scholarly journals Assessment of the macroinvertebrate community of the Vjosa river through non-destructive DNA metabarcoding of preservative ethanol

2021 ◽  
Vol 4 ◽  
Author(s):  
Marie Brasseur ◽  
Simon Vitecek ◽  
Vera Zizka ◽  
Jan Martini ◽  
Remo Wüthrich ◽  
...  
Keyword(s):  
Habitat Diversity ◽  
Molecular Data ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Natural State ◽  
Dna Metabarcoding ◽  
Balkan Region ◽  
Stream Biodiversity ◽  
Dna Template ◽  
Non Destructive

Streams and rivers represent hotspots of biodiversity in their natural state. This biodiversity is declining worldwide due to pollution, exploitation and hydromorphological degradation of these systems. One of the last big, natural rivers in Europe is the Vjosa in the Balkan region. The catchment is characterized by natural flow dynamics, resulting in high habitat diversity and turnover, and hosts several sensitive and endemic species (e.g. Isoperla vjosae). DNA metabarcoding represents a promising approach to characterize this biodiversity but methodological drawbacks such as primer bias or incomplete reference databases limit the application, particularly in taxonomically underexplored regions. Here, we assessed stream biodiversity with a focus on macrozoobenthic (MZB) taxa via a non-destructive, voucher-preserving DNA metabarcoding protocol. In this approach, ethanol used for preservation of multi-habitat samples in the field was used as DNA template, allowing to retain the integrity of the original sample and further comparison of molecular and morphological taxa lists. Samples were taken in spring and autumn 2018 at 48 sites allocated over the Vjosa catchment. The preservative ethanol was filtered through 0.43 um nitrocellulose membranes from which DNA was extracted. Subsequently, a 421 bp fragment of the COI gene was amplified with the primer pair BF2/BR2 and Illumina sequenced. After filtering for sequences with similarity to reference entries >85%, 4,123 OTUs were obtained, of which 921 were identified as MZB taxa. Dipterans and ephemeropterans were most abundant, followed by plecopterans. Some taxa (e.g. molluscs) were not identified due to a known primer bias and >7,000 OTUs could not be assigned above 85 % similarity. Using the here presented voucher-preserving approach allowed us to identify the pitfalls of DNA metabarcoding as tool for biodiversity assessment in taxonomically unexplored regions such as the Vjosa catchment. However, the comparison of specimen abundance data and molecular data showed the power of non-destructive fixative metabarcoding for detecting MZB communities with highly increased taxonomic resolution.

scholarly journals Preliminary analysis of New Zealand scampi (Metanephrops challengeri) diet using metabarcoding

2018 ◽  
Author(s):  
Aimee L van der Reis ◽  
Olivier Laroche ◽  
Andrew G Jeffs ◽  
Shane D Lavery
Keyword(s):  
New Zealand ◽  
Preliminary Analysis ◽  
18S Rrna ◽  
18S Rrna Gene ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Gut Contents ◽  
Rrna Gene ◽  
Diverse Range ◽  
Dna Metabarcoding

Deep sea lobsters are highly valued for seafood and provide the basis of important commercial fisheries in many parts of the world. Despite their economic significance, relatively little is known about their natural diets. Microscopic analyses of foregut content in some species have suffered from low taxonomic resolution, with many of the dietary items difficult to reliably identify as their tissue is easily digested. DNA metabarcoding has the potential to provide greater taxonomic resolution of the diet of the New Zealand scampi (Metanephrops challengeri) through the identification of gut contents, but a number of methodological concerns need to be overcome first to ensure optimum DNA metabarcoding results. In this study, a range of methodological parameters were tested to determine the optimum protocols for DNA metabarcoding, and provide a first view of M. challengeri diet. Several PCR protocols were tested, using two universal primer pairs targeting the 18S rRNA and COI genes, on DNA extracted from both frozen and ethanol preserved samples for both foregut and hindgut digesta. The selection of appropriate DNA polymerases, buffers and methods for reducing PCR inhibitors (including the use of BSA) were found to be critical. Amplification from frozen or ethanol preserved gut contents appeared similarly dependable, but metabarcoding outcomes indicated that the ethanol samples produced better results from the COI gene. The COI gene was found to be more effective than 18S rRNA gene for identifying large eukaryotic taxa from the digesta, however, it was less successfully amplified. The 18S rRNA gene was more easily amplified, but identified mostly smaller marine organisms such as plankton and parasites. This preliminary analysis of the diet of M. challengeri identified a range of species (13,541 reads identified as diet), which included the ghost shark (Hydrolagus novaezealandiae), silver warehou (Seriolella punctate), tall sea pen (Funiculina quadrangularis) and the salp (Ihlea racovitza), suggesting that they have a varied diet, with a high reliance on scavenging a diverse range of pelagic and benthic species from the seafloor.

scholarly journals New insight into the taxonomic resolution of the genus Bythotrephes Leydig (Crustacea: Cladocera) based on molecular data from Central Europe

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Maciej Karpowicz ◽  
Magdalena Świsłocka ◽  
Joanna Moroz ◽  
Łukasz Sługocki
Keyword(s):  
Genetic Analysis ◽  
Central Europe ◽  
Taxonomic Status ◽  
Molecular Data ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Monotypic Genus ◽  
Mtdna Coi ◽  
Xix Century ◽  
Insight Into

AbstractThe taxonomic status of the genus Bythotrephes Leydig (Crustacea: Cladocera) has been debated since the second half of the XIX century. The most widespread view of recent decades has been that Bythotrephes is a monotypic genus, which was support by preliminary molecular data. However, the recent detailed morphological revision of this genus clearly distinguishes at least seven species. Therefore, we performed a multi-lake survey in Central Europe to give new insight into the taxonomic status of Bythotrephes by combining genetic analysis with traditional morphology-based taxonomy. Based on the morphology we identified two species in Central Europe, B. brevimanus and B. lilljeborgi, as well as hybrid forms. For the genetic analysis, we used newly obtained 113 sequences of mtDNA COI gene of the 535-bp length Bythotrephes from Central Europe and sequences downloaded from GenBank. There were no significant differences between all analyzed sequences, which supports the hypothesis that Bythotrephes is a monotypic genus, with only one highly polymorphic species. On the other hand, the results of our work could point out that the COI gene is insufficient to evaluate the taxonomic status of Bythotrephes. Nonetheless, we have identified 29 new haplotypes of mtDNA COI, and one which was the same as the haplotype found in North America and Finland. Furthermore, this haplotype was the source variant from which most other haplotypes were derived.

scholarly journals Testing the potential of a ribosomal 16S marker for DNA metabarcoding of insects

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e1966 ◽  
Author(s):  
Vasco Elbrecht ◽  
Pierre Taberlet ◽  
Tony Dejean ◽  
Alice Valentini ◽  
Philippe Usseglio-Polatera ◽  
...  
Keyword(s):  
Species Level ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Reference Database ◽  
Bias Estimation ◽  
Knowing That ◽  
Dna Metabarcoding ◽  
Local Reference ◽  
Comprehensive Survey ◽  
Reference Databases

Cytochrome c oxidase I (COI) is a powerful marker for DNA barcoding of animals, with good taxonomic resolution and a large reference database. However, when used for DNA metabarcoding, estimation of taxa abundances and species detection are limited due to primer bias caused by highly variable primer binding sites across the COI gene. Therefore, we explored the ability of the 16S ribosomal DNA gene as an alternative metabarcoding marker for species level assessments. Ten bulk samples, each containing equal amounts of tissue from 52 freshwater invertebrate taxa, were sequenced with the Illumina NextSeq 500 system. The 16S primers amplified three more insect species than the Folmer COI primers and amplified more equally, probably due to decreased primer bias. Estimation of biomass might be less biased with 16S than with COI, although variation in read abundances of two orders of magnitudes is still observed. According to these results, the marker choice depends on the scientific question. If the goal is to obtain a taxonomic identification at the species level, then COI is more appropriate due to established reference databases and known taxonomic resolution of this marker, knowing that a greater proportion of insects will be missed using COI Folmer primers. If the goal is to obtain a more comprehensive survey the 16S marker, which requires building a local reference database, or optimised degenerated COI primers could be more appropriate.

scholarly journals Testing the potential of a ribosomal 16S marker for DNA metabarcoding of insects

2016 ◽  
Author(s):  
Vasco Elbrecht ◽  
Pierre Taberlet ◽  
Tony Dejean ◽  
Alice Valentini ◽  
Philippe Usseglio-polatera ◽  
...  
Keyword(s):  
Species Level ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Reference Database ◽  
Knowing That ◽  
Freshwater Invertebrate ◽  
Dna Metabarcoding ◽  
Local Reference ◽  
Comprehensive Survey ◽  
Reference Databases

Cytochrome c oxidase I (COI) is a powerful marker for DNA barcoding of animals, with good taxonomic resolution and a large reference database. However, when used for DNA metabarcoding, estimation of taxa abundances and species detection are limited due to primer bias caused by highly variable primer binding sites across the COI gene. Therefore, we explored the ability of the 16S ribosomal DNA gene as an alternative metabarcoding marker for species level assessments. Ten bulk samples, each containing equal amounts of tissue from 52 freshwater invertebrate taxa, were sequenced with the Illumina NextSeq 500 system. In comparison to COI, the 16S marker amplified more insect species and amplified more equally, probably due to decreased primer bias. Rough estimation of biomass might thus be less biased with 16S than with COI. According to these results, the marker choice depends on the scientific question. If the goal is to obtain a taxonomic identification at the species level, then COI is more appropriate due to established reference databases and known taxonomic resolution of this marker, knowing that a greater proportion of species will be missed using COI Folmer primers. If the goal is to obtain a more comprehensive survey in a context where it is possible to build a local reference database, the 16S marker could be more appropriate.

scholarly journals Preliminary analysis of New Zealand scampi (Metanephrops challengeri) diet using metabarcoding

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e5641 ◽  
Author(s):  
Aimee L. van der Reis ◽  
Olivier Laroche ◽  
Andrew G. Jeffs ◽  
Shane D. Lavery
Keyword(s):  
New Zealand ◽  
Preliminary Analysis ◽  
18S Rrna ◽  
18S Rrna Gene ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Gut Contents ◽  
Rrna Gene ◽  
Diverse Range ◽  
Dna Metabarcoding

Deep sea lobsters are highly valued for seafood and provide the basis of important commercial fisheries in many parts of the world. Despite their economic significance, relatively little is known about their natural diets. Microscopic analyses of foregut content in some species have suffered from low taxonomic resolution, with many of the dietary items difficult to reliably identify as their tissue is easily digested. DNA metabarcoding has the potential to provide greater taxonomic resolution of the diet of the New Zealand scampi (Metanephrops challengeri) through the identification of gut contents, but a number of methodological concerns need to be overcome first to ensure optimum DNA metabarcoding results. In this study, a range of methodological parameters were tested to determine the optimum protocols for DNA metabarcoding, and provide a first view ofM.challengeridiet. Several PCR protocols were tested, using two universal primer pairs targeting the 18S rRNA and COI genes, on DNA extracted from both frozen and ethanol preserved samples for both foregut and hindgut digesta. The selection of appropriate DNA polymerases, buffers and methods for reducing PCR inhibitors (including the use of BSA) were found to be critical. Amplification from frozen or ethanol preserved gut contents appeared similarly dependable. The COI gene was found to be more effective than 18S rRNA gene for identifying large eukaryotic taxa from the digesta; however, it was less successfully amplified. The 18S rRNA gene was more easily amplified, but identified mostly smaller marine organisms such as plankton and parasites. This preliminary analysis of the diet ofM.challengeriidentified a range of species (13,541 reads identified as diet), which included the ghost shark (Hydrolagus novaezealandiae), silver warehou (Seriolella punctata), tall sea pen (Funiculina quadrangularis) and the salp (Ihlea racovitzai), suggesting that they have a varied diet, with a high reliance on scavenging a diverse range of pelagic and benthic species from the seafloor.

scholarly journals DNA Barcoding for Delimitation of Putative Mexican Marine Nematodes Species

Diversity ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 107 ◽  
Author(s):  
Arely Martínez-Arce ◽  
Alberto De Jesús-Navarrete ◽  
Francesca Leasi
Keyword(s):  
Dna Barcoding ◽  
Low Cost ◽  
Nematode Species ◽  
Molecular Data ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Species List ◽  
Nematode Diversity ◽  
Coi Sequences ◽  
Morphological And Molecular Data

Nematode biodiversity is mostly unknown; while about 20,000 nematode species have been described, estimates for species diversity range from 0.1 to 100 million. The study of nematode diversity, like that of meiofaunal organisms in general, has been mostly based on morphology-based taxonomy, a time-consuming and costly task that requires well-trained specialists. This work represents the first study on the taxonomy of Mexican nematodes that integrates morphological and molecular data. We added eleven new records to the Mexican Caribbean nematode species list: Anticomidae sp.1, Catanema sp.1, Enoploides gryphus, Eurystomina sp.1, Haliplectus bickneri, Metachromadora sp.1, Odontophora bermudensis, Oncholaimus sp.1, Onyx litorale, Proplatycoma fleurdelis, and Pontonema cf. simile. We improved the COI database with 57 new sequences from 20 morphotypes. All COI sequences obtained in this work are new entries for the international genetic databases GenBank and BOLD. Among the studied sites, we report the most extensive species record (12 species) at Cozumel. DNA barcoding and species delineation methods supported the occurrence of 20 evolutionary independent entities and confirmed the high taxonomic resolution of the COI gene. Different approaches provided consistent results: ABGD and mPTP methods disentangled 20 entities, whereas Barcode Index Numbers (BINs) recovered 22 genetic species. Results support DNA barcoding being an efficient, fast, and low-cost method to integrate into morphological observations in order to address taxonomical shortfalls in meiofaunal organisms.

scholarly journals Preliminary analysis of New Zealand scampi (Metanephrops challengeri) diet using metabarcoding

2018 ◽  
Author(s):  
Aimee L van der Reis ◽  
Olivier Laroche ◽  
Andrew G Jeffs ◽  
Shane D Lavery
Keyword(s):  
New Zealand ◽  
Preliminary Analysis ◽  
18S Rrna ◽  
18S Rrna Gene ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Gut Contents ◽  
Rrna Gene ◽  
Diverse Range ◽  
Dna Metabarcoding

Deep sea lobsters are highly valued for seafood and provide the basis of important commercial fisheries in many parts of the world. Despite their economic significance, relatively little is known about their natural diets. Microscopic analyses of foregut content in some species have suffered from low taxonomic resolution, with many of the dietary items difficult to reliably identify as their tissue is easily digested. DNA metabarcoding has the potential to provide greater taxonomic resolution of foregut and hindgut contents of the New Zealand (NZ) scampi (Metanephrops challengeri), but a number of methodological concerns need to first be overcome to ensure optimum DNA metabarcoding results. In this study, a range of methodological parameters were trialled to determine the optimum protocols for DNA metabarcoding, and provide a first view of the NZ scampi diet. Several PCR protocols were trialled, using two universal primer pairs targeting the 18S rRNA and COI genes, on DNA extracted from frozen and ethanol preserved samples of both foregut and hindgut digesta. The selection of appropriate DNA polymerases, buffers and methods for reducing PCR inhibitors (including use of BSA) were found to be critical. Amplification from frozen or ethanol preserved gut contents appeared similarly dependable, but metabarcoding results showed that ethanol samples resulted in better results from the COI gene. The COI gene was found to be more effective than 18S rRNA gene for identifying large eukaryotic taxa from the digesta, however, it was less successfully amplified. The 18S rRNA gene was more easily amplified, but identified mostly smaller marine organisms such as plankton and parasites. This preliminary analysis of the diet of the NZ scampi identified a range of species, which included the ghost shark (Hydrolagus novaezealandiae), silver warehou (Seriolella punctate), tall sea pen (Funiculina quadrangularis ) and salp (Ihlea racovitza), suggesting that they have a varied diet, with a high reliance on scavenging a diverse range of pelagic and benthic species from the seafloor.

scholarly journals Testing the potential of a ribosomal 16S marker for DNA metabarcoding of insects

2016 ◽  
Author(s):  
Vasco Elbrecht ◽  
Pierre Taberlet ◽  
Tony Dejean ◽  
Alice Valentini ◽  
Philippe Usseglio-polatera ◽  
...  
Keyword(s):  
Species Level ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Reference Database ◽  
Knowing That ◽  
Freshwater Invertebrate ◽  
Dna Metabarcoding ◽  
Local Reference ◽  
Comprehensive Survey ◽  
Reference Databases

Cytochrome c oxidase I (COI) is a powerful marker for DNA barcoding of animals, with good taxonomic resolution and a large reference database. However, when used for DNA metabarcoding, estimation of taxa abundances and species detection are limited due to primer bias caused by highly variable primer binding sites across the COI gene. Therefore, we explored the ability of the 16S ribosomal DNA gene as an alternative metabarcoding marker for species level assessments. Ten bulk samples, each containing equal amounts of tissue from 52 freshwater invertebrate taxa, were sequenced with the Illumina NextSeq 500 system. In comparison to COI, the 16S marker amplified more insect species and amplified more equally, probably due to decreased primer bias. Rough estimation of biomass might thus be less biased with 16S than with COI. According to these results, the marker choice depends on the scientific question. If the goal is to obtain a taxonomic identification at the species level, then COI is more appropriate due to established reference databases and known taxonomic resolution of this marker, knowing that a greater proportion of species will be missed using COI Folmer primers. If the goal is to obtain a more comprehensive survey in a context where it is possible to build a local reference database, the 16S marker could be more appropriate.

scholarly journals Preliminary analysis of New Zealand scampi (Metanephrops challengeri) diet using metabarcoding

2018 ◽  
Author(s):  
Aimee L van der Reis ◽  
Olivier Laroche ◽  
Andrew G Jeffs ◽  
Shane D Lavery
Keyword(s):  
New Zealand ◽  
Preliminary Analysis ◽  
18S Rrna ◽  
18S Rrna Gene ◽  
Coi Gene ◽  
Taxonomic Resolution ◽  
Gut Contents ◽  
Rrna Gene ◽  
Diverse Range ◽  
Dna Metabarcoding

Deep sea lobsters are highly valued for seafood and provide the basis of important commercial fisheries in many parts of the world. Despite their economic significance, relatively little is known about their natural diets. Microscopic analyses of foregut content in some species have suffered from low taxonomic resolution, with many of the dietary items difficult to reliably identify as their tissue is easily digested. DNA metabarcoding has the potential to provide greater taxonomic resolution of the diet of the New Zealand scampi (Metanephrops challengeri) through the identification of gut contents, but a number of methodological concerns need to be overcome first to ensure optimum DNA metabarcoding results. In this study, a range of methodological parameters were tested to determine the optimum protocols for DNA metabarcoding, and provide a first view of M. challengeri diet. Several PCR protocols were tested, using two universal primer pairs targeting the 18S rRNA and COI genes, on DNA extracted from both frozen and ethanol preserved samples for both foregut and hindgut digesta. The selection of appropriate DNA polymerases, buffers and methods for reducing PCR inhibitors (including the use of BSA) were found to be critical. Amplification from frozen or ethanol preserved gut contents appeared similarly dependable, but metabarcoding outcomes indicated that the ethanol samples produced better results from the COI gene. The COI gene was found to be more effective than 18S rRNA gene for identifying large eukaryotic taxa from the digesta, however, it was less successfully amplified. The 18S rRNA gene was more easily amplified, but identified mostly smaller marine organisms such as plankton and parasites. This preliminary analysis of the diet of M. challengeri identified a range of species (13,541 reads identified as diet), which included the ghost shark (Hydrolagus novaezealandiae), silver warehou (Seriolella punctate), tall sea pen (Funiculina quadrangularis) and the salp (Ihlea racovitza), suggesting that they have a varied diet, with a high reliance on scavenging a diverse range of pelagic and benthic species from the seafloor.

Notes on the molecular taxonomy of the Proclossiana eunomia complex (Lepidoptera, Nymphalidae: Argynnini): analysis of DNA barcodes

2018 ◽  
Vol 8 (1) ◽  
pp. 222-232 ◽  
Author(s):  
R. V. Yakovlev ◽  
N. A. Shapoval ◽  
G. N. Kuftina ◽  
A. V. Kulak ◽  
S. V. Kovalev
Keyword(s):  
Body Size ◽  
Molecular Taxonomy ◽  
Taxonomic Status ◽  
Molecular Data ◽  
Coi Gene ◽  
Wing Pattern ◽  
Molecular Approach ◽  
Dna Barcodes ◽  
Morphological Differences ◽  
Proclossiana Eunomia

The Proclossiana eunomia (Esper, 1799) complex is currently composed of the several subspecies distributed throughout Palaearсtic region and North America. Despite the fact that some of the taxa have differences in wing pattern and body size, previous assumptions on taxonomy not supported by molecular data. Therefore, the identity of certain populations of this complex has remained unclear and the taxonomic status of several recently described taxa is debated. Here, we provide insights into systematics of some Palaearctic members of this group using molecular approach, based on the analysis of the barcoding fragment of the COI gene taking into account known morphological differences.

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