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Transcriptome analysis of Senecavirus A-infected cells: type I interferon is a critical anti-viral factor

ABSTRACT: Purpose: to study the changes in various genes in cells during the early stages of SVA infection. Differentially expressed genes were screened from them, and GO enrichment analysis and KEGG pathway enrichment analysis were performed. This study provides some help for better prevention and control of SVA infection. Methods: Deep sequencing of SVA-infected cells at 6hpi and 12hpi and mock-infected cells were performed, in triplicate. The raw data from the sequencing is filtered using ngqc software and compared to the reference sequence. Differential and enrichment analyses were then performed. PK:mock-infeted groups TP:SVA-infected groups Results: Using RNA-seq technology, more than 46 billion raw reads of each sample were generated, and the proportion of clean reads was >94%. The ratios of clean reads successfully mapped to the swine reference genome in all samples ranged from 90.41% to 95.92%, and the ratios of the uniquely mapped reads were >91.48%. The R2 values of the samples in the same group were >0.99. 1584 genes had a significant difference at 6hpi between SVA- and mock-infected groups and 9785 DEGs were screened at 12hpi, 28 of these DEGs were validated with RT–qPCR. Altered expression of 28 genes was confirmed with RT–qPCR, demonstrating the high degree of sensitivity of the RNA-seq method. At the same time, we selected three important DEGs for WB experiments and analyzed their protein-level alterations. The results were consistent with RNA-seq data, Further confirmation of the accuracy of RNA-SEQ technology Conclusion: Our experiments analyzed in detail the changes in various genes in cells after SVA infection. A series of cytokines, such as interleukins (IL6, TNF-α), chemokines (CCL4, CCL5), and immune-related factors (IFN-α, IFN-β, RSAD2, MX1), are involved in the battle between SVA and the host. We postulated that this innate immune response is the main mean used by the host for the initial response to SVA infection. This study evaluated SVA-induced immune responses and provides information that can be used to investigate the molecular mechanisms of SVA-host interactions.

ORGANISM(S): Sus scrofa

PROVIDER: GSE150242 | GEO | 2020/05/12

REPOSITORIES: GEO

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Project description:Purpose: Post-weaning multisystemic wasting syndrome (PMWS) is a disease associated with porcine circovirus type 2 (PCV2) infection. One main feature of PMWS is seriously decreased lymphocyte in lymphoid tissues. This research aims to investigate the molecular mechanism of evident lymphocyte depletion in spleen from Yorkshire x Landrace (YL) pigs after infection with PCV2. Methods: At the age of 6 weeks, 15 piglets of Laiwu (LW) and YL pigs were assigned randomly into two groups, respectively: 10 infected pigs and 5 uninfected pigs. Spleen tissues of each group were collected at 35 days post infection with PCV2. Three PCV2-infected YL pigs with apparently pathologic characteristics and mock-infected YL pigs were selected for RNA sequencing, respectively. The RNA extraction, library construction and RNA-Seq were performed on the Illumina HiSeq2500 at Beijing BioMarker Technologies (Beijing, China).Differential expression genes (DEGs) were filtrated from the result of transcriptome. The quantitative real-time PCR was performed to validate the expression patterns of candidate DEGs between LW and YL pigs. Results: The clean reads percolated from the raw reads were mapped to Sus scrofa genome (Sscrofa10.2) using Tophat2 software. The efficiency of alignment was about 76% between the six samples and reference genome, and approximate 50% clean reads were perfectly matched to pig genome. Two samples with high R-squared value (T1-T3: 0.98; T5-T6: 0.97) from PCV2 and mock-infected YL pigs, respectively, was performed differential expression analysis through DESeq after biologic repetition correlation detection. With the criterion of log FC (fold change) > 1.5 and of P value < 0.05, 90 significantly differentially expressed genes were identified between the two groups, of which 57 were up-regulated and 33 were down-regulated. Eleven candidate DEGs were confirmed by quantitative real-time PCR, which expression pattern were generally in accordance with the results of RNA-seq. Conclusions: Many DEGs participate in immune response and some of them are involved in cells proliferation or apoptosis, such as CD81, HGF, CXCL13, KLF11, PSMD4, CYCS and ACTC1. The expression changes of these DEGs may cause the lymphocyte depletion in spleen after challenge with PCV2.

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Transcriptome analysis of Senecavirus A-infected cells: type I interferon is a critical anti-viral factor

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