Establishment and application of a rapid visual detection method for Clostridium perfringens in chicken products based on helical loop-mediated isothermal amplification (HAMP).

Abstract

Clostridium perfringens is a significant foodborne pathogen in chicken products. Rapid on-site detection of C. perfringens is crucial for mitigating the incidence of foodborne illnesses by enabling the prompt identification and recall of contaminated food products. A rapid and visual detection method for C. perfringens in chicken products was developed using helical loop-mediated isothermal amplification (HAMP) technology combined with SYBR Green I fluorescent staining. The reaction temperature, time, and reagent concentrations of HAMP technique were optimized firstly. HAMP displayed high specificity, effectively distinguishing C. perfringens from 18 other common pathogens in chicken products. HAMP also exhibited higher sensitivity (78 fg/µL) compared to endpoint PCR and real-time quantitative PCR (qPCR). The detection limit of HAMP for non-enriched samples was 6.8 × 10² CFU/g, which improved to 68 and 6.8 CFU/g after 5 and 10 h of enrichment, respectively. The detection limit of HAMP was lower by 2 and 1 orders of magnitude compared to endpoint PCR and qPCR under the same conditions. On-site testing of commercially available ready-to-eat chicken products showed that HAMP had the same results as traditional culture methods, indicating the significant potential of HAMP for on-site detection of C. perfringens. This method offered a rapid, accurate, and visual means of detecting C. perfringens in chicken products, making it well-suited for on-site testing. This research represented the first use of the HAMP method for detecting C. perfringens.