title: Development and evaluation of a multiplex PCR assay for simultaneous detection of Flavobacterium psychrophilum, Yersinia ruckeri and Aeromonas salmonicida subsp. salmonicida in culture fisheries.
abstract: Bacterial cold water disease, enteric red mouth disease and frunculosis are the common bacterial diseases of fish worldwide. The etiologic agents of these diseases are Flavobacterium (F.) psychrophilum, Yersinia (Y.) ruckeri and Aeromonas (A.) salmonicida subsp. salmonicida, respectively. In this study, a multiplex polymerase chain reaction (m-PCR) method with YER8/10-Fer3/4-FP1/3 primer pairs which can identify these fish pathogens simultaneously was developed and optimized. In optimized conditions, neither false specific nor nonspecific amplification occurred. The detection limits of the m-PCR method using DNA extracts from dilutions of pure cultures of bacteria were 35 pg for Y. ruckeri and F. psychrophilum and 70 pg for A. salmonicida subsp. salmonicida. It was determined that 15 CFU Y. ruckeri and F. psychrophilum and 30 CFU A. salmonicida subsp. salmonicida could be detected by m-PCR developed using genomic DNA extracted from dilutions of the suspensions. The detection limits in the presence of tissue debris were 125 CFU for Y. ruckeri and F. psychrophilum and 250 CFU for A. salmonicida subsp. salmonicida. In conclusion, we submit that the m-PCR method developed and optimized in this study can be used for accurate and rapid identification of these bacteria.
authors: Department of Diseases of Aquatic Animals, Faculty of Veterinary Medicine, Ondokuz Mayis University, 55139 Kurupelit, Samsun, Turkey.
authors: Ertan Emek
authors: EE
authors: Onuk
authors: Alper
authors: A
authors: Ciftci
authors: Arzu
authors: A
authors: Findik
authors: Yuksel
authors: Y
authors: Durmaz
mesh_terms: D048409:Aeromonas salmonicida / Q000235:genetics*; D000818:Animals; D017931:DNA Primers / Q000235:genetics; D005393:Fish Diseases / Q000175:diagnosis* / Q000382:microbiology*; D005399:Fishes; D005417:Flavobacterium / Q000235:genetics*; D016905:Gram-Negative Bacterial Infections / Q000175:diagnosis / Q000662:veterinary*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D044124:Yersinia ruckeri / Q000235:genetics*
chemical_list: D017931:DNA Primers