DNA barcoding of Congolese freshwater fish collections
Erik VerheyenF. BremanCélestin Danadu MizaniM. HanssensA. Ibala-ZambaZoltán NagyGontran SonetJeroen Van HoudtEmmanuel VrevenSoleil Lunkayilakio WamuiniJos Snoeks
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The inland fisheries in tropical Africa face threats both by stress from climate change and by overexploitation. Species are becoming extinct and populations decline at an alarming but poorly understood rate. Many species may face extinction before they can be identified or described. This presents a problem for conservation planning and prioritization, because those species that have not been identified obviously cannot be protected effectively. Caddy and Garibaldi reported that only 65.09% of worldwide fishery captures reported to the FAO for the year 1996 was identified at species level, ranging from about 90% in temperate areas to less than 40% in tropical regions. Surveys into the accuracies of species identifications have not been reported, but a significant percentage of identifications may still be erroneous. The limitations inherent in morphology-based identification systems and the limited pool of taxonomists paved the way for the introduction of new molecular diagnostic tools for effective species identification. Hitherto, a wide variety of protein-based and DNA-based methods have been evaluated for the molecular identification of fish species in Africa. These studies, however, are not comparable for the purposes of species identification because they lack standardization (e.g. different regions of the mitochondrial genome such as cytochrome b and 16S rDNA were used). Hebert et al. (2003) proposed a single gene sequence to discriminate the vast majority of animal species, using a 650-bp fragment of the 5′ end of the mitochondrial cytochrome c oxidase subunit I (COI) gene as a global bioidentification sequence for animals. This technology (DNA barcoding) relies on the observation that the ‘barcode’ sequence divergence within species is typically much lower than the divergence exhibited between species, making it an effective marker for species identification and discovery. The startling efficiency of the method may arise from selective sweeps and the intricacies of mito-nuclear coadaptation, raising the profile of bioenergetics as a possible speciation mechanism.
DNA barcoding has since gained global support as a rapid, accurate, cost-effective, and broadly applicable tool for species identification, particularly with respect to fishes as coordinated by the fish barcode of life campaign. Barcoding has also been adopted by the census of marine life project, a growing global conglomerate of 50 countries engaged in a 10-year initiative to assess and explain the diversity, distribution, and abundance of life in the ocean. Although there has been criticism of both the philosophical and the practical underpinnings of DNA barcoding, its successful application for both species identification and discovery has been demonstrated in many studies, involving many taxonomic groups, for example birds, fish, fish parasites, bats, spiders, crustaceans, nematodes, earthworms, mosquitoes, and diverse arrays of Lepidoptera. In addition, DNA barcoding strategies are now being applied for other groups of organisms including plants, macroalgae, fungi, protists, and bacteria. Furthermore, DNA barcoding has gained wide application in forensic analysis to investigate cases of illegal poaching, separation of species, gut content analysis in ecological studies, food product analysis and market substitution, and Asian medicine trade regulation. DNA barcoding has also been employed to validate the identity of biomaterial collections and cell lines. A sufficient accumulation of DNA barcodes can also help conservation managers to identify interim priority areas for conservation efforts in the absence of species data. Currently, DNA barcode reference library records are available for more than 1 million sequences representing more than 94,000 species on the barcode of life data systems, an informatics workbench aiding the acquisition, storage, analysis, and publication of DNA barcode records. Nearly, 10% of these records comprise marine and freshwater fish species.
The COI divergence and species identification success based on DNA barcodes have been previously assessed for many freshwater fish species, for example in Canada, Mexico and Guatemala, and Brazil. Since, to date, there is no detailed knowledge about the diversity and distribution of freshwater fish species in Nigeria, the aim of this study was to determine whether DNA barcoding can be used as an effective tool to perform unambiguous species identification of freshwater fishes in this region, with a view toward establishing a DNA barcode reference library for utilization in biodiversity assessment and conservation for the entire country.
Fish is a proteinous animal which plays a vital role in the protection and prevention of human diseases. DNA barcoding which uses the 50 region of the mitochondrial cytochrome c oxidase subunit as the target gene was implored as an efficient tool in the identification of fish species in the Enugu Metropolis River (Nike Lake and Abakpa River). 10-20mg fish tissue sample of 18 species were extracted for DNA using Promega kit. The polymerase chain reaction (PCR) was used to amplify short sequences of mitochondrial DNA, which were denatured and analysed by polyacrylamide gel electrophoresis (native PAGE), for detection of single strand conformation. Polymorphism species specific muscle alignment patterns of DNA bands were obtained for Chrischthys sp, Parachinna sp, Ctenopoma sp, Tilapia sp and for a number of Clarias species. Out of the 18 fish species, only 15 fish samples were analysed using their genomic make-up, 4 out of the 15 samples (Parachinna obscura -2) (Clarias sp -2) did not show statistical significant evidence of spatial genetic differentiation in their nucleotides despite the enormous geographical distance separating populations. The morphological studies on this fish species have shown that these lines of evidence are taxonomically important and also partial differences in genomic nucleotide base pairs when noticed. This difference is the polymorphism, which is the key to flagging new specie in a particular genus can be attributed to environmental changes and diversity.
DNA Barcoding
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Overexploitation
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DNA Barcoding
Fresh water fish
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This study represents the comprehensive molecular identification of freshwater fishes of Bangladesh based on a fragment of the cytochrome c oxidase subunit I (COI) gene in the mitochondrial genome. A total of 315 mitochondrial COI barcode sequences were obtained from 153 species,114 genera, 49 families and 16 orders of fishes. The mean length of the sequences was 652 base pairs. For all the samples, %G was significantly lower compared to the other nucleotides and %GC was lower compared to %AT (p-value ˂ 0.05). Also, a significantly lower %GC content was observed in second and third codon position compared to the first one in all the samples (1st>2nd>3rd, p-value˂ 0.05). The average K2P distances within species, genera, families and orders were 0.38%, 7.02%, 12.75% and 18.68%, respectively. The mean interspecific distance was 18-fold higher than the mean intraspecific distance. The K2P neighbor-joining (NJ) trees based on the sequences generally clustered species according to their taxonomic position. A total of 12 species were newly recorded in Bangladesh. High efficiency in species identification were demonstrated in the present study by DNA barcoding, and concluded that COI sequencing can be used as an authentic identification marker for freshwater fish species.
Bangladesh J. Zool. 48(1): 1-19, 2020
DNA Barcoding
Nucleotide diversity
Fresh water fish
Molecular marker
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DNA Barcoding
Marine fish
Identification
Species Identification
Marine species
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Barcode
DNA Barcoding
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Nomenclature for the nameless is one of the successful events after the invention of the DNA barcoding technique in biodiversity research. The ornamental fish species are frequently hunted from the east and northeast India and trafficked with various elusive names and high demanding values. Such illegitimate trading stimulated the severe threats on the native freshwater ecosystems and their indigenous biodiversity. Both traditional taxonomy and DNA barcoding technique successfully identified 11 ornamental fish species from a small riverine system, Murti river; linked up with three protected areas in the northern part of West Bengal. To test the efficacy of DNA barcode data for species identification, the generated sequences were subjected to similarity search results, and Neighbour-Joining tree clustering and genetic divergences. The mean genetic divergence was 21.3% and the interspecific genetic distance was ranging from 17.8% to 28.7% in the studied dataset. The detected high intraspecific genetic divergence in Opsarius barna (14.1%) and Channa gachua (6.4%) in the present dataset suggested further genetic investigation from their known distributions.
DNA Barcoding
Genetic divergence
Ornamental plant
Fresh water fish
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DNA-based identification methods have been shown to have high detection capability and reduced costs compared to traditional methods and can also enable the detection of species that might be missed using traditional methods (e.g. rare species, cryptic species, larval stages). The success of DNA-based identification is dependent on the ‘DNA barcodes’ of target species being present in a barcode reference database. In order to use DNA-based identification methods to assess and monitor UK freshwater arthropods for biodiversity and ecological quality assessments, it is vital that comprehensive reference databases are available. Incomplete reference databases result in many sequences derived from metabarcoding not being assigned to species. Two current projects aim to create collections of high-quality sequences from expertly identified specimens of UK species. The Darwin Tree of Life project aims to sequence the genomes of all the eukaryotic species in Britain and Ireland and FreshBase aims to create a genomic reference collection for UK freshwater invertebrates. The Barcode of Life Data System (BOLD) is one of the main reference databases for animal barcodes. Prioritising the sequencing of UK freshwater arthropod species that are not yet represented in BOLD, would enable more complete identification of UK freshwater biodiversity using metabarcoding and would enable the development of primers to target specific arthropod groups or species. We analysed the coverage of UK freshwater arthropod species in BOLD. Our analyses show that coverage varies between taxonomic groups and large proportions of sequences in some orders are only represented by privately stored sequences in BOLD. Analyses of intra- and inter-specific variation in sequences stored in BOLD show that misidentifications or errors can reduce the barcode gap in some species which could cause difficulties in accurately identifying sequences derived from metabarcoding. Representation in BOLD by specimens from the UK is extremely low and analyses show that high geographic variation in sequences in some species could be important for accurate DNA-based identification of UK species. Our results have implications for prioritising the sequencing of UK freshwater arthropods and for the quality control of stored sequences in order to reduce the occurrence of misidentifications and errors that could impact the accuracy of DNA-based identification.
DNA Barcoding
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Arthropod
Identification
Environmental DNA
Taxonomic rank
Species complex
Freshwater ecosystem
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Barcode
DNA Barcoding
Marine fish
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