The usage of porcine pepsin or other porcine derivatives in food products is a common practice in European, American and certain Asian countries although it creates issues in religious and personnel health concerns. In this study, porcine pepsin was detected using indirect ELISA that involved the anti-pep80510 polyclonal antibody raised against a specific peptide of porcine pepsin, pep80510. The sensitivity of the assay for standard porcine pepsin was 0.008 µg/g. The immunoassay did not cross-react to other animal rennet and milk proteins except for microbial coagulant from Mucor miehie. The recovery of porcine pepsin in spiked cheese curd within the range of CV < 20% while for porcine pepsin in spiked cheese whey the recovery is also within the range of CV% < 20%.
Detection of porcine plasma using indirect ELISA was developed using mAb B4E1 for the prevention of their usage in human food that creates religious and health conflicts. The immunoassay has a CV
Lateral flow devices (LFDs) are straightforward scientific tools that have made substantial advances in recent years. They have been used in many fields including the meat industry to detect disease markers, determine meat freshness or meat species determination. They are, therefore, significant in the research of meat adulteration by mixed animal species, because food component authenticity is a serious concern encompassing health, economic, legal, and religious issues. Pork adulteration is one of the most crucial issues in the global meat industry. In this review, we discuss the various types of LFDs and recent research on the development of LFDs as an authenticity tool for detecting pig additives in meat-based products, and how regulatory authorities could adopt LFDs for their workflows. Despite the benefits of rapidity, simplicity, low cost, high sensitivity, and specificity, researchers face challenges when using LFD as a final confirmation test. Future directions are suggested for globalising the use of LFD as a halal authentication method.
The inclusion of ingredients derived from pigs in highly processed consumer products poses a significant challenge for DNA-targeted analytical enforcement, which could be overcome by using digital PCR. However, most species detection methods use digital PCR to target single-copy nuclear genes, which limits their sensitivity. In this work, we examined the performance of a nanoplate-based digital PCR method that targets multi-copy nuclear (MPRE42) and mitochondrial (Cytb) genes. Poor separation of positive and negative partitions, as well as a 'rain effect' were obtained in the porcine-specific MPRE42 assay. Among the optimization strategies examined, the inclusion of restriction enzymes slightly improved the separation of positive and negative partitions, but a more extensive 'rain effect' was observed. The high copy number of the MPRE42 amplicon is hypothesized to contribute to the saturation of the positive signal. In contrast, the porcine-specific Cytb assay achieved perfect separation of positive and negative partitions with no 'rain effect'. This assay can detect as little as 0.4 pg of pork DNA, with a sensitivity of 0.05% (w/w) in a pork-chicken mixture, proving its applicability for detecting pork in meat and meat-based products. For the MPRE42 assay, potential applications in highly degraded products such as gelatin and lard are anticipated.
The inhibitory properties of novel antimicrobial proteins against food-borne pathogens such as Listeria monocytogenes offer extensive benefits to the food and medical industries. In this study, we have identified antimicrobial proteins from a milk curd-derived bacterial isolate that exhibits antilisterial activity using genome mining and mass spectrometry analysis. The analysis of the draft genome sequence identified the isolate as Paenibacillus polymyxa Kp10, and predicted the presence of antimicrobial paenibacillin, paenilan, paeninodin, sactipeptides, thiazole-oxazole modified microcin, and histone-like DNA binding protein HU encoded in its genome. Interestingly, nanoLC-MS/MS analysis identified two histone-like DNA binding proteins HU as predicted in silico earlier, exhibiting antilisterial activity. Additionally, translation initiation factor IF-1 and 50S ribosomal protein L29 were also discovered by the mass spectrometry in the active fractions. The antilisterial activity of the four proteins was verified through heterologous protein expression and antimicrobial activity assay in vitro. This study has identified structural regulatory proteins from Paenibacillus possessing antilisterial activity with potential future application in the food and medical industries.