Analyses to ensure food safety and quality are more relevant now because of rapid changes in the quantity, diversity and mobility of food. Food-contamination must be determined to maintain health and up-hold laws, as well as for ethical and cultural concerns. Real-time polymerase chain reaction (RT-PCR), a rapid and inexpensive quantitative method to detect the presence of targeted DNA-segments in samples, helps in determining both accidental and intentional adulterations of foods by biological contaminants. This review presents recent developments in theory, techniques, and applications of RT-PCR in food analyses, RT-PCR addresses the limitations of traditional food analyses in terms of sensitivity, range of analytes, multiplexing ability, cost, time, and pointof-care applications. A range of targets, including species of plants or animals which are used as food ingredients, food-borne bacteria or viruses, genetically modified organisms, and allergens, even in highly processed foods can be identified by RT-PCR, even at very low concentrations. Microfluidic RT-PCR eliminates the separate sample-processing step to create opportunities for point-of-care analyses. We also cover the challenges related to using RT-PCR for food analyses, such as the need to further improve sample handling.
Accidental and/or incidental adulterations of foods by porcine ingredients are common in the globalized food processing industry. Food mislabelling and fraudulent substitutions of non-porcine ingredients with porcine ones are objectionable to those who abstain from porcine derived products due to habitual (e.g., vegans and vegetarians), medical (e.g., porcine allergies), legal (fraudulent labelling), economic (e.g., substitution of expensive meat with cheaper pork meat) and cultural or religious grounds (e.g., Islamic and Jewish dietary restrictions). Thus, a strong demand exists for a fast and sensitive method for quantitative sensing of porcine DNA in food. In this study, we are reporting the development of probe-free real-time PCR assay with new primer sets targeting the cytochrome b gene for the fast and sensitive detection of porcine DNA in real food samples. Standard curve was developed with six tenfold dilutions of the DNA standard and the assay successfully detected up to 0.00001 ng/μL of porcine DNA and as low as 0.001 % porcine adulteration in raw pork-chicken binary mixture. The standard curve indicated a linear regression of R 2 value of 0.990 and an efficiency of 92.5 %. The Ct value range for the detected pork DNA from the 35 food samples tested was 16.03-28.76. We confirmed the assay's specificity to porcine DNA against nine non-porcine animal species and 6 vegetables species.
Rapid, sensitive, and selective pathogen detection in food industry is of paramount importance for global health. Currently available diagnostic assays based on polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA), which are time-consuming, complex, and relatively expensive, are ill suited to meet the current requirements for pathogen detection in the food industry. Different isothermal amplification techniques are faster, more sensitive, and more robust compared to other standard pathogen detection methods. Modern techniques are based on molecular diagnostics integrated with various detection modalities such as bioanalytical, electrochemical, and luminescence techniques to ensure more sensitive and specific detection processes. Here, we review conventional and non-conventional techniques for pathogen detection on food industry. We focus mainly on nucleic acid detection integrated with electrochemical biosensors. In addition, the applications of nanoparticle biosensors and colorimetric sensors are reviewed. Lab-on-a-chip (LOC) devices based on design strategies utilizing isothermal amplification for pathogen detection are also discussed.
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