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Staphylococcus aureus is an important and leading cause of foodborne diseases worldwide. Prompt detection and recall of contaminated foods are crucial to prevent untoward health consequences caused by S. aureus. Helix loop-mediated isothermal amplification (HAMP) is an exciting recent addition to the array of available isothermal-based nucleic acid amplification techniques. This study aimed to develop and evaluate a HAMP assay for detecting S. aureus in milk and milk products. The assay is completed in 75 minutes of isothermal temperature incubation (64 ˚C) and dye-based visual interpretation of results based on colour change. The specificity of the developed assay was ascertained using 27 S. aureus and 17 non S. aureus bacterial strains. The analytical sensitivity of the developed HAMP assay was 9.7 fg/µL of pure S. aureus DNA. The detection limit of the HAMP assay in milk (86 CFU/mL) was 1000x greater than the routinely used endpoint PCR (86 × 103 CFU/mL). The practicality of applying the HAMP assay was also assessed by analysing milk and milk product samples (n = 95) obtained from different dairy farms and retail outlets. The developed test is a more rapid, sensitive, and user-friendly method for the high-throughput screening of S. aureus in food samples and may therefore be suitable for field laboratories. To our knowledge, this is the first study to develop and evaluate the HAMP platform for detecting S. aureus. [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]