Development and evaluation of a multiplex PCR assay for simultaneous detection of Flavobacterium psychrophilum, Yersinia ruckeri and Aeromonas salmonicida subsp. salmonicida in culture fisheries.
10.4142/jvs.2010.11.3.235
- Author:
Ertan Emek ONUK
1
;
Alper CIFTCI
;
Arzu FINDIK
;
Yuksel DURMAZ
Author Information
1. Department of Diseases of Aquatic Animals, Faculty of Veterinary Medicine, Ondokuz Mayis University, 55139, Kurupelit, Samsun, Turkey.
- Publication Type:Original Article ; Evaluation Studies ; Research Support, Non-U.S. Gov't
- Keywords:
Aeromonas salmonicida subsp. salmonicida;
Flavobacterium psychrophilum;
multiplex PCR;
Yersinia ruckeri
- MeSH:
Aeromonas salmonicida/*genetics;
Animals;
DNA Primers/genetics;
Fish Diseases/*diagnosis/*microbiology;
Fishes;
Flavobacterium/*genetics;
Gram-Negative Bacterial Infections/diagnosis/*veterinary;
Polymerase Chain Reaction/methods/*veterinary;
Yersinia rucker/*genetics
- From:Journal of Veterinary Science
2010;11(3):235-241
- CountryRepublic of Korea
- Language:English
-
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.
