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.
Highlights
Foot-and-mouth disease (FMD) is a highly contagious vesicular disease of wild and domestic cloven-hoofed animals, cattle, sheep, pigs and goats
Outbreaks can occur in countries that are normally free of FMD: including recently Japan and Korea (2000 and 2010), the United Kingdom (UK: 2001 and 2007), France and The Netherlands (2001) plus Bulgaria (2010–2011)
FMD viruses (FMDVs) isolates were selected from archival stocks held in the WRLFMD (Pirbright Institute, UK) repository at −20 ◦C and contemporary submissions from within the serotype O PanAsia-2 and serotype A Iran-05 lineages, respectively, serotype Asia-1 subgroups 1, 2 and 6, plus representative FMDV-Genome Detected samples of undetermined serotype (FMDV-GD: Ag-ELISA and virus isolation (VI) negative)
Summary
Foot-and-mouth disease (FMD) is a highly contagious vesicular disease of wild and domestic cloven-hoofed animals, cattle, sheep, pigs and goats. It is the most economically important viral disease of domesticated livestock throughout the world today, being endemic in many countries of Africa, Asia and South (D.P. King). There are seven immunologically distinct serotypes: O, A, C, SAT (Southern African Territories) 1, SAT 2, SAT 3 and Asia-1 which encompass a diverse antigenic spectrum of virus strains. The seven serotypes are not distributed around the world (Knowles and Samuel, 2003). Serotypes O and A are widely disseminated but type C viruses have not been detected since 2004, while Asia-1, despite having been detected in Greece, is mainly confined to the Asian continent
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