Additional file 1: of Correction to: Dual RNA-seq reveals viral infections in asthmatic children without respiratory illness which are associated with changes in the airway transcriptome

Table S2. Gene count table for 48 RNA-seq nasal brushing subjects from GALA cohort. (XLSX 5732 kb

Additional file 2: Figure S1. of Dual RNA-seq reveals viral infections in asthmatic children without respiratory illness which are associated with changes in the airway transcriptome

Study design. Figure S2. Genome coverage for viruses detected in Ion Torrent Proton sequenced samples. Figure S3. Comparison of in vivo and in vitro log2 gene expression fold changes by infection status. Figure S4. WGCNA gene correlation dendrogram showing nine detected gene co-expression modules. Figure S5. H&E staining of nasal airway epithelial brushing cell cytospin from a viral-infected subject. (PDF 30208 kb

Additional file 1: of Dual RNA-seq reveals viral infections in asthmatic children without respiratory illness which are associated with changes in the airway transcriptome

Table S1. Clinical characteristics of subjects. Table S2. Gene count table for 48 RNA-seq nasal brushing subjects from GALA cohort. Table S3. Virus-High DESeq differential expression analysis. Table S4. DAVID enrichment for Virus High DEGs. Table S5. Gene count table for HRV in vitro stimulation dataset. Table S6. HRV in vitro DEGs. Table S7. Virus Low Differential Expression Analysis. Table S8. WGCNA gene2module. Table S9. DAVId for modules. Table S10. immune cell enrichments for modules. (XLS 18739 kb

Comparison of CNV allele frequency between Hutterites and CEU.

<p>A histogram of the difference in allele frequency for the non-reference allele between Hutterites and the CEU individuals from the 1000 Genomes Project is shown. On the x-axis are bins for the absolute value of the non-reference allele frequency in the Hutterites minus the allele frequency in CEU. The y-axis represents the number of CNVs in each bin.</p

Association results of 593 CNVs to asthma.

<p>CNVs for association testing in the full Hutterite pedigree were identified in the 16 sequenced genomes. The points for these CNVs are colored based on the results of the whole-genome sequencing to represent whether the variant was observed in cases only (red), control individuals only (blue), or in both case and control individuals (gray). The genomic position is represented on the x-axis and the −log₁₀(p-value) of the nominal association of each CNV to asthma in the full Hutterite pedigree is on the y-axis.</p

Identification of an intronic deletion in <i>NEDD4L</i> associated with asthma.

<p>(<b>A</b>) A representation of the location of the 6 kbp deletion. This deletion occurs in an intron shared by all reported transcripts of this gene (blue). The deletion was identified from paired-end sequence reads and validated by array CGH as shown for one of the sequenced trios. The log₂ ratios of the probes in this region are shown as vertical bars with a log₂ ratio of zero represented by the horizontal line. The red vertical bars in the child and mother indicate negative log₂ ratios and confirm the deletion. (<b>B</b>) The frequency of this deletion in multiple populations is shown in the bar graph with the deletion allele frequency on the y-axis. The error bars represent the standard error on the allele frequency based on the binomial distribution. The Hutterite case (black) and control (gray) frequencies were determined by array CGH; the frequencies for the other populations are as reported by the 1000 Genomes Project.</p

Whole-Genome Sequencing of Individuals from a Founder Population Identifies Candidate Genes for Asthma

<div><p>Asthma is a complex genetic disease caused by a combination of genetic and environmental risk factors. We sought to test classes of genetic variants largely missed by genome-wide association studies (GWAS), including copy number variants (CNVs) and low-frequency variants, by performing whole-genome sequencing (WGS) on 16 individuals from asthma-enriched and asthma-depleted families. The samples were obtained from an extended 13-generation Hutterite pedigree with reduced genetic heterogeneity due to a small founding gene pool and reduced environmental heterogeneity as a result of a communal lifestyle. We sequenced each individual to an average depth of 13-fold, generated a comprehensive catalog of genetic variants, and tested the most severe mutations for association with asthma. We identified and validated 1960 CNVs, 19 nonsense or splice-site single nucleotide variants (SNVs), and 18 insertions or deletions that were out of frame. As follow-up, we performed targeted sequencing of 16 genes in 837 cases and 540 controls of Puerto Rican ancestry and found that controls carry a significantly higher burden of mutations in <i>IL27RA</i> (2.0% of controls; 0.23% of cases; nominal p = 0.004; Bonferroni p = 0.21). We also genotyped 593 CNVs in 1199 Hutterite individuals. We identified a nominally significant association (p = 0.03; Odds ratio (OR) = 3.13) between a 6 kbp deletion in an intron of <i>NEDD4L</i> and increased risk of asthma. We genotyped this deletion in an additional 4787 non-Hutterite individuals (nominal p = 0.056; OR = 1.69). <i>NEDD4L</i> is expressed in bronchial epithelial cells, and conditional knockout of this gene in the lung in mice leads to severe inflammation and mucus accumulation. Our study represents one of the early instances of applying WGS to complex disease with a large environmental component and demonstrates how WGS can identify risk variants, including CNVs and low-frequency variants, largely untested in GWAS.</p></div

Overview of sequenced genomes.

<p>Ideograms of the autosomes and X chromosome are shown. Red bars represent segments of autozygosity observed in five individuals with asthma (parents of the sequenced trios) and yellow triangles represent asthma-specific gene-disruptive SNVs or indels. The regions of autozygosity in multiple cases on chromosomes 1q and 5q were also observed to be autozygous in at least one control individual.</p

Association of intronic deletion in <i>NEDD4L</i> with asthma in additional populations.

<p>*Odds ratio (OR).</p>#<p>Combined association statistics were obtained using the Cochran–Mantel–Haenszel test.</p

Summary of whole-genome sequencing.

<p>*Mean coverage of the genome (NCBI build 36) based on mapped reads.</p>#<p>Identified from read-pair mappings.</p>†<p>Identified with read-depth.</p>‡<p>Used as “reference” sample in the read-depth approach.</p>§<p>Bronchial hyperresponsiveness (BHR).</p