Alignment of the inferred aa sequences of the SH gene.

<p>The potential neurovirulence sites are marked in boxes. Period (.) denotes identity with the majority sequence; (-) denotes areas that were not determined; (*) represents Genotype-strain ID.</p

Amino acid changes associated with neurovirulence in the HN or SH gene.

<p>Amino acid changes associated with neurovirulence in the HN or SH gene.</p

Alignment of the inferred partial aa sequences of the HN gene.

<p>The potential glycosylation sites are shown in shadow and the potential neurovirulence sites are marked in boxes. Period (.) denotes identity with the uppermost sequence; (-) denotes areas that were not determined; (*) represents Genotype-strain ID.</p

Patients and specimens in the study.

<p>Note: OF, Oral fluid; CSF, cerebrospinal fluid; TS, throat swab; TCF, tissue culture fluid; P, parotitis; N, neurological complications; NK: not known; MMR, measles-mumps-rubella vaccine; UK, United Kingdom.</p>*<p>Dominican Republic linked re-infection case in Sweden, childhood infected (Dr. Kari Johansen, Swedish Institute for Infectious Disease Control).</p

The sequences of the primers and probe for HEV, HEV71, and CVA16.

<p>Note:</p><p>1. “a” indicates the sequence position of the primers and probes referring to strain BrCr of HEV71 (GenBank accession number U22521); “b” indicates the sequence position of the primers and probe referring to strain G-10 of CVA16 (GenBank accession number U05876).</p><p>2. The lengths of real-time RT-PCR products for HEV71, CVA16, and HEV were 76, 104, and 194 nt, respectively.</p><p>3. The primers and probes of HEV71, CVA16, and HEV have been applied for national invention patent in China, and the numbers of the patents are ZL200810063097.5, ZL200810121454.9, and ZL200810121453.4, respectively.</p

The real-time RT-PCR results of HEV71, CVA16, and HEVs for different types of HFMD clinical specimens.

<p>Note:</p><p>1. *Among 48 HEVs-positive specimens, one specimen was positive for HEVs, but negative for HEV71 or CVA16, which was identified as Echo11 by virus isolation, RT-PCR, and sequencing for the VP1 gene.</p><p>2. For the positive specimens for HEV71 or CVA16, the results of real-time RT-PCR for HEVs were also positive.</p><p>3. A total of 85 HEV71 and 16 CVA16 were identified using RT-PCR method among the 213 HFMD clinical specimens, which were lower than those of the real-time RT-PCR assay.</p><p>4. The multiplex real-time RT-PCR assay was shown to exhibit 100% specificity in detecting HEV71 and CVA16, which was confirmed by sequencing.</p

Monthly distribution of the 58 hospitalized children with HPIV-3 infection.

<p>Monthly distribution of the 58 hospitalized children with HPIV-3 infection.</p

Human parainfluenza viruses real-time reverse-transcription polymerase chain reaction (RT-PCR) panel primer and probe sequences.

a<p>Primer and probe sequences for HPIV-1 and HPIV-2 are from F. Watzinger et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043893#pone.0043893-Watzinger1" target="_blank">[46]</a>.</p>b<p>Primer and probe sequences for HPIV-3 are fromKate E. Templeton et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043893#pone.0043893-Templeton1" target="_blank">[47]</a>.</p>c<p>Labeled at the 5′ end with FAM and terminally quenched at the 3′ end with Black Hole Quencher-1.</p

Co-infections in HPIV-3 positive Samples.

<p>Co-infections in HPIV-3 positive Samples.</p

Genetic distances among HPIV-3 isolates.

<p>Nucleotide sequence diversity was calculated using Kimura 2-parameter method. The threshold value of 0.045 that divided comparisons between members of the same (phylogenetically defined) cluster and between different clusters is shown as a dashed line.</p