Development and Application of a Quantitative Real‐time Polymerase Chain Reaction Assay for the Detection of Aeromonas salmonicida
2017
Du, Yishuai | Liu, Ying | Xiao, Peng | Meng, Lingjie | Liu, Pengfei
A rapid, economical, specific, and sensitive quantitative real‐time polymerase chain reaction (qPCR) assay coupled with SYBR Green I chemistry was developed for the quantitative detection of Aeromonas salmonicida from farmed Atlantic salmon, Salmo salar, with the symptoms of furunculosis. The set of primers designed from the virulence array protein (vapA) gene was specific to A. salmonicida. Compared with the conventional PCR, qPCR had a lower detection limit of 5.6 copies of the positive plasmids. The standard curve, which showed the relationship between the copies of A. salmonicida and its quantification cycle (Cq) value, could be described as follows: log (copies of A. salmonicida) = −0.3213 Cq + 10.721. The quantitative detection of copies of A. salmonicida in different tissues of the moribund Atlantic salmon showed that A. salmonicida could be detected in all tissues; the spleen contained the largest number of A. salmonicida and then the kidney. These results suggest that the qPCR assay reported here is a specific, sensitive, and quantitative method for detecting A. salmonicida. It can be used for the routine tests of A. salmonicida in local aquaculture enterprise and for the research of infection routes of A. salmonicida to Atlantic salmon.
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