Comparison between nucleotide sequences of selected FMDV samples and the primers and probes.

<p>Panel (A) Sequences of serotype A FMDV samples indicating mismatches with primers and probe of the Reid et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>] serotype A-specific assay. Samples numbered 1495, 1364 and 2809 were not detected in the serotype A-specific assays described by Reid and colleagues [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>]. Panel (B) Sequences of serotype A FMDVs indicating mismatches with primers and probe of the new serotype A-specific assay. Samples numbered 1364 and 2809 were not detected in the new assay. Panel (C) Nucleotide sequences of serotype O FMDV samples indicating mismatches with primer and probe sequences of the Reid et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>] serotype O-specific assay. The samples 1343 and Nzm21 were not detected in this assay.</p

Summary of the RT-qPCR results using serotype O-, A- and Asia-1-specific and pan-FMDV 5'-UTR and 3D assays.

<p>* two samples had high Ct values (38.9 and 37.9)</p><p>** two samples had high Ct values (38.1 and 38.9)</p><p>*** includes sample positive for both serotypes A and Asia-1</p><p>Summary of the RT-qPCR results using serotype O-, A- and Asia-1-specific and pan-FMDV 5'-UTR and 3D assays.</p

Diagnostic sensitivity and specificity of FMDV serotype O specific RT-qPCR assays.

<p>The C_T values obtained in the different RT-qPCR assays for the indicated serotype O virus samples are shown. The genetic relationships between the various strains from West Eurasia, based on VP1 coding sequences, are also indicated.</p

Comparison between nucleotide sequences of selected FMDV samples and the primers and probes.

<p>Panel (A) Sequences of serotype A FMDV samples indicating mismatches with primers and probe of the Reid et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>] serotype A-specific assay. Samples numbered 1495, 1364 and 2809 were not detected in the serotype A-specific assays described by Reid and colleagues [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>]. Panel (B) Sequences of serotype A FMDVs indicating mismatches with primers and probe of the new serotype A-specific assay. Samples numbered 1364 and 2809 were not detected in the new assay. Panel (C) Nucleotide sequences of serotype O FMDV samples indicating mismatches with primer and probe sequences of the Reid et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>] serotype O-specific assay. The samples 1343 and Nzm21 were not detected in this assay.</p

Sequences of primer and probe sets designed for serotyping of West Eurasian FMDVs using RT-qPCR assays.

<p>Sequences of primer and probe sets designed for serotyping of West Eurasian FMDVs using RT-qPCR assays.</p

Comparison of amplification curves of the serotype-specific RT-qPCR assays.

<p>(A) Amplification curves for the six selected serotype O samples obtained with the 1^st generation assay [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>] (+ symbols) and the 2^nd generation serotype O-specific assay described here (round symbols). (B) Amplification curves for the six selected serotype A samples obtained with the 1^st generation assay [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>] (+ symbols) and the 2^nd generation serotype A-specific assay described here (round symbols). (C) Amplification curves for the six selected serotype Asia-1 samples obtained with the 1^st generation assay [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135559#pone.0135559.ref033" target="_blank">33</a>] (+ symbols) and the 2^nd generation serotype Asia-1-specific assay described here (round symbols).</p

Diagnostic sensitivity and specificity of FMDV serotype A-specific RT-qPCR assays.

<p>The C_T values obtained in the different RT-qPCR assays for the indicated serotype A virus samples are shown. The genetic relationships between the various strains from West Eurasia, based on VP1 coding sequences, are also indicated. Note, sample 8 is marked with an * to indicate this sample contained two different serotypes of FMDV.</p

Titration of anti-FMDV serotype Asia 1 antibodies induced following infection with rescued serotype Asia 1 FMDV.

<p>Values are reciprocals of the highest serum dilutions yielding positive results in the blocking ELISA on the indicated day post inoculation (dpi). A (-) indicates samples which were scored negative in the spot test (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014621#pone-0014621-g004" target="_blank">Figure 4B</a>) and N.D. indicates when samples were not collected. Calves 1C1 and 1C5 were inoculated with the rescued serotype Asia 1 FMDV (Asia-1/BAM/AFG/L590/2009) on day 0 and the other calves were maintained in contact with them.</p

Rescue of infectious FMDV from RNA samples isolated from bovine tongue epithelium from suspected cases of FMD in Afghanistan (AFG) and Pakistan (PAK).

<p>The presence of FMDV RNA was determined using real time RT-PCR assays which detect the 5′ UTR or the 3D coding sequence. The serotype of the virus from which the RNA was obtained was determined from VP1 coding sequence determination and comparison with other FMDV strains using BLAST. The success of virus rescue was determined by the presence of CPE (indicated by +++).</p

Titration of anti-FMDV serotype O antibodies induced following infection with rescued serotype O FMDV.

<p>Values are reciprocals of highest serum dilutions yielding positive results in the blocking ELISA on the indicated day post inoculation (dpi). A (-) indicates samples which were scored negative in the spot test (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014621#pone-0014621-g005" target="_blank">Figure 5B</a>) and N.D. indicates when samples were not collected (calf 2C6 was euthanized on day 4). Calves 2C1 and 2C5 were inoculated with serotype O FMDV (O/ISL/PAK/L1573/2009) on day 0 and the other calves were maintained in contact with them.</p