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  • Bioline Scholar: Mystery Foods, Food Authenticity & DNA-Based Testing Methods

    28 March 2013 No comments

    Horsemeat in beef products A 'beef' burger?

    Last night BBC3 broadcast a novel show presented by Rick Edwards and Heydon Prowse, Horsemeat Banquet, a "one-off experiment in which a chef cooks up a horsemeat banquet designed to challenge the prejudice of a group of diners". Set in a dining area that more resembled an abattoir than a usual eating environment, the aim of the show was to address fears and qualms about eating horsemeat by preparing and presenting it in ways similar to that of other more routinely eaten meats. Horse courses included steak tartare, hache (meatballs) and a roast horse sirloin. You can watch the show here on BBC iPlayer but be warned, it's not for the squeamish.

    In addition to serving up horse-courses, the show gathered various freely available high-street fast-foods and tested them with perhaps surprising results. A beef in black-bean sauce and lamb curry tested were found to contain very little if any, actual beef or lamb and one burger contained no beef, but only blood and heart. Another beef dish contained only chicken material, again including blood. Most alarmingly of all, a mystery meat in one lamb curry could not be identified by scientists, despite their best efforts, leading to today's headlines such as, "Dog or cat in our curry: Fears over 'mystery meat' in London takeaway which has baffled experts".

    The reality is that the witting consumption of horsemeat is as much a cultural issue as anything else, as horse is routinely and knowingly eaten in many countries across Europe and Asia; the key here being the idea of witting consumption. Indeed, the issue at the heart of the ongoing horsemeat scandal is unrelated to the knowing consumption of horse or any other meat-based foodstuffs, but instead revolves around the integrity and transparency of more generally accepted beef, lamb, chicken or pork products. As one of the participants in Horsemeat Banquet noted, "At least I know that it's 100% what you tell me it is. at least I know what I'm eating."

    So, if more conventionally consumed meats are not in fact the meats they purport to be, while in some cases claiming to be 100% a particular meat, what in fact are they made of and what are food testing laboratories likely to find next in our food supply chains, Soylent Green?

    "Without a commandment that "thou shall not adulterate," food adulteration has been a crime of the ages." wrote Susan Taylor of the Chicago Times Tribune in a March 13 lifestyle special article on how to avoid doctored food.

    Food authenticity is undoubtedly a 'hot potato' subject of great concern to food authorities and the general public. Horsemeat found in food ranging from Ikea meatballs through to Burger King burgers are just two of the recent high-profile organisations to be tarnished by the international 'horsegate' scandal.

    It is not just meat that has made everyone look harder at what they are (or believe to be) eating. Adulterated olive oil and honey and diluted juices are long-running 'scams'. Even herbs and spices including Saffron (one of the world's most expensive spices) has a 4,000 year history of adulteration. In the US, one study found a third of seafood products were mislabelled.

    Incorrect labelling of foodstuffs is not only commercial fraud, but represents a public health hazard due to the presence of sub-standard commodities unfit for human consumption. Such contaminants include powerful veterinary drugs, toxins, or allergens such as soya and peanuts, which can cause severe allergic reactions and, at worst, could result in death of the unwitting allergic consumer. Food manufacturers have a public duty to be vigilant as the prevalence of allergies is rising at an "alarming rate", according to Allergy UK. In addition to the physical harm caused by contaminated foodstuffs, there are also ethical, religious and legal implications and considerations, such as non-Halal/non-Kosher contaminants in products sold as being Halal or Kosher.

    Not surprisingly authorities have been busy ramping up scientific testing efforts and determining food origins and traceability, which are easily lost in what are often very complex, cross-border supply chains. DNA-based analytical methods such as the Polymerase Chain Reaction (PCR) are used to authenticate meat, seafood and dairy products, as well as foods originating from plants. However, while food testing can easily determine the presence of contaminating material in foodstuffs, tests will only confirm or deny the presence of substances that are actively being sought. In other words, in cases where multiple contaminants are suspected, unless multiplex test methods are used, separate tests must be carried out for each contaminant to ascertain its presence.

    PCR can also be used to detect presence of food allergens, bacterial contamination and genetically modified organisms (GMO) thanks to the method's high specificity and sensitivity. Rapid turnaround time and low running costs are also advantages with PCR and PCR based food testing. With the availability of very high-speed PCR reagents such as MyTaq™, PCR can be performed in just 30-60 minutes, rather than the more usual 2.5-3 hours.

    In this edition of Bioline Scholar we take a look at recent papers in the important area of food testing. Bioline offers a range of widely used and trusted molecular biology reagents and kits perfectly suited to DNA-based food testing.


    Cawthorn, D.-M. et al., (2013). A high incidence of species substitution and mislabelling detected in meat products sold in South Africa. Food Control 32(2), 440-449

    This study from Professor Hoffman at the University of Stellenbosch, found rather startlingly, 68% of meat samples in South Africa contained species not declared in the label, including pork, chicken, and even donkey, goat and water buffalo. Almost a third of the products contained soy and gluten, also not mentioned in the label. No horsemeat, however, was found.

    Lago, F.C. et al., (2013). Authentication of gadoids from highly processed products susceptible to include species mixtures by means of DNA sequencing methods. Eur. Food Res. Tech. 236 (1), 171-180

    The authors developed a unique method to detect species mixtures of the economically important Gadus genus (includes Atlantic Cod) in even highly processed products. The authors performed PCR and sequencing of mitochondrial cytochrome b and subsequent SNP analysis. Such efforts will help assurance of correct labelling.

    Barros, E., (2013). Identification of Mycotoxigenic Fungi Using an Oligonucleotide Microarray. Lab Prot. Fungal. Biol. DOI: 10.1007/978-1-4614-2356-0_52

    Mycotoxins are secondary metabolites produced by fungi that can contaminate food and affect human and animal health. This paper describes an oligonucleotide microarray specific for eleven mycotoxigenic fungi isolated from different food commodities in South Africa.

    dNTPs & BIOTAQ

    Ojer-Usoz, E. et al. (2013). Prevalence of extended-spectrum β-lactamase-producing Enterobacteriaceae in meat products sold in Navarra, Spain. Meat Sci. 93 (2), 316-321

    Resistance to β-lactam antibiotics in Enterobacteriaceae isolates were investigated and found to be highly prevalent in 141 meat products (beef, poultry and pork) purchased in Spain. β-lactamase genes were detected using multiplex PCR and DNA sequencing. Poultry products had the highest prevalence (84%), with E.coli being the predominant bacteria (71.3%).


    Vaagt F., et al., (2013). Loop-mediated isothermal Amplification (LAMP) based Method for rapid Mushroom Species Identification. J. Agric. Food Chem. 61 (8), 1833–1840

    Toxic mushroom species, such as the death cap (Amanita phalloides), are responsible for most mushroom poisonings. In this paper from the Institute of Food Chemistry, University of Hamburg, novel LAMP assays were employed to differentiate even closely related edible and toxic mushroom species.

    Focke, F., et al., (2013). Loop-mediated Isothermal Amplification (LAMP): Methods for Plant Species Identification in Food. J. Agric. Food Chem. DOI: 10.1021/jf304295b

    LAMP-based methods for plant species identification were compared to an existing real-time PCR assay for detecting spices.

    Mayer, F. et al., (2012). Use of Polymorphisms in the γ-Gliadin Gene of Spelt and Wheat as a Tool for Authenticity Control J. Agric. Food Chem. 60 (6), 1350–1357

    Two alternative DNA-based analytical methods (PCR-RFLP and real-time PCR) were developed for detection and quantification of spelt flour "adulteration" with soft wheat using SNPs in γ-Gliadin.

    dNTPs, HyperLadder II

    Vázquez-Sánchez D., et al., (2012). Incidence and characterization of Staphylococcus aureus in fishery products marketed in Galicia (Northwest Spain). Int. J. Food Microbiol. 157 (2), 286-296

    A quarter of 298 fishery products purchased from retail outlets in Galicia, Spain between January 2008 and May 2009 were positive for the presence of Staphylococcus aureus using PCR-RAPD. Putative enterotoxigenic strain counts reached high risk levels in 17 products.

    Horsegate: Horse-meat DNA testing in Beef Products

    15 February 2013 No comments

    On 15 January, four major UK supermarkets withdrew meat-based products in the light of the Food Standards Agency Ireland (FSAI) findings into the contamination of beef with horse and other animal meats.

    The highest content of equine DNA was 29% in one sample, with the vast majority of others tested below <0.2%. Many samples were also found to contain porcine DNA.

    The scandal has resulted in one of the biggest food recalls in UK history, and multiple suppliers across the EU being implicated. To deal with the growing scandal over mislabelled horse-meat, the EU Health Commissioner today urged all members to carry out DNA tests on processed beef for traces of horse-meat for three months from 1 March. Furthermore, tests for the presence of the veterinary medicine phenylbutazone ("bute") were recommended.

    We thought it may be interesting to look at some of the scientific challenges in authenticating presence of animal species in our meat. What types of horse-meat tests are out there? Further, how accurate are current tests in quantifying horse DNA? In fact, one food testing company in the UK this week has urged the food testing industry to not rush into horse DNA testing due to issues surrounding DNA quantification.

    Currently, in the food testing industry there exist two main types of assays for detecting horse meat in beef – either protein-based or DNA-based.

    The ELISA method tests for protein and has a 1-2% detection limit. Separate kits exist for testing either raw or cooked meat. However, due to the contamination being largely unknown, the ELISA method could possibly show inaccurate results.

    With molecular-based testing of equine DNA using PCR, the detection limit is 1% but the limit of detection (i.e. sensitivity) can go down even lower. Both real-time PCR methods and end-point PCR (with subsequent RFLP analysis or sequencing) are suitable for DNA testing of equine DNA. PCR based methods amplify sequences from mitochondrial DNA (mtDNA) which is more highly conserved and widely found in animal species than nuclear DNA. Specific details of primer sequences are not always made available by different companies.

    Due to the wide variation in the amount of mitochondria from cell to cell in different tissue types (varying from 1 mitochondria per cell to 1000’s), it is not possible to accurately quantify the amount of horse DNA in a sample. DNA testing therefore should be seen as being qualitative rather than quantitative, and limitations to their accuracy should be borne in mind when interpreting results or those quoted in the press.

    Bioline high performance molecular biology products have been cited in recent papers investigating contamination of meat products by horse meat, as well as other types of contaminants detected in food-testing.

    BIOTAQAuthentication of species in meat products by genetic techniques.

    BIOTAQ: A high incidence of species substitution and mislabelling detected in meat products sold in south Africa

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