Restriction fragment length polymorphism (RFLP) analysis of open reading frame 5 was developed for typing of Czech strains of porcine reproductive and respiratory syndrome virus (PRRSV). e set of restriction enzymes Acc I, Hae II and SnaB I allowed the differentiation of heterogeneous Czech strains of PRRSV clustered separately in the phylogenetic tree. e high-passage strain V-502 (164) was also differentiated from its parent strain V-502. e same restriction enzymes could distinguish the European-type vaccine strains Porcilis PRRS and Pyrsvac-183, registered in Czech Republic, from the Czech field isolates. e published ORF5 nucleotide sequences allowed us to presume that it will also be possible to distinguish most of European field strains from vaccine strains. PCR-based RFLP analysis can become a valuable tool in epidemiological studies of PRRSV in Europe.

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Abstract. Swine influenza is an acute respiratory disease of swine caused by type A influenza viruses. Before 1998, mainly ''classical'' H1N1 swine influenza viruses (SIVs) were isolated from swine in the United States. Since then, antigenically distinct reassortant H3 and H1 SIVs have been identified as causative agents of respiratory disease in pigs on US farms. Improvement in SIV diagnostics is needed in light of the recently observed rapid evolution of H1 and H3 SIVs and their zoonotic potential. To address this need, real-time reverse transcription-polymerase chain reaction (RT-PCR) assays for the detection of SIVs were developed. A highly sensitive matrix (M) gene-based RT-PCR assay that is able to detect both the H1 and H3 subtypes of SIVs, with a sensitivity per reaction of approximately 2 copies of in vitro-generated M-specific negative-sense RNA molecules and approximately 0.05 TCID 50 in lung lavage of experimentally SIV-infected pigs, was established. This RT-PCR assay can be performed within a few hours and showed a sensitivity of 94% and a specificity of 85% when compared with virus isolation. In addition, H1-, H3-, N1-, and N2-specific primer and probe sets were designed for use in the differentiation of different SIV subtypes. The hemagglutinin (H)-and neuraminidase (N)-specific primer and probe sets were less sensitive than the M-specific assay, although they were found to be specific for their respective viral genes and able to distinguish between their respective SIV subtypes.