DNA testing of food products and ingredients is the most accurate way to identify and authenticate species, detect contaminants or adulterants, and verify label claims. It can help prevent food fraud and may help meet upcoming NOAA traceability regulations for seafood importers. This article discusses how DNA testing can help supply chain and seafood authenticity; the differences in chemical, DNA barcode, and next-generation sequencing (NGS) testing; DNA testing specific to seafood; and DNA testing as part of quality control.
As the global food supply chain becomes increasingly complex and fragmented, the challenges of safeguarding the food supply also increase. One of these challenges is protecting against food fraud and misrepresentation. Food fraud is a growing problem worldwide, costing over $49 billion annually. Economic pressure to provide cheaper food products has contributed to the fraud issue. For example Alaskan pollock being substituted for cod and steelhead trout for salmon, which causes increasingly savvy consumers to look for third-party verification of label claims.
To ensure their products are accurately identified and labeled, and contain only what is listed on the ingredient list, food companies are turning to DNA testing of products and ingredients.
Authenticity Specific to Seafood
According to the U.S. FDA, fresh fish offered in restaurants and retail stores is not always labeled correctly. A lower-cost fish such as tilapia might be labeled as a higher-value fish such as grouper. In other cases, the same fish may be marketed under different regional names or an uncommon fish name may be changed to something more familiar to consumers, even though the common name is a different genus and species (such as lingcod labeled as cod).
To encourage consistent fish labeling, the FDA developed a list of recognized seafood names that companies may use. The FDA also created a single, laboratory-validated method of generating DNA barcodes for the identification of fish for regulatory compliance. This means one set of universally accepted barcode IDs is available for companies to test any seafood.
Under the Federal Food, Drug, and Cosmetic Act, seafood must be labeled truthfully and not mislead consumers. Products that don’t comply are considered misbranded, which can result in FDA regulatory action such as a civil money penalty, no-sale order, seizure, and/or injunction. The law applies to components, packaging, and finished products, so all companies involved in handling fish—manufacturers, packers, distributors, and retailers—are responsible for assuring that they are not dealing in adulterated or misbranded products.
How DNA Testing Can Help
Being able to prove that what is on the label is what is in the product is beneficial for producers, suppliers, retailers, and consumers. It helps producers know that the ingredients they are paying for is what they are receiving.
Testing using an independent lab allows labeling ingredients and finished products as third-party DNA authenticated, which also increases consumer acceptance and allows differentiating products from competitors. In addition, authenticated food and food ingredients reduce the risk of adverse events caused by unidentified ingredients, of litigation, and of regulatory action.
DNA Testing in a Nutshell (or Lobster Shell)
DNA is the genetic code contained in cells of plants, animals, bacteria, and fungi. Parts of DNA can identify a specific individual (as in forensic or medical DNA), a broad group (such as fish), or a specific genus and species (such as tilapia).
DNA testing is appropriate for virtually any material that contains DNA in a wide range of fresh and processed products. The more processed a product is, the smaller or more fragmented the DNA samples available for extraction are. Examples from least to most processed seafood products include fresh fish (like a whole salmon or a tilapia fillet), mixed ingredients (like crab cakes), liquid extract (like fish oil), and dried extracts (like fishmeal).
If the material to be tested never contained cells or if no cell fragments remain in a product (e.g. they were removed through filtration, exposed to extremely high heat, or highly chemically processed), alternative methods to DNA are necessary, and may include chemical or other analytical tests.
Chemical Testing vs. DNA Testing
While chemical testing can provide information about the specific chemical components both in products that contain DNA and those that do not, chemical testing requires having an idea of what the product is. If you think you have salmon, you can run a chemical test to see if it’s salmon—the result is a yes or no. Chemical testing can also identify a known ingredient and its quantity. For example, you can test if the product contains caffeine and if so, how much. In addition, chemical testing is not highly specific, like DNA testing; many different organisms have identical chemical components. For instance, caffeine is found in numerous plants, such as coffee and tea, so it is not useful for species identification.
DNA testing provides the most definitive and specific identification available today. It can identify expected and unexpected adulterants, contaminants, substitutes, and allergens. DNA testing does not require a priori knowledge of what is supposed to be in the product. For instance, it can identify a completely unknown or mislabeled product; this is very important for testing samples from the marketplace that often are misbranded.
DNA Barcoding vs. NGS
DNA barcoding uses one standard genomic region (or gene sequence) to identify a species, similar to how UPC codes identify products when read by barcode scanners. It was developed to identify distinct groups of animals, such as fish versus chicken. NGS can also use the single gene region, but sequence it thousands of times from a single sample. It can also detect and identify allergens, fillers, and contaminants.
Both barcode and NGS methods can:
- Be used for living, fresh, or dried raw material with little or no processing;
- Place unknown or mislabeled species into general categories (tuna or salmon);
- Detect large amounts of adulterants (rice or soy); and
- Identify animal and plant DNA.
DNA barcoding and NGS differ in the use of primers. (A primer is a strand of short nucleic acid sequences, generally about 20 base pairs, that is the basis of DNA replication; it is explained in more detail it the next section.) DNA barcoding uses a single set of universal primers. While NSF’s NGS method uses validated universal primers and proprietary species-specific primers.
NSF uses a proprietary process called specific, targeted NGS that can identify more than 10,000 species of botanicals, animals, fungi, and bacteria in raw ingredients and finished products. These tests can differentiate between the most closely related and most difficult species to identify, including plant hybrids and complex mixtures. Batches can be sampled from any starting material.
NSF’s methods can also detect and identify allergens, fillers, and contaminants. For example, DNA Clean Screen identifies the presence of 11 common fillers and allergens at limits of detection as low as 50 picograms of DNA. These include wheat, soy, pea, corn starch, potato (maltodextrin), rice, peanut, almond, walnut, pecan, and hazelnut. NSF also offers two proprietary genetically modified organism (GMO) screens, which can be used on finished processed seafood products to verify label claims. These tests screen for the top 10 GMO species and the 10 most common GMO events or elements.
Both expected and unexpected contaminants, as well as fillers and allergens, are detected, whether they are in high and or very low abundance (down to a few molecules of DNA). NGS testing also provides relative ratios of DNA sequences to assess the extent of adulteration.
NSF’s technique was developed to work with old, degraded DNA pieces, or very small fragments of DNA. The method uses targeted DNA primers to definitively identify species through a polymerase chain reaction, or PCR, technique that amplifies DNA segments of interest, or those segments that uniquely identify a species. Primers are like a probe that is made specifically to target a particular species to allow replication of DNA. To develop primers capable of identifying species in processed and degraded products, NSF scientists reverse engineered these primers from known DNA sequences of the target species.
Creating primers requires advanced knowledge of the biology, evolutionary history, and DNA of the target and closely related species. NSF developed specific primers using a proprietary database of thousands of validated DNA reference sequences obtained from museum specimens through partnerships with academic institutions and botanical gardens. NSF sequenced the source DNA of these specimens and determined which regions of the genome (genes) are unique identifiers for each of the target species of interest. Next, NSF designed primer sets to amplify the specific gene region of interest to use in NGS.
Identification for Seafood
DNA barcode methods are appropriate for testing and identifying seafood and meat because a single gene region can be used to differentiate and ID a wide range of different animal species. The U.S. FDA has validated a single gene region or barcode, and created specific guidelines for the DNA barcode testing of seafood. Plant species, on the other hand, have more than one gene region critical for identification, and due to hybridization, one plant genus can contain thousands of species, whereas most animal (mammal and fish) genera contain only a few species in comparison.
Unlike DNA barcoding, however, NSF’s NGS can use the single gene region and sequence it many thousands of times from a single sample. Therefore, instead of identifying a single species in a material—which would be appropriate for a whole fish fillet—NGS can identify all elements of processed or ground products (like crab cake or fishmeal), including fillers (such as soy protein, rice flower, and maltodextrin), binder starches, contaminants, undeclared additives, and any other animal species.
DNA Testing in Quality Control
Integrating DNA testing as a standard of quality control can help food companies create a “first line of defense” against adulteration and fraud. Testing all or sample lots of incoming raw materials ensures they are the correct species and free from harmful adulterants, and can also help companies verify suppliers.
The cost of DNA testing is comparable to other analytical testing methods, but is more accurate at detecting adulteration. DNA tests identify over 25 percent of routine samples as adulterated or substituted, most commonly by unexpected species and by closely related species not detected by other testing methods.
Additionally, DNA testing assures that finished products are “pure” by screening for the presence of labeled ingredients and the absence of allergens, fillers, and GMOs. The best way to ensure products are pure is to test raw ingredients, but knowing what is on the shelf is also important.
Dr. Reynaud is global director of scientific innovation for NSF International and co-founder of NSF AuthenTechnologies. She is a botanical taxonomist and geneticist with over 15 years of DNA testing experience. Reach her at firstname.lastname@example.org.