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      Development of tailored real-time RT-PCR assays for the detection and differentiation of serotype O, A and Asia-1 foot-and-mouth disease virus lineages circulating in the Middle East.

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          Abstract

          Rapid and accurate diagnosis is essential for effective control of foot-and-mouth disease (FMD). In countries where FMD is endemic, identification of the serotypes of the causative virus strains is important for vaccine selection and tracing the source of outbreaks. In this study, real-time reverse transcription polymerase chain reaction (rRT-PCR) assays using primer/probe sets designed from the VP1 coding region of the virus genomes were developed for the specific detection of serotype O, A and Asia-1 FMD viruses (FMDVs) circulating in the Middle East. These assays were evaluated using representative field samples of serotype O strains belonging exclusively to the PanAsia-2 lineage, serotype A strains of the Iran-05 lineage and serotype Asia-1 viruses from three relevant sub-groups. When RNA extracted from archival and contemporary field strains was tested using one- or two-step rRT-PCR assays, all three primer/probe sets detected the RNA from homotypic viruses and no cross-reactivity was observed with heterotypic viruses. Similar results were obtained using both single- and multiplex assay formats. Using plasmid standards, the minimum detection level of these tests was found to be lower than two copies. The results illustrate the potential of tailored rRT-PCR tools for the detection and categorization of viruses circulating in the Middle East belonging to distinct subgroups of serotypes O, A and Asia-1. These assays can also overcome the problem of serotyping samples which are found positive by the generic rRT-PCR diagnostic assays but negative by virus isolation and antigen-detection ELISA which would otherwise have to be serotyped by nucleotide sequencing. A similar approach could be used to develop serotyping assays for FMDV strains circulating in other regions of the world.

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          Author and article information

          Journal
          J. Virol. Methods
          Journal of virological methods
          1879-0984
          0166-0934
          Oct 2014
          : 207
          Affiliations
          [1 ] Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, United Kingdom.
          [2 ] Khorasan Razavi Central Veterinary Laboratory, Iran Veterinary Organization, Iran.
          [3 ] National Veterinary Institute, Technical University of Denmark, Lindholm, 4771 Kalvehave, Denmark.
          [4 ] Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, United Kingdom. Electronic address: donald.king@pirbright.ac.uk.
          Article
          S0166-0934(14)00258-4
          10.1016/j.jviromet.2014.07.002
          25016065
          Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

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