Authentication of caprine milk and cheese by commercial qPCR assay

Abstract

The objective of the study was to investigate potential adulteration of commercial caprine milks and cheeses with bovine milk using commercial qPCR assay. The assay comprised of bovine-, ovine-and caprine-specific primers and TaqMan probe and mammalian internal control. Specificity, sensitivity, linearity, reproducibility and efficiency of the bovine assay were tested as well. Specificity was verified by running reaction on the DNA of other milk-producing species (caprine and ovine) and made-up bovine-caprine (v/v) milk mixes. In both experiments, a bovine DNA fragment was amplified whereas no amplification was obtained from the other species. Sensitivity, linearity, reproducibility and efficiency were tested on 10-fold dilution series of 10 ng bovine DNA. The assay has shown good linearity (R 2 = 0.983) within whole range, with efficiency of 86% and excellent reproducibility (SD around the C T for the technical replicates <0.5). The sensitivity was adequate, as calculated LOD and LOQ were 1.44 pg and 2.94 pg of bovine DNA, respectively. Finally, the assay was used to authenticate 5 caprine milk samples and 5 caprine cheese samples, purchased from local supermarkets. Totally, 1 milk sample has shown the fluorescence signal, which exceeded baseline in cycle 39.01 ±0.69. However, the signal was above LOD and LOQ suggesting that there could not be unambiguously declared any adulteration with bovine milk. Amplification of bovine-specific DNA was not observed in the other samples indicating products were not adulterated. The commercial qPCR assay has proved that real-time PCR assays, as well as DNA-based techniques in a general, are the excellent and reliable tools for fighting with frauds in the food industry and protecting the public health.

Assessment of cheese frauds, and relevant detection methods: A systematic review

“… ( Fuselli et al, 2015 ) – Caprine Bovine's milk QPCR ASSAY method was successfully implemented. ( Fekete et al, 2017 ) Brazil Fresh goat cheese Bovine's milk A duplex PCR test was used to test goat cheeses for assessing adulteration. The test could detect 0.5 % (v/v) cow milk.…”

Food safety and quality. Scientific proceedings

“…Ako uvádza Zeleňáková et al (2017), medzi hlavné faktory ovplyvňujúce vznik mikrobiologického nebezpečenstva patria: teplota, pH a vodná aktivita (aw). Pre rýchlosť rastu si mikroorganizmy vyžadujú určité optimálne teploty.…”

“…Po separácii a zafarbení sa vykonáva denzitometrické vyhodnotenie elektroforetických pásov pomocou softvéru (Sienkiewicz, Dogan a Kroh, 2006). Na detekciu falšovania ovčieho syra sú k dispozícii aj iné metódy (Zeleňáková et al, 2016;Fekete et al, 2017;Šnirc et al, 2017). a možno ich použiť v kombinácii s touto metódou na podporu platnosti výsledkov.…”

Detection of adulteration of traditional Slovak bryndza ewe’s cheese with cow’s lump cheese by isoelectric focusing of gamma caseins

“…The presence of modified γ2-casein (with pI = 6.7), which derives from the interaction between γ2-casein and aldehydes coming from microbial metabolism, could be present on the gel . [28] There are several methods available to detect adulteration in ewe's cheese, [29][30][31] and these can be used in combination with this method to validate the results.…”