Development of a novel multiplex-PCR technology for simultaneous detection of five major aquaculture pathogens

Early and precise pathogen identification corresponding disease management are primary concerns in aquaculture. Here, we attempted at diagnostic methods that can simultaneously identify multiple pathogens, where many samples, several concurrent infections to be handled. Hence, a multiplex PCR assay targeting five major aquaculture viz. Vibrio parahaemolyticus, anguillarum, alginolyticus, vulnificus harveyi was developed for the first time. The primers toxR of V. amiB col vvhA vulnificus, topA were applied. Furthermore, reaction included an internal amplification control against prokaryotic 16S rRNA perceive false-negative results. showed 100% specificity 56 unique bacteria. sensitivity 0.25 ng harveyi, 0.5 vulnificus, 1 parahaemolyticus 2 alginolyticus DNA per µL assay. Sensitivity regarding CFU 1.2, 5.2, 10, 5.6×101 3.8×102 µL, respectively. results suggest optimized method applied sensitive specific pathogens through single