title: Detection of North American orthopoxviruses by real time-PCR.
abstract: The prevalence of North American orthopoxviruses in nature is unknown and may be more difficult to ascertain due to wide spread use of vaccinia virus recombinant vaccines in the wild. A real time PCR assay was developed to allow for highly sensitive and specific detection of North American orthopoxvirus DNA in animal tissues and bodily fluids. This method is based on the amplification of a 156 bp sequence within a myristylated protein, highly conserved within the North American orthopoxviruses but distinct from orthologous genes present in other orthopoxviruses. The analytical sensitivity was 1.1 fg for Volepox virus DNA, 1.99 fg for Skunkpox virus DNA, and 6.4 fg for Raccoonpox virus DNA with a 95% confidence interval. Our assay did not cross-react with other orthopoxviruses or ten diverse representatives of the Chordopoxvirinae subfamily. This new assay showed more sensitivity than tissue culture tests, and was capable of differentiating North American orthopoxviruses from other members of Orthopoxvirus. Thus, our assay is a promising tool for highly sensitive and specific detection of North American orthopoxviruses in the United States and abroad.
authors.affiliation: Centers for Disease Control and Prevention, National Center for Emerging and Zoonotic Infectious Diseases, Division of High-Consequence Pathogens and Pathology, Poxvirus and Rabies Branch, Atlanta, GA 30333, USA. hfa5@cdc.gov
authors.forename: Nadia F
authors.initials: NF
authors.lastname: Gallardo-Romero
authors.forename: Andres
authors.initials: A
authors.lastname: Velasco-Villa
authors.forename: Sonja L
authors.initials: SL
authors.lastname: Weiss
authors.forename: Ginny L
authors.initials: GL
authors.lastname: Emerson
authors.forename: Darin S
authors.initials: DS
authors.lastname: Carroll
authors.forename: Christine M
authors.initials: CM
authors.lastname: Hughes
authors.forename: Yu
authors.initials: Y
authors.lastname: Li
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Karem
authors.forename: Inger K
authors.initials: IK
authors.lastname: Damon
authors.forename: Victoria A
authors.initials: VA
authors.lastname: Olson
mesh_terms: D000818:Animals; D003429:Cross Reactions; D009656:North America; D018155:Orthopoxvirus / Q000235:genetics* / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011213:Poxviridae Infections / Q000662:veterinary* / Q000821:virology; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods*