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For the first time, we describe an innovative polymerase spiral reaction (PSR) assay for the rapid, simple, and accurate detection of pig tissues or pork in adulterated meat including heat-treated and processed ones. The PSR assay specifically targeting the mitochondrial cytochrome b (cyt-b) gene of the pig was successfully optimized permitting assay results in 65 min time. The developed detection method was 100% specific amplifying only the cyt-b gene and displaying negative results with all the tested non-pork meats. The sensitivity of the developed PSR (760 fg porcine DNA) was tenfold better than the end-point PCR and able to detect heat-treated (121 °C) and adulterated (0.5% pork in beef) meat and processed pork products such as sausages, salami, meatball, soup, curry, etc. The developed PSR-based method can be used for point-of-care detection with minimum instrumentation and technical expertise to guarantee instant clearance of exported and imported meat products. This is the first time that PSR has been adapted for food authenticity purposes. [ABSTRACT FROM AUTHOR]

Campylobacter jejuni is a widespread zoonotic foodborne pathogen and one of the major concerns in the poultry-based food industry. There is a need for a rapid, robust and equipment free detection system for C. jejuni as the conventional culture method is labour and resource-intensive. In this study, we have developed a simple saltatory rolling circle amplification (SRCA) assay integrating a cost-effective three-step centrifugation-based DNA extraction method for rapid and visual detection of C. jejuni in chicken meat. The specificity of the SRCA assay was established by using 17 C. jejuni and 18 non -C. jejuni strains. SRCA, end-point and real-time PCR were able to detect 50 fg, 5 pg and 500 fg C. jejuni DNA/tube, respectively. In the artificial-spiking study conducted in C. jejuni -free chicken meat, a limit of detection (LoD) of 3 log CFU/g and 2 log CFU/g were attained in 120 min (without enrichment) and 14 h (with 12 h enrichment), respectively. The LoD of the SRCA assay was 10- and 100-fold better than the end-point and real-time PCR, respectively. Therefore, keeping the end-user of resource-limited food analysis laboratories in mind, we have successfully developed a SRCA assay for rapid and accurate detection of C. jejuni in chicken meat. - Developed a novel SRCA assay to detect Campylobacter jejuni in chicken meat. - LoD of 3 log CFU/g and 2 log CFU/g were attained in 120 min and 14 h, respectively. - The method was validated in artificially spiked and naturally contaminated chicken meat samples. - A simple method that enables sophisticated-equipment free operation and visual interpretation of results. [ABSTRACT FROM AUTHOR]

Clostridium perfringens (C. perfringens), a prolific toxin-producing anaerobe is an important foodborne pathogen with a huge public health concern. Rapid and on-site detection of C. perfringens is of specific importance in developing countries. In the present study, saltatory rolling circle amplification (SRCA) assay was developed for culture-independent, rapid and visual detection of C. perfringens and evaluated in meat with pork as a model. The specificity of the SRCA assay was ascertained by using 62 C. perfringens and 18 non- C. perfringens strains. The analytical sensitivity of the developed SRCA, conventional and real-time PCR assays were 80 fg, 800 fg and 800 fg DNA per tube, respectively. The limit of detection of the SRCA assay was 80 CFU/g of pork in the absence of enrichment and 8 CFU/g after short enrichment of 6 h. The detection limits of 80 CFU/g and 8 CFU/g of pork were attained within 120 min and 8 h, respectively. Real-world or field relevancy of the developed assay was evaluated by screening 82 raw and processed pork samples. As the developed assay is simple, user-friendly, cost-effective and sophisticated-equipment free, it would be more suitable for on-site testing of C. perfringens in foods. To our information, this is the first report to apply SRCA for the detection of C. perfringens. [Display omitted] • A novel SRCA assay was developed for rapid and visual detection of Clostridium perfringens in meat. • The analytical sensitivity of the SRCA assay (80 fg) was 10 fold more than the conventional and real-time PCR. • The detection limit of 80 CFU and 8 CFU per gram of meat was attained within 120 min and 8 h, respectively. • This is the first report to use SRCA for the detection of C. perfringens. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is one of the most common and leading causes of food poisoning and also responsible for multifarious diseases in humans. Rapid and on-site detection of S. aureus is of specific importance in developing countries. In the present study, a polymerase spiral reaction (PSR) assay was developed for sensitive, rapid and visual detection of S. aureus and validated in meat using pork as a model. The specificity of the PSR assay was ascertained by using 18 S. aureus and 17 non- S.aureus strains. The PSR assay was 100- and 10-fold more sensitive than conventional end-point PCR and real-time PCR, respectively. The limit of detection of the PSR assay was 19.9 × 103 CFU/g of pork without enrichment and 19.9 CFU/g after 6 h enrichment. The detection of 19.9 CFU/g of pork was attained within 8 h. Real-world or field applicability of the developed assay was evaluated by screening 76 raw and processed pork samples. Seeing its rapidity, simplicity, cost-effectiveness, user-friendliness and sophisticated-equipment free nature, this assay has the prospective to turn into the assay of choice for regular detection of S. aureus in foods in less-advanced laboratories. To our knowledge, this is the first report to use PSR for the detection of S. aureus. • A novel PSR assay was developed for rapid and visual detection of Staphylococcus aureus in meat. • The analytical sensitivity of the PSR assay was 100 and 10 fold more than the conventional and real-time PCR, respectively. • The detection of limit of 19.9 × 103 CFU and 19.9 CFU per gram of meat was attained within 2 h and 8 h, respectively. [ABSTRACT FROM AUTHOR]