Detection of bacterial pathogens in aquaculture samples by DNA microarray analysis

Abstract

Rapid, sensitive, and specific detection of pathogens is important for effective disease control in aquaculture. However, the use of conventional microbiological methods for pathogen detection is often limited by the time required to complete the assays. The aim of this study was to develop a DNA microarray assay to detect 8 species of pathogenic bacteria, i.e., Vibrio harveyi, V. alginolyticus, V. parahaemolyticus, V. anguillarum, V. cholerae, Nocardia seriolae, Aeromonas hydrophila, and Streptococcus iniae, in cultured aquatic animals. Probes were designed according to a variable region of the bacterial 16S rRNA gene and gryB sequences, and 23 probes were finally selected by hybridization with 25 bacterial type cultures. The microarray sensitivity for V. anguillarum detection in healthy fish blood was ≥100CFU/mL. Pathogenic bacteria in blood samples from 21 diseased aquacultural samples were analyzed in a blinded fashion by using the microarray, conventional culture, and 16S rRNA gene sequencing methods; the results of the microarray assay agreed well with those of the conventional culture and 16S rRNA gene sequencing methods. The results of this study show that the microarray method described is suitable for preliminary diagnoses or for confirmation of the main potential pathogens in aquatic animals.