DAO 101:225-234 (2012)  -  doi:10.3354/dao02514

Real-time PCR assays targeting unique DNA sequences of fish-pathogenic Francisella noatunensis subspecies noatunensis and orientalis

Samuel Duodu1,*, Pär Larsson2, Andreas Sjödin2, Esteban Soto3, Mats Forsman2, Duncan J. Colquhoun1

1Section for Bacteriology, Norwegian Veterinary Institute, PO Box 750, Sentrum, 0106 Oslo, Norway
2Swedish Defense Research Agency (FOI), CBRN Defense and Security, Umeå, Sweden
3Ross University−School of Veterinary Medicine, PO Box 334, Basseterre, St. Kitts, West Indies

ABSTRACT: Specific identification and differentiation of the 2 subspecies of the fish pathogen Francisella noatunensis, namely, F. noatunensis subsp. noatunensis and F. noatunensis subsp. orientalis, remains a major diagnostic challenge. Following whole-genome sequencing and analysis of representatives of all major subclades of the genus Francisella, specific genomic regions were identified for each of the subspecies of this fish pathogen. Two specific real-time quantitative PCR assays, directed at hypothetical genes within these regions were developed. Specificity was confirmed by lack of signal and cross-reactivity with the closest relative, F. philomiragia, and other common bacterial fish pathogens. Both assays, used either as monoplex or multiplex, have a limit of detection of 10 genome equivalents. The quantitative sensitivity of the assays was not affected by the presence of kidney tissues or DNA from Atlantic cod Gadus morhua or tilapia Oreochromis sp.


KEY WORDS: Francisella noatunensis · Fish · DNA · qPCR


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Cite this article as: Duodu S, Larsson P, Sjödin A, Soto E, Forsman M, Colquhoun DJ (2012) Real-time PCR assays targeting unique DNA sequences of fish-pathogenic Francisella noatunensis subspecies noatunensis and orientalis. Dis Aquat Org 101:225-234

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