title: Rapid detection and identification of viral and bacterial fish pathogens using a DNA array-based multiplex assay.
abstract: Fish diseases can be caused by a variety of diverse organisms, including bacteria, fungi, viruses and protozoa, and pose a universal threat to the ornamental fish industry and aquaculture. The lack of rapid, accurate and reliable means by which fish pathogens can be detected and identified has been one of the main limitations in fish pathogen diagnosis and fish disease management and has consequently stimulated the search for alternative diagnostic techniques. Here, we describe a method based on multiplex and broad-range PCR amplification combined with DNA array hybridization for the simultaneous detection and identification of all cyprinid herpesviruses (CyHV-1, CyHV-2 and CyHV-3) and some of the most important fish pathogenic Flavobacterium species, including F. branchiophilum, F. columnare and F. psychrophilum. For virus identification, the DNA polymerase and helicase genes were targeted. For bacterial identification, the ribosomal RNA gene was used. The developed methodology permitted 100% specificity for the identification of the target species. Detection sensitivity was equivalent to 10 viral genomes or less than a picogram of bacterial DNA. The utility and power of the array for sensitive pathogen detection and identification in complex samples such as infected tissue is demonstrated in this study.
authors: Laboratory for Process Microbial Ecology and Bioinspirational Management (PME&BIM), Consortium for Industrial Microbiology and Biotechnology (CIMB), Department of Microbial and Molecular Systems (M2S), K.U. Leuven Association, Lessius Mechelen, Campus De Nayer, Sint-Katelijne-Waver, Belgium.bli@scientiaterrae.org
authors: B
authors: B
authors: Lievens
authors: I
authors: I
authors: Frans
authors: C
authors: C
authors: Heusdens
authors: A
authors: A
authors: Justé
authors: S P
authors: SP
authors: Jonstrup
authors: F
authors: F
authors: Lieffrig
authors: K A
authors: KA
authors: Willems
mesh_terms: D000818:Animals; D001424:Bacterial Infections / Q000175:diagnosis / Q000662:veterinary*; D002347:Carps; D004265:DNA Helicases / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000235:genetics; D005393:Fish Diseases / Q000175:diagnosis*; D005398:Fisheries / Q000379:methods*; D045826:Flavobacteriaceae Infections / Q000175:diagnosis / Q000662:veterinary*; D005417:Flavobacterium / Q000235:genetics; D006564:Herpesviridae / Q000235:genetics; D006566:Herpesviridae Infections / Q000175:diagnosis / Q000662:veterinary*; D060885:Multiplex Polymerase Chain Reaction / Q000662:veterinary*; D020411:Oligonucleotide Array Sequence Analysis / Q000662:veterinary*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D012336:RNA, Ribosomal, 16S; D004259:DNA-Directed DNA Polymerase; D004265:DNA Helicases