeng; Liu, Feng

2013-03-01

This study is part of the endeavor to construct a comprehensive DNA barcoding database for common seaweeds in China. Identifications of red seaweeds, which have simple morphology and anatomy, are sometimes difficult solely depending on morphological characteristics. In recent years, DNA barcode technique has become a more and more effective tool to help solve some of the taxonomic difficulties. Some DNA markers such as COI (cytochrome oxidase subunit I) are proposed as standardized DNA barcodes for all seaweed species. In this study, COI, UPA (universal plastid amplicon, domain V of 23S rRNA), and ITS (nuclear internal transcribed spacer) were employed to analyze common species of intertidal red seaweeds in Qingdao (119.3°-121°E, 35.35°-37.09°N). The applicability of using one or a few combined barcodes to identify red seaweed species was tested. The results indicated that COI is a sensitive marker at species level. However, not all the tested species gave PCR amplification products due to lack of the universal primers. The second barcode UPA had effective universal primers but needed to be tested for the effectiveness of resolving closely related species. More than one ITS sequence types were found in some species in this investigation, which might lead to confusion in further analysis. Therefore ITS sequence is not recommended as a universal barcode for seaweeds identification.

DNA barcoding and development of species-specific markers for the identification of tea mosquito bugs (Miridae: Heteroptera) in India.

PubMed

Rebijith, K B; Asokan, R; Kumar, N K Krishna; Srikumar, K K; Ramamurthy, V V; Bhat, P Shivarama

2012-10-01

Rapid, accurate, and timely identification of insects as a group is important and challenging worldwide, as they outnumber all other animals in number and diversity. DNA barcoding is a method for the identification of species in a wide range of animal taxa, which uses the 5' region of the mitochondrial cytochrome c oxidase-I (CO-I). Yet another easy, accurate, and economical method of species discrimination is by developing species-specific markers, which produce specific amplicon for the species in question. The method is handy because it is not limited by life stages, sex, polymorphism, and other factors. Herein, we measured the usefulness of CO-I for the species discrimination of mirids in India viz. Helopeltis antonii Signoret, H. thievora Waterhouse, H. bradyi Waterhouse, and Pachypeltis maesarum Kirkaldy in their various life stages. Furthermore, our study showed the utility of species-specific markers in differentiating H. antonii (295) and H. bradyi (514) regardless of their life stages. Analysis of CO-I gene revealed <1% intraspecific divergence for all four species examined, whereas the interspecific distances ranged from 7 to 13%. This study showed that the DNA barcode and species-specific markers will aid the identification of mirids in India and will stand as a decisive tool in formulating integrated pest management (IPM) strategy, quick identification of invasive and cryptic species, haplotypes, biotypes, and other factors, if any.

DNA barcoding and morphological analysis for rapid identification of most economically important crop-infesting Sunn pests belonging to Eurygaster Laporte, 1833 (Hemiptera, Scutelleridae).

PubMed

Syromyatnikov, Mikhail Y; Golub, Victor B; Kokina, Anastasia V; Victoria A Soboleva; Popov, Vasily N

2017-01-01

The genus Eurygaster Laporte, 1833 includes ten species five of which inhabit the European part of Russia. The harmful species of the genus is E. integriceps . Eurygaster species identification based on the morphological traits is very difficult, while that of the species at the egg or larval stages is extremely difficult or impossible. Eurygaster integriceps , E. maura , and E. testudinaria differ only slightly between each other morphologically, E. maura and E. testudinaria being almost indiscernible. DNA barcoding based on COI sequences have shown that E. integriceps differs significantly from these closely related species, which enables its rapid and accurate identification. Based on COI nucleotide sequences, three species of Sunn pests, E. maura , E. testudinarius , E. dilaticollis , could not be differentiated from each other through DNA barcoding. The difference in the DNA sequences between the COI gene of E. integriceps and COI genes of E. maura and E. testudinarius was more than 4%. In the present study DNA barcoding of two Eurygaster species was performed for the first time on E. integriceps , the most dangerous pest in the genus, and E. dilaticollis that only inhabits natural ecosystems. The PCR-RFLP method was developed in this work for the rapid identification of E. integriceps .

DNA barcoding and morphological analysis for rapid identification of most economically important crop-infesting Sunn pests belonging to Eurygaster Laporte, 1833 (Hemiptera, Scutelleridae)

PubMed Central

Syromyatnikov, Mikhail Y.; Golub, Victor B.; Kokina, Anastasia V.; Victoria A. Soboleva; Popov, Vasily N.

2017-01-01

Abstract The genus Eurygaster Laporte, 1833 includes ten species five of which inhabit the European part of Russia. The harmful species of the genus is E. integriceps. Eurygaster species identification based on the morphological traits is very difficult, while that of the species at the egg or larval stages is extremely difficult or impossible. Eurygaster integriceps, E. maura, and E. testudinaria differ only slightly between each other morphologically, E. maura and E. testudinaria being almost indiscernible. DNA barcoding based on COI sequences have shown that E. integriceps differs significantly from these closely related species, which enables its rapid and accurate identification. Based on COI nucleotide sequences, three species of Sunn pests, E. maura, E. testudinarius, E. dilaticollis, could not be differentiated from each other through DNA barcoding. The difference in the DNA sequences between the COI gene of E. integriceps and COI genes of E. maura and E. testudinarius was more than 4%. In the present study DNA barcoding of two Eurygaster species was performed for the first time on E. integriceps, the most dangerous pest in the genus, and E. dilaticollis that only inhabits natural ecosystems. The PCR-RFLP method was developed in this work for the rapid identification of E. integriceps. PMID:29118620

Testing the performance of a fragment of the COI gene to identify western Palaearctic stag beetle species (Coleoptera, Lucanidae)

PubMed Central

Cox, Karen; Thomaes, Arno; Antonini, Gloria; Zilioli, Michele; De Gelas, Koen; Harvey, Deborah; Solano, Emanuela; Audisio, Paolo; McKeown, Niall; Shaw, Paul; Minetti, Robert; Bartolozzi, Luca; Mergeay, Joachim

2013-01-01

Abstract The taxonomy of stag beetles (Coleoptera: Lucanidae) remains challenging, mainly due to the sexual dimorphism and the strong allometry in males. Such conjecture confounds taxonomic based conservation efforts that are urgently needed due to numerous threats to stag beetle biodiversity. Molecular tools could help solve the problem of identification of the different recognized taxa in the “Lucanus cervus complex” and in some related Palaearctic species. We investigated the potential use of a 670 bp region at the 3’ end of the mitochondrial cytochrome c oxidase subunit I gene (COI) for barcoding purposes (different from the standard COI barcoding region). Well resolved species and subspecies were L. tetraodon, L. cervusakbesianus, L. c. laticornis, as well as the two eastern Asian outgroup taxa L. formosanus and L. hermani. Conversely, certain taxa could not be distinguished from each other based on K2P-distances and tree topologies: L. c. fabiani / L. (P.) barbarossa, L. c. judaicus / an unknown Lucanus species, L. c. cervus / L. c. turcicus / L. c. pentaphyllus / L. (P.) macrophyllus / L. ibericus. The relative roles of phenotypic plasticity, recurrent hybridisation and incomplete lineage sorting underlying taxonomic and phylogenetic discordances are discussed. PMID:24453554

Contrasting morphological and DNA barcode-suggested species boundaries among shallow-water amphipod fauna from the southern European Atlantic coast.

PubMed

Lobo, Jorge; Ferreira, Maria S; Antunes, Ilisa C; Teixeira, Marcos A L; Borges, Luisa M S; Sousa, Ronaldo; Gomes, Pedro A; Costa, Maria Helena; Cunha, Marina R; Costa, Filipe O

2017-02-01

In this study we compared DNA barcode-suggested species boundaries with morphology-based species identifications in the amphipod fauna of the southern European Atlantic coast. DNA sequences of the cytochrome c oxidase subunit I barcode region (COI-5P) were generated for 43 morphospecies (178 specimens) collected along the Portuguese coast which, together with publicly available COI-5P sequences, produced a final dataset comprising 68 morphospecies and 295 sequences. Seventy-five BINs (Barcode Index Numbers) were assigned to these morphospecies, of which 48 were concordant (i.e., 1 BIN = 1 species), 8 were taxonomically discordant, and 19 were singletons. Twelve species had matching sequences (<2% distance) with conspecifics from distant locations (e.g., North Sea). Seven morphospecies were assigned to multiple, and highly divergent, BINs, including specimens of Corophium multisetosum (18% divergence) and Dexamine spiniventris (16% divergence), which originated from sampling locations on the west coast of Portugal (only about 36 and 250 km apart, respectively). We also found deep divergence (4%-22%) among specimens of seven species from Portugal compared to those from the North Sea and Italy. The detection of evolutionarily meaningful divergence among populations of several amphipod species from southern Europe reinforces the need for a comprehensive re-assessment of the diversity of this faunal group.

DNA barcode variability and host plant usage of fruit flies (Diptera: Tephritidae) in Thailand.

PubMed

Kunprom, Chonticha; Pramual, Pairot

2016-10-01

The objectives of this study were to examine the genetic variation in fruit flies (Diptera: Tephritidae) in Thailand and to test the efficiency of the mitochondrial cytochrome c oxidase subunit I (COI) barcoding region for species-level identification. Twelve fruit fly species were collected from 24 host plant species of 13 families. The number of host plant species for each fruit fly species ranged between 1 and 11, with Bactrocera correcta found in the most diverse host plants. A total of 123 COI sequences were obtained from these fruit fly species. Sequences from the NCBI database were also included, for a total of 17 species analyzed. DNA barcoding identification analysis based on the best close match method revealed a good performance, with 94.4% of specimens correctly identified. However, many specimens (3.6%) had ambiguous identification, mostly due to intra- and interspecific overlap between members of the B. dorsalis complex. A phylogenetic tree based on the mitochondrial barcode sequences indicated that all species, except for the members of the B. dorsalis complex, were monophyletic with strong support. Our work supports recent calls for synonymization of these species. Divergent lineages were observed within B. correcta and B. tuberculata, and this suggested that these species need further taxonomic reexamination.

DNA barcoding as a tool for coral reef conservation

NASA Astrophysics Data System (ADS)

Neigel, J.; Domingo, A.; Stake, J.

2007-09-01

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.

Evaluating ethanol-based sample preservation to facilitate use of DNA barcoding in routine freshwater biomonitoring programs using benthic macroinvertebrates.

PubMed

Stein, Eric D; White, Bryan P; Mazor, Raphael D; Miller, Peter E; Pilgrim, Erik M

2013-01-01

Molecular methods, such as DNA barcoding, have the potential to enhance biomonitoring programs worldwide. Altering routinely used sample preservation methods to protect DNA from degradation may pose a potential impediment to application of DNA barcoding and metagenomics for biomonitoring using benthic macroinvertebrates. Using higher volumes or concentrations of ethanol, requirements for shorter holding times, or the need to include additional filtering may increase cost and logistical constraints to existing biomonitoring programs. To address this issue we evaluated the efficacy of various ethanol-based sample preservation methods at maintaining DNA integrity. We evaluated a series of methods that were minimally modified from typical field protocols in order to identify an approach that can be readily incorporated into existing monitoring programs. Benthic macroinvertebrates were collected from a minimally disturbed stream in southern California, USA and subjected to one of six preservation treatments. Ten individuals from five taxa were selected from each treatment and processed to produce DNA barcodes from the mitochondrial gene cytochrome c oxidase I (COI). On average, we obtained successful COI sequences (i.e. either full or partial barcodes) for between 93-99% of all specimens across all six treatments. As long as samples were initially preserved in 95% ethanol, successful sequencing of COI barcodes was not affected by a low dilution ratio of 2∶1, transfer to 70% ethanol, presence of abundant organic matter, or holding times of up to six months. Barcoding success varied by taxa, with Leptohyphidae (Ephemeroptera) producing the lowest barcode success rate, most likely due to poor PCR primer efficiency. Differential barcoding success rates have the potential to introduce spurious results. However, routine preservation methods can largely be used without adverse effects on DNA integrity.

Evaluating Ethanol-based Sample Preservation to Facilitate Use of DNA Barcoding in Routine Freshwater Biomonitoring Programs Using Benthic Macroinvertebrates

PubMed Central

Stein, Eric D.; White, Bryan P.; Mazor, Raphael D.; Miller, Peter E.; Pilgrim, Erik M.

2013-01-01

Molecular methods, such as DNA barcoding, have the potential to enhance biomonitoring programs worldwide. Altering routinely used sample preservation methods to protect DNA from degradation may pose a potential impediment to application of DNA barcoding and metagenomics for biomonitoring using benthic macroinvertebrates. Using higher volumes or concentrations of ethanol, requirements for shorter holding times, or the need to include additional filtering may increase cost and logistical constraints to existing biomonitoring programs. To address this issue we evaluated the efficacy of various ethanol-based sample preservation methods at maintaining DNA integrity. We evaluated a series of methods that were minimally modified from typical field protocols in order to identify an approach that can be readily incorporated into existing monitoring programs. Benthic macroinvertebrates were collected from a minimally disturbed stream in southern California, USA and subjected to one of six preservation treatments. Ten individuals from five taxa were selected from each treatment and processed to produce DNA barcodes from the mitochondrial gene cytochrome c oxidase I (COI). On average, we obtained successful COI sequences (i.e. either full or partial barcodes) for between 93–99% of all specimens across all six treatments. As long as samples were initially preserved in 95% ethanol, successful sequencing of COI barcodes was not affected by a low dilution ratio of 2∶1, transfer to 70% ethanol, presence of abundant organic matter, or holding times of up to six months. Barcoding success varied by taxa, with Leptohyphidae (Ephemeroptera) producing the lowest barcode success rate, most likely due to poor PCR primer efficiency. Differential barcoding success rates have the potential to introduce spurious results. However, routine preservation methods can largely be used without adverse effects on DNA integrity. PMID:23308097

DNA Barcoding of Bemisia tabaci Complex (Hemiptera: Aleyrodidae) Reveals Southerly Expansion of the Dominant Whitefly Species on Cotton in Pakistan

PubMed Central

Ashfaq, Muhammad; Hebert, Paul D. N.; Mirza, M. Sajjad; Khan, Arif M.; Mansoor, Shahid; Shah, Ghulam S.; Zafar, Yusuf

2014-01-01

Background Although whiteflies (Bemisia tabaci complex) are an important pest of cotton in Pakistan, its taxonomic diversity is poorly understood. As DNA barcoding is an effective tool for resolving species complexes and analyzing species distributions, we used this approach to analyze genetic diversity in the B. tabaci complex and map the distribution of B. tabaci lineages in cotton growing areas of Pakistan. Methods/Principal Findings Sequence diversity in the DNA barcode region (mtCOI-5′) was examined in 593 whiteflies from Pakistan to determine the number of whitefly species and their distributions in the cotton-growing areas of Punjab and Sindh provinces. These new records were integrated with another 173 barcode sequences for B. tabaci, most from India, to better understand regional whitefly diversity. The Barcode Index Number (BIN) System assigned the 766 sequences to 15 BINs, including nine from Pakistan. Representative specimens of each Pakistan BIN were analyzed for mtCOI-3′ to allow their assignment to one of the putative species in the B. tabaci complex recognized on the basis of sequence variation in this gene region. This analysis revealed the presence of Asia II 1, Middle East-Asia Minor 1, Asia 1, Asia II 5, Asia II 7, and a new lineage “Pakistan”. The first two taxa were found in both Punjab and Sindh, but Asia 1 was only detected in Sindh, while Asia II 5, Asia II 7 and “Pakistan” were only present in Punjab. The haplotype networks showed that most haplotypes of Asia II 1, a species implicated in transmission of the cotton leaf curl virus, occurred in both India and Pakistan. Conclusions DNA barcodes successfully discriminated cryptic species in B. tabaci complex. The dominant haplotypes in the B. tabaci complex were shared by India and Pakistan. Asia II 1 was previously restricted to Punjab, but is now the dominant lineage in southern Sindh; its southward spread may have serious implications for cotton plantations in this region. PMID:25099936

DNA-barcoding of forensically important blow flies (Diptera: Calliphoridae) in the Caribbean Region

PubMed Central

Agnarsson, Ingi

2017-01-01

Correct identification of forensically important insects, such as flies in the family Calliphoridae, is a crucial step for them to be used as evidence in legal investigations. Traditional identification based on morphology has been effective, but has some limitations when it comes to identifying immature stages of certain species. DNA-barcoding, using COI, has demonstrated potential for rapid and accurate identification of Calliphoridae, however, this gene does not reliably distinguish among some recently diverged species, raising questions about its use for delimitation of species of forensic importance. To facilitate DNA based identification of Calliphoridae in the Caribbean we developed a vouchered reference collection from across the region, and a DNA sequence database, and further added the nuclear ITS2 as a second marker to increase accuracy of identification through barcoding. We morphologically identified freshly collected specimens, did phylogenetic analyses and employed several species delimitation methods for a total of 468 individuals representing 19 described species. Our results show that combination of COI + ITS2 genes yields more accurate identification and diagnoses, and better agreement with morphological data, than the mitochondrial barcodes alone. All of our results from independent and concatenated trees and most of the species delimitation methods yield considerably higher diversity estimates than the distance based approach and morphology. Molecular data support at least 24 distinct clades within Calliphoridae in this study, recovering substantial geographic variation for Lucilia eximia, Lucilia retroversa, Lucilia rica and Chloroprocta idioidea, probably indicating several cryptic species. In sum, our study demonstrates the importance of employing a second nuclear marker for barcoding analyses and species delimitation of calliphorids, and the power of molecular data in combination with a complete reference database to enable identification of taxonomically and geographically diverse insects of forensic importance. PMID:28761780

DNA barcoding of the vegetable leafminer Liriomyza sativae Blanchard (Diptera: Agromyzidae) in Bangladesh

USDA-ARS?s Scientific Manuscript database

DNA barcoding revealed the presence of the polyphagous leafminer pest Liriomyza sativae Blanchard in Bangladesh. DNA barcode sequences for mitochondrial COI were generated for Agromyzidae larvae, pupae and adults collected from field populations across Bangladesh. BLAST sequence similarity searches ...

A DNA Barcoding Based Study to Identify Main Mosquito Species in Taiwan and its Difference from Those in Mainland China.

PubMed

Gao, Bo; Fang, Yiliang; Zhang, Jianqing; Wu, Rongquan; Xu, Baohai; Xie, Lianhui

2017-01-01

Mosquitoes can transmit many types of viruses such as West Nile virus and Zika virus and are responsible for a number of virus-causing diseases including malaria, dengue fever, yellow fever, lymphatic filariasis, and Japanese B encephalitis. On January 19, 2016, the first case of Zika virus infection was identified in Taiwan, which presents the need for studying the mosquito species in the Taiwan Strait and evaluating the risk of the outbreak of this infection. In this study, we have collected 144 mosquito specimens from 42 species belonging to nine genera from both sides of the Taiwan Strait during 2013 and 2014. We then applied the COI DNA Barcoding technique to classify the specimens and performed a phylogenetic analysis to infer the evolutionary history of these mosquitoes. Based on the analyses, we found that though the mosquitoes from different sides of the Taiwan Strait share a lot of commonality, they have a few regional specificities. Our results also suggested a very small divergences (1%~9%) between specimens from the same mosquito species and relatively large divergences (8%~25%) between specimens from different mosquito species. Within the same species, the divergence of specimens from the same region is significantly smaller than that between two regions. A few highly divergent species between Fujian and Taiwan (e.g., An.maculatus and Ae.elsiae) might be formed due to the so-called "cryptic evolutionary events", in which the species has differentiation into cryptic species due to geographical differences without changing morphological characteristics. In conclusion, the phylogenetic analyses showed a very similar taxonomy to the historical one based on morphological characteristics, validating again the application of COI DNA Barcoding technique in classifying mosquito species. However, there are also some inconsistencies between COI DNA Barcoding and historical taxonomy, which points out the differences between mosquito DNA and morphological characteristics and suggests the possibility to improve mosquito taxonomy based on DNA techniques. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Morphology versus DNA barcoding: two sides of the same coin. A case study of Ceutorhynchus erysimi and C. contractus identification.

PubMed

Stepanović, Svetlana; Kosovac, Andrea; Krstić, Oliver; Jović, Jelena; Toševski, Ivo

2016-08-01

Genotyping of 2 well-known weevil species from the genus Ceutorhynchus (Coleoptera: Curculionidae) distributed in west Palearctic, C. erysimi and C. contractus, revealed phenotype versus genotype inconsistencies in a set of 56 specimens (25 C. erysimi and 31 C. contractus) collected from 25 locations in Serbia and Montenegro. An analysis of the mitochondrial cytochrome oxidase subunit I gene (COI), widely used as a barcoding region, and a nuclear gene, elongation factor-1α (EF-1α), revealed stable genetic divergence among these species. The average uncorrected pairwise distances for the COI and EF-1α genes were 3.8%, and 1.3%, respectively, indicating 2 genetically well-segregated species. However, the genetic data were not congruent with the phenotypic characteristics of the studied specimens. In the first place, C. erysimi genotypes were attached to specimens with phenotypic characteristics of C. contractus. Species-specific PCR-RFLP assays for the barcoding gene COI were applied for the molecular identification of 101 additional specimens of both morphospecies (33 C. erysimi and 68 C. contractus) and were found to confirm this incongruity. The discrepancy between the genetic and morphological data raises the question of the accuracy of using a barcoding approach, as it may result in misleading conclusions about the taxonomic position of the studied organism. Additionally, the typological species concept shows considerable weakness when genetic data are not supported with phenotypic characteristics as in case of asymmetric introgression, which may cause certain problems, especially in applied studies such as biological control programs in which the biological properties of the studied organisms are the main focus. © 2015 Institute of Zoology, Chinese Academy of Sciences.

Searching for evidence of selection in avian DNA barcodes.

PubMed

Kerr, Kevin C R

2011-11-01

The barcode of life project has assembled a tremendous number of mitochondrial cytochrome c oxidase I (COI) sequences. Although these sequences were gathered to develop a DNA-based system for species identification, it has been suggested that further biological inferences may also be derived from this wealth of data. Recurrent selective sweeps have been invoked as an evolutionary mechanism to explain limited intraspecific COI diversity, particularly in birds, but this hypothesis has not been formally tested. In this study, I collated COI sequences from previous barcoding studies on birds and tested them for evidence of selection. Using this expanded data set, I re-examined the relationships between intraspecific diversity and interspecific divergence and sampling effort, respectively. I employed the McDonald-Kreitman test to test for neutrality in sequence evolution between closely related pairs of species. Because amino acid sequences were generally constrained between closely related pairs, I also included broader intra-order comparisons to quantify patterns of protein variation in avian COI sequences. Lastly, using 22 published whole mitochondrial genomes, I compared the evolutionary rate of COI against the other 12 protein-coding mitochondrial genes to assess intragenomic variability. I found no conclusive evidence of selective sweeps. Most evidence pointed to an overall trend of strong purifying selection and functional constraint. The COI protein did vary across the class Aves, but to a very limited extent. COI was the least variable gene in the mitochondrial genome, suggesting that other genes might be more informative for probing factors constraining mitochondrial variation within species. © 2011 Blackwell Publishing Ltd.

Diversity in a Cold Hot-Spot: DNA-Barcoding Reveals Patterns of Evolution among Antarctic Demosponges (Class Demospongiae, Phylum Porifera).

PubMed

Vargas, Sergio; Kelly, Michelle; Schnabel, Kareen; Mills, Sadie; Bowden, David; Wörheide, Gert

2015-01-01

The approximately 350 demosponge species that have been described from Antarctica represent a faunistic component distinct from that of neighboring regions. Sponges provide structure to the Antarctic benthos and refuge to other invertebrates, and can be dominant in some communities. Despite the importance of sponges in the Antarctic subtidal environment, sponge DNA barcodes are scarce but can provide insight into the evolutionary relationships of this unique biogeographic province. We sequenced the standard barcoding COI region for a comprehensive selection of sponges collected during expeditions to the Ross Sea region in 2004 and 2008, and produced DNA-barcodes for 53 demosponge species covering about 60% of the species collected. The Antarctic sponge communities are phylogenetically diverse, matching the diversity of well-sampled sponge communities in the Lusitanic and Mediterranean marine provinces in the Temperate Northern Atlantic for which molecular data are readily available. Additionally, DNA-barcoding revealed levels of in situ molecular evolution comparable to those present among Caribbean sponges. DNA-barcoding using the Segregating Sites Algorithm correctly assigned approximately 54% of the barcoded species to the morphologically determined species. A barcode library for Antarctic sponges was assembled and used to advance the systematic and evolutionary research of Antarctic sponges. We provide insights on the evolutionary forces shaping Antarctica's diverse sponge communities, and a barcode library against which future sequence data from other regions or depth strata of Antarctica can be compared. The opportunity for rapid taxonomic identification of sponge collections for ecological research is now at the horizon.

DNA barcoding as an aid for species identification in austral black flies (Insecta: Diptera: Simuliidae).

PubMed

Hernández-Triana, Luis M; Montes De Oca, Fernanda; Prosser, Sean W J; Hebert, Paul D N; Gregory, T Ryan; McMurtrie, Shelley

2017-04-01

In this paper, the utility of a partial sequence of the COI gene, the DNA barcoding region, for the identification of species of black flies in the austral region was assessed. Twenty-eight morphospecies were analyzed: eight of the genus Austrosimulium (four species in the subgenus Austrosimulium s. str., three species in the subgenus Novaustrosimulium, and one species unassigned to subgenus), two of the genus Cnesia, eight of Gigantodax, three of Paracnephia, one of Paraustrosimulium, and six of Simulium (subgenera Morops, Nevermannia, and Pternaspatha). The neighbour-joining tree derived from the DNA barcode sequences grouped most specimens according to species or species groups recognized by morphotaxonomic studies. Intraspecific sequence divergences within morphologically distinct species ranged from 0% to 1.8%, while higher divergences (2%-4.2%) in certain species suggested the presence of cryptic diversity. The existence of well-defined groups within S. simile revealed the likely inclusion of cryptic diversity. DNA barcodes also showed that specimens identified as C. dissimilis, C. nr. pussilla, and C. ornata might be conspecific, suggesting possible synonymy. DNA barcoding combined with a sound morphotaxonomic framework would provide an effective approach for the identification of black flies in the region.

DNA Barcoding Reveals High Cryptic Diversity of the Freshwater Halfbeak Genus Hemirhamphodon from Sundaland

PubMed Central

Zainal Abidin, Muchlisin; Pulungan, Chaidir Parlindungan

2016-01-01

DNA barcoding of the cytochrome oxidase subunit I (COI) gene was utilized to assess the species diversity of the freshwater halfbeak genus Hemirhamphodon. A total of 201 individuals from 46 locations in Peninsular Malaysia, north Borneo (Sarawak) and Sumatra were successfully amplified for 616 base pairs of the COI gene revealing 231 variable and 213 parsimony informative sites. COI gene trees showed that most recognized species form monophyletic clades with high bootstrap support. Pairwise within species comparisons exhibited a wide range of intraspecific diversity from 0.0% to 14.8%, suggesting presence of cryptic diversity. This finding was further supported by barcode gap analysis, ABGD and the constructed COI gene trees. In particular, H. pogonognathus from Kelantan (northeast Peninsular Malaysia) diverged from the other H. pogonognathus groups with distances ranging from 7.8 to 11.8%, exceeding the nearest neighbor taxon. High intraspecific diversity was also observed in H. byssus and H. kuekanthali, but of a lower magnitude. This study also provides insights into endemism and phylogeographic structuring, and limited support for the Paleo-drainage divergence hypothesis as a driver of speciation in the genus Hemirhamphodon. PMID:27657915

Molecular Barcoding of Aquatic Oligochaetes: Implications for Biomonitoring

PubMed Central

Vivien, Régis; Wyler, Sofia; Lafont, Michel; Pawlowski, Jan

2015-01-01

Aquatic oligochaetes are well recognized bioindicators of quality of sediments and water in watercourses and lakes. However, the difficult taxonomic determination based on morphological features compromises their more common use in eco-diagnostic analyses. To overcome this limitation, we investigated molecular barcodes as identification tool for broad range of taxa of aquatic oligochaetes. We report 185 COI and 52 ITS2 rDNA sequences for specimens collected in Switzerland and belonging to the families Naididae, Lumbriculidae, Enchytraeidae and Lumbricidae. Phylogenetic analyses allowed distinguishing 41 lineages separated by more than 10 % divergence in COI sequences. The lineage distinction was confirmed by Automatic Barcode Gap Discovery (ABGD) method and by ITS2 data. Our results showed that morphological identification underestimates the oligochaete diversity. Only 26 of the lineages could be assigned to morphospecies, of which seven were sequenced for the first time. Several cryptic species were detected within common morphospecies. Many juvenile specimens that could not be assigned morphologically have found their home after genetic analysis. Our study showed that COI barcodes performed very well as species identifiers in aquatic oligochaetes. Their easy amplification and good taxonomic resolution might help promoting aquatic oligochaetes as bioindicators for next generation environmental DNA biomonitoring of aquatic ecosystems. PMID:25856230

Profiling Nematode Communities in Unmanaged Flowerbed and Agricultural Field Soils in Japan by DNA Barcode Sequencing

PubMed Central

Morise, Hisashi; Miyazaki, Erika; Yoshimitsu, Shoko; Eki, Toshihiko

2012-01-01

Soil nematodes play crucial roles in the soil food web and are a suitable indicator for assessing soil environments and ecosystems. Previous nematode community analyses based on nematode morphology classification have been shown to be useful for assessing various soil environments. Here we have conducted DNA barcode analysis for soil nematode community analyses in Japanese soils. We isolated nematodes from two different environmental soils of an unmanaged flowerbed and an agricultural field using the improved flotation-sieving method. Small subunit (SSU) rDNA fragments were directly amplified from each of 68 (flowerbed samples) and 48 (field samples) isolated nematodes to determine the nucleotide sequence. Sixteen and thirteen operational taxonomic units (OTUs) were obtained by multiple sequence alignment from the flowerbed and agricultural field nematodes, respectively. All 29 SSU rDNA-derived OTUs (rOTUs) were further mapped onto a phylogenetic tree with 107 known nematode species. Interestingly, the two nematode communities examined were clearly distinct from each other in terms of trophic groups: Animal predators and plant feeders were markedly abundant in the flowerbed soils, in contrast, bacterial feeders were dominantly observed in the agricultural field soils. The data from the flowerbed nematodes suggests a possible food web among two different trophic nematode groups and plants (weeds) in the closed soil environment. Finally, DNA sequences derived from the mitochondrial cytochrome oxidase c subunit 1 (COI) gene were determined as a DNA barcode from 43 agricultural field soil nematodes. These nematodes were assigned to 13 rDNA-derived OTUs, but in the COI gene analysis were assigned to 23 COI gene-derived OTUs (cOTUs), indicating that COI gene-based barcoding may provide higher taxonomic resolution than conventional SSU rDNA-barcoding in soil nematode community analysis. PMID:23284767

DNA barcoding of five common stored-product pest species of genus Cryptolestes (Coleoptera: Laemophloeidae).

PubMed

Wang, Y J; Li, Z H; Zhang, S F; Varadínová, Z; Jiang, F; Kučerová, Z; Stejskal, V; Opit, G; Cao, Y; Li, F J

2014-10-01

Several species of the genus Cryptolestes Ganglbauer, 1899 (Coleoptera: Laemophloeidae) are commonly found in stored products. In this study, five species of Cryptolestes, with almost worldwide distribution, were obtained from laboratories in China, Czech Republic and the USA: Cryptolestes ferrugineus (Stephens, 1831), Cryptolestes pusillus (Schönherr, 1817), Cryptolestes turcicus (Grouvelle, 1876), Cryptolestes pusilloides (Steel & Howe, 1952) and Cryptolestes capensis (Waltl, 1834). Molecular identification based on a 658 bp fragment from the mitochondrial DNA cytochrome c oxidase subunit I (COI) was adopted to overcome some problems of morphological identification of Cryptolestes species. The utility of COI sequences as DNA barcodes in discriminating the five Cryptolestes species was evaluated on adults and larvae by analysing Kimura 2-parameter distances, phylogenetic tree and haplotype networks. The results showed that molecular approaches based on DNA barcodes were able to accurately identify these species. This is the first study using DNA barcoding to identify Cryptolestes species and the gathered DNA sequences will complement the biological barcode database.

DNA barcoding detected improper labelling and supersession of crab food served by restaurants in India.

PubMed

Vartak, Vivek Rohidas; Narasimmalu, Rajendran; Annam, Pavan Kumar; Singh, Dhirendra P; Lakra, Wazir S

2015-01-01

Detection of improper labelling of raw and processed seafood is of global importance for reducing commercial fraud and enhancing food safety. Crabs are crustaceans with intricate morphological as well as genetic divergence among species and are popular as seafood in restaurants. Owing to the high number of crab species available, it can be difficult to identify those included in particular food dishes, thus increasing the chance of supersession. DNA barcoding is an advanced technology for detecting improper food labelling and has been used successfully to authenticate seafood. This study identified 11 edible crab species from India by classical taxonomy and developed molecular barcodes with the cytochrome c oxidase I (COI) gene. These barcodes were used as reference barcodes for detecting any improper labelling of 50 restaurant crab samples. Neighbour-joining tree analysis with COI barcodes showed distinct clusters of restaurant samples with respective reference species. The study demonstrated 100% improper labelling of restaurant samples to cover up acts of inferior crab supersession. DNA barcoding successfully identified 11 edible crabs in accordance with classical taxonomy and discerned improper crab food labelling in restaurants of India. © 2014 Society of Chemical Industry.

First large-scale DNA barcoding assessment of reptiles in the biodiversity hotspot of Madagascar, based on newly designed COI primers.

PubMed

Nagy, Zoltán T; Sonet, Gontran; Glaw, Frank; Vences, Miguel

2012-01-01

DNA barcoding of non-avian reptiles based on the cytochrome oxidase subunit I (COI) gene is still in a very early stage, mainly due to technical problems. Using a newly developed set of reptile-specific primers for COI we present the first comprehensive study targeting the entire reptile fauna of the fourth-largest island in the world, the biodiversity hotspot of Madagascar. Representatives of the majority of Madagascan non-avian reptile species (including Squamata and Testudines) were sampled and successfully DNA barcoded. The new primer pair achieved a constantly high success rate (72.7-100%) for most squamates. More than 250 species of reptiles (out of the 393 described ones; representing around 64% of the known diversity of species) were barcoded. The average interspecific genetic distance within families ranged from a low of 13.4% in the Boidae to a high of 29.8% in the Gekkonidae. Using the average genetic divergence between sister species as a threshold, 41-48 new candidate (undescribed) species were identified. Simulations were used to evaluate the performance of DNA barcoding as a function of completeness of taxon sampling and fragment length. Compared with available multi-gene phylogenies, DNA barcoding correctly assigned most samples to species, genus and family with high confidence and the analysis of fewer taxa resulted in an increased number of well supported lineages. Shorter marker-lengths generally decreased the number of well supported nodes, but even mini-barcodes of 100 bp correctly assigned many samples to genus and family. The new protocols might help to promote DNA barcoding of reptiles and the established library of reference DNA barcodes will facilitate the molecular identification of Madagascan reptiles. Our results might be useful to easily recognize undescribed diversity (i.e. novel taxa), to resolve taxonomic problems, and to monitor the international pet trade without specialized expert knowledge.

First Large-Scale DNA Barcoding Assessment of Reptiles in the Biodiversity Hotspot of Madagascar, Based on Newly Designed COI Primers

PubMed Central

Nagy, Zoltán T.; Sonet, Gontran; Glaw, Frank; Vences, Miguel

2012-01-01

Background DNA barcoding of non-avian reptiles based on the cytochrome oxidase subunit I (COI) gene is still in a very early stage, mainly due to technical problems. Using a newly developed set of reptile-specific primers for COI we present the first comprehensive study targeting the entire reptile fauna of the fourth-largest island in the world, the biodiversity hotspot of Madagascar. Methodology/Principal Findings Representatives of the majority of Madagascan non-avian reptile species (including Squamata and Testudines) were sampled and successfully DNA barcoded. The new primer pair achieved a constantly high success rate (72.7–100%) for most squamates. More than 250 species of reptiles (out of the 393 described ones; representing around 64% of the known diversity of species) were barcoded. The average interspecific genetic distance within families ranged from a low of 13.4% in the Boidae to a high of 29.8% in the Gekkonidae. Using the average genetic divergence between sister species as a threshold, 41–48 new candidate (undescribed) species were identified. Simulations were used to evaluate the performance of DNA barcoding as a function of completeness of taxon sampling and fragment length. Compared with available multi-gene phylogenies, DNA barcoding correctly assigned most samples to species, genus and family with high confidence and the analysis of fewer taxa resulted in an increased number of well supported lineages. Shorter marker-lengths generally decreased the number of well supported nodes, but even mini-barcodes of 100 bp correctly assigned many samples to genus and family. Conclusions/Significance The new protocols might help to promote DNA barcoding of reptiles and the established library of reference DNA barcodes will facilitate the molecular identification of Madagascan reptiles. Our results might be useful to easily recognize undescribed diversity (i.e. novel taxa), to resolve taxonomic problems, and to monitor the international pet trade without specialized expert knowledge. PMID:22479636

The Role of DNA Barcodes in Understanding and Conservation of Mammal Diversity in Southeast Asia

PubMed Central

Francis, Charles M.; Borisenko, Alex V.; Ivanova, Natalia V.; Eger, Judith L.; Lim, Burton K.; Guillén-Servent, Antonio; Kruskop, Sergei V.; Mackie, Iain; Hebert, Paul D. N.

2010-01-01

Background Southeast Asia is recognized as a region of very high biodiversity, much of which is currently at risk due to habitat loss and other threats. However, many aspects of this diversity, even for relatively well-known groups such as mammals, are poorly known, limiting ability to develop conservation plans. This study examines the value of DNA barcodes, sequences of the mitochondrial COI gene, to enhance understanding of mammalian diversity in the region and hence to aid conservation planning. Methodology and Principal Findings DNA barcodes were obtained from nearly 1900 specimens representing 165 recognized species of bats. All morphologically or acoustically distinct species, based on classical taxonomy, could be discriminated with DNA barcodes except four closely allied species pairs. Many currently recognized species contained multiple barcode lineages, often with deep divergence suggesting unrecognized species. In addition, most widespread species showed substantial genetic differentiation across their distributions. Our results suggest that mammal species richness within the region may be underestimated by at least 50%, and there are higher levels of endemism and greater intra-specific population structure than previously recognized. Conclusions DNA barcodes can aid conservation and research by assisting field workers in identifying species, by helping taxonomists determine species groups needing more detailed analysis, and by facilitating the recognition of the appropriate units and scales for conservation planning. PMID:20838635

Species diversity of planktonic gastropods (Pteropoda and Heteropoda) from six ocean regions based on DNA barcode analysis

NASA Astrophysics Data System (ADS)

Jennings, Robert M.; Bucklin, Ann; Ossenbrügger, Holger; Hopcroft, Russell R.

2010-12-01

Pteropods and heteropods are two distinct groups of holoplanktonic gastropods whose species and genetic diversity remain poorly understood, despite their ubiquity in the world's oceans. Some species apparently attain near cosmopolitan distributions, implying long-distance dispersal or cryptic species assemblages. We present the first multi-regional and species-rich molecular dataset of holoplanktonic gastropods, comprising DNA barcodes from the mitochondrial cytochrome c oxidase I subunit gene (COI) from 115 individuals of 41 species sampled from six ocean regions across the globe. Molecular analysis and assessment of barcoding utility supported the validity of several morphological subspecies and forms (e.g. of Creseis virgula and Limacina helicina), while others were not supported (e.g. Cavolinia uncinata). Significant genetic variation was observed among conspecific specimens collected in different geographic regions for some species, particularly in euthecosomatous pteropods. Several species of euthecosomes showed no evidence of genetic separation among distant ocean regions. Overall, we suggest some taxonomic revision of the holoplanktonic gastropods will be required, pending a more complete molecular inventory of these groups.

Implications of Hybridization, NUMTs, and Overlooked Diversity for DNA Barcoding of Eurasian Ground Squirrels

PubMed Central

Ermakov, Oleg A.; Simonov, Evgeniy; Surin, Vadim L.; Titov, Sergey V.; Brandler, Oleg V.; Ivanova, Natalia V.; Borisenko, Alex V.

2015-01-01

The utility of DNA Barcoding for species identification and discovery has catalyzed a concerted effort to build the global reference library; however, many animal groups of economical or conservational importance remain poorly represented. This study aims to contribute DNA barcode records for all ground squirrel species (Xerinae, Sciuridae, Rodentia) inhabiting Eurasia and to test efficiency of this approach for species discrimination. Cytochrome c oxidase subunit 1 (COI) gene sequences were obtained for 97 individuals representing 16 ground squirrel species of which 12 were correctly identified. Taxonomic allocation of some specimens within four species was complicated by geographically restricted mtDNA introgression. Exclusion of individuals with introgressed mtDNA allowed reaching a 91.6% identification success rate. Significant COI divergence (3.5–4.4%) was observed within the most widespread ground squirrel species (Spermophilus erythrogenys, S. pygmaeus, S. suslicus, Urocitellus undulatus), suggesting the presence of cryptic species. A single putative NUMT (nuclear mitochondrial pseudogene) sequence was recovered during molecular analysis; mitochondrial COI from this sample was amplified following re-extraction of DNA. Our data show high discrimination ability of 100 bp COI fragments for Eurasian ground squirrels (84.3%) with no incorrect assessments, underscoring the potential utility of the existing reference librariy for the development of diagnostic ‘mini-barcodes’. PMID:25617768

Molecular taxonomy and identification within the Antarctic genus Trematomus (Notothenioidei, Teleostei): How valuable is barcoding with COI?

NASA Astrophysics Data System (ADS)

Lautredou, A.-C.; Bonillo, C.; Denys, G.; Cruaud, C.; Ozouf-Costaz, C.; Lecointre, G.; Dettai, A.

2010-08-01

The Trematominae are a particularly interesting subfamily within the antarctic suborder Notothenioidei (Teleostei). The 14 closely related species occupy a large range of ecological of niches, extremely useful for evolutionary and biogeography studies in the Antarctic Ocean. But some Trematomus species can be difficult to identify by using morphological criteria, specially young stages and damaged specimens. Molecular identification would therefore be highly useful, however the suitability of the cytochrome oxidase I gene in a barcoding approach needs to be assessed. We evaluated species delineation within the genus Trematomus comparing morphological identification, nuclear markers (the rhodopsin retrogene and a new nuclear marker pkd1: polycystic kidney disease 1) and COI. We show that Trematomus vicarius is not distinguishable from Trematomus bernacchii with the molecular markers used, and neither is Trematomus loennbergii from Trematomus lepidorhinus. We suggest that until this is investigated further, studies including these species list them as T. loennbergii/ T. lepidorhinus group, and keep voucher samples and specimens. Generally, COI gives a congruent result with the rhodopsin retrogene, and except for the previously cited species pairs, COI barcoding is efficient for identification in this group. Moreover pkd1 might not be suitable for a phylogenetic study at this scale for this group.

Genetic diversity of armored scales (Hemiptera: Diaspididae) and soft scales (Hemiptera: Coccidae) in Chile.

PubMed

Amouroux, P; Crochard, D; Germain, J-F; Correa, M; Ampuero, J; Groussier, G; Kreiter, P; Malausa, T; Zaviezo, T

2017-05-17

Scale insects (Sternorrhyncha: Coccoidea) are one of the most invasive and agriculturally damaging insect groups. Their management and the development of new control methods are currently jeopardized by the scarcity of identification data, in particular in regions where no large survey coupling morphological and DNA analyses have been performed. In this study, we sampled 116 populations of armored scales (Hemiptera: Diaspididae) and 112 populations of soft scales (Hemiptera: Coccidae) in Chile, over a latitudinal gradient ranging from 18°S to 41°S, on fruit crops, ornamental plants and trees. We sequenced the COI and 28S genes in each population. In total, 19 Diaspididae species and 11 Coccidae species were identified morphologically. From the 63 COI haplotypes and the 54 28S haplotypes uncovered, and using several DNA data analysis methods (Automatic Barcode Gap Discovery, K2P distance, NJ trees), up to 36 genetic clusters were detected. Morphological and DNA data were congruent, except for three species (Aspidiotus nerii, Hemiberlesia rapax and Coccus hesperidum) in which DNA data revealed highly differentiated lineages. More than 50% of the haplotypes obtained had no high-scoring matches with any of the sequences in the GenBank database. This study provides 63 COI and 54 28S barcode sequences for the identification of Coccoidea from Chile.

Pay Attention to the Overlooked Cryptic Diversity in Existing Barcoding Data: the Case of Mollusca with Character-Based DNA Barcoding.

PubMed

Zou, Shanmei; Li, Qi

2016-06-01

With the global biodiversity crisis, DNA barcoding aims for fast species identification and cryptic species diversity revelation. For more than 10 years, large amounts of DNA barcode data have been accumulating in publicly available databases, most of which were conducted by distance or tree-building methods that have often been argued, especially for cryptic species revelation. In this context, overlooked cryptic diversity may exist in the available barcoding data. The character-based DNA barcoding, however, has a good chance for detecting the overlooked cryptic diversity. In this study, marine mollusk was as the ideal case for detecting the overlooked potential cryptic species from existing cytochrome c oxidase I (COI) sequences with character-based DNA barcode. A total of 1081 COI sequences of mollusks, belonging to 176 species of 25 families of Gastropoda, Cephalopoda, and Lamellibranchia, were conducted by character analysis. As a whole, the character-based barcoding results were consistent with previous distance and tree-building analysis for species discrimination. More importantly, quite a number of species analyzed were divided into distinct clades with unique diagnostical characters. Based on the concept of cryptic species revelation of character-based barcoding, these species divided into separate taxonomic groups might be potential cryptic species. The detection of the overlooked potential cryptic diversity proves that the character-based barcoding mode possesses more advantages of revealing cryptic biodiversity. With the development of DNA barcoding, making the best use of barcoding data is worthy of our attention for species conservation.

A “Rosetta Stone” for metazoan zooplankton: DNA barcode analysis of species diversity of the Sargasso Sea (Northwest Atlantic Ocean)

NASA Astrophysics Data System (ADS)

Bucklin, Ann; Ortman, Brian D.; Jennings, Robert M.; Nigro, Lisa M.; Sweetman, Christopher J.; Copley, Nancy J.; Sutton, Tracey; Wiebe, Peter H.

2010-12-01

Species diversity of the metazoan holozooplankton assemblage of the Sargasso Sea, Northwest Atlantic Ocean, was examined through coordinated morphological taxonomic identification of species and DNA sequencing of a ˜650 base-pair region of mitochondrial cytochrome oxidase I (mtCOI) as a DNA barcode (i.e., short sequence for species recognition and discrimination). Zooplankton collections were made from the surface to 5,000 meters during April, 2006 on the R/V R.H. Brown. Samples were examined by a ship-board team of morphological taxonomists; DNA barcoding was carried out in both ship-board and land-based DNA sequencing laboratories. DNA barcodes were determined for a total of 297 individuals of 175 holozooplankton species in four phyla, including: Cnidaria (Hydromedusae, 4 species; Siphonophora, 47); Arthropoda (Amphipoda, 10; Copepoda, 34; Decapoda, 9; Euphausiacea, 10; Mysidacea, 1; Ostracoda, 27); and Mollusca (Cephalopoda, 8; Heteropoda, 6; Pteropoda, 15); and Chaetognatha (4). Thirty species of fish (Teleostei) were also barcoded. For all seven zooplankton groups for which sufficient data were available, Kimura-2-Parameter genetic distances were significantly lower between individuals of the same species (mean=0.0114; S.D. 0.0117) than between individuals of different species within the same group (mean=0.3166; S.D. 0.0378). This difference, known as the barcode gap, ensures that mtCOI sequences are reliable characters for species identification for the oceanic holozooplankton assemblage. In addition, DNA barcodes allow recognition of new or undescribed species, reveal cryptic species within known taxa, and inform phylogeographic and population genetic studies of geographic variation. The growing database of "gold standard" DNA barcodes serves as a Rosetta Stone for marine zooplankton, providing the key for decoding species diversity by linking species names, morphology, and DNA sequence variation. In light of the pivotal position of zooplankton in ocean food webs, their usefulness as rapid responders to environmental change, and the increasing scarcity of taxonomists, the use of DNA barcodes is an important and useful approach for rapid analysis of species diversity and distribution in the pelagic community.

DNA barcode assessment of Ceramiales (Rhodophyta) in the intertidal zone of the northwestern Yellow Sea

NASA Astrophysics Data System (ADS)

Du, Guoying; Wu, Feifei; Guo, Hao; Xue, Hongfan; Mao, Yunxiang

2015-05-01

A total of 142 specimens of Ceramiales (Rhodophyta) were collected each month from October 2011 to November 2012 in the intertidal zone of the northwestern Yellow Sea. These specimens covered 21 species, 14 genera, and four families. Cluster analyses show that the specimens had a high diversity for the three DNA markers, namely, partial large subunit rRNA gene (LSU), universal plastid amplicon (UPA), and partial mitochondrial cytochrome c oxidase subunit I gene (COI). No intraspecific divergence was found in our collection for these markers, except for a 1-3 bp divergence in the COI of Ceramium kondoi, Symphyocladia latiuscula, and Neosiphonia japonica. Because short DNA markers were used, the phylogenetic relationships of higher taxonomic levels were hard to evaluate with poor branch support. More than half species of our collection failed to find their matched sequences owing to shortage information of DNA barcodes for macroalgae in GenBank or BOLD (Barcode of Life Data) Systems. Three specimens were presumed as Heterosiphonia crispella by cluster analyses on DNA barcodes assisted by morphological identification, which was the first record in the investigated area, implying that it might be a cryptic or invasive species in the coastal area of northwestern Yellow Sea. In the neighbor-joining trees of all three DNA markers, Heterosiphonia japonica converged with Dasya spp. and was distant from the other Heterosiphonia spp., implying that H. japonica had affinities to the genus Dasya. The LSU and UPA markers amplified and sequenced easier than the COI marker across the Ceramiales species, but the COI had a higher ability to discriminate between species.

DNA barcoding of Arctic Ocean holozooplankton for species identification and recognition

NASA Astrophysics Data System (ADS)

Bucklin, Ann; Hopcroft, Russell R.; Kosobokova, Ksenia N.; Nigro, Lisa M.; Ortman, Brian D.; Jennings, Robert M.; Sweetman, Christopher J.

2010-01-01

Zooplankton species diversity and distribution are important measures of environmental change in the Arctic Ocean, and may serve as 'rapid-responders' of climate-induced changes in this fragile ecosystem. The scarcity of taxonomists hampers detailed and up-to-date monitoring of these patterns for the rarer and more problematic species. DNA barcodes (short DNA sequences for species recognition and discovery) provide an alternative approach to accurate identification of known species, and can speed routine analysis of zooplankton samples. During 2004-2008, zooplankton samples were collected during cruises to the central Arctic Ocean and Chukchi Sea. A ˜700 base-pair region of the mitochondrial cytochrome oxidase I (mtCOI) gene was amplified and sequenced for 82 identified specimens of 41 species, including cnidarians (six hydrozoans, one scyphozoan), arthropod crustaceans (five amphipods, 24 copepods, one decapod, and one euphausiid); two chaetognaths; and one nemertean. Phylogenetic analysis used the Neighbor-Joining algorithm with Kimura-2-Parameter (K-2-P) distances, with 1000-fold bootstrapping. K-2-P genetic distances between individuals of the same species ranged from 0.0 to 0.2; genetic distances between species ranged widely from 0.1 to 0.7. The mtCOI gene tree showed monophyly (at 100% bootstrap value) for each of the 26 species for which more than one individual was analyzed. Of seven genera for which more than one species was analyzed, four were shown to be monophyletic; three genera were not resolved. At higher taxonomic levels, only the crustacean order Copepoda was resolved, with bootstrap value of 83%. The mtCOI barcodes accurately discriminated and identified known species of 10 taxonomic groups of Arctic Ocean holozooplankton. A comprehensive DNA barcode database for the estimated 300 described species of Arctic holozooplankton will allow rapid assessment of species diversity and distribution in this climate-vulnerable ocean ecosystem.

Identifying the true oysters (Bivalvia: Ostreidae) with mitochondrial phylogeny and distance-based DNA barcoding.

PubMed

Liu, Jun; Li, Qi; Kong, Lingfeng; Yu, Hong; Zheng, Xiaodong

2011-09-01

Oysters (family Ostreidae), with high levels of phenotypic plasticity and wide geographic distribution, are a challenging group for taxonomists and phylogenetics. As a useful tool for molecular species identification, DNA barcoding might offer significant potential for oyster identification and taxonomy. This study used two mitochondrial fragments, cytochrome c oxidase I (COI) and the large ribosomal subunit (16S rDNA), to assess whether oyster species could be identified by phylogeny and distance-based DNA barcoding techniques. Relationships among species were estimated by the phylogenetic analyses of both genes, and then pairwise inter- and intraspecific genetic divergences were assessed. Species forming well-differentiated clades in the molecular phylogenies were identical for both genes even when the closely related species were included. Intraspecific variability of 16S rDNA overlapped with interspecific divergence. However, average intra- and interspecific genetic divergences for COI were 0-1.4% (maximum 2.2%) and 2.6-32.2% (minimum 2.2%), respectively, indicating the existence of a barcoding gap. These results confirm the efficacy of species identification in oysters via DNA barcodes and phylogenetic analysis. © 2011 Blackwell Publishing Ltd.

Determining Plant – Leaf Miner – Parasitoid Interactions: A DNA Barcoding Approach

PubMed Central

Derocles, Stéphane A. P.; Evans, Darren M.; Nichols, Paul C.; Evans, S. Aifionn; Lunt, David H.

2015-01-01

A major challenge in network ecology is to describe the full-range of species interactions in a community to create highly-resolved food-webs. We developed a molecular approach based on DNA full barcoding and mini-barcoding to describe difficult to observe plant – leaf miner – parasitoid interactions, consisting of animals commonly regarded as agricultural pests and their natural enemies. We tested the ability of universal primers to amplify the remaining DNA inside leaf miner mines after the emergence of the insect. We compared the results of a) morphological identification of adult specimens; b) identification based on the shape of the mines; c) the COI Mini-barcode (130 bp) and d) the COI full barcode (658 bp) fragments to accurately identify the leaf-miner species. We used the molecular approach to build and analyse a tri-partite ecological network of plant – leaf miner – parasitoid interactions. We were able to detect the DNA of leaf-mining insects within their feeding mines on a range of host plants using mini-barcoding primers: 6% for the leaves collected empty and 33% success after we observed the emergence of the leaf miner. We suggest that the low amplification success of leaf mines collected empty was mainly due to the time since the adult emerged and discuss methodological improvements. Nevertheless our approach provided new species-interaction data for the ecological network. We found that the 130 bp fragment is variable enough to identify all the species included in this study. Both COI fragments reveal that some leaf miner species could be composed of cryptic species. The network built using the molecular approach was more accurate in describing tri-partite interactions compared with traditional approaches based on morphological criteria. PMID:25710377

DNA barcoding for the identification of sand fly species (Diptera, Psychodidae, Phlebotominae) in Colombia.

PubMed

Contreras Gutiérrez, María Angélica; Vivero, Rafael J; Vélez, Iván D; Porter, Charles H; Uribe, Sandra

2014-01-01

Sand flies include a group of insects that are of medical importance and that vary in geographic distribution, ecology, and pathogen transmission. Approximately 163 species of sand flies have been reported in Colombia. Surveillance of the presence of sand fly species and the actualization of species distribution are important for predicting risks for and monitoring the expansion of diseases which sand flies can transmit. Currently, the identification of phlebotomine sand flies is based on morphological characters. However, morphological identification requires considerable skills and taxonomic expertise. In addition, significant morphological similarity between some species, especially among females, may cause difficulties during the identification process. DNA-based approaches have become increasingly useful and promising tools for estimating sand fly diversity and for ensuring the rapid and accurate identification of species. A partial sequence of the mitochondrial cytochrome oxidase gene subunit I (COI) is currently being used to differentiate species in different animal taxa, including insects, and it is referred as a barcoding sequence. The present study explored the utility of the DNA barcode approach for the identification of phlebotomine sand flies in Colombia. We sequenced 700 bp of the COI gene from 36 species collected from different geographic localities. The COI barcode sequence divergence within a single species was <2% in most cases, whereas this divergence ranged from 9% to 26.6% among different species. These results indicated that the barcoding gene correctly discriminated among the previously morphologically identified species with an efficacy of nearly 100%. Analyses of the generated sequences indicated that the observed species groupings were consistent with the morphological identifications. In conclusion, the barcoding gene was useful for species discrimination in sand flies from Colombia.

DNA Barcoding for the Identification of Sand Fly Species (Diptera, Psychodidae, Phlebotominae) in Colombia

PubMed Central

Contreras Gutiérrez, María Angélica; Vivero, Rafael J.; Vélez, Iván D.; Porter, Charles H.; Uribe, Sandra

2014-01-01

Sand flies include a group of insects that are of medical importance and that vary in geographic distribution, ecology, and pathogen transmission. Approximately 163 species of sand flies have been reported in Colombia. Surveillance of the presence of sand fly species and the actualization of species distribution are important for predicting risks for and monitoring the expansion of diseases which sand flies can transmit. Currently, the identification of phlebotomine sand flies is based on morphological characters. However, morphological identification requires considerable skills and taxonomic expertise. In addition, significant morphological similarity between some species, especially among females, may cause difficulties during the identification process. DNA-based approaches have become increasingly useful and promising tools for estimating sand fly diversity and for ensuring the rapid and accurate identification of species. A partial sequence of the mitochondrial cytochrome oxidase gene subunit I (COI) is currently being used to differentiate species in different animal taxa, including insects, and it is referred as a barcoding sequence. The present study explored the utility of the DNA barcode approach for the identification of phlebotomine sand flies in Colombia. We sequenced 700 bp of the COI gene from 36 species collected from different geographic localities. The COI barcode sequence divergence within a single species was <2% in most cases, whereas this divergence ranged from 9% to 26.6% among different species. These results indicated that the barcoding gene correctly discriminated among the previously morphologically identified species with an efficacy of nearly 100%. Analyses of the generated sequences indicated that the observed species groupings were consistent with the morphological identifications. In conclusion, the barcoding gene was useful for species discrimination in sand flies from Colombia. PMID:24454877

DNA Barcoding to Improve the Taxonomy of the Afrotropical Hoverflies (Insecta: Diptera: Syrphidae)

PubMed Central

Jordaens, Kurt; Goergen, Georg; Virgilio, Massimiliano; Backeljau, Thierry; Vokaer, Audrey; De Meyer, Marc

2015-01-01

The identification of Afrotropical hoverflies is very difficult because of limited recent taxonomic revisions and the lack of comprehensive identification keys. In order to assist in their identification, and to improve the taxonomy of this group, we constructed a reference dataset of 513 COI barcodes of 90 of the more common nominal species from Ghana, Togo, Benin and Nigeria (W Africa) and added ten publically available COI barcodes from nine nominal Afrotropical species to this (total: 523 COI barcodes; 98 nominal species; 26 genera). The identification accuracy of this dataset was evaluated with three methods (K2P distance-based, Neighbor-Joining (NJ) / Maximum Likelihood (ML) analysis, and using SpeciesIdentifier). Results of the three methods were highly congruent and showed a high identification success. Nine species pairs showed a low (< 0.03) mean interspecific K2P distance that resulted in several incorrect identifications. A high (> 0.03) maximum intraspecific K2P distance was observed in eight species and barcodes of these species not always formed single clusters in the NJ / ML analayses which may indicate the occurrence of cryptic species. Optimal K2P thresholds to differentiate intra- from interspecific K2P divergence were highly different among the three subfamilies (Eristalinae: 0.037, Syrphinae: 0.06, Microdontinae: 0.007–0.02), and among the different general suggesting that optimal thresholds are better defined at the genus level. In addition to providing an alternative identification tool, our study indicates that DNA barcoding improves the taxonomy of Afrotropical hoverflies by selecting (groups of) taxa that deserve further taxonomic study, and by attributing the unknown sex to species for which only one of the sexes is known. PMID:26473612

Identifying Fishes through DNA Barcodes and Microarrays.

PubMed

Kochzius, Marc; Seidel, Christian; Antoniou, Aglaia; Botla, Sandeep Kumar; Campo, Daniel; Cariani, Alessia; Vazquez, Eva Garcia; Hauschild, Janet; Hervet, Caroline; Hjörleifsdottir, Sigridur; Hreggvidsson, Gudmundur; Kappel, Kristina; Landi, Monica; Magoulas, Antonios; Marteinsson, Viggo; Nölte, Manfred; Planes, Serge; Tinti, Fausto; Turan, Cemal; Venugopal, Moleyur N; Weber, Hannes; Blohm, Dietmar

2010-09-07

International fish trade reached an import value of 62.8 billion Euro in 2006, of which 44.6% are covered by the European Union. Species identification is a key problem throughout the life cycle of fishes: from eggs and larvae to adults in fisheries research and control, as well as processed fish products in consumer protection. This study aims to evaluate the applicability of the three mitochondrial genes 16S rRNA (16S), cytochrome b (cyt b), and cytochrome oxidase subunit I (COI) for the identification of 50 European marine fish species by combining techniques of "DNA barcoding" and microarrays. In a DNA barcoding approach, neighbour Joining (NJ) phylogenetic trees of 369 16S, 212 cyt b, and 447 COI sequences indicated that cyt b and COI are suitable for unambiguous identification, whereas 16S failed to discriminate closely related flatfish and gurnard species. In course of probe design for DNA microarray development, each of the markers yielded a high number of potentially species-specific probes in silico, although many of them were rejected based on microarray hybridisation experiments. None of the markers provided probes to discriminate the sibling flatfish and gurnard species. However, since 16S-probes were less negatively influenced by the "position of label" effect and showed the lowest rejection rate and the highest mean signal intensity, 16S is more suitable for DNA microarray probe design than cty b and COI. The large portion of rejected COI-probes after hybridisation experiments (>90%) renders the DNA barcoding marker as rather unsuitable for this high-throughput technology. Based on these data, a DNA microarray containing 64 functional oligonucleotide probes for the identification of 30 out of the 50 fish species investigated was developed. It represents the next step towards an automated and easy-to-handle method to identify fish, ichthyoplankton, and fish products.

Identification, Discrimination, and Discovery of Species of Marine Planktonic Ostracods Using DNA Barcodes.

PubMed

Nigro, Lisa M; Angel, Martin V; Blachowiak-Samolyk, Katarzyna; Hopcroft, Russell R; Bucklin, Ann

2016-01-01

The Ostracoda (Crustacea; Class Ostracoda) is a diverse, frequently abundant, and ecologically important component of the marine zooplankton assemblage. There are more than 200 described species of marine planktonic ostracods, many of which (especially conspecific species) can be identified only by microscopic examination and dissection of fragile morphological characters. Given the complexity of species identification and increasing lack of expert taxonomists, DNA barcodes (short DNA sequences for species discrimination and identification) are particularly useful and necessary. Results are reported from analysis of 210 specimens of 78 species of marine planktonic ostracods, including two novel species, and 51 species for which barcodes have not been previously published. Specimens were collected during 2006 to 2008 from the Atlantic, Indian, and Southern Oceans, Greenland Sea and Gulf of Alaska. Samples were collected from surface to 5,000 m using various collection devices. DNA sequence variation was analyzed for a 598 base-pair region of the mitochondrial cytochrome oxidase subunit I (COI) gene. Kimura-2-Parameter (K2P) genetic distances within described species (mean = 0.010 ± 0.017 SD) were significantly smaller than between species (0.260 + 0.080), excluding eight taxa hypothesized to comprise cryptic species due to morphological variation (especially different size forms) and/or collection from different geographic regions. These taxa showed similar K2P distance values within (0.014 + 0.026) and between (0.221 ± 0.068) species. All K2P distances > 0.1 resulted from comparisons between identified or cryptic species, with no overlap between intra- and interspecific genetic distances. A Neighbor Joining tree resolved nearly all described species analyzed, with multiple sequences forming monophyletic clusters with high bootstrap values (typically 99%). Based on taxonomically and geographically extensive sampling and analysis (albeit with small sample sizes), the COI barcode region was shown to be a valuable character for discrimination, recognition, identification, and discovery of species of marine planktonic ostracods.

Identification, Discrimination, and Discovery of Species of Marine Planktonic Ostracods Using DNA Barcodes

PubMed Central

2016-01-01

The Ostracoda (Crustacea; Class Ostracoda) is a diverse, frequently abundant, and ecologically important component of the marine zooplankton assemblage. There are more than 200 described species of marine planktonic ostracods, many of which (especially conspecific species) can be identified only by microscopic examination and dissection of fragile morphological characters. Given the complexity of species identification and increasing lack of expert taxonomists, DNA barcodes (short DNA sequences for species discrimination and identification) are particularly useful and necessary. Results are reported from analysis of 210 specimens of 78 species of marine planktonic ostracods, including two novel species, and 51 species for which barcodes have not been previously published. Specimens were collected during 2006 to 2008 from the Atlantic, Indian, and Southern Oceans, Greenland Sea and Gulf of Alaska. Samples were collected from surface to 5,000 m using various collection devices. DNA sequence variation was analyzed for a 598 base-pair region of the mitochondrial cytochrome oxidase subunit I (COI) gene. Kimura-2-Parameter (K2P) genetic distances within described species (mean = 0.010 ± 0.017 SD) were significantly smaller than between species (0.260 + 0.080), excluding eight taxa hypothesized to comprise cryptic species due to morphological variation (especially different size forms) and/or collection from different geographic regions. These taxa showed similar K2P distance values within (0.014 + 0.026) and between (0.221 ± 0.068) species. All K2P distances > 0.1 resulted from comparisons between identified or cryptic species, with no overlap between intra- and interspecific genetic distances. A Neighbor Joining tree resolved nearly all described species analyzed, with multiple sequences forming monophyletic clusters with high bootstrap values (typically 99%). Based on taxonomically and geographically extensive sampling and analysis (albeit with small sample sizes), the COI barcode region was shown to be a valuable character for discrimination, recognition, identification, and discovery of species of marine planktonic ostracods. PMID:26730595

Unusually high genetic diversity in COI sequences of Chimarra obscura (Trichoptera: Philopotamidae)

EPA Science Inventory

Chimarra obscura (Walker 1852) is a philopotamid caddisfly found throughout much of North America. Using the COI DNA barcode locus, we have found unexpectedly high amounts of genetic diversity and distances within C. obscura. Of the approximately 150 specimens sampled, we have fo...

DNA Barcode Sequence Identification Incorporating Taxonomic Hierarchy and within Taxon Variability

PubMed Central

Little, Damon P.

2011-01-01

For DNA barcoding to succeed as a scientific endeavor an accurate and expeditious query sequence identification method is needed. Although a global multiple–sequence alignment can be generated for some barcoding markers (e.g. COI, rbcL), not all barcoding markers are as structurally conserved (e.g. matK). Thus, algorithms that depend on global multiple–sequence alignments are not universally applicable. Some sequence identification methods that use local pairwise alignments (e.g. BLAST) are unable to accurately differentiate between highly similar sequences and are not designed to cope with hierarchic phylogenetic relationships or within taxon variability. Here, I present a novel alignment–free sequence identification algorithm–BRONX–that accounts for observed within taxon variability and hierarchic relationships among taxa. BRONX identifies short variable segments and corresponding invariant flanking regions in reference sequences. These flanking regions are used to score variable regions in the query sequence without the production of a global multiple–sequence alignment. By incorporating observed within taxon variability into the scoring procedure, misidentifications arising from shared alleles/haplotypes are minimized. An explicit treatment of more inclusive terminals allows for separate identifications to be made for each taxonomic level and/or for user–defined terminals. BRONX performs better than all other methods when there is imperfect overlap between query and reference sequences (e.g. mini–barcode queries against a full–length barcode database). BRONX consistently produced better identifications at the genus–level for all query types. PMID:21857897

DNA barcode based wildlife forensics for resolving the origin of claw samples using a novel primer cocktail.

PubMed

Khedkar, Gulab D; Abhayankar, Shil Bapurao; Nalage, Dinesh; Ahmed, Shaikh Nadeem; Khedkar, Chandraprakash D

2016-11-01

Excessive wildlife hunting for commercial purposes can have negative impacts on biodiversity and may result in species extinction. To ensure compliance with legal statutes, forensic identification approaches relying on molecular markers may be used to identify the species of origin of animal material from hairs, claw, blood, bone, or meat. Using this approach, DNA sequences from the COI "barcoding" gene have been used to identify material from a number of domesticated animal species. However, many wild species of carnivores still present great challenges in generating COI barcodes using standard "universal" primer pairs. In the work presented here, the mitochondrial COI gene was successfully amplified using a novel primer cocktail, and the products were sequenced to determine the species of twenty one unknown samples of claw material collected as part of forensic wildlife case investigations. Sixteen of the unknown samples were recognized to have originated from either Panthera leo or P. pardus individuals. The remaining five samples could be identified only to the family level due to the absence of reference animal sequences. This is the first report on the use of COI sequences for the identification of P. pardus and P. leo from claw samples as part of forensic investigations in India. The study also highlights the need for adequate reference material to aid in the resolution of suspected cases of illegal wildlife harvesting.

DNA barcoding of fishes of Laguna de Bay, Philippines.

PubMed

Aquino, Luis Miguel G; Tango, Jazzlyn M; Canoy, Reynand Jay C; Fontanilla, Ian Kendrich C; Basiao, Zubaida U; Ong, Perry S; Quilang, Jonas P

2011-08-01

Laguna de Bay, the largest lake in the Philippines, is an important part of the country's fisheries industry. It is also home to a number of endemic fishes including Gobiopterus lacustris (Herre 1927) of family Gobiidae, Leiopotherapon plumbeus (Kner 1864) of family Terapontidae, Zenarchopterus philippinus (Peters 1868) of family Hemiramphidae and Arius manillensis Valenciennes 1840 of family Ariidae. Over the years, a steady decline has been observed in the abundance and diversity of native fishes in the lake due to anthropogenic disturbances. In this study, a total of 71 specimens of 18 different species belonging to 18 genera, 16 families, and seven orders were DNA barcoded using the mitochondrial cytochrome c oxidase subunit I (COI) gene. All of the fish species were discriminated by their COI sequences and one endemic species G. lacustris, showing deep genetic divergence, was highlighted for further taxonomic investigation. Average Kimura 2-parameter genetic distances within species, family, and order were 1.33%, 18.91%, and 24.22%, respectively. These values show that COI divergence increases as taxa become less exclusive. All of the COI sequences obtained were grouped together according to their species designation in the Neighbor-joining tree that was constructed. This study demonstrated that DNA barcoding has great potential as a tool for fast and accurate species identification and also for highlighting species that warrant further taxonomic investigation.

DNA barcoding amphibians and reptiles.

PubMed

Vences, Miguel; Nagy, Zoltán T; Sonet, Gontran; Verheyen, Erik

2012-01-01

Only a few major research programs are currently targeting COI barcoding of amphibians and reptiles (including chelonians and crocodiles), two major groups of tetrapods. Amphibian and reptile species are typically old, strongly divergent, and contain deep conspecific lineages which might lead to problems in species assignment with incomplete reference databases. As far as known, there is no single pair of COI primers that will guarantee a sufficient rate of success across all amphibian and reptile taxa, or within major subclades of amphibians and reptiles, which means that the PCR amplification strategy needs to be adjusted depending on the specific research question. In general, many more amphibian and reptile taxa have been sequenced for 16S rDNA, which for some purposes may be a suitable complementary marker, at least until a more comprehensive COI reference database becomes available. DNA barcoding has successfully been used to identify amphibian larval stages (tadpoles) in species-rich tropical assemblages. Tissue sampling, DNA extraction, and amplification of COI is straightforward in amphibians and reptiles. Single primer pairs are likely to have a failure rate between 5 and 50% if taxa of a wide taxonomic range are targeted; in such cases the use of primer cocktails or subsequent hierarchical usage of different primer pairs is necessary. If the target group is taxonomically limited, many studies have followed a strategy of designing specific primers which then allow an easy and reliable amplification of all samples.

DNA barcoding commercially important fish species of Turkey.

PubMed

Keskın, Emre; Atar, Hasan H

2013-09-01

DNA barcoding was used in the identification of 89 commercially important freshwater and marine fish species found in Turkish ichthyofauna. A total of 1765 DNA barcodes using a 654-bp-long fragment of the mitochondrial cytochrome c oxidase subunit I gene were generated for 89 commercially important freshwater and marine fish species found in Turkish ichthyofauna. These species belong to 70 genera, 40 families and 19 orders from class Actinopterygii, and all were associated with a distinct DNA barcode. Nine and 12 of the COI barcode clusters represent the first species records submitted to the BOLD and GenBank databases, respectively. All COI barcodes (except sequences of first species records) were matched with reference sequences of expected species, according to morphological identification. Average nucleotide frequencies of the data set were calculated as T = 29.7%, C = 28.2%, A = 23.6% and G = 18.6%. Average pairwise genetic distance among individuals were estimated as 0.32%, 9.62%, 17,90% and 22.40% for conspecific, congeneric, confamilial and within order, respectively. Kimura 2-parameter genetic distance values were found to increase with taxonomic level. For most of the species analysed in our data set, there is a barcoding gap, and an overlap in the barcoding gap exists for only two genera. Neighbour-joining trees were drawn based on DNA barcodes and all the specimens clustered in agreement with their taxonomic classification at species level. Results of this study supported DNA barcoding as an efficient molecular tool for a better monitoring, conservation and management of fisheries. © 2013 John Wiley & Sons Ltd.

Identification of marine traditional Chinese medicine dried seahorses in the traditional Chinese medicine market using DNA barcoding.

PubMed

Hou, Feixia; Wen, Longlian; Peng, Cheng; Guo, Jinlin

2018-01-01

Seahorse documented in Chinese pharmacopeia possess important medicinal efficacy and are used as an ingredient in traditional Chinese medicines. The growing international trade threatens the species. DNA barcoding holds a great application potentiality in wildlife conservation and might prevent the illegal trade of threatened species. The COI gene was used to identify seahorse, and nine Hippocampus species were found in the three large traditional Chinese medicines markets of China. All inter-specific genetic variations were larger than 2%. Mean genetic distances between species were 17-fold larger than those within the species. Phylogenetic tree showed that each species clustered in the appropriate branch. All results demonstrated that COI-based barcoding technique could be used to identify seahorse species and played a major role in monitoring the seahorse trade.

Molecular diagnosis of populational variants of Anthonomus grandis (Coleoptera: Curculionidae) in North America.

PubMed

Barr, Norman; Ruiz-Arce, Raul; Obregón, Oscar; De Leon, Rosita; Foster, Nelson; Reuter, Chris; Boratynski, Theodore; Vacek, Don

2013-02-01

The utility of the cytochrome oxidase I (COI) DNA sequence used for DNA barcoding and a Sequence Characterized Amplified Region for diagnosing boll weevil, Anthonomus grandis Boheman, variants was evaluated. Maximum likelihood analysis of COI DNA sequences from 154 weevils collected from the United States and Mexico supports previous evidence for limited gene flow between weevil populations on wild cotton and commercial cotton in northern Mexico and southern United States. The wild cotton populations represent a variant of the species called the thurberia weevil, which is not regarded as a significant pest. The 31 boll weevil COI haplotypes observed in the study form two distinct haplogroups (A and B) that are supported by five fixed nucleotide differences and a phylogenetic analysis. Although wild and commercial cotton populations are closely associated with specific haplogroups, there is not a fixed difference between the thurberia weevil variant and other populations. The Sequence Characterized Amplified Region marker generated a larger number of inconclusive results than the COI gene but also supported evidence of shared genotypes between wild and commercial cotton weevil populations. These methods provide additional markers that can assist in the identification of pest weevil populations but not definitively diagnose samples.

DNA reference libraries of French Guianese mosquitoes for barcoding and metabarcoding

PubMed Central

Leroy, Céline; Guidez, Amandine; Dusfour, Isabelle; Girod, Romain; Dejean, Alain; Murienne, Jérôme

2017-01-01

The mosquito family (Diptera: Culicidae) constitutes the most medically important group of arthropods because certain species are vectors of human pathogens. In some parts of the world, the diversity is so high that the accurate delimitation and/or identification of species is challenging. A DNA-based identification system for all animals has been proposed, the so-called DNA barcoding approach. In this study, our objectives were (i) to establish DNA barcode libraries for the mosquitoes of French Guiana based on the COI and the 16S markers, (ii) to compare distance-based and tree-based methods of species delimitation to traditional taxonomy, and (iii) to evaluate the accuracy of each marker in identifying specimens. A total of 266 specimens belonging to 75 morphologically identified species or morphospecies were analyzed allowing us to delimit 86 DNA clusters with only 21 of them already present in the BOLD database. We thus provide a substantial contribution to the global mosquito barcoding initiative. Our results confirm that DNA barcodes can be successfully used to delimit and identify mosquito species with only a few cases where the marker could not distinguish closely related species. Our results also validate the presence of new species identified based on morphology, plus potential cases of cryptic species. We found that both COI and 16S markers performed very well, with successful identifications at the species level of up to 98% for COI and 97% for 16S when compared to traditional taxonomy. This shows great potential for the use of metabarcoding for vector monitoring and eco-epidemiological studies. PMID:28575090

DNA barcoding coupled to HRM analysis as a new and simple tool for the authentication of Gadidae fish species.

PubMed

Fernandes, Telmo J R; Costa, Joana; Oliveira, M Beatriz P P; Mafra, Isabel

2017-09-01

This work aimed to exploit the use of DNA mini-barcodes combined with high resolution melting (HRM) for the authentication of gadoid species: Atlantic cod (Gadus morhua), Pacific cod (Gadus macrocephalus), Alaska pollock (Theragra chalcogramma) and saithe (Pollachius virens). Two DNA barcode regions, namely cytochrome c oxidase subunit I (COI) and cytochrome b (cytb), were analysed in silico to identify genetic variability among the four species and used, subsequently, to develop a real-time PCR method coupled with HRM analysis. The cytb mini-barcode enabled best discrimination of the target species with a high level of confidence (99.3%). The approach was applied successfully to identify gadoid species in 30 fish-containing foods, 30% of which were not as declared on the label. Herein, a novel approach for rapid, simple and cost-effective discrimination/clustering, as a tool to authenticate Gadidae fish species, according to their genetic relationship, is proposed. Copyright © 2017 Elsevier Ltd. All rights reserved.

Utility of DNA barcoding for rapid and accurate assessment of bat diversity in Malaysia in the absence of formally described species.

PubMed

Wilson, J-J; Sing, K-W; Halim, M R A; Ramli, R; Hashim, R; Sofian-Azirun, M

2014-02-19

Bats are important flagship species for biodiversity research; however, diversity in Southeast Asia is considerably underestimated in the current checklists and field guides. Incorporation of DNA barcoding into surveys has revealed numerous species-level taxa overlooked by conventional methods. Inclusion of these taxa in inventories provides a more informative record of diversity, but is problematic as these species lack formal description. We investigated how frequently documented, but undescribed, bat taxa are encountered in Peninsular Malaysia. We discuss whether a barcode library provides a means of recognizing and recording these taxa across biodiversity inventories. Tissue was sampled from bats trapped at Pasir Raja, Dungun Terengganu, Peninsular Malaysia. The DNA was extracted and the COI barcode region amplified and sequenced. We identified 9 species-level taxa within our samples, based on analysis of the DNA barcodes. Six specimens matched to four previously documented taxa considered candidate species but currently lacking formal taxonomic status. This study confirms the high diversity of bats within Peninsular Malaysia (9 species in 13 samples) and demonstrates how DNA barcoding allows for inventory and documentation of known taxa lacking formal taxonomic status.

Towards a comprehensive barcode library for arctic life - Ephemeroptera, Plecoptera, and Trichoptera of Churchill, Manitoba, Canada

PubMed Central

2009-01-01

Background This study reports progress in assembling a DNA barcode reference library for Ephemeroptera, Plecoptera, and Trichoptera ("EPTs") from a Canadian subarctic site, which is the focus of a comprehensive biodiversity inventory using DNA barcoding. These three groups of aquatic insects exhibit a moderate level of species diversity, making them ideal for testing the feasibility of DNA barcoding for routine biotic surveys. We explore the correlation between the morphological species delineations, DNA barcode-based haplotype clusters delimited by a sequence threshold (2%), and a threshold-free approach to biodiversity quantification--phylogenetic diversity. Results A DNA barcode reference library is built for 112 EPT species for the focal region, consisting of 2277 COI sequences. Close correspondence was found between EPT morphospecies and haplotype clusters as designated using a standard threshold value. Similarly, the shapes of taxon accumulation curves based upon haplotype clusters were very similar to those generated using phylogenetic diversity accumulation curves, but were much more computationally efficient. Conclusion The results of this study will facilitate other lines of research on northern EPTs and also bode well for rapidly conducting initial biodiversity assessments in unknown EPT faunas. PMID:20003245

DNA Barcoding Reveals Hidden Diversity of Sand Flies (Diptera: Psychodidae) at Fine and Broad Spatial Scales in Brazilian Endemic Regions for Leishmaniasis.

PubMed

Rodrigues, Bruno Leite; Carvalho-Costa, Luís Fernando; Pinto, Israel de Souza; Rebêlo, José Manuel Macário

2018-03-17

Sand fly (Diptera: Psychodidae) taxonomy is complex and time-consuming, which hampers epidemiological efforts directed toward controlling leishmaniasis in endemic regions such as northeastern Brazil. Here, we used a fragment of the mitochondrial cytochrome c oxidase I (COI) gene to identify sand fly species in Maranhão State (northeastern Brazil) and to assess cryptic diversity occurring at different spatial scales. For this, we obtained 148 COI sequences of 15 sand fly species (10 genera) from Maranhão (fine spatial scale), and joined them to COI sequences from other Brazilian localities (distant about 2,000 km from Maranhão, broad spatial scale) available in GenBank. We revealed cases of cryptic diversity in sand flies both at fine (Lutzomyia longipalpis (Lutz and Neiva) and Evandromyia termitophila (Martins, Falcão and Silva)) and broad spatial scales (Migonemyia migonei (França), Pressatia choti (Floch and Abonnenc), Psychodopygus davisi (Root), Sciopemyia sordellii (Shannon and Del Ponte), and Bichromomyia flaviscutellata (Mangabeira)). We argue that in the case of Bi. flaviscutellata, the cryptic diversity is associated with a putative new species. Cases in which DNA taxonomy was not as effective as morphological identification possibly involved recent speciation and/or introgressive hybridization, highlighting the need for integrative approaches to identify some sand fly species. Finally, we provide the first barcode sequences for four species (Brumptomyia avellari (Costa Lima), Evandromyia infraspinosa (Mangabeira), Evandromyia evandroi (Costa Lima and Antunes), and Psychodopygus complexus (Mangabeira)), which will be useful for further molecular identification of neotropical species.

DNA Barcoding Identifies Argentine Fishes from Marine and Brackish Waters

PubMed Central

Mabragaña, Ezequiel; Díaz de Astarloa, Juan Martín; Hanner, Robert; Zhang, Junbin; González Castro, Mariano

2011-01-01

Background DNA barcoding has been advanced as a promising tool to aid species identification and discovery through the use of short, standardized gene targets. Despite extensive taxonomic studies, for a variety of reasons the identification of fishes can be problematic, even for experts. DNA barcoding is proving to be a useful tool in this context. However, its broad application is impeded by the need to construct a comprehensive reference sequence library for all fish species. Here, we make a regional contribution to this grand challenge by calibrating the species discrimination efficiency of barcoding among 125 Argentine fish species, representing nearly one third of the known fauna, and examine the utility of these data to address several key taxonomic uncertainties pertaining to species in this region. Methodology/Principal Findings Specimens were collected and morphologically identified during crusies conducted between 2005 and 2008. The standard BARCODE fragment of COI was amplified and bi-directionally sequenced from 577 specimens (mean of 5 specimens/species), and all specimens and sequence data were archived and interrogated using analytical tools available on the Barcode of Life Data System (BOLD; www.barcodinglife.org). Nearly all species exhibited discrete clusters of closely related haplogroups which permitted the discrimination of 95% of the species (i.e. 119/125) examined while cases of shared haplotypes were detected among just three species-pairs. Notably, barcoding aided the identification of a new species of skate, Dipturus argentinensis, permitted the recognition of Genypterus brasiliensis as a valid species and questions the generic assignment of Paralichthys isosceles. Conclusions/Significance This study constitutes a significant contribution to the global barcode reference sequence library for fishes and demonstrates the utility of barcoding for regional species identification. As an independent assessment of alpha taxonomy, barcodes provide robust support for most morphologically based taxon concepts and also highlight key areas of taxonomic uncertainty worthy of reappraisal. PMID:22174860

Taxonomy, distribution and ecology of the order Phyllodocida (Annelida, Polychaeta) in deep-sea habitats around the Iberian margin

NASA Astrophysics Data System (ADS)

Ravara, Ascensão; Ramos, Diana; Teixeira, Marcos A. L.; Costa, Filipe O.; Cunha, Marina R.

2017-03-01

The polychaetes of the order Phyllodocida (excluding Nereidiformia and Phyllodociformia incertae sedis) collected from deep-sea habitats of the Iberian margin (Bay of Biscay, Horseshoe continental rise, Gulf of Cadiz and Alboran Sea), and Atlantic seamounts (Gorringe Bank, Atlantis and Nameless) are reported herein. Thirty-six species belonging to seven families - Acoetidae, Pholoidae, Polynoidae, Sigalionidae, Glyceridae, Goniadidae and Phyllodocidae, were identified. Amended descriptions and/or new illustrations are given for the species Allmaniella setubalensis, Anotochaetonoe michelbhaudi, Lepidasthenia brunnea and Polynoe sp. Relevant taxonomical notes are provided for other seventeen species. Allmaniella setubalensis, Anotochaetonoe michelbhaudi, Harmothoe evei, Eumida longicirrata and Glycera noelae, previously known only from their type localities were found in different deep-water places of the studied areas and constitute new records for the Iberian margin. The geographic distributions and the bathymetric range of thirteen and fifteen species, respectively, are extended. The morphology-based biodiversity inventory was complemented with DNA sequences of the mitochondrial barcode region (COI barcodes) providing a molecular tag for future reference. Twenty new sequences were obtained for nine species in the families Acoetidae, Glyceridae and Polynoidae and for three lineages within the Phylodoce madeirensis complex (Phyllodocidae). A brief analysis of the newly obtained sequences and publicly available COI barcode data for the genera herein reported, highlighted several cases of unclear taxonomic assignments, which need further study.

Linking adults and immatures of South African marine fishes.

PubMed

Steinke, Dirk; Connell, Allan D; Hebert, Paul D N

2016-11-01

The early life-history stages of fishes are poorly known, impeding acquisition of the identifications needed to monitor larval recruitment and year-class strength. A comprehensive database of COI sequences, linked to authoritatively identified voucher specimens, promises to change this situation, representing a significant advance for fisheries science. Barcode records were obtained from 2526 early larvae and pelagic eggs of fishes collected on the inshore shelf within 5 km of the KwaZulu-Natal coast, about 50 km south of Durban, South Africa. Barcodes were also obtained from 3215 adults, representing 946 South African fish species. Using the COI reference library on BOLD based on adults, 89% of the immature fishes could be identified to a species level; they represented 450 species. Most of the uncertain sequences could be assigned to a genus, family, or order; only 92 specimens (4%) were unassigned. Accumulation curves based on inference of phylogenetic diversity indicate near-completeness of the collecting effort. The entire set of adult and larval fishes included 1006 species, representing 43% of all fish species known from South African waters. However, this total included 189 species not previously recorded from this region. The fact that almost 90% of the immatures gained a species identification demonstrates the power and completeness of the DNA barcode reference library for fishes generated during the 10 years of FishBOL.

DNA Barcodes of Asian Houbara Bustard (Chlamydotis undulata macqueenii)

PubMed Central

Arif, Ibrahim A.; Khan, Haseeb A.; Williams, Joseph B.; Shobrak, Mohammad; Arif, Waad I.

2012-01-01

Populations of Houbara Bustards have dramatically declined in recent years. Captive breeding and reintroduction programs have had limited success in reviving population numbers and thus new technological solutions involving molecular methods are essential for the long term survival of this species. In this study, we sequenced the 694 bp segment of COI gene of the four specimens of Asian Houbara Bustard (Chlamydotis undulata macqueenii). We also compared these sequences with earlier published barcodes of 11 individuals comprising different families of the orders Gruiformes, Ciconiiformes, Podicipediformes and Crocodylia (out group). The pair-wise sequence comparison showed a total of 254 variable sites across all the 15 sequences from different taxa. Three of the four specimens of Houbara Bustard had an identical sequence of COI gene and one individual showed a single nucleotide difference (G > A transition at position 83). Within the bustard family (Otididae), comparison among the three species (Asian Houbara Bustard, Great Bustard (Otis tarda) and the Little Bustard (Tetrax tetrax)), representing three different genera, showed 116 variable sites. For another family (Rallidae), the intra-family variable sites among the individuals of four different genera were found to be 146. The COI genetic distances among the 15 individuals varied from 0.000 to 0.431. Phylogenetic analysis using 619 bp nucleotide segment of COI clearly discriminated all the species representing different genera, families and orders. All the four specimens of Houbara Bustard formed a single clade and are clearly separated from other two individuals of the same family (Otis tarda and Tetrax tetrax). The nucleotide sequence of partial segment of COI gene effectively discriminated the closely related species. This is the first study reporting the barcodes of Houbara Bustard and would be helpful in future molecular studies, particularly for the conservation of this threatened bird in Saudi Arabia. PMID:22408462

DNA barcoding discriminates freshwater fishes from southeastern Nigeria and provides river system-level phylogeographic resolution within some species.

PubMed

Nwani, Christopher D; Becker, Sven; Braid, Heather E; Ude, Emmanuel F; Okogwu, Okechukwu I; Hanner, Robert

2011-10-01

Fishes are the main animal protein source for human beings and play a vital role in aquatic ecosystems and food webs. Fish identification can be challenging, especially in the tropics (due to high diversity), and this is particularly true for larval forms or fragmentary remains. DNA barcoding, which uses the 5' region of the mitochondrial cytochrome c oxidase subunit I (COI) as a target gene, is an efficient method for standardized species-level identification for biodiversity assessment and conservation, pending the establishment of reference sequence libraries. In this study, fishes were collected from three rivers in southeastern Nigeria, identified morphologically, and imaged digitally. DNA was extracted, PCR-amplified, and the standard barcode region was bidirectionally sequenced for 363 individuals belonging to 70 species in 38 genera. All specimen provenance data and associated sequence information were recorded in the barcode of life data systems (BOLD; www.barcodinglife.org ). Analytical tools on BOLD were used to assess the performance of barcoding to identify species. Using neighbor-joining distance comparison, the average genetic distance was 60-fold higher between species than within species, as pairwise genetic distance estimates averaged 10.29% among congeners and only 0.17% among conspecifics. Despite low levels of divergence within species, we observed river system-specific haplotype partitioning within eight species (11.4% of all species). Our preliminary results suggest that DNA barcoding is very effective for species identification of Nigerian freshwater fishes.

Barcoding Fauna Bavarica: Myriapoda – a contribution to DNA sequence-based identifications of centipedes and millipedes (Chilopoda, Diplopoda)

PubMed Central

Spelda, Jörg; Reip, Hans S.; Oliveira–Biener, Ulla; Melzer, Roland R.

2011-01-01

Abstract We give a first account of our ongoing barcoding activities on Bavarian myriapods in the framework of the Barcoding Fauna Bavarica project and IBOL, the International Barcode of Life. Having analyzed 126 taxa (including 122 species) belonging to all major German chilopod and diplopod lineages, often using four or more specimens each, at the moment our species stock includes 82% of the diplopods and 65% of the chilopods found in Bavaria, southern Germany. The partial COI sequences allow correct identification of more than 95% of the current set of Bavarian species. Moreover, most of the myriapod orders and families appear as distinct clades in neighbour-joining trees, although the phylogenetic relationships between them are not always depicted correctly. We give examples of (1) high interspecific sequence variability among closely related species; (2) low interspecific variability in some chordeumatidan genera, indicating that recent speciations cannot be resolved with certainty using COI DNA barcodes; (3) high intraspecific variation in some genera, suggesting the existence of cryptic lineages; and (4) the possible polyphyly of some taxa, i.e. the chordeumatidan genus Ochogona. This shows that, in addition to species identification, our data may be useful in various ways in the context of species delimitations, taxonomic revisions and analyses of ongoing speciation processes. PMID:22303099

Comparison of four species-delimitation methods applied to a DNA barcode data set of insect larvae for use in routine bioassessment for use in routine bioassessment

EPA Science Inventory

Species delimitation (grouping individuals into distinct taxonomic groups) is an essential part of evolutionary, conservation, and molecular ecology. Deoxyribonucleic acid (DNA) barcodes, short fragments of the cytochrome c oxidase subunit I (COI) gene, are being used in environm...

DNA barcodes to identify species and explore diversity in the Adelgidae (Insecta: Hemiptera: Aphidoidea)

Treesearch

R.G. Foottit; H.E.L. Maw; N.P. Havill; R.G. Ahern; M.E. Montgomery

2009-01-01

The Adelgidae are relatively small, cryptic insects, exhibiting complex life cycles with parthenogenetic reproduction. Due to these characteristics, the taxonomy of the group is problematic. Here, we test the effectiveness of the standard 658-bp barcode fragment from the 5'-end of the mitochondrial cytochrome c oxidase 1 gene (COI) in...

A new species of the genus Hyalessa (Hemiptera, Cicadidae) from China, with DNA barcoding data and a key to related species.

PubMed

Wang, Xu; Qiu, Yue; Wei, Cong

2016-03-02

One new species of the genus Hyalessa China, H. wangi sp. nov., from Yunnan, China is described. Partial mitochondrial COI gene (DNA barcoding) of this new species is sequenced and uploaded to GenBank. A key to all species of Hyalessa is provided.

Testing DNA barcode performance in 1000 species of European lepidoptera: large geographic distances have small genetic impacts.

PubMed

Huemer, Peter; Mutanen, Marko; Sefc, Kristina M; Hebert, Paul D N

2014-01-01

This study examines the performance of DNA barcodes (mt cytochrome c oxidase 1 gene) in the identification of 1004 species of Lepidoptera shared by two localities (Finland, Austria) that are 1600 km apart. Maximum intraspecific distances for the pooled data were less than 2% for 880 species (87.6%), while deeper divergence was detected in 124 species. Despite such variation, the overall DNA barcode library possessed diagnostic COI sequences for 98.8% of the taxa. Because a reference library based on Finnish specimens was highly effective in identifying specimens from Austria, we conclude that barcode libraries based on regional sampling can often be effective for a much larger area. Moreover, dispersal ability (poor, good) and distribution patterns (disjunct, fragmented, continuous, migratory) had little impact on levels of intraspecific geographic divergence. Furthermore, the present study revealed that, despite the intensity of past taxonomic work on European Lepidoptera, nearly 20% of the species shared by Austria and Finland require further work to clarify their status. Particularly discordant BIN (Barcode Index Number) cases should be checked to ascertain possible explanatory factors such as incorrect taxonomy, hybridization, introgression, and Wolbachia infections.

DNA barcoding of human-biting black flies (Diptera: Simuliidae) in Thailand.

PubMed

Pramual, Pairot; Thaijarern, Jiraporn; Wongpakam, Komgrit

2016-12-01

Black flies (Diptera: Simuliidae) are important insect vectors and pests of humans and animals. Accurate identification, therefore, is important for control and management. In this study, we used mitochondrial cytochrome oxidase I (COI) barcoding sequences to test the efficiency of species identification for the human-biting black flies in Thailand. We used human-biting specimens because they enabled us to link information with previous studies involving the immature stages. Three black fly taxa, Simulium nodosum, S. nigrogilvum and S. doipuiense complex, were collected. The S. doipuiense complex was confirmed for the first time as having human-biting habits. The COI sequences revealed considerable genetic diversity in all three species. Comparisons to a COI sequence library of black flies in Thailand and in a public database indicated a high efficiency for specimen identification for S. nodosum and S. nigrogilvum, but this method was not successful for the S. doipuiense complex. Phylogenetic analyses revealed two divergent lineages in the S. doipuiense complex. Human-biting specimens formed a separate clade from other members of this complex. The results are consistent with the Barcoding Index Number System (BINs) analysis that found six BINs in the S. doipuiense complex. Further taxonomic work is needed to clarify the species status of these human-biting specimens. Copyright © 2016 Elsevier B.V. All rights reserved.

[Principles for molecular identification of traditional Chinese materia medica using DNA barcoding].

PubMed

Chen, Shi-Lin; Yao, Hui; Han, Jian-Ping; Xin, Tian-Yi; Pang, Xiao-Hui; Shi, Lin-Chun; Luo, Kun; Song, Jing-Yuan; Hou, Dian-Yun; Shi, Shang-Mei; Qian, Zhong-Zhi

2013-01-01

Since the research of molecular identification of Chinese Materia Medica (CMM) using DNA barcode is rapidly developing and popularizing, the principle of this method is approved to be listed in the Supplement of the Pharmacopoeia of the People's Republic of China. Based on the study on comprehensive samples, the DNA barcoding systems have been established to identify CMM, i.e. ITS2 as a core barcode and psbA-trnH as a complementary locus for identification of planta medica, and COI as a core barcode and ITS2 as a complementary locus for identification of animal medica. This article introduced the principle of molecular identification of CMM using DNA barcoding and its drafting instructions. Furthermore, its application perspective was discussed.

Identification of Scleractinian Coral Recruits Using Fluorescent Censusing and DNA Barcoding Techniques

PubMed Central

Hsu, Chia-Min; de Palmas, Stéphane; Kuo, Chao-Yang; Denis, Vianney; Chen, Chaolun Allen

2014-01-01

The identification of coral recruits has been problematic due to a lack of definitive morphological characters being available for higher taxonomic resolution. In this study, we tested whether fluorescent detection of coral recruits used in combinations of different DNA-barcoding markers (cytochrome oxidase I gene [COI], open reading frame [ORF], and nuclear Pax-C intron [PaxC]) could be useful for increasing the resolution of coral spat identification in ecological studies. One hundred and fifty settlement plates were emplaced at nine sites on the fringing reefs of Kenting National Park in southern Taiwan between April 2011 and September 2012. A total of 248 living coral spats and juveniles (with basal areas ranging from 0.21 to 134.57 mm2) were detected on the plates with the aid of fluorescent light and collected for molecular analyses. Using the COI DNA barcoding technique, 90.3% (224/248) of coral spats were successfully identified into six genera, including Acropora, Isopora, Montipora, Pocillopora, Porites, and Pavona. PaxC further separated I. cuneata and I. palifera of Isopora from Acropora, and ORF successfully identified the species of Pocillopora (except P. meandrina and P. eydouxi). Moreover, other cnidarian species such as actinarians, zoanthids, and Millepora species were visually found using fluorescence and identified by COI DNA barcoding. This combination of existing approaches greatly improved the taxonomic resolution of early coral life stages, which to date has been mainly limited to the family level based on skeletal identification. Overall, this study suggests important improvements for the identification of coral recruits in ecological studies. PMID:25211345

Identification of scleractinian coral recruits using fluorescent censusing and DNA barcoding techniques.

PubMed

Hsu, Chia-Min; de Palmas, Stéphane; Kuo, Chao-Yang; Denis, Vianney; Chen, Chaolun Allen

2014-01-01

The identification of coral recruits has been problematic due to a lack of definitive morphological characters being available for higher taxonomic resolution. In this study, we tested whether fluorescent detection of coral recruits used in combinations of different DNA-barcoding markers (cytochrome oxidase I gene [COI], open reading frame [ORF], and nuclear Pax-C intron [PaxC]) could be useful for increasing the resolution of coral spat identification in ecological studies. One hundred and fifty settlement plates were emplaced at nine sites on the fringing reefs of Kenting National Park in southern Taiwan between April 2011 and September 2012. A total of 248 living coral spats and juveniles (with basal areas ranging from 0.21 to 134.57 mm(2)) were detected on the plates with the aid of fluorescent light and collected for molecular analyses. Using the COI DNA barcoding technique, 90.3% (224/248) of coral spats were successfully identified into six genera, including Acropora, Isopora, Montipora, Pocillopora, Porites, and Pavona. PaxC further separated I. cuneata and I. palifera of Isopora from Acropora, and ORF successfully identified the species of Pocillopora (except P. meandrina and P. eydouxi). Moreover, other cnidarian species such as actinarians, zoanthids, and Millepora species were visually found using fluorescence and identified by COI DNA barcoding. This combination of existing approaches greatly improved the taxonomic resolution of early coral life stages, which to date has been mainly limited to the family level based on skeletal identification. Overall, this study suggests important improvements for the identification of coral recruits in ecological studies.

Analyzing Mosquito (Diptera: Culicidae) Diversity in Pakistan by DNA Barcoding

PubMed Central

Ashfaq, Muhammad; Hebert, Paul D. N.; Mirza, Jawwad H.; Khan, Arif M.; Zafar, Yusuf; Mirza, M. Sajjad

2014-01-01

Background Although they are important disease vectors mosquito biodiversity in Pakistan is poorly known. Recent epidemics of dengue fever have revealed the need for more detailed understanding of the diversity and distributions of mosquito species in this region. DNA barcoding improves the accuracy of mosquito inventories because morphological differences between many species are subtle, leading to misidentifications. Methodology/Principal Findings Sequence variation in the barcode region of the mitochondrial COI gene was used to identify mosquito species, reveal genetic diversity, and map the distribution of the dengue-vector species in Pakistan. Analysis of 1684 mosquitoes from 491 sites in Punjab and Khyber Pakhtunkhwa during 2010–2013 revealed 32 species with the assemblage dominated by Culex quinquefasciatus (61% of the collection). The genus Aedes (Stegomyia) comprised 15% of the specimens, and was represented by six taxa with the two dengue vector species, Ae. albopictus and Ae. aegypti, dominant and broadly distributed. Anopheles made up another 6% of the catch with An. subpictus dominating. Barcode sequence divergence in conspecific specimens ranged from 0–2.4%, while congeneric species showed from 2.3–17.8% divergence. A global haplotype analysis of disease-vectors showed the presence of multiple haplotypes, although a single haplotype of each dengue-vector species was dominant in most countries. Geographic distribution of Ae. aegypti and Ae. albopictus showed the later species was dominant and found in both rural and urban environments. Conclusions As the first DNA-based analysis of mosquitoes in Pakistan, this study has begun the construction of a barcode reference library for the mosquitoes of this region. Levels of genetic diversity varied among species. Because of its capacity to differentiate species, even those with subtle morphological differences, DNA barcoding aids accurate tracking of vector populations. PMID:24827460

Analyzing mosquito (Diptera: culicidae) diversity in Pakistan by DNA barcoding.

PubMed

Ashfaq, Muhammad; Hebert, Paul D N; Mirza, Jawwad H; Khan, Arif M; Zafar, Yusuf; Mirza, M Sajjad

2014-01-01

Although they are important disease vectors mosquito biodiversity in Pakistan is poorly known. Recent epidemics of dengue fever have revealed the need for more detailed understanding of the diversity and distributions of mosquito species in this region. DNA barcoding improves the accuracy of mosquito inventories because morphological differences between many species are subtle, leading to misidentifications. Sequence variation in the barcode region of the mitochondrial COI gene was used to identify mosquito species, reveal genetic diversity, and map the distribution of the dengue-vector species in Pakistan. Analysis of 1684 mosquitoes from 491 sites in Punjab and Khyber Pakhtunkhwa during 2010-2013 revealed 32 species with the assemblage dominated by Culex quinquefasciatus (61% of the collection). The genus Aedes (Stegomyia) comprised 15% of the specimens, and was represented by six taxa with the two dengue vector species, Ae. albopictus and Ae. aegypti, dominant and broadly distributed. Anopheles made up another 6% of the catch with An. subpictus dominating. Barcode sequence divergence in conspecific specimens ranged from 0-2.4%, while congeneric species showed from 2.3-17.8% divergence. A global haplotype analysis of disease-vectors showed the presence of multiple haplotypes, although a single haplotype of each dengue-vector species was dominant in most countries. Geographic distribution of Ae. aegypti and Ae. albopictus showed the later species was dominant and found in both rural and urban environments. As the first DNA-based analysis of mosquitoes in Pakistan, this study has begun the construction of a barcode reference library for the mosquitoes of this region. Levels of genetic diversity varied among species. Because of its capacity to differentiate species, even those with subtle morphological differences, DNA barcoding aids accurate tracking of vector populations.

Morphological identification and COI barcodes of adult flies help determine species identities of chironomid larvae (Diptera, Chironomidae).

PubMed

Failla, A J; Vasquez, A A; Hudson, P; Fujimoto, M; Ram, J L

2016-02-01

Establishing reliable methods for the identification of benthic chironomid communities is important due to their significant contribution to biomass, ecology and the aquatic food web. Immature larval specimens are more difficult to identify to species level by traditional morphological methods than their fully developed adult counterparts, and few keys are available to identify the larval species. In order to develop molecular criteria to identify species of chironomid larvae, larval and adult chironomids from Western Lake Erie were subjected to both molecular and morphological taxonomic analysis. Mitochondrial cytochrome c oxidase I (COI) barcode sequences of 33 adults that were identified to species level by morphological methods were grouped with COI sequences of 189 larvae in a neighbor-joining taxon-ID tree. Most of these larvae could be identified only to genus level by morphological taxonomy (only 22 of the 189 sequenced larvae could be identified to species level). The taxon-ID tree of larval sequences had 45 operational taxonomic units (OTUs, defined as clusters with >97% identity or individual sequences differing from nearest neighbors by >3%; supported by analysis of all larval pairwise differences), of which seven could be identified to species or 'species group' level by larval morphology. Reference sequences from the GenBank and BOLD databases assigned six larval OTUs with presumptive species level identifications and confirmed one previously assigned species level identification. Sequences from morphologically identified adults in the present study grouped with and further classified the identity of 13 larval OTUs. The use of morphological identification and subsequent DNA barcoding of adult chironomids proved to be beneficial in revealing possible species level identifications of larval specimens. Sequence data from this study also contribute to currently inadequate public databases relevant to the Great Lakes region, while the neighbor-joining analysis reported here describes the application and confirmation of a useful tool that can accelerate identification and bioassessment of chironomid communities.

Morphological identification and COI barcodes of adult flies help determine species identities of chironomid larvae (Diptera, Chironomidae)

USGS Publications Warehouse

Failla, Andrew Joseph; Vasquez, Adrian Amelio; Hudson, Patrick L.; Fujimoto, Masanori; Ram, Jeffrey L.

2016-01-01

Establishing reliable methods for the identification of benthic chironomid communities is important due to their significant contribution to biomass, ecology and the aquatic food web. Immature larval specimens are more difficult to identify to species level by traditional morphological methods than their fully developed adult counterparts, and few keys are available to identify the larval species. In order to develop molecular criteria to identify species of chironomid larvae, larval and adult chironomids from Western Lake Erie were subjected to both molecular and morphological taxonomic analysis. Mitochondrial cytochrome c oxidase I (COI) barcode sequences of 33 adults that were identified to species level by morphological methods were grouped with COI sequences of 189 larvae in a neighbor-joining taxon-ID tree. Most of these larvae could be identified only to genus level by morphological taxonomy (only 22 of the 189 sequenced larvae could be identified to species level). The taxon-ID tree of larval sequences had 45 operational taxonomic units (OTUs, defined as clusters with >97% identity or individual sequences differing from nearest neighbors by >3%; supported by analysis of all larval pairwise differences), of which seven could be identified to species or ‘species group’ level by larval morphology. Reference sequences from the GenBank and BOLD databases assigned six larval OTUs with presumptive species level identifications and confirmed one previously assigned species level identification. Sequences from morphologically identified adults in the present study grouped with and further classified the identity of 13 larval OTUs. The use of morphological identification and subsequent DNA barcoding of adult chironomids proved to be beneficial in revealing possible species level identifications of larval specimens. Sequence data from this study also contribute to currently inadequate public databases relevant to the Great Lakes region, while the neighbor-joining analysis reported here describes the application and confirmation of a useful tool that can accelerate identification and bioassesment of chironomid communities.

Using DNA barcodes to identify a bird involved in a birdstrike at a Chinese airport.

PubMed

Yang, Rong; Wu, Xiaobing; Yan, Peng; Li, Xiaoqiang

2010-10-01

One day at dusk in August, 200X, an airplane was struck by a bird at a Chinese airport (M Airport). After a careful check, some blades of the plane's engine were found to be out of shape and a few feathers and some bloodstains were found in the air intake of the engine. In order to know which species of bird was involved in the birdstrike, firstly we extracted DNA from the bloodstains; secondly, the DNA barcode (portion of COI gene) of the unknown species was amplified by PCR method; thirdly, sequence divergences (K2P differences) of the DNA barcode between the unknown species and a library of 59 common bird species distributed at the airport area were analyzed. Furthermore, a neighbor-joining (NJ) tree based on COI barcodes was created to provide graphic representation of sequence divergences among the species to confirm the identification. The result showed that red-rumped swallow (Hirundo daurica) was involved in the birdstrike incident. Some suggestions to avoid birdstrikes caused by red-rumped swallows were given to the administrative department of M Airport to ensure flying safety.

DNA barcoding for identification of sand fly species (Diptera: Psychodidae) from leishmaniasis-endemic areas of Peru.

PubMed

Nzelu, Chukwunonso O; Cáceres, Abraham G; Arrunátegui-Jiménez, Martín J; Lañas-Rosas, Máximo F; Yañez-Trujillano, Henrry H; Luna-Caipo, Deysi V; Holguín-Mauricci, Carlos E; Katakura, Ken; Hashiguchi, Yoshihisa; Kato, Hirotomo

2015-05-01

Phlebotomine sand flies are the only proven vectors of leishmaniases, a group of human and animal diseases. Accurate knowledge of sand fly species identification is essential in understanding the epidemiology of leishmaniasis and vector control in endemic areas. Classical identification of sand fly species based on morphological characteristics often remains difficult and requires taxonomic expertise. Here, we generated DNA barcodes of the cytochrome c oxidase subunit 1 (COI) gene using 159 adult specimens morphologically identified to be 19 species of sand flies, belonging to 6 subgenera/species groups circulating in Peru, including the vector species. Neighbor-joining (NJ) analysis based on Kimura 2-Parameter genetic distances formed non-overlapping clusters for all species. The levels of intraspecific genetic divergence ranged from 0 to 5.96%, whereas interspecific genetic divergence among different species ranged from 8.39 to 19.08%. The generated COI barcodes could discriminate between all the sand fly taxa. Besides its success in separating known species, we found that DNA barcoding is useful in revealing population differentiation and cryptic diversity, and thus promises to be a valuable tool for epidemiological studies of leishmaniasis. Copyright © 2015 Elsevier B.V. All rights reserved.

Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences☆

PubMed Central

Kowasupat, Chanon; Panijpan, Bhinyo; Laosinchai, Parames; Ruenwongsa, Pintip; Phongdara, Amornrat; Wanna, Warapond; Senapin, Saengchan; Phiwsaiya, Kornsunee

2014-01-01

In Thailand, there are currently five recognized species members of the bubble-nesting Betta genus, namely Betta splendens, B. smaragdina, B. imbellis, B. mahachaiensis and B. siamorientalis. In 2010, we indicated the possibility, based on COI barcoding evidence, that there might be two additional species, albeit cryptic, related to the type-locality B. smaragdina in some provinces in the northeast of Thailand. In the present study, after a more extensive survey of the northeast, and phylogenetic analyses based on COI and ITS1 sequences, the B. smaragdina group may be composed of at least 3 cryptic species members. The phylogenetic positions of these B. smaragdina group members in the bubble-nesting bettas' tree together with those of their congeners have been consolidated by better DNA sequence quality and phylogenetic analyses. With a better supported tree, the species statuses of B. siamorientalis and the Cambodian B. smaragdina-like fish, B. stiktos, are also confirmed. PMID:25606392

Cytochrome C oxidase subunit I barcodes provide an efficient tool for Jinqian Baihua She (Bungarus parvus) authentication

PubMed Central

Chao, Zhi; Liao, Jing; Liang, Zhenbiao; Huang, Suhua; Zhang, Liang; Li, Junde

2014-01-01

Objective: To test the feasibility of DNA barcoding for accurate identification of Jinqian Baihua She and its adulterants. Materials and Methods: Standard cytochrome C oxidase subunit I (COI) gene fragments were sequenced for DNA barcoding of 39 samples from 9 snake species, including Bungarus multicinctus, the officially recognized origin animal by Chinese Pharmacopoeia, and other 8 adulterate species. The aligned sequences, 658 base pairs in length, were analyzed for divergence using the Kimura-2-parameter (K2P) distance model with MEGA5.0. Results: The mean intraspecific K2P distance was 0.0103 and the average interspecific genetic distance was 0.2178 in B. multicinctus, far greater than the minimal interspecific genetic distance of 0.027 recommended for species identification. A neighbor-joining (NJ) tree was constructed, in which each species formed a monophyletic clade with bootstrap supports of 100%. All the data were submitted to Barcode of Life Data system version 3.0 (BOLD, http://www.barcodinglife.org) under the project title “DNA barcoding Bungarus multicinctus and its adulterants”. Ten samples of commercially available crude drugs of JBS were identified using the identification engine provided by BOLD. All the samples were clearly identified at the species level, among which five were found to be the adulterants and identified as Dinodon rufozonatum. Conclusion: DNA barcoding using the standard COI gene fragments provides an effective and accurate means for JBS identification and authentication. PMID:25422545

Pitfalls of Establishing DNA Barcoding Systems in Protists: The Cryptophyceae as a Test Case

PubMed Central

Hoef-Emden, Kerstin

2012-01-01

A DNA barcode is a preferrably short and highly variable region of DNA supposed to facilitate a rapid identification of species. In many protistan lineages, a lack of species-specific morphological characters hampers an identification of species by light or electron microscopy, and difficulties to perform mating experiments in laboratory cultures also do not allow for an identification of biological species. Thus, testing candidate barcode markers as well as establishment of accurately working species identification systems are more challenging than in multicellular organisms. In cryptic species complexes the performance of a potential barcode marker can not be monitored using morphological characters as a feedback, but an inappropriate choice of DNA region may result in artifactual species trees for several reasons. Therefore a priori knowledge of the systematics of a group is required. In addition to identification of known species, methods for an automatic delimitation of species with DNA barcodes have been proposed. The Cryptophyceae provide a mixture of systematically well characterized as well as badly characterized groups and are used in this study to test the suitability of some of the methods for protists. As species identification method the performance of blast in searches against badly to well-sampled reference databases has been tested with COI-5P and 5′-partial LSU rDNA (domains A to D of the nuclear LSU rRNA gene). In addition the performance of two different methods for automatic species delimitation, fixed thresholds of genetic divergence and the general mixed Yule-coalescent model (GMYC), have been examined. The study demonstrates some pitfalls of barcoding methods that have to be taken care of. Also a best-practice approach towards establishing a DNA barcode system in protists is proposed. PMID:22970104

Pitfalls of establishing DNA barcoding systems in protists: the cryptophyceae as a test case.

PubMed

Hoef-Emden, Kerstin

2012-01-01

A DNA barcode is a preferrably short and highly variable region of DNA supposed to facilitate a rapid identification of species. In many protistan lineages, a lack of species-specific morphological characters hampers an identification of species by light or electron microscopy, and difficulties to perform mating experiments in laboratory cultures also do not allow for an identification of biological species. Thus, testing candidate barcode markers as well as establishment of accurately working species identification systems are more challenging than in multicellular organisms. In cryptic species complexes the performance of a potential barcode marker can not be monitored using morphological characters as a feedback, but an inappropriate choice of DNA region may result in artifactual species trees for several reasons. Therefore a priori knowledge of the systematics of a group is required. In addition to identification of known species, methods for an automatic delimitation of species with DNA barcodes have been proposed. The Cryptophyceae provide a mixture of systematically well characterized as well as badly characterized groups and are used in this study to test the suitability of some of the methods for protists. As species identification method the performance of blast in searches against badly to well-sampled reference databases has been tested with COI-5P and 5'-partial LSU rDNA (domains A to D of the nuclear LSU rRNA gene). In addition the performance of two different methods for automatic species delimitation, fixed thresholds of genetic divergence and the general mixed Yule-coalescent model (GMYC), have been examined. The study demonstrates some pitfalls of barcoding methods that have to be taken care of. Also a best-practice approach towards establishing a DNA barcode system in protists is proposed.

Integrative taxonomy supports new candidate fish species in a poorly studied neotropical region: the Jequitinhonha River Basin.

PubMed

Pugedo, Marina Lages; de Andrade Neto, Francisco Ricardo; Pessali, Tiago Casarim; Birindelli, José Luís Olivan; Carvalho, Daniel Cardoso

2016-06-01

Molecular identification through DNA barcoding has been proposed as a way to standardize a global biodiversity identification system using a partial sequence of the mitochondrial COI gene. We applied an integrative approach using DNA barcoding and traditional morphology-based bioassessment to identify fish from a neotropical region possessing low taxonomic knowledge: the Jequitinhonha River Basin (Southeastern Brazil). The Jequitinhonha River Basin (JRB) has a high rate of endemism and is considered an area of high priority for fish conservation, with estimates indicating the presence of around 110 native and non-indigenous species. DNA barcodes were obtained from 260 individuals belonging to 52 species distributed among 35 genera, 21 families and 6 orders, including threatened and rare species such as Rhamdia jequitinhonha and Steindachneridion amblyurum. The mean Kimura two-parameter genetic distances within species, genera and families were: 0.44, 12.16 and 20.58 %, respectively. Mean intraspecific genetic variation ranged from 0 to 11.43 %, and high values (>2 %) were recovered for five species. Species with a deep intraspecific distance, possibly flagging overlooked taxa, were detected within the genus Pimelodella. Fifteen species, only identified to the genus level, had unique BINs, with a nearest neighbor distance over 2 % and therefore, potential new candidate species supported by DNA barcoding. The integrative taxonomy approach using DNA barcoding and traditional taxonomy may be a remedy to taxonomy impediment, accelerating species identification by flagging potential new candidate species and to adequately conserve the megadiverse neotropical ichthyofauna.

Phylogenetic Reconstruction and DNA Barcoding for Closely Related Pine Moth Species (Dendrolimus) in China with Multiple Gene Markers

PubMed Central

Dai, Qing-Yan; Gao, Qiang; Wu, Chun-Sheng; Chesters, Douglas; Zhu, Chao-Dong; Zhang, Ai-Bing

2012-01-01

Unlike distinct species, closely related species offer a great challenge for phylogeny reconstruction and species identification with DNA barcoding due to their often overlapping genetic variation. We tested a sibling species group of pine moth pests in China with a standard cytochrome c oxidase subunit I (COI) gene and two alternative internal transcribed spacer (ITS) genes (ITS1 and ITS2). Five different phylogenetic/DNA barcoding analysis methods (Maximum likelihood (ML)/Neighbor-joining (NJ), “best close match” (BCM), Minimum distance (MD), and BP-based method (BP)), representing commonly used methodology (tree-based and non-tree based) in the field, were applied to both single-gene and multiple-gene analyses. Our results demonstrated clear reciprocal species monophyly for three relatively distant related species, Dendrolimus superans, D. houi, D. kikuchii, as recovered by both single and multiple genes while the phylogenetic relationship of three closely related species, D. punctatus, D. tabulaeformis, D. spectabilis, could not be resolved with the traditional tree-building methods. Additionally, we find the standard COI barcode outperforms two nuclear ITS genes, whatever the methods used. On average, the COI barcode achieved a success rate of 94.10–97.40%, while ITS1 and ITS2 obtained a success rate of 64.70–81.60%, indicating ITS genes are less suitable for species identification in this case. We propose the use of an overall success rate of species identification that takes both sequencing success and assignation success into account, since species identification success rates with multiple-gene barcoding system were generally overestimated, especially by tree-based methods, where only successfully sequenced DNA sequences were used to construct a phylogenetic tree. Non-tree based methods, such as MD, BCM, and BP approaches, presented advantages over tree-based methods by reporting the overall success rates with statistical significance. In addition, our results indicate that the most closely related species D. punctatus, D. tabulaeformis, and D. spectabilis, may be still in the process of incomplete lineage sorting, with occasional hybridizations occurring among them. PMID:22509245

DNA Barcoding of Metazoan Zooplankton Copepods from South Korea

PubMed Central

Ryu, Shi Hyun; Kim, Sang Ki; Lee, Jin Hee; Lim, Young Jin; Lee, Jimin; Jun, Jumin; Kwak, Myounghai; Lee, Young-Sup; Hwang, Jae-Sam; Venmathi Maran, Balu Alagar; Chang, Cheon Young; Kim, Il-Hoi; Hwang, Ui Wook

2016-01-01

Copepods, small aquatic crustaceans, are the most abundant metazoan zooplankton and outnumber every other group of multicellular animals on earth. In spite of ecological and biological importance in aquatic environment, their morphological plasticity, originated from their various lifestyles and their incomparable capacity to adapt to a variety of environments, has made the identification of species challenging, even for expert taxonomists. Molecular approaches to species identification have allowed rapid detection, discrimination, and identification of cryptic or sibling species based on DNA sequence data. We examined sequence variation of a partial mitochondrial cytochrome C oxidase I gene (COI) from 133 copepod individuals collected from the Korean Peninsula, in order to identify and discriminate 94 copepod species covering six copepod orders of Calanoida, Cyclopoida, Harpacticoida, Monstrilloida, Poecilostomatoida and Siphonostomatoida. The results showed that there exists a clear gap with ca. 20 fold difference between the averages of within-specific sequence divergence (2.42%) and that of between-specific sequence divergence (42.79%) in COI, suggesting the plausible utility of this gene in delimitating copepod species. The results showed, with the COI barcoding data among 94 copepod species, that a copepod species could be distinguished from the others very clearly, only with four exceptions as followings: Mesocyclops dissimilis–Mesocyclops pehpeiensis (0.26% K2P distance in percent) and Oithona davisae–Oithona similis (1.1%) in Cyclopoida, Ostrincola japonica–Pseudomyicola spinosus (1.5%) in Poecilostomatoida, and Hatschekia japonica–Caligus quadratus (5.2%) in Siphonostomatoida. Thus, it strongly indicated that COI may be a useful tool in identifying various copepod species and make an initial progress toward the construction of a comprehensive DNA barcode database for copepods inhabiting the Korean Peninsula. PMID:27383475

DNA barcoding and molecular systematics of the benthic and demersal organisms of the CEAMARC survey

NASA Astrophysics Data System (ADS)

Dettai, Agnes; Adamowizc, Sarah J.; Allcock, Louise; Arango, Claudia P.; Barnes, David K. A.; Barratt, Iain; Chenuil, Anne; Couloux, Arnaud; Cruaud, Corinne; David, Bruno; Denis, Françoise; Denys, Gael; Díaz, Angie; Eléaume, Marc; Féral, Jean-Pierre; Froger, Aurélie; Gallut, Cyril; Grant, Rachel; Griffiths, Huw J.; Held, Christoph; Hemery, Lenaïg G.; Hosie, Graham; Kuklinski, Piotr; Lecointre, Guillaume; Linse, Katrin; Lozouet, Pierre; Mah, Christopher; Monniot, Françoise; Norman, Mark D.; O'Hara, Timothy; Ozouf-Costaz, Catherine; Piedallu, Claire; Pierrat, Benjamin; Poulin, Elie; Puillandre, Nicolas; Riddle, Martin; Samadi, Sarah; Saucède, Thomas; Schubart, Christoph; Smith, Peter J.; Stevens, Darren W.; Steinke, Dirk; Strugnell, Jan M.; Tarnowska, K.; Wadley, Victoria; Ameziane, Nadia

2011-08-01

The Dumont d’Urville Sea (East Antarctic region) has been less investigated for DNA barcoding and molecular taxonomy than other parts of the Southern Ocean, such as the Ross Sea and the Antarctic Peninsula. The Collaborative East Antarctic MARine Census (CEAMARC) took place in this area during the austral summer of 2007-2008. The Australian vessel RSV Aurora Australis collected very diverse samples of demersal and benthic organisms. The specimens were sorted centrally, and then distributed to taxonomic experts for molecular and morphological taxonomy and identification, especially barcoding. The COI sequences generated from CEAMARC material provide a sizeable proportion of the Census of Antarctic Marine Life barcodes although the studies are still ongoing, and represent the only source of sequences for a number of species. Barcoding appears to be a valuable method for identification within most groups, despite low divergences and haplotype sharing in a few species, and it is also useful as a preliminary taxonomic exploration method. Several new species are being described. CEAMARC samples have already provided new material for phylogeographic and phylogenetic studies in cephalopods, pycnogonids, teleost fish, crinoids and sea urchins, helping these studies to provide a better insight in the patterns of evolution in the Southern Ocean.

Cytochrome c oxidase subunit 1 barcode data of fish of the Nayband National Park in the Persian Gulf and analysis using meta-data flag several cryptic species.

PubMed

Asgharian, Hosseinali; Sahafi, Homayoun Hosseinzadeh; Ardalan, Aria Ashja; Shekarriz, Shahrokh; Elahi, Elahe

2011-05-01

We provide cytochrome c oxidase subunit 1 (COI) barcode sequences of fishes of the Nayband National Park, Persian Gulf, Iran. Industrial activities, ecological considerations and goals of The Fish Barcode of Life campaign make it crucial that fish species residing in the park be identified. To the best of our knowledge, this is the first report of barcoding data on fishes of the Persian Gulf. We examined 187 individuals representing 76 species, 56 genera and 32 families. The data flagged potentially cryptic species of Gerres filamentosus and Plectorhinchus schotaf. 16S rDNA data on these species are provided. Exclusion of these two potential cryptic species resulted in a mean COI intraspecific distance of 0.18%, and a mean inter- to intraspecific divergence ratio of 66.7. There was no overlap between maximum Kimura 2-parameter distances among conspecifics (1.66%) and minimum distance among congeneric species (6.19%). Barcodes shared among species were not observed. Neighbour-joining analysis showed that most species formed cohesive sequence units with little variation. Finally, the comparison of 16 selected species from this study with meta-data of conspecifics from Australia, India, China and South Africa revealed high interregion divergences and potential existence of six cryptic species. Pairwise interregional comparisons were more informative than global divergence assessments with regard to detection of cryptic variation. Our analysis exemplifies optimal use of the expanding barcode data now becoming available. © 2011 Blackwell Publishing Ltd.

Conflicting patterns of DNA barcoding and taxonomy in the cicada genus Tettigettalna from Southern Europe (Hemiptera: Cicadidae).

PubMed

Nunes, Vera L; Mendes, Raquel; Marabuto, Eduardo; Novais, Bruno M; Hertach, Thomas; Quartau, José A; Seabra, Sofia G; Paulo, Octávio S; Simões, Paula C

2014-01-01

DNA barcodes have great potential to assist in species identification, especially when high taxonomical expertise is required. We investigated the utility of the 5' mitochondrial cytochrome c oxidase I (COI) region to discriminate between 13 European cicada species. These included all nine species currently recognized under the genus Tettigettalna, from which seven are endemic to the southern Iberian Peninsula. These cicadas have species-specific male calling songs but are morphologically very similar. Mean COI divergence between congeners ranged from 0.4% to 10.6%, but this gene was proven insufficient to determine species limits within genus Tettigettalna because a barcoding gap was absent for several of its species, that is, the highest intraspecific distance exceeded the lowest interspecific distance. The genetic data conflicted with current taxonomic classification for T. argentata and T. mariae. Neighbour-joining and Bayesian analyses revealed that T. argentata is geographically structured (clades North and South) and might constitute a species complex together with T. aneabi and T. mariae. The latter diverges very little from the southern clade of T. argentata and shares with it its most common haplotype. T. mariae is often in sympatry with T. argentata but it remains unclear whether introgression or incomplete lineage sorting may be responsible for the sharing of haplotypes. T. helianthemi and T. defauti also show high intraspecific variation that might signal hidden cryptic diversity. These taxonomic conflicts must be re-evaluated with further studies using additional genes and extensive morphological and acoustic analyses. © 2013 John Wiley & Sons Ltd.

First Molecular Identification and Phylogeny of Moroccan Anopheles sergentii (Diptera: Culicidae) Based on Second Internal Transcribed Spencer (ITS2) and Cytochrome c Oxidase I (COI) Sequences.

PubMed

Benabdelkrim Filali, Oumama; Kabine, Mostafa; El Hamouchi, Adil; Lemrani, Meryem; Debboun, Mustapha; Sarih, M'hammed

2018-06-05

Anopheles sergentii known as the "oasis vector" or the "desert malaria vector" is considered the main vector of malaria in the southern parts of Morocco. Its presence in Morocco is confirmed for the first time through sequencing of mitochondrial DNA (mDNA) cytochrome c oxidase subunit I (COI) barcodes and nuclear ribosomal DNA (rDNA) second internal transcribed spacer (ITS2) sequences and direct comparison with specimens of A. sergentii of other countries. The DNA barcodes (n = 39) obtained from A. sergentii collected in 2015 and 2016 showed more diversity with 10 haplotypes, compared with 3 haplotypes obtained from ITS2 sequences (n = 59). Moreover, the comparison using the ITS2 sequences showed closer evolutionary relationship between the Moroccan and Egyptian strains than the Iranian strain. Nevertheless, genetic differences due to geographical segregation were also observed. This study provides the first report on the sequence of rDNA-ITS2 and mtDNA COI, which could be used to better understand the biodiversity of A. sergentii.

"Crown of thorns" of Daphnia: an exceptional inducible defense discovered by DNA barcoding.

PubMed

Laforsch, Christian; Haas, Andreas; Jung, Nina; Schwenk, Klaus; Tollrian, Ralph; Petrusek, Adam

2009-09-01

DNA barcoding has emerged as valuable tool to document global biodiversity. Mitochondrial cytochrome oxidase I (COI) sequences serve as genetic markers to catalogue species richness in the animal kingdom and to identify cryptic and polymorphic animal species. Furthermore, DNA barcoding data serve as a fuel for ecological studies, as they provide the opportunity to unravel species interactions among hosts and parasites, predators and prey, and among competitors in unprecedented detail. In a recent paper we described how DNA barcoding in combination with morphological and ecological data unravelled a striking predator-prey interaction of organisms from temporary aquatic habitats, the predatory notostracan Triops and its prey, cladocerans of the Daphnia atkinsoni complex.

Using DNA barcoding to differentiate invasive Dreissena species (Mollusca, Bivalvia)

PubMed Central

Marescaux, Jonathan; Van Doninck, Karine

2013-01-01

Abstract The zebra mussel (Dreissena polymorpha) and the quagga mussel (Dreissena rostriformis bugensis) are considered as the most competitive invaders in freshwaters of Europe and North America. Although shell characteristics exist to differentiate both species, phenotypic plasticity in the genus Dreissena does not always allow a clear identification. Therefore, the need to find an accurate identification method is essential. DNA barcoding has been proven to be an adequate procedure to discriminate species. The cytochrome c oxidase subunit I mitochondrial gene (COI) is considered as the standard barcode for animals. We tested the use of this gene as an efficient DNA barcode and found that it allow rapid and accurate identification of adult Dreissena individuals. PMID:24453560

Two new and one newly recorded species of Polypedilum Kieffer 1912 with DNA barcodes from Oriental China (Chironomidae: Diptera).

PubMed

Yan, Chuncai; Song, Chao; Liu, Ting; Zhao, Guangjun; Hou, Ziyuan; Cao, Wei; Wang, Xinhua

2017-03-02

Polypedilum (Tripodura) enshiense Song & Wang sp. n. and P. (Tripodura) jianfengense Song & Wang sp. n. are described and illustrated as male imagines from China. P. (Uresipedilum) paraconvictum Yamamoto, Yamamoto & Hirowatari, 2016 is recorded from China for the first time. Cytochrome coxidase subunit I (COI) sequence of above species are uploaded to Barcode of Life Data Systems (BOLD).

DNA Barcoding of genus Hexacentrus in China reveals cryptic diversity within Hexacentrus japonicus (Orthoptera, Tettigoniidae).

PubMed

Guo, Hui-Fang; Guan, Bei; Shi, Fu-Ming; Zhou, Zhi-Jun

2016-01-01

DNA barcoding has been proved successful to provide resolution beyond the boundaries of morphological information. Hence, a study was undertaken to establish DNA barcodes for all morphologically determined Hexacentrus species in China collections. In total, 83 specimens of five Hexacentrus species were barcoded using standard mitochondrial cytochrome c oxidase subunit I (COI) gene. Except for Hexacentrus japonicus, barcode gaps were present in the remaining Hexacentrus species. Taxon ID tree generated seven BOLD's barcode index numbers (BINs), four of which were in agreement with the morphological species. For Hexacentrus japonicus, the maximum intraspecific divergence (4.43%) produced a minimal overlap (0.64%), and 19 specimens were divided into three different BINs. There may be cryptic species within the current Hexacentrus japonicus. This study adds to a growing body of DNA barcodes that have become available for katydids, and shows that a DNA barcoding approach enables the identification of known Hexacentrus species with a very high resolution.

DNA Barcoding of genus Hexacentrus in China reveals cryptic diversity within Hexacentrus japonicus (Orthoptera, Tettigoniidae)

PubMed Central

Guo, Hui-Fang; Guan, Bei; Shi, Fu-Ming; Zhou, Zhi-Jun

2016-01-01

Abstract DNA barcoding has been proved successful to provide resolution beyond the boundaries of morphological information. Hence, a study was undertaken to establish DNA barcodes for all morphologically determined Hexacentrus species in China collections. In total, 83 specimens of five Hexacentrus species were barcoded using standard mitochondrial cytochrome c oxidase subunit I (COI) gene. Except for Hexacentrus japonicus, barcode gaps were present in the remaining Hexacentrus species. Taxon ID tree generated seven BOLD’s barcode index numbers (BINs), four of which were in agreement with the morphological species. For Hexacentrus japonicus, the maximum intraspecific divergence (4.43%) produced a minimal overlap (0.64%), and 19 specimens were divided into three different BINs. There may be cryptic species within the current Hexacentrus japonicus. This study adds to a growing body of DNA barcodes that have become available for katydids, and shows that a DNA barcoding approach enables the identification of known Hexacentrus species with a very high resolution. PMID:27408576

Design of character-based DNA barcode motif for species identification: A computational approach and its validation in fishes.

PubMed

Chakraborty, Mohua; Dhar, Bishal; Ghosh, Sankar Kumar

2017-11-01

The DNA barcodes are generally interpreted using distance-based and character-based methods. The former uses clustering of comparable groups, based on the relative genetic distance, while the latter is based on the presence or absence of discrete nucleotide substitutions. The distance-based approach has a limitation in defining a universal species boundary across the taxa as the rate of mtDNA evolution is not constant throughout the taxa. However, character-based approach more accurately defines this using a unique set of nucleotide characters. The character-based analysis of full-length barcode has some inherent limitations, like sequencing of the full-length barcode, use of a sparse-data matrix and lack of a uniform diagnostic position for each group. A short continuous stretch of a fragment can be used to resolve the limitations. Here, we observe that a 154-bp fragment, from the transversion-rich domain of 1367 COI barcode sequences can successfully delimit species in the three most diverse orders of freshwater fishes. This fragment is used to design species-specific barcode motifs for 109 species by the character-based method, which successfully identifies the correct species using a pattern-matching program. The motifs also correctly identify geographically isolated population of the Cypriniformes species. Further, this region is validated as a species-specific mini-barcode for freshwater fishes by successful PCR amplification and sequencing of the motif (154 bp) using the designed primers. We anticipate that use of such motifs will enhance the diagnostic power of DNA barcode, and the mini-barcode approach will greatly benefit the field-based system of rapid species identification. © 2017 John Wiley & Sons Ltd.

DNA analysis of molluscs from a museum wet collection: a comparison of different extraction methods.

PubMed

Jaksch, Katharina; Eschner, Anita; Rintelen, Thomas V; Haring, Elisabeth

2016-07-18

DNA isolation and PCR amplification from molluscan taxa is considered as problematic because polysaccharides in tissue and mucus presumably co-precipitate with the DNA and inhibit the activity of DNA polymerase. In the present study we tested two common extraction methods on specimens from the mollusc collection of the Natural History Museum Vienna (NHMW). We analysed a broad variety of taxa covering a large temporal span (acquisition years 1877 to 1999), which distinguishes our study from previous ones where mostly fresh material was used. We also took other factors into account: effects of sample age, effects of formaldehyde treatment and taxon-specific problems. We used several primer combinations to amplify amplicons of different lengths of two mitochondrial genes: cytochrome c oxidase subunit 1 (COI) and 16S rRNA gene (16S). Overall PCR success was 43 % in the 576 extractions (including all primer combinations). The smallest amplicon (~240 bp) showed the best results (49 % positive reactions), followed by the 400 bp amplicon (40.5 %). Both short sections yielded significantly better results than the 700 bp long amplicon (27 %). Comparatively, the Gen-ial-First, All-tissue DNA-Kit-extraction method performed significantly better than Promega-Tissue and Hair Extraction Kit. Generally, PCR success is age-dependent. Nonetheless, we were able to obtain the longest amplicon even from 137-year-old material. Importantly, formaldehyde traces did not totally inhibit amplification success, although very high concentrations did. Museum material has gained importance for DNA analysis in recent years, especially for DNA barcoding projects. In some cases, however, the amplification of the standard barcoding region (partial sequence of the COI) is problematic with old material. Our study clearly shows that the COI barcoding region could be amplified in up to 49 % of PCRs (varying with amplicon length), which is, for museum samples, quite a high percentage. The difference between extraction methods was minimal and we recommend using an established kit for a first attempt because experience and routine in handling might be more important than slight performance differences of the various kits. Finally, we identify fixation, storage, sample conservation and documentation of the specimens' history rather than the DNA extraction method to be the most crucial factors for PCR success.

Phylogeny and species delineation in European species of the genus Steganacarus (Acari, Oribatida) using mitochondrial and nuclear markers.

PubMed

Kreipe, Victoria; Corral-Hernández, Elena; Scheu, Stefan; Schaefer, Ina; Maraun, Mark

2015-06-01

Species of the genus Steganacarus are soil-living oribatid mites (Acari, Phthiracaridae) with a ptychoid body. The phylogeny and species status of the species of Steganacarus are not resolved, some authors group all ten German species of Steganacarus within the genus Steganacarus whereas others split them into three subgenera, Steganacarus, Tropacarus and Atropacarus. Additionally, two species, S. magnus and T. carinatus, comprise morphotypes of questionable species status. We investigated the phylogeny and species status of ten European Steganacarus species, i.e. S. applicatus, S. herculeanus, S. magnus forma magna, S. magnus forma anomala, S. spinosus, Tropacarus brevipilus, T. carinatus forma carinata, T. carinatus forma pulcherrima, Atropacarus striculus and Rhacaplacarus ortizi. We used two molecular markers, a 251 bp fragment of the nuclear gene 28S rDNA (D3) and a 477 bp fragment of the mitochondrial COI region. The phylogeny based on a combined analysis of D3 and COI separated four subgenera (Steganacarus, Tropacarus and Atropacarus, Rhacaplacarus) indicating that they form monophyletic groups. The COI region separated all ten species of the genus Steganacarus and showed variation within some species often correlating with the geographic origin of the species. Resolution of the more conserved D3 region was limited, indicating that radiation events are rather recent. Overall, our results indicate that both genes alone cannot be used for phylogeny and barcoding since variation is too low in D3 and too high in COI. However, when used in combination these genes provide reliable insight into the phylogeny, radiation and species status of taxa of the genus Steganacarus.

Relative profile analysis of molecular markers for identification and genetic discrimination of loaches (Pisces, Nemacheilidae).

PubMed

Patil, Tejas Suresh; Tamboli, Asif Shabodin; Patil, Swapnil Mahadeo; Bhosale, Amrut Ravindra; Govindwar, Sanjay Prabhu; Muley, Dipak Vishwanathrao

2016-01-01

Genus Nemacheilus, Nemachilichthys and Schistura belong to the family Nemacheilidae of the order Cypriniformes. The present investigation was undertaken to observe genetic diversity, phylogenetic relationship and to develop a molecular-based tool for taxonomic identification. For this purpose, four different types of molecular markers were utilized in which 29 random amplified polymorphic DNA (RAPD), 25 inter-simple sequence repeat (ISSR) markers, and 10 amplified fragment length polymorphism (AFLP) marker sets were screened and mitochondrial COI gene was sequenced. This study added COI barcodes for the identification of Nemacheilus anguilla, Nemachilichthys rueppelli and Schistura denisoni. RAPD showed higher polymorphism (100%) than the ISSR (93.75-100%) and AFLP (93.86-98.96%). The polymorphic information content (PIC), heterozygosity, multiplex ratio, and gene diversity was observed highest for AFLP primers, whereas the major allele frequency was observed higher for RAPD (0.5556) and lowest for AFLP (0.1667). The COI region of all individuals was successfully amplified and sequenced, which gave a 100% species resolution. Copyright © 2016 Académie des sciences. Published by Elsevier SAS. All rights reserved.

Quantifying Species Diversity with a DNA Barcoding-Based Method: Tibetan Moth Species (Noctuidae) on the Qinghai-Tibetan Plateau

PubMed Central

Jin, Qian; Han, Huilin; Hu, XiMin; Li, XinHai; Zhu, ChaoDong; Ho, Simon Y. W.; Ward, Robert D.; Zhang, Ai-bing

2013-01-01

With the ongoing loss of biodiversity, there is a great need for fast and effective ways to assess species richness and diversity: DNA barcoding provides a powerful new tool for this. We investigated this approach by focusing on the Tibetan plateau, which is one of the world's top biodiversity hotspots. There have been few studies of its invertebrates, although they constitute the vast majority of the region's diversity. Here we investigated species diversity of the lepidopteran family Noctuidae, across different environmental gradients, using measurements based on traditional morphology as well as on DNA barcoding. The COI barcode showed an average interspecific K2P distance of , which is about four times larger than the mean intraspecific distance (). Using six diversity indices, we did not detect any significant differences in estimated species diversity between measurements based on traditional morphology and on DNA barcoding. Furthermore, we found strong positive correlations between them, indicating that barcode-based measures of species diversity can serve as a good surrogate for morphology-based measures in most situations tested. Eastern communities were found to have significantly higher diversity than Western ones. Among 22 environmental factors tested, we found that three (precipitation of driest month, precipitation of driest quarter, and precipitation of coldest quarter) were significantly correlated with species diversity. Our results indicate that these factors could be the key ecological factors influencing the species diversity of the lepidopteran family Noctuidae on the Tibetan plateau. PMID:23741330

Sliding Window Analyses for Optimal Selection of Mini-Barcodes, and Application to 454-Pyrosequencing for Specimen Identification from Degraded DNA

PubMed Central

Boyer, Stephane; Brown, Samuel D. J.; Collins, Rupert A.; Cruickshank, Robert H.; Lefort, Marie-Caroline; Malumbres-Olarte, Jagoba; Wratten, Stephen D.

2012-01-01

DNA barcoding remains a challenge when applied to diet analyses, ancient DNA studies, environmental DNA samples and, more generally, in any cases where DNA samples have not been adequately preserved. Because the size of the commonly used barcoding marker (COI) is over 600 base pairs (bp), amplification fails when the DNA molecule is degraded into smaller fragments. However, relevant information for specimen identification may not be evenly distributed along the barcoding region, and a shorter target can be sufficient for identification purposes. This study proposes a new, widely applicable, method to compare the performance of all potential ‘mini-barcodes’ for a given molecular marker and to objectively select the shortest and most informative one. Our method is based on a sliding window analysis implemented in the new R package SPIDER (Species IDentity and Evolution in R). This method is applicable to any taxon and any molecular marker. Here, it was tested on earthworm DNA that had been degraded through digestion by carnivorous landsnails. A 100 bp region of 16 S rDNA was selected as the shortest informative fragment (mini-barcode) required for accurate specimen identification. Corresponding primers were designed and used to amplify degraded earthworm (prey) DNA from 46 landsnail (predator) faeces using 454-pyrosequencing. This led to the detection of 18 earthworm species in the diet of the snail. We encourage molecular ecologists to use this method to objectively select the most informative region of the gene they aim to amplify from degraded DNA. The method and tools provided here, can be particularly useful (1) when dealing with degraded DNA for which only small fragments can be amplified, (2) for cases where no consensus has yet been reached on the appropriate barcode gene, or (3) to allow direct analysis of short reads derived from massively parallel sequencing without the need for bioinformatic consolidation. PMID:22666489

A Tri-Oceanic Perspective: DNA Barcoding Reveals Geographic Structure and Cryptic Diversity in Canadian Polychaetes

PubMed Central

Carr, Christina M.; Hardy, Sarah M.; Brown, Tanya M.; Macdonald, Tara A.; Hebert, Paul D. N.

2011-01-01

Background Although polychaetes are one of the dominant taxa in marine communities, their distributions and taxonomic diversity are poorly understood. Recent studies have shown that many species thought to have broad distributions are actually a complex of allied species. In Canada, 12% of polychaete species are thought to occur in Atlantic, Arctic, and Pacific Oceans, but the extent of gene flow among their populations has not been tested. Methodology/Principal Findings Sequence variation in a segment of the mitochondrial cytochrome c oxidase I (COI) gene was employed to compare morphological versus molecular diversity estimates, to examine gene flow among populations of widespread species, and to explore connectivity patterns among Canada's three oceans. Analysis of 1876 specimens, representing 333 provisional species, revealed 40 times more sequence divergence between than within species (16.5% versus 0.38%). Genetic data suggest that one quarter of previously recognized species actually include two or more divergent lineages, indicating that richness in this region is currently underestimated. Few species with a tri-oceanic distribution showed genetic cohesion. Instead, large genetic breaks occur between Pacific and Atlantic-Arctic lineages, suggesting their long-term separation. High connectivity among Arctic and Atlantic regions and low connectivity with the Pacific further supports the conclusion that Canadian polychaetes are partitioned into two distinct faunas. Conclusions/Significance Results of this study confirm that COI sequences are an effective tool for species identification in polychaetes, and suggest that DNA barcoding will aid the recognition of species overlooked by the current taxonomic system. The consistent geographic structuring within presumed widespread species suggests that historical range fragmentation during the Pleistocene ultimately increased Canadian polychaete diversity and that the coastal British Columbia fauna played a minor role in Arctic recolonization following deglaciation. This study highlights the value of DNA barcoding for providing rapid insights into species distributions and biogeographic patterns in understudied groups. PMID:21829451

DNA Barcoding of Sigmodontine Rodents: Identifying Wildlife Reservoirs of Zoonoses

PubMed Central

Müller, Lívia; Gonçalves, Gislene L.; Cordeiro-Estrela, Pedro; Marinho, Jorge R.; Althoff, Sérgio L.; Testoni, André. F.; González, Enrique M.; Freitas, Thales R. O.

2013-01-01

Species identification through DNA barcoding is a tool to be added to taxonomic procedures, once it has been validated. Applying barcoding techniques in public health would aid in the identification and correct delimitation of the distribution of rodents from the subfamily Sigmodontinae. These rodents are reservoirs of etiological agents of zoonoses including arenaviruses, hantaviruses, Chagas disease and leishmaniasis. In this study we compared distance-based and probabilistic phylogenetic inference methods to evaluate the performance of cytochrome c oxidase subunit I (COI) in sigmodontine identification. A total of 130 sequences from 21 field-trapped species (13 genera), mainly from southern Brazil, were generated and analyzed, together with 58 GenBank sequences (24 species; 10 genera). Preliminary analysis revealed a 9.5% rate of misidentifications in the field, mainly of juveniles, which were reclassified after examination of external morphological characters and chromosome numbers. Distance and model-based methods of tree reconstruction retrieved similar topologies and monophyly for most species. Kernel density estimation of the distance distribution showed a clear barcoding gap with overlapping of intraspecific and interspecific densities < 1% and 21 species with mean intraspecific distance < 2%. Five species that are reservoirs of hantaviruses could be identified through DNA barcodes. Additionally, we provide information for the description of a putative new species, as well as the first COI sequence of the recently described genus Drymoreomys. The data also indicated an expansion of the distribution of Calomys tener. We emphasize that DNA barcoding should be used in combination with other taxonomic and systematic procedures in an integrative framework and based on properly identified museum collections, to improve identification procedures, especially in epidemiological surveillance and ecological assessments. PMID:24244670

Molecular-based estimate of species number, phylogenetic relationships and divergence times for the genus Stenotaenia (Chilopoda, Geophilomorpha) in the Italian region

PubMed Central

Del Latte, Laura; Bortolin, Francesca; Rota-Stabelli, Omar; Fusco, Giuseppe; Bonato, Lucio

2015-01-01

Abstract Stenotaenia is one of the largest and most widespread genera of geophilid centipedes in the Western Palearctic, with a very uniform morphology and about fifteen species provisionally recognized. For a better understanding of Stenotaenia species-level taxonomy, we have explored the possibility of using molecular data. As a preliminary assay, we sampled twelve populations, mainly from the Italian region, and analyzed partial sequences of the two genes COI and 28S. We employed a DNA-barcoding approach, complemented by a phylogenetic analysis coupled with divergence time estimation. Assuming a barcoding gap of 10–16% K2P pairwise distances, we found evidence for the presence of at least six Stenotaenia species in the Italian region, which started diverging about 50 million years ago, only partially matching with previously recognized species. We found that small-sized oligopodous species belong to a single clade that originated about 33 million years ago, and obtained some preliminary evidence of the related genus Tuoba being nested within Stenotaenia. PMID:26257533

PCR-based assessment of shellfish traceability and sustainability in international Mediterranean seafood markets.

PubMed

Galal-Khallaf, Asmaa; Ardura, Alba; Borrell, Yaisel J; Garcia-Vazquez, Eva

2016-07-01

Two mitochondrial markers (cytochrome oxidase COI and 16S rDNA) were employed for species identification of commercial shellfish from two Mediterranean countries. New COI Barcodes were generated for six species: Pleoticus robustus, Metapenaeopsis barbata, Parapenaeus fissuroides, Hymenopenaeus debilis, Metapenaeus affinis and Sepia aculeata. Biodiversity of the seafood species analyzed was greater in Egypt, with nine crustacean and two cephalopod species found compared with only three crustaceans and three cephalopods in Spain. In total, 17.2% and 15.2% products were mislabeled in Egypt and Spain, respectively. Population decline is a problem for some of the substitute species. Others were exotic and/or invasive in exporters' regions. This study offers the first comparable study of shellfish traceability in these Mediterranean markets. The PCR-based method used in this study proved to be reliable, effective and, therefore, could be employed for routine seafood analysis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Identification of Forensically Important Calliphoridae and Sarcophagidae Species Collected in Korea Using SNaPshot Multiplex System Targeting the Cytochrome c Oxidase Subunit I Gene

PubMed Central

Park, Ji Hye

2018-01-01

Estimation of postmortem interval (PMI) is paramount in modern forensic investigation. After the disappearance of the early postmortem phenomena conventionally used to estimate PMI, entomologic evidence provides important indicators for PMI estimation. The age of the oldest fly larvae or pupae can be estimated to pinpoint the time of oviposition, which is considered the minimum PMI (PMImin). The development rate of insects is usually temperature dependent and species specific. Therefore, species identification is mandatory for PMImin estimation using entomological evidence. The classical morphological identification method cannot be applied when specimens are damaged or have not yet matured. To overcome this limitation, some investigators employ molecular identification using mitochondrial cytochrome c oxidase subunit I (COI) nucleotide sequences. The molecular identification method commonly uses Sanger's nucleotide sequencing and molecular phylogeny, which are complex and time consuming and constitute another obstacle for forensic investigators. In this study, instead of using conventional Sanger's nucleotide sequencing, single-nucleotide polymorphisms (SNPs) in the COI gene region, which are unique between fly species, were selected and targeted for single-base extension (SBE) technology. These SNPs were genotyped using a SNaPshot® kit. Eleven Calliphoridae and seven Sarcophagidae species were covered. To validate this genotyping, fly DNA samples (103 adults, 84 larvae, and 4 pupae) previously confirmed by DNA barcoding were used. This method worked quickly with minimal DNA, providing a potential alternative to conventional DNA barcoding. Consisting of only a few simple electropherogram peaks, the results were more straightforward compared with those of the conventional DNA barcoding produced by Sanger's nucleotide sequencing. PMID:29682531

Morphological description and DNA barcoding of Hydrobaenus majus sp. nov. (Diptera: Chironomidae: Orthocladiinae) from the Russian Far East.

PubMed

Makarchenko, Eugenyi A; Makarchenko, Marina A; Semenchenko, Alexander A

2015-08-14

Illustrated descriptions of adult male, pupa and fourth instar larva, as well as DNA barcoding, of Hydrobaenus majus sp. nov. in comparison with the close related species H. sikhotealinensis Makarchenko et Makarchenko from the Russian Far East are provided. The species-specificity of H. majus sp. nov. COI sequences is analyzed and the sequences are presented as diagnostic characters--molecular markers of H. majus and H. sikhotealinensis.

Maternal mismatches in farmed tilapia strains (Oreochromis spp.) in the Philippines as revealed by mitochondrial COI gene.

PubMed

Ordoñez, June Feliciano F; Ventolero, Minerva Fatimae H; Santos, Mudjekeewis D

2017-07-01

The introduction of genetically enhanced tilapia has significantly boosted the performance of Philippine aquaculture industry. While enhanced strains contribute to the increase in tilapia production, genetic characterization of present tilapia stocks is critical to maintain their quality and to ensure the genetic gains are sustained. To understand and determine the genetic relationship of the genetically enhanced strains produced in the Philippines, mitochondrial cytochrome oxidase subunit I (COI) gene using DNA barcoding approach was analyzed. Specimens representing 10 genetically enhanced strains (GIFT, FaST, GET-EXCEL, GST, SST, COLD, YY-male, GMT, Molobicus, and BEST), three red tilapia (Taiwan red, Florida red, and FAC-red), and two pure lines (initially identified as O. aureus and O. spilurus) were collected, sequenced, and identified using DNA barcoding. Results revealed that farmed tilapias consisted of four different Oreochromis species. As expected, COI could not distinguish individuals at the strain level but surprisingly, mismatch between the species of maternal origin and present-day offspring was observed. This particular result may pose a question on the genetic purity and integrity of the strains being distributed to farmers and suggests a re-evaluation of the effectiveness of major tilapia breeding centers in maintaining their stocks.

Reading Mammal Diversity from Flies: The Persistence Period of Amplifiable Mammal mtDNA in Blowfly Guts (Chrysomya megacephala) and a New DNA Mini-Barcode Target.

PubMed

Lee, Ping-Shin; Sing, Kong-Wah; Wilson, John-James

2015-01-01

Most tropical mammal species are threatened or data-deficient. Data collection is impeded by the traditional monitoring approaches which can be laborious, expensive and struggle to detect cryptic diversity. Monitoring approaches using mammal DNA derived from invertebrates are emerging as cost- and time-effective alternatives. As a step towards development of blowfly-derived DNA as an effective method for mammal monitoring in the biodiversity hotspot of Peninsular Malaysia, our objectives were (i) to determine the persistence period of amplifiable mammal mtDNA in blowfly guts through a laboratory feeding experiment (ii) to design and test primers that can selectively amplify mammal COI DNA mini-barcodes in the presence of high concentrations of blowfly DNA. The persistence period of amplifiable mammal mtDNA in blowfly guts was 24 h to 96 h post-feeding indicating the need for collecting flies within 24 h of capture to detect mammal mtDNA of sufficient quantity and quality. We designed a new primer combination for a COI DNA mini-barcode that did not amplify blowfly DNA and showed 89% amplification success for a dataset of mammals from Peninsular Malaysia. The short (205 bp) DNA mini-barcode could distinguish most mammal species (including separating dark taxa) and is of suitable length for high-throughput sequencing. Our new DNA mini-barcode target and a standardized trapping protocol with retrieval of blowflies every 24 h could point the way forward in the development of blowfly-derived DNA as an effective method for mammal monitoring.

DNA barcoding for species identification in deep-sea clams (Mollusca: Bivalvia: Vesicomyidae).

PubMed

Liu, Jun; Zhang, Haibin

2018-01-15

Deep-sea clams (Bivalvia: Vesicomyidae) have been found in reduced environments over the world oceans, but taxonomy of this group remains confusing at species and supraspecific levels due to their high-morphological similarity and plasticity. In the present study, we collected mitochondrial COI sequences to evaluate the utility of DNA barcoding on identifying vesicomyid species. COI dataset identified 56 well-supported putative species/operational taxonomic units (OTUs), approximately covering half of the extant vesicomyid species. One species (OTU2) was first detected, and may represent a new species. Average distances between species ranged from 1.65 to 29.64%, generally higher than average intraspecific distances (0-1.41%) when excluding Pliocardia sp.10 cf. venusta (average intraspecific distance 1.91%). Local barcoding gap existed in 33 of the 35 species when comparing distances of maximum interspecific and minimum interspecific distances with two exceptions (Abyssogena southwardae and Calyptogena rectimargo-starobogatovi). The barcode index number (BIN) system determined 41 of the 56 species/OTUs, each with a unique BIN, indicating their validity. Three species were found to have two BINs, together with their high level of intraspecific variation, implying cryptic diversity within them. Although fewer 16 S sequences were collected, similar results were obtained. Nineteen putative species were determined and no overlap observed between intra- and inter-specific variation. Implications of DNA barcoding for the Vesicomyidae taxonomy were then discussed. Findings of this study will provide important evidence for taxonomic revision in this problematic clam group, and accelerate the discovery of new vesicomyid species in the future.

Reading Mammal Diversity from Flies: The Persistence Period of Amplifiable Mammal mtDNA in Blowfly Guts (Chrysomya megacephala) and a New DNA Mini-Barcode Target

PubMed Central

Lee, Ping-Shin; Sing, Kong-Wah; Wilson, John-James

2015-01-01

Most tropical mammal species are threatened or data-deficient. Data collection is impeded by the traditional monitoring approaches which can be laborious, expensive and struggle to detect cryptic diversity. Monitoring approaches using mammal DNA derived from invertebrates are emerging as cost- and time-effective alternatives. As a step towards development of blowfly-derived DNA as an effective method for mammal monitoring in the biodiversity hotspot of Peninsular Malaysia, our objectives were (i) to determine the persistence period of amplifiable mammal mtDNA in blowfly guts through a laboratory feeding experiment (ii) to design and test primers that can selectively amplify mammal COI DNA mini-barcodes in the presence of high concentrations of blowfly DNA. The persistence period of amplifiable mammal mtDNA in blowfly guts was 24 h to 96 h post-feeding indicating the need for collecting flies within 24 h of capture to detect mammal mtDNA of sufficient quantity and quality. We designed a new primer combination for a COI DNA mini-barcode that did not amplify blowfly DNA and showed 89% amplification success for a dataset of mammals from Peninsular Malaysia. The short (205 bp) DNA mini-barcode could distinguish most mammal species (including separating dark taxa) and is of suitable length for high-throughput sequencing. Our new DNA mini-barcode target and a standardized trapping protocol with retrieval of blowflies every 24 h could point the way forward in the development of blowfly-derived DNA as an effective method for mammal monitoring. PMID:25898278

Using DNA-barcoding to make the necrobiont beetle family Cholevidae accessible for forensic entomology.

PubMed

Schilthuizen, Menno; Scholte, Cindy; van Wijk, Renske E J; Dommershuijzen, Jessy; van der Horst, Devi; Zu Schlochtern, Melanie Meijer; Lievers, Rik; Groenenberg, Dick S J

2011-07-15

The beetle family Cholevidae (Coleoptera: Staphylinoidea), sometimes viewed as the subfamily Cholevinae of the Leiodidae, consists of some 1700 species worldwide. With the exception of specialized cave-dwelling species and species living in bird and mammal nests and burrows, the species are generalized soil-dwellers that, at least in temperate regions, are mostly found on vertebrate cadavers. Although they have been regularly reported from human corpses, and offer potential because of many species' peak activity in the cold season, they have not been a focus of forensic entomologists so far. This is probably due to their small size and the difficulty in identifying the adults and their larvae. In this paper, we show that DNA-barcoding can help make this group of necrobiont beetles available as a tool for forensic research. We collected 86 specimens of 20 species of the genera Catops, Fissocatops, Apocatops, Choleva, Nargus, Ptomaphagus, and Sciodrepoides from the Netherlands and France and show that a broad "barcoding gap" allows almost all species to be easily and unambiguously identified by the sequence of the "barcoding gene" cytochrome c oxidase I (COI). This opens up the possibility of adding Cholevidae to the set of insect taxa routinely used in forensic entomology. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Quadrulella texcalense sp. nov. from a Mexican desert: An unexpected new environment for hyalospheniid testate amoebae.

PubMed

Pérez-Juárez, Horacio; Serrano-Vázquez, Angélica; Kosakyan, Anush; Mitchell, Edward A D; Rivera Aguilar, Víctor M; Lahr, Daniel J G; Hernández Moreno, Mayra M; Cuellar, Humberto Macías; Eguiarte, Luis E; Lara, Enrique

2017-10-01

Quadrulella (Amoebozoa, Arcellinida, Hyalospheniidae) is a genus of testate amoebae with unmistakable morphology, which secretes characteristic square plates to reinforce the test. They are mainly known from fens and freshwater habitats and have never been documented in deserts. We describe a new species, Quadrulella texcalense, from biological soil crusts in the intertropical desert of Tehuacán (state of Puebla, Mexico). Quadrulella texcalense occurred only at altitudes between 2140 and 2221m.a.s.l., together with the bryophyte genera Pseudocrossidium, Weissia, Bryum, Didymodon, Neohyophyla and Aloina. The soil was extremely dry (moisture of 1.97-2.6%), which contrasts sharply with previous reports for the Quadrulella genus. Single cell mitochondrial cytochrome oxidase I (COI) barcoding of thirteen isolated cells showed an important morphological variability despite having all the same COI barcode sequence. Quadrulella texcalense was placed in a tree containing other Hyalsopheniidae, including a newly barcoded South African species, Q. elegans. Q. texcalense unambiguously branched within genus Quadrulella in a compact clade but with a long branch, suggesting accelerated evolution due to a transition towards a new environment and/or under-sampling. Copyright © 2017 Elsevier GmbH. All rights reserved.

DNA barcodes for dragonflies and damselflies (Odonata) of Mindanao, Philippines.

PubMed

Casas, Princess Angelie S; Sing, Kong-Wah; Lee, Ping-Shin; Nuñeza, Olga M; Villanueva, Reagan Joseph T; Wilson, John-James

2018-03-01

Reliable species identification provides a sounder basis for use of species in the order Odonata as biological indicators and for their conservation, an urgent concern as many species are threatened with imminent extinction. We generated 134 COI barcodes from 36 morphologically identified species of Odonata collected from Mindanao Island, representing 10 families and 19 genera. Intraspecific sequence divergences ranged from 0 to 6.7% with four species showing more than 2%, while interspecific sequence divergences ranged from 0.5 to 23.3% with seven species showing less than 2%. Consequently, no distinct gap was observed between intraspecific and interspecific DNA barcode divergences. The numerous islands of the Philippine archipelago may have facilitated rapid speciation in the Odonata and resulted in low interspecific sequence divergences among closely related groups of species. This study contributes DNA barcodes for 36 morphologically identified species of Odonata reported from Mindanao including 31 species with no previous DNA barcode records.

Molecular characterization of Anopheline (Diptera: Culicidae) mosquitoes from eight geographical locations of Sri Lanka.

PubMed

Weeraratne, Thilini C; Surendran, Sinnathambi N; Reimer, Lisa J; Wondji, Charles S; Perera, M Devika B; Walton, Catherine; Parakrama Karunaratne, S H P

2017-06-02

Genus Anopheles is a major mosquito group of interest in Sri Lanka as it includes vectors of malaria and its members exist as species complexes. Taxonomy of the group is mainly based on morphological features, which are not conclusive and can be easily erased while handling the specimens. A combined effort, using morphology and DNA barcoding (using the markers cytochrome c oxidase subunit I (COI) gene and internal transcribed spacer 2 (ITS2) region, was made during the present study to recognize anophelines collected from eight districts of Sri Lanka for the first time. Cytochrome c oxidase subunit I and ITS2 regions of morphologically identified anopheline mosquitoes from Sri Lanka were sequenced. These sequences together with GenBank sequences were used in phylogenetic tree construction and molecular characterization of mosquitoes. According to morphological identification, the field-collected adult mosquitoes belonged to 15 species, i.e., Anopheles aconitus, Anopheles annularis, Anopheles barbirostris, Anopheles culicifacies, Anopheles jamesii, Anopheles karwari, Anopheles maculatus, Anopheles nigerrimus, Anopheles pallidus, Anopheles peditaeniatus, Anopheles pseudojamesi, Anopheles subpictus, Anopheles tessellatus, Anopheles vagus, and Anopheles varuna. However, analysis of 123 COI sequences (445 bp) (16 clades supported by strong bootstrap value in the neighbour joining tree and inter-specific distances of >3%) showed that there are 16 distinct species. Identity of the morphologically identified species, except An. subpictus, was comparable with the DNA barcoding results. COI sequence analysis showed that morphologically identified An. subpictus is composed of two genetic entities: An. subpictus species A and species B (inter-specific K2P distance 0.128). All the four haplotypes of An. culicifacies discovered during the present study belonged to a single species. ITS2 sequences (542 bp) were obtained for all the species except for An. barbirostris, An. subpictus species B, An. tessellatus, and An. varuna. Each of these sequences was represented by a single species-specific haplotype. The present study reflects the importance and feasibility of COI and ITS2 genetic markers in identifying anophelines and their sibling species, and the significance of integrated systematic approach in mosquito taxonomy. Wide distribution of malaria vectors in the country perhaps indicates the potential for re-emergence of malaria in the country.

Efficacy of DNA barcoding for the species identification of spiders from Western Ghats of India.

PubMed

Gaikwad, Swapnil; Warudkar, Ashwin; Shouche, Yogesh

2017-09-01

DNA barcoding has emerged as an additional tool for taxonomy and as an aid to taxonomic impediments. Due to their extensive morphological variation, spiders are taxonomically challenging. Therefore, all over the world, attempts are being made to DNA barcode species of spiders. Till now no attempts were made to DNA barcode Indian spiders despite their rich diversity. We have generated DNA barcodes for 60 species (n = 112) of spiders for the first time from India. Although only 17 species were correctly identified at the species level, DNA barcoding correctly discriminated 99% of the species studied here. We have also found high intraspecies nucleotide divergence in Plexippus paykulli suggesting cryptic diversity that needs to be studied in detail. Our study also showed non-specific amplification of the Cytochrome Oxidase I (COI) gene of endosymbiont bacteria Wolbachia. However, these cases are very rare and could be resolved by the use of modified or group specific primers.

Mapping global biodiversity connections with DNA barcodes: Lepidoptera of Pakistan.

PubMed

Ashfaq, Muhammad; Akhtar, Saleem; Rafi, Muhammad Athar; Mansoor, Shahid; Hebert, Paul D N

2017-01-01

Sequences from the DNA barcode region of the mitochondrial COI gene are an effective tool for specimen identification and for the discovery of new species. The Barcode of Life Data Systems (BOLD) (www.boldsystems.org) currently hosts 4.5 million records from animals which have been assigned to more than 490,000 different Barcode Index Numbers (BINs), which serve as a proxy for species. Because a fourth of these BINs derive from Lepidoptera, BOLD has a strong capability to both identify specimens in this order and to support studies of faunal overlap. DNA barcode sequences were obtained from 4503 moths from 329 sites across Pakistan, specimens that represented 981 BINs from 52 families. Among 379 species with a Linnaean name assignment, all were represented by a single BIN excepting five species that showed a BIN split. Less than half (44%) of the 981 BINs had counterparts in other countries; the remaining BINs were unique to Pakistan. Another 218 BINs of Lepidoptera from Pakistan were coupled with the 981 from this study before being compared with all 116,768 BINs for this order. As expected, faunal overlap was highest with India (21%), Sri Lanka (21%), United Arab Emirates (20%) and with other Asian nations (2.1%), but it was very low with other continents including Africa (0.6%), Europe (1.3%), Australia (0.6%), Oceania (1.0%), North America (0.1%), and South America (0.1%). This study indicates the way in which DNA barcoding facilitates measures of faunal overlap even when taxa have not been assigned to a Linnean species.

Comparing and combining distance-based and character-based approaches for barcoding turtles.

PubMed

Reid, B N; LE, M; McCord, W P; Iverson, J B; Georges, A; Bergmann, T; Amato, G; Desalle, R; Naro-Maciel, E

2011-11-01

Molecular barcoding can serve as a powerful tool in wildlife forensics and may prove to be a vital aid in conserving organisms that are threatened by illegal wildlife trade, such as turtles (Order Testudines). We produced cytochrome oxidase subunit one (COI) sequences (650 bp) for 174 turtle species and combined these with publicly available sequences for 50 species to produce a data set representative of the breadth of the order. Variability within the barcode region was assessed, and the utility of both distance-based and character-based methods for species identification was evaluated. For species in which genetic material from more than one individual was available (n = 69), intraspecific divergences were 1.3% on average, although divergences greater than the customary 2% barcode threshold occurred within 15 species. High intraspecific divergences could indicate species with a high degree of internal genetic structure or possibly even cryptic species, although introgression is also probable in some of these taxa. Divergences between species of the same genus were 6.4% on average; however, 49 species were <2% divergent from congeners. Low levels of interspecific divergence could be caused by recent evolutionary radiations coupled with the low rates of mtDNA evolution previously observed in turtles. Complementing distance-based barcoding with character-based methods for identifying diagnostic sets of nucleotides provided better resolution in several cases where distance-based methods failed to distinguish species. An online identification engine was created to provide character-based identifications. This study constitutes the first comprehensive barcoding effort for this seriously threatened order. © 2011 Blackwell Publishing Ltd.

Mapping global biodiversity connections with DNA barcodes: Lepidoptera of Pakistan

PubMed Central

Akhtar, Saleem; Rafi, Muhammad Athar; Mansoor, Shahid; Hebert, Paul D. N.

2017-01-01

Sequences from the DNA barcode region of the mitochondrial COI gene are an effective tool for specimen identification and for the discovery of new species. The Barcode of Life Data Systems (BOLD) (www.boldsystems.org) currently hosts 4.5 million records from animals which have been assigned to more than 490,000 different Barcode Index Numbers (BINs), which serve as a proxy for species. Because a fourth of these BINs derive from Lepidoptera, BOLD has a strong capability to both identify specimens in this order and to support studies of faunal overlap. DNA barcode sequences were obtained from 4503 moths from 329 sites across Pakistan, specimens that represented 981 BINs from 52 families. Among 379 species with a Linnaean name assignment, all were represented by a single BIN excepting five species that showed a BIN split. Less than half (44%) of the 981 BINs had counterparts in other countries; the remaining BINs were unique to Pakistan. Another 218 BINs of Lepidoptera from Pakistan were coupled with the 981 from this study before being compared with all 116,768 BINs for this order. As expected, faunal overlap was highest with India (21%), Sri Lanka (21%), United Arab Emirates (20%) and with other Asian nations (2.1%), but it was very low with other continents including Africa (0.6%), Europe (1.3%), Australia (0.6%), Oceania (1.0%), North America (0.1%), and South America (0.1%). This study indicates the way in which DNA barcoding facilitates measures of faunal overlap even when taxa have not been assigned to a Linnean species. PMID:28339501

COI haplotype groups in Mesocriconema (Nematoda: Criconematidae) and their morphospecies associations.

PubMed

Powers, T O; Bernard, E C; Harris, T; Higgins, R; Olson, M; Lodema, M; Mullin, P; Sutton, L; Powers, K S

2014-07-03

Without applying an a priori bias for species boundaries, specimen identities in the plant-parasitic nematode genus Mesocriconema were evaluated by examining mitochondrial COI nucleotide sequences, morphology, and biogeography. A total of 242 specimens that morphologically conformed to the genus were individually photographed, measured, and amplified by a PCR primer set to preserve the linkage between specimen morphology and a specific DNA barcode sequence. Specimens were extracted from soil samples representing 45 locations across 23 ecoregions in North America. Dendrograms constructed by neighbor-joining, maximum likelihood, and Bayesian Inference using a 721-bp COI barcode were used to group COI haplotypes. Each tree-building approach resulted in 24 major haplotype groups within the dataset. The distinctiveness of these groups was evaluated by node support, genetic distance, absence of intermediates, and several measures of distinctiveness included in software used for the exploration of species boundaries. Five of the 24 COI haplotype groups corresponded to morphologically characterized, Linnaean species. Morphospecies conforming to M. discus, Discocriconemella inarata, M. rusticum, M. onoense, and M. kirjanovae were represented by groups composed of multiple closely related or identical COI haplotypes. In other cases, morphospecies names could be equally applied to multiple haplotype groups that were genetically distant from each other. Identification based on morphology alone resulted in M. curvatum and M. ornatum species designations applied to seven and three groups, respectively. Morphological characters typically used for species level identification were demonstrably variable within haplotype groups, suggesting caution in assigning species names based on published compendia that solely consider morphological characters. Morphospecies classified as M. xenoplax formed a monophyletic group composed of seven genetically distinct COI subgroups. The species Discocriconemella inarata is transferred to Mesocriconema inaratum based on its phylogenetic position on the COI tree as well as previous phylogenetic analyses using 18S, ITS1, and cytochrome b nucleotide sequences. This study indicates that some of the species considered cosmopolitan in their distribution are actually multispecies polyphyletic groupings and an accurate assessment of Mesocriconema species distributions will benefit from molecular determination of haplotype relationships. The groups revealed by COI analysis should provide a useful framework for the evaluation of additional Mesocriconema species and will improve the reliability of designating taxonomic units in studies of nematode biodiversity. 

Supervised DNA Barcodes species classification: analysis, comparisons and results

PubMed Central

2014-01-01

Background Specific fragments, coming from short portions of DNA (e.g., mitochondrial, nuclear, and plastid sequences), have been defined as DNA Barcode and can be used as markers for organisms of the main life kingdoms. Species classification with DNA Barcode sequences has been proven effective on different organisms. Indeed, specific gene regions have been identified as Barcode: COI in animals, rbcL and matK in plants, and ITS in fungi. The classification problem assigns an unknown specimen to a known species by analyzing its Barcode. This task has to be supported with reliable methods and algorithms. Methods In this work the efficacy of supervised machine learning methods to classify species with DNA Barcode sequences is shown. The Weka software suite, which includes a collection of supervised classification methods, is adopted to address the task of DNA Barcode analysis. Classifier families are tested on synthetic and empirical datasets belonging to the animal, fungus, and plant kingdoms. In particular, the function-based method Support Vector Machines (SVM), the rule-based RIPPER, the decision tree C4.5, and the Naïve Bayes method are considered. Additionally, the classification results are compared with respect to ad-hoc and well-established DNA Barcode classification methods. Results A software that converts the DNA Barcode FASTA sequences to the Weka format is released, to adapt different input formats and to allow the execution of the classification procedure. The analysis of results on synthetic and real datasets shows that SVM and Naïve Bayes outperform on average the other considered classifiers, although they do not provide a human interpretable classification model. Rule-based methods have slightly inferior classification performances, but deliver the species specific positions and nucleotide assignments. On synthetic data the supervised machine learning methods obtain superior classification performances with respect to the traditional DNA Barcode classification methods. On empirical data their classification performances are at a comparable level to the other methods. Conclusions The classification analysis shows that supervised machine learning methods are promising candidates for handling with success the DNA Barcoding species classification problem, obtaining excellent performances. To conclude, a powerful tool to perform species identification is now available to the DNA Barcoding community. PMID:24721333

DNA Barcoding of the parasitoid wasp subfamily Doryctinae (Hymenoptera: Braconidae) from Chamela, Mexico.

PubMed

Gutiérrez-Arellano, Daniela; Gutiérrez-Arellano, Claudia Renata; Zaldívar-Riverón, Alejandro

2015-01-01

Background and aims. The Doryctinae is a considerably diverse, poorly studied group of parasitoid wasps and one of the most diverse subfamilies within Braconidae. Taxonomic knowledge of this group remains highly incomplete, specially in the tropics. In Mexico, it has been reported as the subfamily with the highest number of recorded genera. A preliminary Barcoding study carried out in the Chamela region, located near the Mexican pacific coast in Jalisco, identified 185 barcoding species of Dorytinae assigned to 19 identified doryctine genera. This work updates the later study, representing a three years effort to assess the species richness of this subfamily for the Chamela region. Materials and methods. Ten collecting field trips of 5 to 10 days each were carried out from June 2009 to May 2011. A 2% divergence criterion using the BIN system implemented in BOLD was followed in order to establish species boundaries among the specimens that were collected. Results and conclusions. A total of 961 specimens were collected, from which 883 COI sequences were obtained. The sequences generated corresponded to 289 barcoding species and 30 identified genera. The most speciose genera were Heterospilus Haliday (170 spp.), Ecphylus Förster (19 spp.), Allorhogas Gahan (15 spp.) and Callihormius Ashmead (14 spp.). Addition of previously collected material increased the diversity of the subfamily in the region to 34 genera and 290 species. Paraphyly of Heterospilus with respect to Neoheterospilus and Heterospathius was again recovered. Twenty new species and two new genera (Sabinita Belokobylskij, Zaldívar-Riverón et Martínez, Ficobolus Martínez, Belokobylskij et Zaldívar-Riverón) have been described so far from the material collected in this work.

DNA Barcoding of the parasitoid wasp subfamily Doryctinae (Hymenoptera: Braconidae) from Chamela, Mexico

PubMed Central

Gutiérrez-Arellano, Daniela; Gutiérrez-Arellano, Claudia Renata

2015-01-01

Abstract Background and aims. The Doryctinae is a considerably diverse, poorly studied group of parasitoid wasps and one of the most diverse subfamilies within Braconidae. Taxonomic knowledge of this group remains highly incomplete, specially in the tropics. In Mexico, it has been reported as the subfamily with the highest number of recorded genera. A preliminary Barcoding study carried out in the Chamela region, located near the Mexican pacific coast in Jalisco, identified 185 barcoding species of Dorytinae assigned to 19 identified doryctine genera. This work updates the later study, representing a three years effort to assess the species richness of this subfamily for the Chamela region. Materials and methods. Ten collecting field trips of 5 to 10 days each were carried out from June 2009 to May 2011. A 2% divergence criterion using the BIN system implemented in BOLD was followed in order to establish species boundaries among the specimens that were collected. Results and conclusions. A total of 961 specimens were collected, from which 883 COI sequences were obtained. The sequences generated corresponded to 289 barcoding species and 30 identified genera. The most speciose genera were Heterospilus Haliday (170 spp.), Ecphylus Förster (19 spp.), Allorhogas Gahan (15 spp.) and Callihormius Ashmead (14 spp.). Addition of previously collected material increased the diversity of the subfamily in the region to 34 genera and 290 species. Paraphyly of Heterospilus with respect to Neoheterospilus and Heterospathius was again recovered. Twenty new species and two new genera (Sabinita Belokobylskij, Zaldívar-Riverón et Martínez, Ficobolus Martínez, Belokobylskij et Zaldívar-Riverón) have been described so far from the material collected in this work. PMID:26023287

Squalius namak, a new chub from Lake Namak basin in Iran (Teleostei: Cyprinidae).

PubMed

Khaefi, Roozbehan; Esmaeili, Hamid Reza; Sayyadzadeh, Golnaz; Geiger, Matthias F; Freyhof, Jörg

2016-09-19

Squalius namak, new species, from the endorheic Lake Namak and Kavir basins in Iran, is distinguished from the species of the genus Squalius in the Persian Gulf and the southern Caspian Sea basins by having a wide and thick symphysial knob on the lower jaw, a convex posterior anal-fin margin, a bold, dark-grey or brown, roundish or crescent-shaped blotch at the posterior tip of each flank scale and orange caudal-, anal- and pelvic-fin rays in life. Squalius namak is also characterized by four fixed, diagnostic nucleotide substitutions in the mtDNA COI barcode region.

Composition and Genetic Diversity of Mosquitoes (Diptera: Culicidae) on Islands and Mainland Shores of Kenya’s Lakes Victoria and Baringo

PubMed Central

Ajamma, Yvonne Ukamaka; Villinger, Jandouwe; Omondi, David; Salifu, Daisy; Onchuru, Thomas Ogao; Njoroge, Laban; Muigai, Anne W. T.; Masiga, Daniel K.

2016-01-01

The Lake Baringo and Lake Victoria regions of Kenya are associated with high seroprevalence of mosquito-transmitted arboviruses. However, molecular identification of potential mosquito vector species, including morphologically identified ones, remains scarce. To estimate the diversity, abundance, and distribution of mosquito vectors on the mainland shores and adjacent inhabited islands in these regions, we collected and morphologically identified adult and immature mosquitoes and obtained the corresponding sequence variation at cytochrome c oxidase 1 (COI) and internal transcribed spacer region 2 (ITS2) gene regions. A total of 63 species (including five subspecies) were collected from both study areas, 47 of which have previously been implicated as disease vectors. Fourteen species were found only on island sites, which are rarely included in mosquito diversity surveys. We collected more mosquitoes, yet with lower species composition, at Lake Baringo (40,229 mosquitoes, 32 species) than at Lake Victoria (22,393 mosquitoes, 54 species). Phylogenetic analysis of COI gene sequences revealed Culex perexiguus and Cx. tenagius that could not be distinguished morphologically. Most Culex species clustered into a heterogeneous clade with closely related sequences, while Culex pipiens clustered into two distinct COI and ITS2 clades. These data suggest limitations in current morphological identification keys. This is the first DNA barcode report of Kenyan mosquitoes. To improve mosquito species identification, morphological identifications should be supported by their molecular data, while diversity surveys should target both adults and immatures. The diversity of native mosquito disease vectors identified in this study impacts disease transmission risks to humans and livestock. PMID:27402888

Molecular taxonomy of the two Leishmania vectors Lutzomyia umbratilis and Lutzomyia anduzei (Diptera: Psychodidae) from the Brazilian Amazon

PubMed Central

2013-01-01

Background Lutzomyia umbratilis (a probable species complex) is the main vector of Leishmania guyanensis in the northern region of Brazil. Lutzomyia anduzei has been implicated as a secondary vector of this parasite. These species are closely related and exhibit high morphological similarity in the adult stage; therefore, they have been wrongly identified, both in the past and in the present. This shows the need for employing integrated taxonomy. Methods With the aim of gathering information on the molecular taxonomy and evolutionary relationships of these two vectors, 118 sequences of 663 base pairs (barcode region of the mitochondrial DNA cytochrome oxidase I – COI) were generated from 72 L. umbratilis and 46 L. anduzei individuals captured, respectively, in six and five localities of the Brazilian Amazon. The efficiency of the barcode region to differentiate the L. umbratilis lineages I and II was also evaluated. The data were analyzed using the pairwise genetic distances matrix and the Neighbor-Joining (NJ) tree, both based on the Kimura Two Parameter (K2P) evolutionary model. Results The analyses resulted in 67 haplotypes: 32 for L. umbratilis and 35 for L. anduzei. The mean intra-specific genetic distance was 0.008 (0.002 to 0.010 for L. umbratilis; 0.008 to 0.014 for L. anduzei), whereas the mean interspecific genetic distance was 0.044 (0.041 to 0.046), supporting the barcoding gap. Between the L. umbratilis lineages I and II, it was 0.009 to 0.010. The NJ tree analysis strongly supported monophyletic clades for both L. umbratilis and L. anduzei, whereas the L. umbratilis lineages I and II formed two poorly supported monophyletic subclades. Conclusions The barcode region clearly separated the two species and may therefore constitute a valuable tool in the identification of the sand fly vectors of Leishmania in endemic leishmaniasis areas. However, the barcode region had not enough power to separate the two lineages of L. umbratilis, likely reflecting incipient species that have not yet reached the status of distinct species. PMID:24021095

Molecular taxonomy of the two Leishmania vectors Lutzomyia umbratilis and Lutzomyia anduzei (Diptera: Psychodidae) from the Brazilian Amazon.

PubMed

Scarpassa, Vera Margarete; Alencar, Ronildo Baiatone

2013-09-11

Lutzomyia umbratilis (a probable species complex) is the main vector of Leishmania guyanensis in the northern region of Brazil. Lutzomyia anduzei has been implicated as a secondary vector of this parasite. These species are closely related and exhibit high morphological similarity in the adult stage; therefore, they have been wrongly identified, both in the past and in the present. This shows the need for employing integrated taxonomy. With the aim of gathering information on the molecular taxonomy and evolutionary relationships of these two vectors, 118 sequences of 663 base pairs (barcode region of the mitochondrial DNA cytochrome oxidase I - COI) were generated from 72 L. umbratilis and 46 L. anduzei individuals captured, respectively, in six and five localities of the Brazilian Amazon. The efficiency of the barcode region to differentiate the L. umbratilis lineages I and II was also evaluated. The data were analyzed using the pairwise genetic distances matrix and the Neighbor-Joining (NJ) tree, both based on the Kimura Two Parameter (K2P) evolutionary model. The analyses resulted in 67 haplotypes: 32 for L. umbratilis and 35 for L. anduzei. The mean intra-specific genetic distance was 0.008 (0.002 to 0.010 for L. umbratilis; 0.008 to 0.014 for L. anduzei), whereas the mean interspecific genetic distance was 0.044 (0.041 to 0.046), supporting the barcoding gap. Between the L. umbratilis lineages I and II, it was 0.009 to 0.010. The NJ tree analysis strongly supported monophyletic clades for both L. umbratilis and L. anduzei, whereas the L. umbratilis lineages I and II formed two poorly supported monophyletic subclades. The barcode region clearly separated the two species and may therefore constitute a valuable tool in the identification of the sand fly vectors of Leishmania in endemic leishmaniasis areas. However, the barcode region had not enough power to separate the two lineages of L. umbratilis, likely reflecting incipient species that have not yet reached the status of distinct species.

Rapid and accurate taxonomic classification of insect (class Insecta) cytochrome c oxidase subunit 1 (COI) DNA barcode sequences using a naïve Bayesian classifier

PubMed Central

Porter, Teresita M; Gibson, Joel F; Shokralla, Shadi; Baird, Donald J; Golding, G Brian; Hajibabaei, Mehrdad

2014-01-01

Current methods to identify unknown insect (class Insecta) cytochrome c oxidase (COI barcode) sequences often rely on thresholds of distances that can be difficult to define, sequence similarity cut-offs, or monophyly. Some of the most commonly used metagenomic classification methods do not provide a measure of confidence for the taxonomic assignments they provide. The aim of this study was to use a naïve Bayesian classifier (Wang et al. Applied and Environmental Microbiology, 2007; 73: 5261) to automate taxonomic assignments for large batches of insect COI sequences such as data obtained from high-throughput environmental sequencing. This method provides rank-flexible taxonomic assignments with an associated bootstrap support value, and it is faster than the blast-based methods commonly used in environmental sequence surveys. We have developed and rigorously tested the performance of three different training sets using leave-one-out cross-validation, two field data sets, and targeted testing of Lepidoptera, Diptera and Mantodea sequences obtained from the Barcode of Life Data system. We found that type I error rates, incorrect taxonomic assignments with a high bootstrap support, were already relatively low but could be lowered further by ensuring that all query taxa are actually present in the reference database. Choosing bootstrap support cut-offs according to query length and summarizing taxonomic assignments to more inclusive ranks can also help to reduce error while retaining the maximum number of assignments. Additionally, we highlight gaps in the taxonomic and geographic representation of insects in public sequence databases that will require further work by taxonomists to improve the quality of assignments generated using any method.

Filling reference gaps via assembling DNA barcodes using high-throughput sequencing—moving toward barcoding the world

PubMed Central

Zhou, Chengran

2017-01-01

Abstract Over the past decade, biodiversity researchers have dedicated tremendous efforts to constructing DNA reference barcodes for rapid species registration and identification. Although analytical cost for standard DNA barcoding has been significantly reduced since early 2000, further dramatic reduction in barcoding costs is unlikely because Sanger sequencing is approaching its limits in throughput and chemistry cost. Constraints in barcoding cost not only led to unbalanced barcoding efforts around the globe, but also prevented high-throughput sequencing (HTS)–based taxonomic identification from applying binomial species names, which provide crucial linkages to biological knowledge. We developed an Illumina-based pipeline, HIFI-Barcode, to produce full-length Cytochrome c oxidase subunit I (COI) barcodes from pooled polymerase chain reaction amplicons generated by individual specimens. The new pipeline generated accurate barcode sequences that were comparable to Sanger standards, even for different haplotypes of the same species that were only a few nucleotides different from each other. Additionally, the new pipeline was much more sensitive in recovering amplicons at low quantity. The HIFI-Barcode pipeline successfully recovered barcodes from more than 78% of the polymerase chain reactions that didn’t show clear bands on the electrophoresis gel. Moreover, sequencing results based on the single molecular sequencing platform Pacbio confirmed the accuracy of the HIFI-Barcode results. Altogether, the new pipeline can provide an improved solution to produce full-length reference barcodes at about one-tenth of the current cost, enabling construction of comprehensive barcode libraries for local fauna, leading to a feasible direction for DNA barcoding global biomes. PMID:29077841

Filling reference gaps via assembling DNA barcodes using high-throughput sequencing-moving toward barcoding the world.

PubMed

Liu, Shanlin; Yang, Chentao; Zhou, Chengran; Zhou, Xin

2017-12-01

Over the past decade, biodiversity researchers have dedicated tremendous efforts to constructing DNA reference barcodes for rapid species registration and identification. Although analytical cost for standard DNA barcoding has been significantly reduced since early 2000, further dramatic reduction in barcoding costs is unlikely because Sanger sequencing is approaching its limits in throughput and chemistry cost. Constraints in barcoding cost not only led to unbalanced barcoding efforts around the globe, but also prevented high-throughput sequencing (HTS)-based taxonomic identification from applying binomial species names, which provide crucial linkages to biological knowledge. We developed an Illumina-based pipeline, HIFI-Barcode, to produce full-length Cytochrome c oxidase subunit I (COI) barcodes from pooled polymerase chain reaction amplicons generated by individual specimens. The new pipeline generated accurate barcode sequences that were comparable to Sanger standards, even for different haplotypes of the same species that were only a few nucleotides different from each other. Additionally, the new pipeline was much more sensitive in recovering amplicons at low quantity. The HIFI-Barcode pipeline successfully recovered barcodes from more than 78% of the polymerase chain reactions that didn't show clear bands on the electrophoresis gel. Moreover, sequencing results based on the single molecular sequencing platform Pacbio confirmed the accuracy of the HIFI-Barcode results. Altogether, the new pipeline can provide an improved solution to produce full-length reference barcodes at about one-tenth of the current cost, enabling construction of comprehensive barcode libraries for local fauna, leading to a feasible direction for DNA barcoding global biomes. © The Authors 2017. Published by Oxford University Press.

20 years since the introduction of DNA barcoding: from theory to application.

PubMed

Fišer Pečnikar, Živa; Buzan, Elena V

2014-02-01

Traditionally, taxonomic identification has relied upon morphological characters. In the last two decades, molecular tools based on DNA sequences of short standardised gene fragments, termed DNA barcodes, have been developed for species discrimination. The most common DNA barcode used in animals is a fragment of the cytochrome c oxidase (COI) mitochondrial gene, while for plants, two chloroplast gene fragments from the RuBisCo large subunit (rbcL) and maturase K (matK) genes are widely used. Information gathered from DNA barcodes can be used beyond taxonomic studies and will have far-reaching implications across many fields of biology, including ecology (rapid biodiversity assessment and food chain analysis), conservation biology (monitoring of protected species), biosecurity (early identification of invasive pest species), medicine (identification of medically important pathogens and their vectors) and pharmacology (identification of active compounds). However, it is important that the limitations of DNA barcoding are understood and techniques continually adapted and improved as this young science matures.

Evaluation of the efficacy of twelve mitochondrial protein-coding genes as barcodes for mollusk DNA barcoding.

PubMed

Yu, Hong; Kong, Lingfeng; Li, Qi

2016-01-01

In this study, we evaluated the efficacy of 12 mitochondrial protein-coding genes from 238 mitochondrial genomes of 140 molluscan species as potential DNA barcodes for mollusks. Three barcoding methods (distance, monophyly and character-based methods) were used in species identification. The species recovery rates based on genetic distances for the 12 genes ranged from 70.83 to 83.33%. There were no significant differences in intra- or interspecific variability among the 12 genes. The monophyly and character-based methods provided higher resolution than the distance-based method in species delimitation. Especially in closely related taxa, the character-based method showed some advantages. The results suggested that besides the standard COI barcode, other 11 mitochondrial protein-coding genes could also be potentially used as a molecular diagnostic for molluscan species discrimination. Our results also showed that the combination of mitochondrial genes did not enhance the efficacy for species identification and a single mitochondrial gene would be fully competent.

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

PubMed Central

Sonet, Gontran; Jordaens, Kurt; Braet, Yves; Bourguignon, Luc; Dupont, Eréna; Backeljau, Thierry; De Meyer, Marc; Desmyter, Stijn

2013-01-01

Abstract 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. PMID:24453564

New or little-known species of Chaetocladius s. str. Kieffer, 1911 (Diptera: Chironomidae: Orthocladiinae) from the Amur River basin (Russian Far East).

PubMed

Makarchenko, Eugenyi A; Makarchenko, Marina A; Semenchenko, Alexander A

2017-03-27

Chironomids of the subgenus Chaetocladius s. str. from the Amur River basin are revised using both morphological characters and molecular data. Three new species, C. egorych sp. nov., C. lopatinskiy sp. nov. and C. yavorskayae sp. nov., are described and figured. The pupa of C. fedotkin is described for the first time. Adult males of C. ligni and C. piger, little-known in the Far East, are redescribed and annotated, and key to males of the Chaetocladius s. str. from the Amur River basin is provided. A reference 658 bp barcode sequence from a fragment of the mitochondrial gene cytochrome oxidase I (COI) was used as a tool for species delimitation. Comparisons with corresponding regions of COI between 5 species in the subgenus produced K2P genetic distances of 8.3-12.6%, values well associated with interspecific variation. Molecular data were also used for the reconstruction of the phylogenetic relationships within the subgenus Chaetocladius s. str.

Use of mitochondrial COI gene for the identification of family Salticidae and Lycosidae of spiders.

PubMed

Naseem, Sajida; Tahir, Hafiz Muhammad

2018-01-01

In recent years, DNA barcoding has become quite popular for molecular identification of species because it is simple, quick and an affordable method. Present study was conducted to identify spiders of most abundant families, i.e. Salticidae and Lycosidae from citrus orchards in Sargodha district using DNA barcoding. A total of 160 specimens were subjected to DNA barcoding but, sequences up to 600 bp were recovered for 156 specimens. This molecular approach proved helpful to assign the exact taxon to those specimens which were misidentified through morphological characters in the study. We were succeeded to discriminate six species of Lycosidae and nine species of Salticidae through DNA barcoding. Results revealed the presence of clear barcode gap (discontinuity in intra- and inter-specific divergences) for members of both families. Furthermore, the maximum intra-specific divergence was less than NN (nearest neighbour) distance for all species. This suggested the reliability of DNA barcoding for spider's identification up to species level. We got 98% success in our study. It is concluded from present study that DNA barcoding is more reliable tool especially for immature spiders, when morphological characters are ambiguous.

Reliable DNA Barcoding Performance Proved for Species and Island Populations of Comoran Squamate Reptiles

PubMed Central

Hawlitschek, Oliver; Nagy, Zoltán T.; Berger, Johannes; Glaw, Frank

2013-01-01

In the past decade, DNA barcoding became increasingly common as a method for species identification in biodiversity inventories and related studies. However, mainly due to technical obstacles, squamate reptiles have been the target of few barcoding studies. In this article, we present the results of a DNA barcoding study of squamates of the Comoros archipelago, a poorly studied group of oceanic islands close to and mostly colonized from Madagascar. The barcoding dataset presented here includes 27 of the 29 currently recognized squamate species of the Comoros, including 17 of the 18 endemic species. Some species considered endemic to the Comoros according to current taxonomy were found to cluster with non-Comoran lineages, probably due to poorly resolved taxonomy. All other species for which more than one barcode was obtained corresponded to distinct clusters useful for species identification by barcoding. In most species, even island populations could be distinguished using barcoding. Two cryptic species were identified using the DNA barcoding approach. The obtained barcoding topology, a Bayesian tree based on COI sequences of 5 genera, was compared with available multigene topologies, and in 3 cases, major incongruences between the two topologies became evident. Three of the multigene studies were initiated after initial screening of a preliminary version of the barcoding dataset presented here. We conclude that in the case of the squamates of the Comoros Islands, DNA barcoding has proven a very useful and efficient way of detecting isolated populations and promising starting points for subsequent research. PMID:24069192

DNA Barcodes for the FIshes of the Narmada, One of India’s Longest Rivers

PubMed Central

Khedkar, Gulab Dattarao; Jamdade, Rahul; Naik, Suresh; David, Lior; Haymer, David

2014-01-01

This study describes the species diversity of fishes of the Narmada River in India. A total of 820 fish specimens were collected from 17 sampling locations across the whole river basin. Fish were taxonomically classified into one of 90 possible species based on morphological characters, and then DNA barcoding was employed using COI gene sequences as a supplemental identification method. A total of 314 different COI sequences were generated, and specimens were confirmed to belong to 85 species representing 63 genera, 34 families and 10 orders. Findings of this study include the identification of five putative cryptic or sibling species and 43 species not previously known from the Narmada River basin. Five species are endemic to India and three are introduced species that had not been previously reported to occur in the Narmada River. Conversely, 43 species previously reported to occur in the Narmada were not found. Genetic diversity and distance values were generated for all of the species within genera, families and orders using Kimura’s 2 parameter distance model followed by the construction of a Neighbor Joining tree. High resolution clusters generated in NJ trees aided the groupings of species corresponding to their genera and families which are in confirmation to the values generated by Automatic Barcode Gap Discovery bioinformatics platform. This aided to decide a threshold value for the discrimination of species boundary from the Narmada River. This study provides an important validation of the use of DNA barcode sequences for monitoring species diversity and changes within complex ecosystems such as the Narmada River. PMID:24991801

DNA barcodes for the fishes of the Narmada, one of India's longest rivers.

PubMed

Khedkar, Gulab Dattarao; Jamdade, Rahul; Naik, Suresh; David, Lior; Haymer, David

2014-01-01

This study describes the species diversity of fishes of the Narmada River in India. A total of 820 fish specimens were collected from 17 sampling locations across the whole river basin. Fish were taxonomically classified into one of 90 possible species based on morphological characters, and then DNA barcoding was employed using COI gene sequences as a supplemental identification method. A total of 314 different COI sequences were generated, and specimens were confirmed to belong to 85 species representing 63 genera, 34 families and 10 orders. Findings of this study include the identification of five putative cryptic or sibling species and 43 species not previously known from the Narmada River basin. Five species are endemic to India and three are introduced species that had not been previously reported to occur in the Narmada River. Conversely, 43 species previously reported to occur in the Narmada were not found. Genetic diversity and distance values were generated for all of the species within genera, families and orders using Kimura's 2 parameter distance model followed by the construction of a Neighbor Joining tree. High resolution clusters generated in NJ trees aided the groupings of species corresponding to their genera and families which are in confirmation to the values generated by Automatic Barcode Gap Discovery bioinformatics platform. This aided to decide a threshold value for the discrimination of species boundary from the Narmada River. This study provides an important validation of the use of DNA barcode sequences for monitoring species diversity and changes within complex ecosystems such as the Narmada River.

Potential for DNA-based identification of Great Lakes fauna: match and mismatch between taxa inventories and DNA barcode libraries.

PubMed

Trebitz, Anett S; Hoffman, Joel C; Grant, George W; Billehus, Tyler M; Pilgrim, Erik M

2015-07-22

DNA-based identification of mixed-organism samples offers the potential to greatly reduce the need for resource-intensive morphological identification, which would be of value both to bioassessment and non-native species monitoring. The ability to assign species identities to DNA sequences found depends on the availability of comprehensive DNA reference libraries. Here, we compile inventories for aquatic metazoans extant in or threatening to invade the Laurentian Great Lakes and examine the availability of reference mitochondrial COI DNA sequences (barcodes) in the Barcode of Life Data System for them. We found barcode libraries largely complete for extant and threatening-to-invade vertebrates (100% of reptile, 99% of fish, and 92% of amphibian species had barcodes). In contrast, barcode libraries remain poorly developed for precisely those organisms where morphological identification is most challenging; 46% of extant invertebrates lacked reference barcodes with rates especially high among rotifers, oligochaetes, and mites. Lack of species-level identification for many aquatic invertebrates also is a barrier to matching DNA sequences with physical specimens. Attaining the potential for DNA-based identification of mixed-organism samples covering the breadth of aquatic fauna requires a concerted effort to build supporting barcode libraries and voucher collections.

Potential for DNA-based identification of Great Lakes fauna: match and mismatch between taxa inventories and DNA barcode libraries

NASA Astrophysics Data System (ADS)

Trebitz, Anett S.; Hoffman, Joel C.; Grant, George W.; Billehus, Tyler M.; Pilgrim, Erik M.

2015-07-01

DNA-based identification of mixed-organism samples offers the potential to greatly reduce the need for resource-intensive morphological identification, which would be of value both to bioassessment and non-native species monitoring. The ability to assign species identities to DNA sequences found depends on the availability of comprehensive DNA reference libraries. Here, we compile inventories for aquatic metazoans extant in or threatening to invade the Laurentian Great Lakes and examine the availability of reference mitochondrial COI DNA sequences (barcodes) in the Barcode of Life Data System for them. We found barcode libraries largely complete for extant and threatening-to-invade vertebrates (100% of reptile, 99% of fish, and 92% of amphibian species had barcodes). In contrast, barcode libraries remain poorly developed for precisely those organisms where morphological identification is most challenging; 46% of extant invertebrates lacked reference barcodes with rates especially high among rotifers, oligochaetes, and mites. Lack of species-level identification for many aquatic invertebrates also is a barrier to matching DNA sequences with physical specimens. Attaining the potential for DNA-based identification of mixed-organism samples covering the breadth of aquatic fauna requires a concerted effort to build supporting barcode libraries and voucher collections.

Taxonomic challenges in freshwater fishes: a mismatch between morphology and DNA barcoding in fish of the north-eastern part of the Congo basin.

PubMed

Decru, Eva; Moelants, Tuur; De Gelas, Koen; Vreven, Emmanuel; Verheyen, Erik; Snoeks, Jos

2016-01-01

This study evaluates the utility of DNA barcoding to traditional morphology-based species identifications for the fish fauna of the north-eastern Congo basin. We compared DNA sequences (COI) of 821 samples from 206 morphologically identified species. Best match, best close match and all species barcoding analyses resulted in a rather low identification success of 87.5%, 84.5% and 64.1%, respectively. The ratio 'nearest-neighbour distance/maximum intraspecific divergence' was lower than 1 for 26.1% of the samples, indicating possible taxonomic problems. In ten genera, belonging to six families, the number of species inferred from mtDNA data exceeded the number of species identified using morphological features; and in four cases indications of possible synonymy were detected. Finally, the DNA barcodes confirmed previously known identification problems within certain genera of the Clariidae, Cyprinidae and Mormyridae. Our results underscore the large number of taxonomic problems lingering in the taxonomy of the fish fauna of the Congo basin and illustrate why DNA barcodes will contribute to future efforts to compile a reliable taxonomic inventory of the Congo basin fish fauna. Therefore, the obtained barcodes were deposited in the reference barcode library of the Barcode of Life Initiative. © 2015 John Wiley & Sons Ltd.

A revision of the Schinia Volupia (Fitch) species complex Lepidoptera: Noctuidae: Heliothinae)

USDA-ARS?s Scientific Manuscript database

DNA barcode analysis of cytochrome oxidase I (COI) could not differentiate between the species of the Schinia volupia (Fitch) complex including, S. volupia, S. masoni Smith, S. fulleri (McElvare), S. sanrafaeli (Opler), S. miniana (Grote), and S. biforma Smith. Genitalic characters could only differ...

Composition and Genetic Diversity of Mosquitoes (Diptera: Culicidae) on Islands and Mainland Shores of Kenya's Lakes Victoria and Baringo.

PubMed

Ajamma, Yvonne Ukamaka; Villinger, Jandouwe; Omondi, David; Salifu, Daisy; Onchuru, Thomas Ogao; Njoroge, Laban; Muigai, Anne W T; Masiga, Daniel K

2016-11-01

The Lake Baringo and Lake Victoria regions of Kenya are associated with high seroprevalence of mosquito-transmitted arboviruses. However, molecular identification of potential mosquito vector species, including morphologically identified ones, remains scarce. To estimate the diversity, abundance, and distribution of mosquito vectors on the mainland shores and adjacent inhabited islands in these regions, we collected and morphologically identified adult and immature mosquitoes and obtained the corresponding sequence variation at cytochrome c oxidase 1 (COI) and internal transcribed spacer region 2 (ITS2) gene regions. A total of 63 species (including five subspecies) were collected from both study areas, 47 of which have previously been implicated as disease vectors. Fourteen species were found only on island sites, which are rarely included in mosquito diversity surveys. We collected more mosquitoes, yet with lower species composition, at Lake Baringo (40,229 mosquitoes, 32 species) than at Lake Victoria (22,393 mosquitoes, 54 species). Phylogenetic analysis of COI gene sequences revealed Culex perexiguus and Cx tenagius that could not be distinguished morphologically. Most Culex species clustered into a heterogeneous clade with closely related sequences, while Culex pipiens clustered into two distinct COI and ITS2 clades. These data suggest limitations in current morphological identification keys. This is the first DNA barcode report of Kenyan mosquitoes. To improve mosquito species identification, morphological identifications should be supported by their molecular data, while diversity surveys should target both adults and immatures. The diversity of native mosquito disease vectors identified in this study impacts disease transmission risks to humans and livestock. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America.

Identifying krill eggs in the central California current using novel multiplex PCR primers: Applications and limitations

NASA Astrophysics Data System (ADS)

Carrion, C. N.; Slesinger, E.; Marinovic, B.

2016-02-01

Euphausiids, otherwise known as krill, are an important link between primary producers and higher trophic levels within the central California current upwelling system. Euphausia pacifica and Thysanoessa spinifera, two of the most common euphausiid species along the central California coast, are both broadcast spawners and have some overlap in habitat, e.g. near marine life hotspots like the Monterey Bay and Gulf of Farallones. Species composition of euphausiid egg population within these regions is currently unknown. Distinct morphological differences between their eggs are lost once the egg dies or is preserved via formalin, alcohol, or freezing. In this project we designed genus specific DNA primers (mtCOI) for use in a multiplex PCR to distinguish among spawned euphausiid eggs of Euphausia spp. and Thysanoessa spp. in central California current surface waters. Effective and ineffective application of primers in a multiplex versus single-plex PCR is discussed, with an emphasis on primer design limitations in reference to the available barcoded regions of mitochondrial cytochrome oxidase subunit I (mtCOI) for each species in GenBank. This new protocol expands current monitoring efforts into sampling a non-swimming portion of the population which has the potential to improve euphausiid biomass estimates.

Molecular Identification of Sibling Species of Sclerodermus (Hymenoptera: Bethylidae) That Parasitize Buprestid and Cerambycid Beetles by Using Partial Sequences of Mitochondrial DNA Cytochrome Oxidase Subunit 1 and 28S Ribosomal RNA Gene

PubMed Central

Jiang, Yuan; Yang, Zhongqi; Wang, Xiaoyi; Hou, Yuxia

2015-01-01

The species belonging to Sclerodermus (Hymenoptera: Bethylidae) are currently the most important insect natural enemies of wood borer pests, mainly buprestid and cerambycid beetles, in China. However, some sibling species of this genus are very difficult to distinguish because of their similar morphological features. To address this issue, we conducted phylogenetic and genetic analyses of cytochrome oxidase subunit I (COI) and 28S RNA gene sequences from eight species of Sclerodermus reared from different wood borer pests. The eight sibling species were as follows: S. guani Xiao et Wu, S. sichuanensis Xiao, S. pupariae Yang et Yao, and Sclerodermus spp. (Nos. 1–5). A 594-bp fragment of COI and 750-bp fragment of 28S were subsequently sequenced. For COI, the G-C content was found to be low in all the species, averaging to about 30.0%. Sequence divergences (Kimura-2-parameter distances) between congeneric species averaged to 4.5%, and intraspecific divergences averaged to about 0.09%. Further, the maximum sequence divergences between congeneric species and Sclerodermus sp. (No. 5) averaged to about 16.5%. All 136 samples analyzed were included in six reciprocally monophyletic clades in the COI neighbor-joining (NJ) tree. The NJ tree inferred from the 28S rRNA sequence yielded almost identical results, but the samples from S. guani, S. sichuanensis, S. pupariae, and Sclerodermus spp. (Nos. 1–4) clustered together and only Sclerodermus sp. (No. 5) clustered separately. Our findings indicate that the standard barcode region of COI can be efficiently used to distinguish morphologically similar Sclerodermus species. Further, we speculate that Sclerodermus sp. (No. 5) might be a new species of Sclerodermus. PMID:25782000

DNA barcoding Neotropical fishes: recent advances from the Pampa Plain, Argentina.

PubMed

Rosso, J J; Mabragaña, E; Castro, M González; de Astarloa, J M Díaz

2012-11-01

The fish fauna of the Pampa Plain, the southernmost distribution range of many Neotropical species, was barcoded in this study. COI sequences were analysed by means of distance (K2P/NJ) and character-based (ML) models, as well as the Barcode Index Number (BIN). K2P/NJ analysis was able to discriminate among all previously identified species while also revealing the likely occurrence of two cryptic species that were further supported by BIN and ML analyses. On the other hand, both BIN and ML were not able to discriminate between two species of Rineloricaria. Despite the small genetic divergence between A. cf. pampa and A. eigenmanniorum, a tight array of haplotypes was observed for each species in both the distance and character-based methods. Deep intraspecific divergences were detected in Cnesterodon decemmaculatus (5%) and Salminus brasiliensis (6%). For Salminus brasiliensis, these findings were further supported by character-based (ML) evidence and meristic and morphological data. Our results also showed that Pampa Plain representatives of Salminus brasiliensis, Rhamdia quelen, Hoplias malabaricus, Synbranchus marmoratus, Australoheros facetus, Oligosarcus jenynsii and Corydoras paleatus differed by more than 3% from their conspecifics from other parts of South America. Overall, this study was able to highlight the likely occurrence of a cryptic species in Salminus brasiliensis and also illustrate the strong geographical structure in the COI sequence composition of seven fish species from South America. © 2012 Blackwell Publishing Ltd.

Comparison of manual and semi-automatic DNA extraction protocols for the barcoding characterization of hematophagous louse flies (Diptera: Hippoboscidae).

PubMed

Gutiérrez-López, Rafael; Martínez-de la Puente, Josué; Gangoso, Laura; Soriguer, Ramón C; Figuerola, Jordi

2015-06-01

The barcoding of life initiative provides a universal molecular tool to distinguish animal species based on the amplification and sequencing of a fragment of the subunit 1 of the cytochrome oxidase (COI) gene. Obtaining good quality DNA for barcoding purposes is a limiting factor, especially in studies conducted on small-sized samples or those requiring the maintenance of the organism as a voucher. In this study, we compared the number of positive amplifications and the quality of the sequences obtained using DNA extraction methods that also differ in their economic costs and time requirements and we applied them for the genetic characterization of louse flies. Four DNA extraction methods were studied: chloroform/isoamyl alcohol, HotShot procedure, Qiagen DNeasy(®) Tissue and Blood Kit and DNA Kit Maxwell(®) 16LEV. All the louse flies were morphologically identified as Ornithophila gestroi and a single COI-based haplotype was identified. The number of positive amplifications did not differ significantly among DNA extraction procedures. However, the quality of the sequences was significantly lower for the case of the chloroform/isoamyl alcohol procedure with respect to the rest of methods tested here. These results may be useful for the genetic characterization of louse flies, leaving most of the remaining insect as a voucher. © 2015 The Society for Vector Ecology.

Identification of Neoceratitis asiatica (Becker) (Diptera: Tephritidae) based on morphological characteristics and DNA barcode.

PubMed

Guo, Shaokun; He, Jia; Zhao, Zihua; Liu, Lijun; Gao, Liyuan; Wei, Shuhua; Guo, Xiaoyu; Zhang, Rong; Li, Zhihong

2017-12-12

Neoceratitis asiatica (Becker), which especially infests wolfberry (Lycium barbarum L.), could cause serious economic losses every year in China, especially to organic wolfberry production. In some important wolfberry plantings, it is difficult and time-consuming to rear the larvae or pupae to adults for morphological identification. Molecular identification based on DNA barcode is a solution to the problem. In this study, 15 samples were collected from Ningxia, China. Among them, five adults were identified according to their morphological characteristics. The utility of mitochondrial DNA (mtDNA) cytochrome c oxidase I (COI) gene sequence as DNA barcode in distinguishing N. asiatica was evaluated by analysing Kimura 2-parameter distances and phylogenetic trees. There were significant differences between intra-specific and inter-specific genetic distances according to the barcoding gap analysis. The uncertain larval and pupal samples were within the same cluster as N. asiatica adults and formed sister cluster to N. cyanescens. A combination of morphological and molecular methods enabled accurate identification of N. asiatica. This is the first study using DNA barcode to identify N. asiatica and the obtained DNA sequences will be added to the DNA barcode database.

Mosquitoes of eastern Amazonian Ecuador: biodiversity, bionomics and barcodes

PubMed Central

Linton, Yvonne-Marie; Pecor, James E; Porter, Charles H; Mitchell, Luke Brett; Garzón-Moreno, Andrés; Foley, Desmond H; Pecor, David Brooks; Wilkerson, Richard C

2013-01-01

Two snapshot surveys to establish the diversity and ecological preferences of mosquitoes (Diptera: Culicidae) in the terra firme primary rain forest surrounding the Tiputini Biodiversity Station in the UNESCO Yasuní Biosphere Reserve of eastern Amazonian Ecuador were carried out in November 1998 and May 1999. The mosquito fauna of this region is poorly known; the focus of this study was to obtain high quality link-reared specimens that could be used to unequivocally confirm species level diversity through integrated systematic study of all life stages and DNA sequences. A total of 2,284 specimens were preserved; 1,671 specimens were link-reared with associated immature exuviae, all but 108 of which are slide mounted. This study identified 68 unique taxa belonging to 17 genera and 27 subgenera. Of these, 12 are new to science and 37 comprise new country records. DNA barcodes [658-bp of the mtDNA cytochrome c oxidase ( COI ) I gene] are presented for 58 individuals representing 20 species and nine genera. DNA barcoding proved useful in uncovering and confirming new species and we advocate an integrated systematics approach to biodiversity studies in future. Associated bionomics of all species collected are discussed. An updated systematic checklist of the mosquitoes of Ecuador (n = 179) is presented for the first time in 60 years. PMID:24473809

Indigenous species barcode database improves the identification of zooplankton

PubMed Central

Yang, Jianghua; Zhang, Wanwan; Sun, Jingying; Xie, Yuwei; Zhang, Yimin; Burton, G. Allen; Yu, Hongxia

2017-01-01

Incompleteness and inaccuracy of DNA barcode databases is considered an important hindrance to the use of metabarcoding in biodiversity analysis of zooplankton at the species-level. Species barcoding by Sanger sequencing is inefficient for organisms with small body sizes, such as zooplankton. Here mitochondrial cytochrome c oxidase I (COI) fragment barcodes from 910 freshwater zooplankton specimens (87 morphospecies) were recovered by a high-throughput sequencing platform, Ion Torrent PGM. Intraspecific divergence of most zooplanktons was < 5%, except Branchionus leydign (Rotifer, 14.3%), Trichocerca elongate (Rotifer, 11.5%), Lecane bulla (Rotifer, 15.9%), Synchaeta oblonga (Rotifer, 5.95%) and Schmackeria forbesi (Copepod, 6.5%). Metabarcoding data of 28 environmental samples from Lake Tai were annotated by both an indigenous database and NCBI Genbank database. The indigenous database improved the taxonomic assignment of metabarcoding of zooplankton. Most zooplankton (81%) with barcode sequences in the indigenous database were identified by metabarcoding monitoring. Furthermore, the frequency and distribution of zooplankton were also consistent between metabarcoding and morphology identification. Overall, the indigenous database improved the taxonomic assignment of zooplankton. PMID:28977035

Instances of erroneous DNA barcoding of metazoan invertebrates: Are universal cox1 gene primers too "universal"?

PubMed

Mioduchowska, Monika; Czyż, Michał Jan; Gołdyn, Bartłomiej; Kur, Jarosław; Sell, Jerzy

2018-01-01

The cytochrome c oxidase subunit I (cox1) gene is the main mitochondrial molecular marker playing a pivotal role in phylogenetic research and is a crucial barcode sequence. Folmer's "universal" primers designed to amplify this gene in metazoan invertebrates allowed quick and easy barcode and phylogenetic analysis. On the other hand, the increase in the number of studies on barcoding leads to more frequent publishing of incorrect sequences, due to amplification of non-target taxa, and insufficient analysis of the obtained sequences. Consequently, some sequences deposited in genetic databases are incorrectly described as obtained from invertebrates, while being in fact bacterial sequences. In our study, in which we used Folmer's primers to amplify COI sequences of the crustacean fairy shrimp Branchipus schaefferi (Fischer 1834), we also obtained COI sequences of microbial contaminants from Aeromonas sp. However, when we searched the GenBank database for sequences closely matching these contaminations we found entries described as representatives of Gastrotricha and Mollusca. When these entries were compared with other sequences bearing the same names in the database, the genetic distance between the incorrect and correct sequences amplified from the same species was c.a. 65%. Although the responsibility for the correct molecular identification of species rests on researchers, the errors found in already published sequences data have not been re-evaluated so far. On the basis of the standard sampling technique we have estimated with 95% probability that the chances of finding incorrectly described metazoan sequences in the GenBank depend on the systematic group, and variety from less than 1% (Mollusca and Arthropoda) up to 6.9% (Gastrotricha). Consequently, the increasing popularity of DNA barcoding and metabarcoding analysis may lead to overestimation of species diversity. Finally, the study also discusses the sources of the problems with amplification of non-target sequences.

With a little help from DNA barcoding: investigating the diversity of Gastropoda from the Portuguese coast

PubMed Central

Borges, Luísa M. S.; Hollatz, Claudia; Lobo, Jorge; Cunha, Ana M.; Vilela, Ana P.; Calado, Gonçalo; Coelho, Rita; Costa, Ana C.; Ferreira, Maria S. G.; Costa, Maria H.; Costa, Filipe O.

2016-01-01

The Gastropoda is one of the best studied classes of marine invertebrates. Yet, most species have been delimited based on morphology only. The application of DNA barcodes has shown to be greatly useful to help delimiting species. Therefore, sequences of the cytochrome c oxidase I gene from 108 specimens of 34 morpho-species were used to investigate the molecular diversity within the gastropods from the Portuguese coast. To the above dataset, we added available COI-5P sequences of taxonomically close species, in a total of 58 morpho-species examined. There was a good match between ours and sequences from independent studies, in public repositories. We found 32 concordant (91.4%) out of the 35 Barcode Index Numbers (BINs) generated from our sequences. The application of a ranking system to the barcodes yield over 70% with top taxonomic congruence, while 14.2% of the species barcodes had insufficient data. In the majority of the cases, there was a good concordance between morphological identification and DNA barcodes. Nonetheless, the discordance between morphological and molecular data is a reminder that even the comparatively well-known European marine gastropods can benefit from being probed using the DNA barcode approach. Discordant cases should be reviewed with more integrative studies. PMID:26876495

With a little help from DNA barcoding: investigating the diversity of Gastropoda from the Portuguese coast.

PubMed

Borges, Luísa M S; Hollatz, Claudia; Lobo, Jorge; Cunha, Ana M; Vilela, Ana P; Calado, Gonçalo; Coelho, Rita; Costa, Ana C; Ferreira, Maria S G; Costa, Maria H; Costa, Filipe O

2016-02-15

The Gastropoda is one of the best studied classes of marine invertebrates. Yet, most species have been delimited based on morphology only. The application of DNA barcodes has shown to be greatly useful to help delimiting species. Therefore, sequences of the cytochrome c oxidase I gene from 108 specimens of 34 morpho-species were used to investigate the molecular diversity within the gastropods from the Portuguese coast. To the above dataset, we added available COI-5P sequences of taxonomically close species, in a total of 58 morpho-species examined. There was a good match between ours and sequences from independent studies, in public repositories. We found 32 concordant (91.4%) out of the 35 Barcode Index Numbers (BINs) generated from our sequences. The application of a ranking system to the barcodes yield over 70% with top taxonomic congruence, while 14.2% of the species barcodes had insufficient data. In the majority of the cases, there was a good concordance between morphological identification and DNA barcodes. Nonetheless, the discordance between morphological and molecular data is a reminder that even the comparatively well-known European marine gastropods can benefit from being probed using the DNA barcode approach. Discordant cases should be reviewed with more integrative studies.

A DNA barcode library for ground beetles (Insecta, Coleoptera, Carabidae) of Germany: The genus Bembidion Latreille, 1802 and allied taxa

PubMed Central

Raupach, Michael J.; Hannig, Karsten; Morinière, Jérome; Hendrich, Lars

2016-01-01

Abstract As molecular identification method, DNA barcoding based on partial cytochrome c oxidase subunit 1 (COI) sequences has been proven to be a useful tool for species determination in many insect taxa including ground beetles. In this study we tested the effectiveness of DNA barcodes to discriminate species of the ground beetle genus Bembidion and some closely related taxa of Germany. DNA barcodes were obtained from 819 individuals and 78 species, including sequences from previous studies as well as more than 300 new generated DNA barcodes. We found a 1:1 correspondence between BIN and traditionally recognized species for 69 species (89%). Low interspecific distances with maximum pairwise K2P values below 2.2% were found for three species pairs, including two species pairs with haplotype sharing (Bembidion atrocaeruleum/Bembidion varicolor and Bembidion guttula/Bembidion mannerheimii). In contrast to this, deep intraspecific sequence divergences with distinct lineages were revealed for two species (Bembidion geniculatum/Ocys harpaloides). Our study emphasizes the use of DNA barcodes for the identification of the analyzed ground beetles species and represents an important step in building-up a comprehensive barcode library for the Carabidae in Germany and Central Europe as well. PMID:27408547

Half of the European fruit fly species barcoded (Diptera, Tephritidae); a feasibility test for molecular identification

PubMed Central

Smit, John; Reijnen, Bastian; Stokvis, Frank

2013-01-01

Abstract A feasibility test of molecular identification of European fruit flies (Diptera: Tephritidae) based on COI barcode sequences has been executed. A dataset containing 555 sequences of 135 ingroup species from three subfamilies and 42 genera and one single outgroup species has been analysed. 73.3% of all included species could be identified based on their COI barcode gene, based on similarity and distances. The low success rate is caused by singletons as well as some problematic groups: several species groups within the genus Terellia and especially the genus Urophora. With slightly more than 100 sequences – almost 20% of the total – this genus alone constitutes the larger part of the failure for molecular identification for this dataset. Deleting the singletons and Urophora results in a success-rate of 87.1% of all queries and 93.23% of the not discarded queries as correctly identified. Urophora is of special interest due to its economic importance as beneficial species for weed control, therefore it is desirable to have alternative markers for molecular identification. We demonstrate that the success of DNA barcoding for identification purposes strongly depends on the contents of the database used to BLAST against. Especially the necessity of including multiple specimens per species of geographically distinct populations and different ecologies for the understanding of the intra- versus interspecific variation is demonstrated. Furthermore thresholds and the distinction between true and false positives and negatives should not only be used to increase the reliability of the success of molecular identification but also to point out problematic groups, which should then be flagged in the reference database suggesting alternative methods for identification. PMID:24453563

Barcoding Sponges: An Overview Based on Comprehensive Sampling

PubMed Central

Vargas, Sergio; Schuster, Astrid; Sacher, Katharina; Büttner, Gabrielle; Schätzle, Simone; Läuchli, Benjamin; Hall, Kathryn; Hooper, John N. A.; Erpenbeck, Dirk; Wörheide, Gert

2012-01-01

Background Phylum Porifera includes ∼8,500 valid species distributed world-wide in aquatic ecosystems ranging from ephemeral fresh-water bodies to coastal environments and the deep-sea. The taxonomy and systematics of sponges is complicated, and morphological identification can be both time consuming and erroneous due to phenotypic convergence and secondary losses, etc. DNA barcoding can provide sponge biologists with a simple and rapid method for the identification of samples of unknown taxonomic membership. The Sponge Barcoding Project (www.spongebarcoding.org), the first initiative to barcode a non-bilaterian metazoan phylum, aims to provide a comprehensive DNA barcode database for Phylum Porifera. Methodology/Principal Findings ∼7,400 sponge specimens have been extracted, and amplification of the standard COI barcoding fragment has been attempted for approximately 3,300 museum samples with ∼25% mean amplification success. Based on this comprehensive sampling, we present the first report on the workflow and progress of the sponge barcoding project, and discuss some common pitfalls inherent to the barcoding of sponges. Conclusion A DNA-barcoding workflow capable of processing potentially large sponge collections has been developed and is routinely used for the Sponge Barcoding Project with success. Sponge specific problems such as the frequent co-amplification of non-target organisms have been detected and potential solutions are currently under development. The initial success of this innovative project have already demonstrated considerable refinement of sponge systematics, evaluating morphometric character importance, geographic phenotypic variability, and the utility of the standard barcoding fragment for Porifera (despite its conserved evolution within this basal metazoan phylum). PMID:22802937

Identification and reassessment of the specific status of some tropical freshwater midges (Diptera: Chironomidae) using DNA barcode data.

PubMed

Pramual, Pairot; Simwisat, Kusumart; Martin, Jon

2016-01-28

Chironomidae are a highly diverse group of insects. Members of this family are often included in programs monitoring the health of freshwater ecosystems. However, a difficulty in morphological identification, particularly of larval stages is the major obstacle to this application. In this study, we tested the efficiency of mitochondrial cytochrome c oxidase I (COI) sequences as the DNA barcoding region for species identification of Chironomidae in Thailand. The results revealed 14 species with a high success rate (>90%) for the correct species identification, which suggests the potential usefulness of the technique. However, some morphological species possess high (>3%) intraspecific genetic divergence that suggests these species could be species complexes and need further morphological or cytological examination. Sequence-based species delimitation analyses indicated that most specimens identified as Chironomus kiiensis, Tokunaga 1936, in Japan are conspecific with C. striatipennis, Kieffer 1912, although a small number form a separate cluster. A review of the descriptions of Kiefferulus tainanus (Kieffer 1912) and its junior synonym, K. biroi (Kieffer 1918), following our results, suggests that this synonymy is probably not correct and that K. tainanus occurs in Japan, China and Singapore, while K. biroi occurs in India and Thailand. Our results therefore revealed the usefulness of DNA barcoding for correct species identification of Chironomidae, particularly the immature stages. In addition, DNA barcodes could also uncover hidden diversity that can guide further taxonomic study, and offer a more efficient way to identify species than morphological analysis where large numbers of specimens are involved, provided the identifications of DNA barcodes in the databases are correct. Our studies indicate that this is not the case, and we identify cases of misidentifications for C. flaviplumus, Tokunaga 1940 and K. tainanus.

DNA barcoding to identify species of phlebotomine sand fly (Diptera: Psychodidae) in the mixed leishmaniasis focus of the Colombian Caribbean.

PubMed

Romero-Ricardo, Luis; Lastre-Meza, Natalia; Pérez-Doria, Alveiro; Bejarano, Eduar E

2016-07-01

Identification of the species of phlebotomine sand flies present in each focus of leishmaniasis is necessary to incriminate vectors and implement vector control strategies. Although the cytochrome oxidase I (COI) gene has been proposed as a barcode for the identification of animal species, less than 20% of New World phlebotomines have been characterized to date. In this study DNA barcoding was used to identify phlebotomine species of the mixed leishmaniasis focus in the Colombian Caribbean by means of three evolutionary models: Kimura's two parameter (K2P) nucleotide substitution model, that of (Tamura and Nei, 1993) (TN93) and proportional sequence divergence (p-distances). A 681bp sequence of the COI gene was obtained from 66 individuals belonging to 19 species of the genus Lutzomyia (Lu. abonnenci, Lu. atroclavata, Lu. bicolor, Lu. carpenteri, Lu. cayennensis cayennensis, Lu. dubitans, Lu. evansi, Lu. gomezi, Lu. gorbitzi, Lu. longipalpis, Lu. micropyga, Lu. migonei, Lu. panamensis, Lu. (Psathyromyia) sp., Lu. rangeliana, Lu. serrana, Lu. shannoni, Lu. trinidadensis and Lu. venezuelensis) and one of Brumptomyia (B. mesai). The genetic divergence values for TN93 among individuals of the same species fluctuated up to 3.2% (vs. 2.9% for K2P and 2.8% for p-distances), while the values between species ranged from 8.8-43.7% (vs. 6.8-19.6% for K2P and 6.6-17.4% for p-distances). A dendrogram constructed by means of the Neighbor-Joining method grouped phlebotomines into 20 clusters according to species, with bootstrap values of up to 100% in those with more than one individual. However, loss of the phylogenetic signal of the gene COI was observed at the supraspecific level as a consequence of substitutional saturation. In conclusion, irrespective of the evolutionary model selected, all phlebotomines were correctly assigned to species, showing 100% concordance with morphological identification. Copyright © 2016 Elsevier B.V. All rights reserved.

Moorea BIOCODE barcode library as a tool for understanding predator-prey interactions: insights into the diet of common predatory coral reef fishes

NASA Astrophysics Data System (ADS)

Leray, M.; Boehm, J. T.; Mills, S. C.; Meyer, C. P.

2012-06-01

Identifying species involved in consumer-resource interactions is one of the main limitations in the construction of food webs. DNA barcoding of prey items in predator guts provides a valuable tool for characterizing trophic interactions, but the method relies on the availability of reference sequences to which prey sequences can be matched. In this study, we demonstrate that the COI sequence library of the Moorea BIOCODE project, an ecosystem-level barcode initiative, enables the identification of a large proportion of semi-digested fish, crustacean and mollusks found in the guts of three Hawkfish and two Squirrelfish species. While most prey remains lacked diagnostic morphological characters, 94% of the prey found in 67 fishes had >98% sequence similarity with BIOCODE reference sequences. Using this species-level prey identification, we demonstrate how DNA barcoding can provide insights into resource partitioning, predator feeding behaviors and the consequences of predation on ecosystem function.

Integrating DNA barcodes and morphology for species delimitation in the Corynoneura group (Diptera: Chironomidae: Orthocladiinae).

PubMed

Silva, F L; Wiedenbrug, S

2014-02-01

In this study, we use DNA barcodes for species delimitation to solve taxonomic conflicts in 86 specimens of 14 species belonging to the Corynoneura group (Diptera: Chironomidae: Orthocladiinae), from the Atlantic Forest, Brazil. Molecular analysis of cytochrome c-oxidase subunit I (COI) gene sequences supported 14 cohesive species groups, of which two similar groups were subsequently associated with morphological variation at the pupal stage. Eleven species previously described based on morphological criteria were linked to DNA markers. Furthermore, there is the possibility that there may be cryptic species within the Corynoneura group, since one group of species presented internal grouping, although no morphological divergence was observed. Our results support DNA-barcoding as an excellent tool for species delimitation in groups where taxonomy by means of morphology is difficult or even impossible.

Can DNA barcoding accurately discriminate megadiverse Neotropical freshwater fish fauna?

PubMed Central

2013-01-01

Background The megadiverse Neotropical freshwater ichthyofauna is the richest in the world with approximately 6,000 recognized species. Interestingly, they are distributed among only 17 orders, and almost 80% of them belong to only three orders: Characiformes, Siluriformes and Perciformes. Moreover, evidence based on molecular data has shown that most of the diversification of the Neotropical ichthyofauna occurred recently. These characteristics make the taxonomy and identification of this fauna a great challenge, even when using molecular approaches. In this context, the present study aimed to test the effectiveness of the barcoding methodology (COI gene) to identify the mega diverse freshwater fish fauna from the Neotropical region. For this purpose, 254 species of fishes were analyzed from the Upper Parana River basin, an area representative of the larger Neotropical region. Results Of the 254 species analyzed, 252 were correctly identified by their barcode sequences (99.2%). The main K2P intra- and inter-specific genetic divergence values (0.3% and 6.8%, respectively) were relatively low compared with similar values reported in the literature, reflecting the higher number of closely related species belonging to a few higher taxa and their recent radiation. Moreover, for 84 pairs of species that showed low levels of genetic divergence (<2%), application of a complementary character-based nucleotide diagnostic approach proved useful in discriminating them. Additionally, 14 species displayed high intra-specific genetic divergence (>2%), pointing to at least 23 strong candidates for new species. Conclusions Our study is the first to examine a large number of freshwater fish species from the Neotropical area, including a large number of closely related species. The results confirmed the efficacy of the barcoding methodology to identify a recently radiated, megadiverse fauna, discriminating 99.2% of the analyzed species. The power of the barcode sequences to identify species, even with low interspecific divergence, gives us an idea of the distribution of inter-specific genetic divergence in these megadiverse fauna. The results also revealed hidden genetic divergences suggestive of reproductive isolation and putative cryptic speciation in some species (23 candidates for new species). Finally, our study constituted an important contribution to the international Barcoding of Life (iBOL.org) project, providing barcode sequences for use in identification of these species by experts and non-experts, and allowing them to be available for use in other applications. PMID:23497346

Molecular Detection of Chilo infuscatellus

PubMed Central

Wang, Weizhong; Wang, Rong; Zheng, Haotian; Gao, Sanji

2017-01-01

Abstract The Chilo infuscatellus (Snellen; Lepidoptera: Pyralidae) is the main pest of sugarcane in China. The shortage of easily distinguishable morphological characters especially in early stage make it challenging to diagnosed and promptly take steps for pest management. In the present study, we described a PCR method for the molecular identification of based on barcode region of COI sequences between C. infuscatellus and four other sugarcane borer species. A 285bp fragment was successfully amplified from all life stages and different geographical populations. Sensitivity tests revealed that diagnostic bands were generated as low as the DNA template concentration of 5 ng/µl. Our work demonstrated a rapid and accurate way for the molecular diagnosis of C. infuscatellus. PMID:29117377

Comprehensive DNA barcoding of the herpetofauna of Germany.

PubMed

Hawlitschek, O; Morinière, J; Dunz, A; Franzen, M; Rödder, D; Glaw, F; Haszprunar, G

2016-01-01

We present the first comprehensive DNA barcoding study of German reptiles and amphibians representing likewise the first on the European herpetofauna. A total of 248 barcodes for all native species and subspecies in the country and a few additional taxa were obtained in the framework of the projects 'Barcoding Fauna Bavarica' (BFB) and 'German Barcode of Life' (GBOL). In contrast to many invertebrate groups, the success rate of the identification of mitochondrial lineages representing species via DNA barcode was almost 100% because no cases of Barcode Index Number (BIN) sharing were detected within German native reptiles and amphibians. However, as expected, a reliable identification of the hybridogenetic species complex in the frog genus Pelophylax was not possible. Deep conspecific lineages resulting in the identification of more than one BIN were found in Lissotriton vulgaris, Natrix natrix and the hybridogenetic Pelophylax complex. A high variety of lineages with different BINs was also found in the barcodes of wall lizards (Podarcis muralis), confirming the existence of many introduced lineages and the frequent occurrence of multiple introductions. Besides the reliable species identification of all life stages and even of tissue remains, our study highlights other potential applications of DNA barcoding concerning German amphibians and reptiles, such as the detection of allochthonous lineages, monitoring of gene flow and also noninvasive sampling via environmental DNA. DNA barcoding based on COI has now proven to be a reliable and efficient tool for studying most amphibians and reptiles as it is already for many other organism groups in zoology. © 2015 John Wiley & Sons Ltd.

Forensically Relevant Blow Flies in Lebanon Survey and Identification Using Molecular Markers (Diptera: Calliphoridae).

PubMed

Shayya, Salman; Debruyne, Régis; Nel, André; Azar, Dany

2018-05-12

Calliphoridae are among the first insects associated to decomposing animal remains. We have collected 1,841 specimens of three calliphorid genera: Calliphora, Lucilia, and Chrysomya, from different Lebanese localities as a first step in implementing a database of insects of forensic relevance for the country. Blow-flies are crucial for the estimation of the postmortem interval. DNA-based identification is a rapid and accurate method, often used for morphologically similar species, especially for immatures or incomplete specimens. In this study, we test the suitability of three genetic markers to identify adults and immature stages of calliphorids, viz., mitochondrial cytochrome c oxidase subunit I (COI) barcode, a region including partial sequences of mitochondrial Cyt-b-tRNAser-ND1, and second internal transcribed spacer (ITS2) region of nuclear ribosomal DNA. Forty Lebanese specimens of various developmental stages (egg, larva, wandering third instar, pupa, newly emerged adult, and mature adult) were identified among the three calliphorid genera: Calliphora, Lucilia, and Chrysomya, and compared with published sequences to confirm their specific assignation. Phylogenetic analyses showed the robustness of ITS2 and COI to identify calliphorids at species level. Nevertheless, ITS2 failed to discriminate Lucilia caesar (Linnaeus) (Diptera, Calliphoridae) from Lucilia illustris (Meigen) (Diptera, Calliphoridae), and COI had a similar issue with Lucilia sericata (Meigen) (Diptera, Calliphoridae) and Lucilia cuprina (Wiedemann) (Diptera, Calliphoridae). Thus, these two markers are complementary. This work contributes new nucleotide sequences for Lebanon. It is a first step in implementing a molecular database of forensic relevant insects for the country.

Diversity of planktonic fish larvae along a latitudinal gradient in the Eastern Atlantic Ocean estimated through DNA barcodes

PubMed Central

Morote, Elvira; Kochzius, Marc; Garcia-Vazquez, Eva

2016-01-01

Mid-trophic pelagic fish are essential components of marine ecosystems because they represent the link between plankton and higher predators. Moreover, they are the basis of the most important fisheries resources; for example, in African waters. In this study, we have sampled pelagic fish larvae in the Eastern Atlantic Ocean along a latitudinal gradient between 37°N and 2°S. We have employed Bongo nets for plankton sampling and sorted visually fish and fish larvae. Using the cytochrome oxidase I gene (COI) as a DNA barcode, we have identified 44 OTUs down to species level that correspond to 14 families, with Myctophidae being the most abundant. A few species were cosmopolitan and others latitude-specific, as was expected. The latitudinal pattern of diversity did not exhibit a temperate-tropical cline; instead, it was likely correlated with environmental conditions with a decline in low-oxygen zones. Importantly, gaps and inconsistencies in reference DNA databases impeded accurate identification to the species level of 49% of the individuals. Fish sampled from tropical latitudes and some orders, such as Perciformes, Myctophiformes and Stomiiformes, were largely unidentified due to incomplete references. Some larvae were identified based on morphology and COI analysis for comparing time and costs employed from each methodology. These results suggest the need of reinforcing DNA barcoding reference datasets of Atlantic bathypelagic tropical fish that, as main prey of top predators, are crucial for ecosystem-based management of fisheries resources. PMID:27761307

DNA barcoding as a useful tool in the systematic study of wild bees of the tribe Augochlorini (Hymenoptera: Halictidae).

PubMed

González-Vaquero, Rocío Ana; Roig-Alsina, Arturo; Packer, Laurence

2016-10-01

Special care is needed in the delimitation and identification of halictid bee species, which are renowned for being morphologically monotonous. Corynura Spinola and Halictillus Moure (Halictidae: Augochlorini) contain species that are key elements in southern South American ecosystems. These bees are very difficult to identify due to close morphological similarity among species and high sexual dimorphism. We analyzed 170 barcode-compliant COI sequences from 19 species. DNA barcodes were useful to confirm gender associations and to detect two new cryptic species. Interspecific distances were significantly higher than those reported for other bees. Maximum intraspecific divergence was less than 1% in 14 species. Barcode index numbers (BINs) were useful to identify putative species that need further study. More than one BIN was assigned to five species. The name Corynura patagonica (Cockerell) probably refers to two cryptic species. The results suggest that Corynura and Halictillus species can be identified using DNA barcodes. The sequences of the species included in this study can be used as a reference to assess the identification of unknown specimens. This study provides additional support for the use of DNA barcodes in bee taxonomy and the identification of specimens, which is particularly relevant in insects of ecological importance such as pollinators.

Machine Learned Replacement of N-Labels for Basecalled Sequences in DNA Barcoding.

PubMed

Ma, Eddie Y T; Ratnasingham, Sujeevan; Kremer, Stefan C

2018-01-01

This study presents a machine learning method that increases the number of identified bases in Sanger Sequencing. The system post-processes a KB basecalled chromatogram. It selects a recoverable subset of N-labels in the KB-called chromatogram to replace with basecalls (A,C,G,T). An N-label correction is defined given an additional read of the same sequence, and a human finished sequence. Corrections are added to the dataset when an alignment determines the additional read and human agree on the identity of the N-label. KB must also rate the replacement with quality value of in the additional read. Corrections are only available during system training. Developing the system, nearly 850,000 N-labels are obtained from Barcode of Life Datasystems, the premier database of genetic markers called DNA Barcodes. Increasing the number of correct bases improves reference sequence reliability, increases sequence identification accuracy, and assures analysis correctness. Keeping with barcoding standards, our system maintains an error rate of percent. Our system only applies corrections when it estimates low rate of error. Tested on this data, our automation selects and recovers: 79 percent of N-labels from COI (animal barcode); 80 percent from matK and rbcL (plant barcodes); and 58 percent from non-protein-coding sequences (across eukaryotes).

Development of a multilocus-based approach for sponge (phylum Porifera) identification: refinement and limitations.

PubMed

Yang, Qi; Franco, Christopher M M; Sorokin, Shirley J; Zhang, Wei

2017-02-02

For sponges (phylum Porifera), there is no reliable molecular protocol available for species identification. To address this gap, we developed a multilocus-based Sponge Identification Protocol (SIP) validated by a sample of 37 sponge species belonging to 10 orders from South Australia. The universal barcode COI mtDNA, 28S rRNA gene (D3-D5), and the nuclear ITS1-5.8S-ITS2 region were evaluated for their suitability and capacity for sponge identification. The highest Bit Score was applied to infer the identity. The reliability of SIP was validated by phylogenetic analysis. The 28S rRNA gene and COI mtDNA performed better than the ITS region in classifying sponges at various taxonomic levels. A major limitation is that the databases are not well populated and possess low diversity, making it difficult to conduct the molecular identification protocol. The identification is also impacted by the accuracy of the morphological classification of the sponges whose sequences have been submitted to the database. Re-examination of the morphological identification further demonstrated and improved the reliability of sponge identification by SIP. Integrated with morphological identification, the multilocus-based SIP offers an improved protocol for more reliable and effective sponge identification, by coupling the accuracy of different DNA markers.

Development of a multilocus-based approach for sponge (phylum Porifera) identification: refinement and limitations

PubMed Central

Yang, Qi; Franco, Christopher M. M.; Sorokin, Shirley J.; Zhang, Wei

2017-01-01

For sponges (phylum Porifera), there is no reliable molecular protocol available for species identification. To address this gap, we developed a multilocus-based Sponge Identification Protocol (SIP) validated by a sample of 37 sponge species belonging to 10 orders from South Australia. The universal barcode COI mtDNA, 28S rRNA gene (D3–D5), and the nuclear ITS1-5.8S-ITS2 region were evaluated for their suitability and capacity for sponge identification. The highest Bit Score was applied to infer the identity. The reliability of SIP was validated by phylogenetic analysis. The 28S rRNA gene and COI mtDNA performed better than the ITS region in classifying sponges at various taxonomic levels. A major limitation is that the databases are not well populated and possess low diversity, making it difficult to conduct the molecular identification protocol. The identification is also impacted by the accuracy of the morphological classification of the sponges whose sequences have been submitted to the database. Re-examination of the morphological identification further demonstrated and improved the reliability of sponge identification by SIP. Integrated with morphological identification, the multilocus-based SIP offers an improved protocol for more reliable and effective sponge identification, by coupling the accuracy of different DNA markers. PMID:28150727

[Identification of original species of Mantidis Oötheca (Sangpiaoxiao) based on DNA barcoding].

PubMed

Wang, Xi; Hou, Fei-xia; Wang, Yi-xuan; Wang, Yu-xian; Li, Jun-de; Yuan, Yuan; Peng, Cheng; Guo, Jin-lin

2015-10-01

Both market research and literature reports both found that the ootheca of mantodea was all used as medicine. However, Chinese Pharmacopoeia only records the ootheca of three mantis species. The clinical use of ootheca unrecorded in Chinese Pharmacopoeia, will pose potential risks to drug safety. It's urgent to identify the origin of Mantidis Oötheca. The current researches about original animal in Mantidis Oötheca are based on morphology and unanimous. DNA barcoding fill gaps of the traditional morphological identification, which is widely used in animal classification studies. This study first use DNA barcoding to analyze genetic distance among different Mantidis Oötheca types, align COI sequences between mantis and Mantidis Oötheca and construct the phylogeny tree. The result confirmed that Tenodera sinensis and Hierodula patellifera were the origin insects of Tuanpiaoxiao and Heipiaoxiao, respectively, and Statilia maculate and Mantis religiosa were the origin insects of Changpiaoxiao.

Reading the Complex Skipper Butterfly Fauna of One Tropical Place

PubMed Central

Janzen, Daniel H.; Hallwachs, Winnie; Burns, John M.; Hajibabaei, Mehrdad; Bertrand, Claudia; Hebert, Paul D. N.

2011-01-01

Background An intense, 30-year, ongoing biodiversity inventory of Lepidoptera, together with their food plants and parasitoids, is centered on the rearing of wild-caught caterpillars in the 120,000 terrestrial hectares of dry, rain, and cloud forest of Area de Conservacion Guanacaste (ACG) in northwestern Costa Rica. Since 2003, DNA barcoding of all species has aided their identification and discovery. We summarize the process and results for a large set of the species of two speciose subfamilies of ACG skipper butterflies (Hesperiidae) and emphasize the effectiveness of barcoding these species (which are often difficult and time-consuming to identify). Methodology/Principal Findings Adults are DNA barcoded by the Biodiversity Institute of Ontario, Guelph, Canada; and they are identified by correlating the resulting COI barcode information with more traditional information such as food plant, facies, genitalia, microlocation within ACG, caterpillar traits, etc. This process has found about 303 morphologically defined species of eudamine and pyrgine Hesperiidae breeding in ACG (about 25% of the ACG butterfly fauna) and another 44 units indicated by distinct barcodes (n = 9,094), which may be additional species and therefore may represent as much as a 13% increase. All but the members of one complex can be identified by their DNA barcodes. Conclusions/Significance Addition of DNA barcoding to the methodology greatly improved the inventory, both through faster (hence cheaper) accurate identification of the species that are distinguishable without barcoding, as well as those that require it, and through the revelation of species “hidden” within what have long been viewed as single species. Barcoding increased the recognition of species-level specialization. It would be no more appropriate to ignore barcode data in a species inventory than it would be to ignore adult genitalia variation or caterpillar ecology. PMID:21857895

Inferring the Provenance of an Alien Species with DNA Barcodes: The Neotropical Butterfly Dryas iulia in Thailand

PubMed Central

Burg, Noah A.; Pradhan, Ashman; Gonzalez, Rebecca M.; Morban, Emely Z.; Zhen, Erica W.; Sakchoowong, Watana; Lohman, David J.

2014-01-01

The Neotropical butterfly Dryas iulia has been collected from several locations in Thailand and Malaysia since 2007, and has been observed breeding in the wild, using introduced Passiflora foetida as a larval host plant. The butterfly is bred by a butterfly house in Phuket, Thailand, for release at weddings and Buddhist ceremonies, and we hypothesized that this butterfly house was the source of wild, Thai individuals. We compared wing patterns and COI barcodes from two, wild Thai populations with individuals obtained from this butterfly house. All Thai individuals resemble the subspecies D. iulia modesta, and barcodes from wild and captive Thai specimens were identical. This unique, Thai barcode was not found in any of the 30 specimens sampled from the wild in the species' native range, but is most similar to specimens from Costa Rica, where many exporting butterfly farms are located. These data implicate the butterfly house as the source of Thailand's wild D. iulia populations, which are currently so widespread that eradication efforts are unlikely to be successful. PMID:25119899

Inferring the provenance of an alien species with DNA barcodes: the neotropical butterfly Dryas iulia in Thailand.

PubMed

Burg, Noah A; Pradhan, Ashman; Gonzalez, Rebecca M; Morban, Emely Z; Zhen, Erica W; Sakchoowong, Watana; Lohman, David J

2014-01-01

The Neotropical butterfly Dryas iulia has been collected from several locations in Thailand and Malaysia since 2007, and has been observed breeding in the wild, using introduced Passiflora foetida as a larval host plant. The butterfly is bred by a butterfly house in Phuket, Thailand, for release at weddings and Buddhist ceremonies, and we hypothesized that this butterfly house was the source of wild, Thai individuals. We compared wing patterns and COI barcodes from two, wild Thai populations with individuals obtained from this butterfly house. All Thai individuals resemble the subspecies D. iulia modesta, and barcodes from wild and captive Thai specimens were identical. This unique, Thai barcode was not found in any of the 30 specimens sampled from the wild in the species' native range, but is most similar to specimens from Costa Rica, where many exporting butterfly farms are located. These data implicate the butterfly house as the source of Thailand's wild D. iulia populations, which are currently so widespread that eradication efforts are unlikely to be successful.

DNA Barcoding Identifies Illegal Parrot Trade.

PubMed

Gonçalves, Priscila F M; Oliveira-Marques, Adriana R; Matsumoto, Tania E; Miyaki, Cristina Y

2015-01-01

Illegal trade threatens the survival of many wild species, and molecular forensics can shed light on various questions raised during the investigation of cases of illegal trade. Among these questions is the identity of the species involved. Here we report a case of a man who was caught in a Brazilian airport trying to travel with 58 avian eggs. He claimed they were quail eggs, but authorities suspected they were from parrots. The embryos never hatched and it was not possible to identify them based on morphology. As 29% of parrot species are endangered, the identity of the species involved was important to establish a stronger criminal case. Thus, we identified the embryos' species based on the analyses of mitochondrial DNA sequences (cytochrome c oxidase subunit I gene [COI] and 16S ribosomal DNA). Embryonic COI sequences were compared with those deposited in BOLD (The Barcode of Life Data System) while their 16S sequences were compared with GenBank sequences. Clustering analysis based on neighbor-joining was also performed using parrot COI and 16S sequences deposited in BOLD and GenBank. The results, based on both genes, indicated that 57 embryos were parrots (Alipiopsitta xanthops, Ara ararauna, and the [Amazona aestiva/A. ochrocephala] complex), and 1 was an owl. This kind of data can help criminal investigations and to design species-specific anti-poaching strategies, and demonstrate how DNA sequence analysis in the identification of bird species is a powerful conservation tool. © The American Genetic Association 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Molecular Approach to the Identification of Fish in the South China Sea

PubMed Central

Zhang, Junbin; Hanner, Robert

2012-01-01

Background DNA barcoding is one means of establishing a rapid, accurate, and cost-effective system for the identification of species. It involves the use of short, standard gene targets to create sequence profiles of known species against sequences of unknowns that can be matched and subsequently identified. The Fish Barcode of Life (FISH-BOL) campaign has the primary goal of gathering DNA barcode records for all the world's fish species. As a contribution to FISH-BOL, we examined the degree to which DNA barcoding can discriminate marine fishes from the South China Sea. Methodology/Principal Findings DNA barcodes of cytochrome oxidase subunit I (COI) were characterized using 1336 specimens that belong to 242 species fishes from the South China Sea. All specimen provenance data (including digital specimen images and geospatial coordinates of collection localities) and collateral sequence information were assembled using Barcode of Life Data System (BOLD; www.barcodinglife.org). Small intraspecific and large interspecific differences create distinct genetic boundaries among most species. In addition, the efficiency of two mitochondrial genes, 16S rRNA (16S) and cytochrome b (cytb), and one nuclear ribosomal gene, 18S rRNA (18S), was also evaluated for a few select groups of species. Conclusions/Significance The present study provides evidence for the effectiveness of DNA barcoding as a tool for monitoring marine biodiversity. Open access data of fishes from the South China Sea can benefit relative applications in ecology and taxonomy. PMID:22363454

Unexpectedly High Levels of Cryptic Diversity Uncovered by a Complete DNA Barcoding of Reptiles of the Socotra Archipelago.

PubMed

Vasconcelos, Raquel; Montero-Mendieta, Santiago; Simó-Riudalbas, Marc; Sindaco, Roberto; Santos, Xavier; Fasola, Mauro; Llorente, Gustavo; Razzetti, Edoardo; Carranza, Salvador

2016-01-01

Few DNA barcoding studies of squamate reptiles have been conducted. Due to the significance of the Socotra Archipelago (a UNESCO Natural World Heritage site and a biodiversity hotspot) and the conservation interest of its reptile fauna (94% endemics), we performed the most comprehensive DNA barcoding study on an island group to date to test its applicability to specimen identification and species discovery. Reptiles constitute Socotra's most important vertebrate fauna, yet their taxonomy remains under-studied. We successfully DNA-barcoded 380 individuals of all 31 presently recognized species. The specimen identification success rate is moderate to high, and almost all species presented local barcoding gaps. The unexpected high levels of intra-specific variability found within some species suggest cryptic diversity. Species richness may be under-estimated by 13.8-54.4%. This has implications in the species' ranges and conservation status that should be considered for conservation planning. Other phylogenetic studies using mitochondrial and nuclear markers are congruent with our results. We conclude that, despite its reduced length (663 base pairs), cytochrome c oxidase 1, COI, is very useful for specimen identification and for detecting intra-specific diversity, and has a good phylogenetic signal. We recommend DNA barcoding to be applied to other biodiversity hotspots for quickly and cost-efficiently flagging species discovery, preferentially incorporated into an integrative taxonomic framework.

Unexpectedly High Levels of Cryptic Diversity Uncovered by a Complete DNA Barcoding of Reptiles of the Socotra Archipelago

PubMed Central

Simó-Riudalbas, Marc; Sindaco, Roberto; Santos, Xavier; Fasola, Mauro; Llorente, Gustavo; Razzetti, Edoardo; Carranza, Salvador

2016-01-01

Few DNA barcoding studies of squamate reptiles have been conducted. Due to the significance of the Socotra Archipelago (a UNESCO Natural World Heritage site and a biodiversity hotspot) and the conservation interest of its reptile fauna (94% endemics), we performed the most comprehensive DNA barcoding study on an island group to date to test its applicability to specimen identification and species discovery. Reptiles constitute Socotra’s most important vertebrate fauna, yet their taxonomy remains under-studied. We successfully DNA-barcoded 380 individuals of all 31 presently recognized species. The specimen identification success rate is moderate to high, and almost all species presented local barcoding gaps. The unexpected high levels of intra-specific variability found within some species suggest cryptic diversity. Species richness may be under-estimated by 13.8–54.4%. This has implications in the species’ ranges and conservation status that should be considered for conservation planning. Other phylogenetic studies using mitochondrial and nuclear markers are congruent with our results. We conclude that, despite its reduced length (663 base pairs), cytochrome c oxidase 1, COI, is very useful for specimen identification and for detecting intra-specific diversity, and has a good phylogenetic signal. We recommend DNA barcoding to be applied to other biodiversity hotspots for quickly and cost-efficiently flagging species discovery, preferentially incorporated into an integrative taxonomic framework. PMID:26930572

Three new species of Rectiostoma Becker, 1982 (Lepidoptera: Gelechioidea: Depressariidae) from Area de Conservación Guanacaste, northwestern Costa Rica

USDA-ARS?s Scientific Manuscript database

We describe three new species of Rectiostoma Becker, 1982 from Costa Rica: R. annemayae Heikkilä and Metz spec. nov., R. eowilsoni Heikkilä and Metz spec. nov. and R. philipmayi Heikkilä and Metz spec. nov. We used a data set of DNA COI-barcodes accumulated for Lepidoptera collected at Area de Conse...

Technical Note: "Mitochondrial and nuclear DNA approaches for reliable identification of Lucilia (Diptera, Calliphoridae) species of forensic interest from Southern Europe".

PubMed

GilArriortua, Maite; Saloña-Bordas, Marta I; Cainé, Laura M; Pinheiro, Fátima; M de Pancorbo, Marian

2015-12-01

In forensic entomology, rapid and unambiguous identification of blowfly species is a critical prerequisite for accurately estimating the post-mortem interval (PMI). The conventional diagnosis of cadaveric entomofauna based on external characters is hampered by the morphological similarities between species, especially in immature stages. Genetic analysis has been shown to allow precise and reliable diagnosis and delimitation of insect species. Nevertheless, the taxonomy of some species remains unresolved. This study was focused on improving the effectiveness and accuracy of analysis based on the widely used cytochrome c oxidase subunit I barcode region (COI barcode, 658 bp), complemented by other mitochondrial and nuclear regions, such as cytochrome b (Cyt-b, 307 bp) and the second internal transcribed spacer (ITS2, 310-331 bp), for the identification of Southern European blowflies. We analyzed a total of 209 specimens, collected from 38 human corpses, belonging to three Calliphoridae genera and seven species: Chrysomya (Ch. albiceps), Calliphora (C. vicina and C. vomitoria), and Lucilia (L. sericata, L. ampullacea, L. caesar and L. illustris). These species are the most common PMI indicators in Portugal. The results revealed that unambiguous separation of species of the Lucilia genus requires different loci from the barcode region. Furthermore, we conclude that the ITS2 (310-331 bp) molecular marker is a promising diagnostic tool because its inter-specific discriminatory power enables unequivocal and consistent distinctions to be made, even between closely related species (L. caesar-L. illustris). This work also contributes new genetic data that may be of interest in performing species diagnosis for Southern European blowflies. Notably, to the best of our knowledge, we provide the first records of the Cyt-b (307 bp) locus for L. illustris and the ITS2 (310-331 bp) region for Iberian Peninsula Lucilia species. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Species-Level Para- and Polyphyly in DNA Barcode Gene Trees: Strong Operational Bias in European Lepidoptera

PubMed Central

Mutanen, Marko; Kivelä, Sami M.; Vos, Rutger A.; Doorenweerd, Camiel; Ratnasingham, Sujeevan; Hausmann, Axel; Huemer, Peter; Dincă, Vlad; van Nieukerken, Erik J.; Lopez-Vaamonde, Carlos; Vila, Roger; Aarvik, Leif; Decaëns, Thibaud; Efetov, Konstantin A.; Hebert, Paul D. N.; Johnsen, Arild; Karsholt, Ole; Pentinsaari, Mikko; Rougerie, Rodolphe; Segerer, Andreas; Tarmann, Gerhard; Zahiri, Reza; Godfray, H. Charles J.

2016-01-01

The proliferation of DNA data is revolutionizing all fields of systematic research. DNA barcode sequences, now available for millions of specimens and several hundred thousand species, are increasingly used in algorithmic species delimitations. This is complicated by occasional incongruences between species and gene genealogies, as indicated by situations where conspecific individuals do not form a monophyletic cluster in a gene tree. In two previous reviews, non-monophyly has been reported as being common in mitochondrial DNA gene trees. We developed a novel web service “Monophylizer” to detect non-monophyly in phylogenetic trees and used it to ascertain the incidence of species non-monophyly in COI (a.k.a. cox1) barcode sequence data from 4977 species and 41,583 specimens of European Lepidoptera, the largest data set of DNA barcodes analyzed from this regard. Particular attention was paid to accurate species identification to ensure data integrity. We investigated the effects of tree-building method, sampling effort, and other methodological issues, all of which can influence estimates of non-monophyly. We found a 12% incidence of non-monophyly, a value significantly lower than that observed in previous studies. Neighbor joining (NJ) and maximum likelihood (ML) methods yielded almost equal numbers of non-monophyletic species, but 24.1% of these cases of non-monophyly were only found by one of these methods. Non-monophyletic species tend to show either low genetic distances to their nearest neighbors or exceptionally high levels of intraspecific variability. Cases of polyphyly in COI trees arising as a result of deep intraspecific divergence are negligible, as the detected cases reflected misidentifications or methodological errors. Taking into consideration variation in sampling effort, we estimate that the true incidence of non-monophyly is ∼23%, but with operational factors still being included. Within the operational factors, we separately assessed the frequency of taxonomic limitations (presence of overlooked cryptic and oversplit species) and identification uncertainties. We observed that operational factors are potentially present in more than half (58.6%) of the detected cases of non-monophyly. Furthermore, we observed that in about 20% of non-monophyletic species and entangled species, the lineages involved are either allopatric or parapatric—conditions where species delimitation is inherently subjective and particularly dependent on the species concept that has been adopted. These observations suggest that species-level non-monophyly in COI gene trees is less common than previously supposed, with many cases reflecting misidentifications, the subjectivity of species delimitation or other operational factors. PMID:27288478

[DNA barcoding and its utility in commonly-used medicinal snakes].

PubMed

Huang, Yong; Zhang, Yue-yun; Zhao, Cheng-jian; Xu, Yong-li; Gu, Ying-le; Huang, Wen-qi; Lin, Kui; Li, Li

2015-03-01

Identification accuracy of traditional Chinese medicine is crucial for the traditional Chinese medicine research, production and application. DNA barcoding based on the mitochondrial gene coding for cytochrome c oxidase subunit I (COI), are more and more used for identification of traditional Chinese medicine. Using universal barcoding primers to sequence, we discussed the feasibility of DNA barcoding method for identification commonly-used medicinal snakes (a total of 109 samples belonging to 19 species 15 genera 6 families). The phylogenetic trees using Neighbor-joining were constructed. The results indicated that the mean content of G + C(46.5%) was lower than that of A + T (53.5%). As calculated by Kimera-2-parameter model, the mean intraspecies genetic distance of Trimeresurus albolabris, Ptyas dhumnades and Lycodon rufozonatus was greater than 2%. Further phylogenetic relationship results suggested that identification of one sample of T. albolabris was erroneous. The identification of some samples of P. dhumnades was also not correct, namely originally P. korros was identified as P. dhumnades. Factors influence on intraspecific genetic distance difference of L. rufozonatus need to be studied further. Therefore, DNA barcoding for identification of medicinal snakes is feasible, and greatly complements the morphological classification method. It is necessary to further study in identification of traditional Chinese medicine.

DNA barcoding and real-time PCR detection of Bactrocera xanthodes (Tephritidae: Diptera) complex.

PubMed

Li, D; Waite, D W; Gunawardana, D N; McCarthy, B; Anderson, D; Flynn, A; George, S

2018-05-06

Immature fruit fly stages of the family Tephritidae are commonly intercepted on breadfruit from Pacific countries at the New Zealand border but are unable to be identified to the species level using morphological characters. Subsequent molecular identification showed that they belong to Bactrocera xanthodes, which is part of a species complex that includes Bactrocera paraxanthodes, Bactrocera neoxanthodes and an undescribed species. To establish a more reliable molecular identification system for B. xanthodes, a reference database of DNA barcode sequences for the 5'-fragment of COI gene region was constructed for B. xanthodes from Fiji, Samoa and Tonga. To better understand the species complex, B. neoxanthodes from Vanuatu and B. paraxanthodes from New Caledonia were also barcoded. Using the results of this analysis, real-time TaqMan polymerase chain reaction (PCR) assays for the detection of B. xanthodes complex and for the three individual species of the complex were developed and validated. The assay showed high specificity for the target species, with no cross-reaction observed for closely related organisms. Each of the real-time PCR assays is sensitive, detecting the target sequences at concentrations as low as ten copies µl-1 and can be used as either singleplex or multiplex formats. This real-time PCR assay for B. xanthodes has been successfully applied at the borders in New Zealand, leading to the rapid identification of intercepted Tephritidae eggs and larvae. The developed assays will be useful biosecurity tools for rapid detection of species in the B. xanthodes complex worldwide.

DNA barcoding of euryglossine bees and the description of new species of Euhesma Michener (Hymenoptera, Colletidae, Euryglossinae)

PubMed Central

Hogendoorn, Katja; Stevens, Mark; Leijs, Remko

2015-01-01

Abstract This paper launches an open access DNA barcoding project “AUSBS” under the Barcoding of Life Datasystems (BOLD). The aims of the project are to help scientists who lack the necessary morphological knowledge to identify known species using molecular markers, to aid native bee specialists with the recognition of species groups that morphologically are difficult to define, and, eventually, to assist with the recognition of new species among known species. Using integrative taxonomy, i.e. morphological comparison to type specimens in Australian museum collections combined with phylogenetic analysis of a fragment of the mitochondrial DNA cytochrome c oxidase subunit I (mtCOI) gene sequences led to the recognition of four new species of Euhesma Michener (Hymenoptera: Colletidae: Euryglossini) collected during intensive surveys in remote Australian conservation areas, which are described. The new species are Euhesma micans, Euhesma lyngouriae, and Euhesma aulaca in a species group associated with Eremophila flowers, and Euhesma albamala in the walkeriana species group. PMID:26448713

Tetrodotoxin detection and species identification of pufferfish in retail roasted fish fillet by DNA barcoding in China.

PubMed

Li, Nan; Shen, Qing; Wang, Jiahui; Han, Chunhui; Ji, Rong; Li, Fengqin; Jiang, Tao

2015-01-01

This study identifies the pufferfish species and detects tetrodotoxin (TTX) in roasted fish fillet samples collected in Beijing, Qingdao and Xiamen, China. The cytochrome c oxidase I (COI) gene was used as the target gene for identification of the pufferfish species in the samples. Enzyme-linked immunosorbent assay (ELISA) screened the TTX levels in samples that had been detected as containing pufferfish by DNA barcode. A total of 125 samples were identified by DNA barcodes; 32 (26%) samples contained pufferfish composition and, among them, 26 (81%) were the highly toxic species Lagocephalus lunaris. All 32 samples containing the pufferfish composition were positive for TTX with levels ranging from 100 to 63,800 ng g(-1). Most of the 32 samples contained the highly toxic L. lunaris. Based on the results, we suggest that the monitoring of roasted fish fillet should be strengthened and the processing procedures should be standardised to minimise TTX poisoning caused by pufferfish.

A review of the Cercyon Leach (Coleoptera, Hydrophilidae, Sphaeridiinae) of the Greater Antilles

PubMed Central

Arriaga-Varela, Emmanuel; Seidel, Matthias; Deler-Hernández, Albert; Viktor Senderov; Fikáček, Martin

2017-01-01

Abstract The representatives of the genus Cercyon Leach occurring in the Greater Antilles are reviewed. Ten species are recorded, of which five are described here as new: C. gimmeli sp. n. (Dominican Republic), C. armatipenis sp. n. (Dominican Republic), C. taino sp. n. (Dominican Republic), C. sklodowskae sp. n. (Jamaica) and C. spiniventris sp. n. (Dominican Republic). Diagnoses and detailed distributional data are also provided for C. floridanus Horn, 1890 (distributed in southeastern United States of America and Cayman Islands), C. insularis Chevrolat, 1863 (endemic to the Antilles) C. praetextatus (Say, 1825) (widely distributed in the New World incl. Greater Antilles), C. quisquilius (Linnaeus, 1761) (an adventive species of Paleartic origin) and C. nigriceps (Marshall, 1802) (an adventive species probably of Oriental origin). Cercyon armatipenis, C. gimmeli, C. taino form a group of closely related species only distinguishable by male genitalia and DNA sequences. A key to the Great Antillean Cercyon is provided and important diagnostic characters are illustrated. The larvae of C. insularis and C. taino were associated with adults using COI barcode sequences, illustrated and diagnosed. Full occurrence data, additional images and COI barcode sequences were submitted to open access on-line depositories in an effort to provide access to complete data. PMID:28769720

Exploring Genetic Divergence in a Species-Rich Insect Genus Using 2790 DNA Barcodes

PubMed Central

Lin, Xiaolong; Stur, Elisabeth; Ekrem, Torbjørn

2015-01-01

DNA barcoding using a fragment of the mitochondrial cytochrome c oxidase subunit 1 gene (COI) has proven to be successful for species-level identification in many animal groups. However, most studies have been focused on relatively small datasets or on large datasets of taxonomically high-ranked groups. We explore the quality of DNA barcodes to delimit species in the diverse chironomid genus Tanytarsus (Diptera: Chironomidae) by using different analytical tools. The genus Tanytarsus is the most species-rich taxon of tribe Tanytarsini (Diptera: Chironomidae) with more than 400 species worldwide, some of which can be notoriously difficult to identify to species-level using morphology. Our dataset, based on sequences generated from own material and publicly available data in BOLD, consist of 2790 DNA barcodes with a fragment length of at least 500 base pairs. A neighbor joining tree of this dataset comprises 131 well separated clusters representing 121 morphological species of Tanytarsus: 77 named, 16 unnamed and 28 unidentified theoretical species. For our geographically widespread dataset, DNA barcodes unambiguously discriminate 94.6% of the Tanytarsus species recognized through prior morphological study. Deep intraspecific divergences exist in some species complexes, and need further taxonomic studies using appropriate nuclear markers as well as morphological and ecological data to be resolved. The DNA barcodes cluster into 120–242 molecular operational taxonomic units (OTUs) depending on whether Objective Clustering, Automatic Barcode Gap Discovery (ABGD), Generalized Mixed Yule Coalescent model (GMYC), Poisson Tree Process (PTP), subjective evaluation of the neighbor joining tree or Barcode Index Numbers (BINs) are used. We suggest that a 4–5% threshold is appropriate to delineate species of Tanytarsus non-biting midges. PMID:26406595

A Multilocus Species Delimitation Reveals a Striking Number of Species of Coralline Algae Forming Maerl in the OSPAR Maritime Area

PubMed Central

Pardo, Cristina; Lopez, Lua; Peña, Viviana; Hernández-Kantún, Jazmin; Le Gall, Line; Bárbara, Ignacio; Barreiro, Rodolfo

2014-01-01

Maerl beds are sensitive biogenic habitats built by an accumulation of loose-lying, non-geniculate coralline algae. While these habitats are considered hot-spots of marine biodiversity, the number and distribution of maerl-forming species is uncertain because homoplasy and plasticity of morphological characters are common. As a result, species discrimination based on morphological features is notoriously challenging, making these coralline algae the ideal candidates for a DNA barcoding study. Here, mitochondrial (COI-5P DNA barcode fragment) and plastidial (psbA gene) sequence data were used in a two-step approach to delimit species in 224 collections of maerl sampled from Svalbard (78°96’N) to the Canary Islands (28°64’N) that represented 10 morphospecies from four genera and two families. First, the COI-5P dataset was analyzed with two methods based on distinct criteria (ABGD and GMYC) to delineate 16 primary species hypotheses (PSHs) arranged into four major lineages. Second, chloroplast (psbA) sequence data served to consolidate these PSHs into 13 secondary species hypotheses (SSHs) that showed biologically plausible ranges. Using several lines of evidence (e.g. morphological characters, known species distributions, sequences from type and topotype material), six SSHs were assigned to available species names that included the geographically widespread Phymatolithon calcareum, Lithothamnion corallioides, and L. glaciale; possible identities of other SSHs are discussed. Concordance between SSHs and morphospecies was minimal, highlighting the convenience of DNA barcoding for an accurate identification of maerl specimens. Our survey indicated that a majority of maerl forming species have small distribution ranges and revealed a gradual replacement of species with latitude. PMID:25111057

A new record of ponyfish Deveximentum megalolepis (Perciformes: Leiognathidae) in Beibu Gulf of China

NASA Astrophysics Data System (ADS)

Ju, Yuman; Song, Na; Chen, Guobao; Sun, Dianrong; Han, Zhiqiang; Gao, Tianxiang

2017-06-01

A new record ponyfish, Deveximentum megalolepis Mochizuki and Hayashi, 1989, was documented based on its morphological characteristics and DNA barcode. Fifty specimens were collected from Beibu Gulf of China and identified as D. megalolepis by morphological characterization. The coloration, meristic traits, and morphometric measurements were consistent with previously published records. In general, it is a silver-white, laterally compressed and deep bodied ponyfish with 6-9 rows of scales on cheek; scale rows above lateral line 6-8; scale rows below lateral line 14-17. Mitochondrial cytochrome oxidase I subunit (COI) gene fragment was sequenced for phylogenetic analysis. There is no sequence variation of COI gene between the specimens collected in this study. The genetic distances between D. megalolepis and other congeneric species range from 3.6% to 14.0%, which were greater than the threshold for fish species delimitation. The COI sequence analysis also supported the validity of D. megalolepis at genetic level. However, the genetic distance between Chinese and Philippine individuals was about 1.2% and they formed two lineages in gene tree, which may be caused by the geographical distance.

Three new brackish-water thalassocypridine species (Crustacea: Ostracoda: Paracyprididae) from the Ryukyu Islands, southwestern Japan.

PubMed

Hiruta, Shimpei F; Kakui, Keiichi

2016-09-21

We describe three new species of brackish-water ostracods representing two genera in the ostracod tribe Thalassocypridini from mangrove forests in the Ryukyu Islands, subtropical southwestern Japan, and provide their barcoding sequences for the mitochondrial cytochrome c oxidase subunit I (COI) gene. Mangalocypria ryukyuensis sp. nov. was found on Okinawa Island. We also found a Mangalocypria population on Ishigaki Island that was morphologically identical to M. ryukyuensis on Okinawa, but an individual differed by 4.7% in COI sequence (K2P distance) from an individual from Okinawa. This is the first record for Japan of a species in Mangalocypria. Paracypria longiseta sp. nov., obtained from Okinawa Island, is similar to Pontoparta hartmanni. Paracypria plumosa sp. nov. from Ishigaki Island is similar to Pa. adnata described from Yakushima Island, Japan. The COI genetic distance between individuals of Pa. longiseta and Pa. plumosa was roughly as large as that between either of these species and individuals in the Mangalocypria populations. Our study underscores that genera in Thalassocypridini may not represent natural groups, and that this tribe needs taxonomic revision.

A DNA microarray for identification of selected Korean birds based on mitochondrial cytochrome c oxidase I gene sequences.

PubMed

Chung, In-Hyuk; Yoo, Hye Sook; Eah, Jae-Yong; Yoon, Hyun-Kyu; Jung, Jin-Wook; Hwang, Seung Yong; Kim, Chang-Bae

2010-10-01

DNA barcoding with the gene encoding cytochrome c oxidase I (COI) in the mitochondrial genome has been proposed as a standard marker to identify and discover animal species. Some migratory wild birds are suspected of transmitting avian influenza and pose a threat to aircraft safety because of bird strikes. We have previously reported the COI gene sequences of 92 Korean bird species. In the present study, we developed a DNA microarray to identify 17 selected bird species on the basis of nucleotide diversity. We designed and synthesized 19 specific oligonucleotide probes; these probes were arrayed on a silylated glass slide. The length of the probes was 19-24 bps. The COI sequences amplified from the tissues of the selected birds were labeled with a fluorescent probe for microarray hybridization, and unique hybridization patterns were detected for each selected species. These patterns may be considered diagnostic patterns for species identification. This microarray system will provide a sensitive and a high-throughput method for identification of Korean birds.

Starting a DNA barcode reference library for shallow water polychaetes from the southern European Atlantic coast.

PubMed

Lobo, Jorge; Teixeira, Marcos A L; Borges, Luisa M S; Ferreira, Maria S G; Hollatz, Claudia; Gomes, Pedro T; Sousa, Ronaldo; Ravara, Ascensão; Costa, Maria H; Costa, Filipe O

2016-01-01

Annelid polychaetes have been seldom the focus of dedicated DNA barcoding studies, despite their ecological relevance and often dominance, particularly in soft-bottom estuarine and coastal marine ecosystems. Here, we report the first assessment of the performance of DNA barcodes in the discrimination of shallow water polychaete species from the southern European Atlantic coast, focusing on specimens collected in estuaries and coastal ecosystems of Portugal. We analysed cytochrome oxidase I DNA barcodes (COI-5P) from 164 specimens, which were assigned to 51 morphospecies. To our data set from Portugal, we added available published sequences selected from the same species, genus or family, to inspect for taxonomic congruence among studies and collection location. The final data set comprised 290 specimens and 79 morphospecies, which generated 99 Barcode Index Numbers (BINs) within Barcode of Life Data Systems (BOLD). Among these, 22 BINs were singletons, 47 other BINs were concordant, confirming the initial identification based on morphological characters, and 30 were discordant, most of which consisted on multiple BINs found for the same morphospecies. Some of the most prominent cases in the latter category include Hediste diversicolor (O.F. Müller, 1776) (7), Eulalia viridis (Linnaeus, 1767) (2) and Owenia fusiformis (delle Chiaje, 1844) (5), all of them reported from Portugal and frequently used in ecological studies as environmental quality indicators. Our results for these species showed discordance between molecular lineages and morphospecies, or added additional relatively divergent lineages. The potential inaccuracies in environmental assessments, where underpinning polychaete species diversity is poorly resolved or clarified, demand additional and extensive investigation of the DNA barcode diversity in this group, in parallel with alpha taxonomy efforts. © 2015 John Wiley & Sons Ltd.

Mitochondrial heteroplasmy and DNA barcoding in Hawaiian Hylaeus (Nesoprosopis) bees (Hymenoptera: Colletidae).

PubMed

Magnacca, Karl N; Brown, Mark J F

2010-06-11

The past several years have seen a flurry of papers seeking to clarify the utility and limits of DNA barcoding, particularly in areas such as species discovery and paralogy due to nuclear pseudogenes. Heteroplasmy, the coexistence of multiple mitochondrial haplotypes in a single organism, has been cited as a potentially serious problem for DNA barcoding but its effect on identification accuracy has not been tested. In addition, few studies of barcoding have tested a large group of closely-related species with a well-established morphological taxonomy. In this study we examine both of these issues, by densely sampling the Hawaiian Hylaeus bee radiation. Individuals from 21 of the 49 a priori morphologically-defined species exhibited coding sequence heteroplasmy at levels of 1-6% or more. All homoplasmic species were successfully identified by COI using standard methods of analysis, but only 71% of heteroplasmic species. The success rate in identifying heteroplasmic species was increased to 86% by treating polymorphisms as character states rather than ambiguities. Nuclear pseudogenes (numts) were also present in four species, and were distinguishable from heteroplasmic sequences by patterns of nucleotide and amino acid change. Heteroplasmy significantly decreased the reliability of species identification. In addition, the practical issue of dealing with large numbers of polymorphisms- and resulting increased time and labor required - makes the development of DNA barcode databases considerably more complex than has previously been suggested. The impact of heteroplasmy on the utility of DNA barcoding as a bulk specimen identification tool will depend upon its frequency across populations, which remains unknown. However, DNA barcoding is still likely to remain an important identification tool for those species that are difficult or impossible to identify through morphology, as is the case for the ecologically important solitary bee fauna.

Mitochondrial heteroplasmy and DNA barcoding in Hawaiian Hylaeus (Nesoprosopis) bees (Hymenoptera: Colletidae)

PubMed Central

2010-01-01

Background The past several years have seen a flurry of papers seeking to clarify the utility and limits of DNA barcoding, particularly in areas such as species discovery and paralogy due to nuclear pseudogenes. Heteroplasmy, the coexistence of multiple mitochondrial haplotypes in a single organism, has been cited as a potentially serious problem for DNA barcoding but its effect on identification accuracy has not been tested. In addition, few studies of barcoding have tested a large group of closely-related species with a well-established morphological taxonomy. In this study we examine both of these issues, by densely sampling the Hawaiian Hylaeus bee radiation. Results Individuals from 21 of the 49 a priori morphologically-defined species exhibited coding sequence heteroplasmy at levels of 1-6% or more. All homoplasmic species were successfully identified by COI using standard methods of analysis, but only 71% of heteroplasmic species. The success rate in identifying heteroplasmic species was increased to 86% by treating polymorphisms as character states rather than ambiguities. Nuclear pseudogenes (numts) were also present in four species, and were distinguishable from heteroplasmic sequences by patterns of nucleotide and amino acid change. Conclusions Heteroplasmy significantly decreased the reliability of species identification. In addition, the practical issue of dealing with large numbers of polymorphisms- and resulting increased time and labor required - makes the development of DNA barcode databases considerably more complex than has previously been suggested. The impact of heteroplasmy on the utility of DNA barcoding as a bulk specimen identification tool will depend upon its frequency across populations, which remains unknown. However, DNA barcoding is still likely to remain an important identification tool for those species that are difficult or impossible to identify through morphology, as is the case for the ecologically important solitary bee fauna. PMID:20540728

Species-Level Para- and Polyphyly in DNA Barcode Gene Trees: Strong Operational Bias in European Lepidoptera.

PubMed

Mutanen, Marko; Kivelä, Sami M; Vos, Rutger A; Doorenweerd, Camiel; Ratnasingham, Sujeevan; Hausmann, Axel; Huemer, Peter; Dincă, Vlad; van Nieukerken, Erik J; Lopez-Vaamonde, Carlos; Vila, Roger; Aarvik, Leif; Decaëns, Thibaud; Efetov, Konstantin A; Hebert, Paul D N; Johnsen, Arild; Karsholt, Ole; Pentinsaari, Mikko; Rougerie, Rodolphe; Segerer, Andreas; Tarmann, Gerhard; Zahiri, Reza; Godfray, H Charles J

2016-11-01

The proliferation of DNA data is revolutionizing all fields of systematic research. DNA barcode sequences, now available for millions of specimens and several hundred thousand species, are increasingly used in algorithmic species delimitations. This is complicated by occasional incongruences between species and gene genealogies, as indicated by situations where conspecific individuals do not form a monophyletic cluster in a gene tree. In two previous reviews, non-monophyly has been reported as being common in mitochondrial DNA gene trees. We developed a novel web service "Monophylizer" to detect non-monophyly in phylogenetic trees and used it to ascertain the incidence of species non-monophyly in COI (a.k.a. cox1) barcode sequence data from 4977 species and 41,583 specimens of European Lepidoptera, the largest data set of DNA barcodes analyzed from this regard. Particular attention was paid to accurate species identification to ensure data integrity. We investigated the effects of tree-building method, sampling effort, and other methodological issues, all of which can influence estimates of non-monophyly. We found a 12% incidence of non-monophyly, a value significantly lower than that observed in previous studies. Neighbor joining (NJ) and maximum likelihood (ML) methods yielded almost equal numbers of non-monophyletic species, but 24.1% of these cases of non-monophyly were only found by one of these methods. Non-monophyletic species tend to show either low genetic distances to their nearest neighbors or exceptionally high levels of intraspecific variability. Cases of polyphyly in COI trees arising as a result of deep intraspecific divergence are negligible, as the detected cases reflected misidentifications or methodological errors. Taking into consideration variation in sampling effort, we estimate that the true incidence of non-monophyly is ∼23%, but with operational factors still being included. Within the operational factors, we separately assessed the frequency of taxonomic limitations (presence of overlooked cryptic and oversplit species) and identification uncertainties. We observed that operational factors are potentially present in more than half (58.6%) of the detected cases of non-monophyly. Furthermore, we observed that in about 20% of non-monophyletic species and entangled species, the lineages involved are either allopatric or parapatric-conditions where species delimitation is inherently subjective and particularly dependent on the species concept that has been adopted. These observations suggest that species-level non-monophyly in COI gene trees is less common than previously supposed, with many cases reflecting misidentifications, the subjectivity of species delimitation or other operational factors. © The Author(s) 2016. Published by Oxford University Press, on behalf of the Society of Systematic Biologists.

Species From Feces: Order-Wide Identification of Chiroptera From Guano and Other Non-Invasive Genetic Samples

PubMed Central

Williamson, Charles H. D.; Sanchez, Daniel E.; Sobek, Colin J.; Chambers, Carol L.

2016-01-01

Bat guano is a relatively untapped reservoir of information, having great utility as a DNA source because it is often available at roosts even when bats are not and is an easy type of sample to collect from a difficult-to-study mammalian order. Recent advances from microbial community studies in primer design, sequencing, and analysis enable fast, accurate, and cost-effective species identification. Here, we borrow from this discipline to develop an order-wide DNA mini-barcode assay (Species from Feces) based on a segment of the mitochondrial gene cytochrome c oxidase I (COI). The assay works effectively with fecal DNA and is conveniently transferable to low-cost, high-throughput Illumina MiSeq technology that also allows simultaneous pairing with other markers. Our PCR primers target a region of COI that is highly discriminatory among Chiroptera (92% species-level identification of barcoded species), and are sufficiently degenerate to allow hybridization across diverse bat taxa. We successfully validated our system with 54 bat species across both suborders. Despite abundant arthropod prey DNA in guano, our primers were highly specific to bats; no arthropod DNA was detected in thousands of feces run on Sanger and Illumina platforms. The assay is extendable to fecal pellets of unknown age as well as individual and pooled guano, to allow for individual (using singular fecal pellets) and community (using combined pellets collected from across long-term roost sites) analyses. We developed a searchable database (http://nau.edu/CEFNS/Forestry/Research/Bats/Search-Tool/) that allows users to determine the discriminatory capability of our markers for bat species of interest. Our assay has applications worldwide for examining disease impacts on vulnerable species, determining species assemblages within roosts, and assessing the presence of bat species that are vulnerable or facing extinction. The development and analytical pathways are rapid, reliable, and inexpensive, and can be applied to ecology and conservation studies of other taxa. PMID:27654850

Species From Feces: Order-Wide Identification of Chiroptera From Guano and Other Non-Invasive Genetic Samples.

PubMed

Walker, Faith M; Williamson, Charles H D; Sanchez, Daniel E; Sobek, Colin J; Chambers, Carol L

Bat guano is a relatively untapped reservoir of information, having great utility as a DNA source because it is often available at roosts even when bats are not and is an easy type of sample to collect from a difficult-to-study mammalian order. Recent advances from microbial community studies in primer design, sequencing, and analysis enable fast, accurate, and cost-effective species identification. Here, we borrow from this discipline to develop an order-wide DNA mini-barcode assay (Species from Feces) based on a segment of the mitochondrial gene cytochrome c oxidase I (COI). The assay works effectively with fecal DNA and is conveniently transferable to low-cost, high-throughput Illumina MiSeq technology that also allows simultaneous pairing with other markers. Our PCR primers target a region of COI that is highly discriminatory among Chiroptera (92% species-level identification of barcoded species), and are sufficiently degenerate to allow hybridization across diverse bat taxa. We successfully validated our system with 54 bat species across both suborders. Despite abundant arthropod prey DNA in guano, our primers were highly specific to bats; no arthropod DNA was detected in thousands of feces run on Sanger and Illumina platforms. The assay is extendable to fecal pellets of unknown age as well as individual and pooled guano, to allow for individual (using singular fecal pellets) and community (using combined pellets collected from across long-term roost sites) analyses. We developed a searchable database (http://nau.edu/CEFNS/Forestry/Research/Bats/Search-Tool/) that allows users to determine the discriminatory capability of our markers for bat species of interest. Our assay has applications worldwide for examining disease impacts on vulnerable species, determining species assemblages within roosts, and assessing the presence of bat species that are vulnerable or facing extinction. The development and analytical pathways are rapid, reliable, and inexpensive, and can be applied to ecology and conservation studies of other taxa.

New taxonomy and old collections: integrating DNA barcoding into the collection curation process.

PubMed

Puillandre, N; Bouchet, P; Boisselier-Dubayle, M-C; Brisset, J; Buge, B; Castelin, M; Chagnoux, S; Christophe, T; Corbari, L; Lambourdière, J; Lozouet, P; Marani, G; Rivasseau, A; Silva, N; Terryn, Y; Tillier, S; Utge, J; Samadi, S

2012-05-01

Because they house large biodiversity collections and are also research centres with sequencing facilities, natural history museums are well placed to develop DNA barcoding best practices. The main difficulty is generally the vouchering system: it must ensure that all data produced remain attached to the corresponding specimen, from the field to publication in articles and online databases. The Museum National d'Histoire Naturelle in Paris is one of the leading laboratories in the Marine Barcode of Life (MarBOL) project, which was used as a pilot programme to include barcode collections for marine molluscs and crustaceans. The system is based on two relational databases. The first one classically records the data (locality and identification) attached to the specimens. In the second one, tissue-clippings, DNA extractions (both preserved in 2D barcode tubes) and PCR data (including primers) are linked to the corresponding specimen. All the steps of the process [sampling event, specimen identification, molecular processing, data submission to Barcode Of Life Database (BOLD) and GenBank] are thus linked together. Furthermore, we have developed several web-based tools to automatically upload data into the system, control the quality of the sequences produced and facilitate the submission to online databases. This work is the result of a joint effort from several teams in the Museum National d'Histoire Naturelle (MNHN), but also from a collaborative network of taxonomists and molecular systematists outside the museum, resulting in the vouchering so far of ∼41,000 sequences and the production of ∼11,000 COI sequences. © 2012 Blackwell Publishing Ltd.

Taxonomy 2.0: Sequencing of old type specimens supports the description of two new species of the Lasiocampa decolorata group from Morocco (Lepidoptera, Lasiocampidae).

PubMed

Speidel, Wolfgang; Hausmann, Axel; Müller, Günter C; Kravchenko, Vasiliy; Mooser, Josef; Witt, Thomas J; Khallaayoune, Khalid; Prosser, Sean; Hebert, Paul D N

2015-08-11

The type of Lasiocampa decolorata (KLUG, 1830), collected in 1820, was successfully barcoded to generate a 658bp COI-fragment after 194 years. The resulting molecular data allowed the description of two closely related species from Morocco: Lasiocampa hannae SPEIDEL, MOOSER & WITT sp. n. from the Anti Atlas and Lasiocampa editae SPEIDEL, MOOSER & WITT sp. n. from the High Atlas.

DNA barcoding reveals that the common cupped oyster in Taiwan is the Portuguese oyster Crassostrea angulata (Ostreoida; Ostreidae), not C. gigas

NASA Astrophysics Data System (ADS)

Hsiao, Sheng-Tai; Chuang, Shin-Chang; Chen, Kao-Sung; Ho, Ping-Ho; Wu, Chi-Lun; Chen, Chaolun Allen

2016-09-01

The Pacific cupped oyster, Crassostrea gigas, is one of the major aquacultural shellfish species that has been introduced to Europe and America from its native source in the West Pacific. In Taiwan, the cultivated cupped oysters along the west coast have been identified as C. gigas for over centuries; however, several molecular phylogenetic studies have cast doubt upon the existence of this species in Taiwan and adjacent waters. Indeed, our analyses of mitochondrial cytochrome oxidase I (COI) sequences from 313 Crassostrea collected from 12 locations along Taiwanese and southern Chinese coastlines confirm that all samples were the Portuguese oyster, C. angulata, rather than C. gigas. Multiple lines of evidence, including haplotypic and nucleotide diversity of the COI gene, demographic history, and population genetics, suggest that Taiwanese C. angulata is unique, probably experienced a sudden population expansion after the Last Glacial Maxima around 20,000 years ago, and has a significantly limited genetic connectivity across the Taiwan Strait. Our study applies an extended sampling and DNA barcoding to confirm the absence of C. gigas in natural and cultivated populations in Taiwan and southern China, where we only found C. angulata. We highlight the importance of conserving the gene pool of the C. angulata population in Taiwan, particularly considering the current threats by large-scale environmental disturbances such as marine pollution, habitat destruction, and climate change.

DNA barcoding reveals that the common cupped oyster in Taiwan is the Portuguese oyster Crassostrea angulata (Ostreoida; Ostreidae), not C. gigas

PubMed Central

Hsiao, Sheng-Tai; Chuang, Shin-Chang; Chen, Kao-Sung; Ho, Ping-Ho; Wu, Chi-Lun; Chen, Chaolun Allen

2016-01-01

The Pacific cupped oyster, Crassostrea gigas, is one of the major aquacultural shellfish species that has been introduced to Europe and America from its native source in the West Pacific. In Taiwan, the cultivated cupped oysters along the west coast have been identified as C. gigas for over centuries; however, several molecular phylogenetic studies have cast doubt upon the existence of this species in Taiwan and adjacent waters. Indeed, our analyses of mitochondrial cytochrome oxidase I (COI) sequences from 313 Crassostrea collected from 12 locations along Taiwanese and southern Chinese coastlines confirm that all samples were the Portuguese oyster, C. angulata, rather than C. gigas. Multiple lines of evidence, including haplotypic and nucleotide diversity of the COI gene, demographic history, and population genetics, suggest that Taiwanese C. angulata is unique, probably experienced a sudden population expansion after the Last Glacial Maxima around 20,000 years ago, and has a significantly limited genetic connectivity across the Taiwan Strait. Our study applies an extended sampling and DNA barcoding to confirm the absence of C. gigas in natural and cultivated populations in Taiwan and southern China, where we only found C. angulata. We highlight the importance of conserving the gene pool of the C. angulata population in Taiwan, particularly considering the current threats by large-scale environmental disturbances such as marine pollution, habitat destruction, and climate change. PMID:27666088

DNA barcoding reveals that the common cupped oyster in Taiwan is the Portuguese oyster Crassostrea angulata (Ostreoida; Ostreidae), not C. gigas.

PubMed

Hsiao, Sheng-Tai; Chuang, Shin-Chang; Chen, Kao-Sung; Ho, Ping-Ho; Wu, Chi-Lun; Chen, Chaolun Allen

2016-09-26

The Pacific cupped oyster, Crassostrea gigas, is one of the major aquacultural shellfish species that has been introduced to Europe and America from its native source in the West Pacific. In Taiwan, the cultivated cupped oysters along the west coast have been identified as C. gigas for over centuries; however, several molecular phylogenetic studies have cast doubt upon the existence of this species in Taiwan and adjacent waters. Indeed, our analyses of mitochondrial cytochrome oxidase I (COI) sequences from 313 Crassostrea collected from 12 locations along Taiwanese and southern Chinese coastlines confirm that all samples were the Portuguese oyster, C. angulata, rather than C. gigas. Multiple lines of evidence, including haplotypic and nucleotide diversity of the COI gene, demographic history, and population genetics, suggest that Taiwanese C. angulata is unique, probably experienced a sudden population expansion after the Last Glacial Maxima around 20,000 years ago, and has a significantly limited genetic connectivity across the Taiwan Strait. Our study applies an extended sampling and DNA barcoding to confirm the absence of C. gigas in natural and cultivated populations in Taiwan and southern China, where we only found C. angulata. We highlight the importance of conserving the gene pool of the C. angulata population in Taiwan, particularly considering the current threats by large-scale environmental disturbances such as marine pollution, habitat destruction, and climate change.

New identification of the moray eel Gymnothorax minor (Temminck & Schlegel, 1846) in China (Anguilliformes, Muraenidae).

PubMed

Li, Yuan; Zhang, Liyan; Zhao, Linlin; Feng, Ji; Loh, Karhoe; Zheng, Xinqing; Lin, Longshan

2018-01-01

A new identification of Gymnothorax minor (Temminck & Schlegel, 1846) is documented based on morphological characteristics and DNA barcoding. Sixty-one individuals of G. minor were collected from the East China Sea and the South China Sea. This species was previously reported as Gymnothorax reticularis Bloch, 1795 in China because of the similarity in external shape and color. Gymnothorax minor can be easily distinguished from G. reticularis by its color pattern of 18-20 irregular dark brown vertical bars and the body having scattered small brown spots. Additionally, the teeth are uniserial on both jaws, and the vertebrae number 137-139. By combining congener sequences of the cytochrome oxidase I (COI) gene from GenBank, two groups were detected among all the COI sequences of the currently named G. minor , which further indicated that two valid species were present based on genetic distance. A divergence also occurred on the number of vertebrae between the northern and southern populations. The phylogenetic and morphological analysis strongly supports that the northern and southern populations of G. minor are two different species. Furthermore, the distribution area of the northern G. minor has expanded southward to 5°15'N in the South China Sea. More specimens of G. minor and G. reticularis are crucial in order to define their geographical distribution boundaries and provide the correct DNA barcoding.

PCR cycles above routine numbers do not compromise high-throughput DNA barcoding results.

PubMed

Vierna, J; Doña, J; Vizcaíno, A; Serrano, D; Jovani, R

2017-10-01

High-throughput DNA barcoding has become essential in ecology and evolution, but some technical questions still remain. Increasing the number of PCR cycles above the routine 20-30 cycles is a common practice when working with old-type specimens, which provide little amounts of DNA, or when facing annealing issues with the primers. However, increasing the number of cycles can raise the number of artificial mutations due to polymerase errors. In this work, we sequenced 20 COI libraries in the Illumina MiSeq platform. Libraries were prepared with 40, 45, 50, 55, and 60 PCR cycles from four individuals belonging to four species of four genera of cephalopods. We found no relationship between the number of PCR cycles and the number of mutations despite using a nonproofreading polymerase. Moreover, even when using a high number of PCR cycles, the resulting number of mutations was low enough not to be an issue in the context of high-throughput DNA barcoding (but may still remain an issue in DNA metabarcoding due to chimera formation). We conclude that the common practice of increasing the number of PCR cycles should not negatively impact the outcome of a high-throughput DNA barcoding study in terms of the occurrence of point mutations.

Systematics of spiny predatory katydids (Tettigoniidae: Listroscelidinae) from the Brazilian Atlantic Forest based on morphology and molecular data.

PubMed

Fialho, Verônica Saraiva; Chamorro-Rengifo, Juliana; Lopes-Andrade, Cristiano; Yotoko, Karla Suemy Clemente

2014-01-01

Listroscelidinae (Orthoptera: Tettigoniidae) are insectivorous Pantropical katydids whose taxonomy presents a long history of controversy, with several genera incertae sedis. This work focused on species occurring in the Brazilian Atlantic Forest, one of the world's most threatened biomes. We examined material deposited in scientific collections and visited 15 conservation units from Rio de Janeiro to southern Bahia between November 2011 and January 2012, catching 104 specimens from 10 conservation units. Based on morphological and molecular data we redefined Listroscelidini, adding a new tribe, new genus and eight new species to the subfamily. Using morphological analysis, we redescribed and added new geographic records for six species, synonymized two species and built a provisional identification key for the Atlantic Forest Listroscelidinae. Molecular results suggest two new species and a new genus to be described, possibly by the fission of the genus Hamayulus. We also proposed a 500 bp region in the final portion of the COI to be used as a molecular barcode. Our data suggest that the Atlantic Forest Listroscelidinae are seriously endangered, because they occur in highly preserved forest remnants, show high rates of endemism and have a narrow geographic distribution. Based on our results, we suggest future collection efforts must take into account the molecular barcode data to accelerate species recognition.

DNA barcoding of Mobulid Ray Gill Rakers for Implementing CITES on Elasmobranch in China

PubMed Central

Zeng, Yan; Wu, Zhongze; Zhang, Chunguang; Meng, Zhibin; Jiang, Zhigang; Zhang, Jie

2016-01-01

The Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES) has been counted on for conserving threatened marine fish since it regulates the commercial international trade of these species. Implementation of the international treaty for Mantas included on CITES Appendix II is challenging due to insufficient information on species identification and markets management. To fill the gap in such aspects, we identified five species of Mobulid rays (Mobula spps. and Manta spp) by using COI and NADH2 mtDNA markers in dried ray gill rakers from Chinese markets, namely, Mobula japonica (representing 54.8% of the sample set), M. tarapacana (14.4%), M. kuhlii (13.3%), M. thurstoni (6.4%), along with Manta birostris (11.2%; CITES Appendix II). The utilization and conservation statuses of these species were discussed. Based on combination of DNA barcodes and key morphological characters, we developed a three-step process for identifying the gill rakers of Mobulid rays which has been adopted by frontline enforcement in China. We hope that our work can serve as a foundation and basis to reinforce objectives of international treaties, regulation of consumer-driven markets, regional cooperation, and national fishery management on endangered elasmobranchs in China as well as related countries. PMID:27876850

DNA barcoding of Mobulid Ray Gill Rakers for Implementing CITES on Elasmobranch in China.

PubMed

Zeng, Yan; Wu, Zhongze; Zhang, Chunguang; Meng, Zhibin; Jiang, Zhigang; Zhang, Jie

2016-11-23

The Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES) has been counted on for conserving threatened marine fish since it regulates the commercial international trade of these species. Implementation of the international treaty for Mantas included on CITES Appendix II is challenging due to insufficient information on species identification and markets management. To fill the gap in such aspects, we identified five species of Mobulid rays (Mobula spps. and Manta spp) by using COI and NADH2 mtDNA markers in dried ray gill rakers from Chinese markets, namely, Mobula japonica (representing 54.8% of the sample set), M. tarapacana (14.4%), M. kuhlii (13.3%), M. thurstoni (6.4%), along with Manta birostris (11.2%; CITES Appendix II). The utilization and conservation statuses of these species were discussed. Based on combination of DNA barcodes and key morphological characters, we developed a three-step process for identifying the gill rakers of Mobulid rays which has been adopted by frontline enforcement in China. We hope that our work can serve as a foundation and basis to reinforce objectives of international treaties, regulation of consumer-driven markets, regional cooperation, and national fishery management on endangered elasmobranchs in China as well as related countries.

A multiplex PCR mini-barcode assay to identify processed shark products in the global trade

PubMed Central

Fields, Andrew; Abercrombie, Debra; Feldheim, Kevin; Shea, Stanley K. H.; Chapman, Demian D.

2017-01-01

Protecting sharks from overexploitation has become global priority after widespread population declines have occurred. Tracking catches and trade on a species-specific basis has proven challenging, in part due to difficulties in identifying processed shark products such as fins, meat, and liver oil. This has hindered efforts to implement regulations aimed at promoting sustainable use of commercially important species and protection of imperiled species. Genetic approaches to identify shark products exist but are typically based on sequencing or amplifying large DNA regions and may fail to work on heavily processed products in which DNA is degraded. Here, we describe a novel multiplex PCR mini-barcode assay based on two short fragments of the cytochrome oxidase I (COI) gene. This assay can identify to species all sharks currently listed on the Convention of International Trade of Endangered Species (CITES) and most shark species present in the international trade. It achieves species diagnosis based on a single PCR and one to two downstream DNA sequencing reactions. The assay is capable of identifying highly processed shark products including fins, cooked shark fin soup, and skin-care products containing liver oil. This is a straightforward and reliable identification method for data collection and enforcement of regulations implemented for certain species at all governance levels. PMID:29020095

A multiplex PCR mini-barcode assay to identify processed shark products in the global trade.

PubMed

Cardeñosa, Diego; Fields, Andrew; Abercrombie, Debra; Feldheim, Kevin; Shea, Stanley K H; Chapman, Demian D

2017-01-01

Protecting sharks from overexploitation has become global priority after widespread population declines have occurred. Tracking catches and trade on a species-specific basis has proven challenging, in part due to difficulties in identifying processed shark products such as fins, meat, and liver oil. This has hindered efforts to implement regulations aimed at promoting sustainable use of commercially important species and protection of imperiled species. Genetic approaches to identify shark products exist but are typically based on sequencing or amplifying large DNA regions and may fail to work on heavily processed products in which DNA is degraded. Here, we describe a novel multiplex PCR mini-barcode assay based on two short fragments of the cytochrome oxidase I (COI) gene. This assay can identify to species all sharks currently listed on the Convention of International Trade of Endangered Species (CITES) and most shark species present in the international trade. It achieves species diagnosis based on a single PCR and one to two downstream DNA sequencing reactions. The assay is capable of identifying highly processed shark products including fins, cooked shark fin soup, and skin-care products containing liver oil. This is a straightforward and reliable identification method for data collection and enforcement of regulations implemented for certain species at all governance levels.

Species diversity and phylogeographical affinities of the Branchiopoda (Crustacea) of Churchill, Manitoba, Canada.

PubMed

Jeffery, Nicholas W; Elías-Gutiérrez, Manuel; Adamowicz, Sarah J

2011-01-01

The region of Churchill, Manitoba, contains a wide variety of habitats representative of both the boreal forest and arctic tundra and has been used as a model site for biodiversity studies for nearly seven decades within Canada. Much previous work has been done in Churchill to study the Daphnia pulex species complex in particular, but no study has completed a wide-scale survey on the crustacean species that inhabit Churchill's aquatic ecosystems using molecular markers. We have employed DNA barcoding to study the diversity of the Branchiopoda (Crustacea) in a wide variety of freshwater habitats and to determine the likely origins of the Churchill fauna following the last glaciation. The standard animal barcode marker (COI) was sequenced for 327 specimens, and a 3% divergence threshold was used to delineate potential species. We found 42 provisional and valid branchiopod species from this survey alone, including several cryptic lineages, in comparison with the 25 previously recorded from previous ecological works. Using published sequence data, we explored the phylogeographic affinities of Churchill's branchiopods, finding that the Churchill fauna apparently originated from all directions from multiple glacial refugia (including southern, Beringian, and high arctic regions). Overall, these microcrustaceans are very diverse in Churchill and contain multiple species complexes. The present study introduces among the first sequences for some understudied genera, for which further work is required to delineate species boundaries and develop a more complete understanding of branchiopod diversity over a larger spatial scale.

First record of Stegomyia albopicta in Turkey determined by active ovitrap surveillance and DNA barcoding.

PubMed

Oter, Kerem; Gunay, Filiz; Tuzer, Erkut; Linton, Yvonne-Marie; Bellini, Romeo; Alten, Bulent

2013-10-01

Despite its confirmed establishment in neighboring Greece and Bulgaria, the presence of the Oriental invasive species Stegomyia albopicta (Skuse) (=Aedes albopictus) has never been confirmed in Turkey. Active surveillance for this container-breeding species was carried out using oviposition traps at 15 discrete sites in the towns of Ipsala (n=8 sites), Kesan (n=5) (Edirne District), and Malkara (n=2) (Tekirdag District) in the Thrace region of northwestern Turkey, from May 23 through November 10, 2011. Eggs collected were reared to the fourth larval instar and adult stages where possible to facilitate integrated morphological and molecular species identification. DNA barcodes (658 bp of the mitochondrial cytochrome c oxidase I [COI] gene) were compared with all four potentially invasive Stegomyia species: St. aegypti, St. albopicta, St. cretina, and St. japonica. Sequences generated for samples collected in Thrace Region were herein confirmed as St. albopicta, the first record of this vector species in Turkey. Eggs of St. albopicta were detected in two discrete localities: (1) In the grounds of a restaurant in Kesan (in week 36), and (2) in the customs area of the Turkish-Greek border at Ipsala (in weeks 32 and 38). Multiple detection of St. albopicta eggs indicates the possible establishment of the species in northwestern Turkey. Finding this important disease vector has implications for public health and requires the implementation of active vector monitoring programs and targeted vector suppression strategies to limit the spread of this invasive vector species in Turkey.

Species Diversity and Phylogeographical Affinities of the Branchiopoda (Crustacea) of Churchill, Manitoba, Canada

PubMed Central

Jeffery, Nicholas W.; Elías-Gutiérrez, Manuel; Adamowicz, Sarah J.

2011-01-01

The region of Churchill, Manitoba, contains a wide variety of habitats representative of both the boreal forest and arctic tundra and has been used as a model site for biodiversity studies for nearly seven decades within Canada. Much previous work has been done in Churchill to study the Daphnia pulex species complex in particular, but no study has completed a wide-scale survey on the crustacean species that inhabit Churchill's aquatic ecosystems using molecular markers. We have employed DNA barcoding to study the diversity of the Branchiopoda (Crustacea) in a wide variety of freshwater habitats and to determine the likely origins of the Churchill fauna following the last glaciation. The standard animal barcode marker (COI) was sequenced for 327 specimens, and a 3% divergence threshold was used to delineate potential species. We found 42 provisional and valid branchiopod species from this survey alone, including several cryptic lineages, in comparison with the 25 previously recorded from previous ecological works. Using published sequence data, we explored the phylogeographic affinities of Churchill's branchiopods, finding that the Churchill fauna apparently originated from all directions from multiple glacial refugia (including southern, Beringian, and high arctic regions). Overall, these microcrustaceans are very diverse in Churchill and contain multiple species complexes. The present study introduces among the first sequences for some understudied genera, for which further work is required to delineate species boundaries and develop a more complete understanding of branchiopod diversity over a larger spatial scale. PMID:21610864

Universal Plant DNA Barcode Loci May Not Work in Complex Groups: A Case Study with Indian Berberis Species

PubMed Central

Roy, Sribash; Tyagi, Antariksh; Shukla, Virendra; Kumar, Anil; Singh, Uma M.; Chaudhary, Lal Babu; Datt, Bhaskar; Bag, Sumit K.; Singh, Pradhyumna K.; Nair, Narayanan K.; Husain, Tariq; Tuli, Rakesh

2010-01-01

Background The concept of DNA barcoding for species identification has gained considerable momentum in animals because of fairly successful species identification using cytochrome oxidase I (COI). In plants, matK and rbcL have been proposed as standard barcodes. However, barcoding in complex genera is a challenging task. Methodology and Principal Findings We investigated the species discriminatory power of four reportedly most promising plant DNA barcoding loci (one from nuclear genome- ITS, and three from plastid genome- trnH-psbA, rbcL and matK) in species of Indian Berberis L. (Berberidaceae) and two other genera, Ficus L. (Moraceae) and Gossypium L. (Malvaceae). Berberis species were delineated using morphological characters. These characters resulted in a well resolved species tree. Applying both nucleotide distance and nucleotide character-based approaches, we found that none of the loci, either singly or in combinations, could discriminate the species of Berberis. ITS resolved all the tested species of Ficus and Gossypium and trnH-psbA resolved 82% of the tested species in Ficus. The highly regarded matK and rbcL could not resolve all the species. Finally, we employed amplified fragment length polymorphism test in species of Berberis to determine their relationships. Using ten primer pair combinations in AFLP, the data demonstrated incomplete species resolution. Further, AFLP analysis showed that there was a tendency of the Berberis accessions to cluster according to their geographic origin rather than species affiliation. Conclusions/Significance We reconfirm the earlier reports that the concept of universal barcode in plants may not work in a number of genera. Our results also suggest that the matK and rbcL, recommended as universal barcode loci for plants, may not work in all the genera of land plants. Morphological, geographical and molecular data analyses of Indian species of Berberis suggest probable reticulate evolution and thus barcode markers may not work in this case. PMID:21060687

A Molecular Phylogeny of Anopheles Annulipes (Diptera: Culicidae) sensu lato: The Most Species-Rich Anopheline Complex

DTIC Science & Technology

2006-09-27

Maximum parsimony; Sibling species; Species complex; Myxomatosis ; DNA barcoding; Australia; Papua New Guinea; ITS2; COI; COII; EF-11. Introduction... myxomatosis to con- trol rabbits (Fenner and RatcliVe, 1965). Chris Green used data from cross-matings and the band- ing pattern of polytene chromosomes to... myxomatosis based on distribution but more sam- pling is required to conWrm this. Many of the sampling locations in this study and the allozyme study of

Simultaneous identification and DNA barcoding of six Eimeria species infecting turkeys using PCR primers targeting the mitochondrial cytochrome c oxidase subunit I (mtCOI) locus.

PubMed

Hafeez, Mian A; Shivaramaiah, Srichaitanya; Dorsey, Kristi Moore; Ogedengbe, Mosun E; El-Sherry, Shiem; Whale, Julia; Cobean, Julie; Barta, John R

2015-05-01

Species-specific PCR primers targeting the mitochondrial cytochrome c oxidase subunit I (mtCOI) locus were generated that allow for the specific identification of the most common Eimeria species infecting turkeys (i.e., Eimeria adenoeides, Eimeria meleagrimitis, Eimeria gallopavonis, Eimeria meleagridis, Eimeria dispersa, and Eimeria innocua). PCR reaction chemistries were optimized with respect to divalent cation (MgCl2) and dNTP concentrations, as well as PCR cycling conditions (particularly anneal temperature for primers). Genomic DNA samples from single oocyst-derived lines of six Eimeria species were tested to establish specificity and sensitivity of these newly designed primer pairs. A mixed 60-ng total DNA sample containing 10 ng of each of the six Eimeria species was used as DNA template to demonstrate specific amplification of the correct product using each of the species-specific primer pairs. Ten nanograms of each of the five non-target Eimeria species was pooled to provide a non-target, control DNA sample suitable to test the specificity of each primer pair. The amplifications of the COI region with species-specific primer pairs from pooled samples yielded products of expected sizes (209 to 1,012 bp) and no amplification of non-target Eimeria sp. DNA was detected using the non-target, control DNA samples. These primer pairs specific for Eimeria spp. of turkeys did not amplify any of the seven Eimeria species infecting chickens. The newly developed PCR primers can be used as a diagnostic tool capable of specifically identifying six turkey Eimeria species; additionally, sequencing of the PCR amplification products yields sequence-based genotyping data suitable for identification and molecular phylogenetics.

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

PubMed Central

Takiya, Daniela M.; Nessimian, Jorge L.

2016-01-01

Metrichia is assigned to the Ochrotrichiinae, a group of almost exclusively Neotropical microcaddisflies. Metrichia comprises 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. acuminata sp. nov., M. azul sp. nov., M. bonita sp. nov., M. bracui sp. nov., M. caraca sp. nov., M. circuliforme sp. nov., M. curta sp. nov., M. farofa sp. nov., M. forceps sp. nov., M. formosinha sp. nov., M. goiana sp. nov., M. itabaiana sp. nov., M. longissima sp. nov., M. peluda sp. nov., M. rafaeli sp. nov., M. simples sp. nov., M. talhada sp. nov., M. tere sp. nov., M. ubajara sp. nov., and M. vulgaris sp. nov. DNA barcode sequences (577 bp of the mitochondrial gene COI) were generated for 13 of the new species and two previously known species of Metrichia resulting in 64 sequences. In addition, COI sequences were obtained for other genera of Ochrotrichiinae (Angrisanoia, Nothotrichia, Ochrotrichia, Ragatrichia, and Rhyacopsyche). 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 among Metrichia sequences (highest intraspecific divergence: 4.8%; lowest interspecific divergence: 12.6%). Molecular analyses also allowed the association of larvae and adults of Metrichia bonita sp. 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 of Metrichia species, 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. PMID:27169001

Using DNA barcodes for assessing diversity in the family Hybotidae (Diptera, Empidoidea)

PubMed Central

Nagy, Zoltán T.; Sonet, Gontran; Mortelmans, Jonas; Vandewynkel, Camille; Grootaert, Patrick

2013-01-01

Abstract Empidoidea is one of the largest extant lineages of flies, but phylogenetic relationships among species of this group are poorly investigated and global diversity remains scarcely assessed. In this context, one of the most enigmatic empidoid families is Hybotidae. Within the framework of a pilot study, we barcoded 339 specimens of Old World hybotids belonging to 164 species and 22 genera (plus two Empis as outgroups) and attempted to evaluate whether patterns of intra- and interspecific divergences match the current taxonomy. We used a large sampling of diverse Hybotidae. The material came from the Palaearctic (Belgium, France, Portugal and Russian Caucasus), the Afrotropic (Democratic Republic of the Congo) and the Oriental realms (Singapore and Thailand). Thereby, we optimized lab protocols for barcoding hybotids. Although DNA barcodes generally well distinguished recognized taxa, the study also revealed a number of unexpected phenomena: e.g., undescribed taxa found within morphologically very similar or identical specimens, especially when geographic distance was large; some morphologically distinct species showed no genetic divergence; or different pattern of intraspecific divergence between populations or closely related species. Using COI sequences and simple Neighbour-Joining tree reconstructions, the monophyly of many species- and genus-level taxa was well supported, but more inclusive taxonomical levels did not receive significant bootstrap support. We conclude that in hybotids DNA barcoding might be well used to identify species, when two main constraints are considered. First, incomplete barcoding libraries hinder efficient (correct) identification. Therefore, extra efforts are needed to increase the representation of hybotids in these databases. Second, the spatial scale of sampling has to be taken into account, and especially for widespread species or species complexes with unclear taxonomy, an integrative approach has to be used to clarify species boundaries and identities. PMID:24453562

Existence of species complex largely reduced barcoding success for invasive species of Tephritidae: a case study in Bactrocera spp.

PubMed

Jiang, F; Jin, Q; Liang, L; Zhang, A B; Li, Z H

2014-11-01

Fruit flies in the family Tephritidae are the economically important pests that have many species complexes. DNA barcoding has gradually been verified as an effective tool for identifying species in a wide range of taxonomic groups, and there are several publications on rapid and accurate identification of fruit flies based on this technique; however, comprehensive analyses of large and new taxa for the effectiveness of DNA barcoding for fruit flies identification have been rare. In this study, we evaluated the COI barcode sequences for the diagnosis of fruit flies using 1426 sequences for 73 species of Bactrocera distributed worldwide. Tree-based [neighbour-joining (NJ)]; distance-based, such as Best Match (BM), Best Close Match (BCM) and Minimum Distance (MD); and character-based methods were used to evaluate the barcoding success rates obtained with maintaining the species complex in the data set, treating a species complex as a single taxon unit, and removing the species complex. Our results indicate that the average divergence between species was 14.04% (0.00-25.16%), whereas within a species this was 0.81% (0.00-9.71%); the existence of species complexes largely reduced the barcoding success for Tephritidae, for example relatively low success rates (74.4% based on BM and BCM and 84.8% based on MD) were obtained when the sequences from species complexes were included in the analysis, whereas significantly higher success rates were achieved if the species complexes were treated as a single taxon or removed from the data set - BM (98.9%), BCM (98.5%) and MD (97.5%), or BM (98.1%), BCM (97.4%) and MD (98.2%). © 2014 John Wiley & Sons Ltd.

Morphological and genetic analyses of the first record of longrakered trevally, Ulua mentalis (Perciformes: Carangidae) and of the pinjalo snapper, Pinjalo pinjalo (Perciformes: Lutjanidae) in the Odisha coast, Bay of Bengal.

PubMed

Barik, Tapan K; Swain, Surya N; Sahu, Bijayalaxmi; Tripathy, Bibarani; Acharya, Usha R

2018-05-01

Identification of fish species have so far been carried out mostly by classical morpho-taxonomy. In the present study, however, an attempt has been taken to identify two species of fishes Ulua mentalis and Pinjalo pinjalo of order Perciformes which happens to be the first record in Odisha coast Bay of Bengal, India during the year 2015, using DNA barcoding technique for reconfirmation over conventional morpho-taxonomy. During recent past, study of molecular-taxonomical profile of mitochondrial DNA in general and Cytochrome Oxidase subunit I (COI) gene in particular has gained enormous importance for accurate identification of species. In the present study, the partial COI sequence of Ulua mentalis and Pinjalo pinjalo were generated. Analysis using the COI gene produced phylogenetic trees in concurrence with other multi gene studies and we came across the identical phylogenetic relationship considering Neighbor-Joining and Maximum Likelihood tree. Moreover, these molecular data set further testified in Bayesian framework to reevaluate the exact taxonomic groupings within the family. Surprisingly, Ulua mentalis and Pinjalo pinjalo seems to be closely related to their sister taxa.

Revision of torrent mites (Parasitengona, Torrenticolidae, Torrenticola) of the United States and Canada: 90 descriptions, molecular phylogenetics, and a key to species

PubMed Central

Fisher, J. Ray; Fisher, Danielle M.; Skvarla, Michael J.; Nelson, Whitney A.; Dowling, Ashley P.G.

2017-01-01

Abstract The descriptive biology of torrent mites (Parasitengona: Torrenticolidae: Torrenticola) of North America (north of Mexico) is investigated using integrative methods. Material examined includes approximately 2,300 specimens from nearly 500 localities across the United States and Canada, and a few collections in Mexico and Central America. Species hypotheses are derived from a phylogenetic analysis of the barcoding region of cytochrome c oxidase subunit 1 (COI) for 476 specimens and supported with morphology and biogeography. Relationships between species are examined with a combined analysis of COI and two expansion regions (D2–3) of the large ribosomal subunit (28S rDNA) for 57 specimens. All previously described species from the US and Canada are examined. Our results indicate the need to synonymize four species: T. mercedensis (Marshall, 1943) is a junior synonym of T. sierrensis (Marshall, 1943); T. rectiforma Habeeb, 1974 is a junior synonym of T. ellipsoidalis (Marshall, 1943); T. neoconnexa Habeeb, 1957 is a junior synonym of T. magnexa Habeeb, 1955; and T. esbelta Cramer, 1992 is a junior synonym of T. boettgeri KO Viets, 1977. We describe 66 new species and re-describe all previously described regional species. Our findings indicate that total diversity of Torrenticola in the United States and Canada comprises 90 species, 57 known from the east and 33 from the west. We organize these species into four species complexes that include 13 identification groups. An additional 13 species do not fit within an identification group. The southern Appalachians are suspected to contain the highest concentration of remaining undescribed diversity. A key is provided to all known species in the US and Canada. PMID:29118596

Species diversity in the marine microturbellarian Astrotorhynchus bifidus sensu lato (Platyhelminthes: Rhabdocoela) from the Northeast Pacific Ocean.

PubMed

Van Steenkiste, Niels W L; Herbert, Elizabeth R; Leander, Brian S

2018-03-01

Increasing evidence suggests that many widespread species of meiofauna are in fact regional complexes of (pseudo-)cryptic species. This knowledge has challenged the 'Everything is Everywhere' hypothesis and also partly explains the meiofauna paradox of widespread nominal species with limited dispersal abilities. Here, we investigated species diversity within the marine microturbellarian Astrotorhynchus bifidus sensu lato in the Northeast Pacific Ocean. We used a multiple-evidence approach combining multi-gene (18S, 28S, COI) phylogenetic analyses, several single-gene and multi-gene species delimitation methods, haplotype networks and conventional taxonomy to designate Primary Species Hypotheses (PSHs). This included the development of rhabdocoel-specific COI barcode primers, which also have the potential to aid in species identification and delimitation in other rhabdocoels. Secondary Species Hypotheses (SSHs) corresponding to morphospecies and pseudo-cryptic species were then proposed based on the minimum consensus of different PSHs. Our results showed that (a) there are at least five species in the A. bifidus complex in the Northeast Pacific Ocean, four of which can be diagnosed based on stylet morphology, (b) the A. bifidus complex is a mixture of sympatric and allopatric species with regional and/or subglobal distributions, (c) sympatry occurs on local (sample sites), regional (Northeastern Pacific) and subglobal (Northern Atlantic, Arctic, Northeastern Pacific) scales. Mechanisms for this co-occurrence are still poorly understood, but we hypothesize they could include habitat differentiation (spatial and/or seasonal) and life history characteristics such as sexual selection and dispersal abilities. Our results also suggest the need for improved sampling and exploration of molecular markers to accurately map gene flow and broaden our understanding of species diversity and distribution of microturbellarians in particular and meiofauna in general. Copyright © 2017 Elsevier Inc. All rights reserved.

Resolving species delimitation within the genus Bunopus Blanford, 1874 (Squamata: Gekkonidae) in Iran using DNA barcoding approach.

PubMed

Khosravani, Azar; Rastegar-Pouyani, Eskandar; Rastegar-Pouyani, Nasrullah; Oraie, Hamzeh; Papenfuss, Theodore J

2017-12-19

Mitochondrial COI sequences were used to investigate species delimitation within the genus Bunopus in Iran. A dataset with a final sequence length of 633 nucleotides including 100 specimens from 31 geographically distant localities across Iran were generated. The result demonstrated that two major clades with strong support can be identified within the genus Bunopus in Iran. Clade A includes Bunopus crassicaudus and two new entities, eastern populations (subclade A2,1) and Shahdad populations (subclade A2,2). The second clade comprises western and southwestern populations (subclade B1,1), Arabian populations (subclade B1,2) and south and southeast populations in Iran, to which Bunopus tuberculatus (subclade B2) is assigned. In addition to Bunopus crassicaudus and B. tuberculatus, three new candidate species in Iran can easily be identified based on the DNA barcoding approach.

New species and phylogenetic relationships of the spider genus Coptoprepes using morphological and sequence data (Araneae: Anyphaenidae).

PubMed

Barone, Mariana L; Werenkraut, Victoria; Ramírez, Martín J

2016-10-17

We present evidence from the standard cytochrome c oxidase subunit I (COI) barcoding marker and from new collections, showing that the males and females of C. ecotono Werenkraut & Ramírez were mismatched, and describe the female of that species for the first time. An undescribed male from Chile is assigned to the new species Coptoprepes laudani, together with the female that was previously thought as C. ecotono. The matching of sexes is justified after a dual cladistics analysis of morphological and sequence data in combination. New locality data and barcoding sequences are provided for other species of Coptoprepes, all endemic of the temperate forests of Chile and adjacent Argentina. Although morphology and sequences are not conclusive on the relationships of Coptoprepes species, the sequence data suggests that the species without a retrolateral tibial apophysis may belong to an independent lineage.

Integrated Taxonomy Reveals Hidden Diversity in Northern Australian Fishes: A New Species of Seamoth (Genus Pegasus)

PubMed Central

Osterhage, Deborah; Pogonoski, John J.; Appleyard, Sharon A.; White, William T.

2016-01-01

Fishes are one of the most intensively studied marine taxonomic groups yet cryptic species are still being discovered. An integrated taxonomic approach is used herein to delineate and describe a new cryptic seamoth (genus Pegasus) from what was previously a wide-ranging species. Preliminary mitochondrial DNA barcoding indicated possible speciation in Pegasus volitans specimens collected in surveys of the Torres Strait and Great Barrier Reef off Queensland in Australia. Morphological and meristic investigations found key differences in a number of characters between P. volitans and the new species, P. tetrabelos. Further mt DNA barcoding of both the COI and the slower mutating 16S genes of additional specimens provided strong support for two separate species. Pegasus tetrabelos and P. volitans are sympatric in northern Australia and were frequently caught together in trawls at the same depths. PMID:26934529

DNA barcoding simplifies environmental risk assessment of genetically modified crops in biodiverse regions.

PubMed

Nzeduru, Chinyere V; Ronca, Sandra; Wilkinson, Mike J

2012-01-01

Transgenes encoding for insecticidal crystal (Cry) proteins from the soil-dwelling bacterium Bacillus Thuringiensis have been widely introduced into Genetically Modified (GM) crops to confer protection against insect pests. Concern that these transgenes may also harm beneficial or otherwise valued insects (so-called Non Target Organisms, NTOs) represents a major element of the Environmental Risk Assessments (ERAs) used by all countries prior to commercial release. Compiling a comprehensive list of potentially susceptible NTOs is therefore a necessary part of an ERA for any Cry toxin-containing GM crop. In partly-characterised and biodiverse countries, NTO identification is slowed by the need for taxonomic expertise and time to enable morphological identifications. This limitation represents a potentially serious barrier to timely adoption of GM technology in some developing countries. We consider Bt Cry1A cowpea (Vigna unguiculata) in Nigeria as an exemplar to demonstrate how COI barcoding can provide a simple and cost-effective means of addressing this problem. Over a period of eight weeks, we collected 163 insects from cowpea flowers across the agroecological and geographic range of the crop in Nigeria. These individuals included 32 Operational Taxonomic Units (OTUs) spanning four Orders and that could mostly be assigned to genus or species level. They included 12 Lepidopterans and two Coleopterans (both potentially sensitive to different groups of Cry proteins). Thus, barcode-assisted diagnoses were highly harmonised across groups (typically to genus or species level) and so were insensitive to expertise or knowledge gaps. Decisively, the entire study was completed within four months at a cost of less than 10,000 US$. The broader implications of the findings for food security and the capacity for safe adoption of GM technology are briefly explored.

Identification of Belgian mosquito species (Diptera: Culicidae) by DNA barcoding.

PubMed

Versteirt, V; Nagy, Z T; Roelants, P; Denis, L; Breman, F C; Damiens, D; Dekoninck, W; Backeljau, T; Coosemans, M; Van Bortel, W

2015-03-01

Since its introduction in 2003, DNA barcoding has proven to be a promising method for the identification of many taxa, including mosquitoes (Diptera: Culicidae). Many mosquito species are potential vectors of pathogens, and correct identification in all life stages is essential for effective mosquito monitoring and control. To use DNA barcoding for species identification, a reliable and comprehensive reference database of verified DNA sequences is required. Hence, DNA sequence diversity of mosquitoes in Belgium was assessed using a 658 bp fragment of the mitochondrial cytochrome oxidase I (COI) gene, and a reference data set was established. Most species appeared as well-supported clusters. Intraspecific Kimura 2-parameter (K2P) distances averaged 0.7%, and the maximum observed K2P distance was 6.2% for Aedes koreicus. A small overlap between intra- and interspecific K2P distances for congeneric sequences was observed. Overall, the identification success using best match and the best close match criteria were high, that is above 98%. No clear genetic division was found between the closely related species Aedes annulipes and Aedes cantans, which can be confused using morphological identification only. The members of the Anopheles maculipennis complex, that is Anopheles maculipennis s.s. and An. messeae, were weakly supported as monophyletic taxa. This study showed that DNA barcoding offers a reliable framework for mosquito species identification in Belgium except for some closely related species. © 2014 John Wiley & Sons Ltd.

A reliable DNA barcode reference library for the identification of the North European shelf fish fauna.

PubMed

Knebelsberger, Thomas; Landi, Monica; Neumann, Hermann; Kloppmann, Matthias; Sell, Anne F; Campbell, Patrick D; Laakmann, Silke; Raupach, Michael J; Carvalho, Gary R; Costa, Filipe O

2014-09-01

Valid fish species identification is an essential step both for fundamental science and fisheries management. The traditional identification is mainly based on external morphological diagnostic characters, leading to inconsistent results in many cases. Here, we provide a sequence reference library based on mitochondrial cytochrome c oxidase subunit I (COI) for a valid identification of 93 North Atlantic fish species originating from the North Sea and adjacent waters, including many commercially exploited species. Neighbour-joining analysis based on K2P genetic distances formed nonoverlapping clusters for all species with a ≥99% bootstrap support each. Identification was successful for 100% of the species as the minimum genetic distance to the nearest neighbour always exceeded the maximum intraspecific distance. A barcoding gap was apparent for the whole data set. Within-species distances ranged from 0 to 2.35%, while interspecific distances varied between 3.15 and 28.09%. Distances between congeners were on average 51-fold higher than those within species. The validation of the sequence library by applying BOLDs barcode index number (BIN) analysis tool and a ranking system demonstrated high taxonomic reliability of the DNA barcodes for 85% of the investigated fish species. Thus, the sequence library presented here can be confidently used as a benchmark for identification of at least two-thirds of the typical fish species recorded for the North Sea. © 2014 John Wiley & Sons Ltd.

Application of DNA barcoding in biodiversity studies of shallow-water octocorals: molecular proxies agree with morphological estimates of species richness in Palau

NASA Astrophysics Data System (ADS)

McFadden, C. S.; Brown, A. S.; Brayton, C.; Hunt, C. B.; van Ofwegen, L. P.

2014-06-01

The application of DNA barcoding to anthozoan cnidarians has been hindered by their slow rates of mitochondrial gene evolution and the failure to identify alternative molecular markers that distinguish species reliably. Among octocorals, however, multilocus barcodes can distinguish up to 70 % of morphospecies, thereby facilitating the identification of species that are ecologically important but still very poorly known taxonomically. We tested the ability of these imperfect DNA barcodes to estimate species richness in a biodiversity survey of the shallow-water octocoral fauna of Palau using multilocus ( COI, mtMutS, 28S rDNA) sequences obtained from 305 specimens representing 38 genera of octocorals. Numbers and identities of species were estimated independently (1) by a taxonomic expert using morphological criteria and (2) by assigning sequences to molecular operational taxonomic units (MOTUs) using predefined genetic distance thresholds. Estimated numbers of MOTUs ranged from 73 to 128 depending on the barcode and distance threshold applied, bracketing the estimated number of 118 morphospecies. Concordance between morphospecies identifications and MOTUs ranged from 71 to 75 % and differed little among barcodes. For the speciose and ecologically dominant genus Sinularia, however, we were able to identify 95 % of specimens correctly simply by comparing mtMutS sequences and in situ photographs of colonies to an existing vouchered database. Because we lack a clear understanding of species boundaries in most of these taxa, numbers of morphospecies and MOTUs are both estimates of the true species diversity, and we cannot currently determine which is more accurate. Our results suggest, however, that the two methods provide comparable estimates of species richness for shallow-water Indo-Pacific octocorals. Use of molecular barcodes in biodiversity surveys will facilitate comparisons of species richness and composition among localities and over time, data that do not currently exist for any octocoral community.

A Transcontinental Challenge — A Test of DNA Barcode Performance for 1,541 Species of Canadian Noctuoidea (Lepidoptera)

PubMed Central

Zahiri, Reza; Lafontaine, J. Donald; Schmidt, B. Christian; deWaard, Jeremy R.; Zakharov, Evgeny V.; Hebert, Paul D. N.

2014-01-01

This study provides a first, comprehensive, diagnostic use of DNA barcodes for the Canadian fauna of noctuoids or “owlet” moths (Lepidoptera: Noctuoidea) based on vouchered records for 1,541 species (99.1% species coverage), and more than 30,000 sequences. When viewed from a Canada-wide perspective, DNA barcodes unambiguously discriminate 90% of the noctuoid species recognized through prior taxonomic study, and resolution reaches 95.6% when considered at a provincial scale. Barcode sharing is concentrated in certain lineages with 54% of the cases involving 1.8% of the genera. Deep intraspecific divergence exists in 7.7% of the species, but further studies are required to clarify whether these cases reflect an overlooked species complex or phylogeographic variation in a single species. Non-native species possess higher Nearest-Neighbour (NN) distances than native taxa, whereas generalist feeders have lower NN distances than those with more specialized feeding habits. We found high concordance between taxonomic names and sequence clusters delineated by the Barcode Index Number (BIN) system with 1,082 species (70%) assigned to a unique BIN. The cases of discordance involve both BIN mergers and BIN splits with 38 species falling into both categories, most likely reflecting bidirectional introgression. One fifth of the species are involved in a BIN merger reflecting the presence of 158 species sharing their barcode sequence with at least one other taxon, and 189 species with low, but diagnostic COI divergence. A very few cases (13) involved species whose members fell into both categories. Most of the remaining 140 species show a split into two or three BINs per species, while Virbia ferruginosa was divided into 16. The overall results confirm that DNA barcodes are effective for the identification of Canadian noctuoids. This study also affirms that BINs are a strong proxy for species, providing a pathway for a rapid, accurate estimation of animal diversity. PMID:24667847

Genetic assessment of leech species from yak (Bos grunniens) in the tract of Northeast India.

PubMed

Chatterjee, Nilkantha; Dhar, Bishal; Bhattarcharya, Debasis; Deori, Sourabh; Doley, Juwar; Bam, Joken; Das, Pranab J; Bera, Asit K; Deb, Sitangshu M; Devi, Ningthoujam Neelima; Paul, Rajesh; Malvika, Sorokhaibam; Ghosh, Sankar Kumar

2018-01-01

Yak is an iconic symbol of Tibet and high altitudes of Northeast India. It is highly cherished for milk, meat, and skin. However, yaks suffer drastic change in milk production, weight loss, etc, when infested by parasites. Among them, infestation by leeches is a serious problem in the Himalayan belt of Northeast India. The parasite feeds on blood externally or from body orifices, like nasopharynx, oral, rectum, etc. But there has been limited data about the leech species infesting the yak in that region because of the difficulties in morphological identification due to plasticity of the body, changes in shape, and surface structure and thus, warrants for the molecular characterization of leech. In anticipation, this study would be influential in proper identification of leech species infesting yak track and also helpful in inventorying of leech species in Northeast India. Here, we investigated, through combined approach of molecular markers and morphological parameters for the identification of leech species infesting yak. The DNA sequences of COI barcode fragment, 18S and 28S rDNA, were analyzed for species identification. The generated sequences were subjected to similarity match in global database and analyzed further through Neighbour-Joining, K2P distance based as well as ML approach. Among the three markers, only COI was successful in delineating species whereas the 18S and 28S failed to delineate the species. Our study confirmed the presence of the species from genus Hirudinaria, Haemadipsa, Whitmania, and one species Myxobdella annandalae, which has not been previously reported from this region.

A DNA barcode library for Germany's mayflies, stoneflies and caddisflies (Ephemeroptera, Plecoptera and Trichoptera).

PubMed

Morinière, Jérôme; Hendrich, Lars; Balke, Michael; Beermann, Arne J; König, Tobias; Hess, Monika; Koch, Stefan; Müller, Reinhard; Leese, Florian; Hebert, Paul D N; Hausmann, Axel; Schubart, Christoph D; Haszprunar, Gerhard

2017-11-01

Mayflies, stoneflies and caddisflies (Ephemeroptera, Plecoptera and Trichoptera) are prominent representatives of aquatic macroinvertebrates, commonly used as indicator organisms for water quality and ecosystem assessments. However, unambiguous morphological identification of EPT species, especially their immature life stages, is a challenging, yet fundamental task. A comprehensive DNA barcode library based upon taxonomically well-curated specimens is needed to overcome the problematic identification. Once available, this library will support the implementation of fast, cost-efficient and reliable DNA-based identifications and assessments of ecological status. This study represents a major step towards a DNA barcode reference library as it covers for two-thirds of Germany's EPT species including 2,613 individuals belonging to 363 identified species. As such, it provides coverage for 38 of 44 families (86%) and practically all major bioindicator species. DNA barcode compliant sequences (≥500 bp) were recovered from 98.74% of the analysed specimens. Whereas most species (325, i.e., 89.53%) were unambiguously assigned to a single Barcode Index Number (BIN) by its COI sequence, 38 species (18 Ephemeroptera, nine Plecoptera and 11 Trichoptera) were assigned to a total of 89 BINs. Most of these additional BINs formed nearest neighbour clusters, reflecting the discrimination of geographical subclades of a currently recognized species. BIN sharing was uncommon, involving only two species pairs of Ephemeroptera. Interestingly, both maximum pairwise and nearest neighbour distances were substantially higher for Ephemeroptera compared to Plecoptera and Trichoptera, possibly indicating older speciation events, stronger positive selection or faster rate of molecular evolution. © 2017 John Wiley & Sons Ltd.

DNA barcoding using skin exuviates can improve identification and biodiversity studies of snakes.

PubMed

Khedkar, Trupti; Sharma, Rashmi; Tiknaik, Anita; Khedkar, Gulab; Naikwade, Bhagwat S; Ron, Tetsuzan Benny; Haymer, David

2016-01-01

Snakes represent a taxonomically underdeveloped group of animals in India with a lack of experts and incomplete taxonomic descriptions being the main deterrents to advances in this area. Molecular taxonomic approaches using DNA barcoding could aid in snake identification as well as studies of biodiversity. Here a non-invasive sampling method using DNA barcoding is tested using skin exuviates. Taxonomically authenticated samples were collected and tested for validation and comparisons to unknown snake exuviate samples. This approach was also used to construct the first comprehensive study targeting the snake species from Maharashtra state in India. A total of 92 skin exuviate samples were collected and tested for this study. Of these, 81 samples were successfully DNA barcoded and compared with unknown samples for assignment of taxonomic identity. Good quality DNA was obtained irrespective of age and quality of the exuviate material, and all unknown samples were successfully identified. A total of 23 species of snakes were identified, six of which were in the list of Endangered species (Red Data Book). Intra- and inter-specific distance values were also calculated, and these were sufficient to allow discrimination among species and between species without ambiguity in most cases. Two samples were suspected to represent cryptic species based on deep K2P divergence values (>3%), and one sample could be identified to the genus level only. Eleven samples failed to amplify COI sequences, suggesting the need for alternative PCR primer pairs. This study clearly documents how snake skin exuviates can be used for DNA barcoding, estimates of diversity and population genetic structuring in a noninvasive manner.

Quality Assurance in the Polio Laboratory. Cell Sensitivity and Cell Authentication Assays.

PubMed

Dunn, Glynis

2016-01-01

The accuracy of poliovirus surveillance is largely dependent on the quality of the cell lines used for virus isolation, which is the foundation of poliovirus diagnostic work. Many cell lines are available for the isolation of enteroviruses, whilst genetically modified L20B cells can be used as a diagnostic tool for the identification of polioviruses. To be confident that cells can consistently isolate the virus of interest, it is necessary to have a quality assurance system in place, which will ensure that the cells in use are not contaminated with other cell lines or microorganisms and that they remain sensitive to the viruses being studied.The sensitivity of cell lines can be assessed by the regular testing of a virus standard of known titer in the cell lines used for virus isolation. The titers obtained are compared to previously obtained titers in the same assay, so that any loss of sensitivity can be detected.However, the detection of cell line cross contamination is more difficult. DNA bar coding is a technique that uses a short DNA sequence from a standardized position in the genome as a molecular diagnostic assay for species-level identification. For almost all groups of higher animals, the cytochrome c oxidase subunit 1 of mitochondrial DNA (CO1) is emerging as the standard barcode region. This region is 648 nucleotide base pairs long in most phylogenetic groups and is flanked by regions of conserved sequences, making it relatively easy to isolate and analyze. DNA barcodes vary among individuals of the same species to a very minor degree (generally less than 1-2 %), and a growing number of studies have shown that the COI sequences of even closely related species differ by several per cent, making it possible to identify different species with high confidence.

First results of the German Barcode of Life (GBOL) – Myriapoda project: Cryptic lineages in German Stenotaenia linearis (Koch, 1835) (Chilopoda, Geophilomorpha)

PubMed Central

Wesener, Thomas; Voigtländer, Karin; Decker, Peter; Oeyen, Jan Philip; Spelda, Jörg; Lindner, Norman

2015-01-01

Abstract As part of the German Barcode of Life (GBOL) Myriapoda program, which aims to sequence the COI barcoding fragment for 2000 specimens of Germany’s 200 myriapod species in the near future, 44 sequences of the centipede order Geophilomorpha are analyzed. The analyses are limited to the genera Geophilus Leach, 1814 and Stenotaenia Koch, 1847 and include a total of six species. A special focus is Stenotaenia, of which 19 specimens from southern, western and eastern Germany could be successfully sequenced. The Stenotaenia data shows the presence of three to four vastly different (13.7–16.7% p-distance) lineages of the genus in Germany. At least two of the three lineages show a wide distribution across Germany, only the lineage including topotypes of Stenotaenia linearis shows a more restricted distribution in southern Germany. In a maximum likelihood phylogenetic analysis the Italian species Stenotaenia ‘sorrentina’ (Attems, 1903) groups with the different German Stenotaenia linearis clades. The strongly different Stenotaenia linearis lineages within Germany, independent of geography, are a strong hint for the presence of additional, cryptic Stenotaenia species in Germany. PMID:26257532

Revision of the species of Lytopylus from Area de Conservación Guanacaste, northwestern Costa Rica (Hymenoptera, Braconidae, Agathidinae)

PubMed Central

Kang, Ilgoo; Chapman, Eric G.; Janzen, Daniel H.; Hallwachs, Winnie; Tanya Dapkey; Smith, M. Alex; Sharkey, Michael J.

2017-01-01

Abstract Thirty two new species of Lytopylus (Agathidinae) are described with image plates for each species: Lytopylus alejandromasisi sp. n., Lytopylus alfredomainieri sp. n., Lytopylus anamariamongeae sp. n., Lytopylus angelagonzalezae sp. n., Lytopylus cesarmorai sp. n., Lytopylus eddysanchezi sp. n., Lytopylus eliethcantillanoae sp. n., Lytopylus ericchapmani sp. n., Lytopylus gahyunae sp. n., Lytopylus gisukae sp. n., Lytopylus guillermopereirai sp. n., Lytopylus gustavoindunii sp. n., Lytopylus hartmanguidoi sp. n., Lytopylus hernanbravoi sp. n., Lytopylus hokwoni sp. n., Lytopylus ivanniasandovalae sp. n., Lytopylus johanvalerioi sp. n., Lytopylus josecortesi sp. n., Lytopylus luisgaritai sp. n., Lytopylus mariamartachavarriae sp. n., Lytopylus miguelviquezi sp. n., Lytopylus motohasegawai sp. n., Lytopylus okchunae sp. n., Lytopylus pablocobbi sp. n., Lytopylus robertofernandezi sp. n., Lytopylus rogerblancoi sp. n., Lytopylus salvadorlopezi sp. n., Lytopylus sangyeoni sp. n., Lytopylus sarahmeierottoae sp. n., Lytopylus sergiobermudezi sp. n., Lytopylus sigifredomarini sp. n., and Lytopylus youngcheae sp. n. A dichotomous key and a link to an electronic, interactive key are included. All specimens were reared from Lepidoptera larvae collected in Area de Conservación Guanacaste (ACG) and all are associated with ecological information including host caterpillar, collection date, eclosion date, caterpillar food plant, and locality. Neighbor-joining and maximum likelihood analyses of the barcode region of the mitochondrial cytochrome c oxidase subunit I gene (COI DNA barcode) were conducted to aid in species delimitation. PMID:29308026

Systematics of Spiny Predatory Katydids (Tettigoniidae: Listroscelidinae) from the Brazilian Atlantic Forest Based on Morphology and Molecular Data

PubMed Central

Fialho, Verônica Saraiva; Chamorro-Rengifo, Juliana; Lopes-Andrade, Cristiano; Yotoko, Karla Suemy Clemente

2014-01-01

Listroscelidinae (Orthoptera: Tettigoniidae) are insectivorous Pantropical katydids whose taxonomy presents a long history of controversy, with several genera incertae sedis. This work focused on species occurring in the Brazilian Atlantic Forest, one of the world's most threatened biomes. We examined material deposited in scientific collections and visited 15 conservation units from Rio de Janeiro to southern Bahia between November 2011 and January 2012, catching 104 specimens from 10 conservation units. Based on morphological and molecular data we redefined Listroscelidini, adding a new tribe, new genus and eight new species to the subfamily. Using morphological analysis, we redescribed and added new geographic records for six species, synonymized two species and built a provisional identification key for the Atlantic Forest Listroscelidinae. Molecular results suggest two new species and a new genus to be described, possibly by the fission of the genus Hamayulus. We also proposed a 500 bp region in the final portion of the COI to be used as a molecular barcode. Our data suggest that the Atlantic Forest Listroscelidinae are seriously endangered, because they occur in highly preserved forest remnants, show high rates of endemism and have a narrow geographic distribution. Based on our results, we suggest future collection efforts must take into account the molecular barcode data to accelerate species recognition. PMID:25118712

Resurrection of New Caledonian maskray Neotrygon trigonoides (Myliobatoidei: Dasyatidae) from synonymy with N. kuhlii, based on cytochrome-oxidase I gene sequences and spotting patterns.

PubMed

Borsa, Philippe; Arlyza, Irma S; Chen, Wei-Jen; Durand, Jean-Dominique; Meekan, Mark G; Shen, Kang-Ning

2013-04-01

The maskray from New Caledonia, Neotrygon trigonoides Castelnau, 1873, has been recently synonymized with the blue-spotted maskray, N. kuhlii (Müller and Henle, 1841), a species with wide Indo-West Pacific distribution, but the reasons for this are unclear. Blue-spotted maskray specimens were collected from the Indian Ocean (Tanzania, Sumatra) and the Coral Triangle (Indonesia, Taiwan, and West Papua), and N. trigonoides specimens were collected from New Caledonia (Coral-Sea). Their partial COI gene sequences were generated to expand the available DNA-barcode database on this species, which currently comprises homologous sequences from Ningaloo Reef, the Coral Triangle and the Great Barrier Reef (Coral-Sea). Spotting patterns were also compared across regions. Haplotypes from the Coral-Sea formed a haplogroup phylogenetically distinct from all other haplotypes sampled in the Indo-West Pacific. No clear-cut geographic composition relative to DNA-barcodes or spotting patterns was apparent in N. kuhlii samples across the Indian Ocean and the Coral Triangle. The New Caledonian maskray had spotting patterns markedly different from all the other samples. This, added to a substantial level of net nucleotide divergence (2.6%) with typical N. kuhlii justifies considering the New Caledonian maskray as a separate species, for which we propose to resurrect the name Neotrygon trigonoides. Copyright © 2013. Published by Elsevier SAS.

Revision of the Hawaiian psyllid genus Swezeyana, with descriptions of seven new species (Hemiptera, Psylloidea, Triozidae).

PubMed

Percy, Diana M

2018-01-01

The endemic Hawaiian genus Swezeyana Caldwell, 1940 is highly distinctive due to the extremely long genal processes. In addition, some of the immatures are ornamented with extraordinary tubercles and tentacles. Two Swezeyana species are redescribed, and seven new species are described, bringing the total number of species in the genus to nine. All species are hosted by a single, endemic host plant, Planchonella sandwicensis (Sapotaceae), which is distributed across all major islands in the archipelago. The majority of Swezeyana species are single island endemics. A sister taxon pair is found sympatrically on the same individual plants on Kauai, and putative sister or at least closely related species are also found sympatrically on Oahu and Hawaii, suggesting these taxa may have diversified in sympatry. However, there is no observed ecological niche partitioning, despite some striking morphological diversity, as all Swezeyana species have free-living immatures that are found on the leaf surface, and therefore no apparent biological shifts are coincident with occupying the same host plant. Two species groups are represented by strikingly different female terminalia structure and endoskeletal development, although ovipositor structure is very similar between the two groups. Mitochondrial DNA barcodes (COI and cytB) are provided for eight of the nine species. A phylogenetic analysis of the mitochondrial barcode regions indicates species relationships within Swezeyana and provides a comparison of genetic divergence with other Hawaiian endemic genera.

Morphological characters and DNA barcoding of Syngnathus schlegeli in the coastal waters of China

NASA Astrophysics Data System (ADS)

Chen, Zhi; Zhang, Yan; Han, Zhiqiang; Song, Na; Gao, Tianxiang

2018-03-01

A Syngnathus species widely distributed in Chinese seas was permanently identified as Syngnathus acus by native ichthyologists, but the taxonomic description about this species was inadequate and lacking conclusively molecular evidence. To identify this species, 357 individuals of this species from the coastal waters of Dandong, Yantai, Qingdao and Zhoushan were collected and measured. Morphological results showed that these slender specimens were mainly brownish, usually mottled with pale. Standard length ranged from 117 mm to 213 mm with an average length of 180.3 mm. The above characters were consistent with S. schlegeli distributed in Japan but colored differently from and much smaller than typical S. acus reported in Europe. Thus, morphological studies revealed that this species was previously misidentified as S. acus and might be S. schlegeli in reality. In addition, a fragment of cytochrome oxidase subunit I ( COI) gene of mitochondrial DNA was also sequenced for species identification, and 15 COI sequences belonging to different Syngnathus species were also used for the molecular identification. COI sequences of our specimens had the minimum genetic distance from recognized S. schlegeli from Japan and clustered with it firstly. The phylogenetic analysis similarly suggested that the species previously identified as S. acus in the coastal waters of China was S. schlegeli actually.

Discovering hidden biodiversity: the use of complementary monitoring of fish diet based on DNA barcoding in freshwater ecosystems.

PubMed

Jo, Hyunbin; Ventura, Marc; Vidal, Nicolas; Gim, Jeong-Soo; Buchaca, Teresa; Barmuta, Leon A; Jeppesen, Erik; Joo, Gea-Jae

2016-01-01

Ecological monitoring contributes to the understanding of complex ecosystem functions. The diets of fish reflect the surrounding environment and habitats and may, therefore, act as useful integrating indicators of environmental status. It is, however, often difficult to visually identify items in gut contents to species level due to digestion of soft-bodied prey beyond visual recognition, but new tools rendering this possible are now becoming available. We used a molecular approach to determine the species identities of consumed diet items of an introduced generalist feeder, brown trout (Salmo trutta), in 10 Tasmanian lakes and compared the results with those obtained from visual quantification of stomach contents. We obtained 44 unique taxa (OTUs) belonging to five phyla, including seven classes, using the barcode of life approach from cytochrome oxidase I (COI). Compared with visual quantification, DNA analysis showed greater accuracy, yielding a 1.4-fold higher number of OTUs. Rarefaction curve analysis showed saturation of visually inspected taxa, while the curves from the DNA barcode did not saturate. The OTUs with the highest proportions of haplotypes were the families of terrestrial insects Formicidae, Chrysomelidae, and Torbidae and the freshwater Chironomidae. Haplotype occurrence per lake was negatively correlated with lake depth and transparency. Nearly all haplotypes were only found in one fish gut from a single lake. Our results indicate that DNA barcoding of fish diets is a useful and complementary method for discovering hidden biodiversity.

Who's there? - First morphological and DNA barcoding catalogue of the shallow Hawai'ian sponge fauna.

PubMed

Núñez Pons, Laura; Calcinai, Barbara; Gates, Ruth D

2017-01-01

The sponge fauna has been largely overlooked in the Archipelago of Hawai'i, notwithstanding the paramount role of this taxon in marine ecosystems. The lack of knowledge about Porifera populations inhabiting the Hawai'ian reefs limits the development of ecological studies aimed at understanding the functioning of these marine systems. Consequently, this project addresses this gap by describing the most representative sponge species in the shallow waters of the enigmatic bay of Kane'ohe Bay, in O'ahu Island. A total of 30 species (28 demosponges and two calcareous sponges) living associated to the reef structures are here reported. Six of these species are new records to the Hawai'ian Porifera catalogue and are suspected to be recent introductions to these islands. Morphological descriptions of the voucher specimens are provided, along with sequencing data of two partitions involving the mitochondrial cytochrome oxidase subunit 1 (COI) marker and a fragment covering partial (18S and 28S) and full (ITS-1, 5.8S and ITS-2) nuclear ribosomal genes. Species delimitations based on genetic distances were calculated to valitate how taxonomic assignments from DNA barcoding aligned with morphological identifications. Of the 60 sequences submitted to GenBank ~88% are the first sequencing records for the corresponding species and genetic marker. This work compiles the first catalogue combining morphological characters with DNA barcoding of Hawai'ian sponges, and contributes to the repository of public databases through the Sponge Barcoding Project initiative.

Who’s there? – First morphological and DNA barcoding catalogue of the shallow Hawai’ian sponge fauna

PubMed Central

Gates, Ruth D.

2017-01-01

The sponge fauna has been largely overlooked in the Archipelago of Hawai’i, notwithstanding the paramount role of this taxon in marine ecosystems. The lack of knowledge about Porifera populations inhabiting the Hawai’ian reefs limits the development of ecological studies aimed at understanding the functioning of these marine systems. Consequently, this project addresses this gap by describing the most representative sponge species in the shallow waters of the enigmatic bay of Kane’ohe Bay, in O’ahu Island. A total of 30 species (28 demosponges and two calcareous sponges) living associated to the reef structures are here reported. Six of these species are new records to the Hawai’ian Porifera catalogue and are suspected to be recent introductions to these islands. Morphological descriptions of the voucher specimens are provided, along with sequencing data of two partitions involving the mitochondrial cytochrome oxidase subunit 1 (COI) marker and a fragment covering partial (18S and 28S) and full (ITS-1, 5.8S and ITS-2) nuclear ribosomal genes. Species delimitations based on genetic distances were calculated to valitate how taxonomic assignments from DNA barcoding aligned with morphological identifications. Of the 60 sequences submitted to GenBank ~88% are the first sequencing records for the corresponding species and genetic marker. This work compiles the first catalogue combining morphological characters with DNA barcoding of Hawai’ian sponges, and contributes to the repository of public databases through the Sponge Barcoding Project initiative. PMID:29267311

Integrative taxonomy of Leptonetela spiders (Araneae, Leptonetidae), with descriptions of 46 new species

PubMed Central

Wang, Chun-Xia; Xu, Xin; Li, Shu-Qiang

2017-01-01

Extreme environments, such as subterranean habitats, are suspected to be responsible for morphologically inseparable cryptic or sibling species and can bias biodiversity assessment. A DNA barcode is a short, standardized DNA sequence used for taxonomic purposes and has the potential to lessen the challenges presented by a biotic inventory. Here, we investigate the diversity of the genus Leptonetela Kratochvíl, 1978 that is endemic to karst systems in Eurasia using DNA barcoding. We analyzed 624 specimens using one mitochondrial gene fragment (COI). The results show that DNA barcoding is an efficient and rapid species identification method in this genus. DNA barcoding gap and automatic barcode gap discovery (ABGD) analyses indicated the existence of 90 species, a result consistent with previous taxonomic hypotheses, and supported the existence of extreme male pedipalpal tibial spine and median apophysis polymorphism in Leptonetela species, with direct implications for the taxonomy of the group and its diversity. Based on the molecular and morphological evidence, we delimit and diagnose 90 Leptonetela species, including the type species Leptonetela kanellisi(Deeleman-Reinhold, 1971). Forty-six of them are previously undescribed. The female of Leptonetela zhai Wang & Li, 2011 is reported for the first time. Leptonetela tianxinensis (Tong & Li, 2008) comb. nov. is transferred from the genus Leptoneta Simon, 1872;the genus Guineta Lin & Li, 2010 syn. nov. is a junior synonym of Leptonetela; Leptonetela gigachela(Lin & Li, 2010) comb. nov. is transferred from Guineta. The genus Sinoneta Lin & Li, 2010 syn. nov. is a junior synonym of Leptonetela; Leptonetela notabilis(Lin & Li, 2010) comb. nov. and Leptonetela sexdigiti(Lin & Li, 2010) comb. nov. are transferred from Sinoneta; Leptonetela sanchahe Wang & Li nom. nov. is proposed as a replacement name for Sinoneta palmata(Chen et al, 2010) because Leptonetela palmata is preoccupied. PMID:29280363

Integrative taxonomy of Leptonetela spiders (Araneae, Leptonetidae), with descriptions of 46 new species.

PubMed

Wang, Chun-Xia; Xu, Xin; Li, Shu-Qiang

2017-11-18

Extreme environments, such as subterranean habitats, are suspected to be responsible for morphologically inseparable cryptic or sibling species and can bias biodiversity assessment. A DNA barcode is a short, standardized DNA sequence used for taxonomic purposes and has the potential to lessen the challenges presented by a biotic inventory. Here, we investigate the diversity of the genus Leptonetela Kratochvíl, 1978 that is endemic to karst systems in Eurasia using DNA barcoding. We analyzed 624 specimens using one mitochondrial gene fragment ( COI ). The results show that DNA barcoding is an efficient and rapid species identification method in this genus. DNA barcoding gap and automatic barcode gap discovery (ABGD) analyses indicated the existence of 90 species, a result consistent with previous taxonomic hypotheses, and supported the existence of extreme male pedipalpal tibial spine and median apophysis polymorphism in Leptonetela species, with direct implications for the taxonomy of the group and its diversity. Based on the molecular and morphological evidence, we delimit and diagnose 90 Leptonetela species, including the type species Leptonetela kanellisi (Deeleman-Reinhold, 1971). Forty-six of them are previously undescribed. The female of Leptonetela zhai Wang & Li, 2011 is reported for the first time. Leptonetela tianxinensis (Tong & Li, 2008) comb. nov. is transferred from the genus Leptoneta Simon, 1872; the genus Guineta Lin & Li, 2010 syn. nov. is a junior synonym of Leptonetela; Leptonetela gigachela (Lin & Li, 2010) comb. nov. is transferred from Guineta . The genus Sinoneta Lin & Li, 2010 syn. nov. is a junior synonym of Leptonetela ; Leptonetela notabilis (Lin & Li, 2010) comb. nov. and Leptonetela sexdigiti (Lin & Li, 2010) comb. nov. are transferred from Sinoneta ; Leptonetela sanchahe Wang & Li nom. nov. is proposed as a replacement name for Sinoneta palmata (Chen et al., 2010) because Leptonetela palmata is preoccupied.

Mayaweckelia troglomorpha, a new subterranean amphipod species from Yucatán state, México (Amphipoda, Hadziidae)

PubMed Central

Angyal, Dorottya; Solís, Efraín Chávez; Magaña, Benjamín; Balázs, Gergely; Simoes, Nuno

2018-01-01

Abstract A detailed description of a new stygobiont species of the amphipod family Hadziidae, Mayaweckelia troglomorpha Angyal, sp. n. is given, based on material collected in four cenotes of Yucatán federal state, México. Morphology was studied under light microscopy and with scanning electron microscopy. Morphological description is complemented with mitochondrial cytochrome c oxidase subunit I (COI) sequences as barcodes, with affinities to the related taxa and with notes on the species’ ecology. Using COI Bayesian inference and genetic distance analyses, we show that the closest relative of the new species is M. cenoticola, forming a monophyletic group referring to the genus Mayaweckelia. Based on the available sequences, we also revealed that Mayaweckelia and Tuluweckelia are sister genera, standing close to the third Yucatán subterranean genus, Bahadzia. The data gathered on the habitat, distribution, abundance, and ecology will contribute to the conservation planning for M. troglomorpha Angyal, sp. n. PMID:29674859

Mayaweckelia troglomorpha, a new subterranean amphipod species from Yucatán state, México (Amphipoda, Hadziidae).

PubMed

Angyal, Dorottya; Solís, Efraín Chávez; Magaña, Benjamín; Balázs, Gergely; Simoes, Nuno

2018-01-01

A detailed description of a new stygobiont species of the amphipod family Hadziidae, Mayaweckelia troglomorpha Angyal, sp. n. is given, based on material collected in four cenotes of Yucatán federal state, México. Morphology was studied under light microscopy and with scanning electron microscopy. Morphological description is complemented with mitochondrial cytochrome c oxidase subunit I (COI) sequences as barcodes, with affinities to the related taxa and with notes on the species' ecology. Using COI Bayesian inference and genetic distance analyses, we show that the closest relative of the new species is M. cenoticola , forming a monophyletic group referring to the genus Mayaweckelia . Based on the available sequences, we also revealed that Mayaweckelia and Tuluweckelia are sister genera, standing close to the third Yucatán subterranean genus, Bahadzia . The data gathered on the habitat, distribution, abundance, and ecology will contribute to the conservation planning for M. troglomorpha Angyal, sp. n.

A regional approach to plant DNA barcoding provides high species resolution of sedges (Carex and Kobresia, Cyperaceae) in the Canadian Arctic Archipelago.

PubMed

Clerc-Blain, Jessica L E; Starr, Julian R; Bull, Roger D; Saarela, Jeffery M

2010-01-01

Previous research on barcoding sedges (Carex) suggested that basic searches within a global barcoding database would probably not resolve more than 60% of the world's some 2000 species. In this study, we take an alternative approach and explore the performance of plant DNA barcoding in the Carex lineage from an explicitly regional perspective. We characterize the utility of a subset of the proposed protein-coding and noncoding plastid barcoding regions (matK, rpoB, rpoC1, rbcL, atpF-atpH, psbK-psbI) for distinguishing species of Carex and Kobresia in the Canadian Arctic Archipelago, a clearly defined eco-geographical region representing 1% of the Earth's landmass. Our results show that matK resolves the greatest number of species of any single-locus (95%), and when combined in a two-locus barcode, it provides 100% species resolution in all but one combination (matK + atpFH) during unweighted pair-group method with arithmetic mean averages (UPGMA) analyses. Noncoding regions were equally or more variable than matK, but as single markers they resolve substantially fewer taxa than matK alone. When difficulties with sequencing and alignment due to microstructural variation in noncoding regions are also considered, our results support other studies in suggesting that protein-coding regions are more practical as barcoding markers. Plastid DNA barcodes are an effective identification tool for species of Carex and Kobresia in the Canadian Arctic Archipelago, a region where the number of co-existing closely related species is limited. We suggest that if a regional approach to plant DNA barcoding was applied on a global scale, it could provide a solution to the generally poor species resolution seen in previous barcoding studies. © 2009 Blackwell Publishing Ltd.

Evidence of new species for malaria vector Anopheles nuneztovari sensu lato in the Brazilian Amazon region.

PubMed

Scarpassa, Vera Margarete; Cunha-Machado, Antonio Saulo; Saraiva, José Ferreira

2016-04-12

Anopheles nuneztovari sensu lato comprises cryptic species in northern South America, and the Brazilian populations encompass distinct genetic lineages within the Brazilian Amazon region. This study investigated, based on two molecular markers, whether these lineages might actually deserve species status. Specimens were collected in five localities of the Brazilian Amazon, including Manaus, Careiro Castanho and Autazes, in the State of Amazonas; Tucuruí, in the State of Pará; and Abacate da Pedreira, in the State of Amapá, and analysed for the COI gene (Barcode region) and 12 microsatellite loci. Phylogenetic analyses were performed using the maximum likelihood (ML) approach. Intra and inter samples genetic diversity were estimated using population genetics analyses, and the genetic groups were identified by means of the ML, Bayesian and factorial correspondence analyses and the Bayesian analysis of population structure. The Barcode region dataset (N = 103) generated 27 haplotypes. The haplotype network suggested three lineages. The ML tree retrieved five monophyletic groups. Group I clustered all specimens from Manaus and Careiro Castanho, the majority of Autazes and a few from Abacate da Pedreira. Group II clustered most of the specimens from Abacate da Pedreira and a few from Autazes and Tucuruí. Group III clustered only specimens from Tucuruí (lineage III), strongly supported (97 %). Groups IV and V clustered specimens of A. nuneztovari s.s. and A. dunhami, strongly (98 %) and weakly (70 %) supported, respectively. In the second phylogenetic analysis, the sequences from GenBank, identified as A. goeldii, clustered to groups I and II, but not to group III. Genetic distances (Kimura-2 parameters) among the groups ranged from 1.60 % (between I and II) to 2.32 % (between I and III). Microsatellite data revealed very high intra-population genetic variability. Genetic distances showed the highest and significant values (P = 0.005) between Tucuruí and all the other samples, and between Abacate da Pedreira and all the other samples. Genetic distances, Bayesian (Structure and BAPS) analyses and FCA suggested three distinct biological groups, supporting the barcode region results. The two markers revealed three genetic lineages for A. nuneztovari s.l. in the Brazilian Amazon region. Lineages I and II may represent genetically distinct groups or species within A. goeldii. Lineage III may represent a new species, distinct from the A. goeldii group, and may be the most ancestral in the Brazilian Amazon. They may have differences in Plasmodium susceptibility and should therefore be investigated further.

Evaluation of candidate barcoding markers in Orinus (Poaceae).

PubMed

Su, X; Liu, Y P; Chen, Z; Chen, K L

2016-04-26

Orinus is an alpine endemic genus of Poaceae. Because of the imperfect specimens, high level of intraspecific morphological variability, and homoplasies of morphological characters, it is relatively difficult to delimitate species of Orinus by using morphology alone. To this end, the DNA barcoding has shown great potential in identifying species. The present study is the first attempt to test the feasibility of four proposed DNA barcoding markers (matK, rbcL, trnH-psbA, and ITS) in identifying four currently revised species of Orinus from the Qinghai-Tibetan Plateau (QTP). Among all the single-barcode candidates, the differentiation power was the highest for the nuclear internal transcribed spacer (ITS), while the chloroplast barcodes matK (M), rbcL (R), and trnH-psbA (H) could not identify the species. Meanwhile, the differentiation efficiency of the nuclear ITS (I) was also higher than any two- or three-locus combination of chloroplast barcodes, or even a combination of ITS and any chloroplast barcode except H + I and R + I. All the combinations of chloroplast barcodes plus the nuclear ITS, H + I, and R + I differentiated the highest portion of species. The highest differentiation rate for the barcodes or barcode combinations examined here was 100% (H + I and R + I). In summary, this case study showed that the nuclear ITS region represents a more promising barcode than any maternally inherited chloroplast region or combination of chloroplast regions in differentiating Orinus species from the QTP. Moreover, combining the ITS region with chloroplast regions may improve the barcoding success rate.

DNA barcoding reveals species level divergence between populations of the microhylid frog genus Arcovomer (Anura: Microhylidae) in the Atlantic Rainforest of southeastern Brazil.

PubMed

Jennings, W Bryan; Wogel, Henrique; Bilate, Marcos; Salles, Rodrigo de O L; Buckup, Paulo A

2016-09-01

The microhylid frogs belonging to the genus Arcovomer have been reported from lowland Atlantic Rainforest in the Brazilian states of Espírito Santo, Rio de Janeiro, and São Paulo. Here, we use DNA barcoding to assess levels of genetic divergence between apparently isolated populations in Espírito Santo and Rio de Janeiro. Our mtDNA data consisting of cytochrome oxidase subunit I (COI) nucleotide sequences reveals 13.2% uncorrected and 30.4% TIM2 + I + Γ corrected genetic divergences between these two populations. This level of divergence exceeds the suggested 10% uncorrected divergence threshold for elevating amphibian populations to candidate species using this marker, which implies that the Espírito Santo population is a species distinct from Arcovomer passarellii. Calibration of our model-corrected sequence divergence estimates suggests that the time of population divergence falls between 12 and 29 million years ago.

Characterisation of Asian Snakehead Murrel Channa striata (Channidae) in Malaysia: An Insight into Molecular Data and Morphological Approach

PubMed Central

Song, Li Min; Munian, Kaviarasu; Abd Rashid, Zulkafli; Bhassu, Subha

2013-01-01

Conservation is imperative for the Asian snakeheads Channa striata, as the species has been overfished due to its high market demand. Using maternal markers (mitochondrial cytochrome c oxidase subunit 1 gene (COI)), we discovered that evolutionary forces that drove population divergence did not show any match between the genetic and morphological divergence pattern. However, there is evidence of incomplete divergence patterns between the Borneo population and the populations from Peninsular Malaysia. This supports the claim of historical coalescence of C. striata during Pleistocene glaciations. Ecological heterogeneity caused high phenotypic variance and was not correlated with genetic variance among the populations. Spatial conservation assessments are required to manage different stock units. Results on DNA barcoding show no evidence of cryptic species in C. striata in Malaysia. The newly obtained sequences add to the database of freshwater fish DNA barcodes and in future will provide information relevant to identification of species. PMID:24396312

Molecular Diet Analysis of Two African Free-Tailed Bats (Molossidae) Using High Throughput Sequencing

PubMed Central

Bohmann, Kristine; Monadjem, Ara; Lehmkuhl Noer, Christina; Rasmussen, Morten; Zeale, Matt R. K.; Clare, Elizabeth; Jones, Gareth; Willerslev, Eske; Gilbert, M. Thomas P.

2011-01-01

Given the diversity of prey consumed by insectivorous bats, it is difficult to discern the composition of their diet using morphological or conventional PCR-based analyses of their faeces. We demonstrate the use of a powerful alternate tool, the use of the Roche FLX sequencing platform to deep-sequence uniquely 5′ tagged insect-generic barcode cytochrome c oxidase I (COI) fragments, that were PCR amplified from faecal pellets of two free-tailed bat species Chaerephon pumilus and Mops condylurus (family: Molossidae). Although the analyses were challenged by the paucity of southern African insect COI sequences in the GenBank and BOLD databases, similarity to existing collections allowed the preliminary identification of 25 prey families from six orders of insects within the diet of C. pumilus, and 24 families from seven orders within the diet of M. condylurus. Insects identified to families within the orders Lepidoptera and Diptera were widely present among the faecal samples analysed. The two families that were observed most frequently were Noctuidae and Nymphalidae (Lepidoptera). Species-level analysis of the data was accomplished using novel bioinformatics techniques for the identification of molecular operational taxonomic units (MOTU). Based on these analyses, our data provide little evidence of resource partitioning between sympatric M. condylurus and C. pumilus in the Simunye region of Swaziland at the time of year when the samples were collected, although as more complete databases against which to compare the sequences are generated this may have to be re-evaluated. PMID:21731749

Limnodrilus sulphurensis n. sp., from a sulfur cave in Colorado, USA, with notes on the morphologically similar L. profundicola (Clitellata, Naididae, Tubificinae).

PubMed

Fend, Steven V; Liu, Yingkui; Steinmann, David; Giere, Olav; Barton, Hazel A; Luiszer, Fred; Erséus, Christer

2016-01-18

A new species of the tubificine genus Limnodrilus is described and COI barcoded from Sulphur Cave and associated springs in Colorado, USA. The habitats are characterized by high sulfide concentrations. The new species, L. sulphurensis, is distinguished from all congeners by the elongate, nearly parallel teeth of chaetae in its anterior segments. It has a penis sheath resembling that of L. profundicola; consequently, museum specimens and new collections are examined here to resolve some of the taxonomic confusion surrounding that widespread, but uncommon species.

Three Taxa in One: Cryptic Diversity in the Black Fly Simulium nobile (Diptera: Simuliidae) in Southeast Asia.

PubMed

Low, Van Lun; Takaoka, Hiroyuki; Pramual, Pairot; Adler, Peter H; Ya'cob, Zubaidah; Chen, Chee Dhang; Yotopranoto, Subagyo; Zaid, Adnan; Hadi, Upik Kesumawati; Lardizabal, Maria Lourdes; Nasruddin-Roshidi, Affan; Sofian-Azirun, Mohd

2016-07-01

We access the molecular diversity of the black fly Simulium nobile De Mejiere, using the universal cytochrome c oxidase subunit I (COI) barcoding gene, across its distributional range in Southeast Asia. Our phylogenetic analyses recovered three well-supported mitochondrial lineages of S. nobile, suggesting the presence of cryptic species. Lineage A is composed of a population from Sabah, East Malaysia (Borneo); lineage B represents the type population from Java, Indonesia; and lineage C includes populations from the mainland of Southeast Asia (Peninsular Malaysia and Thailand). The genetic variation of lineage C on the mainland is greater than that of lineages A and B on the islands of Borneo and Java. Our study highlights the value of a molecular approach in assessing species status of simuliids in geographically distinct regions. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

The Hidden Diversity of Zanclea Associated with Scleractinians Revealed by Molecular Data

PubMed Central

Montano, Simone; Maggioni, Davide; Arrigoni, Roberto; Seveso, Davide; Puce, Stefania; Galli, Paolo

2015-01-01

Scleractinian reef corals have recently been acknowledged as the most numerous host group found in association with hydroids belonging to the Zanclea genus. However, knowledge of the molecular phylogenetic relationships among Zanclea species associated with scleractinians is just beginning. This study, using the nuclear 28S rDNA region and the fast-evolving mitochondrial 16S rRNA and COI genes, provides the most comprehensive phylogenetic reconstruction of the genus Zanclea with a particular focus on the genetic diversity among Zanclea specimens associated with 13 scleractinian genera. The monophyly of Zanclea associated with scleractinians was strongly supported in all nuclear and mitochondrial phylogenetic reconstructions. Furthermore, a combined mitochondrial 16S and COI phylogenetic tree revealed a multitude of hidden molecular lineages within this group (Clades I, II, III, V, VI, VII, and VIII), suggesting the existence of both host-generalist and genus-specific lineages of Zanclea associated with scleractinians. In addition to Z. gallii living in association with the genus Acropora, we discovered four well-supported lineages (Clades I, II, III, and VII), each one forming a strict association with a single scleractinian genus, including sequences of Zanclea associated with Montipora from two geographically separated areas (Maldives and Taiwan). Two host-generalist Zanclea lineages were also observed, and one of them was formed by Zanclea specimens symbiotic with seven scleractinian genera (Clade VIII). We also found that the COI gene allows the recognition of separated hidden lineages in agreement with the commonly recommended mitochondrial 16S as a DNA barcoding gene for Hydrozoa and shows reasonable potential for phylogenetic and evolutionary analyses in the genus Zanclea. Finally, as no DNA sequences are available for the majority of the nominal Zanclea species known, we note that they will be necessary to elucidate the diversity of the Zanclea-scleractinian association. PMID:26207903

Taxonomic Notes on Nasutitermes and Bulbitermes (Termitidae, Nasutitermitinae) from the Sunda region of Southeast Asia based on morphological and molecular characters

PubMed Central

Syaukani; Thompson, Graham J.

2011-01-01

Abstract The Sunda region of Southeastern Asia is rich in termite fauna, but termites from this region have been poorly described. In this study, we described eight species from two diverse genera from this region, and from the family Termitidae. We describe Bulbitermes 4 spp. and Nasutitermes 4 spp. from new field collections. Where possible we examine original holotype specimens, and describe the essential morphological characters for soldier and worker castes. We devise two new bifurcating keys to guide the field identification of each species. In addition, we develop a nucleotide sequence profile for the COI gene. From this molecular character matrix, we use Neighbour-Joining analysis to test the monophyly of each morphospecies and genus. We find that the morphological and molecular characters are highly concordant, whereby all taxa appear to represent distinct molecular clades. For termites, there is therefore agreement between the morphological taxonomic characters used to sort species from a bifurcating key and the molecular taxonomic characters used to sort species on a bifurcating tree. This joint analysis suggests that DNA barcoding holds considerable promise for termite taxonomy, especially for diverse clades like Bulbitermes and Nasutitermes for which a global morphological key would be intractable. PMID:22287894

DNA barcode analysis of butterfly species from Pakistan points towards regional endemism

PubMed Central

Ashfaq, Muhammad; Akhtar, Saleem; Khan, Arif M; Adamowicz, Sarah J; Hebert, Paul D N

2013-01-01

DNA barcodes were obtained for 81 butterfly species belonging to 52 genera from sites in north-central Pakistan to test the utility of barcoding for their identification and to gain a better understanding of regional barcode variation. These species represent 25% of the butterfly fauna of Pakistan and belong to five families, although the Nymphalidae were dominant, comprising 38% of the total specimens. Barcode analysis showed that maximum conspecific divergence was 1.6%, while there was 1.7–14.3% divergence from the nearest neighbour species. Barcode records for 55 species showed <2% sequence divergence to records in the Barcode of Life Data Systems (BOLD), but only 26 of these cases involved specimens from neighbouring India and Central Asia. Analysis revealed that most species showed little incremental sequence variation when specimens from other regions were considered, but a threefold increase was noted in a few cases. There was a clear gap between maximum intraspecific and minimum nearest neighbour distance for all 81 species. Neighbour-joining cluster analysis showed that members of each species formed a monophyletic cluster with strong bootstrap support. The barcode results revealed two provisional species that could not be clearly linked to known taxa, while 24 other species gained their first coverage. Future work should extend the barcode reference library to include all butterfly species from Pakistan as well as neighbouring countries to gain a better understanding of regional variation in barcode sequences in this topographically and climatically complex region. PMID:23789612

A DNA barcode for land plants.

PubMed

2009-08-04

DNA barcoding involves sequencing a standard region of DNA as a tool for species identification. However, there has been no agreement on which region(s) should be used for barcoding land plants. To provide a community recommendation on a standard plant barcode, we have compared the performance of 7 leading candidate plastid DNA regions (atpF-atpH spacer, matK gene, rbcL gene, rpoB gene, rpoC1 gene, psbK-psbI spacer, and trnH-psbA spacer). Based on assessments of recoverability, sequence quality, and levels of species discrimination, we recommend the 2-locus combination of rbcL+matK as the plant barcode. This core 2-locus barcode will provide a universal framework for the routine use of DNA sequence data to identify specimens and contribute toward the discovery of overlooked species of land plants.

A DNA barcode for land plants

PubMed Central

Hollingsworth, Peter M.; Forrest, Laura L.; Spouge, John L.; Hajibabaei, Mehrdad; Ratnasingham, Sujeevan; van der Bank, Michelle; Chase, Mark W.; Cowan, Robyn S.; Erickson, David L.; Fazekas, Aron J.; Graham, Sean W.; James, Karen E.; Kim, Ki-Joong; Kress, W. John; Schneider, Harald; van AlphenStahl, Jonathan; Barrett, Spencer C.H.; van den Berg, Cassio; Bogarin, Diego; Burgess, Kevin S.; Cameron, Kenneth M.; Carine, Mark; Chacón, Juliana; Clark, Alexandra; Clarkson, James J.; Conrad, Ferozah; Devey, Dion S.; Ford, Caroline S.; Hedderson, Terry A.J.; Hollingsworth, Michelle L.; Husband, Brian C.; Kelly, Laura J.; Kesanakurti, Prasad R.; Kim, Jung Sung; Kim, Young-Dong; Lahaye, Renaud; Lee, Hae-Lim; Long, David G.; Madriñán, Santiago; Maurin, Olivier; Meusnier, Isabelle; Newmaster, Steven G.; Park, Chong-Wook; Percy, Diana M.; Petersen, Gitte; Richardson, James E.; Salazar, Gerardo A.; Savolainen, Vincent; Seberg, Ole; Wilkinson, Michael J.; Yi, Dong-Keun; Little, Damon P.

2009-01-01

DNA barcoding involves sequencing a standard region of DNA as a tool for species identification. However, there has been no agreement on which region(s) should be used for barcoding land plants. To provide a community recommendation on a standard plant barcode, we have compared the performance of 7 leading candidate plastid DNA regions (atpF–atpH spacer, matK gene, rbcL gene, rpoB gene, rpoC1 gene, psbK–psbI spacer, and trnH–psbA spacer). Based on assessments of recoverability, sequence quality, and levels of species discrimination, we recommend the 2-locus combination of rbcL+matK as the plant barcode. This core 2-locus barcode will provide a universal framework for the routine use of DNA sequence data to identify specimens and contribute toward the discovery of overlooked species of land plants. PMID:19666622

Identifying species of moths (Lepidoptera) from Baihua Mountain, Beijing, China, using DNA barcodes

PubMed Central

Liu, Xiao F; Yang, Cong H; Han, Hui L; Ward, Robert D; Zhang, Ai-bing

2014-01-01

DNA barcoding has become a promising means for the identification of organisms of all life-history stages. Currently, distance-based and tree-based methods are most widely used to define species boundaries and uncover cryptic species. However, there is no universal threshold of genetic distance values that can be used to distinguish taxonomic groups. Alternatively, DNA barcoding can deploy a “character-based” method, whereby species are identified through the discrete nucleotide substitutions. Our research focuses on the delimitation of moth species using DNA-barcoding methods. We analyzed 393 Lepidopteran specimens belonging to 80 morphologically recognized species with a standard cytochrome c oxidase subunit I (COI) sequencing approach, and deployed tree-based, distance-based, and diagnostic character-based methods to identify the taxa. The tree-based method divided the 393 specimens into 79 taxa (species), and the distance-based method divided them into 84 taxa (species). Although the diagnostic character-based method found only 39 so-identifiable species in the 80 species, with a reduction in sample size the accuracy rate substantially improved. For example, in the Arctiidae subset, all 12 species had diagnostics characteristics. Compared with traditional morphological method, molecular taxonomy performed well. All three methods enable the rapid delimitation of species, although they have different characteristics and different strengths. The tree-based and distance-based methods can be used for accurate species identification and biodiversity studies in large data sets, while the character-based method performs well in small data sets and can also be used as the foundation of species-specific biochips. PMID:25360280

Potential efficacy of mitochondrial genes for animal DNA barcoding: a case study using eutherian mammals.

PubMed

Luo, Arong; Zhang, Aibing; Ho, Simon Yw; Xu, Weijun; Zhang, Yanzhou; Shi, Weifeng; Cameron, Stephen L; Zhu, Chaodong

2011-01-28

A well-informed choice of genetic locus is central to the efficacy of DNA barcoding. Current DNA barcoding in animals involves the use of the 5' half of the mitochondrial cytochrome oxidase 1 gene (CO1) to diagnose and delimit species. However, there is no compelling a priori reason for the exclusive focus on this region, and it has been shown that it performs poorly for certain animal groups. To explore alternative mitochondrial barcoding regions, we compared the efficacy of the universal CO1 barcoding region with the other mitochondrial protein-coding genes in eutherian mammals. Four criteria were used for this comparison: the number of recovered species, sequence variability within and between species, resolution to taxonomic levels above that of species, and the degree of mutational saturation. Based on 1,179 mitochondrial genomes of eutherians, we found that the universal CO1 barcoding region is a good representative of mitochondrial genes as a whole because the high species-recovery rate (> 90%) was similar to that of other mitochondrial genes, and there were no significant differences in intra- or interspecific variability among genes. However, an overlap between intra- and interspecific variability was still problematic for all mitochondrial genes. Our results also demonstrated that any choice of mitochondrial gene for DNA barcoding failed to offer significant resolution at higher taxonomic levels. We suggest that the CO1 barcoding region, the universal DNA barcode, is preferred among the mitochondrial protein-coding genes as a molecular diagnostic at least for eutherian species identification. Nevertheless, DNA barcoding with this marker may still be problematic for certain eutherian taxa and our approach can be used to test potential barcoding loci for such groups.

Potential efficacy of mitochondrial genes for animal DNA barcoding: a case study using eutherian mammals

PubMed Central

2011-01-01

Background A well-informed choice of genetic locus is central to the efficacy of DNA barcoding. Current DNA barcoding in animals involves the use of the 5' half of the mitochondrial cytochrome oxidase 1 gene (CO1) to diagnose and delimit species. However, there is no compelling a priori reason for the exclusive focus on this region, and it has been shown that it performs poorly for certain animal groups. To explore alternative mitochondrial barcoding regions, we compared the efficacy of the universal CO1 barcoding region with the other mitochondrial protein-coding genes in eutherian mammals. Four criteria were used for this comparison: the number of recovered species, sequence variability within and between species, resolution to taxonomic levels above that of species, and the degree of mutational saturation. Results Based on 1,179 mitochondrial genomes of eutherians, we found that the universal CO1 barcoding region is a good representative of mitochondrial genes as a whole because the high species-recovery rate (> 90%) was similar to that of other mitochondrial genes, and there were no significant differences in intra- or interspecific variability among genes. However, an overlap between intra- and interspecific variability was still problematic for all mitochondrial genes. Our results also demonstrated that any choice of mitochondrial gene for DNA barcoding failed to offer significant resolution at higher taxonomic levels. Conclusions We suggest that the CO1 barcoding region, the universal DNA barcode, is preferred among the mitochondrial protein-coding genes as a molecular diagnostic at least for eutherian species identification. Nevertheless, DNA barcoding with this marker may still be problematic for certain eutherian taxa and our approach can be used to test potential barcoding loci for such groups. PMID:21276253

Antispila oinophylla new species (Lepidoptera, Heliozelidae), a new North American grapevine leafminer invading Italian vineyards: taxonomy, DNA barcodes and life cycle

PubMed Central

van Nieukerken, Erik J.; Wagner, David L.; Baldessari, Mario; Mazzon, Luca; Angeli, Gino; Girolami, Vicenzo; Duso, Carlo; Doorenweerd, Camiel

2012-01-01

Abstract A grapevine leafminer Antispila oinophylla van Nieukerken & Wagner, sp. n., is described both from eastern North America (type locality: Georgia) and as a new important invader in North Italian vineyards (Trentino and Veneto Region) since 2006. The species is closely related to, and previously confused with Antispila ampelopsifoliella Chambers, 1874, a species feeding on Virginia creeper Parthenocissus quinquefolia (L.) Planchon., and both are placed in an informal Antispila ampelopsifoliella group. Wing pattern, genitalia, and DNA barcode data all confirm the conspecificity of native North American populations and Italian populations. COI barcodes differ by only 0–1.23%, indicating that the Italian populations are recently established from eastern North America. The new species feeds on various wild Vitis species in North America, on cultivated Vitis vinifera L. in Italy, and also on Parthenocissus quinquefolia in Italy. North American Antispila feeding on Parthenocissus include at least two other species, one of which is Antispila ampelopsifoliella. Morphology and biology of the new species are contrasted with those of North American Antispila Hübner, 1825 species and European Holocacista rivillei (Stainton, 1855). The source population of the introduction is unknown, but cases with larvae or pupae, attached to imported plants, are a likely possibility. DNA barcodes of the three European grapevine leafminers and those of all examined Heliozelidae are highly diagnostic. North American Vitaceae-feeding Antispila form two species complexes and include several as yet unnamed taxa. The identity of three out of the four previously described North American Vitaceae-feeding species cannot be unequivocally determined without further revision, but these are held to be different from Antispila oinophylla. In Italy the biology of Antispila oinophylla was studied in a vineyard in the Trento Province (Trentino-Alto Adige Region) in 2008 and 2009. Mature larvae overwinter inside their cases, fixed to vine trunks or training stakes. The first generation flies in June. An additional generation occurs from mid-August onwards. The impact of the pest in this vineyard was significant with more than 90% of leaves infested in mid-summer. Since the initial discovery in 2006, the pest spread to several additional Italian provinces, in 2010 the incidence of infestation was locally high in commercial vineyards. Preliminary phylogenetic analyses suggest that Antispila is paraphyletic, and that the Antispila ampelopsifoliella group is related to Coptodisca Walsingham, 1895, Holocacista Walsingham & Durrant, 1909 and Antispilina Hering, 1941, all of which possess reduced wing venation. Vitaceae may be the ancestral hostplant family for modern Heliozelidae. PMID:22408380

DNA barcode analysis of butterfly species from Pakistan points towards regional endemism.

PubMed

Ashfaq, Muhammad; Akhtar, Saleem; Khan, Arif M; Adamowicz, Sarah J; Hebert, Paul D N

2013-09-01

DNA barcodes were obtained for 81 butterfly species belonging to 52 genera from sites in north-central Pakistan to test the utility of barcoding for their identification and to gain a better understanding of regional barcode variation. These species represent 25% of the butterfly fauna of Pakistan and belong to five families, although the Nymphalidae were dominant, comprising 38% of the total specimens. Barcode analysis showed that maximum conspecific divergence was 1.6%, while there was 1.7-14.3% divergence from the nearest neighbour species. Barcode records for 55 species showed <2% sequence divergence to records in the Barcode of Life Data Systems (BOLD), but only 26 of these cases involved specimens from neighbouring India and Central Asia. Analysis revealed that most species showed little incremental sequence variation when specimens from other regions were considered, but a threefold increase was noted in a few cases. There was a clear gap between maximum intraspecific and minimum nearest neighbour distance for all 81 species. Neighbour-joining cluster analysis showed that members of each species formed a monophyletic cluster with strong bootstrap support. The barcode results revealed two provisional species that could not be clearly linked to known taxa, while 24 other species gained their first coverage. Future work should extend the barcode reference library to include all butterfly species from Pakistan as well as neighbouring countries to gain a better understanding of regional variation in barcode sequences in this topographically and climatically complex region. © 2013 The Authors. Molecular Ecology Resources published by John Wiley & Sons Ltd.

Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi

USDA-ARS?s Scientific Manuscript database

Six DNA regions were evaluated in a multi-national, multi-laboratory consortium as potential DNA barcodes for Fungi, the second largest kingdom of eukaryotic life. The region of the mitochondrial cytochrome c oxidase subunit 1 used as the animal barcode was excluded as a potential marker, because it...

Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi

PubMed Central

Schoch, Conrad L.; Seifert, Keith A.; Huhndorf, Sabine; Robert, Vincent; Spouge, John L.; Levesque, C. André; Chen, Wen; Bolchacova, Elena; Voigt, Kerstin; Crous, Pedro W.; Miller, Andrew N.; Wingfield, Michael J.; Aime, M. Catherine; An, Kwang-Deuk; Bai, Feng-Yan; Barreto, Robert W.; Begerow, Dominik; Bergeron, Marie-Josée; Blackwell, Meredith; Boekhout, Teun; Bogale, Mesfin; Boonyuen, Nattawut; Burgaz, Ana R.; Buyck, Bart; Cai, Lei; Cai, Qing; Cardinali, G.; Chaverri, Priscila; Coppins, Brian J.; Crespo, Ana; Cubas, Paloma; Cummings, Craig; Damm, Ulrike; de Beer, Z. Wilhelm; de Hoog, G. Sybren; Del-Prado, Ruth; Dentinger, Bryn; Diéguez-Uribeondo, Javier; Divakar, Pradeep K.; Douglas, Brian; Dueñas, Margarita; Duong, Tuan A.; Eberhardt, Ursula; Edwards, Joan E.; Elshahed, Mostafa S.; Fliegerova, Katerina; Furtado, Manohar; García, Miguel A.; Ge, Zai-Wei; Griffith, Gareth W.; Griffiths, K.; Groenewald, Johannes Z.; Groenewald, Marizeth; Grube, Martin; Gryzenhout, Marieka; Guo, Liang-Dong; Hagen, Ferry; Hambleton, Sarah; Hamelin, Richard C.; Hansen, Karen; Harrold, Paul; Heller, Gregory; Herrera, Cesar; Hirayama, Kazuyuki; Hirooka, Yuuri; Ho, Hsiao-Man; Hoffmann, Kerstin; Hofstetter, Valérie; Högnabba, Filip; Hollingsworth, Peter M.; Hong, Seung-Beom; Hosaka, Kentaro; Houbraken, Jos; Hughes, Karen; Huhtinen, Seppo; Hyde, Kevin D.; James, Timothy; Johnson, Eric M.; Johnson, Joan E.; Johnston, Peter R.; Jones, E.B. Gareth; Kelly, Laura J.; Kirk, Paul M.; Knapp, Dániel G.; Kõljalg, Urmas; Kovács, Gábor M.; Kurtzman, Cletus P.; Landvik, Sara; Leavitt, Steven D.; Liggenstoffer, Audra S.; Liimatainen, Kare; Lombard, Lorenzo; Luangsa-ard, J. Jennifer; Lumbsch, H. Thorsten; Maganti, Harinad; Maharachchikumbura, Sajeewa S. N.; Martin, María P.; May, Tom W.; McTaggart, Alistair R.; Methven, Andrew S.; Meyer, Wieland; Moncalvo, Jean-Marc; Mongkolsamrit, Suchada; Nagy, László G.; Nilsson, R. Henrik; Niskanen, Tuula; Nyilasi, Ildikó; Okada, Gen; Okane, Izumi; Olariaga, Ibai; Otte, Jürgen; Papp, Tamás; Park, Duckchul; Petkovits, Tamás; Pino-Bodas, Raquel; Quaedvlieg, William; Raja, Huzefa A.; Redecker, Dirk; Rintoul, Tara L.; Ruibal, Constantino; Sarmiento-Ramírez, Jullie M.; Schmitt, Imke; Schüßler, Arthur; Shearer, Carol; Sotome, Kozue; Stefani, Franck O.P.; Stenroos, Soili; Stielow, Benjamin; Stockinger, Herbert; Suetrong, Satinee; Suh, Sung-Oui; Sung, Gi-Ho; Suzuki, Motofumi; Tanaka, Kazuaki; Tedersoo, Leho; Telleria, M. Teresa; Tretter, Eric; Untereiner, Wendy A.; Urbina, Hector; Vágvölgyi, Csaba; Vialle, Agathe; Vu, Thuy Duong; Walther, Grit; Wang, Qi-Ming; Wang, Yan; Weir, Bevan S.; Weiß, Michael; White, Merlin M.; Xu, Jianping; Yahr, Rebecca; Yang, Zhu L.; Yurkov, Andrey; Zamora, Juan-Carlos; Zhang, Ning; Zhuang, Wen-Ying; Schindel, David

2012-01-01

Six DNA regions were evaluated as potential DNA barcodes for Fungi, the second largest kingdom of eukaryotic life, by a multinational, multilaboratory consortium. The region of the mitochondrial cytochrome c oxidase subunit 1 used as the animal barcode was excluded as a potential marker, because it is difficult to amplify in fungi, often includes large introns, and can be insufficiently variable. Three subunits from the nuclear ribosomal RNA cistron were compared together with regions of three representative protein-coding genes (largest subunit of RNA polymerase II, second largest subunit of RNA polymerase II, and minichromosome maintenance protein). Although the protein-coding gene regions often had a higher percent of correct identification compared with ribosomal markers, low PCR amplification and sequencing success eliminated them as candidates for a universal fungal barcode. Among the regions of the ribosomal cistron, the internal transcribed spacer (ITS) region has the highest probability of successful identification for the broadest range of fungi, with the most clearly defined barcode gap between inter- and intraspecific variation. The nuclear ribosomal large subunit, a popular phylogenetic marker in certain groups, had superior species resolution in some taxonomic groups, such as the early diverging lineages and the ascomycete yeasts, but was otherwise slightly inferior to the ITS. The nuclear ribosomal small subunit has poor species-level resolution in fungi. ITS will be formally proposed for adoption as the primary fungal barcode marker to the Consortium for the Barcode of Life, with the possibility that supplementary barcodes may be developed for particular narrowly circumscribed taxonomic groups. PMID:22454494

Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi.

PubMed

Schoch, Conrad L; Seifert, Keith A; Huhndorf, Sabine; Robert, Vincent; Spouge, John L; Levesque, C André; Chen, Wen

2012-04-17

Six DNA regions were evaluated as potential DNA barcodes for Fungi, the second largest kingdom of eukaryotic life, by a multinational, multilaboratory consortium. The region of the mitochondrial cytochrome c oxidase subunit 1 used as the animal barcode was excluded as a potential marker, because it is difficult to amplify in fungi, often includes large introns, and can be insufficiently variable. Three subunits from the nuclear ribosomal RNA cistron were compared together with regions of three representative protein-coding genes (largest subunit of RNA polymerase II, second largest subunit of RNA polymerase II, and minichromosome maintenance protein). Although the protein-coding gene regions often had a higher percent of correct identification compared with ribosomal markers, low PCR amplification and sequencing success eliminated them as candidates for a universal fungal barcode. Among the regions of the ribosomal cistron, the internal transcribed spacer (ITS) region has the highest probability of successful identification for the broadest range of fungi, with the most clearly defined barcode gap between inter- and intraspecific variation. The nuclear ribosomal large subunit, a popular phylogenetic marker in certain groups, had superior species resolution in some taxonomic groups, such as the early diverging lineages and the ascomycete yeasts, but was otherwise slightly inferior to the ITS. The nuclear ribosomal small subunit has poor species-level resolution in fungi. ITS will be formally proposed for adoption as the primary fungal barcode marker to the Consortium for the Barcode of Life, with the possibility that supplementary barcodes may be developed for particular narrowly circumscribed taxonomic groups.

Mesobiotus philippinicus sp. nov., the first limnoterrestrial tardigrade from the Philippines.

PubMed

Mapalo, Marc A; Stec, Daniel; Mirano-Bascos, Denise; Michalczyk, Łukasz

2016-06-20

The limnoterrestrial tardigrade fauna of the Philippines is completely unknown. In this paper, we describe the first ever limnoterrestrial water bear species from this southeast Asian country, Mesobiotus philippinicus sp. nov., found in a moss sample collected in Quezon City. Apart from morphometrics and imaging in light microscopy, we also analysed the new species under scanning electron microscope and sequenced four DNA markers differing in mutation rates, three nuclear (18S rRNA, 28S rRNA, and ITS-2) and one mitochondrial (COI). This allowed not only a detailed description but also provided barcodes to aid future species identification. The new species belongs to the harmsworthi group and is most similar to M. diffusus (Binda & Pilato, 1987), M. pseudocoronatus (Pilato et al., 2006), M. montanus (Murray, 1910) and M. mottai (Binda & Pilato, 1994), but differs from these species by whorled egg processes and dimensions of some morphometric traits. The 28S rRNA, ITS-2 and COI sequences presented in this paper are the first published DNA sequences for the genus Mesobiotus.

The taxonomic status of Japanese threadfin bream Nemipterus japonicus (Bloch, 1791) (Perciformes: Nemipteridae) with a redescription of this species from the south china sea based on morphology and DNA barcodes

NASA Astrophysics Data System (ADS)

Ning, Ping; Sha, Zhongli; Hebert, Paul D. N.; Russell, Barry

2015-02-01

Because of its importance as a food source, Nemipterus japonicus (Bloch, 1791) (Nemipteridae) or Japanese threadfin bream is the best studied of these taxa, and numerous investigations have examined its fisheries, its biology and biochemistry. Despite such intensive work, the taxonomic status of N. japonicus has never been seriously questioned and it is regarded as a common species, widely distributed throughout the Indo-Western Pacific Ocean. In fact, Bloch's description of the type specimen of N. japonicus has ambiguous collection data and lacks a designation for the type locality, though it is probably Java. In this paper, DNA barcode results based on COI gene support the existence of two geographically separated lineages of the Japanese threadfin bream, both being an Indian Ocean and western Pacific lineage, with 2.7% sequence divergence, and the results indicate a possible existing of some cryptic species. The two lineages also possess a diagnostic difference in their belly color, with specimens in the South China Sea having a silver belly, while those from the Indian Ocean isolate specimen have a yellow coloration. Based upon new collections from the South China Sea, this species from the western Pacific is morphologically redescribed and its details of DNA barcode diversity are shown for the future investigations.

Using DNA Barcodes to Identify Road-Killed Animals in Two Atlantic Forest Nature Reserves, Brazil

PubMed Central

Klippel, Angélica H.; Oliveira, Pablo V.; Britto, Karollini B.; Freire, Bárbara F.; Moreno, Marcel R.; dos Santos, Alexandre R.; Banhos, Aureo; Paneto, Greiciane G.

2015-01-01

Road mortality is the leading source of biodiversity loss in the world, especially due to fragmentation of natural habitats and loss of wildlife. The survey of the main species victims of roadkill is of fundamental importance for the better understanding of the problem, being necessary, for this, the correct species identification. The aim of this study was to verify if DNA barcodes can be applied to identify road-killed samples that often cannot be determined morphologically. For this purpose, 222 vertebrate samples were collected in a stretch of the BR-101 highway that crosses two Discovery Coast Atlantic Forest Natural Reserves, the Sooretama Biological Reserve and the Vale Natural Reserve, in Espírito Santo, Brazil. The mitochondrial COI gene was amplified, sequenced and confronted with the BOLD database. It was possible to identify 62.16% of samples, totaling 62 different species, including Pyrrhura cruentata, Chaetomys subspinosus, Puma yagouaroundi and Leopardus wiedii considered Vulnerable in the National Official List of Species of Endangered Wildlife. The most commonly identified animals were a bat (Molossus molossus), an opossum (Didelphis aurita) and a frog (Trachycephalus mesophaeus) species. Only one reptile was identified using the technique, probably due to lack of reference sequences in BOLD. These data may contribute to a better understanding of the impact of roads on species biodiversity loss and to introduce the DNA barcode technique to road ecology scenarios. PMID:26244644

A new Heraclides swallowtail (Lepidoptera, Papilionidae) from North America is recognized by the pattern on its neck

PubMed Central

Shiraiwa, Kojiro; Cong, Qian; Grishin, Nick V.

2014-01-01

Abstract Heraclides rumiko Shiraiwa & Grishin, sp. n. is described from southwestern United States, Mexico, and Central America (type locality: USA, Texas, Duval County). It is closely allied to Heraclides cresphontes (Cramer, 1777) and the two species are sympatric in central Texas. The new species is diagnosed by male genitalia and exhibits a nearly 3% difference from Heraclides cresphontes in the COI DNA barcode sequence of mitochondrial DNA. The two Heraclides species can usually be told apart by the shape and size of yellow spots on the neck, by the wing shape, and the details of wing patterns. “Western Giant Swallowtail” is proposed as the English name for Heraclides rumiko. To stabilize nomenclature, neotype for Papilio cresphontes Cramer, 1777, an eastern United States species, is designated from Brooklyn, New York, USA; and lectotype for Papilio thoas Linnaeus, 1771 is designated from Suriname. We sequenced DNA barcodes and ID tags of nearly 400 Papilionini specimens completing coverage of all Heraclides species. Comparative analyses of DNA barcodes, genitalia, and facies suggest that Heraclides oviedo (Gundlach, 1866), reinstated status, is a species-level taxon rather than a subspecies of Heraclides thoas (Linnaeus, 1771); and Heraclides pallas (G. Gray, [1853]), reinstated status, with its subspecies Heraclides Papilio bajaensis (J. Brown & Faulkner, 1992), comb. n., and Heraclides anchicayaensis Constantino, Le Crom & Salazar, 2002, stat. n., are not conspecific with Heraclides astyalus (Godart, 1819). PMID:25610342

Evolutionary Diversification of Banded Tube-Dwelling Anemones (Cnidaria; Ceriantharia; Isarachnanthus) in the Atlantic Ocean

PubMed Central

Stampar, Sergio N.; Maronna, Maximiliano M.; Vermeij, Mark J. A.; Silveira, Fabio L. d.; Morandini, André C.

2012-01-01

The use of molecular data for species delimitation in Anthozoa is still a very delicate issue. This is probably due to the low genetic variation found among the molecular markers (primarily mitochondrial) commonly used for Anthozoa. Ceriantharia is an anthozoan group that has not been tested for genetic divergence at the species level. Recently, all three Atlantic species described for the genus Isarachnanthus of Atlantic Ocean, were deemed synonyms based on morphological simmilarities of only one species: Isarachnanthus maderensis. Here, we aimed to verify whether genetic relationships (using COI, 16S, ITS1 and ITS2 molecular markers) confirmed morphological affinities among members of Isarachnanthus from different regions across the Atlantic Ocean. Results from four DNA markers were completely congruent and revealed that two different species exist in the Atlantic Ocean. The low identification success and substantial overlap between intra and interspecific COI distances render the Anthozoa unsuitable for DNA barcoding, which is not true for Ceriantharia. In addition, genetic divergence within and between Ceriantharia species is more similar to that found in Medusozoa (Hydrozoa and Scyphozoa) than Anthozoa and Porifera that have divergence rates similar to typical metazoans. The two genetic species could also be separated based on micromorphological characteristics of their cnidomes. Using a specimen of Isarachnanthus bandanensis from Pacific Ocean as an outgroup, it was possible to estimate the minimum date of divergence between the clades. The cladogenesis event that formed the species of the Atlantic Ocean is estimated to have occured around 8.5 million years ago (Miocene) and several possible speciation scenarios are discussed. PMID:22815928

Evolutionary diversification of banded tube-dwelling anemones (Cnidaria; Ceriantharia; Isarachnanthus) in the Atlantic Ocean.

PubMed

Stampar, Sergio N; Maronna, Maximiliano M; Vermeij, Mark J A; Silveira, Fabio L d; Morandini, André C

2012-01-01

The use of molecular data for species delimitation in Anthozoa is still a very delicate issue. This is probably due to the low genetic variation found among the molecular markers (primarily mitochondrial) commonly used for Anthozoa. Ceriantharia is an anthozoan group that has not been tested for genetic divergence at the species level. Recently, all three Atlantic species described for the genus Isarachnanthus of Atlantic Ocean, were deemed synonyms based on morphological simmilarities of only one species: Isarachnanthus maderensis. Here, we aimed to verify whether genetic relationships (using COI, 16S, ITS1 and ITS2 molecular markers) confirmed morphological affinities among members of Isarachnanthus from different regions across the Atlantic Ocean. Results from four DNA markers were completely congruent and revealed that two different species exist in the Atlantic Ocean. The low identification success and substantial overlap between intra and interspecific COI distances render the Anthozoa unsuitable for DNA barcoding, which is not true for Ceriantharia. In addition, genetic divergence within and between Ceriantharia species is more similar to that found in Medusozoa (Hydrozoa and Scyphozoa) than Anthozoa and Porifera that have divergence rates similar to typical metazoans. The two genetic species could also be separated based on micromorphological characteristics of their cnidomes. Using a specimen of Isarachnanthus bandanensis from Pacific Ocean as an outgroup, it was possible to estimate the minimum date of divergence between the clades. The cladogenesis event that formed the species of the Atlantic Ocean is estimated to have occured around 8.5 million years ago (Miocene) and several possible speciation scenarios are discussed.

How many species and under what names? Using DNA barcoding and GenBank data for west Central African amphibian conservation

PubMed Central

Mulcahy, Daniel G.; Vanthomme, Hadrien; Tobi, Elie; Wynn, Addison H.; Zimkus, Breda M.; McDiarmid, Roy W.

2017-01-01

Development projects in west Central Africa are proceeding at an unprecedented rate, often with little concern for their effects on biodiversity. In an attempt to better understand potential impacts of a road development project on the anuran amphibian community, we conducted a biodiversity assessment employing multiple methodologies (visual encounter transects, auditory surveys, leaf litter plots and pitfall traps) to inventory species prior to construction of a new road within the buffer zone of Moukalaba-Doudou National Park, Gabon. Because of difficulties in morphological identification and taxonomic uncertainty of amphibian species observed in the area, we integrated a DNA barcoding analysis into the project to improve the overall quality and accuracy of the species inventory. Based on morphology alone, 48 species were recognized in the field and voucher specimens of each were collected. We used tissue samples from specimens collected at our field site, material available from amphibians collected in other parts of Gabon and the Republic of Congo to initiate a DNA barcode library for west Central African amphibians. We then compared our sequences with material in GenBank for the genera recorded at the study site to assist in identifications. The resulting COI and 16S barcode library allowed us to update the number of species documented at the study site to 28, thereby providing a more accurate assessment of diversity and distributions. We caution that because sequence data maintained in GenBank are often poorly curated by the original submitters and cannot be amended by third-parties, these data have limited utility for identification purposes. Nevertheless, the use of DNA barcoding is likely to benefit biodiversity inventories and long-term monitoring, particularly for taxa that can be difficult to identify based on morphology alone; likewise, inventory and monitoring programs can contribute invaluable data to the DNA barcode library and the taxonomy of complex groups. Our methods provide an example of how non-taxonomists and parataxonomists working in understudied parts of the world with limited geographic sampling and comparative morphological material can use DNA barcoding and publicly available sequence data (GenBank) to rapidly identify the number of species and assign tentative names to aid in urgent conservation management actions and contribute to taxonomic resolution. PMID:29131846

Forensic botany II, DNA barcode for land plants: Which markers after the international agreement?

PubMed

Ferri, G; Corradini, B; Ferrari, F; Santunione, A L; Palazzoli, F; Alu', M

2015-03-01

The ambitious idea of using a short piece of DNA for large-scale species identification (DNA barcoding) is already a powerful tool for scientists and the application of this standard technique seems promising in a range of fields including forensic genetics. While DNA barcoding enjoyed a remarkable success for animal identification through cytochrome c oxidase I (COI) analysis, the attempts to identify a single barcode for plants remained a vain hope for a longtime. From the beginning, the Consortium for the Barcode of Life (CBOL) showed a lack of agreement on a core plant barcode, reflecting the diversity of viewpoints. Different research groups advocated various markers with divergent set of criteria until the recent publication by the CBOL-Plant Working Group. After a four-year effort, in 2009 the International Team concluded to agree on standard markers promoting a multilocus solution (rbcL and matK), with 70-75% of discrimination to the species level. In 2009 our group firstly proposed the broad application of DNA barcoding principles as a tool for identification of trace botanical evidence through the analysis of two chloroplast loci (trnH-psbA and trnL-trnF) in plant species belonging to local flora. Difficulties and drawbacks that were encountered included a poor coverage of species in specific databases and the lack of authenticated reference sequences for the selected markers. Successful preliminary results were obtained providing an approach to progressively identify unknown plant specimens to a given taxonomic rank, usable by any non-specialist botanist or in case of a shortage of taxonomic expertise. Now we considered mandatory to update and to compare our previous findings with the new selected plastid markers (matK+rbcL), taking into account forensic requirements. Features of all the four loci (the two previously analyzed trnH-psbA+trnL-trnF and matK+rbcL) were compared singly and in multilocus solutions to assess the most suitable combination for forensic botany. Based on obtained results, we recommend the adoption of a two-locus combination with rbcL+trnH-psbA plastid markers, which currently best satisfies forensic needs for botanical species identification. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Collecting in collections: a PCR strategy and primer set for DNA barcoding of decades-old dried museum specimens.

PubMed

Mitchell, Andrew

2015-09-01

Natural history museums are vastly underutilized as a source of material for DNA analysis because of perceptions about the limitations of DNA degradation in older specimens. Despite very few exceptions, most DNA barcoding projects, which aim to obtain sequence data from all species, generally use specimens collected specifically for that purpose, instead of the wealth of identified material in museums, constrained by the lack of suitable PCR methods. Any techniques that extend the utility of museum specimens for DNA analysis therefore are highly valuable. This study first tested the effects of specimen age and PCR amplicon size on PCR success rates in pinned insect specimens, then developed a PCR primer set and amplification strategy allowing greatly increased utilization of older museum specimens for DNA barcoding. PCR success rates compare favourably with the few published studies utilizing similar aged specimens, and this new strategy has the advantage of being easily automated for high-throughput laboratory workflows. The strategy uses hemi-nested, degenerate, M13-tailed PCR primers to amplify two overlapping amplicons, using two PCRs per amplicon (i.e. four PCRs per DNA sample). Initial PCR products are reamplified using an internal primer and a M13 primer. Together the two PCR amplicons yield 559 bp of the COI gene from Coleoptera, Lepidoptera, Diptera, Hemiptera, Odonata and presumably also other insects. BARCODE standard-compliant data were recovered from 67% (56 of 84) of specimens up to 25 years old, and 51% (102 of 197) of specimens up to 55 years old. Given the time, cost and specialist expertise required for fieldwork and identification, 'collecting in collections' is a viable alternative allowing researchers to capitalize on the knowledge captured by curation work in decades past. © 2015 John Wiley & Sons Ltd.

Molecular systematics of the freshwater stingrays (myliobatiformes: potamotrygonidae) of the Amazon, Orinoco, Magdalena, Esequibo, Caribbean, and Maracaibo basins (Colombia - Venezuela): evidence from three mitochondrial genes.

PubMed

Garcia, David Alejandro; Lasso, Carlos Andres; Morales, Monica; Caballero, Susana Josefina

2016-11-01

Lack of adequate information about the taxonomic and evolutionary relationships, ecology, biology, and distribution of several species belonging to the family Potamotrygonidae makes these species vulnerable to anthropic activities, including commercial overexploitation for the ornamental fish market. The aim of this study was to investigate the systematic relationships among genera and species belonging to this family by analyses of three mitochondrial gene regions. Samples were collected from the main river basins in Colombia and Venezuela for four genera and seven species of the family, as well as for what appear to be unidentified species. Three mitochondrial molecular markers COI, Cytb, and ATP6 were amplified and sequenced. Maximum likelihood and Bayesian inference analysis were performed to obtain topologies for each marker and for a concatenated dataset including the three genes. Small dataset may compromise some methods estimations of sequence divergence in the ATP6 marker. Monophyly of the four genera in Potamotrygonidae was confirmed and phylogenetic relationships among members of the Potamotrygon genus were not clearly resolved. However, results obtained with the molecular marker Cytb appear to offer a good starting point to differentiate among genera and species as a tool that could be used for barcoding. The application of this gene as a barcode could be applied for management and regulation of extraction practices for these genera. Sequencing complete mitochondrial genomes would be the next step for testing evolutionary hypothesis among these genera. Population structure analyses should be undertaken for Paratrygon, Potamotrygon magdalenae and motoro.

The complete mitochondrial genome of Percocypris pingi (Teleostei, Cypriniformes).

PubMed

Li, Yanping; Wang, Jinjin; Peng, Zuogang

2013-02-01

Percocypris pingi is an endemic and economic fish species only found in the upper Yangtze River basin in China. It has become endangered in recent years due to overfishing and/or dam construction. However, the available genetic data are still scarce for this species. Here, we sequenced the complete mitochondrial genome sequence of P. pingi using long polymerase chain reactions. The complete mitogenome sequence has 16,586 bp and contains the usual 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA (tRNA) genes, and 1 control region, the gene composition and order of which are similar to most of other vertebrates. Most mitochondrial genes except ND6 and eight tRNAs are encoded on the heavy strand. The overall base composition of the heavy strand is 30.9% A, 25.7% T, 26.6% C, and 16.8% G with a slight AT bias of 56.6%. There are seven regions of gene overlaps totaling 23 bp and 11 intergenic spacer regions totaling 35 bp. Combined with the COI barcoding region sequences of other 25 cyprinids, the phylogenetic position of P. pingi was estimated using neighbor-joining method. The results showed that P. pingi had a close phylogenetic relationship with the species from genus Schizothorax. This mitogenome sequence data of P. pingi would provide the fundamental genetic data for further conservation genetic studies for this endangered fish species.

Species boundaries of Gulf of Mexico vestimentiferans (Polychaeta, Siboglinidae) inferred from mitochondrial genes

NASA Astrophysics Data System (ADS)

Pia Miglietta, Maria; Hourdez, Stephane; Cowart, Dominique A.; Schaeffer, Stephen W.; Fisher, Charles

2010-11-01

At least six morphospecies of vestimentiferan tubeworms are associated with cold seeps in the Gulf of Mexico (GOM). The physiology and ecology of the two best-studied species from depths above 1000 m in the upper Louisiana slope (Lamellibrachia luymesi and Seepiophila jonesi) are relatively well understood. The biology of one rare species from the upper slope (escarpiid sp. nov.) and three morphospecies found at greater depths in the GOM (Lamellibrachia sp. 1, L. sp. 2, and Escarpia laminata) are not as well understood. Here we address species distributions and boundaries of cold-seep tubeworms using phylogenetic hypotheses based on two mitochondrial genes. Fragments of the mitochondrial large ribosomal subunit rDNA (16S) and cytochrome oxidase subunit I (COI) genes were sequenced for 167 vestimentiferans collected from the GOM and analyzed in the context of other seep vestimentiferans for which sequence data were available. The analysis supported five monophyletic clades of vestimentiferans in the GOM. Intra-clade variation in both genes was very low, and there was no apparent correlation between the within-clade diversity and collection depth or location. Two of the morphospecies of Lamellibrachia from different depths in the GOM could not be distinguished by either mitochondrial gene. Similarly, E. laminata could not be distinguished from other described species of Escarpia from either the west coast of Africa or the eastern Pacific using COI. We suggest that the mitochondrial COI and 16S genes have little utility as barcoding markers for seep vestimentiferan tubeworms.

Application of DNA barcodes in wildlife conservation in Tropical East Asia.

PubMed

Wilson, John-James; Sing, Kong-Wah; Lee, Ping-Shin; Wee, Alison K S

2016-10-01

Over the past 50 years, Tropical East Asia has lost more biodiversity than any tropical region. Tropical East Asia is a megadiverse region with an acute taxonomic impediment. DNA barcodes are short standardized DNA sequences used for taxonomic purposes and have the potential to lessen the challenges of biodiversity inventory and assessments in regions where they are most needed. We reviewed DNA barcoding efforts in Tropical East Asia relative to other tropical regions. We suggest DNA barcodes (or metabarcodes from next-generation sequencers) may be especially useful for characterizing and connecting species-level biodiversity units in inventories encompassing taxa lacking formal description (particularly arthropods) and in large-scale, minimal-impact approaches to vertebrate monitoring and population assessments through secondary sources of DNA (invertebrate derived DNA and environmental DNA). We suggest interest and capacity for DNA barcoding are slowly growing in Tropical East Asia, particularly among the younger generation of researchers who can connect with the barcoding analogy and understand the need for new approaches to the conservation challenges being faced. © 2016 Society for Conservation Biology.

DNA barcoding for conservation, seed banking and ecological restoration of Acacia in the Midwest of Western Australia.

PubMed

Nevill, Paul G; Wallace, Mark J; Miller, Joseph T; Krauss, Siegfried L

2013-11-01

We used DNA barcoding to address an important conservation issue in the Midwest of Western Australia, working on Australia's largest genus of flowering plant. We tested whether or not currently recommended plant DNA barcoding regions (matK and rbcL) were able to discriminate Acacia taxa of varying phylogenetic distances, and ultimately identify an ambiguously labelled seed collection from a mine-site restoration project. Although matK successfully identified the unknown seed as the rare and conservation priority listed A. karina, and was able to resolve six of the eleven study species, this region was difficult to amplify and sequence. In contrast, rbcL was straightforward to recover and align, but could not determine the origin of the seed and only resolved 3 of the 11 species. Other chloroplast regions (rpl32-trnL, psbA-trnH, trnL-F and trnK) had mixed success resolving the studied taxa. In general, species were better resolved in multilocus data sets compared to single-locus data sets. We recommend using the formal barcoding regions supplemented with data from other plastid regions, particularly rpl32-trnL, for barcoding in Acacia. Our study demonstrates the novel use of DNA barcoding for seed identification and illustrates the practical potential of DNA barcoding for the growing discipline of restoration ecology. © 2013 John Wiley & Sons Ltd.

Evaluation of single and multilocus DNA barcodes towards species delineation in complex tree genus Terminalia

PubMed Central

Mishra, Priyanka; Kumar, Amit; Nagireddy, Akshitha; Shukla, Ashutosh K.

2017-01-01

DNA barcoding is used as a universal tool for delimiting species boundaries in taxonomically challenging groups, with different plastid and nuclear regions (rbcL, matK, ITS and psbA-trnH) being recommended as primary DNA barcodes for plants. We evaluated the feasibility of using these regions in the species-rich genus Terminalia, which exhibits various overlapping morphotypes with pantropical distribution, owing to its complex taxonomy. Terminalia bellerica and T. chebula are ingredients of the famous Ayurvedic Rasayana formulation Triphala, used for detoxification and rejuvenation. High demand for extracted phytochemicals as well as the high trade value of several species renders mandatory the need for the correct identification of traded plant material. Three different analytical methods with single and multilocus barcoding regions were tested to develop a DNA barcode reference library from 222 individuals representing 41 Terminalia species. All the single barcodes tested had a lower discriminatory power than the multilocus regions, and the combination of matK+ITS had the highest resolution rate (94.44%). The average intra-specific variations (0.0188±0.0019) were less than the distance to the nearest neighbour (0.106±0.009) with matK and ITS. Distance-based Neighbour Joining analysis outperformed the character-based Maximum Parsimony method in the identification of traded species such as T. arjuna, T. chebula and T. tomentosa, which are prone to adulteration. rbcL was shown to be a highly conservative region with only 3.45% variability between all of the sequences. The recommended barcode combination, rbcL+matK, failed to perform in the genus Terminalia. Considering the complexity of resolution observed with single regions, the present study proposes the combination of matK+ITS as the most successful barcode in Terminalia. PMID:28829803

Efficiency of ITS Sequences for DNA Barcoding in Passiflora (Passifloraceae)

PubMed Central

Giudicelli, Giovanna Câmara; Mäder, Geraldo; de Freitas, Loreta Brandão

2015-01-01

DNA barcoding is a technique for discriminating and identifying species using short, variable, and standardized DNA regions. Here, we tested for the first time the performance of plastid and nuclear regions as DNA barcodes in Passiflora. This genus is a largely variable, with more than 900 species of high ecological, commercial, and ornamental importance. We analyzed 1034 accessions of 222 species representing the four subgenera of Passiflora and evaluated the effectiveness of five plastid regions and three nuclear datasets currently employed as DNA barcodes in plants using barcoding gap, applied similarity-, and tree-based methods. The plastid regions were able to identify less than 45% of species, whereas the nuclear datasets were efficient for more than 50% using “best match” and “best close match” methods of TaxonDNA software. All subgenera presented higher interspecific pairwise distances and did not fully overlap with the intraspecific distance, and similarity-based methods showed better results than tree-based methods. The nuclear ribosomal internal transcribed spacer 1 (ITS1) region presented a higher discrimination power than the other datasets and also showed other desirable characteristics as a DNA barcode for this genus. Therefore, we suggest that this region should be used as a starting point to identify Passiflora species. PMID:25837628

Molecular evidence of undescribed Ceratonova sp. (Cnidaria: Myxosporea) in the freshwater polychaete, Manayunkia speciosa, from western Lake Erie

USGS Publications Warehouse

Malakauskas, David M.; Snipes, Robert Benjamin; Thompson, Ann M.; Schloesser, Donald W.

2016-01-01

We used PCR to screen pooled individuals of Manayunkia speciosa from western Lake Erie, Michigan, USA for myxosporean parasites. Amplicons from positive PCRs were sequenced and showed a Ceratonova species in an estimated 1.1% (95% CI = 0.46%, 1.8%) of M. speciosa individuals. We sequenced 18S, ITS1, 5.8S, ITS2 and most of the 28S rDNA regions of this Ceratonova sp., and part of the protein-coding EF2 gene. Phylogenetic analyses of ribosomal and EF2 sequences showed the Lake Erie Ceratonova sp. is most similar to, but genetically distinct from, Ceratonova shasta. Marked interspecific polymorphism in all genes examined, including the ITS barcoding genes, along with geographic location suggests this is an undescribed Ceratonova species. COI sequences showed M. speciosa individuals in Michigan and California are the same species. These findings represent a third parasite in the genus Ceratonovapotentially hosted by M. speciosa.

Validation of the ITS2 region as a novel DNA barcode for identifying medicinal plant species.

PubMed

Chen, Shilin; Yao, Hui; Han, Jianping; Liu, Chang; Song, Jingyuan; Shi, Linchun; Zhu, Yingjie; Ma, Xinye; Gao, Ting; Pang, Xiaohui; Luo, Kun; Li, Ying; Li, Xiwen; Jia, Xiaocheng; Lin, Yulin; Leon, Christine

2010-01-07

The plant working group of the Consortium for the Barcode of Life recommended the two-locus combination of rbcL+matK as the plant barcode, yet the combination was shown to successfully discriminate among 907 samples from 550 species at the species level with a probability of 72%. The group admits that the two-locus barcode is far from perfect due to the low identification rate, and the search is not over. Here, we compared seven candidate DNA barcodes (psbA-trnH, matK, rbcL, rpoC1, ycf5, ITS2, and ITS) from medicinal plant species. Our ranking criteria included PCR amplification efficiency, differential intra- and inter-specific divergences, and the DNA barcoding gap. Our data suggest that the second internal transcribed spacer (ITS2) of nuclear ribosomal DNA represents the most suitable region for DNA barcoding applications. Furthermore, we tested the discrimination ability of ITS2 in more than 6600 plant samples belonging to 4800 species from 753 distinct genera and found that the rate of successful identification with the ITS2 was 92.7% at the species level. The ITS2 region can be potentially used as a standard DNA barcode to identify medicinal plants and their closely related species. We also propose that ITS2 can serve as a novel universal barcode for the identification of a broader range of plant taxa.

Testing DNA barcodes in closely related species of Curcuma (Zingiberaceae) from Myanmar and China.

PubMed

Chen, Juan; Zhao, Jietang; Erickson, David L; Xia, Nianhe; Kress, W John

2015-03-01

The genus Curcuma L. is commonly used as spices, medicines, dyes and ornamentals. Owing to its economic significance and lack of clear-cut morphological differences between species, this genus is an ideal case for developing DNA barcodes. In this study, four chloroplast DNA regions (matK, rbcL, trnH-psbA and trnL-F) and one nuclear region (ITS2) were generated for 44 Curcuma species and five species from closely related genera, represented by 96 samples. PCR amplification success rate, intra- and inter-specific genetic distance variation and the correct identification percentage were taken into account to assess candidate barcode regions. PCR and sequence success rate were high in matK (89.7%), rbcL (100%), trnH-psbA (100%), trnL-F (95.7%) and ITS2 (82.6%) regions. The results further showed that four candidate chloroplast barcoding regions (matK, rbcL, trnH-psbA and trnL-F) yield no barcode gaps, indicating that the genus Curcuma represents a challenging group for DNA barcoding. The ITS2 region presented large interspecific variation and provided the highest correct identification rates (46.7%) based on BLASTClust method among the five regions. However, the ITS2 only provided 7.9% based on NJ tree method. An increase in discriminatory power needs the development of more variable markers. © 2014 John Wiley & Sons Ltd.

Presence of the tunicate Asterocarpa humilis on ship hulls and aquaculture facilities in the coast of the Biobío Region, south central Chile.

PubMed

Pinochet, Javier; Leclerc, Jean-Charles; Brante, Antonio; Daguin-Thiébaut, Claire; Díaz, Christian; Tellier, Florence; Viard, Frédérique

2017-01-01

Non-native ascidians are important members of the fouling community associated with artificial substrata and man-made structures. Being efficient fouling species, they are easily spread by human-mediated transports (e.g., with aquaculture trade and maritime transports). This is exemplified by the ascidian Asterocarpa humilis which displays a wide distribution in the Southern Hemisphere and has been recently reported in the Northern Hemisphere (NW Europe). In continental Chile, its first report dates back from 2000 for the locality of Antofagasta (23°S). Although there was no evidence about the vectors of introduction and spread, nor the source, some authors suggested maritime transport by ship hulls and aquaculture devices as putative introduction pathways and vectors. In the present study, we report for the first time the presence of A. humilis on the hull of an international ship in a commercial port in Concepción bay (36°S), south central Chile. We also found one individual associated to a seashell farm, 70 km far from Concepción bay. Further individuals were subsequently identified within Concepción bay: one juvenile settled upon international harbor pilings and a dozen individuals along aquaculture seashell longlines. For the first specimens sampled, species identification was ascertained using both morphological criteria and molecular barcoding, using the mitochondrial gene cytochrome c oxidase subunit I (COI) and a nuclear gene (ribosomal RNA 18S). The nuclear 18S gene and the mitochondrial gene COI clearly assigned the specimens to A. humilis, confirming our morphological identification. Two haplotypes were obtained with COI corresponding to haplotypes previously obtained with European and Northern Chilean specimens. The present study thus reports for the first time the presence of A. humilis in the Araucanian ecoregion, documenting the apparent expansion of this non-native tunicate in Chile over 2,000 km, spanning over three ecoregions. In addition we reveal the potential implication of the international maritime transport as a vector of spread of this species along the Eastern Pacific coast, and the putative role of aquaculture facilities in promoting local establishments of non-native tunicates.

The resurrection of Charybdis (Gonioinfradens) giardi (Nobili, 1905), newly recorded from the SE Mediterranean Sea.

PubMed

Galil, Bella S; Douek, Jacob; Gevili, Roy; Goren, Menachem; Yudkovsky, Yana; Paz, Guy; Rinekvich, Baruch

2018-01-15

A single adult specimen of Gonioinfradens giardi, a portunid crab known from the Red Sea, Gulf of Oman and Arabian Gulf, was recently collected off the southern Israeli coast, in the southeastern Mediterranean Sea. Morphological characters, as well as molecular analyses based on the mitochondrial barcoding gene cytochrome oxidase sub unit I (COI), support its distinction from the widely distributed G. paucidentata. Therefore, G. giardi is reinstated as a valid species, and withdrawn from its synonymy with G. paucidentata. Previous Mediterranean records of the latter species are misidentifications and should be referred to G. giardi. The species is described, illustrated, and differentiated from its cogener.

Morphological redescription and DNA barcoding of Linevitshia prima Makarchenko, 1987 (Diptera: Chironomidae: Diamesinae) from Amur River basin (Russian Far East), with notes on systematics of the genus.

PubMed

Makarchenko, Eugenyi A; Semenchenko, Alexander A

2014-10-10

Additions and corrections to the diagnosis of the genus Linevitshia for male adult, pupa and larva are given, and systematic position of the genus is discussed. Illustrated redescription of adult male and first description of 4^th instar larva of L. prima Makarchenko from Amur River basin are provided. Comparison of data based on a new material with those of L. yezoensis Endo showed that the latter name is a junior synonym of L. prima. The species-specificity of L. prima COI sequences is analyzed and the sequences are presented as diagnostic characters-molecular markers of L. prima.

Ultra-deep sequencing enables high-fidelity recovery of biodiversity for bulk arthropod samples without PCR amplification

PubMed Central

2013-01-01

Background Next-generation-sequencing (NGS) technologies combined with a classic DNA barcoding approach have enabled fast and credible measurement for biodiversity of mixed environmental samples. However, the PCR amplification involved in nearly all existing NGS protocols inevitably introduces taxonomic biases. In the present study, we developed new Illumina pipelines without PCR amplifications to analyze terrestrial arthropod communities. Results Mitochondrial enrichment directly followed by Illumina shotgun sequencing, at an ultra-high sequence volume, enabled the recovery of Cytochrome c Oxidase subunit 1 (COI) barcode sequences, which allowed for the estimation of species composition at high fidelity for a terrestrial insect community. With 15.5 Gbp Illumina data, approximately 97% and 92% were detected out of the 37 input Operational Taxonomic Units (OTUs), whether the reference barcode library was used or not, respectively, while only 1 novel OTU was found for the latter. Additionally, relatively strong correlation between the sequencing volume and the total biomass was observed for species from the bulk sample, suggesting a potential solution to reveal relative abundance. Conclusions The ability of the new Illumina PCR-free pipeline for DNA metabarcoding to detect small arthropod specimens and its tendency to avoid most, if not all, false positives suggests its great potential in biodiversity-related surveillance, such as in biomonitoring programs. However, further improvement for mitochondrial enrichment is likely needed for the application of the new pipeline in analyzing arthropod communities at higher diversity. PMID:23587339

Validation of the ITS2 Region as a Novel DNA Barcode for Identifying Medicinal Plant Species

PubMed Central

Chen, Shilin; Yao, Hui; Han, Jianping; Liu, Chang; Song, Jingyuan; Shi, Linchun; Zhu, Yingjie; Ma, Xinye; Gao, Ting; Pang, Xiaohui; Luo, Kun; Li, Ying; Li, Xiwen; Jia, Xiaocheng; Lin, Yulin; Leon, Christine

2010-01-01

Background The plant working group of the Consortium for the Barcode of Life recommended the two-locus combination of rbcL + matK as the plant barcode, yet the combination was shown to successfully discriminate among 907 samples from 550 species at the species level with a probability of 72%. The group admits that the two-locus barcode is far from perfect due to the low identification rate, and the search is not over. Methodology/Principal Findings Here, we compared seven candidate DNA barcodes (psbA-trnH, matK, rbcL, rpoC1, ycf5, ITS2, and ITS) from medicinal plant species. Our ranking criteria included PCR amplification efficiency, differential intra- and inter-specific divergences, and the DNA barcoding gap. Our data suggest that the second internal transcribed spacer (ITS2) of nuclear ribosomal DNA represents the most suitable region for DNA barcoding applications. Furthermore, we tested the discrimination ability of ITS2 in more than 6600 plant samples belonging to 4800 species from 753 distinct genera and found that the rate of successful identification with the ITS2 was 92.7% at the species level. Conclusions The ITS2 region can be potentially used as a standard DNA barcode to identify medicinal plants and their closely related species. We also propose that ITS2 can serve as a novel universal barcode for the identification of a broader range of plant taxa. PMID:20062805

Opening the treasure chest: A DNA-barcoding primer set for most higher taxa of Central European birds and mammals from museum collections

PubMed Central

Schäffer, Sylvia; Zachos, Frank E.

2017-01-01

DNA-barcoding is a rapidly developing method for efficiently identifying samples to species level by means of short standard DNA sequences. However, reliable species assignment requires the availability of a comprehensive DNA barcode reference library, and hence numerous initiatives aim at generating such barcode databases for particular taxa or geographic regions. Historical museum collections represent a potentially invaluable source for the DNA-barcoding of many taxa. This is particularly true for birds and mammals, for which collecting fresh (voucher) material is often very difficult to (nearly) impossible due to the special animal welfare and conservation regulations that apply to vertebrates in general, and birds and mammals in particular. Moreover, even great efforts might not guarantee sufficiently complete sampling of fresh material in a short period of time. DNA extracted from historical samples is usually degraded, such that only short fragments can be amplified, rendering the recovery of the barcoding region as a single fragment impossible. Here, we present a new set of primers that allows the efficient amplification and sequencing of the entire barcoding region in most higher taxa of Central European birds and mammals in six overlapping fragments, thus greatly increasing the value of historical museum collections for generating DNA barcode reference libraries. Applying our new primer set in recently established NGS protocols promises to further increase the efficiency of barcoding old bird and mammal specimens. PMID:28358863

Opening the treasure chest: A DNA-barcoding primer set for most higher taxa of Central European birds and mammals from museum collections.

PubMed

Schäffer, Sylvia; Zachos, Frank E; Koblmüller, Stephan

2017-01-01

DNA-barcoding is a rapidly developing method for efficiently identifying samples to species level by means of short standard DNA sequences. However, reliable species assignment requires the availability of a comprehensive DNA barcode reference library, and hence numerous initiatives aim at generating such barcode databases for particular taxa or geographic regions. Historical museum collections represent a potentially invaluable source for the DNA-barcoding of many taxa. This is particularly true for birds and mammals, for which collecting fresh (voucher) material is often very difficult to (nearly) impossible due to the special animal welfare and conservation regulations that apply to vertebrates in general, and birds and mammals in particular. Moreover, even great efforts might not guarantee sufficiently complete sampling of fresh material in a short period of time. DNA extracted from historical samples is usually degraded, such that only short fragments can be amplified, rendering the recovery of the barcoding region as a single fragment impossible. Here, we present a new set of primers that allows the efficient amplification and sequencing of the entire barcoding region in most higher taxa of Central European birds and mammals in six overlapping fragments, thus greatly increasing the value of historical museum collections for generating DNA barcode reference libraries. Applying our new primer set in recently established NGS protocols promises to further increase the efficiency of barcoding old bird and mammal specimens.

Locating and decoding barcodes in fuzzy images captured by smart phones

NASA Astrophysics Data System (ADS)

Deng, Wupeng; Hu, Jiwei; Liu, Quan; Lou, Ping

2017-07-01

With the development of barcodes for commercial use, people's requirements for detecting barcodes by smart phone become increasingly pressing. The low quality of barcode image captured by mobile phone always affects the decoding and recognition rates. This paper focuses on locating and decoding EAN-13 barcodes in fuzzy images. We present a more accurate locating algorithm based on segment length and high fault-tolerant rate algorithm for decoding barcodes. Unlike existing approaches, location algorithm is based on the edge segment length of EAN -13 barcodes, while our decoding algorithm allows the appearance of fuzzy region in barcode image. Experimental results are performed on damaged, contaminated and scratched digital images, and provide a quite promising result for EAN -13 barcode location and decoding.

Genetic diversity among populations of Antarctic springtails (Collembola) within the Mackay Glacier ecotone.

PubMed

Beet, Clare R; Hogg, Ian D; Collins, Gemma E; Cowan, Don A; Wall, Diana H; Adams, Byron J

2016-09-01

Climate changes are likely to have major influences on the distribution and abundance of Antarctic terrestrial biota. To assess arthropod distribution and diversity within the Ross Sea region, we examined mitochondrial DNA (COI) sequences for three currently recognized species of springtail (Collembola) collected from sites in the vicinity, and to the north of, the Mackay Glacier (77°S). This area acts as a transition between two biogeographic regions (northern and southern Victoria Land). We found populations of highly divergent individuals (5%-11.3% intraspecific sequence divergence) for each of the three putative springtail species, suggesting the possibility of cryptic diversity. Based on molecular clock estimates, these divergent lineages are likely to have been isolated for 3-5 million years. It was during this time that the Western Antarctic Ice Sheet (WAIS) was likely to have completely collapsed, potentially facilitating springtail dispersal via rafting on running waters and open seaways. The reformation of the WAIS would have isolated newly established populations, with subsequent dispersal restricted by glaciers and ice-covered areas. Given the currently limited distributions for these genetically divergent populations, any future changes in species' distributions can be easily tracked through the DNA barcoding of springtails from within the Mackay Glacier ecotone.

INTERNAL TRANSCRIBED SPACER (ITS), AN IDEAL DNA BARCODE FOR SPECIES DISCRIMINATION IN CRAWFURDIA WALL. (GENTIANACEAE).

PubMed

Zhang, Dequan; Jiang, Bei; Duan, Lizhen; Zhou, Nong

2016-01-01

DNA barcoding is a technique used to identify species based on species-specific differences in short regions of their DNA. It is widely used in species discrimination of medicinal plants and traditional medicines. In the present study, four potential DNA barcodes, namely rbcL , matK , trnH-psbA and ITS (nuclear ribosomal internal transcribed spacer) were adopted for species discrimination in Crawfurdia Wall (Genetiaceae). Identification ability of these DNA barcodes and combinations were evaluated using three classic methods (Distance, Blast and Tree-Building). As a result, ITS, trnH-psbA and rbcL regions showed great universality for a success rate of 100%; whereas matK was disappointing for which only 65% samples gained useful DNA sequences. ITS region, which could clearly and effectively identify the five species in Crawfurdia , performed very well in this study. On the contrary, trnH-psbA and rbcL performed poorly in discrimination among these species. ITS marker was an ideal DNA barcode in Crawfurdia and it should be incorporated into one of the core barcodes for seed plants.

Evidence of natural Wolbachia infections and molecular identification of field populations of Culex pipiens complex (Diptera: Culicidae) mosquitoes in western Turkey.

PubMed

Morçiçek, Burçin; Taskin, Belgin Gocmen; Doğaç, Ersin; Doğaroğlu, Taylan; Taskin, Vatan

2018-06-01

Establishing reliable risk projection information about the distribution pattern of members of the Culex pipiens complex is of particular interest, as these mosquitoes are competent vectors for certain disease-causing pathogens. Wolbachia, a maternally inherited bacterial symbiont, are distributed in various arthropod species and can induce cytoplasmic incompatibility, i.e., reduced egg hatch, in certain crosses. It is being considered as a tool for population control of mosquito disease vectors. The Aegean region is characterized by highly populated, rural, and agricultural areas and is also on the route of the migratory birds. In this study, a fragment of the 658 bp of the mitochondrial cytochrome c oxidase subunit 1 (COI) gene, which includes the barcode region, was employed to differentiate Cx. pipiens complex species found in this region. Moreover, for the first time, the prevalence of Wolbachia endobacteria in these natural populations was examined using PCR amplification of a specific wsp gene. Our results revealed a widespread (more than 90%, n=121) presence of the highly efficient West Nile virus vector Cx. quinquefasciatus in the region. We also found that Wolbachia infection is widespread; the average prevalence was 62% in populations throughout the region. This study provided valuable information about the composition of Cx. pipiens complex mosquitoes and the prevalence of Wolbachia infection in these populations in the Aegean region. This information will be helpful in tracking mosquito-borne diseases and designing and implementing Wolbachia-based control strategies in the region. © 2018 The Society for Vector Ecology.

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

PubMed

Frantine-Silva, W; Sofia, S H; Orsi, M L; Almeida, F S

2015-09-01

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. © 2015 John Wiley & Sons Ltd.

Sushi barcoding in the UK: another kettle of fish

PubMed Central

Taylor, Sasha-Ann; Di Muri, Cristina; Hankard, Elizabeth A.; Towne, Jessica A.; Watson, Mhairi

2016-01-01

Although the spread of sushi restaurants in the European Union and United States is a relatively new phenomenon, they have rapidly become among the most popular food services globally. Recent studies indicate that they can be associated with very high levels (>70%) of fish species substitution. Based on indications that the European seafood retail sector may currently be under better control than its North American counterpart, here we investigated levels of seafood labelling accuracy in sushi bars and restaurants across England. We used the COI barcoding gene to screen samples of tuna, eel, and a variety of other products characterised by less visually distinctive ‘white flesh’. Moderate levels of substitution were found (10%), significantly lower than observed in North America, which lends support to the argument that public awareness, policy and governance of seafood labels is more effective in the European Union. Nevertheless, the results highlight that current labelling practice in UK restaurants lags behind the level of detail implemented in the retail sector, which hinders consumer choice, with potentially damaging economic, health and environmental consequences. Specifically, critically endangered species of tuna and eel continue to be sold without adequate information to consumers. PMID:27069819

Genetic distance of Malaysian mousedeer based on mitochondrial DNA cytochrome oxidase I (COI) and D-loop region sequences

NASA Astrophysics Data System (ADS)

Bakar, Mohamad-Azam Akmal Abu; Rovie-Ryan, Jeffrine Japning; Ampeng, Ahmad; Yaakop, Salmah; Nor, Shukor Md; Md-Zain, Badrul Munir

2018-04-01

Mousedeer is one of the primitive mammals that can be found mainly in Southeast-Asia region. There are two species of mousedeer in Malaysia which are Tragulus kanchil and Tragulus napu. Both species can be distinguish by size, coat coloration, and throat pattern but clear diagnosis still cannot be found. The objective of the study is to show the genetic distance relationship between T. kanchil and T. napu and their population based on mitochondrial DNA (mtDNA) cytochrome oxidase I (COI) and D-loop region. There are 42 sample of mousedeer were used in this study collected by PERHILITAN from different locality. Another 29 D-loop sequence were retrieved from Genbank for comparative analysis. All sample were amplified using universal primer and species-specific primer for COI and D-loop genes via PCR process. The amplified sequences were analyzed to determine genetic distance of T. kanchil and T. napu. From the analysis, the average genetic distance between T. kanchil and T. napu based on locus COI and D-loop were 0.145 and 0.128 respectively. The genetic distance between populations of T. kanchil based on locus COI was between 0.003-0.013. For locus D-loop, genetic distance analysis showed distance in relationship between west-coast populations to east-coast population of T. kanchil. COI and D-loop mtDNA region provided a clear picture on the relationship within the mousedeer species. Last but not least, conservation effort toward protecting this species can be done by study the molecular genetics and prevent the extinction of this species.

A new species of Oxynetra from Mexico (Hesperiidae, Pyrginae, Pyrrhopygini).

PubMed

Warren, Andrew D; Grishin, Nick V

2017-01-01

Oxynetra aureopecta sp. n. is described from the Sierra Madre Oriental of east-central Mexico. Visually similar to Mesoamerican O. hopfferi Staudinger, 1888 in having five orange bands on the abdomen above, it is diagnosed by orange forecoxae and palpi beneath, narrower forewing hyaline bands and a prominent 6% difference in the COI DNA barcode sequence. It is the northernmost representative of the hopfferi species group that also includes O. stangelandi Grishin & Burns, 2013, characterized by a single-banded abdomen and currently known only from the Area de Conservación Guanacaste in northwestern Costa Rica. Both O. hopfferi and O. stangelandi possess white forecoxae and ventral palpi. This new discovery brings the total number of Oxynetra C. & R. Felder, 1862 species to five.

Taxonomic history and invasion biology of two Phyllonorycter leaf miners (Lepidoptera: Gracillariidae) with links to taxonomic and molecular datasets.

PubMed

De Prins, Jurate; De Prins, Willy; De Coninck, Eliane; Kawahara, Akito Y; Milton, Megan A; Hebert, Paul D N

2013-01-01

This paper deals with two European species, Phyllonorycter mespilella (Hübner, 1805) and P. trifasciella (Haworth, 1828), that have colonized the subtropical Canary Islands. The Rosaceae leaf miner, P. mespilella, is recorded for the first time from Lanzarote and La Palma, while the Caprifoliaceae leaf miner, P. trifasciella, is recorded from Tenerife. We present the diagnoses of these species based on morphology, a preliminary DNA barcode (COI) library of congeneric and con-familial species, and discuss the taxonomic position of the colonizers within the blancardella and trifasciella species groups. The recent intensification of anthropogenic disturbance likely accounts for their range expansion, an event that may impact the relict flora present on the Canary Islands.

How Many Loci Does it Take to DNA Barcode a Crocus?

PubMed Central

Seberg, Ole; Petersen, Gitte

2009-01-01

Background DNA barcoding promises to revolutionize the way taxonomists work, facilitating species identification by using small, standardized portions of the genome as substitutes for morphology. The concept has gained considerable momentum in many animal groups, but the higher plant world has been largely recalcitrant to the effort. In plants, efforts are concentrated on various regions of the plastid genome, but no agreement exists as to what kinds of regions are ideal, though most researchers agree that more than one region is necessary. One reason for this discrepancy is differences in the tests that are used to evaluate the performance of the proposed regions. Most tests have been made in a floristic setting, where the genetic distance and therefore the level of variation of the regions between taxa is large, or in a limited set of congeneric species. Methodology and Principal Findings Here we present the first in-depth coverage of a large taxonomic group, all 86 known species (except two doubtful ones) of crocus. Even six average-sized barcode regions do not identify all crocus species. This is currently an unrealistic burden in a barcode context. Whereas most proposed regions work well in a floristic context, the majority will – as is the case in crocus – undoubtedly be less efficient in a taxonomic setting. However, a reasonable but less than perfect level of identification may be reached – even in a taxonomic context. Conclusions/Significance The time is ripe for selecting barcode regions in plants, and for prudent examination of their utility. Thus, there is no reason for the plant community to hold back the barcoding effort by continued search for the Holy Grail. We must acknowledge that an emerging system will be far from perfect, fraught with problems and work best in a floristic setting. PMID:19240801

How many loci does it take to DNA barcode a crocus?

PubMed

Seberg, Ole; Petersen, Gitte

2009-01-01

DNA barcoding promises to revolutionize the way taxonomists work, facilitating species identification by using small, standardized portions of the genome as substitutes for morphology. The concept has gained considerable momentum in many animal groups, but the higher plant world has been largely recalcitrant to the effort. In plants, efforts are concentrated on various regions of the plastid genome, but no agreement exists as to what kinds of regions are ideal, though most researchers agree that more than one region is necessary. One reason for this discrepancy is differences in the tests that are used to evaluate the performance of the proposed regions. Most tests have been made in a floristic setting, where the genetic distance and therefore the level of variation of the regions between taxa is large, or in a limited set of congeneric species. Here we present the first in-depth coverage of a large taxonomic group, all 86 known species (except two doubtful ones) of crocus. Even six average-sized barcode regions do not identify all crocus species. This is currently an unrealistic burden in a barcode context. Whereas most proposed regions work well in a floristic context, the majority will--as is the case in crocus--undoubtedly be less efficient in a taxonomic setting. However, a reasonable but less than perfect level of identification may be reached--even in a taxonomic context. The time is ripe for selecting barcode regions in plants, and for prudent examination of their utility. Thus, there is no reason for the plant community to hold back the barcoding effort by continued search for the Holy Grail. We must acknowledge that an emerging system will be far from perfect, fraught with problems and work best in a floristic setting.

Giant worms chez moi! Hammerhead flatworms (Platyhelminthes, Geoplanidae, Bipalium spp., Diversibipalium spp.) in metropolitan France and overseas French territories

PubMed Central

Winsor, Leigh; Gey, Delphine; Gros, Pierre; Thévenot, Jessica

2018-01-01

Background Species of the genera Bipalium and Diversibipalium, or bipaliines, are giants among land planarians (family Geoplanidae), reaching length of 1 m; they are also easily distinguished from other land flatworms by the characteristic hammer shape of their head. Bipaliines, which have their origin in warm parts of Asia, are invasive species, now widespread worldwide. However, the scientific literature is very scarce about the widespread repartition of these species, and their invasion in European countries has not been studied. Methods In this paper, on the basis of a four year survey based on citizen science, which yielded observations from 1999 to 2017 and a total of 111 records, we provide information about the five species present in Metropolitan France and French overseas territories. We also investigated the molecular variability of cytochrome-oxidase 1 (COI) sequences of specimens. Results Three species are reported from Metropolitan France: Bipalium kewense, Diversibipalium multilineatum, and an unnamed Diversibipalium ‘black’ species. We also report the presence of B. kewense from overseas territories, such as French Polynesia (Oceania), French Guiana (South America), the Caribbean French islands of Martinique, Guadeloupe, Saint Martin and Saint Barthélemy, and Montserrat (Central America), and La Réunion island (off South-East Africa). For B. vagum, observations include French Guiana, Guadeloupe, Martinique, Saint Barthélemy, Saint Martin, Montserrat, La Réunion, and Florida (USA). A probable new species, Diversibipalium sp. ‘blue,’ is reported from Mayotte Island (off South–East Africa). B. kewense, B. vagum and D. multilineatum each showed 0% variability in their COI sequences, whatever their origin, suggesting that the specimens are clonal, and that sexual reproduction is probably absent. COI barcoding was efficient in identifying species, with differences over 10% between species; this suggests that barcoding can be used in the future for identifying these invasive species. In Metropolitan south–west France, a small area located in the Department of Pyrénées-Atlantiques was found to be a hot-spot of bipaliine biodiversity and abundance for more than 20 years, probably because of the local mild weather. Discussion The present findings strongly suggest that the species present in Metropolitan France and overseas territories should be considered invasive alien species. Our numerous records in the open in Metropolitan France raise questions: as scientists, we were amazed that these long and brightly coloured worms could escape the attention of scientists and authorities in a European developed country for such a long time; improved awareness about land planarians is certainly necessary. PMID:29844951

rbcL and matK earn two thumbs up as the core DNA barcode for ferns.

PubMed

Li, Fay-Wei; Kuo, Li-Yaung; Rothfels, Carl J; Ebihara, Atsushi; Chiou, Wen-Liang; Windham, Michael D; Pryer, Kathleen M

2011-01-01

DNA barcoding will revolutionize our understanding of fern ecology, most especially because the accurate identification of the independent but cryptic gametophyte phase of the fern's life history--an endeavor previously impossible--will finally be feasible. In this study, we assess the discriminatory power of the core plant DNA barcode (rbcL and matK), as well as alternatively proposed fern barcodes (trnH-psbA and trnL-F), across all major fern lineages. We also present plastid barcode data for two genera in the hyperdiverse polypod clade--Deparia (Woodsiaceae) and the Cheilanthes marginata group (currently being segregated as a new genus of Pteridaceae)--to further evaluate the resolving power of these loci. Our results clearly demonstrate the value of matK data, previously unavailable in ferns because of difficulties in amplification due to a major rearrangement of the plastid genome. With its high sequence variation, matK complements rbcL to provide a two-locus barcode with strong resolving power. With sequence variation comparable to matK, trnL-F appears to be a suitable alternative barcode region in ferns, and perhaps should be added to the core barcode region if universal primer development for matK fails. In contrast, trnH-psbA shows dramatically reduced sequence variation for the majority of ferns. This is likely due to the translocation of this segment of the plastid genome into the inverted repeat regions, which are known to have a highly constrained substitution rate. Our study provides the first endorsement of the two-locus barcode (rbcL+matK) in ferns, and favors trnL-F over trnH-psbA as a potential back-up locus. Future work should focus on gathering more fern matK sequence data to facilitate universal primer development.

Assessment of mangroves from Goa, west coast India using DNA barcode.

PubMed

Saddhe, Ankush Ashok; Jamdade, Rahul Arvind; Kumar, Kundan

2016-01-01

Mangroves are salt-tolerant forest ecosystems of tropical and subtropical intertidal regions. They are among most productive, diverse, biologically important ecosystem and inclined toward threatened system. Identification of mangrove species is of critical importance in conserving and utilizing biodiversity, which apparently hindered by a lack of taxonomic expertise. In recent years, DNA barcoding using plastid markers rbcL and matK has been suggested as an effective method to enrich traditional taxonomic expertise for rapid species identification and biodiversity inventories. In the present study, we performed assessment of available 14 mangrove species of Goa, west coast India based on core DNA barcode markers, rbcL and matK. PCR amplification success rate, intra- and inter-specific genetic distance variation and the correct identification percentage were taken into account to assess candidate barcode regions. PCR and sequence success rate were high in rbcL (97.7 %) and matK (95.5 %) region. The two candidate chloroplast barcoding regions (rbcL, matK) yielded barcode gaps. Our results clearly demonstrated that matK locus assigned highest correct identification rates (72.09 %) based on TaxonDNA Best Match criteria. The concatenated rbcL + matK loci were able to adequately discriminate all mangrove genera and species to some extent except those in Rhizophora, Sonneratia and Avicennia. Our study provides the first endorsement of the species resolution among mangroves using plastid genes with few exceptions. Our future work will be focused on evaluation of other barcode markers to delineate complete resolution of mangrove species and identification of putative hybrids.

Complete DNA barcode reference library for a country's butterfly fauna reveals high performance for temperate Europe

PubMed Central

Dincă, Vlad; Zakharov, Evgeny V.; Hebert, Paul D. N.; Vila, Roger

2011-01-01

DNA barcoding aims to accelerate species identification and discovery, but performance tests have shown marked differences in identification success. As a consequence, there remains a great need for comprehensive studies which objectively test the method in groups with a solid taxonomic framework. This study focuses on the 180 species of butterflies in Romania, accounting for about one third of the European butterfly fauna. This country includes five eco-regions, the highest of any in the European Union, and is a good representative for temperate areas. Morphology and DNA barcodes of more than 1300 specimens were carefully studied and compared. Our results indicate that 90 per cent of the species form barcode clusters allowing their reliable identification. The remaining cases involve nine closely related species pairs, some whose taxonomic status is controversial or that hybridize regularly. Interestingly, DNA barcoding was found to be the most effective identification tool, outperforming external morphology, and being slightly better than male genitalia. Romania is now the first country to have a comprehensive DNA barcode reference database for butterflies. Similar barcoding efforts based on comprehensive sampling of specific geographical regions can act as functional modules that will foster the early application of DNA barcoding while a global system is under development. PMID:20702462

Determination of Genetic Diversity in Chilo partellus, Busseola fusca, and Spodoptera frugiperda Infesting Sugarcane in Southern Malawi Using DNA Barcodes.

PubMed

Kasambala Donga, Trust; Meadow, Richard

2018-06-22

Sugarcane is one of the most valuable crops in the world. Native and exotic Lepidopteran stemborers significantly limit sugarcane production. However, the identity and genetic diversity of stemborers infesting sugarcane in Malawi is unknown. The main objectives for this study were to identify and determine genetic diversity in stemborers infesting sugarcane in Malawi. We conducted field surveys between June 2016 and March 2017 in the Lower Shire Valley district of Chikwawa and Nsanje, southern Malawi. Molecular identification was based amplification the partial cytochrome oxidase subunit I (COI) gene region. Phylogenetic trees for sequences were generated and published GenBank accessions for each species were constructed. We found that Malawi Busseola fusca (Lepidoptera: Noctuidae) specimens belonged to clade II, Spodoptera frugiperda sp. 1 (Lepidoptera: Noctuidae) and Chilo partellus (Lepidoptera: Crambidae) were infesting sugarcane. Interspecific divergence ranged from 8.7% to 15.3%. Intraspecific divergence was highest for B. fusca , 3.6%. There were eight haplotypes for B. fusca , three for S. frugiperda and three for C. partellus . The importance of accurate species identification and genetic diversity on stemborer management is presented.

Effectiveness of ITS and sub-regions as DNA barcode markers for the identification of Basidiomycota (Fungi).

PubMed

Badotti, Fernanda; de Oliveira, Francislon Silva; Garcia, Cleverson Fernando; Vaz, Aline Bruna Martins; Fonseca, Paula Luize Camargos; Nahum, Laila Alves; Oliveira, Guilherme; Góes-Neto, Aristóteles

2017-02-23

Fungi are among the most abundant and diverse organisms on Earth. However, a substantial amount of the species diversity, relationships, habitats, and life strategies of these microorganisms remain to be discovered and characterized. One important factor hindering progress is the difficulty in correctly identifying fungi. Morphological and molecular characteristics have been applied in such tasks. Later, DNA barcoding has emerged as a new method for the rapid and reliable identification of species. The nrITS region is considered the universal barcode of Fungi, and the ITS1 and ITS2 sub-regions have been applied as metabarcoding markers. In this study, we performed a large-scale analysis of all the available Basidiomycota sequences from GenBank. We carried out a rigorous trimming of the initial dataset based in methodological principals of DNA Barcoding. Two different approaches (PCI and barcode gap) were used to determine the performance of the complete ITS region and sub-regions. For most of the Basidiomycota genera, the three genomic markers performed similarly, i.e., when one was considered a good marker for the identification of a genus, the others were also; the same results were observed when the performance was insufficient. However, based on barcode gap analyses, we identified genomic markers that had a superior identification performance than the others and genomic markers that were not indicated for the identification of some genera. Notably, neither the complete ITS nor the sub-regions were useful in identifying 11 of the 113 Basidiomycota genera. The complex phylogenetic relationships and the presence of cryptic species in some genera are possible explanations of this limitation and are discussed. Knowledge regarding the efficiency and limitations of the barcode markers that are currently used for the identification of organisms is crucial because it benefits research in many areas. Our study provides information that may guide researchers in choosing the most suitable genomic markers for identifying Basidiomycota species.

Application of DNA Barcodes in Asian Tropical Trees--A Case Study from Xishuangbanna Nature Reserve, Southwest China.

PubMed

Huang, Xiao-cui; Ci, Xiu-qin; Conran, John G; Li, Jie

2015-01-01

Within a regional floristic context, DNA barcoding is more useful to manage plant diversity inventories on a large scale and develop valuable conservation strategies. However, there are no DNA barcode studies from tropical areas of China, which represents one of the biodiversity hotspots around the world. A DNA barcoding database of an Asian tropical trees with high diversity was established at Xishuangbanna Nature Reserve, Yunnan, southwest China using rbcL and matK as standard barcodes, as well as trnH-psbA and ITS as supplementary barcodes. The performance of tree species identification success was assessed using 2,052 accessions from four plots belonging to two vegetation types in the region by three methods: Neighbor-Joining, Maximum-Likelihood and BLAST. We corrected morphological field identification errors (9.6%) for the three plots using rbcL and matK based on Neighbor-Joining tree. The best barcode region for PCR and sequencing was rbcL (97.6%, 90.8%), followed by trnH-psbA (93.6%, 85.6%), while matK and ITS obtained relative low PCR and sequencing success rates. However, ITS performed best for both species (44.6-58.1%) and genus (72.8-76.2%) identification. With trnH-psbA slightly less effective for species identification. The two standard barcode rbcL and matK gave poor results for species identification (24.7-28.5% and 31.6-35.3%). Compared with other studies from comparable tropical forests (e.g. Cameroon, the Amazon and India), the overall performance of the four barcodes for species identification was lower for the Xishuangbanna Nature Reserve, possibly because of species/genus ratios and species composition between these tropical areas. Although the core barcodes rbcL and matK were not suitable for species identification of tropical trees from Xishuangbanna Nature Reserve, they could still help with identification at the family and genus level. Considering the relative sequence recovery and the species identification performance, we recommend the use of trnH-psbA and ITS in combination as the preferred barcodes for tropical tree species identification in China.

Integrative revision of the giant pill-millipede genus Sphaeromimus from Madagascar, with the description of seven new species (Diplopoda, Sphaerotheriida, Arthrosphaeridae)

PubMed Central

Wesener, Thomas; Le, Daniel Minh-Tu; Loria, Stephanie F.

2014-01-01

Abstract The Malagasy giant pill-millipede genus Sphaeromimus de Saussure & Zehntner, 1902 is revised. Seven new species, S. titanus sp. n., S. vatovavy sp. n., S. lavasoa sp. n., S. andohahela sp. n., S. ivohibe sp. n., S. saintelucei sp. n., and S. andrahomana sp. n. were discovered, in one case with the help of sequence data, in the rainforests of southeastern Madagascar. The species are described using light- and scanning electron microscopy. A key to all 10 species of the genus is presented. All but one (S. andohahela) of the newly discovered species are microendemics each occurring in isolated forest fragments. The mitochondrial COI barcoding gene was amplified and sequenced for 18 Sphaeromimus specimens, and a dataset containing COI sequences of 28 specimens representing all Sphaeromimus species (except S. vatovavy) was analyzed. All species are genetically monophyletic. Interspecific uncorrected genetic distances were moderate (4–10%) to high (18–25%), whereas intraspecific variation is low (0–3.5%). Sequence data allowed the correct identification of three colour morphs of S. musicus, as well as the identity of a cave specimen, which although aberrant in its morphology and colouration, was genetically identical to the holotype of S. andrahoma. PMID:25009417

Exploring the utility of DNA barcoding in species delimitation of Polypedilum (Tripodura) non-biting midges (Diptera: Chironomidae).

PubMed

Song, Chao; Wang, Qian; Zhang, Ruilei; Sun, Bingjiao; Wang, Xinhua

2016-02-16

In this study, we tested the utility of the mitochondrial gene cytochrome c oxidase subunit 1 (CO1) as the barcode region to deal with taxonomical problems of Polypedilum (Tripodura) non-biting midges (Diptera: Chironomidae). The 114 DNA barcodes representing 27 morphospecies are divided into 33 well separated clusters based on both Neighbor Joining and Maximum Likelihood methods. DNA barcodes revealed an 82% success rate in matching with morphospecies. The selected DNA barcode data support 37-64 operational taxonomic units (OTUs) based on the methods of Automatic Barcode Gap Discovery (ABGD) and Poisson Tree Process (PTP). Furthermore, a priori species based on consistent phenotypic variations were attested by molecular analysis, and a taxonomical misidentification of barcode sequences from GenBank was found. We could not observe a distinct barcode gap but an overlap ranged from 9-12%. Our results supported DNA barcoding as an ideal method to detect cryptic species, delimit sibling species, and associate different life stages in non-biting midges.

Critical factors for assembling a high volume of DNA barcodes

PubMed Central

Hajibabaei, Mehrdad; deWaard, Jeremy R; Ivanova, Natalia V; Ratnasingham, Sujeevan; Dooh, Robert T; Kirk, Stephanie L; Mackie, Paula M; Hebert, Paul D.N

2005-01-01

Large-scale DNA barcoding projects are now moving toward activation while the creation of a comprehensive barcode library for eukaryotes will ultimately require the acquisition of some 100 million barcodes. To satisfy this need, analytical facilities must adopt protocols that can support the rapid, cost-effective assembly of barcodes. In this paper we discuss the prospects for establishing high volume DNA barcoding facilities by evaluating key steps in the analytical chain from specimens to barcodes. Alliances with members of the taxonomic community represent the most effective strategy for provisioning the analytical chain with specimens. The optimal protocols for DNA extraction and subsequent PCR amplification of the barcode region depend strongly on their condition, but production targets of 100K barcode records per year are now feasible for facilities working with compliant specimens. The analysis of museum collections is currently challenging, but PCR cocktails that combine polymerases with repair enzyme(s) promise future success. Barcode analysis is already a cost-effective option for species identification in some situations and this will increasingly be the case as reference libraries are assembled and analytical protocols are simplified. PMID:16214753

Fixation, description and DNA barcode of a neotype for Botryllus schlosseri (Pallas, 1766) (Tunicata, Ascidiacea).

PubMed

Brunetti, Riccardo; Manni, Lucia; Mastrototaro, Francesco; Gissi, Carmela; Gasparini, Fabio

2017-11-22

Botryllus schlosseri is a widespread colonial ascidian commonly considered cosmopolitan and amply used as model for researches ranging from developmental biology to immunobiology. Recently, molecular data lead to hypothesize that the species named B. schlosseri may consist of more than a single taxon. Indeed, five highly divergent clades, named A-E, have been genetically identified and are referred as cryptic species. In this context, and lacking both a type and a detailed morphological description, we believe that it is necessary, as a taxonomic reference point, to designate a neotype and re-describe the species. Therefore, a sample from the Lagoon of Venice (Adriatic Sea, Italy) was deposited as neotype in the Natural History Museum of Venice (Italy), preserved both in formalin and in 90% ethanol. Here we provide a morphological description of the suggested neotype of B. schlosseri that takes into account several developmental stages (oozooid, zooid of first blastogenic generations, and mature zooid) and is carefully compared with the previous descriptions of samples coming from other European and non-European localities. Finally, we associate our morphological description to a "DNA barcode", consisting in a long fragment of the mitochondrial COI gene. Our description is associated to clade A, although at now we cannot guarantee that this association is univocal.

A new species of Oxynetra from Mexico (Hesperiidae, Pyrginae, Pyrrhopygini)

PubMed Central

Warren, Andrew D.; Grishin, Nick V.

2017-01-01

Abstract Oxynetra aureopecta sp. n. is described from the Sierra Madre Oriental of east-central Mexico. Visually similar to Mesoamerican O. hopfferi Staudinger, 1888 in having five orange bands on the abdomen above, it is diagnosed by orange forecoxae and palpi beneath, narrower forewing hyaline bands and a prominent 6% difference in the COI DNA barcode sequence. It is the northernmost representative of the hopfferi species group that also includes O. stangelandi Grishin & Burns, 2013, characterized by a single-banded abdomen and currently known only from the Area de Conservación Guanacaste in northwestern Costa Rica. Both O. hopfferi and O. stangelandi possess white forecoxae and ventral palpi. This new discovery brings the total number of Oxynetra C. & R. Felder, 1862 species to five. PMID:28769639

rbcL and matK Earn Two Thumbs Up as the Core DNA Barcode for Ferns

PubMed Central

Li, Fay-Wei; Kuo, Li-Yaung; Rothfels, Carl J.; Ebihara, Atsushi; Chiou, Wen-Liang; Windham, Michael D.; Pryer, Kathleen M.

2011-01-01

Background DNA barcoding will revolutionize our understanding of fern ecology, most especially because the accurate identification of the independent but cryptic gametophyte phase of the fern's life history—an endeavor previously impossible—will finally be feasible. In this study, we assess the discriminatory power of the core plant DNA barcode (rbcL and matK), as well as alternatively proposed fern barcodes (trnH-psbA and trnL-F), across all major fern lineages. We also present plastid barcode data for two genera in the hyperdiverse polypod clade—Deparia (Woodsiaceae) and the Cheilanthes marginata group (currently being segregated as a new genus of Pteridaceae)—to further evaluate the resolving power of these loci. Principal Findings Our results clearly demonstrate the value of matK data, previously unavailable in ferns because of difficulties in amplification due to a major rearrangement of the plastid genome. With its high sequence variation, matK complements rbcL to provide a two-locus barcode with strong resolving power. With sequence variation comparable to matK, trnL-F appears to be a suitable alternative barcode region in ferns, and perhaps should be added to the core barcode region if universal primer development for matK fails. In contrast, trnH-psbA shows dramatically reduced sequence variation for the majority of ferns. This is likely due to the translocation of this segment of the plastid genome into the inverted repeat regions, which are known to have a highly constrained substitution rate. Conclusions Our study provides the first endorsement of the two-locus barcode (rbcL+matK) in ferns, and favors trnL-F over trnH-psbA as a potential back-up locus. Future work should focus on gathering more fern matK sequence data to facilitate universal primer development. PMID:22028918

Identification of Dendrobium species by a candidate DNA barcode sequence: the chloroplast psbA-trnH intergenic region.

PubMed

Yao, Hui; Song, Jing-Yuan; Ma, Xin-Ye; Liu, Chang; Li, Ying; Xu, Hong-Xi; Han, Jian-Ping; Duan, Li-Sheng; Chen, Shi-Lin

2009-05-01

DNA barcoding is a novel technology that uses a standard DNA sequence to facilitate species identification. Although a consensus has not been reached regarding which DNA sequences can be used as the best plant barcodes, the psbA-trnH spacer region has been tested extensively in recent years. In this study, we hypothesize that the psbA-trnH spacer regions are also effective barcodes for Dendrobium species. We have sequenced the chloroplast psbA-trnH intergenic spacers of 17 Dendrobium species to test this hypothesis. The sequences were found to be significantly different from those of other species, with percentages of variation ranging from 0.3 % to 2.3 % and an average of 1.2 %. In contrast, the intraspecific variation among the Dendrobium species studied ranged from 0 % to 0.1 %. The sequence difference between the psbA-trnH sequences of 17 Dendrobium species and one Bulbophyllum odoratissimum ranged from 2.0 % to 3.1 %, with an average of 2.5 %. Our results support the notion that the psbA-trnH intergenic spacer region could be used as a barcode to distinguish various Dendrobium species and to differentiate Dendrobium species from other adulterating species. Copyright Georg Thieme Verlag KG Stuttgart. New York.

Barcode ITS2: a useful tool for identifying Trachelospermum jasminoides and a good monitor for medicine market.

PubMed

Yu, Ning; Wei, Yu-Long; Zhang, Xin; Zhu, Ning; Wang, Yan-Li; Zhu, Yue; Zhang, Hai-Ping; Li, Fen-Mei; Yang, Lan; Sun, Jia-Qi; Sun, Ai-Dong

2017-07-11

Trachelospermum jasminoides is commonly used in traditional Chinese medicine. However, the use of the plant's local alternatives is frequent, causing potential clinical problems. The T. jasminoides sold in the medicine market is commonly dried and sliced, making traditional identification methods difficult. In this study, the ITS2 region was evaluated on 127 sequences representing T. jasminoides and its local alternatives according to PCR and sequencing rates, intra- and inter-specific divergences, secondary structure, and discrimination capacity. Results indicated the 100% success rates of PCR and sequencing and the obvious presence of a barcoding gap. Results of BLAST 1, nearest distance and neighbor-joining tree methods showed that barcode ITS2 could successfully identify all the texted samples. The secondary structures of the ITS2 region provided another dimensionality for species identification. Two-dimensional images were obtained for better and easier identification. Previous studies on DNA barcoding concentrated more on the same family, genus, or species. However, an ideal barcode should be variable enough to identify closely related species. Meanwhile, the barcodes should also be conservative in identifying distantly related species. This study highlights the application of barcode ITS2 in solving practical problems in the distantly related local alternatives of medical plants.

DNA Barcoding for Species Identification of Insect Skins: A Test on Chironomidae (Diptera) Pupal Exuviae

PubMed Central

Ekrem, Torbjørn; Stur, Elisabeth

2017-01-01

Abstract Chironomidae (Diptera) pupal exuviae samples are commonly used for biological monitoring of aquatic habitats. DNA barcoding has proved useful for species identification of chironomid life stages containing cellular tissue, but the barcoding success of chironomid pupal exuviae is unknown. We assessed whether standard DNA barcoding could be efficiently used for species identification of chironomid pupal exuviae when compared with morphological techniques and if there were differences in performance between temperate and tropical ecosystems, subfamilies, and tribes. PCR, sequence, and identification success differed significantly between geographic regions and taxonomic groups. For Norway, 27 out of 190 (14.2%) of pupal exuviae resulted in high-quality chironomid sequences that match species. For Costa Rica, 69 out of 190 (36.3%) Costa Rican pupal exuviae resulted in high-quality sequences, but none matched known species. Standard DNA barcoding of chironomid pupal exuviae had limited success in species identification of unknown specimens due to contaminations and lack of matching references in available barcode libraries, especially from Costa Rica. Therefore, we recommend future biodiversity studies that focus their efforts on understudied regions, to simultaneously use morphological and molecular identification techniques to identify all life stages of chironomids and populate the barcode reference library with identified sequences.

DNA barcoding of Cuban freshwater fishes: evidence for cryptic species and taxonomic conflicts.

PubMed

Lara, Ariagna; Ponce de León, José Luis; Rodríguez, Rodet; Casane, Didier; Côté, Guillaume; Bernatchez, Louis; García-Machado, Erik

2010-05-01

Despite ongoing efforts to protect species and ecosystems in Cuba, habitat degradation, overuse and introduction of alien species have posed serious challenges to native freshwater fish species. In spite of the accumulated knowledge on the systematics of this freshwater ichthyofauna, recent results suggested that we are far from having a complete picture of the Cuban freshwater fish diversity. It is estimated that 40% of freshwater Cuban fish are endemic; however, this number may be even higher. Partial sequences (652 bp) of the mitochondrial gene COI (cytochrome c oxidase subunit I) were used to barcode 126 individuals, representing 27 taxonomically recognized species in 17 genera and 10 families. Analysis was based on Kimura 2-parameter genetic distances, and for four genera a character-based analysis (population aggregation analysis) was also used. The mean conspecific, congeneric and confamiliar genetic distances were 0.6%, 9.1% and 20.2% respectively. Molecular species identification was in concordance with current taxonomical classification in 96.4% of cases, and based on the neighbour-joining trees, in all but one instance, members of a given genera clustered within the same clade. Within the genus Gambusia, genetic divergence analysis suggests that there may be at least four cryptic species. In contrast, low genetic divergence and a lack of diagnostic sites suggest that Rivulus insulaepinorum may be conspecific with Rivulus cylindraceus. Distance and character-based analysis were completely concordant, suggesting that they complement species identification. Overall, the results evidenced the usefulness of the DNA barcodes for cataloguing Cuban freshwater fish species and for identifying those groups that deserve further taxonomic attention. © 2009 Blackwell Publishing Ltd.

Integrating feeding behavior, ecological data, and DNA barcoding to identify developmental differences in invertebrate foraging strategies in wild white-faced capuchins (Cebus capucinus).

PubMed

Mallott, Elizabeth K; Garber, Paul A; Malhi, Ripan S

2017-02-01

Invertebrate foraging strategies in nonhuman primates often require complex extractive foraging or prey detection techniques. As these skills take time to master, juveniles may have reduced foraging efficiency or concentrate their foraging efforts on easier to acquire prey than adults. We use DNA barcoding, behavioral observations, and ecological data to assess age-based differences in invertebrate prey foraging strategies in a group of white-faced capuchins (Cebus capucinus) in northeastern Costa Rica. Invertebrate availability was monitored using canopy traps and sweep netting. Fecal samples were collected from adult female, adult male, and juvenile white-faced capuchins (n = 225). COI mtDNA sequences were compared with known sequences in GenBank and the Barcode of Life Database. Frequencies of Lepidoptera and Hymenoptera consumption were higher in juveniles than in adults. A significantly smaller proportion of juvenile fecal samples contained Gryllidae and Cercopidae sequences, compared with adults (0% and 4.2% vs. 4.6% and 12.5%), and a significantly larger proportion contained Tenthredinidae, Culicidae, and Crambidae (5.6%, 9.7%, and 5.6% vs. 1.3%, 0.7%, and 1.3%). Juveniles spent significantly more time feeding and foraging than adults, and focused their foraging efforts on prey that require different skills to capture or extract. Arthropod availability was not correlated with foraging efficiency, and the rate of consumption of specific orders of invertebrates was not correlated with the availability of those same taxa. Our data support the hypothesis that juveniles are concentrating their foraging efforts on different prey than adults, potentially focusing their foraging efforts on more easily acquired types of prey. © 2016 Wiley Periodicals, Inc.

DNA Barcoding of Shark Meats Identify Species Composition and CITES-Listed Species from the Markets in Taiwan

PubMed Central

Liu, Shang-Yin Vanson; Chan, Chia-Ling Carynn; Lin, Oceana; Hu, Chieh-Shen; Chen, Chaolun Allen

2013-01-01

Background An increasing awareness of the vulnerability of sharks to exploitation by shark finning has contributed to a growing concern about an unsustainable shark fishery. Taiwan’s fleet has the 4th largest shark catch in the world, accounting for almost 6% of the global figures. Revealing the diversity of sharks consumed by Taiwanese is important in designing conservation plans. However, fins make up less than 5% of the total body weight of a shark, and their bodies are sold as filets in the market, making it difficult or impossible to identify species using morphological traits. Methods In the present study, we adopted a DNA barcoding technique using a 391-bp fragment of the mitochondrial cytochrome oxidase I (COI) gene to examine the diversity of shark filets and fins collected from markets and restaurants island-wide in Taiwan. Results Amongst the 548 tissue samples collected and sequenced, 20 major clusters were apparent by phylogenetic analyses, each of them containing individuals belonging to the same species (most with more than 95% bootstrap values), corresponding to 20 species of sharks. Additionally, Alopias pelagicus, Carcharhinus falciformis, Isurus oxyrinchus, and Prionace glauca consisted of 80% of the samples we collected, indicating that these species might be heavily consumed in Taiwan. Approximately 5% of the tissue samples used in this study were identified as species listed in CITES Appendix II, including two species of Sphyrna, C. longimanus and Carcharodon carcharias. Conclusion DNA barcoding provides an alternative method for understanding shark species composition when species-specific data is unavailable. Considering the global population decline, stock assessments of Appendix II species and highly consumed species are needed to accomplish the ultimate goal of shark conservation. PMID:24260209

DNA barcoding of shark meats identify species composition and CITES-listed species from the markets in Taiwan.

PubMed

Liu, Shang-Yin Vanson; Chan, Chia-Ling Carynn; Lin, Oceana; Hu, Chieh-Shen; Chen, Chaolun Allen

2013-01-01

An increasing awareness of the vulnerability of sharks to exploitation by shark finning has contributed to a growing concern about an unsustainable shark fishery. Taiwan's fleet has the 4th largest shark catch in the world, accounting for almost 6% of the global figures. Revealing the diversity of sharks consumed by Taiwanese is important in designing conservation plans. However, fins make up less than 5% of the total body weight of a shark, and their bodies are sold as filets in the market, making it difficult or impossible to identify species using morphological traits. In the present study, we adopted a DNA barcoding technique using a 391-bp fragment of the mitochondrial cytochrome oxidase I (COI) gene to examine the diversity of shark filets and fins collected from markets and restaurants island-wide in Taiwan. Amongst the 548 tissue samples collected and sequenced, 20 major clusters were apparent by phylogenetic analyses, each of them containing individuals belonging to the same species (most with more than 95% bootstrap values), corresponding to 20 species of sharks. Additionally, Alopias pelagicus, Carcharhinus falciformis, Isurus oxyrinchus, and Prionace glauca consisted of 80% of the samples we collected, indicating that these species might be heavily consumed in Taiwan. Approximately 5% of the tissue samples used in this study were identified as species listed in CITES Appendix II, including two species of Sphyrna, C. longimanus and Carcharodon carcharias. DNA barcoding provides an alternative method for understanding shark species composition when species-specific data is unavailable. Considering the global population decline, stock assessments of Appendix II species and highly consumed species are needed to accomplish the ultimate goal of shark conservation.

Testing the utility of matK and ITS DNA regions for discrimination of Allium species

USDA-ARS?s Scientific Manuscript database

Molecular phylogenetic analysis of the genus Allium L. has been mainly based on the nucleotide sequences of ITS region. In 2009 matK and rbcL were accepted as a two-locus DNA barcode to classify plant species by the Consortium for the Barcode of Life (CBOL) Plant Working Group. MatK region has been ...

Discriminating plants using the DNA barcode rbcLb: an appraisal based on a large data set.

PubMed

Dong, Wenpan; Cheng, Tao; Li, Changhao; Xu, Chao; Long, Ping; Chen, Chumming; Zhou, Shiliang

2014-03-01

The ideal DNA barcode for plants remains to be discovered, and the candidate barcode rbcL has been met with considerable skepticism since its proposal. In fact, the variability within this gene has never been fully explored across all plant groups from algae to flowering plants, and its performance as a barcode has not been adequately tested. By analysing all of the rbcL sequences currently available in GenBank, we attempted to determine how well a region of rbcL performs as a barcode in species discrimination. We found that the rbcLb region was more variable than the frequently used rbcLa region. Both universal and plant group-specific primers were designed to amplify rbcLb, and the performance of rbcLa and rbcLb was tested in several ways. Using blast, both regions successfully identified all families and nearly all genera; however, the successful species identification rates varied significantly among plant groups, ranging from 24.58% to 85.50% for rbcLa and from 36.67% to 90.89% for rbcLb. Successful species discrimination ranged from 5.19% to 96.33% for rbcLa and from 22.09% to 98.43% for rbcLb in species-rich families, and from 0 to 88.73% for rbcLa and from 2.04% to 100% for rbcLb in species-rich genera. Both regions performed better for lower plants than for higher plants, although rbcLb performed significantly better than rbcLa overall, particularly for angiosperms. Considering the applicability across plants, easy and unambiguous alignment, high primer universality, high sequence quality and high species discrimination power for lower plants, we suggest rbcLb as a universal plant barcode. © 2013 John Wiley & Sons Ltd.

A checklist of the bats of Peninsular Malaysia and progress towards a DNA barcode reference library.

PubMed

Lim, Voon-Ching; Ramli, Rosli; Bhassu, Subha; Wilson, John-James

2017-01-01

Several published checklists of bat species have covered Peninsular Malaysia as part of a broader region and/or in combination with other mammal groups. Other researchers have produced comprehensive checklists for specific localities within the peninsula. To our knowledge, a comprehensive checklist of bats specifically for the entire geopolitical region of Peninsular Malaysia has never been published, yet knowing which species are present in Peninsular Malaysia and their distributions across the region are crucial in developing suitable conservation plans. Our literature search revealed that 110 bat species have been documented in Peninsular Malaysia; 105 species have precise locality records while five species lack recent and/or precise locality records. We retrieved 18 species from records dated before the year 2000 and seven species have only ever been recorded once. Our search of Barcode of Life Datasystems (BOLD) found that 86 (of the 110) species have public records of which 48 species have public DNA barcodes available from bats sampled in Peninsular Malaysia. Based on Neighbour-Joining tree analyses and the allocation of DNA barcodes to Barcode Index Number system (BINs) by BOLD, several DNA barcodes recorded under the same species name are likely to represent distinct taxa. We discuss these cases in detail and highlight the importance of further surveys to determine the occurences and resolve the taxonomy of particular bat species in Peninsular Malaysia, with implications for conservation priorities.

Molecular Identification of Dendrobium Species (Orchidaceae) Based on the DNA Barcode ITS2 Region and Its Application for Phylogenetic Study.

PubMed

Feng, Shangguo; Jiang, Yan; Wang, Shang; Jiang, Mengying; Chen, Zhe; Ying, Qicai; Wang, Huizhong

2015-09-11

The over-collection and habitat destruction of natural Dendrobium populations for their commercial medicinal value has led to these plants being under severe threat of extinction. In addition, many Dendrobium plants are similarly shaped and easily confused during the absence of flowering stages. In the present study, we examined the application of the ITS2 region in barcoding and phylogenetic analyses of Dendrobium species (Orchidaceae). For barcoding, ITS2 regions of 43 samples in Dendrobium were amplified. In combination with sequences from GenBank, the sequences were aligned using Clustal W and genetic distances were computed using MEGA V5.1. The success rate of PCR amplification and sequencing was 100%. There was a significant divergence between the inter- and intra-specific genetic distances of ITS2 regions, while the presence of a barcoding gap was obvious. Based on the BLAST1, nearest distance and TaxonGAP methods, our results showed that the ITS2 regions could successfully identify the species of most Dendrobium samples examined; Second, we used ITS2 as a DNA marker to infer phylogenetic relationships of 64 Dendrobium species. The results showed that cluster analysis using the ITS2 region mainly supported the relationship between the species of Dendrobium established by traditional morphological methods and many previous molecular analyses. To sum up, the ITS2 region can not only be used as an efficient barcode to identify Dendrobium species, but also has the potential to contribute to the phylogenetic analysis of the genus Dendrobium.

Molecular Identification of Dendrobium Species (Orchidaceae) Based on the DNA Barcode ITS2 Region and Its Application for Phylogenetic Study

PubMed Central

Feng, Shangguo; Jiang, Yan; Wang, Shang; Jiang, Mengying; Chen, Zhe; Ying, Qicai; Wang, Huizhong

2015-01-01

The over-collection and habitat destruction of natural Dendrobium populations for their commercial medicinal value has led to these plants being under severe threat of extinction. In addition, many Dendrobium plants are similarly shaped and easily confused during the absence of flowering stages. In the present study, we examined the application of the ITS2 region in barcoding and phylogenetic analyses of Dendrobium species (Orchidaceae). For barcoding, ITS2 regions of 43 samples in Dendrobium were amplified. In combination with sequences from GenBank, the sequences were aligned using Clustal W and genetic distances were computed using MEGA V5.1. The success rate of PCR amplification and sequencing was 100%. There was a significant divergence between the inter- and intra-specific genetic distances of ITS2 regions, while the presence of a barcoding gap was obvious. Based on the BLAST1, nearest distance and TaxonGAP methods, our results showed that the ITS2 regions could successfully identify the species of most Dendrobium samples examined; Second, we used ITS2 as a DNA marker to infer phylogenetic relationships of 64 Dendrobium species. The results showed that cluster analysis using the ITS2 region mainly supported the relationship between the species of Dendrobium established by traditional morphological methods and many previous molecular analyses. To sum up, the ITS2 region can not only be used as an efficient barcode to identify Dendrobium species, but also has the potential to contribute to the phylogenetic analysis of the genus Dendrobium. PMID:26378526

Application of DNA Barcodes in Asian Tropical Trees – A Case Study from Xishuangbanna Nature Reserve, Southwest China

PubMed Central

Conran, John G.; Li, Jie

2015-01-01

Background Within a regional floristic context, DNA barcoding is more useful to manage plant diversity inventories on a large scale and develop valuable conservation strategies. However, there are no DNA barcode studies from tropical areas of China, which represents one of the biodiversity hotspots around the world. Methodology and Principal Findings A DNA barcoding database of an Asian tropical trees with high diversity was established at Xishuangbanna Nature Reserve, Yunnan, southwest China using rbcL and matK as standard barcodes, as well as trnH–psbA and ITS as supplementary barcodes. The performance of tree species identification success was assessed using 2,052 accessions from four plots belonging to two vegetation types in the region by three methods: Neighbor-Joining, Maximum-Likelihood and BLAST. We corrected morphological field identification errors (9.6%) for the three plots using rbcL and matK based on Neighbor-Joining tree. The best barcode region for PCR and sequencing was rbcL (97.6%, 90.8%), followed by trnH–psbA (93.6%, 85.6%), while matK and ITS obtained relative low PCR and sequencing success rates. However, ITS performed best for both species (44.6–58.1%) and genus (72.8–76.2%) identification. With trnH–psbA slightly less effective for species identification. The two standard barcode rbcL and matK gave poor results for species identification (24.7–28.5% and 31.6–35.3%). Compared with other studies from comparable tropical forests (e.g. Cameroon, the Amazon and India), the overall performance of the four barcodes for species identification was lower for the Xishuangbanna Nature Reserve, possibly because of species/genus ratios and species composition between these tropical areas. Conclusions/Significance Although the core barcodes rbcL and matK were not suitable for species identification of tropical trees from Xishuangbanna Nature Reserve, they could still help with identification at the family and genus level. Considering the relative sequence recovery and the species identification performance, we recommend the use of trnH–psbA and ITS in combination as the preferred barcodes for tropical tree species identification in China. PMID:26121045

DNA barcodes for Mexican Cactaceae, plants under pressure from wild collecting.

PubMed

Yesson, Chris; Bárcenas, Rolando T; Hernández, Héctor M; Ruiz-Maqueda, María de la Luz; Prado, Alberto; Rodríguez, Víctor M; Hawkins, Julie A

2011-09-01

DNA barcodes could be a useful tool for plant conservation. Of particular importance is the ability to identify unknown plant material, such as from customs seizures of illegally collected specimens. Mexican cacti are an example of a threatened group, under pressure because of wild collection for the xeriscaping trade and private collectors. Mexican cacti also provide a taxonomically and geographically coherent group with which to test DNA barcodes. Here, we sample the matK barcode for 528 species of Cactaceae including approximately 75% of Mexican species and test the utility of the matK region for species-level identification. We find that the matK DNA barcode can be used to identify uniquely 77% of species sampled, and 79-87% of species of particular conservation importance. However, this is far below the desired rate of 95% and there are significant issues for PCR amplification because of the variability of primer sites. Additionally, we test the nuclear ITS regions for the cactus subfamily Opuntioideae and for the genus Ariocarpus (subfamily Cactoideae). We observed higher rates of variation for ITS (86% unique for Opuntioideae sampled) but a much lower PCR success, encountering significant intra-individual polymorphism in Ariocarpus precluding the use of this marker in this taxon. We conclude that the matK region should provide useful information as a DNA barcode for Cactaceae if the problems with primers can be addressed, but matK alone is not sufficiently variable to achieve species-level identification. Additional complementary regions should be investigated as ITS is shown to be unsuitable. © 2011 Blackwell Publishing Ltd.

A DNA Mini-Barcoding System for Authentication of Processed Fish Products.

PubMed

Shokralla, Shadi; Hellberg, Rosalee S; Handy, Sara M; King, Ian; Hajibabaei, Mehrdad

2015-10-30

Species substitution is a form of seafood fraud for the purpose of economic gain. DNA barcoding utilizes species-specific DNA sequence information for specimen identification. Previous work has established the usability of short DNA sequences-mini-barcodes-for identification of specimens harboring degraded DNA. This study aims at establishing a DNA mini-barcoding system for all fish species commonly used in processed fish products in North America. Six mini-barcode primer pairs targeting short (127-314 bp) fragments of the cytochrome c oxidase I (CO1) DNA barcode region were developed by examining over 8,000 DNA barcodes from species in the U.S. Food and Drug Administration (FDA) Seafood List. The mini-barcode primer pairs were then tested against 44 processed fish products representing a range of species and product types. Of the 44 products, 41 (93.2%) could be identified at the species or genus level. The greatest mini-barcoding success rate found with an individual primer pair was 88.6% compared to 20.5% success rate achieved by the full-length DNA barcode primers. Overall, this study presents a mini-barcoding system that can be used to identify a wide range of fish species in commercial products and may be utilized in high throughput DNA sequencing for authentication of heavily processed fish products.

Species discovery and diversity in Lobocriconema (Criconematidae: Nematoda) and related plant-parasitic nematodes from North American ecoregions.

PubMed

Powers, T O; Bernard, E C; Harris, T; Higgins, R; Olson, M; Olson, S; Lodema, M; Matczyszyn, J; Mullin, P; Sutton, L; Powers, K S

2016-03-03

There are many nematode species that, following formal description, are seldom mentioned again in the scientific literature. Lobocriconema thornei and L. incrassatum are two such species, described from North American forests, respectively 37 and 49 years ago. In the course of a 3-year nematode biodiversity survey of North American ecoregions, specimens resembling Lobocriconema species appeared in soil samples from both grassland and forested sites. Using a combination of molecular and morphological analyses, together with a set of species delimitation approaches, we have expanded the known range of these species, added to the species descriptions, and discovered a related group of species that form a monophyletic group with the two described species. In this study, 148 specimens potentially belonging to the genus Lobocriconema were isolated from soil, individually measured, digitally imaged, and DNA barcoded using a 721 bp region of cytochrome oxidase subunit 1 (COI). One-third of the specimens were also analyzed using amplified DNA from the 3' region of the small subunit ribosomal RNA gene (18SrDNA) and the adjacent first internal transcribed spacer (ITS1). Eighteen mitochondrial haplotype groups, falling into four major clades, were identified by well-supported nodes in Bayesian and maximum likelihood trees and recognized as distinct lineages by species delimitation metrics. Discriminant function analysis of a set of morphological characters indicated that the major clades in the dataset possessed a strong morphological signal that decreased in comparisons of haplotype groups within clades. Evidence of biogeographic and phylogeographic patterns was apparent in the dataset. COI haplotype diversity was high in the southern Appalachian Mountains and Gulf Coast states and lessened in northern temperate forests. Lobocriconema distribution suggests the existence of phylogeographic patterns associated with recolonization of formerly glaciated regions by eastern deciduous forest, but definitive glacial refugia for this group of plant parasitic nematodes have yet to be identified. Unlike agricultural pest species of plant-parasitic nematodes, there is little evidence of long-distance dispersal in Lobocriconema as revealed by haplotype distribution. Most haplotype groups were characterized by low levels of intragroup genetic variation and large genetic distances between haplotype groups. The localization of nematode haplotypes together with their characteristic plant communities could provide insight into the historical formation of these belowground biotic communities.

DNA barcodes for 1/1000 of the animal kingdom.

PubMed

Hebert, Paul D N; Dewaard, Jeremy R; Landry, Jean-François

2010-06-23

This study reports DNA barcodes for more than 1300 Lepidoptera species from the eastern half of North America, establishing that 99.3 per cent of these species possess diagnostic barcode sequences. Intraspecific divergences averaged just 0.43 per cent among this assemblage, but most values were lower. The mean was elevated by deep barcode divergences (greater than 2%) in 5.1 per cent of the species, often involving the sympatric occurrence of two barcode clusters. A few of these cases have been analysed in detail, revealing species overlooked by the current taxonomic system. This study also provided a large-scale test of the extent of regional divergence in barcode sequences, indicating that geographical differentiation in the Lepidoptera of eastern North America is small, even when comparisons involve populations as much as 2800 km apart. The present results affirm that a highly effective system for the identification of Lepidoptera in this region can be built with few records per species because of the limited intra-specific variation. As most terrestrial and marine taxa are likely to possess a similar pattern of population structure, an effective DNA-based identification system can be developed with modest effort.

Microgeographic Genetic Variation of the Malaria Vector Anopheles darlingi Root (Diptera: Culicidae) from Córdoba and Antioquia, Colombia

PubMed Central

Gutiérrez, Lina A.; Gómez, Giovan F.; González, John J.; Castro, Martha I.; Luckhart, Shirley; Conn, Jan E.; Correa, Margarita M.

2010-01-01

Anopheles darlingi is an important vector of Plasmodium spp. in several malaria-endemic regions of Colombia. This study was conducted to test genetic variation of An. darlingi at a microgeographic scale (approximately 100 km) from localities in Córdoba and Antioquia states, in western Colombia, to better understand the potential contribution of population genetics to local malaria control programs. Microsatellite loci: nuclear white and cytochrome oxidase subunit I (COI) gene sequences were analyzed. The northern white gene lineage was exclusively distributed in Córdoba and Antioquia and shared COI haplotypes were highly represented in mosquitoes from both states. COI analyses showed these An. darlingi are genetically closer to Central American populations than southern South American populations. Overall microsatellites and COI analysis showed low to moderate genetic differentiation among populations in northwestern Colombia. Given the existence of high gene flow between An. darlingi populations of Córdoba and Antioquia, integrated vector control strategies could be developed in this region of Colombia. PMID:20595475

The necessity of DNA taxonomy to reveal cryptic diversity and spatial distribution of meiofauna, with a focus on Nemertea.

PubMed

Leasi, Francesca; Norenburg, Jon L

2014-01-01

Meiofauna represent one of the most abundant and diverse communities in marine benthic ecosystems. However, an accurate assessment of diversity at the level of species has been and remains challenging for these microscopic organisms. Therefore, for many taxa, especially the soft body forms such as nemerteans, which often lack clear diagnostic morphological traits, DNA taxonomy is an effective means to assess species diversity. Morphological taxonomy of Nemertea is well documented as complicated by scarcity of unambiguous character states and compromised by diagnoses of a majority of species (and higher clades) being inadequate or based on ambiguous characters and character states. Therefore, recent studies have advocated for the primacy of molecular tools to solve the taxonomy of this group. DNA taxonomy uncovers possible hidden cryptic species, provides a coherent means to systematize taxa in definite clades, and also reveals possible biogeographic patterns. Here, we analyze diversity of nemertean species by considering the barcode region of the mitochondrial gene Cytochrome Oxidase subunit I (COI) and different species delineation approaches in order to infer evolutionarily significant units. In the aim to uncover actual diversity of meiofaunal nemerteans across different sites in Central America, COI sequences were obtained for specimens assigned here to the genera Cephalothrix, Ototyphlonemertes, and Tetrastemma-like worms, each commonly encountered in our sampling. Additional genetic, taxonomic, and geographic data of other specimens belonging to these genera were added from GenBank. Results are consistent across different DNA taxonomy approaches, and revealed (i) the presence of several hidden cryptic species and (ii) numerous potential misidentifications due to traditional taxonomy. (iii) We additionally test a possible biogeographic pattern of taxonomic units revealed by this study, and, except for a few cases, the putative species seem not to be widely distributed, in contrast to what traditional taxonomy would suggest for the recognized morphotypes.

Molecular identification and phylogenetic analysis of important medicinal plant species in genus Paeonia based on rDNA-ITS, matK, and rbcL DNA barcode sequences.

PubMed

Kim, W J; Ji, Y; Choi, G; Kang, Y M; Yang, S; Moon, B C

2016-08-05

This study was performed to identify and analyze the phylogenetic relationship among four herbaceous species of the genus Paeonia, P. lactiflora, P. japonica, P. veitchii, and P. suffruticosa, using DNA barcodes. These four species, which are commonly used in traditional medicine as Paeoniae Radix and Moutan Radicis Cortex, are pharmaceutically defined in different ways in the national pharmacopoeias in Korea, Japan, and China. To authenticate the different species used in these medicines, we evaluated rDNA-internal transcribed spacers (ITS), matK and rbcL regions, which provide information capable of effectively distinguishing each species from one another. Seventeen samples were collected from different geographic regions in Korea and China, and DNA barcode regions were amplified using universal primers. Comparative analyses of these DNA barcode sequences revealed species-specific nucleotide sequences capable of discriminating the four Paeonia species. Among the entire sequences of three barcodes, marker nucleotides were identified at three positions in P. lactiflora, eleven in P. japonica, five in P. veitchii, and 25 in P. suffruticosa. Phylogenetic analyses also revealed four distinct clusters showing homogeneous clades with high resolution at the species level. The results demonstrate that the analysis of these three DNA barcode sequences is a reliable method for identifying the four Paeonia species and can be used to authenticate Paeoniae Radix and Moutan Radicis Cortex at the species level. Furthermore, based on the assessment of amplicon sizes, inter/intra-specific distances, marker nucleotides, and phylogenetic analysis, rDNA-ITS was the most suitable DNA barcode for identification of these species.

A checklist of the bats of Peninsular Malaysia and progress towards a DNA barcode reference library

PubMed Central

Ramli, Rosli; Bhassu, Subha

2017-01-01

Several published checklists of bat species have covered Peninsular Malaysia as part of a broader region and/or in combination with other mammal groups. Other researchers have produced comprehensive checklists for specific localities within the peninsula. To our knowledge, a comprehensive checklist of bats specifically for the entire geopolitical region of Peninsular Malaysia has never been published, yet knowing which species are present in Peninsular Malaysia and their distributions across the region are crucial in developing suitable conservation plans. Our literature search revealed that 110 bat species have been documented in Peninsular Malaysia; 105 species have precise locality records while five species lack recent and/or precise locality records. We retrieved 18 species from records dated before the year 2000 and seven species have only ever been recorded once. Our search of Barcode of Life Datasystems (BOLD) found that 86 (of the 110) species have public records of which 48 species have public DNA barcodes available from bats sampled in Peninsular Malaysia. Based on Neighbour-Joining tree analyses and the allocation of DNA barcodes to Barcode Index Number system (BINs) by BOLD, several DNA barcodes recorded under the same species name are likely to represent distinct taxa. We discuss these cases in detail and highlight the importance of further surveys to determine the occurences and resolve the taxonomy of particular bat species in Peninsular Malaysia, with implications for conservation priorities. PMID:28742835

Testing the Efficacy of DNA Barcodes for Identifying the Vascular Plants of Canada.

PubMed

Braukmann, Thomas W A; Kuzmina, Maria L; Sills, Jesse; Zakharov, Evgeny V; Hebert, Paul D N

2017-01-01

Their relatively slow rates of molecular evolution, as well as frequent exposure to hybridization and introgression, often make it difficult to discriminate species of vascular plants with the standard barcode markers (rbcL, matK, ITS2). Previous studies have examined these constraints in narrow geographic or taxonomic contexts, but the present investigation expands analysis to consider the performance of these gene regions in discriminating the species in local floras at sites across Canada. To test identification success, we employed a DNA barcode reference library with sequence records for 96% of the 5108 vascular plant species known from Canada, but coverage varied from 94% for rbcL to 60% for ITS2 and 39% for matK. Using plant lists from 27 national parks and one scientific reserve, we tested the efficacy of DNA barcodes in identifying the plants in simulated species assemblages from six biogeographic regions of Canada using BLAST and mothur. Mean pairwise distance (MPD) and mean nearest taxon distance (MNTD) were strong predictors of barcode performance for different plant families and genera, and both metrics supported ITS2 as possessing the highest genetic diversity. All three genes performed strongly in assigning the taxa present in local floras to the correct genus with values ranging from 91% for rbcL to 97% for ITS2 and 98% for matK. However, matK delivered the highest species discrimination (~81%) followed by ITS2 (~72%) and rbcL (~44%). Despite the low number of plant taxa in the Canadian Arctic, DNA barcodes had the least success in discriminating species from this biogeographic region with resolution ranging from 36% with rbcL to 69% with matK. Species resolution was higher in the other settings, peaking in the Woodland region at 52% for rbcL and 87% for matK. Our results indicate that DNA barcoding is very effective in identifying Canadian plants to a genus, and that it performs well in discriminating species in regions where floristic diversity is highest.

Testing the Efficacy of DNA Barcodes for Identifying the Vascular Plants of Canada

PubMed Central

Kuzmina, Maria L.; Sills, Jesse; Zakharov, Evgeny V.; Hebert, Paul D. N.

2017-01-01

Their relatively slow rates of molecular evolution, as well as frequent exposure to hybridization and introgression, often make it difficult to discriminate species of vascular plants with the standard barcode markers (rbcL, matK, ITS2). Previous studies have examined these constraints in narrow geographic or taxonomic contexts, but the present investigation expands analysis to consider the performance of these gene regions in discriminating the species in local floras at sites across Canada. To test identification success, we employed a DNA barcode reference library with sequence records for 96% of the 5108 vascular plant species known from Canada, but coverage varied from 94% for rbcL to 60% for ITS2 and 39% for matK. Using plant lists from 27 national parks and one scientific reserve, we tested the efficacy of DNA barcodes in identifying the plants in simulated species assemblages from six biogeographic regions of Canada using BLAST and mothur. Mean pairwise distance (MPD) and mean nearest taxon distance (MNTD) were strong predictors of barcode performance for different plant families and genera, and both metrics supported ITS2 as possessing the highest genetic diversity. All three genes performed strongly in assigning the taxa present in local floras to the correct genus with values ranging from 91% for rbcL to 97% for ITS2 and 98% for matK. However, matK delivered the highest species discrimination (~81%) followed by ITS2 (~72%) and rbcL (~44%). Despite the low number of plant taxa in the Canadian Arctic, DNA barcodes had the least success in discriminating species from this biogeographic region with resolution ranging from 36% with rbcL to 69% with matK. Species resolution was higher in the other settings, peaking in the Woodland region at 52% for rbcL and 87% for matK. Our results indicate that DNA barcoding is very effective in identifying Canadian plants to a genus, and that it performs well in discriminating species in regions where floristic diversity is highest. PMID:28072819

The Application of DNA Barcodes for the Identification of Marine Crustaceans from the North Sea and Adjacent Regions.

PubMed

Raupach, Michael J; Barco, Andrea; Steinke, Dirk; Beermann, Jan; Laakmann, Silke; Mohrbeck, Inga; Neumann, Hermann; Kihara, Terue C; Pointner, Karin; Radulovici, Adriana; Segelken-Voigt, Alexandra; Wesse, Christina; Knebelsberger, Thomas

2015-01-01

During the last years DNA barcoding has become a popular method of choice for molecular specimen identification. Here we present a comprehensive DNA barcode library of various crustacean taxa found in the North Sea, one of the most extensively studied marine regions of the world. Our data set includes 1,332 barcodes covering 205 species, including taxa of the Amphipoda, Copepoda, Decapoda, Isopoda, Thecostraca, and others. This dataset represents the most extensive DNA barcode library of the Crustacea in terms of species number to date. By using the Barcode of Life Data Systems (BOLD), unique BINs were identified for 198 (96.6%) of the analyzed species. Six species were characterized by two BINs (2.9%), and three BINs were found for the amphipod species Gammarus salinus Spooner, 1947 (0.4%). Intraspecific distances with values higher than 2.2% were revealed for 13 species (6.3%). Exceptionally high distances of up to 14.87% between two distinct but monophyletic clusters were found for the parasitic copepod Caligus elongatus Nordmann, 1832, supporting the results of previous studies that indicated the existence of an overlooked sea louse species. In contrast to these high distances, haplotype-sharing was observed for two decapod spider crab species, Macropodia parva Van Noort & Adema, 1985 and Macropodia rostrata (Linnaeus, 1761), underlining the need for a taxonomic revision of both species. Summarizing the results, our study confirms the application of DNA barcodes as highly effective identification system for the analyzed marine crustaceans of the North Sea and represents an important milestone for modern biodiversity assessment studies using barcode sequences.

Molecular species delimitation and its implications for species descriptions using desmosomatid and nannoniscid isopods from the VEMA fracture zone as example taxa

NASA Astrophysics Data System (ADS)

Brix, Saskia; Bober, Simon; Tschesche, Claudia; Kihara, Terue-Cristina; Driskell, Amy; Jennings, Robert M.

2018-02-01

We found 72 species for COI and 45 for 16 S by species delimitation among 186 (from 195 extracted) desmosomatid (144) and nannoniscid (42) sequenced specimens of a total of > 400 specimens for both families. Multiple "discovery"-type species delimitation methods were used, so that consistency across methods could be assessed: The ABGD analysis detected a barcode gap of 3-6% for COI and 4-6% for 16 S, in the whole dataset. Most putative species have a horizontally limited distribution along the Vema fracture zone, although the details depend in part on the interpretation of species delimitation analyses. Putative species were mostly restricted to the eastern or western Vema fracture zone, with only eight crossing the complete Vema fracture zone. Our data suggest that even robustly-sampled species exhibited small ranges; the range estimates calculable from present data were around 500 km, and three were on the order of 1000-2500 km. We chose an abundant, but geographically restricted species (Eugerdella egoni Tschesche and Brix sp. nov.) collected at a single site in the Vema transform fault, and two species (Prochelator barnacki Bober and Brix sp. nov. and Whoia sockei Brix and Kihara sp. nov.) with a broad, but disjunct distribution in the Vema fracture zone for taxonomic description.

Epizoanthus spp. associations revealed using DNA markers: a case study from Kochi, Japan.

PubMed

Reimer, James Davis; Hirose, Mamiko; Nishisaka, Taiki; Sinniger, Frederic; Itani, Gyo

2010-09-01

Zoanthids (Cnidaria, Hexacorallia) of the genus Epizoanthus are often found in association with other marine invertebrates, including gastropods and hermit crabs. However, little information exists on the specificity and nature of these associations due to a lack of investigation into Epizoanthus species diversity, and the taxonomy of Epizoanthus is therefore confused. In this study, analyses of morphological data (tentacle number, polyp size, etc) and molecular data (mitochondrial cytochrome oxidase subunit 1 = COI, 16S ribosomal DNA = 16S rDNA) were used to examine Epizoanthus specimens from Tosa Bay, Kochi, Japan. The Epizoanthus specimens were found on both live gastropods (Gemmula unedo) and hermit crabs (Paguristes palythophilus) inhabiting G. unedo and G. cosmoi shells. While morphological analyses did not show clear differences between examined specimens, both COI and mt 16S rDNA clearly divided the specimens into two groups, one associated only with hermit crabs (= Epizoanthus sp. C), and another associated only with living gastropods (= Epizoanthus sp. S). Unexpectedly, DNA sequences from both groups did not match with two previously reported Epizoanthus species from Japan (E. indicus, E. ramosus), indicating they both may be undescribed species. These results highlight the utility of DNA "barcoding" of unknown zoanthids, and will provide a foundation for re-examinations of Epizoanthus species diversity and specificity, which will be critical in understanding the evolution of these unique marine invertebrates.

Intraspecific phylogeography of Lasmigona subviridis (Bivalvia: Unionidae): Conservation implications of range discontinuity

USGS Publications Warehouse

King, T.L.; Eackles, M.S.; Gjetvaj, B.; Hoeh, W.R.

1999-01-01

A nucleotide sequence analysis of the first internal transcribed spacer region (ITS-1) between the 5.8S and 18S ribosomal DNA genes (640 bp) and cytochrome c oxidase subunit I (COI) of mitochondrial DNA (mtDNA) (576 bp) was conducted for the freshwater bivalve Lasmigona subviridis and three congeners to determine the utility of these regions in identifying phylogeographic and phylogenetic structure. Sequence analysis of the ITS-1 region indicated a zone of discontinuity in the genetic population structure between a group of L. subviridis populations inhabiting the Susquehanna and Potomac Rivers and more southern populations. Moreover, haplotype patterns resulting from variation in the COI region suggested an absence of gene exchange between tributaries within two different river drainages, as well as between adjacent rivers systems. The authors recommend that the northern and southern populations, which are reproductively isolated and constitute evolutionarily significant lineages, be managed as separate conservation units. Results from the COI region suggest that, in some cases, unionid relocations should be avoided between tributaries of the same drainage because these populations may have been reproductively isolated for thousands of generations. Therefore, unionid bivalves distributed among discontinuous habitats (e.g. Atlantic slope drainages) potentially should be considered evolutionarily distinct. The DNA sequence divergences observed in the nuclear and mtDNA regions among the Lasmigona species were congruent, although the level of divergence in the COI region was up to three times greater. The genus Lasmigona, as represented by the four species surveyed in this study, may not be monophyletic.

Exploring the Leaf Beetle Fauna (Coleoptera: Chrysomelidae) of an Ecuadorian Mountain Forest Using DNA Barcoding.

PubMed

Thormann, Birthe; Ahrens, Dirk; Marín Armijos, Diego; Peters, Marcell K; Wagner, Thomas; Wägele, Johann W

2016-01-01

Tropical mountain forests are hotspots of biodiversity hosting a huge but little known diversity of insects that is endangered by habitat destruction and climate change. Therefore, rapid assessment approaches of insect diversity are urgently needed to complement slower traditional taxonomic approaches. We empirically compare different DNA-based species delimitation approaches for a rapid biodiversity assessment of hyperdiverse leaf beetle assemblages along an elevational gradient in southern Ecuador and explore their effect on species richness estimates. Based on a COI barcode data set of 674 leaf beetle specimens (Coleoptera: Chrysomelidae) of 266 morphospecies from three sample sites in the Podocarpus National Park, we employed statistical parsimony analysis, distance-based clustering, GMYC- and PTP-modelling to delimit species-like units and compared them to morphology-based (parataxonomic) species identifications. The four different approaches for DNA-based species delimitation revealed highly similar numbers of molecular operational taxonomic units (MOTUs) (n = 284-289). Estimated total species richness was considerably higher than the sampled amount, 414 for morphospecies (Chao2) and 469-481 for the different MOTU types. Assemblages at different elevational levels (1000 vs. 2000 m) had similar species numbers but a very distinct species composition for all delimitation methods. Most species were found only at one elevation while this turnover pattern was even more pronounced for DNA-based delimitation. Given the high congruence of DNA-based delimitation results, probably due to the sampling structure, our study suggests that when applied to species communities on a regionally limited level with high amount of rare species (i.e. ~50% singletons), the choice of species delimitation method can be of minor relevance for assessing species numbers and turnover in tropical insect communities. Therefore, DNA-based species delimitation is confirmed as a valuable tool for evaluating biodiversity of hyperdiverse insect communities, especially when exact taxonomic identifications are missing.

Exploring the Leaf Beetle Fauna (Coleoptera: Chrysomelidae) of an Ecuadorian Mountain Forest Using DNA Barcoding

PubMed Central

Thormann, Birthe; Ahrens, Dirk; Marín Armijos, Diego; Peters, Marcell K.; Wagner, Thomas; Wägele, Johann W.

2016-01-01

Background Tropical mountain forests are hotspots of biodiversity hosting a huge but little known diversity of insects that is endangered by habitat destruction and climate change. Therefore, rapid assessment approaches of insect diversity are urgently needed to complement slower traditional taxonomic approaches. We empirically compare different DNA-based species delimitation approaches for a rapid biodiversity assessment of hyperdiverse leaf beetle assemblages along an elevational gradient in southern Ecuador and explore their effect on species richness estimates. Methodology/Principal Findings Based on a COI barcode data set of 674 leaf beetle specimens (Coleoptera: Chrysomelidae) of 266 morphospecies from three sample sites in the Podocarpus National Park, we employed statistical parsimony analysis, distance-based clustering, GMYC- and PTP-modelling to delimit species-like units and compared them to morphology-based (parataxonomic) species identifications. The four different approaches for DNA-based species delimitation revealed highly similar numbers of molecular operational taxonomic units (MOTUs) (n = 284–289). Estimated total species richness was considerably higher than the sampled amount, 414 for morphospecies (Chao2) and 469–481 for the different MOTU types. Assemblages at different elevational levels (1000 vs. 2000 m) had similar species numbers but a very distinct species composition for all delimitation methods. Most species were found only at one elevation while this turnover pattern was even more pronounced for DNA-based delimitation. Conclusions/Significance Given the high congruence of DNA-based delimitation results, probably due to the sampling structure, our study suggests that when applied to species communities on a regionally limited level with high amount of rare species (i.e. ~50% singletons), the choice of species delimitation method can be of minor relevance for assessing species numbers and turnover in tropical insect communities. Therefore, DNA-based species delimitation is confirmed as a valuable tool for evaluating biodiversity of hyperdiverse insect communities, especially when exact taxonomic identifications are missing. PMID:26849826

Conflict-of-interest disclosure at medical journals in Japan: a nationwide survey of the practices of journal secretariats

PubMed Central

Kojima, Takako; Green, Joseph; Barron, J Patrick

2015-01-01

Objectives Medical journals in Japan generally have appropriate policies regarding disclosure of conflicts of interest (COI). However, COI management depends on the staff members of each journal's editorial secretariat. This study's objectives were to find out (A) whether COI disclosure and the journal's role in it are clearly understood by the journal’s secretariat staff, (B) how much experience the editorial secretariat has in actually handling issues related to disclosure and (C) what kind of help or support they need. Setting and design In January 2014, questionnaires were sent to the editorial secretariats of journal-publishing societies belonging to the Japanese Association of Medical Sciences (JAMS). Participants The response rate was 100%, and the respondents represented 121 journals published by the 118 JAMS member societies (at the time of the survey). Primary and secondary outcome measures Information was collected on the history of COI policies and on how those policies were implemented. At the end of the questionnaire, there was an open-ended call for comments. Results Compulsory COI disclosure began between 2010 and 2013 for 60.3% of the journals (73/121). Handling of COI issues was not uniform: 17.4% (21/121) of respondents do not pursue cases of dubious disclosure, and 47.9% (58/121) do not require COI disclosures from editorial board members. Very few of the editorial secretariats had clearly-stated consequences for violations of COI-disclosure policy (33/121, 27.3%), and only 28.9% offered COI education (35/121). Respondents’ comments indicated that uniform, easily-searchable guidance regarding COI policies and implementation would be welcome. Conclusions Although commitment is widespread, policy implementation is inconsistent and COI experience is lacking. Clear, easy-to-use guidelines are desired by many societies. The JAMS is to be commended for supporting this country-wide investigation; other countries and regions are encouraged to perform similar investigations to respond to needs regarding COI management. PMID:26310399

Using barcoded Zika virus to assess virus population structure in vitro and in Aedes aegypti mosquitoes.

PubMed

Weger-Lucarelli, James; Garcia, Selene M; Rückert, Claudia; Byas, Alex; O'Connor, Shelby L; Aliota, Matthew T; Friedrich, Thomas C; O'Connor, David H; Ebel, Gregory D

2018-06-20

Arboviruses such as Zika virus (ZIKV, Flaviviridae; Flavivirus) must replicate in both mammalian and insect hosts possessing strong immune defenses. Accordingly, transmission between and replication within hosts involves genetic bottlenecks, during which viral population size and genetic diversity may be significantly reduced. To help quantify these bottlenecks and their effects, we constructed 4 "barcoded" ZIKV populations that theoretically contain thousands of barcodes each. After identifying the most diverse barcoded virus, we passaged this virus 3 times in 2 mammalian and mosquito cell lines and characterized the population using deep sequencing of the barcoded region of the genome. C6/36 maintain higher barcode diversity, even after 3 passages, than Vero. Additionally, field-caught mosquitoes exposed to the virus to assess bottlenecks in a natural host. A progressive reduction in barcode diversity occurred throughout systemic infection of these mosquitoes. Differences in bottlenecks during systemic spread were observed between different populations of Aedes aegypti. Copyright © 2018. Published by Elsevier Inc.

Systematics of testudacarine torrent mites (Acari, Hydrachnidia, Torrenticolidae) with descriptions of 13 new species from North America

PubMed Central

O’Neill, Joseph C.; Fisher, J. Ray; Nelson, Whitney A.; Skvarla, Micheal J.; Fisher, Danielle M.; Dowling, Ashley P. G.

2016-01-01

Abstract Thirteen new species of North American Testudacarus (Torrenticolidae: Testudacarinae) are described: Testudacarus deceptivus O’Neill & Dowling, sp. n., Testudacarus hitchensi O’Neill & Dowling, sp. n., Testudacarus harrisi O’Neill & Dowling, sp. n., Testudacarus dennetti O’Neill & Dowling, sp. n., Testudacarus dawkinsi O’Neill & Dowling, sp. n., Testudacarus radwellae O’Neill & Dowling, sp. n., Testudacarus kirkwoodae O’Neill & Dowling, sp. n., Testudacarus hyporhynchus O’Neill & Dowling, sp. n., Testudacarus smithi O’Neill & Dowling, sp. n., Testudacarus rollerae O’Neill & Dowling, sp. n., Testudacarus elongatus O’Neill & Dowling, sp. n., Testudacarus rectangulatus O’Neill & Dowling, sp. n., and Testudacarus oblongatus O’Neill & Dowling, sp. n. Testudacarus vulgaris Habeeb, 1954 is resurrected from synonymy with Testudacarus minimus and redescribed. Debsacarus (Habeeb, 1961), Testudacarus americanus Marshall, 1943, and Testudacarus minimus Marshall, 1943 are redescribed. All redescriptions are from original types. Species delimination was accomplished through examination of morphology, biogeography, and molecular phylogenetics of the barcoding region of COI. Other species are addressed and a key to world species is presented. For Testudacarinae, this represents the first published: 1) descriptions from multiple specimens (i.e. intraspecific variation); 2) colored photographs; 3) explicit illustrations and discussion of sexual dimorphism within the subfamily; 4) genetic data. A comprehensive testudacarine reference list is also included. PMID:27199586

Jasmonate perception by inositol-phosphate-potentiated COI1-JAZ co-receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sheard, Laura B; Tan, Xu; Mao, Haibin

2011-11-07

Jasmonates are a family of plant hormones that regulate plant growth, development and responses to stress. The F-box protein CORONATINE INSENSITIVE 1 (COI1) mediates jasmonate signalling by promoting hormone-dependent ubiquitylation and degradation of transcriptional repressor JAZ proteins. Despite its importance, the mechanism of jasmonate perception remains unclear. Here we present structural and pharmacological data to show that the true Arabidopsis jasmonate receptor is a complex of both COI1 and JAZ. COI1 contains an open pocket that recognizes the bioactive hormone (3R,7S)-jasmonoyl-l-isoleucine (JA-Ile) with high specificity. High-affinity hormone binding requires a bipartite JAZ degron sequence consisting of a conserved {alpha}-helix formore » COI1 docking and a loop region to trap the hormone in its binding pocket. In addition, we identify a third critical component of the jasmonate co-receptor complex, inositol pentakisphosphate, which interacts with both COI1 and JAZ adjacent to the ligand. Our results unravel the mechanism of jasmonate perception and highlight the ability of F-box proteins to evolve as multi-component signalling hubs.« less

A DNA mini-barcode for land plants.

PubMed

Little, Damon P

2014-05-01

Small portions of the barcode region - mini-barcodes - may be used in place of full-length barcodes to overcome DNA degradation for samples with poor DNA preservation. 591,491,286 rbcL mini-barcode primer combinations were electronically evaluated for PCR universality, and two novel highly universal sets of priming sites were identified. Novel and published rbcL mini-barcode primers were evaluated for PCR amplification [determined with a validated electronic simulation (n = 2765) and empirically (n = 188)], Sanger sequence quality [determined empirically (n = 188)], and taxonomic discrimination [determined empirically (n = 30,472)]. PCR amplification for all mini-barcodes, as estimated by validated electronic simulation, was successful for 90.2-99.8% of species. Overall Sanger sequence quality for mini-barcodes was very low - the best mini-barcode tested produced sequences of adequate quality (B20 ≥ 0.5) for 74.5% of samples. The majority of mini-barcodes provide correct identifications of families in excess of 70.1% of the time. Discriminatory power noticeably decreased at lower taxonomic levels. At the species level, the discriminatory power of the best mini-barcode was less than 38.2%. For samples believed to contain DNA from only one species, an investigator should attempt to sequence, in decreasing order of utility and probability of success, mini-barcodes F (rbcL1/rbcLB), D (F52/R193) and K (F517/R604). For samples believed to contain DNA from more than one species, an investigator should amplify and sequence mini-barcode D (F52/R193). © 2013 John Wiley & Sons Ltd.

Identification of Fabaceae plants using the DNA barcode matK.

PubMed

Gao, Ting; Sun, Zhiying; Yao, Hui; Song, Jingyuan; Zhu, Yingjie; Ma, Xinye; Chen, Shilin

2011-01-01

In this study, we tested the applicability of the core DNA barcode MATK for identifying species within the Fabaceae family. Based on an evaluation of genetic variation, DNA barcoding gaps, and species discrimination power, MATK is a useful barcode for Fabaceae species. Of 1355 plant samples collected from 1079 species belonging to 409 diverse genera, MATK precisely identified approximately 80 % and 96 % of them at the species and genus levels, respectively. Therefore, our research indicates that the MATK region is a valuable marker for plant species within Fabaceae. © Georg Thieme Verlag KG Stuttgart · New York.

Lineage divergence detected in the malaria vector Anopheles marajoara (Diptera: Culicidae) in Amazonian Brazil

PubMed Central

2010-01-01

Background Cryptic species complexes are common among anophelines. Previous phylogenetic analysis based on the complete mtDNA COI gene sequences detected paraphyly in the Neotropical malaria vector Anopheles marajoara. The "Folmer region" detects a single taxon using a 3% divergence threshold. Methods To test the paraphyletic hypothesis and examine the utility of the Folmer region, genealogical trees based on a concatenated (white + 3' COI sequences) dataset and pairwise differentiation of COI fragments were examined. The population structure and demographic history were based on partial COI sequences for 294 individuals from 14 localities in Amazonian Brazil. 109 individuals from 12 localities were sequenced for the nDNA white gene, and 57 individuals from 11 localities were sequenced for the ribosomal DNA (rDNA) internal transcribed spacer 2 (ITS2). Results Distinct A. marajoara lineages were detected by combined genealogical analysis and were also supported among COI haplotypes using a median joining network and AMOVA, with time since divergence during the Pleistocene (<100,000 ya). COI sequences at the 3' end were more variable, demonstrating significant pairwise differentiation (3.82%) compared to the more moderate 2.92% detected by the Folmer region. Lineage 1 was present in all localities, whereas lineage 2 was restricted mainly to the west. Mismatch distributions for both lineages were bimodal, likely due to multiple colonization events and spatial expansion (~798 - 81,045 ya). There appears to be gene flow within, not between lineages, and a partial barrier was detected near Rio Jari in Amapá state, separating western and eastern populations. In contrast, both nDNA data sets (white gene sequences with or without the retention of the 4th intron, and ITS2 sequences and length) detected a single A. marajoara lineage. Conclusions Strong support for combined data with significant differentiation detected in the COI and absent in the nDNA suggest that the divergence is recent, and detectable only by the faster evolving mtDNA. A within subgenus threshold of >2% may be more appropriate among sister taxa in cryptic anopheline complexes than the standard 3%. Differences in demographic history and climatic changes may have contributed to mtDNA lineage divergence in A. marajoara. PMID:20929572

A genetic fingerprint of Amphipoda from Icelandic waters – the baseline for further biodiversity and biogeography studies

PubMed Central

Jażdżewska, Anna M.; Corbari, Laure; Driskell, Amy; Frutos, Inmaculada; Havermans, Charlotte; Hendrycks, Ed; Hughes, Lauren; Lörz, Anne-Nina; Bente Stransky; Tandberg, Anne Helene S.; Vader, Wim; Brix, Saskia

2018-01-01

Abstract Amphipods constitute an abundant part of Icelandic deep-sea zoobenthos yet knowledge of the diversity of this fauna, particularly at the molecular level, is scarce. The present work aims to use molecular methods to investigate genetic variation of the Amphipoda sampled during two IceAGE collecting expeditions. The mitochondrial cytochrome oxidase subunit 1 (COI) of 167 individuals originally assigned to 75 morphospecies was analysed. These targeted morhospecies were readily identifiable by experts using light microscopy and representative of families where there is current ongoing taxonomic research. The study resulted in 81 Barcode Identity Numbers (BINs) (of which >90% were published for the first time), while Automatic Barcode Gap Discovery revealed the existence of 78 to 83 Molecular Operational Taxonomic Units (MOTUs). Six nominal species (Rhachotropis helleri, Arrhis phyllonyx, Deflexilodes tenuirostratus, Paroediceros propinquus, Metopa boeckii, Astyra abyssi) appeared to have a molecular variation higher than the 0.03 threshold of both p-distance and K2P usually used for amphipod species delineation. Conversely, two Oedicerotidae regarded as separate morphospecies clustered together with divergences in the order of intraspecific variation. The incongruence between the BINs associated with presently identified species and the publicly available data of the same taxa was observed in case of Paramphithoe hystrix and Amphilochus manudens. The findings from this research project highlight the necessity of supporting molecular studies with thorough morphology species analyses. PMID:29472762

Range and Frequency of Africanized Honey Bees in California (USA)

PubMed Central

Kono, Yoshiaki; Kohn, Joshua R.

2015-01-01

Africanized honey bees entered California in 1994 but few accounts of their northward expansion or their frequency relative to European honey bees have been published. We used mitochondrial markers and morphometric analyses to determine the prevalence of Africanized honeybees in San Diego County and their current northward progress in California west of the Sierra Nevada crest. The northernmost African mitotypes detected were approximately 40 km south of Sacramento in California’s central valley. In San Diego County, 65% of foraging honey bee workers carry African mitochondria and the estimated percentage of Africanized workers using morphological measurements is similar (61%). There was no correlation between mitotype and morphology in San Diego County suggesting Africanized bees result from bidirectional hybridization. Seventy percent of feral hives, but only 13% of managed hives, sampled in San Diego County carried the African mitotype indicating that a large fraction of foraging workers in both urban and rural San Diego County are feral. We also found a single nucleotide polymorphism at the DNA barcode locus COI that distinguishes European and African mitotypes. The utility of this marker was confirmed using 401 georeferenced honey bee sequences from the worldwide Barcode of Life Database. Future censuses can determine whether the current range of the Africanized form is stable, patterns of introgression at nuclear loci, and the environmental factors that may limit the northern range of the Africanized honey bee. PMID:26361047

DNA barcoding and wing morphometrics to distinguish three Aedes vectors in Thailand.

PubMed

Sumruayphol, Suchada; Apiwathnasorn, Chamnarn; Ruangsittichai, Jiraporn; Sriwichai, Patchara; Attrapadung, Siriluck; Samung, Yudthana; Dujardin, Jean-Pierre

2016-07-01

Aedes aegypti (Diptera: Culicidae) (L.), Ae. albopictus (Skuse), and Ae. scutellaris (Walker) are important mosquito vectors of dengue and chikungunya viruses. They are morphologically similar and sympatric in some parts of their distribution; therefore, there is a risk of incorrect morphological identification. Any confusion could have a negative impact on epidemiological studies or control strategies. Therefore, we explored two modern tools to supplement current morphological identification: DNA barcoding and geometric morphometric analyses. Field larvae were reared to adults and carefully classified based on morphological traits. The genetic analysis was based on the 658bp each of 30COI sequences. Some Culex spp., Mansonia bonneae, were included as outgroups, and inclusion of a few other Aedes spp. facilitated phylogenetic inference of the relationship between Ae. albopictus and Ae. scutellaris. The two species were separated by an average interspecific divergence of 0.123 (0.119-0.127). Morphometric examination included landmark- (392 specimens) and outline-based (317 specimens) techniques. The shape of the wing showed different discriminating power based on sex and digitizing technique. This is the first time that Ae. scutellaris and Ae. albopictus have been compared using these two techniques. We confirm that these morphologically close species are valid, and that geometric morphometrics can considerably increase the reliability of morphological identification. Copyright © 2016 Elsevier B.V. All rights reserved.

A new record of blue-spotted seabream Pagrus caeruleostictus from Chinese coastal waters documented from morphology and DNA barcoding

NASA Astrophysics Data System (ADS)

Chen, Yongxia; Liu, Jing; Wu, Renxie

2015-03-01

A new record of Pagrus caeruleostictus (Valenciennes, 1830), collected from the Beibu Gulf, South China Sea in April 2013, was documented based on morphology and cytochrome oxidase I subunit (COI) gene barcoding analyses. It can be distinguished by a combination of the following characteristics: head scaled to above eyes; cheeks with 5 or 6 rows of scales; lateral line scales 51-52; 5 rows of scales above the lateral line; 4 and 6 cuspidate teeth in front of upper and lower jaws, respectively, followed by 2 rows of blunter teeth posteriorly; gill rakers on first arch 12 to 15; D XI-XII +9-11; A III +8-9; the first two dorsal spines very short, the third to fifth extended, filamentous in the young; the first pelvic ray filamentous; silvery pink with dark blue spots on back and sides; caudal fin pinkish; other fins bluish or pinkish; the blue spots generally disappear in large specimens. The present report of P. caeruleostictus suggests that its distribution in Chinese coastal waters may be attributed to human effort, since this species is unlikely to have expanded naturally from the eastern Atlantic to the South China Sea, with no other records from the Indian or Pacific Oceans. We recommend that a precautionary approach should be adopted for the management of P. caeruleostictus.

A report on identification of sequence polymorphism in barcode region of six commercially important Cymbopogon species.

PubMed

Bishoyi, Ashok Kumar; Kavane, Aarti; Sharma, Anjali; Geetha, K A

2017-02-01

CYMBOPOGON: is an important member of grass family Poaceae, cultivated for essential oils which have greater medicinal and industrial value. Taxonomic identification of Cymbopogon species is determined mainly by morphological markers, odour of essential oils and concentration of bioactive compounds present in the oil matrices which are highly influenced by environment. Authenticated molecular marker based taxonomical identification is also lacking in the genus; hence effort was made to evaluate potential DNA barcode loci in six commercially important Cymbopogon species for their individual discrimination and authentication at the species level. Four widely used DNA barcoding regions viz., ITS 1 & ITS 2 spacers, matK, psbA-trnH and rbcL were taken for the study. Gene sequences of the same or related genera of the concerned loci were mined from NCBI domain and primers were designed and validated for barcode loci amplification. Out of the four loci studied, sequences from matK and ITS spacer loci revealed 0.46% and 5.64% nucleotide sequence diversity, respectively whereas the other two loci i.e., psbA-trnH and rbcL showed 100% sequence homology. The newly developed primers can be used for barcode loci amplification in the genus Cymbopogon. The identified Single Nucleotide Polymorphisms from the studied sequences may be used as barcodes for the six Cymbopogon species. The information generated can also be utilized for barcode development of the genus by including more number of Cymbopgon species in future.

Ultra-barcoding in cacao (Theobroma spp.; Malvaceae) using whole chloroplast genomes and nuclear ribosomal DNA.

PubMed

Kane, Nolan; Sveinsson, Saemundur; Dempewolf, Hannes; Yang, Ji Yong; Zhang, Dapeng; Engels, Johannes M M; Cronk, Quentin

2012-02-01

To reliably identify lineages below the species level such as subspecies or varieties, we propose an extension to DNA-barcoding using next-generation sequencing to produce whole organellar genomes and substantial nuclear ribosomal sequence. Because this method uses much longer versions of the traditional DNA-barcoding loci in the plastid and ribosomal DNA, we call our approach ultra-barcoding (UBC). We used high-throughput next-generation sequencing to scan the genome and generate reliable sequence of high copy number regions. Using this method, we examined whole plastid genomes as well as nearly 6000 bases of nuclear ribosomal DNA sequences for nine genotypes of Theobroma cacao and an individual of the related species T. grandiflorum, as well as an additional publicly available whole plastid genome of T. cacao. All individuals of T. cacao examined were uniquely distinguished, and evidence of reticulation and gene flow was observed. Sequence variation was observed in some of the canonical barcoding regions between species, but other regions of the chloroplast were more variable both within species and between species, as were ribosomal spacers. Furthermore, no single region provides the level of data available using the complete plastid genome and rDNA. Our data demonstrate that UBC is a viable, increasingly cost-effective approach for reliably distinguishing varieties and even individual genotypes of T. cacao. This approach shows great promise for applications where very closely related or interbreeding taxa must be distinguished.

Evaluation of suitable DNA regions for molecular identification of high value medicinal plants in genus Kaempferia.

PubMed

Osathanunkul, Maslin; Dheeranupattana, Srisulak; Rotarayanont, Siriphron; Sookkhee, Siriwoot; Osathanunkul, Khukrit; Madesis, Panagiotis

2017-12-02

DNA barcoding coupled high resolution melting (Bar-HRM) is an emerging method for species discrimination based on DNA dissociation kinetics. The aim of this work was to evaluate the suitability of different primer sets, derived from selected DNA regions, for Bar-HRM analysis of species in Kaempferia (Zingiberaceae). Four primer pairs were evaluated (rbcL, rpoC, trnL and ITS1). It was observed that the ITS1 barcode was the most useful DNA barcoding region overall for species discrimination out of all of the regions and primers assessed. Thus, the primer pair derived from the ITS1 region was the single most effective region for the identification of the tested species, whereas the rbcL primer pair gave the lowest resolution. Our Bar-HRM developed here would not only be useful for identification of Kaempferia plant specimens lacking essential parts for morphological identification but will be useful for authenticating products in powdered form of a high value medicinal species Kaempferia parviflora, in particular.

A two-locus global DNA barcode for land plants: the coding rbcL gene complements the non-coding trnH-psbA spacer region.

PubMed

Kress, W John; Erickson, David L

2007-06-06

A useful DNA barcode requires sufficient sequence variation to distinguish between species and ease of application across a broad range of taxa. Discovery of a DNA barcode for land plants has been limited by intrinsically lower rates of sequence evolution in plant genomes than that observed in animals. This low rate has complicated the trade-off in finding a locus that is universal and readily sequenced and has sufficiently high sequence divergence at the species-level. Here, a global plant DNA barcode system is evaluated by comparing universal application and degree of sequence divergence for nine putative barcode loci, including coding and non-coding regions, singly and in pairs across a phylogenetically diverse set of 48 genera (two species per genus). No single locus could discriminate among species in a pair in more than 79% of genera, whereas discrimination increased to nearly 88% when the non-coding trnH-psbA spacer was paired with one of three coding loci, including rbcL. In silico trials were conducted in which DNA sequences from GenBank were used to further evaluate the discriminatory power of a subset of these loci. These trials supported the earlier observation that trnH-psbA coupled with rbcL can correctly identify and discriminate among related species. A combination of the non-coding trnH-psbA spacer region and a portion of the coding rbcL gene is recommended as a two-locus global land plant barcode that provides the necessary universality and species discrimination.

Detecting very low allele fraction variants using targeted DNA sequencing and a novel molecular barcode-aware variant caller.

PubMed

Xu, Chang; Nezami Ranjbar, Mohammad R; Wu, Zhong; DiCarlo, John; Wang, Yexun

2017-01-03

Detection of DNA mutations at very low allele fractions with high accuracy will significantly improve the effectiveness of precision medicine for cancer patients. To achieve this goal through next generation sequencing, researchers need a detection method that 1) captures rare mutation-containing DNA fragments efficiently in the mix of abundant wild-type DNA; 2) sequences the DNA library extensively to deep coverage; and 3) distinguishes low level true variants from amplification and sequencing errors with high accuracy. Targeted enrichment using PCR primers provides researchers with a convenient way to achieve deep sequencing for a small, yet most relevant region using benchtop sequencers. Molecular barcoding (or indexing) provides a unique solution for reducing sequencing artifacts analytically. Although different molecular barcoding schemes have been reported in recent literature, most variant calling has been done on limited targets, using simple custom scripts. The analytical performance of barcode-aware variant calling can be significantly improved by incorporating advanced statistical models. We present here a highly efficient, simple and scalable enrichment protocol that integrates molecular barcodes in multiplex PCR amplification. In addition, we developed smCounter, an open source, generic, barcode-aware variant caller based on a Bayesian probabilistic model. smCounter was optimized and benchmarked on two independent read sets with SNVs and indels at 5 and 1% allele fractions. Variants were called with very good sensitivity and specificity within coding regions. We demonstrated that we can accurately detect somatic mutations with allele fractions as low as 1% in coding regions using our enrichment protocol and variant caller.

The Application of DNA Barcodes for the Identification of Marine Crustaceans from the North Sea and Adjacent Regions

PubMed Central

Raupach, Michael J.; Barco, Andrea; Steinke, Dirk; Beermann, Jan; Laakmann, Silke; Mohrbeck, Inga; Neumann, Hermann; Kihara, Terue C.; Pointner, Karin; Radulovici, Adriana; Segelken-Voigt, Alexandra; Wesse, Christina; Knebelsberger, Thomas

2015-01-01

During the last years DNA barcoding has become a popular method of choice for molecular specimen identification. Here we present a comprehensive DNA barcode library of various crustacean taxa found in the North Sea, one of the most extensively studied marine regions of the world. Our data set includes 1,332 barcodes covering 205 species, including taxa of the Amphipoda, Copepoda, Decapoda, Isopoda, Thecostraca, and others. This dataset represents the most extensive DNA barcode library of the Crustacea in terms of species number to date. By using the Barcode of Life Data Systems (BOLD), unique BINs were identified for 198 (96.6%) of the analyzed species. Six species were characterized by two BINs (2.9%), and three BINs were found for the amphipod species Gammarus salinus Spooner, 1947 (0.4%). Intraspecific distances with values higher than 2.2% were revealed for 13 species (6.3%). Exceptionally high distances of up to 14.87% between two distinct but monophyletic clusters were found for the parasitic copepod Caligus elongatus Nordmann, 1832, supporting the results of previous studies that indicated the existence of an overlooked sea louse species. In contrast to these high distances, haplotype-sharing was observed for two decapod spider crab species, Macropodia parva Van Noort & Adema, 1985 and Macropodia rostrata (Linnaeus, 1761), underlining the need for a taxonomic revision of both species. Summarizing the results, our study confirms the application of DNA barcodes as highly effective identification system for the analyzed marine crustaceans of the North Sea and represents an important milestone for modern biodiversity assessment studies using barcode sequences. PMID:26417993

Applying plant DNA barcodes to identify species of Parnassia (Parnassiaceae).

PubMed

Yang, Jun-Bo; Wang, Yi-Ping; Möller, Michael; Gao, Lian-Ming; Wu, Ding

2012-03-01

DNA barcoding is a technique to identify species by using standardized DNA sequences. In this study, a total of 105 samples, representing 30 Parnassia species, were collected to test the effectiveness of four proposed DNA barcodes (rbcL, matK, trnH-psbA and ITS) for species identification. Our results demonstrated that all four candidate DNA markers have a maximum level of primer universality and sequencing success. As a single DNA marker, the ITS region provided the highest species resolution with 86.7%, followed by trnH-psbA with 73.3%. The combination of the core barcode regions, matK+rbcL, gave the lowest species identification success (63.3%) among any combination of multiple markers and was found unsuitable as DNA barcode for Parnassia. The combination of ITS+trnH-psbA achieved the highest species discrimination with 90.0% resolution (27 of 30 sampled species), equal to the four-marker combination and higher than any two or three marker combination including rbcL or matK. Therefore, matK and rbcL should not be used as DNA barcodes for the species identification of Parnassia. Based on the overall performance, the combination of ITS+trnH-psbA is proposed as the most suitable DNA barcode for identifying Parnassia species. DNA barcoding is a useful technique and provides a reliable and effective mean for the discrimination of Parnassia species, and in combination with morphology-based taxonomy, will be a robust approach for tackling taxonomically complex groups. In the light of our findings, we found among the three species not identified a possible cryptic speciation event in Parnassia. © 2011 Blackwell Publishing Ltd.

DNA barcoding for identification of consumer-relevant mushrooms: A partial solution for product certification?

PubMed

Raja, Huzefa A; Baker, Timothy R; Little, Jason G; Oberlies, Nicholas H

2017-01-01

One challenge in the dietary supplement industry is confirmation of species identity for processed raw materials, i.e. those modified by milling, drying, or extraction, which move through a multilevel supply chain before reaching the finished product. This is particularly difficult for samples containing fungal mycelia, where processing removes morphological characteristics, such that they do not present sufficient variation to differentiate species by traditional techniques. To address this issue, we have demonstrated the utility of DNA barcoding to verify the taxonomic identity of fungi found commonly in the food and dietary supplement industry; such data are critical for protecting consumer health, by assuring both safety and quality. By using DNA barcoding of nuclear ribosomal internal transcribed spacer (ITS) of the rRNA gene with fungal specific ITS primers, ITS barcodes were generated for 33 representative fungal samples, all of which could be used by consumers for food and/or dietary supplement purposes. In the majority of cases, we were able to sequence the ITS region from powdered mycelium samples, grocery store mushrooms, and capsules from commercial dietary supplements. After generating ITS barcodes utilizing standard procedures accepted by the Consortium for the Barcode of Life, we tested their utility by performing a BLAST search against authenticate published ITS sequences in GenBank. In some cases, we also downloaded published, homologous sequences of the ITS region of fungi inspected in this study and examined the phylogenetic relationships of barcoded fungal species in light of modern taxonomic and phylogenetic studies. We anticipate that these data will motivate discussions on DNA barcoding based species identification as applied to the verification/certification of mushroom-containing dietary supplements. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Species-specific identification from incomplete sampling: applying DNA barcodes to monitoring invasive solanum plants.

PubMed

Zhang, Wei; Fan, Xiaohong; Zhu, Shuifang; Zhao, Hong; Fu, Lianzhong

2013-01-01

Comprehensive sampling is crucial to DNA barcoding, but it is rarely performed because materials are usually unavailable. In practice, only a few rather than all species of a genus are required to be identified. Thus identification of a given species using a limited sample is of great importance in current application of DNA barcodes. Here, we selected 70 individuals representing 48 species from each major lineage of Solanum, one of the most species-rich genera of seed plants, to explore whether DNA barcodes can provide reliable specific-species discrimination in the context of incomplete sampling. Chloroplast genes ndhF and trnS-trnG and the nuclear gene waxy, the commonly used markers in Solanum phylogeny, were selected as the supplementary barcodes. The tree-building and modified barcode gap methods were employed to assess species resolution. The results showed that four Solanum species of quarantine concern could be successfully identified through the two-step barcoding sampling strategy. In addition, discrepancies between nuclear and cpDNA barcodes in some samples demonstrated the ability to discriminate hybrid species, and highlights the necessity of using barcode regions with different modes of inheritance. We conclude that efficient phylogenetic markers are good candidates as the supplementary barcodes in a given taxonomic group. Critically, we hypothesized that a specific-species could be identified from a phylogenetic framework using incomplete sampling-through this, DNA barcoding will greatly benefit the current fields of its application.

DNA barcode goes two-dimensions: DNA QR code web server.

PubMed

Liu, Chang; Shi, Linchun; Xu, Xiaolan; Li, Huan; Xing, Hang; Liang, Dong; Jiang, Kun; Pang, Xiaohui; Song, Jingyuan; Chen, Shilin

2012-01-01

The DNA barcoding technology uses a standard region of DNA sequence for species identification and discovery. At present, "DNA barcode" actually refers to DNA sequences, which are not amenable to information storage, recognition, and retrieval. Our aim is to identify the best symbology that can represent DNA barcode sequences in practical applications. A comprehensive set of sequences for five DNA barcode markers ITS2, rbcL, matK, psbA-trnH, and CO1 was used as the test data. Fifty-three different types of one-dimensional and ten two-dimensional barcode symbologies were compared based on different criteria, such as coding capacity, compression efficiency, and error detection ability. The quick response (QR) code was found to have the largest coding capacity and relatively high compression ratio. To facilitate the further usage of QR code-based DNA barcodes, a web server was developed and is accessible at http://qrfordna.dnsalias.org. The web server allows users to retrieve the QR code for a species of interests, convert a DNA sequence to and from a QR code, and perform species identification based on local and global sequence similarities. In summary, the first comprehensive evaluation of various barcode symbologies has been carried out. The QR code has been found to be the most appropriate symbology for DNA barcode sequences. A web server has also been constructed to allow biologists to utilize QR codes in practical DNA barcoding applications.

Land plants and DNA barcodes: short-term and long-term goals.

PubMed

Chase, Mark W; Salamin, Nicolas; Wilkinson, Mike; Dunwell, James M; Kesanakurthi, Rao Prasad; Haider, Nadia; Haidar, Nadia; Savolainen, Vincent

2005-10-29

Land plants have had the reputation of being problematic for DNA barcoding for two general reasons: (i) the standard DNA regions used in algae, animals and fungi have exceedingly low levels of variability and (ii) the typically used land plant plastid phylogenetic markers (e.g. rbcL, trnL-F, etc.) appear to have too little variation. However, no one has assessed how well current phylogenetic resources might work in the context of identification (versus phylogeny reconstruction). In this paper, we make such an assessment, particularly with two of the markers commonly sequenced in land plant phylogenetic studies, plastid rbcL and internal transcribed spacers of the large subunits of nuclear ribosomal DNA (ITS), and find that both of these DNA regions perform well even though the data currently available in GenBank/EBI were not produced to be used as barcodes and BLAST searches are not an ideal tool for this purpose. These results bode well for the use of even more variable regions of plastid DNA (such as, for example, psbA-trnH) as barcodes, once they have been widely sequenced. In the short term, efforts to bring land plant barcoding up to the standards being used now in other organisms should make swift progress. There are two categories of DNA barcode users, scientists in fields other than taxonomy and taxonomists. For the former, the use of mitochondrial and plastid DNA, the two most easily assessed genomes, is at least in the short term a useful tool that permits them to get on with their studies, which depend on knowing roughly which species or species groups they are dealing with, but these same DNA regions have important drawbacks for use in taxonomic studies (i.e. studies designed to elucidate species limits). For these purposes, DNA markers from uniparentally (usually maternally) inherited genomes can only provide half of the story required to improve taxonomic standards being used in DNA barcoding. In the long term, we will need to develop more sophisticated barcoding tools, which would be multiple, low-copy nuclear markers with sufficient genetic variability and PCR-reliability; these would permit the detection of hybrids and permit researchers to identify the 'genetic gaps' that are useful in assessing species limits.

Land plants and DNA barcodes: short-term and long-term goals

PubMed Central

Chase, Mark W; Salamin, Nicolas; Wilkinson, Mike; Dunwell, James M; Kesanakurthi, Rao Prasad; Haidar, Nadia; Savolainen, Vincent

2005-01-01

Land plants have had the reputation of being problematic for DNA barcoding for two general reasons: (i) the standard DNA regions used in algae, animals and fungi have exceedingly low levels of variability and (ii) the typically used land plant plastid phylogenetic markers (e.g. rbcL, trnL-F, etc.) appear to have too little variation. However, no one has assessed how well current phylogenetic resources might work in the context of identification (versus phylogeny reconstruction). In this paper, we make such an assessment, particularly with two of the markers commonly sequenced in land plant phylogenetic studies, plastid rbcL and internal transcribed spacers of the large subunits of nuclear ribosomal DNA (ITS), and find that both of these DNA regions perform well even though the data currently available in GenBank/EBI were not produced to be used as barcodes and BLAST searches are not an ideal tool for this purpose. These results bode well for the use of even more variable regions of plastid DNA (such as, for example, psbA-trnH) as barcodes, once they have been widely sequenced. In the short term, efforts to bring land plant barcoding up to the standards being used now in other organisms should make swift progress. There are two categories of DNA barcode users, scientists in fields other than taxonomy and taxonomists. For the former, the use of mitochondrial and plastid DNA, the two most easily assessed genomes, is at least in the short term a useful tool that permits them to get on with their studies, which depend on knowing roughly which species or species groups they are dealing with, but these same DNA regions have important drawbacks for use in taxonomic studies (i.e. studies designed to elucidate species limits). For these purposes, DNA markers from uniparentally (usually maternally) inherited genomes can only provide half of the story required to improve taxonomic standards being used in DNA barcoding. In the long term, we will need to develop more sophisticated barcoding tools, which would be multiple, low-copy nuclear markers with sufficient genetic variability and PCR-reliability; these would permit the detection of hybrids and permit researchers to identify the ‘genetic gaps’ that are useful in assessing species limits. PMID:16214746

DNA barcoding for molecular identification of Demodex based on mitochondrial genes.

PubMed

Hu, Li; Yang, YuanJun; Zhao, YaE; Niu, DongLing; Yang, Rui; Wang, RuiLing; Lu, Zhaohui; Li, XiaoQi

2017-12-01

There has been no widely accepted DNA barcode for species identification of Demodex. In this study, we attempted to solve this issue. First, mitochondrial cox1-5' and 12S gene fragments of Demodex folloculorum, D. brevis, D. canis, and D. caprae were amplified, cloned, and sequenced for the first time; intra/interspecific divergences were computed and phylogenetic trees were reconstructed. Then, divergence frequency distribution plots of those two gene fragments were drawn together with mtDNA cox1-middle region and 16S obtained in previous studies. Finally, their identification efficiency was evaluated by comparing barcoding gap. Results indicated that 12S had the higher identification efficiency. Specifically, for cox1-5' region of the four Demodex species, intraspecific divergences were less than 2.0%, and interspecific divergences were 21.1-31.0%; for 12S, intraspecific divergences were less than 1.4%, and interspecific divergences were 20.8-26.9%. The phylogenetic trees demonstrated that the four Demodex species clustered separately, and divergence frequency distribution plot showed that the largest intraspecific divergence of 12S (1.4%) was less than cox1-5' region (2.0%), cox1-middle region (3.1%), and 16S (2.8%). The barcoding gap of 12S was 19.4%, larger than cox1-5' region (19.1%), cox1-middle region (11.3%), and 16S (13.0%); the interspecific divergence span of 12S was 6.2%, smaller than cox1-5' region (10.0%), cox1-middle region (14.1%), and 16S (11.4%). Moreover, 12S has a moderate length (517 bp) for sequencing at once. Therefore, we proposed mtDNA 12S was more suitable than cox1 and 16S to be a DNA barcode for classification and identification of Demodex at lower category level.

High-accuracy biodistribution analysis of adeno-associated virus variants by double barcode sequencing.

PubMed

Marsic, Damien; Méndez-Gómez, Héctor R; Zolotukhin, Sergei

2015-01-01

Biodistribution analysis is a key step in the evaluation of adeno-associated virus (AAV) capsid variants, whether natural isolates or produced by rational design or directed evolution. Indeed, when screening candidate vectors, accurate knowledge about which tissues are infected and how efficiently is essential. We describe the design, validation, and application of a new vector, pTR-UF50-BC, encoding a bioluminescent protein, a fluorescent protein and a DNA barcode, which can be used to visualize localization of transduction at the organism, organ, tissue, or cellular levels. In addition, by linking capsid variants to different barcoded versions of the vector and amplifying the barcode region from various tissue samples using barcoded primers, biodistribution of viral genomes can be analyzed with high accuracy and efficiency.

Molecular Authentication of the Traditional Medicinal Plant "Lakshman Booti" (Smithia conferta Sm.) and Its Adulterants through DNA Barcoding.

PubMed

Umdale, Suraj D; Kshirsagar, Parthraj R; Lekhak, Manoj M; Gaikwad, Nikhil B

2017-07-01

Smithia conferta Sm. is an annual herb widely used in Indian traditional medical practice and commonly known as "Lakshman booti" in Sanskrit. Morphological resemblance among the species of genus Smithia Aiton . leads to inaccurate identification and adulteration. This causes inconsistent therapeutic effects and also affects the quality of herbal medicine. This study aimed to generate potential barcode for authentication of S. conferta and its adulterants through DNA barcoding technique. Genomic DNA extracted from S. conferta and its adulterants was used as templates for polymerase chain reaction amplification of the barcoding regions. The amplicons were directed for sequencing, and species identification was conducted using BLASTn and unweighted pair-group method with arithmetic mean trees. In addition, the secondary structures of internal transcribed spacer (ITS) 2 region were predicted. The nucleotide sequence of ITS provides species-specific single nucleotide polymorphisms and sequence divergence (22%) than psb A- trn H (10.9%) and rbc L (3.1%) sequences. The ITS barcode indicates that S. conferta and Smithia sensitiva are closely related compared to other species. ITS is the most applicable barcode for molecular authentication of S. conferta , and further chloroplast barcodes should be tested for phylogenetic analysis of genus Smithia. The present investigation is the first effort of utilization of DNA barcode for molecular authentication of S. conferta and its adulterants. Also, this study expanded the application of the ITS2 sequence data in the authentication. The ITS has been proved as a potential and reliable candidate barcode for the authentication of S. conferta . Abbreviations used: BLASTn: Basic Local Alignment Search Tool for Nucleotide; MEGA: Molecular Evolutionary Genetic Analysis; EMBL: European Molecular Biology Laboratory; psb A- trn H: Photosystem II protein D1- stuctural RNA: His tRNA gene; rbcL: Ribulose 1,5 bi-phosphate carboxylase/oxygenase large subunit gene.

Use of DNA barcodes to identify flowering plants.

PubMed

Kress, W John; Wurdack, Kenneth J; Zimmer, Elizabeth A; Weigt, Lee A; Janzen, Daniel H

2005-06-07

Methods for identifying species by using short orthologous DNA sequences, known as "DNA barcodes," have been proposed and initiated to facilitate biodiversity studies, identify juveniles, associate sexes, and enhance forensic analyses. The cytochrome c oxidase 1 sequence, which has been found to be widely applicable in animal barcoding, is not appropriate for most species of plants because of a much slower rate of cytochrome c oxidase 1 gene evolution in higher plants than in animals. We therefore propose the nuclear internal transcribed spacer region and the plastid trnH-psbA intergenic spacer as potentially usable DNA regions for applying barcoding to flowering plants. The internal transcribed spacer is the most commonly sequenced locus used in plant phylogenetic investigations at the species level and shows high levels of interspecific divergence. The trnH-psbA spacer, although short ( approximately 450-bp), is the most variable plastid region in angiosperms and is easily amplified across a broad range of land plants. Comparison of the total plastid genomes of tobacco and deadly nightshade enhanced with trials on widely divergent angiosperm taxa, including closely related species in seven plant families and a group of species sampled from a local flora encompassing 50 plant families (for a total of 99 species, 80 genera, and 53 families), suggest that the sequences in this pair of loci have the potential to discriminate among the largest number of plant species for barcoding purposes.

Assessing universality of DNA barcoding in geographically isolated selected desert medicinal species of Fabaceae and Poaceae

PubMed Central

Hussain, Fatma; Ahmed, Nisar; Ghorbani, Abdolbaset

2018-01-01

In pursuit of developing fast and accurate species-level molecular identification methods, we tested six DNA barcodes, namely ITS2, matK, rbcLa, ITS2+matK, ITS2+rbcLa, matK+rbcLa and ITS2+matK+rbcLa, for their capacity to identify frequently consumed but geographically isolated medicinal species of Fabaceae and Poaceae indigenous to the desert of Cholistan. Data were analysed by BLASTn sequence similarity, pairwise sequence divergence in TAXONDNA, and phylogenetic (neighbour-joining and maximum-likelihood trees) methods. Comparison of six barcode regions showed that ITS2 has the highest number of variable sites (209/360) for tested Fabaceae and (106/365) Poaceae species, the highest species-level identification (40%) in BLASTn procedure, distinct DNA barcoding gap, 100% correct species identification in BM and BCM functions of TAXONDNA, and clear cladding pattern with high nodal support in phylogenetic trees in both families. ITS2+matK+rbcLa followed ITS2 in its species-level identification capacity. The study was concluded with advocating the DNA barcoding as an effective tool for species identification and ITS2 as the best barcode region in identifying medicinal species of Fabaceae and Poaceae. Current research has practical implementation potential in the fields of pharmaco-vigilance, trade of medicinal plants and biodiversity conservation. PMID:29576968

Conflict-of-interest disclosure at medical journals in Japan: a nationwide survey of the practices of journal secretariats.

PubMed

Kojima, Takako; Green, Joseph; Barron, J Patrick

2015-08-26

Medical journals in Japan generally have appropriate policies regarding disclosure of conflicts of interest (COI). However, COI management depends on the staff members of each journal's editorial secretariat. This study's objectives were to find out (A) whether COI disclosure and the journal's role in it are clearly understood by the journal's secretariat staff, (B) how much experience the editorial secretariat has in actually handling issues related to disclosure and (C) what kind of help or support they need. In January 2014, questionnaires were sent to the editorial secretariats of journal-publishing societies belonging to the Japanese Association of Medical Sciences (JAMS). The response rate was 100%, and the respondents represented 121 journals published by the 118 JAMS member societies (at the time of the survey). Information was collected on the history of COI policies and on how those policies were implemented. At the end of the questionnaire, there was an open-ended call for comments. Compulsory COI disclosure began between 2010 and 2013 for 60.3% of the journals (73/121). Handling of COI issues was not uniform: 17.4% (21/121) of respondents do not pursue cases of dubious disclosure, and 47.9% (58/121) do not require COI disclosures from editorial board members. Very few of the editorial secretariats had clearly-stated consequences for violations of COI-disclosure policy (33/121, 27.3%), and only 28.9% offered COI education (35/121). Respondents' comments indicated that uniform, easily-searchable guidance regarding COI policies and implementation would be welcome. Although commitment is widespread, policy implementation is inconsistent and COI experience is lacking. Clear, easy-to-use guidelines are desired by many societies. The JAMS is to be commended for supporting this country-wide investigation; other countries and regions are encouraged to perform similar investigations to respond to needs regarding COI management. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

The first initiative of DNA barcoding of ornamental plants from Egypt and potential applications in horticulture industry

PubMed Central

Ashfaq, Muhammad; Ali, Hayssam M.; Yessoufou, Kowiyou

2017-01-01

DNA barcoding relies on short and standardized gene regions to identify species. The agricultural and horticultural applications of barcoding such as for marketplace regulation and copyright protection remain poorly explored. This study examines the effectiveness of the standard plant barcode markers (matK and rbcL) for the identification of plant species in private and public nurseries in northern Egypt. These two markers were sequenced from 225 specimens of 161 species and 62 plant families of horticultural importance. The sequence recovery was similar for rbcL (96.4%) and matK (84%), but the number of specimens assigned correctly to the respective genera and species was lower for rbcL (75% and 29%) than matK (85% and 40%). The combination of rbcL and matK brought the number of correct generic and species assignments to 83.4% and 40%, respectively. Individually, the efficiency of both markers varied among different plant families; for example, all palm specimens (Arecaceae) were correctly assigned to species while only one individual of Asteraceae was correctly assigned to species. Further, barcodes reliably assigned ornamental horticultural and medicinal plants correctly to genus while they showed a lower or no success in assigning these plants to species and cultivars. For future, we recommend the combination of a complementary barcode (e.g. ITS or trnH-psbA) with rbcL + matK to increase the performance of taxa identification. By aiding species identification of horticultural crops and ornamental palms, the analysis of the barcode regions will have large impact on horticultural industry. PMID:28199378

The first initiative of DNA barcoding of ornamental plants from Egypt and potential applications in horticulture industry.

PubMed

O Elansary, Hosam; Ashfaq, Muhammad; Ali, Hayssam M; Yessoufou, Kowiyou

2017-01-01

DNA barcoding relies on short and standardized gene regions to identify species. The agricultural and horticultural applications of barcoding such as for marketplace regulation and copyright protection remain poorly explored. This study examines the effectiveness of the standard plant barcode markers (matK and rbcL) for the identification of plant species in private and public nurseries in northern Egypt. These two markers were sequenced from 225 specimens of 161 species and 62 plant families of horticultural importance. The sequence recovery was similar for rbcL (96.4%) and matK (84%), but the number of specimens assigned correctly to the respective genera and species was lower for rbcL (75% and 29%) than matK (85% and 40%). The combination of rbcL and matK brought the number of correct generic and species assignments to 83.4% and 40%, respectively. Individually, the efficiency of both markers varied among different plant families; for example, all palm specimens (Arecaceae) were correctly assigned to species while only one individual of Asteraceae was correctly assigned to species. Further, barcodes reliably assigned ornamental horticultural and medicinal plants correctly to genus while they showed a lower or no success in assigning these plants to species and cultivars. For future, we recommend the combination of a complementary barcode (e.g. ITS or trnH-psbA) with rbcL + matK to increase the performance of taxa identification. By aiding species identification of horticultural crops and ornamental palms, the analysis of the barcode regions will have large impact on horticultural industry.

Confirmation of cisco spawning in Chaumont Bay, Lake Ontario using an egg pumping device

USGS Publications Warehouse

George, Ellen M.; Stott, Wendylee; Young, Brian; Karboski, Curtis T.; Crabtree, Darran L.; Roseman, Edward; Rudstam, Lars G.

2017-01-01

Cisco Coregonus artedi, a historically abundant and commercially important fish in the Great Lakes, have declined drastically in the last century due to the impacts of invasive species, overfishing, and habitat degradation. Chaumont Bay, New York is believed to contain one of the last remaining spawning populations of cisco in Lake Ontario although direct evidence of spawning has remained elusive. We document cisco spawning in Chaumont Bay for the first time in decades through the use of an egg pumping device specifically developed to sample through lake ice. Forty-one eggs were identified as cisco using genetic barcoding of the mitochondrial cytochrome c oxidase I (COI) gene. Cisco eggs were associated with shallow, rocky shoals. Contemporary knowledge of spawning behavior is an important step toward the successful restoration of cisco in Lake Ontario and across the Great Lakes.

Redescription of two subterranean amphipods Niphargus molnari Méhely, 1927 and Niphargus gebhardti Schellenberg, 1934 (Amphipoda, Niphargidae) and their phylogenetic position

PubMed Central

Angyal, Dorottya; Balázs, Gergely; Zakšek, Valerija; Krízsik, Virág; Fišer, Cene

2015-01-01

Abstract A detailed redescription of two endemic, cave-dwelling niphargid species of the Hungarian Mecsek Mts., Niphargus molnari Méhely, 1927 and Niphargus gebhardti Schellenberg, 1934 is given based on newly collected material. Morphology was studied under light microscopy and with scanning electon microscopy. Morphological descriptions are complemented with mitochondrial cytochrome c oxidase subunit I (COI) sequences as barcodes for both species and with notes on their ecology. Using three independent molecular markers we showed that Niphargus gebhardti belongs to the clade distributed between Central and Eastern Europe, whereas phylogenetic relationship of Niphargus molnari to the rest of Niphargus species is not clear. The two species from the Mecsek Mts. are phylogenetically not closely related. Both species need to be treated as vulnerable according to IUCN Red List of Threatened Species. PMID:26175603

Indoors forensic entomology: colonization of human remains in closed environments by specific species of sarcosaprophagous flies.

PubMed

Pohjoismäki, Jaakko L O; Karhunen, Pekka J; Goebeler, Sirkka; Saukko, Pekka; Sääksjärvi, Ilari E

2010-06-15

Fly species that are commonly recovered on human corpses concealed in houses or other dwellings are often dependent on human created environments and might have special features in their biology that allow them to colonize indoor cadavers. In this study we describe nine typical cases involving forensically relevant flies on human remains found indoors in southern Finland. Eggs, larvae and puparia were reared to adult stage and determined to species. Of the five species found the most common were Lucilia sericata Meigen, Calliphora vicina Robineau-Desvoidy and Protophormia terraenovae Robineau-Desvoidy. The flesh fly Sarcophaga caerulescens Zetterstedt is reported for the first time to colonize human cadavers inside houses and a COI gene sequence based DNA barcode is provided for it to help facilitate identification in the future. Fly biology, colonization speed and the significance of indoors forensic entomological evidence are discussed. (c) 2010 Elsevier Ireland Ltd. All rights reserved.

A new member of the genus Isotomurus from the Kuril Islands (Collembola: Isotomidae): returning to the problem of "colour pattern species".

PubMed

Potapov, Mikhail; Porco, David; Deharveng, Louis

2018-03-14

Colour pattern is the most common character to identify species in several large genera of Collembola. Its use often raises problems due to various and poorly investigated extent of chromatic variability among species. Isotomurus festus sp. nov. is here described from Kunashir Isl. (the Kuriles, the Far East of Russia). The species, a member of the 'antennalis' group, is characterized by the lack of trichobothria and slender claws, but is greatly variable in coloration. DNA barcoding (COI) results supports that all the colour forms encountered belong to the same species. While colour pattern has been shown to be the most reliable character for species identification in several Entomobryidae genera, it might not be the case in Isotomurus Börner, 1903, the sole large Isotomidae genus where colour pattern is routinely used for taxonomy.

A review of the North American genus Epimartyria (Lepidoptera, Micropterigidae) with a discussion of the larval plastron

PubMed Central

Davis, Donald R.; Landry, Jean-François

2012-01-01

Abstract The indigenous North American micropterigid genus Epimartyria Walsingham,1898 is revised. Three species are recognized, including Epimartyria auricrinella Walsingham, 1898 which occurs widely over much of the northeastern United States and Canada, a new species, Epimartyria bimaculella Davis & Landry from northwestern United States and Canada, and Epimartyria pardella (Walsingham, 1880) from northern California to northern Oregon. The larva of Epimartyria auricrinella is described in detail, supplemented with illustrations of the external structure of the larval integument. The larval plastron is described and illustrated for Epimartyria, and this is compared with the plastrons of Neomicropteryx Issiki, 1931 and Micropterix Hübner, 1825. COI barcode sequences show that the three species are genetically distinct, congruent with morphological differences. Marked haplotype divergence within some Epimartyria auricrinella populations appears to be unrelated to morphology, geography or phenology. PMID:22573948

Using herbarium-derived DNAs to assemble a large-scale DNA barcode library for the vascular plants of Canada.

PubMed

Kuzmina, Maria L; Braukmann, Thomas W A; Fazekas, Aron J; Graham, Sean W; Dewaard, Stephanie L; Rodrigues, Anuar; Bennett, Bruce A; Dickinson, Timothy A; Saarela, Jeffery M; Catling, Paul M; Newmaster, Steven G; Percy, Diana M; Fenneman, Erin; Lauron-Moreau, Aurélien; Ford, Bruce; Gillespie, Lynn; Subramanyam, Ragupathy; Whitton, Jeannette; Jennings, Linda; Metsger, Deborah; Warne, Connor P; Brown, Allison; Sears, Elizabeth; Dewaard, Jeremy R; Zakharov, Evgeny V; Hebert, Paul D N

2017-12-01

Constructing complete, accurate plant DNA barcode reference libraries can be logistically challenging for large-scale floras. Here we demonstrate the promise and challenges of using herbarium collections for building a DNA barcode reference library for the vascular plant flora of Canada. Our study examined 20,816 specimens representing 5076 of 5190 vascular plant species in Canada (98%). For 98% of the specimens, at least one of the DNA barcode regions was recovered from the plastid loci rbcL and matK and from the nuclear ITS2 region. We used beta regression to quantify the effects of age, type of preservation, and taxonomic affiliation (family) on DNA sequence recovery. Specimen age and method of preservation had significant effects on sequence recovery for all markers, but influenced some families more (e.g., Boraginaceae) than others (e.g., Asteraceae). Our DNA barcode library represents an unparalleled resource for metagenomic and ecological genetic research working on temperate and arctic biomes. An observed decline in sequence recovery with specimen age may be associated with poor primer matches, intragenomic variation (for ITS2), or inhibitory secondary compounds in some taxa.

Using herbarium-derived DNAs to assemble a large-scale DNA barcode library for the vascular plants of Canada1

PubMed Central

Kuzmina, Maria L.; Braukmann, Thomas W. A.; Fazekas, Aron J.; Graham, Sean W.; Dewaard, Stephanie L.; Rodrigues, Anuar; Bennett, Bruce A.; Dickinson, Timothy A.; Saarela, Jeffery M.; Catling, Paul M.; Newmaster, Steven G.; Percy, Diana M.; Fenneman, Erin; Lauron-Moreau, Aurélien; Ford, Bruce; Gillespie, Lynn; Subramanyam, Ragupathy; Whitton, Jeannette; Jennings, Linda; Metsger, Deborah; Warne, Connor P.; Brown, Allison; Sears, Elizabeth; Dewaard, Jeremy R.; Zakharov, Evgeny V.; Hebert, Paul D. N.

2017-01-01

Premise of the study: Constructing complete, accurate plant DNA barcode reference libraries can be logistically challenging for large-scale floras. Here we demonstrate the promise and challenges of using herbarium collections for building a DNA barcode reference library for the vascular plant flora of Canada. Methods: Our study examined 20,816 specimens representing 5076 of 5190 vascular plant species in Canada (98%). For 98% of the specimens, at least one of the DNA barcode regions was recovered from the plastid loci rbcL and matK and from the nuclear ITS2 region. We used beta regression to quantify the effects of age, type of preservation, and taxonomic affiliation (family) on DNA sequence recovery. Results: Specimen age and method of preservation had significant effects on sequence recovery for all markers, but influenced some families more (e.g., Boraginaceae) than others (e.g., Asteraceae). Discussion: Our DNA barcode library represents an unparalleled resource for metagenomic and ecological genetic research working on temperate and arctic biomes. An observed decline in sequence recovery with specimen age may be associated with poor primer matches, intragenomic variation (for ITS2), or inhibitory secondary compounds in some taxa. PMID:29299394

Does a global DNA barcoding gap exist in Annelida?

PubMed

Kvist, Sebastian

2016-05-01

Accurate identification of unknown specimens by means of DNA barcoding is contingent on the presence of a DNA barcoding gap, among other factors, as its absence may result in dubious specimen identifications - false negatives or positives. Whereas the utility of DNA barcoding would be greatly reduced in the absence of a distinct and sufficiently sized barcoding gap, the limits of intraspecific and interspecific distances are seldom thoroughly inspected across comprehensive sampling. The present study aims to illuminate this aspect of barcoding in a comprehensive manner for the animal phylum Annelida. All cytochrome c oxidase subunit I sequences (cox1 gene; the chosen region for zoological DNA barcoding) present in GenBank for Annelida, as well as for "Polychaeta", "Oligochaeta", and Hirudinea separately, were downloaded and curated for length, coverage and potential contaminations. The final datasets consisted of 9782 (Annelida), 5545 ("Polychaeta"), 3639 ("Oligochaeta"), and 598 (Hirudinea) cox1 sequences and these were either (i) used as is in an automated global barcoding gap detection analysis or (ii) further analyzed for genetic distances, separated into bins containing intraspecific and interspecific comparisons and plotted in a graph to visualize any potential global barcoding gap. Over 70 million pairwise genetic comparisons were made and results suggest that although there is a tendency towards separation, no distinct or sufficiently sized global barcoding gap exists in either of the datasets rendering future barcoding efforts at risk of erroneous specimen identifications (but local barcoding gaps may still exist allowing for the identification of specimens at lower taxonomic ranks). This seems to be especially true for earthworm taxa, which account for fully 35% of the total number of interspecific comparisons that show 0% divergence.

The chaperonin-60 universal target is a barcode for bacteria that enables de novo assembly of metagenomic sequence data.

PubMed

Links, Matthew G; Dumonceaux, Tim J; Hemmingsen, Sean M; Hill, Janet E

2012-01-01

Barcoding with molecular sequences is widely used to catalogue eukaryotic biodiversity. Studies investigating the community dynamics of microbes have relied heavily on gene-centric metagenomic profiling using two genes (16S rRNA and cpn60) to identify and track Bacteria. While there have been criteria formalized for barcoding of eukaryotes, these criteria have not been used to evaluate gene targets for other domains of life. Using the framework of the International Barcode of Life we evaluated DNA barcodes for Bacteria. Candidates from the 16S rRNA gene and the protein coding cpn60 gene were evaluated. Within complete bacterial genomes in the public domain representing 983 species from 21 phyla, the largest difference between median pairwise inter- and intra-specific distances ("barcode gap") was found from cpn60. Distribution of sequence diversity along the ∼555 bp cpn60 target region was remarkably uniform. The barcode gap of the cpn60 universal target facilitated the faithful de novo assembly of full-length operational taxonomic units from pyrosequencing data from a synthetic microbial community. Analysis supported the recognition of both 16S rRNA and cpn60 as DNA barcodes for Bacteria. The cpn60 universal target was found to have a much larger barcode gap than 16S rRNA suggesting cpn60 as a preferred barcode for Bacteria. A large barcode gap for cpn60 provided a robust target for species-level characterization of data. The assembly of consensus sequences for barcodes was shown to be a reliable method for the identification and tracking of novel microbes in metagenomic studies.

Author disclosure of conflict of interest in vascular surgery journals.

PubMed

Forbes, Thomas L

2011-09-01

Advances in vascular surgery are increasingly technology-driven, and the relationships between surgeons and the medical device industry can be complex. This study reviewed conflict of interest (COI) disclosure in the vascular surgery journals regarding several selected technology-driven topics, including endovascular stent grafts (EV), carotid artery stenting (CAS), and peripheral arterial interventions (PI), to suggest further directions. Authors' COI disclosures were reviewed from all clinical papers published in 2008 and 2009 in each of six vascular surgery journals, and pertaining to three selected topics (EV, CAS, and PI). Rate of COI disclosure was evaluated as a function of journal, topic, article type (randomized trial, case series, case report, review, or meta-analysis), and authors' region of origin. Secondarily, consistency of authors' disclosure was evaluated by reviewing papers by the same author and of the same topic. Six hundred thirty-five papers were reviewed from the six journals. A COI was declared in 125 (19.7%) of these papers. This rate differed between journals (range, 3.2%-34.1%; P < .0001). Rate of disclosure did not differ between topics (range, 12.8%-21.2%; P = .12), article type (range, 14.7%-30%; P = .28), or region of origin (range, 0%-33.3%; P = .09). There were 116 instances of the same author writing papers describing the same general topic. COI disclosure was consistent in the majority of these instances (72.4%), but inconsistent in 32 cases (27.6%). The most common (P = .006) inconsistencies involved the same type of article in different journals (46.9%), or in the same journal (25%). Rates of disclosure of COI, and inconsistencies in disclosure in the vascular surgery literature are at least partially due to differences in journals' reporting policies, while a smaller proportion of these inconsistencies are due to individual author behavior. Journals should adopt a consistent requirement for a separate COI declaration where all relevant financial arrangements are disclosed. Copyright © 2011 Society for Vascular Surgery. Published by Mosby, Inc. All rights reserved.

A Two-Locus Global DNA Barcode for Land Plants: The Coding rbcL Gene Complements the Non-Coding trnH-psbA Spacer Region

PubMed Central

Kress, W. John; Erickson, David L.

2007-01-01

Background A useful DNA barcode requires sufficient sequence variation to distinguish between species and ease of application across a broad range of taxa. Discovery of a DNA barcode for land plants has been limited by intrinsically lower rates of sequence evolution in plant genomes than that observed in animals. This low rate has complicated the trade-off in finding a locus that is universal and readily sequenced and has sufficiently high sequence divergence at the species-level. Methodology/Principal Findings Here, a global plant DNA barcode system is evaluated by comparing universal application and degree of sequence divergence for nine putative barcode loci, including coding and non-coding regions, singly and in pairs across a phylogenetically diverse set of 48 genera (two species per genus). No single locus could discriminate among species in a pair in more than 79% of genera, whereas discrimination increased to nearly 88% when the non-coding trnH-psbA spacer was paired with one of three coding loci, including rbcL. In silico trials were conducted in which DNA sequences from GenBank were used to further evaluate the discriminatory power of a subset of these loci. These trials supported the earlier observation that trnH-psbA coupled with rbcL can correctly identify and discriminate among related species. Conclusions/Significance A combination of the non-coding trnH-psbA spacer region and a portion of the coding rbcL gene is recommended as a two-locus global land plant barcode that provides the necessary universality and species discrimination. PMID:17551588

ITS1: a DNA barcode better than ITS2 in eukaryotes?

PubMed

Wang, Xin-Cun; Liu, Chang; Huang, Liang; Bengtsson-Palme, Johan; Chen, Haimei; Zhang, Jian-Hui; Cai, Dayong; Li, Jian-Qin

2015-05-01

A DNA barcode is a short piece of DNA sequence used for species determination and discovery. The internal transcribed spacer (ITS/ITS2) region has been proposed as the standard DNA barcode for fungi and seed plants and has been widely used in DNA barcoding analyses for other biological groups, for example algae, protists and animals. The ITS region consists of both ITS1 and ITS2 regions. Here, a large-scale meta-analysis was carried out to compare ITS1 and ITS2 from three aspects: PCR amplification, DNA sequencing and species discrimination, in terms of the presence of DNA barcoding gaps, species discrimination efficiency, sequence length distribution, GC content distribution and primer universality. In total, 85 345 sequence pairs in 10 major groups of eukaryotes, including ascomycetes, basidiomycetes, liverworts, mosses, ferns, gymnosperms, monocotyledons, eudicotyledons, insects and fishes, covering 611 families, 3694 genera, and 19 060 species, were analysed. Using similarity-based methods, we calculated species discrimination efficiencies for ITS1 and ITS2 in all major groups, families and genera. Using Fisher's exact test, we found that ITS1 has significantly higher efficiencies than ITS2 in 17 of the 47 families and 20 of the 49 genera, which are sample-rich. By in silico PCR amplification evaluation, primer universality of the extensively applied ITS1 primers was found superior to that of ITS2 primers. Additionally, shorter length of amplification product and lower GC content was discovered to be two other advantages of ITS1 for sequencing. In summary, ITS1 represents a better DNA barcode than ITS2 for eukaryotic species. © 2014 John Wiley & Sons Ltd.

DNA barcoding unmasks overlooked diversity improving knowledge on the composition and origins of the Churchill algal flora.

PubMed

Saunders, Gary W; McDevit, Daniel C

2013-03-16

Sampling expeditions to Churchill in the Canadian subarctic were completed with the aim of compiling a molecular-assisted survey of the macroalgal flora (seaweeds) for comparison to published accounts for this area, which are based on morphological identifications. Further, because the Churchill region was covered by ice until recently (~10,000 before present), the current algal flora has had to migrate from adjacent waters into that region. We used our DNA barcode data to predict the relative contribution of the North Atlantic and North Pacific floras (Likely Source Region) in repopulating the Churchill region following the most recent glacial retreat. We processed 422 collections representing ~50 morpho-species, which is the approximate number reported for this region, and generated DNA barcode data for 346 of these. In contrast to the morpho-species count, we recovered 57 genetic groups indicating overlooked species (this despite failing to generate barcode data for six of the ~50 morpho-species). However, we additionally uncovered numerous inconsistencies between the species that are currently listed in the Churchill flora (again as a result of overlooked species diversity, but combined with taxonomic confusion) and those identified following our molecular analyses including eight new records and another 17 genetic complexes in need of further study. Based on a comparison of DNA barcode data from the Churchill flora to collections from the contiguous Atlantic and Pacific floras we estimate that minimally 21% (possibly as much as 44%) of the Churchill flora was established by migration from the Pacific region with the balance of species arriving from the Atlantic (predominantly North American populations) following the last glacial retreat. Owing to difficulties associated with the morphological identification of macroalgae, our results indicate that current comprehension of the Canadian Arctic flora is weak. We consider that morphology-based field-identifications are ill-advised in carrying out floristic and ecological surveys for macroalgae and that much of the current data, at least for the Canadian Arctic, should be used with caution. Our efforts to use DNA barcode data to identify the most Likely Source Regions for macroalgal species currently found in Churchill suggests that migration from both the Atlantic and the Pacific have played important roles in establishing the Canadian Arctic flora. This result has significance for understanding both the current and future biodiversity and biogeography of macroalgae in these waters.

Efficient DNA barcode regions for classifying Piper species (Piperaceae).

PubMed

Chaveerach, Arunrat; Tanee, Tawatchai; Sanubol, Arisa; Monkheang, Pansa; Sudmoon, Runglawan

2016-01-01

Piper species are used for spices, in traditional and processed forms of medicines, in cosmetic compounds, in cultural activities and insecticides. Here barcode analysis was performed for identification of plant parts, young plants and modified forms of plants. Thirty-six Piper species were collected and the three barcode regions, matK , rbcL and psbA - trnH spacer, were amplified, sequenced and aligned to determine their genetic distances. For intraspecific genetic distances, the most effective values for the species identification ranged from no difference to very low distance values. However, Piper betle had the highest values at 0.386 for the matK region. This finding may be due to Piper betle being an economic and cultivated species, and thus is supported with growth factors, which may have affected its genetic distance. The interspecific genetic distances that were most effective for identification of different species were from the matK region and ranged from a low of 0.002 in 27 paired species to a high of 0.486. Eight species pairs, Piper kraense and Piper dominantinervium , Piper magnibaccum and Piper kraense , Piper phuwuaense and Piper dominantinervium , Piper phuwuaense and Piper kraense , Piper pilobracteatum and Piper dominantinervium , Piper pilobracteatum and Piper kraense , Piper pilobracteatum and Piper phuwuaense and Piper sylvestre and Piper polysyphonum , that presented a genetic distance of 0.000 and were identified by independently using each of the other two regions. Concisely, these three barcode regions are powerful for further efficient identification of the 36 Piper species.

Efficient DNA barcode regions for classifying Piper species (Piperaceae)

PubMed Central

Chaveerach, Arunrat; Tanee, Tawatchai; Sanubol, Arisa; Monkheang, Pansa; Sudmoon, Runglawan

2016-01-01

Abstract Piper species are used for spices, in traditional and processed forms of medicines, in cosmetic compounds, in cultural activities and insecticides. Here barcode analysis was performed for identification of plant parts, young plants and modified forms of plants. Thirty-six Piper species were collected and the three barcode regions, matK, rbcL and psbA-trnH spacer, were amplified, sequenced and aligned to determine their genetic distances. For intraspecific genetic distances, the most effective values for the species identification ranged from no difference to very low distance values. However, Piper betle had the highest values at 0.386 for the matK region. This finding may be due to Piper betle being an economic and cultivated species, and thus is supported with growth factors, which may have affected its genetic distance. The interspecific genetic distances that were most effective for identification of different species were from the matK region and ranged from a low of 0.002 in 27 paired species to a high of 0.486. Eight species pairs, Piper kraense and Piper dominantinervium, Piper magnibaccum and Piper kraense, Piper phuwuaense and Piper dominantinervium, Piper phuwuaense and Piper kraense, Piper pilobracteatum and Piper dominantinervium, Piper pilobracteatum and Piper kraense, Piper pilobracteatum and Piper phuwuaense and Piper sylvestre and Piper polysyphonum, that presented a genetic distance of 0.000 and were identified by independently using each of the other two regions. Concisely, these three barcode regions are powerful for further efficient identification of the 36 Piper species. PMID:27829794

A distinct group of north European Aedes vexans as determined by mitochondrial and nuclear markers.

PubMed

Lilja, T; Troell, K; Kirik, H; Lindström, A

2018-01-16

The floodwater mosquito Aedes (Aedimorphus) vexans (Meigen, 1830) (Diptera: Culicidae) is common in several areas of Sweden and is predicted to become more abundant in the wake of expected changes in precipitation and temperature caused by climate change. As well as being a nuisance, Ae. vexans can act as a vector of over 30 viruses. In the event of an outbreak of disease caused by a vector-borne virus, knowledge of the distribution, population structure and intermixing of populations from different locations will help direct resources to target locations to prevent spread of the pathogen. The present study analysed individual Ae. vexans from eight locations throughout Sweden. Based on the mitochondrial cytochrome oxidase I (COI) marker, a subset of the analysed mosquitoes cluster apart from the other samples. Similarly, two nuclear loci were sequenced and the same phylogenetic structure observed. These results indicate that this group represents a reproductively isolated population among Ae. vexans. Comparisons with COI sequences held in the Barcode of Life Database (BoLD) for Ae. vexans from around the world show that specimens collected in Belgium and Estonia group together with the Swedish group, suggesting that this genotype is present throughout northern Europe. These results suggest there is a cryptic taxonomic unit related to Ae. vexans in northern Europe. © 2018 The Authors. Medical and Veterinary Entomology published by John Wiley & Sons Ltd on behalf of Royal Entomological Society.

Phylogenetics of Australasian gall flies (Diptera: Fergusoninidae): Evolutionary patterns of host-shifting and gall morphology.

PubMed

Scheffer, S J; Davies, K A; Taylor, G S; Thornhill, A H; Lewis, M L; Winkler, I S; Yeates, D K; Purcell, M F; Makinson, J; Giblin-Davis, R M

2017-10-01

This study investigated host-specificity and phylogenetic relationships in Australian galling flies, Fergusonina Malloch (Diptera: Fergusoninidae), in order to assess diversity and explore the evolutionary history of host plant affiliation and gall morphology. A DNA barcoding approach using COI data from 203 Fergusonina specimens from 5gall types on 56 host plant species indicated 85 presumptive fly species. These exhibited a high degree of host specificity; of the 40 species with multiple representatives, each fed only on a single host genus, 29 (72.5%) were strictly monophagous, and 11 (27.5%) were reared from multiple closely related hosts. COI variation within species was not correlated with either sample size or geographic distance. However variation was greater within oligophagous species, consistent with expectations of the initial stages of host-associated divergence during speciation. Phylogenetic analysis using both nuclear and mitochondrial genes revealed host genus-restricted clades but also clear evidence of multiple colonizations of both host plant genus and host species. With the exception of unilocular peagalls, evolution of gall type was somewhat constrained, but to a lesser degree than host plant association. Unilocular peagalls arose more often than any other gall type, were primarily located at the tips of the phylogeny, and did not form clades comprising more than a few species. For ecological reasons, species of this gall type are predicted to harbor substantially less genetic variation than others, possibly reducing evolutionary flexibility resulting in reduced diversification in unilocular gallers. Published by Elsevier Inc.

Evaluating the feasibility of using candidate DNA barcodes in discriminating species of the large Asteraceae family.

PubMed

Gao, Ting; Yao, Hui; Song, Jingyuan; Zhu, Yingjie; Liu, Chang; Chen, Shilin

2010-10-26

Five DNA regions, namely, rbcL, matK, ITS, ITS2, and psbA-trnH, have been recommended as primary DNA barcodes for plants. Studies evaluating these regions for species identification in the large plant taxon, which includes a large number of closely related species, have rarely been reported. The feasibility of using the five proposed DNA regions was tested for discriminating plant species within Asteraceae, the largest family of flowering plants. Among these markers, ITS2 was the most useful in terms of universality, sequence variation, and identification capability in the Asteraceae family. The species discriminating power of ITS2 was also explored in a large pool of 3,490 Asteraceae sequences that represent 2,315 species belonging to 494 different genera. The result shows that ITS2 correctly identified 76.4% and 97.4% of plant samples at the species and genus levels, respectively. In addition, ITS2 displayed a variable ability to discriminate related species within different genera. ITS2 is the best DNA barcode for the Asteraceae family. This approach significantly broadens the application of DNA barcoding to resolve classification problems in the family Asteraceae at the genera and species levels.

Disentangling Vector-Borne Transmission Networks: A Universal DNA Barcoding Method to Identify Vertebrate Hosts from Arthropod Bloodmeals

PubMed Central

Alcaide, Miguel; Rico, Ciro; Ruiz, Santiago; Soriguer, Ramón; Muñoz, Joaquín; Figuerola, Jordi

2009-01-01

Emerging infectious diseases represent a challenge for global economies and public health. About one fourth of the last pandemics have been originated by the spread of vector-borne pathogens. In this sense, the advent of modern molecular techniques has enhanced our capabilities to understand vector-host interactions and disease ecology. However, host identification protocols have poorly profited of international DNA barcoding initiatives and/or have focused exclusively on a limited array of vector species. Therefore, ascertaining the potential afforded by DNA barcoding tools in other vector-host systems of human and veterinary importance would represent a major advance in tracking pathogen life cycles and hosts. Here, we show the applicability of a novel and efficient molecular method for the identification of the vertebrate host's DNA contained in the midgut of blood-feeding arthropods. To this end, we designed a eukaryote-universal forward primer and a vertebrate-specific reverse primer to selectively amplify 758 base pairs (bp) of the vertebrate mitochondrial Cytochrome c Oxidase Subunit I (COI) gene. Our method was validated using both extensive sequence surveys from the public domain and Polymerase Chain Reaction (PCR) experiments carried out over specimens from different Classes of vertebrates (Mammalia, Aves, Reptilia and Amphibia) and invertebrate ectoparasites (Arachnida and Insecta). The analysis of mosquito, culicoid, phlebotomie, sucking bugs, and tick bloodmeals revealed up to 40 vertebrate hosts, including 23 avian, 16 mammalian and one reptilian species. Importantly, the inspection and analysis of direct sequencing electropherograms also assisted the resolving of mixed bloodmeals. We therefore provide a universal and high-throughput diagnostic tool for the study of the ecology of haematophagous invertebrates in relation to their vertebrate hosts. Such information is crucial to support the efficient management of initiatives aimed at reducing epidemiologic risks of arthropod vector-borne pathogens, a priority for public health. PMID:19768113

Use of DNA barcodes to identify flowering plants

PubMed Central

Kress, W. John; Wurdack, Kenneth J.; Zimmer, Elizabeth A.; Weigt, Lee A.; Janzen, Daniel H.

2005-01-01

Methods for identifying species by using short orthologous DNA sequences, known as “DNA barcodes,” have been proposed and initiated to facilitate biodiversity studies, identify juveniles, associate sexes, and enhance forensic analyses. The cytochrome c oxidase 1 sequence, which has been found to be widely applicable in animal barcoding, is not appropriate for most species of plants because of a much slower rate of cytochrome c oxidase 1 gene evolution in higher plants than in animals. We therefore propose the nuclear internal transcribed spacer region and the plastid trnH-psbA intergenic spacer as potentially usable DNA regions for applying barcoding to flowering plants. The internal transcribed spacer is the most commonly sequenced locus used in plant phylogenetic investigations at the species level and shows high levels of interspecific divergence. The trnH-psbA spacer, although short (≈450-bp), is the most variable plastid region in angiosperms and is easily amplified across a broad range of land plants. Comparison of the total plastid genomes of tobacco and deadly nightshade enhanced with trials on widely divergent angiosperm taxa, including closely related species in seven plant families and a group of species sampled from a local flora encompassing 50 plant families (for a total of 99 species, 80 genera, and 53 families), suggest that the sequences in this pair of loci have the potential to discriminate among the largest number of plant species for barcoding purposes. PMID:15928076

Applying DNA Barcodes to Identify Closely Related Species of Ferns: A Case Study of the Chinese Adiantum (Pteridaceae)

PubMed Central

Wen, Jun; Ebihara, Atsushi; Li, De-Zhu

2016-01-01

DNA barcoding is a fast-developing technique to identify species by using short and standard DNA sequences. Universal selection of DNA barcodes in ferns remains unresolved. In this study, five plastid regions (rbcL, matK, trnH-psbA, trnL-F and rps4-trnS) and eight nuclear regions (ITS, pgiC, gapC, LEAFY, ITS2, IBR3_2, DET1, and SQD1_1) were screened and evaluated in the fern genus Adiantum from China and neighboring areas. Due to low primer universality (matK) and/or the existence of multiple copies (ITS), the commonly used barcodes matK and ITS were not appropriate for Adiantum. The PCR amplification rate was extremely low in all nuclear genes except for IBR3_2. rbcL had the highest PCR amplification rate (94.33%) and sequencing success rate (90.78%), while trnH-psbA had the highest species identification rate (75%). With the consideration of discriminatory power, cost-efficiency and effort, the two-barcode combination of rbcL+ trnH-psbA seems to be the best choice for barcoding Adiantum, and perhaps basal polypod ferns in general. The nuclear IBR3_2 showed 100% PCR amplification success rate in Adiantum, however, it seemed that only diploid species could acquire clean sequences without cloning. With cloning, IBR3_2 can successfully distinguish cryptic species and hybrid species from their related species. Because hybridization and allopolyploidy are common in ferns, we argue for including a selected group of nuclear loci as barcodes, especially via the next-generation sequencing, as it is much more efficient to obtain single-copy nuclear loci without the cloning procedure. PMID:27603700

Non-Enzymatic Detection of Bacterial Genomic DNA Using the Bio-Barcode Assay

PubMed Central

Hill, Haley D.; Vega, Rafael A.; Mirkin, Chad A.

2011-01-01

The detection of bacterial genomic DNA through a non-enzymatic nanomaterials based amplification method, the bio-barcode assay, is reported. The assay utilizes oligonucleotide functionalized magnetic microparticles to capture the target of interest from the sample. A critical step in the new assay involves the use of blocking oligonucleotides during heat denaturation of the double stranded DNA. These blockers bind to specific regions of the target DNA upon cooling, and prevent the duplex DNA from re-hybridizing, which allows the particle probes to bind. Following target isolation using the magnetic particles, oligonucleotide functionalized gold nanoparticles act as target recognition agents. The oligonucleotides on the nanoparticle (barcodes) act as amplification surrogates. The barcodes are then detected using the Scanometric method. The limit of detection for this assay was determined to be 2.5 femtomolar, and this is the first demonstration of a barcode type assay for the detection of double stranded, genomic DNA. PMID:17927207

DNA barcodes reveal microevolutionary signals in fire response trait in two legume genera

PubMed Central

Bello, Abubakar; Daru, Barnabas H.; Stirton, Charles H.; Chimphango, Samson B. M.; van der Bank, Michelle; Maurin, Olivier; Muasya, A. Muthama

2015-01-01

Large-scale DNA barcoding provides a new technique for species identification and evaluation of relationships across various levels (populations and species) and may reveal fundamental processes in recently diverged species. Here, we analysed DNA sequence variation in the recently diverged legumes from the Psoraleeae (Fabaceae) occurring in the Cape Floristic Region (CFR) of southern Africa to test the utility of DNA barcodes in species identification and discrimination. We further explored the phylogenetic signal on fire response trait (reseeding and resprouting) at species and generic levels. We showed that Psoraleoid legumes of the CFR exhibit a barcoding gap yielding the combination of matK and rbcLa (matK + rbcLa) data set as a better barcode than single regions. We found a high score (100 %) of correct identification of individuals to their respective genera but a very low score (<50 %) in identifying them to species. We found a considerable match (54 %) between genetic species and morphologically delimited species. We also found that different lineages showed a weak but significant phylogenetic conservatism in their response to fire as reseeders or resprouters, with more clustering of resprouters than would be expected by chance. These novel microevolutionary patterns might be acting continuously over time to produce multi-scale regularities of biodiversity. This study provides the first insight into the DNA barcoding campaign of land plants in species identification and detection of the phylogenetic signal in recently diverged lineages of the CFR. PMID:26507570

Quantitative Tracking of Salmonella Enteritidis Transmission Routes Using Barcode-Tagged Isogenic Strains in Chickens: Proof-of-Concept Study

PubMed Central

Yang, Yichao; Ricke, Steven C.; Tellez, Guillermo; Kwon, Young Min

2017-01-01

Salmonella is an important foodborne bacterial pathogen, however, a fundamental understanding on Salmonella transmission routes within a poultry flock remains unclear. In this study, a series of barcode-tagged strains were constructed by inserting six random nucleotides into a functionally neutral region on the chromosome of S. Enteritidis as a tool for quantitative tracking of Salmonella transmission in chickens. Six distinct barcode-tagged strains were used for infection or contamination at either low dose (103 CFUs; three strains) or high dose (105 CFUs; three strains) in three independent experiments (Experiment 1 oral gavage; Experiment 2 contaminated feed; Experiment 3 contaminated water). For all chick experiments, cecal and foot-wash samples were collected from a subset of the chickens at days 7 or/and 14, from which genomic DNA was extracted and used to amplify the barcode regions. After the resulting PCR amplicons were pooled and analyzed by MiSeq sequencing, a total of approximately 1.5 million reads containing the barcode sequences were analyzed to determine the relative frequency of every barcode-tagged strain in each sample. In Experiment 1, the high dose of oral infection was correlated with greater dominance of the strains in the ceca of the respective seeder chickens and also in the contact chickens yet at lesser degrees. When chicks were exposed to contaminated feed (Experiment 2) or water (Experiment 3), there were no clear patterns of the barcode-tagged strains in relation to the dosage, except that the strains introduced at low dose required a longer time to colonize the ceca with contaminated feed. Most foot-wash samples contained only one to three strains for the majority of the samples, suggesting potential existence of an unknown mechanism(s) for strain exclusion. These results demonstrated the proof of concept of using barcode tagged to investigate transmission dynamics of Salmonella in chickens in a quantitative manner. PMID:28261587

First comparative insight into the architecture of COI mitochondrial minicircle molecules of dicyemids reveals marked inter-species variation.

PubMed

Catalano, Sarah R; Whittington, Ian D; Donnellan, Stephen C; Bertozzi, Terry; Gillanders, Bronwyn M

2015-07-01

Dicyemids, poorly known parasites of benthic cephalopods, are one of the few phyla in which mitochondrial (mt) genome architecture departs from the typical ~16 kb circular metazoan genome. In addition to a putative circular genome, a series of mt minicircles that each comprises the mt encoded units (I-III) of the cytochrome c oxidase complex have been reported. Whether the structure of the mt minicircles is a consistent feature among dicyemid species is unknown. Here we analyse the complete cytochrome c oxidase subunit I (COI) minicircle molecule, containing the COI gene and an associated non-coding region (NCR), for ten dicyemid species, allowing for first time comparisons between species of minicircle architecture, NCR function and inferences of minicircle replication. Divergence in COI nucleotide sequences between dicyemid species was high (average net divergence = 31.6%) while within species diversity was lower (average net divergence = 0.2%). The NCR and putative 5' section of the COI gene were highly divergent between dicyemid species (average net nucleotide divergence of putative 5' COI section = 61.1%). No tRNA genes were found in the NCR, although palindrome sequences with the potential to form stem-loop structures were identified in some species, which may play a role in transcription or other biological processes.

The D1-D2 region of the large subunit ribosomal DNA as barcode for ciliates.

PubMed

Stoeck, T; Przybos, E; Dunthorn, M

2014-05-01

Ciliates are a major evolutionary lineage within the alveolates, which are distributed in nearly all habitats on our planet and are an essential component for ecosystem function, processes and stability. Accurate identification of these unicellular eukaryotes through, for example, microscopy or mating type reactions is reserved to few specialists. To satisfy the demand for a DNA barcode for ciliates, which meets the standard criteria for DNA barcodes defined by the Consortium for the Barcode of Life (CBOL), we here evaluated the D1-D2 region of the ribosomal DNA large subunit (LSU-rDNA). Primer universality for the phylum Ciliophora was tested in silico with available database sequences as well as in the laboratory with 73 ciliate species, which represented nine of 12 ciliate classes. Primers tested in this study were successful for all tested classes. To test the ability of the D1-D2 region to resolve conspecific and congeneric sequence divergence, 63 Paramecium strains were sampled from 24 mating species. The average conspecific D1-D2 variation was 0.18%, whereas congeneric sequence divergence averaged 4.83%. In pairwise genetic distance analyses, we identified a D1-D2 sequence divergence of <0.6% as an ideal threshold to discriminate Paramecium species. Using this definition, only 3.8% of all conspecific and 3.9% of all congeneric sequence comparisons had the potential of false assignments. Neighbour-joining analyses inferred monophyly for all taxa but for two Paramecium octaurelia strains. Here, we present a protocol for easy DNA amplification of single cells and voucher deposition. In conclusion, the presented data pinpoint the D1-D2 region as an excellent candidate for an official CBOL barcode for ciliated protists. © 2013 John Wiley & Sons Ltd.

DNA barcoding and microsatellites help species delimitation and hybrid identification in endangered galaxiid fishes.

PubMed

Vanhaecke, Delphine; Garcia de Leaniz, Carlos; Gajardo, Gonzalo; Young, Kyle; Sanzana, Jose; Orellana, Gabriel; Fowler, Daniel; Howes, Paul; Monzon-Arguello, Catalina; Consuegra, Sofia

2012-01-01

The conservation of data deficient species is often hampered by inaccurate species delimitation. The galaxiid fishes Aplochiton zebra and Aplochiton taeniatus are endemic to Patagonia (and for A. zebra the Falkland Islands), where they are threatened by invasive salmonids. Conservation of Aplochiton is complicated because species identification is hampered by the presence of resident as well as migratory ecotypes that may confound morphological discrimination. We used DNA barcoding (COI, cytochrome b) and a new developed set of microsatellite markers to investigate the relationships between A. zebra and A. taeniatus and to assess their distributions and relative abundances in Chilean Patagonia and the Falkland Islands. Results from both DNA markers were 100% congruent and revealed that phenotypic misidentification was widespread, size-dependent, and highly asymmetric. While all the genetically classified A. zebra were correctly identified as such, 74% of A. taeniatus were incorrectly identified as A. zebra, the former species being more widespread than previously thought. Our results reveal, for the first time, the presence in sympatry of both species, not only in Chilean Patagonia, but also in the Falkland Islands, where A. taeniatus had not been previously described. We also found evidence of asymmetric hybridisation between female A. taeniatus and male A. zebra in areas where invasive salmonids have become widespread. Given the potential consequences that species misidentification and hybridisation can have for the conservation of these endangered species, we advocate the use of molecular markers in order to reduce epistemic uncertainty.

Multilocus inference of species trees and DNA barcoding.

PubMed

Mallo, Diego; Posada, David

2016-09-05

The unprecedented amount of data resulting from next-generation sequencing has opened a new era in phylogenetic estimation. Although large datasets should, in theory, increase phylogenetic resolution, massive, multilocus datasets have uncovered a great deal of phylogenetic incongruence among different genomic regions, due both to stochastic error and to the action of different evolutionary process such as incomplete lineage sorting, gene duplication and loss and horizontal gene transfer. This incongruence violates one of the fundamental assumptions of the DNA barcoding approach, which assumes that gene history and species history are identical. In this review, we explain some of the most important challenges we will have to face to reconstruct the history of species, and the advantages and disadvantages of different strategies for the phylogenetic analysis of multilocus data. In particular, we describe the evolutionary events that can generate species tree-gene tree discordance, compare the most popular methods for species tree reconstruction, highlight the challenges we need to face when using them and discuss their potential utility in barcoding. Current barcoding methods sacrifice a great amount of statistical power by only considering one locus, and a transition to multilocus barcodes would not only improve current barcoding methods, but also facilitate an eventual transition to species-tree-based barcoding strategies, which could better accommodate scenarios where the barcode gap is too small or inexistent.This article is part of the themed issue 'From DNA barcodes to biomes'. © 2016 The Authors.

DNA Barcode Goes Two-Dimensions: DNA QR Code Web Server

PubMed Central

Li, Huan; Xing, Hang; Liang, Dong; Jiang, Kun; Pang, Xiaohui; Song, Jingyuan; Chen, Shilin

2012-01-01

The DNA barcoding technology uses a standard region of DNA sequence for species identification and discovery. At present, “DNA barcode” actually refers to DNA sequences, which are not amenable to information storage, recognition, and retrieval. Our aim is to identify the best symbology that can represent DNA barcode sequences in practical applications. A comprehensive set of sequences for five DNA barcode markers ITS2, rbcL, matK, psbA-trnH, and CO1 was used as the test data. Fifty-three different types of one-dimensional and ten two-dimensional barcode symbologies were compared based on different criteria, such as coding capacity, compression efficiency, and error detection ability. The quick response (QR) code was found to have the largest coding capacity and relatively high compression ratio. To facilitate the further usage of QR code-based DNA barcodes, a web server was developed and is accessible at http://qrfordna.dnsalias.org. The web server allows users to retrieve the QR code for a species of interests, convert a DNA sequence to and from a QR code, and perform species identification based on local and global sequence similarities. In summary, the first comprehensive evaluation of various barcode symbologies has been carried out. The QR code has been found to be the most appropriate symbology for DNA barcode sequences. A web server has also been constructed to allow biologists to utilize QR codes in practical DNA barcoding applications. PMID:22574113

DNA barcodes of the native ray-finned fishes in Taiwan.

PubMed

Chang, Chia-Hao; Shao, Kwang-Tsao; Lin, Han-Yang; Chiu, Yung-Chieh; Lee, Mao-Ying; Liu, Shih-Hui; Lin, Pai-Lei

2017-07-01

Species identification based on the DNA sequence of a fragment of the cytochrome c oxidase subunit I gene in the mitochondrial genome, DNA barcoding, is widely applied to assist in sustainable exploitation of fish resources and the protection of fish biodiversity. The aim of this study was to establish a reliable barcoding reference database of the native ray-finned fishes in Taiwan. A total of 2993 individuals, belonging to 1245 species within 637 genera, 184 families and 29 orders of ray-finned fishes and representing approximately 40% of the recorded ray-finned fishes in Taiwan, were PCR amplified at the barcode region and bidirectionally sequenced. The mean length of the 2993 barcodes is 549 bp. Mean congeneric K2P distance (15.24%) is approximately 10-fold higher than the mean conspecific one (1.51%), but approximately 1.4-fold less than the mean genetic distance between families (20.80%). The Barcode Index Number (BIN) discordance report shows that 2993 specimens represent 1275 BINs and, among them, 86 BINs are singletons, 570 BINs are taxonomically concordant, and the other 619 BINs are taxonomically discordant. Barcode gap analysis also revealed that more than 90% of the collected fishes in this study can be discriminated by DNA barcoding. Overall, the barcoding reference database established by this study reveals the need for taxonomic revisions and voucher specimen rechecks, in addition to assisting in the management of Taiwan's fish resources and diversity. © 2016 John Wiley & Sons Ltd.

DNA Barcoding of Freshwater Fishes of Indo-Myanmar Biodiversity Hotspot.

PubMed

Barman, Anindya Sundar; Singh, Mamta; Singh, Soibam Khogen; Saha, Himadri; Singh, Yumlembam Jackie; Laishram, Martina; Pandey, Pramod Kumar

2018-06-05

To develop an effective conservation and management strategy, it is required to assess the biodiversity status of an ecosystem, especially when we deal with Indo-Myanmar biodiversity hotspot. Importance of this reaches to an entirely different level as the hotspot represents the area of high endemism which is under continuous threat. Therefore, the need of the present study was conceptualized, dealing with molecular assessment of the fish fauna of Indo-Myanmar region, which covers the Indian states namely, Manipur, Meghalaya, Mizoram, and Nagaland. A total of 363 specimens, representing 109 species were collected and barcoded from the different rivers and their tributaries of the region. The analyses performed in the present study, i.e. Kimura 2-Parameter genetic divergence, Neighbor-Joining, Automated Barcode Gap Discovery and Bayesian Poisson Tree Processes suggest that DNA barcoding is an efficient and reliable tool for species identification. Most of the species were clearly delineated. However, presence of intra-specific and inter-specific genetic distance overlap in few species, revealed the existence of putative cryptic species. A reliable DNA barcode reference library, established in our study provides an adequate knowledge base to the groups of non-taxonomists, researchers, biodiversity managers and policy makers in sketching effective conservation measures for this ecosystem.

Closely-related taxa influence woody species discrimination via DNA barcoding: evidence from global forest dynamics plots.

PubMed

Pei, Nancai; Erickson, David L; Chen, Bufeng; Ge, Xuejun; Mi, Xiangcheng; Swenson, Nathan G; Zhang, Jin-Long; Jones, Frank A; Huang, Chun-Lin; Ye, Wanhui; Hao, Zhanqing; Hsieh, Chang-Fu; Lum, Shawn; Bourg, Norman A; Parker, John D; Zimmerman, Jess K; McShea, William J; Lopez, Ida C; Sun, I-Fang; Davies, Stuart J; Ma, Keping; Kress, W John

2015-10-12

To determine how well DNA barcodes from the chloroplast region perform in forest dynamics plots (FDPs) from global CTFS-ForestGEO network, we analyzed DNA barcoding sequences of 1277 plant species from a wide phylogenetic range (3 FDPs in tropics, 5 in subtropics and 5 in temperate zone) and compared the rates of species discrimination (RSD). We quantified RSD by two DNA barcode combinations (rbcL + matK and rbcL + matK + trnH-psbA) using a monophyly-based method (GARLI). We defined two indexes of closely-related taxa (Gm/Gt and S/G ratios) and correlated these ratios with RSD. The combination of rbcL + matK averagely discriminated 88.65%, 83.84% and 72.51% at the local, regional and global scales, respectively. An additional locus trnH-psbA increased RSD by 2.87%, 1.49% and 3.58% correspondingly. RSD varied along a latitudinal gradient and were negatively correlated with ratios of closely-related taxa. Successes of species discrimination generally depend on scales in global FDPs. We suggested that the combination of rbcL + matK + trnH-psbA is currently applicable for DNA barcoding-based phylogenetic studies on forest communities.

Barcode Identifiers as a Practical Tool for Reliable Species Assignment of Medically Important Black Yeast Species

PubMed Central

Heinrichs, Guido; de Hoog, G. Sybren

2012-01-01

Herpotrichiellaceous black yeasts and relatives comprise severe pathogens flanked by nonpathogenic environmental siblings. Reliable identification by conventional methods is notoriously difficult. Molecular identification is hampered by the sequence variability in the internal transcribed spacer (ITS) domain caused by difficult-to-sequence homopolymeric regions and by poor taxonomic attribution of sequences deposited in GenBank. Here, we present a potential solution using short barcode identifiers (27 to 50 bp) based on ITS2 ribosomal DNA (rDNA), which allows unambiguous definition of species-specific fragments. Starting from proven sequences of ex-type and authentic strains, we were able to describe 103 identifiers. Multiple BLAST searches of these proposed barcode identifiers in GenBank revealed uniqueness for 100 taxonomic entities, whereas the three remaining identifiers each matched with two entities, but the species of these identifiers could easily be discriminated by differences in the remaining ITS regions. Using the proposed barcode identifiers, a 4.1-fold increase of 100% matches in GenBank was achieved in comparison to the classical approach using the complete ITS sequences. The proposed barcode identifiers will be made accessible for the diagnostic laboratory in a permanently updated online database, thereby providing a highly practical, reliable, and cost-effective tool for identification of clinically important black yeasts and relatives. PMID:22785187

A New Species of Tiger Pleco Panaqolus (Siluriformes: Loricariidae) from the Xingu Basin, Brazil

PubMed Central

Melo de Sousa, Leandro

2016-01-01

Panaqolus tankei is described from the Xingu River, Brazil. The new species is diagnosed from P. albomaculatus, P. dentex, P. nix, P. nocturnus, and P. koko by its color pattern consisting of dark and light diagonal bars on the body and bands on the fins (vs. body and fins without bars or bands); from P. albivermis, P. maccus, and P. purusiensis by the width of the dark bars being more or less the same of the light bars (vs. dark bars at least two or three times wider than light bars) and from P. changae by the absence of vermiculation on the head (vs. vermiculation present on head). The new species differs from P. gnomus by the orientation of the bars from posterodorsal to anteroventral direction (vs. anterodorsal to posteroventral direction), and from P. claustellifer by the orientation of the bands in the dorsal fin that are not parallel to the margin (vs. parallel to the margin). The barcoding region (COI) was sequenced for the new species, sequences were deposited in GenBank and were compared with congeners from other drainages. With regard to the current construction of a hydroelectric power plant (a so-called mega dam) in the Xingu River, herewith we increase knowledge of the river Xingu’s ichthyofauna and, thus improve the assessment of the impacts of that construction on the river. PMID:27828981

Conquered from the deep sea? A new deep-sea isopod species from the Antarctic shelf shows pattern of recent colonization.

PubMed

Riehl, Torben; Kaiser, Stefanie

2012-01-01

The Amundsen Sea, Antarctica, is amongst the most rapidly changing environments of the world. Its benthic inhabitants are barely known and the BIOPEARL 2 project was one of the first to biologically explore this region. Collected during this expedition, Macrostylis roaldi sp. nov. is described as the first isopod discovered on the Amundsen-Sea shelf. Amongst many characteristic features, the most obvious characters unique for M. roaldi are the rather short pleotelson and short operculum as well as the trapezoid shape of the pleotelson in adult males. We used DNA barcodes (COI) and additional mitochondrial markers (12S, 16S) to reciprocally illuminate morphological results and nucleotide variability. In contrast to many other deep-sea isopods, this species is common and shows a wide distribution. Its range spreads from Pine Island Bay at inner shelf right to the shelf break and across 1,000 m bathymetrically. Its gene pool is homogenized across space and depth. This is indicative for a genetic bottleneck or a recent colonization history. Our results suggest further that migratory or dispersal capabilities of some species of brooding macrobenthos have been underestimated. This might be relevant for the species' potential to cope with effects of climate change. To determine where this species could have survived the last glacial period, alternative refuge possibilities are discussed.

A New Species of Tiger Pleco Panaqolus (Siluriformes: Loricariidae) from the Xingu Basin, Brazil.

PubMed

Cramer, Christian Andreas; Melo de Sousa, Leandro

2016-01-01

Panaqolus tankei is described from the Xingu River, Brazil. The new species is diagnosed from P. albomaculatus, P. dentex, P. nix, P. nocturnus, and P. koko by its color pattern consisting of dark and light diagonal bars on the body and bands on the fins (vs. body and fins without bars or bands); from P. albivermis, P. maccus, and P. purusiensis by the width of the dark bars being more or less the same of the light bars (vs. dark bars at least two or three times wider than light bars) and from P. changae by the absence of vermiculation on the head (vs. vermiculation present on head). The new species differs from P. gnomus by the orientation of the bars from posterodorsal to anteroventral direction (vs. anterodorsal to posteroventral direction), and from P. claustellifer by the orientation of the bands in the dorsal fin that are not parallel to the margin (vs. parallel to the margin). The barcoding region (COI) was sequenced for the new species, sequences were deposited in GenBank and were compared with congeners from other drainages. With regard to the current construction of a hydroelectric power plant (a so-called mega dam) in the Xingu River, herewith we increase knowledge of the river Xingu's ichthyofauna and, thus improve the assessment of the impacts of that construction on the river.

Taxonomic review of Argentine mackerel Scomber japonicus (Houttuyn, 1782) by phylogenetic analysis

PubMed Central

Trucco, María Inés; Buratti, Claudio César

2017-01-01

Taxonomically, Argentine mackerels were first considered as Scomber japonicus marplatensis and later as Scomber japonicus Houttuyn 1782, although, in the last years, different studies have suggested that South Atlantic mackerel species belongs to Scomber colias Gmelin 1789. These latter results, incorporated in the main fish databases (FishBase and Catalog of Fishes), promoted a phylogenetic study using cytochrome c oxidase I (COI) gene sequences taken from the Barcode of Life (FISH-BOL) database. Thus, 76 sequences of S. japonicus, S. colias, S. australasicus and S. scombrus from different regions were used; including 3 from Sarda sarda as outgroup. Among S. japonicus selected sequences are those corresponding to the Argentine mackerels collected in 2007. Phylogenetic trees were obtained by neighbor joining and maximum likelihood methods and a network of haplotypes was reconstructed to analyze the relationship between species. The results showed the clear differentiation of S. australasicus, S. scombrus and S. japonicus from the Pacific while S. japonicus from Argentina was included in the S. colias group, with genetic differences corresponding to conspecific populations (0.1%). Four of the five Argentine specimens shared the same haplotype with S. colias, and none were shared with S. japonicus from the Pacific. These results suggest that the current specific name of Argentine mackerel S. japonicus should be changed to S. colias, in agreement with several genetic studies carried out with species of the genus Scomber. PMID:29071283

DNA sequence database as a tool to identify decapod crustaceans on the São Paulo coastline.

PubMed

Mantelatto, Fernando L; Terossi, Mariana; Negri, Mariana; Buranelli, Raquel C; Robles, Rafael; Magalhães, Tatiana; Tamburus, Ana Francisca; Rossi, Natália; Miyazaki, Mayara J

2017-09-05

DNA barcoding has emerged as an efficient tool for taxonomy and other biodiversity fields. The vast and speciose group of decapod crustaceans is not an exception in the current scenario and comparing short DNA fragments has enabled researchers to overcome some taxonomic impediments to help broadening knowledge on the diversity of this group of crustaceans. Brazil is considered as an important area in terms of global marine biodiversity and some regions stand out in terms of decapod fauna, such as the São Paulo coastline. Thus, the aim of this study is to obtain sequences of the mitochondrial markers (COI and 16S) for decapod crustaceans distributed at the São Paulo coastline and to test the accuracy of these markers for species identification from this region by comparing our sequences to those already present in the GenBank database. We sampled along almost the 300 km of the São Paulo coastline from estuaries to offshore islands during the development of a multidisciplinary research project that took place for 5 years. All the species were processed to obtain the DNA sequences. The diversity of the decapod fauna on the São Paulo coastline comprises at least 404 species. We were able to collect 256 of those species and sequence of at least one of the target genes from 221. By testing the accuracy of these two DNA markers as a tool for identification, we were able to check our own identifications, including new records in GenBank, spot potential mistakes in GenBank, and detect potential new species.

Aedes albopictus (Skuse, 1894) infected with the American-Asian genotype of dengue type 2 virus in Medellín suggests its possible role as vector of dengue fever in Colombia.

PubMed

Gómez-Palacio, Andrés; Suaza-Vasco, Juan; Castaño, Sandra; Triana, Omar; Uribe, Sandra

2017-03-29

Aedes aegypti and Ae. albopictus are recognized vectors of dengue, yellow fever, chikungunya and Zika arboviruses in several countries worldwide. In Colombia, Ae. albopictus geographical distribution has increased to include highly populated cities such as Cali and Medellín. Although this species has been frequently found in urban and semi-urban zones in the country, its role as vector of the dengue fever is poorly known. To identify the presence of Ae. albopictus specimens naturally infected with dengue virus collected in Medellín. Insects were collected in the Universidad Nacional de Colombia campus in Medellín. Individuals were classified as Ae. albopictus and confirmed by DNA barcode region analysis. Mosquitoes were processed for dengue virus identification, and a fragment of the NS3 gen was sequenced and compared with DENV-2 genotypes reported in the literature. Sequence analysis of COI indicated Ae. albopictus individuals were similar to those recently reported in Colombia, and genetically close to those from other regions worldwide. Among the pools tested one was positive for DENV-2, and the NS3 analysis indicated it belonged to the Asian-American clade. We report the presence Ae. albopictus naturally infected with the Asian-American genotype of DENV-2 in Colombia. The presence of Ae. albopictus specimens carrying the most common genotype infecting humans in a highly populated city such as Medellín indicates its potential role as dengue vector in Colombia and highlights the relevance of including it in current vector surveillance strategies.

Services of DNA barcoding in different fields.

PubMed

Muhammad Tahir, Hafiz; Akhtar, Samreen

2016-11-01

DNA barcoding is a new master key for species identification and has greatly accelerated the pace of species discovery. In this novel and cost-effective technique, a short DNA sequence from a standard region of mitochondrial "CO1" gene called "barcode" is used. At present, researchers all over the world are utilizing this powerful tool for investigating biodiversity, differentiating cryptic species, testing food authenticity, identifying parasites, vectors, insect pests, and predators, monitoring of illegal trade of animals and their products, and identifying forensically important insects. In addition, this technique can potentially be used to monitor quality of drinking water, quickly identify the indicator species of lakes, rivers, and streams, identify species with harmful attributes or medicinal properties, monitor smuggling of endangered plants and animals and their products, and disease investigations. Despite non-favorable criticism from a few researchers, DNA barcoding has achieved immense popularity in the scientific community, especially among biologists. The present review provides an overview of DNA barcoding and its practical applications. The limitation, future prospective and main informative platforms for DNA barcoding have also been discussed.

Increasing global participation in genetics research through DNA barcoding.

PubMed

Adamowicz, Sarah J; Steinke, Dirk

2015-12-01

DNA barcoding--the sequencing of short, standardized DNA regions for specimen identification and species discovery--has promised to facilitate rapid access to biodiversity knowledge by diverse users. Here, we advance our opinion that increased global participation in genetics research is beneficial, both to scientists and for science, and explore the premise that DNA barcoding can help to democratize participation in genetics research. We examine publication patterns (2003-2014) in the DNA barcoding literature and compare trends with those in the broader, related domain of genomics. While genomics is the older and much larger field, the number of nations contributing to the published literature is similar between disciplines. Meanwhile, DNA barcoding exhibits a higher pace of growth in the number of publications as well as greater evenness among nations in their proportional contribution to total authorships. This exploration revealed DNA barcoding to be a highly international discipline, with growing participation by researchers in especially biodiverse nations. We briefly consider several of the challenges that may hinder further participation in genetics research, including access to training and molecular facilities as well as policy relating to the movement of genetic resources.

DNA barcoding of vouchered xylarium wood specimens of nine endangered Dalbergia species.

PubMed

Yu, Min; Jiao, Lichao; Guo, Juan; Wiedenhoeft, Alex C; He, Tuo; Jiang, Xiaomei; Yin, Yafang

2017-12-01

ITS2+ trnH - psbA was the best combination of DNA barcode to resolve the Dalbergia wood species studied. We demonstrate the feasibility of building a DNA barcode reference database using xylarium wood specimens. The increase in illegal logging and timber trade of CITES-listed tropical species necessitates the development of unambiguous identification methods at the species level. For these methods to be fully functional and deployable for law enforcement, they must work using wood or wood products. DNA barcoding of wood has been promoted as a promising tool for species identification; however, the main barrier to extensive application of DNA barcoding to wood is the lack of a comprehensive and reliable DNA reference library of barcodes from wood. In this study, xylarium wood specimens of nine Dalbergia species were selected from the Wood Collection of the Chinese Academy of Forestry and DNA was then extracted from them for further PCR amplification of eight potential DNA barcode sequences (ITS2, matK, trnL, trnH-psbA, trnV-trnM1, trnV-trnM2, trnC-petN, and trnS-trnG). The barcodes were tested singly and in combination for species-level discrimination ability by tree-based [neighbor-joining (NJ)] and distance-based (TaxonDNA) methods. We found that the discrimination ability of DNA barcodes in combination was higher than any single DNA marker among the Dalbergia species studied, with the best two-marker combination of ITS2+trnH-psbA analyzed with NJ trees performing the best (100% accuracy). These barcodes are relatively short regions (<350 bp) and amplification reactions were performed with high success (≥90%) using wood as the source material, a necessary factor to apply DNA barcoding to timber trade. The present results demonstrate the feasibility of using vouchered xylarium specimens to build DNA barcoding reference databases.

A DNA Barcode Library for North American Ephemeroptera: Progress and Prospects

PubMed Central

Webb, Jeffrey M.; Jacobus, Luke M.; Funk, David H.; Zhou, Xin; Kondratieff, Boris; Geraci, Christy J.; DeWalt, R. Edward; Baird, Donald J.; Richard, Barton; Phillips, Iain; Hebert, Paul D. N.

2012-01-01

DNA barcoding of aquatic macroinvertebrates holds much promise as a tool for taxonomic research and for providing the reliable identifications needed for water quality assessment programs. A prerequisite for identification using barcodes is a reliable reference library. We gathered 4165 sequences from the barcode region of the mitochondrial cytochrome c oxidase subunit I gene representing 264 nominal and 90 provisional species of mayflies (Insecta: Ephemeroptera) from Canada, Mexico, and the United States. No species shared barcode sequences and all can be identified with barcodes with the possible exception of some Caenis. Minimum interspecific distances ranged from 0.3–24.7% (mean: 12.5%), while the average intraspecific divergence was 1.97%. The latter value was inflated by the presence of very high divergences in some taxa. In fact, nearly 20% of the species included two or three haplotype clusters showing greater than 5.0% sequence divergence and some values are as high as 26.7%. Many of the species with high divergences are polyphyletic and likely represent species complexes. Indeed, many of these polyphyletic species have numerous synonyms and individuals in some barcode clusters show morphological attributes characteristic of the synonymized species. In light of our findings, it is imperative that type or topotype specimens be sequenced to correctly associate barcode clusters with morphological species concepts and to determine the status of currently synonymized species. PMID:22666447

The chloroplast psbK-psbI intergenic region, a potential genetic marker for broad sectional relationships in Anthurium

USDA-ARS?s Scientific Manuscript database

Nuclear and chloroplast genetic markers have been extensively used for plant identification and molecular taxonomy studies. The efficacy of genetic markers to be used as DNA barcodes is under constant evaluation and improvement, with identification of new barcodes that provide greater resolution an...

Evaluating the feasibility of using candidate DNA barcodes in discriminating species of the large Asteraceae family

PubMed Central

2010-01-01

Background Five DNA regions, namely, rbcL, matK, ITS, ITS2, and psbA-trnH, have been recommended as primary DNA barcodes for plants. Studies evaluating these regions for species identification in the large plant taxon, which includes a large number of closely related species, have rarely been reported. Results The feasibility of using the five proposed DNA regions was tested for discriminating plant species within Asteraceae, the largest family of flowering plants. Among these markers, ITS2 was the most useful in terms of universality, sequence variation, and identification capability in the Asteraceae family. The species discriminating power of ITS2 was also explored in a large pool of 3,490 Asteraceae sequences that represent 2,315 species belonging to 494 different genera. The result shows that ITS2 correctly identified 76.4% and 97.4% of plant samples at the species and genus levels, respectively. In addition, ITS2 displayed a variable ability to discriminate related species within different genera. Conclusions ITS2 is the best DNA barcode for the Asteraceae family. This approach significantly broadens the application of DNA barcoding to resolve classification problems in the family Asteraceae at the genera and species levels. PMID:20977734

Standardized molecular diagnostic tool for the identification of cryptic species within the Bemisia tabaci complex.

PubMed

Elfekih, Samia; Tay, Wee Tek; Gordon, Karl; Court, Leon N; De Barro, Paul J

2018-01-01

The whitefly Bemisia tabaci complex harbours over 40 cryptic species that have been placed in 11 phylogenetically distinct clades based on the molecular characterization of partial mitochondrial DNA COI (mtCOI) gene region. Four cryptic species are currently within the invasive clade, i.e. MED, MEAM1, MEAM2 and IO. Correct identification of these species is a critical step towards implementing reliable measures for plant biosecurity and border protection; however, no standardized B. tabaci-specific primers are currently available which has caused inconsistencies in the species identification processes. We report three sets of polymerase chain reaction (PCR) primers developed to amplify the mtCOI region which can be used for genotyping MED, MEAM1 and IO species, and tested these primers on 91 MED, 35 MEAM1 and five IO individuals. PCR and sequencing of amplicons identified a total of 21, six and one haplotypes in MED, MEAM1 and IO respectively, of which six haplotypes were new to the B. tabaci database. These primer pairs enabled standardization and robust molecular species identification via mtCOI screening of the targeted invasive cryptic species and will improve quarantine decisions. Use of this diagnostic tool could be extended to other species within the complex. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Molecular Identification of Shark Meat From Local Markets in Southern Brazil Based on DNA Barcoding: Evidence for Mislabeling and Trade of Endangered Species

PubMed Central

Almerón-Souza, Fernanda; Sperb, Christian; Castilho, Carolina L.; Figueiredo, Pedro I. C. C.; Gonçalves, Leonardo T.; Machado, Rodrigo; Oliveira, Larissa R.; Valiati, Victor H.; Fagundes, Nelson J. R.

2018-01-01

Elasmobranchs, the group of cartilaginous fishes that include sharks and rays, are especially vulnerable to overfishing due to low fecundity and late sexual maturation. A significant number of elasmobranch species are currently overexploited or threatened by fisheries activities. Additionally, several recent reports have indicated that there has been a reduction in regional elasmobranch population sizes. Brazil is an important player in elasmobranch fisheries and one of the largest importers of shark meat. However, carcasses entering the shark meat market have usually had their fins and head removed, which poses a challenge to reliable species identification based on the morphology of captured individuals. This is further complicated by the fact that the internal Brazilian market trades several different elasmobranch species under a common popular name: “cação.” The use of such imprecise nomenclature, even among governmental agencies, is problematic for both controlling the negative effects of shark consumption and informing the consumer about the origins of the product. In this study, we used DNA barcoding (mtDNA, COI gene) to identify, at the species level, “cação” samples available in local markets from Southern Brazil. We collected 63 samples traded as “cação,” which we found to correspond to 20 different species. These included two teleost species: Xiphias gladius (n = 1) and Genidens barbus (n = 6), and 18 species from seven elasmobranch orders (Carcharhiniformes, n = 42; Squaliformes, n = 3; Squatiniformes, n = 2; Rhinopristiformes, n = 4; Myliobatiformes, n = 3; Rajiformes, n = 1; and Torpediniformes, n = 1). The most common species in our sample were Prionace glauca (n = 15) and Sphyrna lewini (n = 14), while all other species were represented by four samples or less. Considering IUCN criteria, 47% of the elasmobranch species found are threatened at the global level, while 53% are threatened and 47% are critically endangered in Brazil. These results underline that labeling the meat of any shark species as “cação” is problematic for monitoring catch allocations from the fishing industry and discourages consumer engagement in conservationist practices through informed decision-making. PMID:29755504

Molecular Identification of Shark Meat From Local Markets in Southern Brazil Based on DNA Barcoding: Evidence for Mislabeling and Trade of Endangered Species.

PubMed

Almerón-Souza, Fernanda; Sperb, Christian; Castilho, Carolina L; Figueiredo, Pedro I C C; Gonçalves, Leonardo T; Machado, Rodrigo; Oliveira, Larissa R; Valiati, Victor H; Fagundes, Nelson J R

2018-01-01

Elasmobranchs, the group of cartilaginous fishes that include sharks and rays, are especially vulnerable to overfishing due to low fecundity and late sexual maturation. A significant number of elasmobranch species are currently overexploited or threatened by fisheries activities. Additionally, several recent reports have indicated that there has been a reduction in regional elasmobranch population sizes. Brazil is an important player in elasmobranch fisheries and one of the largest importers of shark meat. However, carcasses entering the shark meat market have usually had their fins and head removed, which poses a challenge to reliable species identification based on the morphology of captured individuals. This is further complicated by the fact that the internal Brazilian market trades several different elasmobranch species under a common popular name: "cação." The use of such imprecise nomenclature, even among governmental agencies, is problematic for both controlling the negative effects of shark consumption and informing the consumer about the origins of the product. In this study, we used DNA barcoding (mtDNA, COI gene) to identify, at the species level, "cação" samples available in local markets from Southern Brazil. We collected 63 samples traded as "cação," which we found to correspond to 20 different species. These included two teleost species: Xiphias gladius ( n = 1) and Genidens barbus ( n = 6), and 18 species from seven elasmobranch orders (Carcharhiniformes, n = 42; Squaliformes, n = 3; Squatiniformes, n = 2; Rhinopristiformes, n = 4; Myliobatiformes, n = 3; Rajiformes, n = 1; and Torpediniformes, n = 1). The most common species in our sample were Prionace glauca ( n = 15) and Sphyrna lewini ( n = 14), while all other species were represented by four samples or less. Considering IUCN criteria, 47% of the elasmobranch species found are threatened at the global level, while 53% are threatened and 47% are critically endangered in Brazil. These results underline that labeling the meat of any shark species as "cação" is problematic for monitoring catch allocations from the fishing industry and discourages consumer engagement in conservationist practices through informed decision-making.

Morphometric characteristics and COI haplotype diversity of Arctodiaptomus spinosus (Copedoda) populations in soda pans in Hungary.

PubMed

Forró, László; Nédli, Judit; Csata, Enikő; Krízsik, Virág; Balogh, Csilla; G-Tóth, László

2017-09-01

Arctodiaptomus spinosus (Daday, 1891) is a characteristic species of the soda pan zooplankton in the Great Hungarian Plain. The biogeographical distribution of the species is interesting, since its range expands from the Pannonian Biogeographic region to the other side of the Carpathians, occurring in saline lakes in Eastern Anatolia, Armenia, Iran and in temporary waters in Ukraine. Our investigations focused on the morphometric characteristics and the COI haplotype diversity of four Hungarian populations in the Kiskunság area. We detected substantial morphological differences between the Böddi-szék population and the rest of the sampling sites, however considerable differences were not observable in the COI haplotypes in the populations. The 20 animals investigated for COI haplotypes belonged to the same haplotype network. Tajima's D indicated departures from the neutral Wright - Fisher population model and suggested population expansion. The genetic composition of Arctodiaptomus spinosus populations in the Kiskunság area is rather uniform.

DNA barcodes from four loci provide poor resolution of taxonomic groups in the genus Crataegus.

PubMed

Zarrei, Mehdi; Talent, Nadia; Kuzmina, Maria; Lee, Jeanette; Lund, Jensen; Shipley, Paul R; Stefanović, Saša; Dickinson, Timothy A

2015-04-28

DNA barcodes can facilitate identification of organisms especially when morphological characters are limited or unobservable. To what extent this potential is realized in specific groups of plants remains to be determined. Libraries of barcode sequences from well-studied authoritatively identified plants represented by herbarium voucher specimens are needed in order for DNA barcodes to serve their intended purpose, where this is possible, and to understand the reasons behind their failure to do so, when this occurs. We evaluated four loci, widely regarded as universal DNA barcodes for plants, for their utility in hawthorn species identification. Three plastid regions, matK, rbcLa and psbA-trnH, and the internal transcribed spacer 2 (ITS2) of nuclear ribosomal DNA discriminate only some of the species of Crataegus that can be recognized on the basis of their morphology etc. This is, in part, because in Rosaceae tribe Maleae most individual plastid loci yield relatively little taxonomic resolution and, in part, because the effects of allopolyploidization have not been eliminated by concerted evolution of the ITS regions. Although individual plastid markers provided generally poor resolution of taxonomic groups in Crataegus, a few species were notable exceptions. In contrast, analyses of concatenated sequences of the 3 plastid barcode loci plus 11 additional plastid loci gave a well-resolved maternal phylogeny. In the ITS2 tree, different individuals of some species formed groups with taxonomically unrelated species. This is a sign of lineage sorting due to incomplete concerted evolution in ITS2. Incongruence between the ITS2 and plastid trees is best explained by hybridization between different lineages within the genus. In aggregate, limited between-species variation in plastid loci, hybridization and a lack of concerted evolution in ITS2 all combine to limit the utility of standard barcoding markers in Crataegus. These results have implications for authentication of hawthorn materials in natural health products. Published by Oxford University Press on behalf of the Annals of Botany Company.

DNA barcodes from four loci provide poor resolution of taxonomic groups in the genus Crataegus

PubMed Central

Zarrei, Mehdi; Talent, Nadia; Kuzmina, Maria; Lee, Jeanette; Lund, Jensen; Shipley, Paul R.; Stefanović, Saša; Dickinson, Timothy A.

2015-01-01

DNA barcodes can facilitate identification of organisms especially when morphological characters are limited or unobservable. To what extent this potential is realized in specific groups of plants remains to be determined. Libraries of barcode sequences from well-studied authoritatively identified plants represented by herbarium voucher specimens are needed in order for DNA barcodes to serve their intended purpose, where this is possible, and to understand the reasons behind their failure to do so, when this occurs. We evaluated four loci, widely regarded as universal DNA barcodes for plants, for their utility in hawthorn species identification. Three plastid regions, matK, rbcLa and psbA-trnH, and the internal transcribed spacer 2 (ITS2) of nuclear ribosomal DNA discriminate only some of the species of Crataegus that can be recognized on the basis of their morphology etc. This is, in part, because in Rosaceae tribe Maleae most individual plastid loci yield relatively little taxonomic resolution and, in part, because the effects of allopolyploidization have not been eliminated by concerted evolution of the ITS regions. Although individual plastid markers provided generally poor resolution of taxonomic groups in Crataegus, a few species were notable exceptions. In contrast, analyses of concatenated sequences of the 3 plastid barcode loci plus 11 additional plastid loci gave a well-resolved maternal phylogeny. In the ITS2 tree, different individuals of some species formed groups with taxonomically unrelated species. This is a sign of lineage sorting due to incomplete concerted evolution in ITS2. Incongruence between the ITS2 and plastid trees is best explained by hybridization between different lineages within the genus. In aggregate, limited between-species variation in plastid loci, hybridization and a lack of concerted evolution in ITS2 all combine to limit the utility of standard barcoding markers in Crataegus. These results have implications for authentication of hawthorn materials in natural health products. PMID:25926325

Identification of processed Chinese medicinal materials using DNA mini-barcoding.

PubMed

Song, Ming; Dong, Gang-Qiang; Zhang, Ya-Qin; Liu, Xia; Sun, Wei

2017-07-01

Most of Chinese medicinal herbs are subjected to traditional processing procedures, including stir-frying, charring, steaming, boiling, and calcining before they are released into dispensaries. The marketing and identification of processed medicinal materials is a growing issue in the marketplace. However, conventional methods of identification have limitations, while DNA mini-barcoding, based on the sequencing of a short-standardized region, has received considerable attention as a new potential means to identify processed medicinal materials. In the present study, six DNA barcode loci including ITS2, psbA-trnH, rbcL, matK, trnL (UAA) intron and its P6 loop, were employed for the authentication of 45 processed samples belonging to 15 species. We evaluated the amplification efficiency of each locus. We also examined the identification accuracy of the potential mini-barcode locus, of trnL (UAA) intron P6 loop. Our results showed that the five primary barcode loci were successfully amplified in only 8.89%-20% of the processed samples, while the amplification rates of the trnL (UAA) intron P6 loop were higher, at 75.56% successful amplification. We compared the mini-barcode sequences with Genbank using the Blast program. The analysis showed that 45.23% samples could be identified to genus level, while only one sample could be identified to the species level. We conclude that trnL (UAA) p6 loop is a candidate mini-barcode that has shown its potential and may become a universal mini-barcode as complementary barcode for authenticity testing and will play an important role in medicinal materials control. Copyright © 2017 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

DNA Barcoding the Geometrid Fauna of Bavaria (Lepidoptera): Successes, Surprises, and Questions

PubMed Central

Hausmann, Axel; Haszprunar, Gerhard; Hebert, Paul D. N.

2011-01-01

Background The State of Bavaria is involved in a research program that will lead to the construction of a DNA barcode library for all animal species within its territorial boundaries. The present study provides a comprehensive DNA barcode library for the Geometridae, one of the most diverse of insect families. Methodology/Principal Findings This study reports DNA barcodes for 400 Bavarian geometrid species, 98 per cent of the known fauna, and approximately one per cent of all Bavarian animal species. Although 98.5% of these species possess diagnostic barcode sequences in Bavaria, records from neighbouring countries suggest that species-level resolution may be compromised in up to 3.5% of cases. All taxa which apparently share barcodes are discussed in detail. One case of modest divergence (1.4%) revealed a species overlooked by the current taxonomic system: Eupithecia goossensiata Mabille, 1869 stat.n. is raised from synonymy with Eupithecia absinthiata (Clerck, 1759) to species rank. Deep intraspecific sequence divergences (>2%) were detected in 20 traditionally recognized species. Conclusions/Significance The study emphasizes the effectiveness of DNA barcoding as a tool for monitoring biodiversity. Open access is provided to a data set that includes records for 1,395 geometrid specimens (331 species) from Bavaria, with 69 additional species from neighbouring regions. Taxa with deep intraspecific sequence divergences are undergoing more detailed analysis to ascertain if they represent cases of cryptic diversity. PMID:21423340

Barcoding Fauna Bavarica: 78% of the Neuropterida Fauna Barcoded!

PubMed Central

Morinière, Jérome; Hendrich, Lars; Hausmann, Axel; Hebert, Paul; Haszprunar, Gerhard; Gruppe, Axel

2014-01-01

This publication provides the first comprehensive DNA barcode data set for the Neuropterida of Central Europe, including 80 of the 102 species (78%) recorded from Bavaria (Germany) and three other species from nearby regions (Austria, France and the UK). Although the 286 specimens analyzed had a heterogeneous conservation history (60% dried; 30% in 80% EtOH; 10% fresh specimens in 95% EtOH), 237 (83%) generated a DNA barcode. Eleven species (13%) shared a BIN, but three of these taxa could be discriminated through barcodes. Four pairs of closely allied species shared barcodes including Chrysoperla pallida Henry et al., 2002 and C. lucasina Lacroix, 1912; Wesmaelius concinnus (Stephens, 1836) and W. quadrifasciatus (Reuter, 1894); Hemerobius handschini Tjeder, 1957 and H. nitidulus Fabricius, 1777; and H. atrifrons McLachlan, 1868 and H. contumax Tjeder, 1932. Further studies are needed to test the possible synonymy of these species pairs or to determine if other genetic markers permit their discrimination. Our data highlight five cases of potential cryptic diversity within Bavarian Neuropterida: Nineta flava (Scopoli, 1763), Sympherobius pygmaeus (Rambur, 1842), Sisyra nigra (Retzius, 1783), Semidalis aleyrodiformis (Stephens, 1836) and Coniopteryx pygmaea Enderlein, 1906 are each split into two or three BINs. The present DNA barcode library not only allows the identification of adult and larval stages, but also provides valuable information for alpha-taxonomy, and for ecological and evolutionary research. PMID:25286434

A comprehensive DNA barcode database for Central European beetles with a focus on Germany: adding more than 3500 identified species to BOLD.

PubMed

Hendrich, Lars; Morinière, Jérôme; Haszprunar, Gerhard; Hebert, Paul D N; Hausmann, Axel; Köhler, Frank; Balke, Michael

2015-07-01

Beetles are the most diverse group of animals and are crucial for ecosystem functioning. In many countries, they are well established for environmental impact assessment, but even in the well-studied Central European fauna, species identification can be very difficult. A comprehensive and taxonomically well-curated DNA barcode library could remedy this deficit and could also link hundreds of years of traditional knowledge with next generation sequencing technology. However, such a beetle library is missing to date. This study provides the globally largest DNA barcode reference library for Coleoptera for 15 948 individuals belonging to 3514 well-identified species (53% of the German fauna) with representatives from 97 of 103 families (94%). This study is the first comprehensive regional test of the efficiency of DNA barcoding for beetles with a focus on Germany. Sequences ≥500 bp were recovered from 63% of the specimens analysed (15 948 of 25 294) with short sequences from another 997 specimens. Whereas most specimens (92.2%) could be unambiguously assigned to a single known species by sequence diversity at CO1, 1089 specimens (6.8%) were assigned to more than one Barcode Index Number (BIN), creating 395 BINs which need further study to ascertain if they represent cryptic species, mitochondrial introgression, or simply regional variation in widespread species. We found 409 specimens (2.6%) that shared a BIN assignment with another species, most involving a pair of closely allied species as 43 BINs were involved. Most of these taxa were separated by barcodes although sequence divergences were low. Only 155 specimens (0.97%) show identical or overlapping clusters. © 2014 John Wiley & Sons Ltd.

Designation of a neotype and redescription of Hesione reticulata von Marenzeller, 1879 from Japan (Annelida, Hesionidae).

PubMed

Jimi, Naoto; Salazar-Vallejo, Sergio I; Kajihara, Hiroshi

2017-01-01

The hesionid polychaete Hesione reticulata von Marenzeller, 1879 was described from Enoshima Island, Japan and has been recorded also from the Red Sea. Depending on researchers, it has been regarded as either a distinct species or synonymous with older established ones. The type specimen has been lost. In order to clarify its taxonomic status, Hesione reticulata is herein redescribed, illustrated, and a neotype is proposed based on recent material collected near the type locality. The diagnostic features include the presence of several dorsal, discontinuous longitudinal bands, interrupted by pale segmental spots; prostomium with tiny antennae; a tuberculated dorsal integument; acicular lobes double; and neurochaetal blades with guards approaching the distal tooth. The dorsal color pattern in life enables a clear distinction from similar species such as Hesione intertexta Grube, 1878 amongst others. Mitochondrial COI barcoding sequences are deposited in the DNA Data Bank of Japan. A key to Hesione species from Japan is also included.

A case study on the labeling of bottarga produced in Sardinia from ovaries of grey mullets (Mugil cephalus and Mugil capurrii) caught in Eastern Central Atlantic coasts

PubMed Central

Piras, Pierluigi; Sardu, Francesco; Meloni, Domenico; Riina, Maria Vittoria; Beltramo, Chiara; Acutis, Pier Luigi

2018-01-01

The aim of this case study is to show how traditional and molecular methods can be employed to identify the Mugilidae species currently used in Sardinia (Italy) to produce the traditional bottarga for the processing of their ovaries. A total of six specimens of Mugil cephalus (n=3) and Mugil capurrii (n=3) were subjected to external morphology and meristic measurements. Subsequently, tissue samples of white muscle and ovaries from three individuals per species were underwent PCR-sequencing assay of mitochondrial DNA cytochrome oxidase subunit I (COI). The external morphology and meristic characters showed a sufficient level of reliability in the identification between the two species. At the same time, the molecular techniques showed the discriminatory power and confirmed the correct species identification in all the sampling units. DNA barcoding may be an effective aid to traditional taxonomy and can facilitate accurate species identification among the Mugilidae. PMID:29732322

Preliminary Characterization of Mitochondrial Genome of Melipona scutellaris, a Brazilian Stingless Bee

PubMed Central

Silverio, Manuella Souza; Rodovalho, Vinícius de Rezende; Bonetti, Ana Maria; de Oliveira, Guilherme Corrêa; Cuadros-Orellana, Sara; Ueira-Vieira, Carlos; Rodrigues dos Santos, Anderson

2014-01-01

Bees are manufacturers of relevant economical products and have a pollinator role fundamental to ecosystems. Traditionally, studies focused on the genus Melipona have been mostly based on behavioral, and social organization and ecological aspects. Only recently the evolutionary history of this genus has been assessed using molecular markers, including mitochondrial genes. Even though these studies have shed light on the evolutionary history of the Melipona genus, a more accurate picture may emerge when full nuclear and mitochondrial genomes of Melipona species become available. Here we present the assembly, annotation, and characterization of a draft mitochondrial genome of the Brazilian stingless bee Melipona scutellaris using Melipona bicolor as a reference organism. Using Illumina MiSeq data, we achieved the annotation of all protein coding genes, as well as the genes for the two ribosomal subunits (16S and 12S) and transfer RNA genes as well. Using the COI sequence as a DNA barcode, we found that M. cramptoni is the closest species to M. scutellaris. PMID:25019088

Preliminary characterization of mitochondrial genome of Melipona scutellaris, a Brazilian stingless bee.

PubMed

Silverio, Manuella Souza; Rodovalho, Vinícius de Rezende; Bonetti, Ana Maria; de Oliveira, Guilherme Corrêa; Cuadros-Orellana, Sara; Ueira-Vieira, Carlos; Rodrigues dos Santos, Anderson

2014-01-01

Bees are manufacturers of relevant economical products and have a pollinator role fundamental to ecosystems. Traditionally, studies focused on the genus Melipona have been mostly based on behavioral, and social organization and ecological aspects. Only recently the evolutionary history of this genus has been assessed using molecular markers, including mitochondrial genes. Even though these studies have shed light on the evolutionary history of the Melipona genus, a more accurate picture may emerge when full nuclear and mitochondrial genomes of Melipona species become available. Here we present the assembly, annotation, and characterization of a draft mitochondrial genome of the Brazilian stingless bee Melipona scutellaris using Melipona bicolor as a reference organism. Using Illumina MiSeq data, we achieved the annotation of all protein coding genes, as well as the genes for the two ribosomal subunits (16S and 12S) and transfer RNA genes as well. Using the COI sequence as a DNA barcode, we found that M. cramptoni is the closest species to M. scutellaris.

Host-ant specificity of endangered large blue butterflies (Phengaris spp., Lepidoptera: Lycaenidae) in Japan.

PubMed

Ueda, Shouhei; Komatsu, Takashi; Itino, Takao; Arai, Ryusuke; Sakamoto, Hironori

2016-11-03

Large blue butterflies, Phengaris (Maculinea), are an important focus of endangered-species conservation in Eurasia. Later-instar Phengaris caterpillars live in Myrmica ant nests and exploit the ant colony's resources, and they are specialized to specific host-ant species. For example, local extinction of P. arion in the U. K. is thought to have been due to the replacement of its host-ant species with a less-suitable congener, as a result of changes in habitat. In Japan, Myrmica kotokui hosts P. teleius and P. arionides caterpillars. We recently showed, however, that the morphological species M. kotokui actually comprises four genetic clades. Therefore, to determine to which group of ants the hosts of these two Japanese Phengaris species belong, we used mitochondrial COI-barcoding of M. kotokui specimens from colonies in the habitats of P. teleius and P. arionides to identify the ant clade actually parasitized by the caterpillars of each species. We found that these two butterfly species parasitize different ant clades within M. kotokui.

Sea cucumber species identification of family Caudinidae from Surabaya based on morphological and mitochondrial DNA evidence

NASA Astrophysics Data System (ADS)

Amin, Muhammad Hilman Fu'adil; Pidada, Ida Bagus Rai; Sugiharto, Widyatmoko, Johan Nuari; Irawan, Bambang

2016-03-01

Species identification and taxonomy of sea cucumber remains a challenge problem in some taxa. Caudinidae family of sea cucumber was comerciallized in Surabaya, and it was used as sea cucumber chips. Members of Caudinid sea cucumber have similiar morphology, so it is hard to identify this sea cucumber only from morphological appearance. DNA barcoding is useful method to overcome this problem. The aim of this study was to determine Caudinid specimen of sea cucumber in East Java by morphological and molecular approach. Sample was collected from east coast of Surabaya, then preserved in absolute ethanol. After DNA isolation, Cytochrome Oxydase I (COI) gene amplification was performed using Echinoderm universal primer and PCR product was sequenced. Sequencing result was analyzed and identified in NCBI database using BLAST. Results showed that Caudinid specimen in have closely related to Acaudina molpadioides sequence in GenBank with 86% identity. Morphological data, especially based on ossicle, also showed that the specimen is Acaudina molpadioides.

Filling the BINs of life: Report of an amphibian and reptile survey of the Tanintharyi (Tenasserim) Region of Myanmar, with DNA barcode data.

PubMed

Mulcahy, Daniel G; Lee, Justin L; Miller, Aryeh H; Chand, Mia; Thura, Myint Kyaw; Zug, George R

2018-01-01

Despite threats of species extinctions, taxonomic crises, and technological advances in genomics and natural history database informatics, we are still distant from cataloguing all of the species of life on earth. Amphibians and reptiles are no exceptions; in fact new species are described nearly every day and many species face possible extinction. The number of described species continues to climb as new areas of the world are explored and as species complexes are examined more thoroughly. The use of DNA barcoding provides a mechanism for rapidly estimating the number of species at a given site and has the potential to record all of the species of life on Earth. Though DNA barcoding has its caveats, it can be useful to estimate the number of species in a more systematic and efficient manner, to be followed in combination with more traditional, morphology-based identifications and species descriptions. Herein, we report the results of a voucher-based herpetological expedition to the Tanintharyi (Tenasserim) Region of Myanmar, enhanced with DNA barcode data. Our main surveys took place in the currently proposed Tanintharyi National Park. We combine our results with photographs and observational data from the Chaung-nauk-pyan forest reserve. Additionally, we provide the first checklist of amphibians and reptiles of the region, with species based on the literature and museum. Amphibians, anurans in particular, are one of the most poorly known groups of vertebrates in terms of taxonomy and the number of known species, particularly in Southeast Asia. Our rapid-assessment program combined with DNA barcoding and use of Barcode Index Numbers (BINs) of voucher specimens reveals the depth of taxonomic diversity in the southern Tanintharyi herpetofauna even though only a third of the potential amphibians and reptiles were seen. A total of 51 putative species (one caecilian, 25 frogs, 13 lizards, 10 snakes, and two turtles) were detected, several of which represent potentially undescribed species. Several of these species were detected by DNA barcode data alone. Furthermore, five species were recorded for the first time in Myanmar, two amphibians (Ichthyophis cf. kohtaoensis and Chalcorana eschatia ) and three snakes ( Ahaetulla mycterizans , Boiga dendrophila , and Boiga drapiezii ).