Rapid and differential diagnosis of foot-and-mouth disease, swine vesicular disease, and vesicular stomatitis by a new multiplex RT-PCR assay

A highly sensitive and specific one-step multiplex RT-PCR assay has been developed and standardised for the simultaneous and differential detection of the most important vesicular viruses affecting livestock: foot-and-mouth disease virus (FMDV), swine vesicular disease virus (SVDV), and vesicular stomatitis virus (VSV). The method uses three primer sets, each one specific for the corresponding virus, selected to detect of all serotypes of FMD and VS. The detection range was confirmed by examination of a collection of 31 isolates of the three target viruses. The specificity of the assay was also demonstrated by testing other related viruses, uninfected cell line cultures and healthy pig tissues. The testing of blood and serum samples from animals infected experimentally proved that the method can be useful for early diagnosis of the diseases, even before the first vesicular lesions are visualized in the infected pigs. An assessment of the performance of the multiplex RT-PCR was carried out using a panel of more than 100 samples from animals infected experimentally, showing the suitability of the method for a rapid (less than 6h), sensitive and specific differential diagnosis in clinical samples. Additionally, a uniplex RT-PCR for VSV, that amplifies the two viral serotypes, was also developed and tested as a rapid tool for the diagnosis of this vesicular disease.