title: Molecular double-check strategy for the identification and characterization of Suid herpesvirus 1.
abstract: Large scale vaccination with glycoprotein E (gE)-deleted marker vaccines and the rapid and reliable differentiation of wild-type and marker vaccine strains are important aspects in eradication programs for Suid herpesvirus 1 [SuHV-1, syn. Aujeszky's disease virus (ADV) or pseudorabies virus (PrV)]. Therefore, two multiplex real-time PCR (qPCR) assays for the genetic differentiation of wild-type and gE-deleted vaccine SuHV-1 strains have been developed. In the first multiplex qPCR SuHV-1 gB-gene specific detection was combined with a gE-gene specific assay and an internal control based on heterologous DNA. In the second system, a SuHV-1 UL19 (major capsid protein gene) assay, a different gE-gene specific assay and an internal control based on the beta-actin gene were combined. The gB-gene, UL19 as well as both gE-gene specific assays had an analytical sensitivity of less than 10 genome copies per reaction in the respective multiplex approaches. A series of reference strains including field isolates obtained from domestic and wild animals, and gE-deleted SuHV-1 were reliably detected, while genetically related non-SuHV-1 herpesviruses tested negative. Both newly developed triplex SuHV-1-specific qPCR assays are specific and sensitive methods for the rapid genetic differentiation of wild-type viruses and gE-deleted vaccine strains in a single reaction.
authors.affiliation: Institute of Diagnostic Virology, Friedrich-Loeffler-Institut, Südufer 10, D-17493 Greifswald-Insel Riems, Germany.
authors.forename: Kerstin
authors.initials: K
authors.lastname: Wernike
authors.affiliation: Institute of Diagnostic Virology, Friedrich-Loeffler-Institut, Südufer 10, D-17493 Greifswald-Insel Riems, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Beer
authors.affiliation: Institute of Molecular Virology and Cell Biology, Friedrich-Loeffler-Institut, Südufer 10, D-17493 Greifswald-Insel Riems, Germany.
authors.forename: Conrad M
authors.initials: CM
authors.lastname: Freuling
authors.affiliation: Institute of Molecular Virology and Cell Biology, Friedrich-Loeffler-Institut, Südufer 10, D-17493 Greifswald-Insel Riems, Germany.
authors.forename: Barbara
authors.initials: B
authors.lastname: Klupp
authors.affiliation: Institute of Molecular Virology and Cell Biology, Friedrich-Loeffler-Institut, Südufer 10, D-17493 Greifswald-Insel Riems, Germany.
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Mettenleiter
authors.affiliation: Institute of Molecular Virology and Cell Biology, Friedrich-Loeffler-Institut, Südufer 10, D-17493 Greifswald-Insel Riems, Germany. Electronic address: thomas.mueller@fli.bund.de.
authors.forename: Thomas
authors.initials: T
authors.lastname: Müller
authors.affiliation: Institute of Diagnostic Virology, Friedrich-Loeffler-Institut, Südufer 10, D-17493 Greifswald-Insel Riems, Germany.
authors.forename: Bernd
authors.initials: B
authors.lastname: Hoffmann
mesh_terms: D017353:Gene Deletion; D011558:Herpesvirus 1, Suid / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014765:Viral Vaccines / Q000235:genetics; D014773:Virology / Q000379:methods*
chemical_list: D014765:Viral Vaccines
keywords: Aujeszky's disease; Multiplex PCR; Pseudorabies; Pseudorabies virus; Real-time PCR; Suid herpesvirus 1