Detection of non-pathogenic and pathogenic populations of Vibrio parahaemolyticus in various samples by the conventional, quantitative and droplet digital PCRs.

Abstract

In this study, three generations of polymerase chain reaction (PCR) assays: (i) conventional PCR, (ii) qPCR and (iii) droplet digital PCR (ddPCR), were systematically tested for their abilities to detect non-pathogenic and pathogenic populations of Vibrio parahaemolyticus. The limit of detection (LOD) for the ddPCR was 1.1pg/µL of purified DNA, followed by the qPCR (5.6pg/µL) and the conventional PCR (8.8pg/µL). Regarding the LOD for V. parahaemolyticus cells, the ddPCR assay was able to detect 29 cells, followed by the conventional PCR assay (58 cells) and the qPCR assay (115 cells). Regarding the sensitivities to detect this pathogen from PCR inhibition prone samples (naturally contaminated mussels), the ddPCR assay significantly outperformed the conventional PCR and qPCR. The ddPCR assay was able to consistently detect non-pathogenic and pathogenic populations of V. parahaemolyticus from naturally contaminated mussels, indicating its tolerance to various PCR inhibitors. This study also revealed the significant difference between conventional PCR and qPCR. The conventional PCR assay showed significantly greater sensitivity than that of the qPCR assay in detecting V. parahaemolyticus in crude samples, whereas the qPCR assay showed better sensitivity in detecting the presence of V. parahaemolyticus in purified DNA samples.