4.4 Article

A duplex real-time PCR assay based on TaqMan technology for simultaneous detection and differentiation of canine adenovirus types 1 and 2

Journal

JOURNAL OF VIROLOGICAL METHODS
Volume 234, Issue -, Pages 1-6

Publisher

ELSEVIER SCIENCE BV
DOI: 10.1016/j.jviromet.2016.03.011

Keywords

Dog; Canine adenovirus 1; Canine adenovirus 2; Duplex real-time PCR

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Canine adenoviruses are a major cause of disease in dogs, coyotes, red foxes and wolves, as well as in other carnivores and marine mammals. Canine adenovirus type 1 (CAdV-1) and canine adenovirus type 2 (CAdV-2) cause infectious canine hepatitis (ICH) and infectious tracheobronchitis (ITB), respectively. In this study, a duplex real-time PCR assay for simultaneous detection and characterisation of CAdV-1 and CAdV-2 was developed by using a single primer pair and virus-specific probes. The assay was validated testing standard DNAs produced on purpose and clinical samples of various matrices known to be positive for CAdV-1, CAdV-2 or both viruses. Precise calculation of DNA loads in samples containing a wide range of viral amounts was allowed by generating a standard curve for absolute quantification. The assay was proven to be highly specific, since no cross-reactions with the different CAdV type was observed, and sensitive, being able to detect less than 10 copies of CAdV-1/CAdV-2 DNA. The low intra-assay and interassay coefficient of variations demonstrated a high repeatability, thus confirming the potential use of this assay for quantitative detection of CAdV-1 and CAdV-2 for rapid diagnosis and epidemiological investigations. (C) 2016 Elsevier B.V. All rights reserved.

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