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12309451865?profile=RESIZE_710xThe modern food industry is a rapidly evolving sector with complex supply chains that utilises a wide variety of analytical tools to support food integrity and authenticity.

Next Generation Sequencing (NGS)-based approaches represent an emerging analytical technology that is growing within the food sector, providing  the potential to not only screen and test input materials, but also the production process and end products. 

Photo by Braňo on Unsplash

This questionnaire is part of a UK Department for Environment, Food and Rural Affairs (Defra) funded project tasked with reviewing the application of NGS technology to food authenticity testing and supporting harmonisation and the development of standards for NGS approaches applicable to the verification of food authenticity. The questionnaire is targeted at individuals involved in the food and associated diagnostics sectors, including technology developers, suppliers and official controls.

Your participation in this questionnaire will directly help inform the direction of the project and contribute to guidance within the sector.

Please do not provide any information that could be used to identify you. Thank you for participating in our questionnaire which should only take around 15 minutes to complete. Your feedback is important.

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We would be very grateful if you could complete the questionnaire by 5 January 2024.
Thank you for participating in this questionnaire.

UK National Measurement Laboratory at LGC

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Traditional cheeses, with protected names (PDO/PGI) are sold at a premium and hence can be susceptible to substitution and fraud. This review reports the available authentication methods, either chemical, physical, or DNA-based methods, currently used for origin authentication, highlighting their principle, reported application to cheese geographical origin authentication, performance, and respective advantages and limits. Isotope and elemental fingerprinting showed consistent accuracy in origin authentication. Other chemical and physical methods, such as near-infrared spectroscopy and NMR, require more studies and larger sampling to assess their discriminative power. Emerging DNA-based methods, such as metabarcoding, showed good potential for origin authentication. However, metagenomics, providing a more in-depth view of the cheese microbiota (up to the strain level) is an interesting approach to authenticate the origin of cheeses, but is still in its early development.

Read the full open access paper.

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Oregano has been identified as one of the herbs most susceptible to adulteration. Methods based on DNA, spectroscopic analysis and even microscopy have already been used. In this paper, a new approach for authentication of oregano, which combines metabarcoding by NGS (next generation sequencing) and metabolomics/chemometrics by NMR, has been developed. The industry standard for oregano permits only 2% extraneous matter. A previous survey on oregano has shown that the most common plant adulterants are olive, sweet marjoram and myrtle leaves, and non-leaf plant material. In this study, 92 oregano, 38 sweet marjoram, and 2 olive leaf samples from 6 different countries in total were used. Metabarcoding by NGS was used to identify the nature of oregano products and possible adulterations. Metabolomic profiles obtained by NMR correlated well with oregano species and their regional origin. Using chemometric analysis, it was possible to quantify of the percentage of an adulterant with error rates of 3–7%.

Read the open access paper here

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In this study, the Thermo Scientific™ NGS Food Authenticity Workflow was used to analyse spices and herbs.  

Reference samples were analysed to verify the specificity, and spikings down to 1% (w/w) allowed verification of its sensitivity including in complex mixtures of five different spices and/or herbs. 

272 commercial samples were collected in Asian and European markets.  78% of the commercial samples were compliant with the declared content, whereas the rest were shown to contain undeclared species that were in a few cases allergenic or potentially toxic. 

The researchers conclude that the overall workflow is user-friendly and straightforward, which makes it simple to use and facilitates data interpretation.

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 Photo by Ratul Ghosh on Unsplash











Photo by Alice Pasqual on Unsplash

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Herbs are high value ingredients, which are vulnerable to adulteration and fraud. Confirmatory methods based on DNA analysis have shown to be the most useful in investigating herb adulteration. In this study, a customised database and bioinformatics pipeline was developed based on a DNA barcoding metagenomics approach to herbal species identification. The pipeline performance was tested with publicly available datasets, as well as, newly sequenced herbal plants and products. The usefulness of metagenomics is limited by the availability of reference sequences and the need for sequencing depth.  However, this method shows promise for evaluating the authenticity of different herbal products provided that it is further refined to increase the qualitative and quantitative accuracy.

You can read tihe full open access paper

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This project (FA0160)  builds on two previous feasibility studies, and develops the principle of using a food microbiome as an authenticity marker for the development of methodology that discriminates provenance on the basis of its microbial fingerprint. It also exploits the latest NGS (Next Generation Sequencing) technology, which permits a large number of microbiological groups to be identified to create the microbial profile. Further details of the project and its results were published in our October 2021 Newsletter. The final report is available on the Defra (Departmnt of the Environment, Food and Rural Affairs) website, and the link has also been added to the website's list of research reports.

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9549834470?profile=RESIZE_400xForeword by the Government Chemist

Next Generation Sequencing (NGS) is a powerful tool for rapidly and cost-effectively identifying and characterising plant, animal and microbial species present in mixed food samples.

The application of NGS to food authenticity, adulteration and safety testing is a constantly evolving field with its own unique set of challenges that need to be explored. Further work needs to be conducted to better understand the performance characteristics and establish relevant performance criteria and metrics, to enable results generated in different laboratories to be compared and interpreted with equal confidence.

Following concerns raised from food industry members on the use of NGS for the quantitative determination of food ingredients, the Government Chemist engaged with Defra’s Authenticity Methodology Working Group (AMWG) [1] and its Technical Sub-Group (AMWG-TSG), resulting in the AMWG producing a view [2] on the use of NGS for food authenticity testing [3].

Download Defra’s Authenticity Methodology Working Group’s view on the use of Next Generation Sequencing for food authenticity testing

[1] An independent expert group that provides scientific and technical advice to support Defra’s food authenticity programme.

[2] The views/opinions expressed by AMWG were correct at the time of the note (November 2020).

[3] Government Chemist representatives: Selvarani Elahi, Deputy Government Chemist, is the Chair of AMWG and Dr Malcolm Burns, Head of GMO unit, Principal Scientist and Special Advisor to the Government Chemist, is a Member of AMWG; they both participated in the AMWG-TSG meeting on NGS and subsequent discussions, inputting into the AMWG view on NGS.

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9242579475?profile=RESIZE_400xHerbs and spices are vulnerable to adulteration, and the problem has only been compounded by the Covid pandemic. However, NGS is giving food manufacturers, retailers, and regulators, the tools they need to help combat on food fraud. Rather than returning a simple positive/ negative result, NGS reliably detects and reports multi-species DNA in even the most complex of samples, including herbs and spices, which allows food analyst laboratories to screen for thousands of species in one test, and get same-day results.

Read the article here

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A Ph.D thesis from the University of Milan-Bicocca in English is publicly available. The thesis gives a good overview of DNA barcoding, NGS (Next Generation Sequencing) and metabarcoding, and isothermal nucleic acid amplification. The research carried out looked at applying DNA barcoding to processed foods, which required smaller DNA fragments. However, the approach is not suitable for mixed species samples, and NGS and metabarcoding approaches were more successful. Finally, an isothermal amplification assay was applied to authenticate truffles.

You can read the 253 page thesis here   

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Sanger sequencing (DNA barcoding) is a robust method for species identification. However, it is not always suitable for species identification of processd mixed species products. Chinese researchers have developed an NGS method based on the amplification and sequencing of shorter 16S rRNA DNA sequences. The assay was developed using a mixture of 8 salmon species, which were all correctly identified even when the species was presented as low as 1%(w/w). It was tested with a market survey of 32 commercial salmon products. Sanger sequencing was used on single species unprocessed products and NGS on mixed species products, which was also cross validated with a real-time PCR assay. The survey revealed that 50% of the samples were mislabelled.

Read the abstract here

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This article in New Food Magazine discusses how Next Generation Sequencing (NGS) and Whole Gene Sequencing (WGS) can assist in detecting and identifying contamination, and also playing an important role in assuring traceability of food products when combined with blockchain along the supply chain. Technology improvements have meant that NGS and WGS have high throughputs at much lower cost than before, and NGS machine can now be used for WGS as well.

The genomic information derived by these techniques on pathogenic bacterial contamination when combined with data such as the date and place of findings, can help track down the exact sources of contamination and therefore avoid large scale recalls of food products. The role of NGS in obtaining DNA traceability combined with blockchain permits products all the way along the supply chain to be traced back to their original raw materials whether that be plants or animals. 

Read the article here

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4439874075?profile=RESIZE_400xA novel method to identify animal species in complex or adulterated processed meat products has been developed by Chinese researchers, which combines a cytochrome oxidase I (COI) mini-barcode with next-generation sequencing (NGS).  A universal primer based on 140 sequences from 51 edible animal species was designed. A mixture of 12 species raw meat samples (beef, water buffalo, pork, sheep, chicken, partridge, grass carp, silver carp, blue scad, tile fish, pomfret, and prawn) were identified both with the clone sequencing and also with the mini-barcode (136 bp) sequence combined NGS method. The NGS method was superior in accuracy, sensitivity, and detection efficiency compared to the clone sequencing method, The edible animal species were identified both in the mixed raw samples and 7 heavily processed food products (different meatballs - beef, pork, fish, and shrimp), 'modified' beefsteak, sausages, and Chinese sausages. Moreover, some unlabelled species and dubious contamination were also detected as well, leading to stringent cleaning procedures.

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This paper reports the performance of a commercial NGS method that has been evaluated as an untargeted tool to identify meat species. The method was tested on pure meat samples, and all species were correctly identified including several exotic species. Closely related meat species were also correctly differentiated. Species were successfully detected and identified in mixtures down to 1% (w/w). The reliability of the method was further confirmed on several proficiency test samples, and promising quantification data were obtained. Finally, 45 minced meat samples sourced from local European and Asian markets were analysed, and 18% of them showed cases of adulteration with undeclared meat species.

3722359646?profile=RESIZE_710x Read the abstract here

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FSAI has worked with a commercial laboratory (Identigen) over the past two years to adapt NGS (next generation sequencing), so that it can be used as a DNA screening tool to check that the composition of the food matches what is stated on the product’s labelling or descripion. FSAI screened 45 plant-based foods and food supplements from Irish health food shops and supermarkets. It looked for the presence of all plant species in the selected products and identified 14 food products for further investigation that may contain undeclared plant species. Of these 14 products, one was confirmed to contain undeclared mustard at significant levels, which is an allergen that should be declared. Another product (oregano) was found to contain DNA from two undeclared plant species, one at significant levels. A third product was found to have no DNA from the plant species declared on the label, but instead rice DNA was identified. All three products are under further investigation. FSAI will apply the same technology for the screening of meat, poultry and fish products.

 Read the FSAI Press Release and the full report

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In this study, NGS (next generation sequencing) using the Ion Torrent semiconductor platform was applied to identify meat species in several highly processed and complex meat products and meat derived broths (a döner kebab, a beef/pork paté, a meat based filling of tortellini, one instant granular preparation of broth stock made from meat, and two ready-to-use meat broths from different producers). Sequence analysis of reads from 6 libraries detected expected and unexpected meat species in the products. A measure of poor hygienic practice during production of the analysed products could be inferred using the number of human reads. In conclusion, NGS data is useful for authentication of highly processed products.

Read the abstract at: NGS of meat products

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Honey is a high value food which suffers from food fraud, and it is important to have methods to verify its floral and geographical origin. Italian researchers isolated DNA from nine honeys (six monofloral honeys produced in Italy, two polyfloral honeys produced in East Europe and Chile respectively, and one honeydew honey), and PCR amplified for a chloroplast trnL barcoding fragment. The amplicons were sequenced and the data bioinformatically analysed against a database of 150,000 botanical entries. A total of 254 botanical groups were identified from the nine analysed samples.

The prevalent expected botanical origin was confirmed in five out of six monofloral honeys. The plant signature of the labelled lime tree blossom honey did not confirm the expected botanical prevalence. The botanical composition of monofloral and polyfloral honey samples was useful to infer their geographical origin. 

Read the abstract at: NGS authentication of honey

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