Real-Time PCR Assay for the Detection and Quantification of Roe Deer to Detect Food Adulteration-Interlaboratory Validation Involving Laboratories in Austria, Germany, and Switzerland.

Game meat products are particularly prone to be adulterated by replacing game meat with cheaper meat species. Recently, we have presented a real-time polymerase chain reaction (PCR) assay for the identification and quantification of roe deer in food. Quantification of the roe deer content in % (w/w) was achieved relatively by subjecting the DNA isolates to a reference real-time PCR assay in addition to the real-time PCR assay for roe deer. Aiming at harmonizing analytical methods for food authentication across EU Member States, the real-time PCR assay for roe deer has been tested in an interlaboratory ring trial including 14 laboratories from Austria, Germany, and Switzerland. Participating laboratories obtained aliquots of DNA isolates from a meat mixture containing 24.8% (w/w) roe deer in pork, roe deer meat, and 12 meat samples whose roe deer content was not disclosed. Performance characteristics included amplification efficiency, level of detection (LOD95%), repeatability, reproducibility, and accuracy of quantitative results. With a relative reproducibility standard deviation ranging from 13.35 to 25.08% (after outlier removal) and recoveries ranging from 84.4 to 114.3%, the real-time PCR assay was found to be applicable for the detection and quantification of roe deer in raw meat samples to detect food adulteration.

Development and validation of a TaqMan real-time PCR assay for the identification and quantification of roe deer (Capreolus capreolus) in food to detect food adulteration.

In order to protect the consumer from meat adulteration it is necessary to identify and quantify the meat content in foodstuffs. Game meat is particularly susceptible for fraudulent labeling since it is more valuable than meat from domestic animals. The paper presents a TaqMan real-time PCR assay for the quantitative determination of roe deer in meat products. The assay developed does not show cross-reactivity with 23 animal and 43 plant species tested and is therefore specific for roe deer. The amplification efficiency determined by analyzing serially diluted roe deer DNA extracts was found to be 93.9%. For quantifying the roe deer content in % (w/w), a reference system based on the myostatin gene was used. The quantification strategy was validated by determining the roe deer content in model meat mixtures and a model sausage. In addition, the real-time PCR assay was applied to the analysis of commercially available meat products.

Authenticity control of game meat products--a single method to detect and quantify adulteration of fallow deer (Dama dama), red deer (Cervus elaphus) and sika deer (Cervus nippon) by real-time PCR.

This contribution presents a single real-time PCR assay allowing the determination of the deer content (the sum of fallow deer (Dama dama), red deer (Cervus elaphus) and sika deer (Cervus nippon)) in meat products to detect food adulteration. The PCR assay does not show cross-reactivity with 20 animal species and 43 botanical species potentially contained in game meat products. The limit of quantification is 0.5% for fallow deer and red deer and 0.1% for sika deer. The deer content in meat products is determined by relating the concentration obtained with the deer PCR assay to that obtained with a reference system which amplifies mammals and poultry DNA. The analysis of binary meat mixtures with pork, a meat mixture containing equal amounts of fallow deer, red deer and sika deer in pork and a model game sausage showed that the quantification approach is very accurate (systematic error generally <25%).

High resolution melting (HRM) analysis of DNA--its role and potential in food analysis.

DNA based methods play an increasing role in food safety control and food adulteration detection. Recent papers show that high resolution melting (HRM) analysis is an interesting approach. It involves amplification of the target of interest in the presence of a saturation dye by the polymerase chain reaction (PCR) and subsequent melting of the amplicons by gradually increasing the temperature. Since the melting profile depends on the GC content, length, sequence and strand complementarity of the product, HRM analysis is highly suitable for the detection of single-base variants and small insertions or deletions. The review gives an introduction into HRM analysis, covers important aspects in the development of an HRM analysis method and describes how HRM data are analysed and interpreted. Then we discuss the potential of HRM analysis based methods in food analysis, i.e. for the identification of closely related species and cultivars and the identification of pathogenic microorganisms.