Location: Virus and Prion ResearchTitle: Transcriptomic analysis of 3D air-liquid interface porcine respiratory epithelial cells culture infected with the betacoronavirus PHEV
|SARLO DAVILA, KAITLYN - Oak Ridge Institute For Science And Education (ORISE)|
|NELLI, RAHUL - Iowa State Veterinary Laboratory|
|MORA-DIAZ, JUAN CARLOS - Iowa State Veterinary Laboratory|
|SANG, YONGMING - Tennessee State University|
|GIMENEZ-LIROLA, LUIS G - Iowa State Veterinary Laboratory|
Submitted to: Swine Disease Conference Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: 10/26/2022
Publication Date: 12/3/2022
Citation: Sarlo Davila, K.M., Nelli, R., Mora-Diaz, J., Sang, Y., Miller, L.C., Gimenez-Lirola, L. 2022. Transcriptomic analysis of 3D air-liquid interface porcine respiratory epithelial cells culture infected with the betacoronavirus PHEV. Swine Disease Conference Proceedings. NAPRRS-NC229 Proceedings.
Technical Abstract: This study is a continuation of our previous studies with air-liquid interface porcine respiratory epithelial cells (ALI-PRECs) inoculated with the neurotropic betacoronavirus porcine hemagglutinating encephalomyelitis virus (PHEV) which primarily infects and replicates in the upper respiratory tract of swine. Here, we present the mechanisms driving the early innate immune modulation in ALI-PRECs following PHEV infection. Following infection with PHEV, total RNA from ALI-PRECs were collected at 24h post-inoculation (hpi), 36hpi, and 48hpi. After determining the quality of RNA, RNA-seq analysis was performed at the Iowa State University Genomics center (library preparation and sequencing) using the Illumina Hiseq 6000. The sequences were aligned on the Sscrofa 11.1 genome with HiSat2 utilizing default parameters for paired end reads. Differential gene expression (DEG) was performed using DeSeq2. Gene ids were based on Ensembl ids and were converted to gene names using the g:Convert function of g:Profiler. The number of significant (FDR < 0.15) differential expressed genes (DEGs) increased with time post infection with 112 DEGs at 24hpi, 163 DEG at 36hpi, and 179 DEGs at 48hpi. As expected, DEGs involved in the Type-1 interferon response were consistent in all three timepoints based on gene ontology terms that were functionally clustered. DEGs related to chemokines were observed at 36hpi and 48hpi, while DEGs related ciliary function were expressed at 48hpi. Using Qiagen Ingenuity Pathway Analysis (IPA), this study has identified significant networks, top functions and canonical pathways associated with DEGs. Among several predicted networks, hypercytokinemia/ hyperchemokinemia in the pathogenesis of influenza pathway, was the most significant canonical pathway at all three time-points. Predicted dysregulation of DEGs involved with RIG-I-like receptors (RLRs), IRF-3/IRF-7 and transcription factors such as STAT1, AP1 were noticed as early as 24 hpi. Their activation is predicted to activate several DEGs involved in proinflammatory response that includes CCL5 and CXCL10 via the NFkB signaling pathway. In summary, RNA-seq analysis performed in this study further supports our previous results that ALI-PRECs are excellent ex vivo infection model to study PHEV and porcine respiratory pathogens. PHEV, indeed disrupts the homeostasis of porcine respiratory epithelia by altering the ciliary function and inducing antiviral, proinflammatory cytokine and chemokine responses.