A fluorescence and colorimetric dual-mode LAMP method for detection of Vibrio parahaemolyticus

Abstract

Conventional nucleic acid detection technology for foodborne pathogens relies on advanced analyzers and professional technicians, but in areas with limited resources, this method limits the rapid detection of pathogens, which highlights the importance of developing a method suitable for on-site detection. A dual-mode detection method was proposed for Vibrio parahaemolyticus (V. parahaemolyticus) based on loop-mediated isothermal amplification (LAMP), which integrates fluorescence detection and visual colorimetric detection. This method was developed for instant on-site detection. When the experimental conditions are available, the fluorescence detection method can be selected for quantitative determination, and when the resources are limited, the visual colorimetric method can be used for qualitative determination. The dual-mode LAMP method has been shown to achieve comparable sensitivity to real-time fluorescent LAMP, with the capacity to detect 1 pg μL⁻¹ of V. parahaemolyticus DNA with high specificity. In order to enhance the objectivity of detection, an RGB image analysis method was developed, and a simple judgment strategy was proposed: when the B/G value is less than 1.3 and the R/G value is less than 0.8, the result can be judged as positive. In addition, the real samples were tested, and the accuracy rate of the visual colorimetric detection mode was found to be 96%. The RGB analysis method reduced the subjective error of naked eye colorimetry, and the accuracy rate was equivalent to that of the real-time fluorescence mode, reaching 98%. This method provides a novel and sensitive idea for on-site instant detection.