Heatmap describing number of contigs identified in each pool after their characterization and classification into taxonomic groups.

<p>Rows correspond to pooled samples whilst columns to families mapped at least to one sample. Numbers within each cell represent the number of sequences that had at least a positive BLAST hit to into known species and passed all the selection criteria. The colours range from yellow to red (low to high abundance respectively); green means that sequences were not detected for that group.</p

Summary of the sequences produced for each pool of serum samples in the sequencing experiment.

<p>All read counts correspond to total values, and the paired-reads real counts are half the values shown in the table. PE: paired-end reads; SE: single-end reads.</p

Summary information for contigs longer than 1,500 bp that were found in the pooled samples and assigned to the <i>Anelloviridae</i> family.

<p>The number and letter codes from the first column (Code) correspond to those in the blank bullets shown on some of the branches of the phylogenetic tree from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0185911#pone.0185911.g003" target="_blank">Fig 3</a>. Those without codes were placed directly on the tree, as they defined new branches.</p

Summary of similarity searches for those detected from the HEV and Ai+ImSP pools.

<p>The first column corresponds to the numbers in the black bullets shown on some of the branches of the <i>Hepeviridae</i> phylogenetic tree from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0185911#pone.0185911.g002" target="_blank">Fig 2</a>.</p

Phylogenetic tree of <i>Hepeviridae</i> based on complete genomes, including the main members of genotype 3.

<p>Numbers in blank bullets correspond to contigs identified in the HEV and Ai+ImSP pools (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0185911#pone.0185911.t002" target="_blank">Table 2</a>); they are located beside the reference sequence where specific individual alignments of sequenced fragments over the same region in the reference sequences generated an equivalent tree topology (further results available from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0185911#pone.0185911.s001" target="_blank">S1 Supporting Information</a>). Labels within the square brackets define the species subtype. Small numbers on the tree branches show the bootstrap score of those branches.</p

Identification of sapovirus GV.2, astrovirus VA3 and novel anelloviruses in serum from patients with acute hepatitis of unknown aetiology

<div><p>Hepatitis is a general term meaning inflammation of the liver, which can be caused by a variety of viruses. However, a substantial number of cases remain with unknown aetiology. We analysed the serum of patients with clinical signs of hepatitis using a metagenomics approach to characterize their viral species composition. Four pools of patients with hepatitis without identified aetiological agents were evaluated. Additionally, one pool of patients with hepatitis E (HEV) and pools of healthy volunteers were included as controls. A high diversity of anelloviruses, including novel sequences, was found in pools from patients with hepatitis of unknown aetiology. Moreover, viruses recently associated with gastroenteritis as sapovirus GV.2 and astrovirus VA3 were also detected only in those pools. Besides, most of the HEV genome was recovered from the HEV pool. Finally, GB virus C and human endogenous retrovirus were found in the HEV and healthy pools. Our study provides an overview of the virome in serum from hepatitis patients suggesting a potential role of these viruses not previously described in cases of hepatitis. However, further epidemiologic studies are necessary to confirm their contribution to the development of hepatitis.</p></div

Upregulation of proinflammatory genes in EB capsules compared to clear capsules.

<p>Expression levels of RNA in six types of brain tissue samples, uninfected brain tissue distant from cysts (no cysts), clear pericystic brain tissue in untreated pigs (UT Clear) and at 48h post treatment (PZQ 48 Clear) and EB capsules in untreated pigs (UT Blue) and at 48h (PZQ 48 h Blue) and 120h post treatment (PZQ 120 h Blue) were quantified using real-time reverse transcriptase polymerase chain reaction (RT-PCR) assays. Gene expression levels are depicted as fold increase of studied RNA over reference (housekeeping) RNA (18S rRNA), TNF-α (A; number [n] of samples for the indicated types of capsules are shown below each bar) and IL-6 (B). The bars represent means and CI₉₅ for the group. Statistically significant differences in levels of gene expression are indicated by asterisks (p<0.05, Mann-Whitney U test). Asterisks represent level of significance: *: p<0.05; **: p<0.01; ***: p<0.005.</p

Increased expression of fibrosis/granuloma promoting genes in EB and clear capsules.

<p>Methods and tissues examined are identical to those analyzed in Figs. <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003577#pntd.0003577.g003" target="_blank">3</a>–<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003577#pntd.0003577.g005" target="_blank">5</a>. Shown are four genes representative of fibrosis and granuloma promoting molecules: MMP1 (A; number [n] of samples for the indicated types of capsules are shown below each bar; n = 7, 6, 3, 6, 6, respectively), MMP9 (B), TIMP1 (C) and TIMP2 (D). Statistically significant differences in levels of gene expression are indicated by asterisks (p<0.05, Mann-Whitney U test). Asterisks represent level of significance: *: p<0.05; **: p<0.01; ***: p<0.005.</p

Upregulation of both Th1 and Th2 markers, IFN-γ and IL-13, in blue capsules following PZQ treatment.

<p>Expression levels of RNA in brain tissues and EB-stained and clear capsules for IFN-γ (A; number [n] of samples for the indicated types of capsules are shown below each bar), and IL-13 (B). In each graph, the bars represent means and CI₉₅ for the group. Statistically significant differences in levels of gene expression are indicated by asterisks (p<0.05, Mann-Whitney U test). Asterisks represent level of significance: *: p<0.05; **: p<0.01; ***: p<0.005.</p

EB capsules show increased inflammation and cyst wall damage compared to clear capsules.

<p>Cysts were harvested from the brains of infected untreated pigs (n = 3; see <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003577#sec002" target="_blank">Methods</a>) or pigs treated with PZQ 48h (n = 4) or 120h (n = 4) earlier. For each cyst, scores reflecting the degree and extent of the inflammatory cell infiltrates (A) and cyst wall damage (B) at 48h (PZQ 48h) and 120h (PZQ 120h) after PZQ treatment (see <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003577#sec002" target="_blank">Methods</a> and <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003577#pntd.0003577.s002" target="_blank">S1 Fig.</a>) are shown (open squares refer to clear capsules and closed squares, to EB capsules, bar = mean). Statistical significance in comparison between groups is represented by asterisks (*: p<0.05; **: p<0.01; ***: p<0.005).</p