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Evaluation of Sensitivity and Applicability of a Droplet Digital Pcr Assay for Simultaneous Detecting Pseudomonas aeruginosa and Pseudomonas fragi in Foods
Huang, J.; Zhai, L.; Wang, J.; Sun, X.; Wang, B.; Wei, Z. An Evaluation of the Sensitivity and Applicability of a Droplet Digital Polymerase Chain Reaction Assay to Simultaneously Detect Pseudomonas aeruginosa and Pseudomonas fragi in Foods. Foods2024, 13, 1453.
Huang, J.; Zhai, L.; Wang, J.; Sun, X.; Wang, B.; Wei, Z. An Evaluation of the Sensitivity and Applicability of a Droplet Digital Polymerase Chain Reaction Assay to Simultaneously Detect Pseudomonas aeruginosa and Pseudomonas fragi in Foods. Foods 2024, 13, 1453.
Huang, J.; Zhai, L.; Wang, J.; Sun, X.; Wang, B.; Wei, Z. An Evaluation of the Sensitivity and Applicability of a Droplet Digital Polymerase Chain Reaction Assay to Simultaneously Detect Pseudomonas aeruginosa and Pseudomonas fragi in Foods. Foods2024, 13, 1453.
Huang, J.; Zhai, L.; Wang, J.; Sun, X.; Wang, B.; Wei, Z. An Evaluation of the Sensitivity and Applicability of a Droplet Digital Polymerase Chain Reaction Assay to Simultaneously Detect Pseudomonas aeruginosa and Pseudomonas fragi in Foods. Foods 2024, 13, 1453.
Abstract
Achieving effective control over microbial contamination necessitates the precise and concurrent identification of numerous pathogens. In this research, we have devised a remarkably sensitive duplex droplet digital PCR (dddPCR) reaction system to simultaneously detect Pseudomonas aeruginosa (P. aeruginosa) and Pseudomonas fragi (P. fragi). Employing comparative genomics, we identified four genes of P. fragi. By specific analysis, RS22680 gene was selected as the detection target of P. fragi and LasR gene was chosed as P. aeruginosa, which were applied to construct a dddPCR reaction. In terms of specificity, sensitivity and anti-interference ability, the constructed dddPCR detection system was verified and analyzed. The assay showed excellent sensitivity and applicability, as evidenced by a limit of detection of 100 CFU/mL. When the concentration of natural background bacteria in milk or fresh meat was 100 times that of the target detection bacteria, the method was still capable of completing the absolute quantification. In the simulation of actual sample contamination, P. aeruginosa could be detected after 3 h of enrichment culture, and P. fragi could be detected after 6 h. The established ddPCR detection system exhibits exceptional performance, serving as a foundation for the simultaneous detection of various pathogenic bacteria in food products.
Keywords
Pseudomonas aeruginosa; Pseudomonas fragi; simultaneous detection; droplet digital PCR
Subject
Biology and Life Sciences, Food Science and Technology
Copyright:
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
