Identification Of Seafood Species Research Articles

Electrophoretic Methods for Identifying the Species of Seafood and Its Derivatives

The identification of the species of seafood and their products, whether they are fresh or cooked, is one of the key concerns of food regulations in many countries that have a significant intake of seafood. In point of fact, a commercial fraud happens when a species that is less value is intentionally substituted for a species that is more valuable, and a sanitary fraud takes place when a product that is potentially harmful is introduced into the market. A primary responsibility of veterinary inspection of seafood products is the detection of harmful species with the aim of removing them from the retail trade (Council Directive 91/493/ECC). Two efficient methods for seafood species identification are molecular biology methods and protein electrophoresis. Classic electrophoretic methods have been used for a long time to authenticate seafood; they are simple, accurate, and inexpensive compared to molecular biological methods, which are the wave of the future in food safety labs. The purpose of this article is to offer an overview of the electrophoretic methods commonly used to identify different species of seafood, including sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE), native or urea-isoelectric focusing electrophoresis (IEF), two-dimensional electrophoresis (2-DE), and capillary electrophoresis (CE).

DNA Barcoding of Commercial Three High Value Red Sea Fishes from the Egyptian Market

Fish play an essential role in food security and are regarded as one of Egypt's primary food resources and, therefore, significantly im-pact the national economy. The three main sources of fish production in Egypt: marine (Red and Mediterranean seas), lakes and the River Nile, due to the elevated incidence of species substitution at the global level, precise identification of seafood species on the mar-kets considered a solution for food safety con-trol institutions and human consumer protec-tion. Mislabeling occurs when one species is substituted for another. Different species of red sea marine fish were collected from the Egyp-tian market. According to the Arabic name in the fish market, Samples searched for English and Latin family names in (www.fishdata-base.org). Commercial samples were success-fully extracted and confirmed with 1.5% aga-rose gel electrophoresis. The result showed that the mtDNA gene cytochrome c oxidase subunit I (COI) amplification was successful for all samples with high concentration, it was purified and sequenced for both directions and blasted using the NCBI database. Resulting in the identification of three commercially essen-tial and expensive fish samples, Baghbaghan (parrotfish), Hamour (Greasy Grouper) and kahaya (Spiny squirrelfish). One of them cor-rectly identified according to the market name, while the other two are recorded mistakenly under another name.

Application of DNA barcoding and metabarcoding for species identification in salmon products

ABSTRACT Mislabelling is a significant manifestation of food fraud. Traditional Sanger sequencing technology is the gold standard for seafood species identification. However, this method is not suitable for analysing processed samples that may contain more than one species. This study tested the feasibility of next-generation sequencing in identifying mixed salmon products. Salmon samples containing up to eight species were amplified using 16S rRNA mini-barcode primers, and sequenced on an Illumina HiSeq2500 platform. All species were accurately identified, and mixtures as low as 1% (w/w) could be detected. Furthermore, this study conducted a market survey of 32 products labelled as salmon. For pure and mixed fish products, Sanger and next-generation sequencing techniques were respectively used for species identification, and for NGS results, we also used real-time PCR method to cross-validate the mixed products to further verify the accuracy of the DNA metabarcoding technology established in this study. DNA barcoding and metabarcoding of commercial salmon food products revealed the presence of mislabelling in 16 of 32 (50%) samples. The developed DNA barcoding and metabarcoding methods are useful for the identification of salmon species in food and can be used for quality control of various types of salmon products.

Molecular identification of dried squid products sold in China using DNA barcoding and SYBR green real time PCR

ABSTRACT Dried squid products are popular in China as a snack food, side dishes, or refreshments, and the market appeal can be reflected by the high price that occasionally reaches 497 RMB per kg. However, the absence of harmonisation around the definition of squid, as well as the problems with visual inspection for processed seafood products, make alternative species substitution for dried squid products a frequent occurrence. The aim of the present study was to apply a DNA barcoding approach for species identification of 48 dried squid products collected from the largest online shopping platform in China. Moreover, we also developed a novel SYBR green real-time PCR assay (simplex and duplex followed by a melting curve analysis) specific for Illex argentinus and Todarodes pacificus based on cytochrome C oxidase subunit I (COI) gene. Results highlighted the successful DNA extraction and PCR amplification of a 655 bp COI gene fragment from all products. A maximum similarity value in the range of 98-100% was obtained for all readable sequences using the BOLD and BLAST public databases and four species (Dosidicus gigas, Uroteuthis edulis, I. argentinus, and T. pacificus) were identified. The specificity of the designed primer sets was confirmed against 23 non-target species, and the newly developed methods were successfully applied to screen I. argentinus and T. pacificus in dried squid products. Overall, DNA barcoding is a robust tool for seafood species identification and the novel method is effective in screening I. argentinus and T. pacificus in food products.

Occurrence of Microplastics in Commercial Seafood under the Perspective of the Human Food Chain. A Review.

The occurrence of microplastics in the marine ecosystem and aquatic organisms, their trophic transfer along the food web, and the identification of seafood species as suitable indicators have become a research priority. Despite the high quantity of research in this field, a comparison between the available data and an appropriate risk assessment remains difficult. In this perspective, as an innovative approach, the association of the feeding strategies of commercial seafood and the microplastic level was considered. Further research to assess the occurrence of microplastics in the marine food web, the long-term effects on animals and humans, and the health implications is needed.

Comparative Evaluation of Web Page and Label Presentation for Imported Seafood Products Sold on Chinese E-Commerce Platform and Molecular Identification Using DNA Barcoding

With the expansion of e-commerce, an increasing number of Chinese consumers are turning to online markets to purchase foreign seafood. When buying seafood online, customers cannot physically evaluate the product, and the market Web page instead of the seafood label conveys all of the product information. However, specific regulations concerning the information presented on the Web page have not been created, which may foster seafood fraud and misdescription. Because mislabeling of seafood has become a widely reported issue in the Chinese offline market, the online scenario must be investigated comprehensively. This study focused on various seafood products that originated from 20 countries and were sold by one of the largest e-commerce companies in China. For each country, only the product with the greatest overall monthly transaction volume was selected, and 5 samples were purchased per product for a total of 100 samples. The Web page description (including the heading of the Web page and the description of the commodity) and the label of the received products were compared to evaluate the description consistency. DNA barcoding technology was used for seafood species identification, and the scientific names retrieved from the sequence analysis after consulting the Barcode of Life Data systems and GenBank were compared with the expected species, genus, and family to determine the description authenticity. Only 25% of the samples had consistent descriptions on the Web page and on the label of the received product. Most of the inconsistency originated from the geographical origin, and only four products (G10, G50, G19, and G69) had inconsistent species, genus, and family descriptions. Molecular analysis revealed that in 65% of samples the species was correctly described. The online seafood market presents challenges regarding seafood fraud and opportunities for seafood species substitution.

Multi-instrument Evaluation of a Real-time PCR Assay for Identification of Atlantic Salmon: a Case Study on the Use of a Pre-packaged Kit for Rapid Seafood Species Identification

Protecting the seafood supply chain from species substitution is critical for economic, health, and conservation reasons. DNA-based methods represent an effective means to detect species substitution, but current methods can be time consuming or costly, and require specialized instruments and operators. Real-time PCR provides an alternative that can be performed quickly, and in some cases even on-site. The use of commercial kits reduces the expertise required by the operator and therefore increases accessibility to testing. This potentially increases the likelihood of adoption into the supply chain, but only if the kits are robust across multiple operators, instruments, and samples. In this study, the InstantID™ Atlantic salmon kits were tested on a variety of instruments with market samples of fresh, frozen, smoked, and canned Atlantic salmon. Results were repeatable across all samples and instruments tested. This kit, and others like it that have undergone appropriate evaluation, represents a means for expanded access to testing for industry or regulators to screen seafood for species authenticity. Portable equipment can bring testing on-site, further reducing analysis time.

Mini DNA-barcode as molecular marker for heavily processed hairtail fish products authentication

Fish species mislabeling, wrongful description, and substitution by species with a lower value are problems that occur due to globalization of fish markets. Establishment of fishery products and seafood species identification methods becomes an important task for Indonesian fishery research laboratories and seafood control authorities. Therefore, various DNA barcode markers must be tested to determine which markers are suitable for rapid, simple, and applicable methods. This study aimed to determine the reliability of full-length DNA barcode and mini DNA-barcodes for raw fillets and numerous cooked Indonesian hairtails. The results showed all DNA barcode markers were successfully applied to all of the samples with the species level assignment of 97-100%. Our study proposed a practical molecular marker for the monitoring of quality and food safety assurance of important hairtail products.

DNA barcoding of fish species reveals low rate of package mislabeling in Qatar.

DNA barcoding technique has made it possible to authenticate various species used for food and medicinal purposes. In the identification of seafood species, studies are concentrated in North America, Europe, and Asia. Elsewhere, including countries in the Middle East and North Africa, studies of this sort are scarce. This study focuses on packaged fresh or minimally processed fish fillet available at eight major supermarket chains in Qatar. A cocktail of eight primers attached with M13 tails established for fish species identification was adopted to facilitate PCR and sequencing. Sequences were compared with those available in the Barcode of Life Databases (BOLD Systems) and BLAST in NCBI databases. Among the 62 unique fish packages with resolved sequences, only three are confirmed to be mislabeled, at a rate of about 5%. Two of the substituted species are high value items while the third species was replaced by another, equally low-cost species. The relatively low rate of mislabeling in the samples is perhaps a result of strict local food safety regulations, which may have led to high consistency between the package labels and their contents.

DNA barcoding for the verification of supplier's compliance in the seafood chain: How the lab can support companies in ensuring traceability.

Food Business Operators (FBOs) rely on laboratory analysis to ensure seafood traceability. DNA barcoding and Forensically Informative Nucleotide Sequencing may represent a support within self-checking programs finalized to suppliers’ qualification and products identity certification. The present study aimed at verifying the usefulness of a decisional procedure (decision tree) set up at the FishLab (Department of Veterinary Sciences, University of Pisa, Italy) for seafood species identification by DNA analysis, to cope with FBOs’ needs. The decision tree was applied to the analysis of 182 seafood (fish and molluscs) products, conferred to the FishLab by different FBOs between 2014 and 2015 as result of their self-checking activities. The analysis relied on a standard COI gene fragment eventually integrated by the analysis of alternative or supportive molecular targets (cytb and 16S rRNA). It also included a mini-DNA barcoding approach for processed products. Overall, 96.2% of the samples were unambiguously identified at species level using the elective target alone (92.4%) or a multi-target approach (3.8%). The lack of species identification (3.8%) was attributable to the absence of reference sequences or to the low resolution of the molecular targets. Nonetheless, all the molecular results were deemed adequate to evaluate the sample’s compliance to the label information. Non-compliances were highlighted in 18.1% of the products. The protocol was proven as an effective supportive tool for the seafood identity verification within the supply chain self-checking activities. In addition, a considerable fraud rate was confirmed and the species most frequently involved in substitution were pointed out.

Rapid Seafood Species Identification Using Chip-Based Capillary Electrophoresis and Protein Pattern Matching.

Authenticity is crucial to the seafood industry, as substitution and mislabeling have important economic, environmental, and food safety consequences. To address this problem, protein profiling and software algorithm techniques were developed to classify fish muscle samples by species. The method uses water-based protein extraction, chip-based microfluidic electrophoresis (Agilent 2100 Bioanalyzer) for the analysis of high abundance fish muscle proteins, and a novel data analysis method for species-specific protein pattern recognition. The method's performance in distinguishing commercially important fish from commonly reported substitutions was evaluated using sensitivity, specificity, and accuracy determinations with all three performance measures at >98% for common substitutions. This study demonstrates that uncooked seafood products of commercially important species of catfish, snapper, and grouper can be rapidly distinguished from commonly substituted species with a high level of confidence. A tiered testing approach to seafood species verification by sequentially applying a rapid screening method and DNA testing is proposed to more effectively ensure accurate product labeling.

DNA barcoding reveals a high level of mislabeling in Egyptian fish fillets

Accurate identification of seafood species in the markets is a growing concern due to the high incidence of species substitution at international level. It is a prime priority for governments to be able to identify the already processed fish products (fish fillets). In this context, DNA barcoding was applied to ascertaining species in fish fillets (tilapia, Nile perch and panga) purchased from Egyptian markets. Ninety commercial samples were analyzed. Sequencing of a short fragment of mitochondrial cytochrome oxidase I (COI) gene revealed 33.3% species substitution in the fish fillets analyzed, 50% Nile perch (Lates niloticus) and 50% basa fish (Pangasius bocourti) being replaced by imported Vietnamese tra fish (Pangasianodon hypophthalmus). These results demonstrate that DNA barcoding is a reliable tool for detecting fish products adulteration in Egypt. We recommend its use for control and law enforcement of seafood quality.

Genetic diversity of ivory shell (Babylonia areolata) in Taiwan and identification of species using DNA-based assays

For better traceability of seafood products in Taiwan, we need to effectively test product quality during the processes of identification of seafood species. The aim of this study was to analyse gene diversity and the methods of identification of high-value seafood, ivory shell (Babylonia areolata), on the Penghu Island in Taiwan using molecular marker technology and to build a relevant molecular database. Thirty-three samples of B. areolata and the samples of 5 other Babylonia species, including Babylonia feicheni, Babylonia spirata spirata, Babylonia perforata and Babylonia formosae formosae, from cultivation and from the wild were tested using the inter-simple sequence repeat (ISSR) method, mitochondrial DNA analysis, PCR-single-strand conformation polymorphism (PCR-SSCP) and PCR-denaturing gradient gel electrophoresis (PCR-DGGE). The results showed that the primers ISSR3, ISSR7 and ISSR13 of the ISSR method and cytochrome C oxidase subunit 1 (coxI) gene analysis have a good discriminatory power in inter-species and intra-species tests. In conclusion, ISSR, PCR-DGGE and PCR-SSCP with coxI analysis can be used for the screening and identification of B. areolata species. Furthermore, these molecular methods could be useful for the identification of other types of seafood.

Food authentication of commercially‐relevant shrimp and prawn species: From classical methods to Foodomics

Although seafood species identification has traditionally relied on morphological analysis, sometimes this is difficult to apply for the differentiation among penaeid shrimps owing to their phenotypic similarities and to the frequent removal of external carapace during processing. The objective of this review is to provide an updated and extensive overview on the molecular methods for shrimp and prawn species authentication, in which several omics approaches based on protein and DNA analysis are described. DNA-based methods include the amplification by PCR of different genes, commonly the mitochondrial 16S ribosomal RNA and cytochrome oxidase I genes. A recently described method based on RFLP coupled to PCR turned out to be particularly interesting for species differentiation and origin identification. Protein analysis methods for the characterization and detection of species-specific peptides are also summarized, emphasizing some novel proteomics-based approaches, such as phyloproteomics, peptide fragmentation, and species-specific peptide detection by HPLC coupled to multiple reaction monitoring (MRM) MS, the latter representing the fastest method described to date for species authentication in food.

Advances in DNA-Based Techniques for the Detection of Seafood Species Substitution on the Commercial Market

Increased worldwide trade and processing of seafood has increased the potential for species substitution on the commercial market. To detect and prevent species substitution, several deoxyribonucleic acid (DNA)-based methods have been developed that can be used to identify species in a variety of food types. For large-scale applications, such as regulatory screening, these methods must be rapid, cost-effective, reliable, and have high potential for automation. This review highlights recent technological advances in DNA-based identification methods, with a focus on seafood species identification in automated, high-throughput settings. Advances in DNA isolation methods include silica-based columns for use in high-throughput operations and magnetic bead particles for increased and targeted recovery of DNA. The three most widely used methods for seafood species identification (polymerase chain reaction [PCR] sequencing, PCR-restriction fragment length polymorphism, and species-specific PCR) will be discussed, with a focus on the incorporation of technologies such as rapid PCR cycling, microfluidic chips, and real-time PCR. Emerging methods, including DNA microarrays and next-generation sequencing will also be explored for their potential to identify seafood species on a large scale. Overall, many of the technological advances discussed here offer complementary properties that will enable species identification in a variety of settings and with a range of products.

Selected tandem mass spectrometry ion monitoring for the fast identification of seafood species

Selected tandem mass spectrometry (MS/MS) ion monitoring (SMIM) is the most suitable scanning mode to detect known peptides in complex samples when an ion-trap mass spectrometer is the instrument used for the analysis. In this mode, the MS detector is programmed to perform continuous MS/MS scans on one or more selected precursors, either during a selected time interval, or along the whole chromatographic run. MS/MS spectra are recorded, so virtual multiple reaction monitoring chromatogram traces for the different fragment ions can be plotted. In this work, a shotgun proteomics approach was applied to the detection of previously characterized species-specific peptides from different seafood species. The proposed methodology makes use of high intensity focused ultrasound-assisted trypsin digestion for ultra fast sample preparation, peptide separation and identification by reverse phase capillary LC coupled to an ion-trap working in the SMIM scanning mode. This methodology was applied to the differential classification of seven commercial, closely related, species of Decapoda shrimps proving to be an excellent tool for seafood product authentication, which may be used by fisheries and manufacturers to provide a fast and effective identification of the specimens, guaranteeing the quality and safety of foodstuffs to consumers.

Occurrence of palytoxin-group toxins in seafood and future strategies to complement the present state of the art

Palytoxin (PlTX) and palytoxin-like (PlTX-like) compounds in seafood have been raising scientific concern in the last years. The constant increase in record numbers of the causative dinoflagellates of the genus Ostreopsis together with the large spatial expansion of this genus has led to intensification of research towards optimization of methods for determination of PlTX presence and toxicity. In this context, identification of seafood species which could possibly contain PlTXs constitutes an important issue for public health protection. In the present paper, worldwide occurrence of PlTX-like compounds in seafood is reviewed, while potential future strategies are discussed. PlTX has been reported to be present in several species of fish, crustaceans, molluscs and echinoderms. In one occasion, PlTX has been identified in freshwater puffer fish whereas all other records of PlTXs refer to marine species and have been recorded in latitudes approximately between 43°N and 15°S. PlTX determination in seafood has relied on different methodologies (mainly LC-MS, mouse bioassay and hemolysis neutralization assay) that have evolved over time. Future recommendations include systematic screening of PlTX in those species and areas where PlTX has already been recorded implementing updated methodologies.

DNA-based methods for identification of seafood species

With the development of molecular biotechnology, methods for identification of seafood species are developed from protein to DNA. At present, the main DNA-based methods for species identification are FINS, PCR-RFLP, and specific-PCR, which have been used to identify the species of fresh, frozen, and pickled or canned seafood. However, qualitative and quantitative methods for identification of the mixed seafood species remain to be resolved. The gene databases play an important role in identifying species and are valuable information resources for identification of seafood species. In this paper, recent progresses of major DNA-based methods for identification of seafood species are reviewed and the perspectives of this field are discussed.