Transcriptional profiling of feline infectious peritonitis virus infection in CRFK cells and in PBMCs from FIP diagnosed cats

Abstract

BackgroundFeline Infectious Peritonitis (FIP) is a lethal systemic disease, caused by the FIP Virus (FIPV); a virulent mutant of Feline Enteric Coronavirus (FECV). Currently, the viruses virulence determinants and host gene expressions during FIPV infection are not fully understood.MethodsRNA sequencing of Crandell Rees Feline Kidney (CRFK) cells, infected with FIPV strain 79–1146 at 3 hours post infection (h.p.i), were sequenced using the Illumina next generation sequencing approach. Bioinformatic’s analysis, based on Felis catus 2X annotated shotgun reference genome, using CLC bio Genome Workbench mapped both control and infected cell reads to 18899 genes out of 19046 annotated genes. Kal’s Z test statistical analysis was used to analyse the differentially expressed genes from the infected CRFK cells. Real time RT-qPCR was developed for further transcriptional profiling of three genes (PD-1, PD-L1 and A3H) in infected CRFK cells and Peripheral Blood Mononuclear Cells (PBMCs) from healthy and FIP-diseased cats.ResultsBased on Kal’s Z-test, with False Discovery Rate (FDR) <0.05 and >1.99 fold change on gene expressions, a total of 61 genes were differentially expressed by both samples, where 44 genes were up-regulated and the remainder were down-regulated. Most genes were closely clustered together, suggesting a homogeneous expression. The majority of the genes that were significantly regulated, were those associated with monocytes-macrophage and Th1 cell functions, and the regulation of apoptosis. Real time RT-qPCR developed focusing on 2 up-regulated genes (PD-L1 and A3H) together with an apoptosis associated gene PD-1 expressions in FIPV infected CRFK cells and in PBMCs from healthy and FIP diagnosed cats produced concordant results with transcriptome data.ConclusionThe possible roles of these genes, and their importance in feline coronaviruses infection, are discussed.

Apoptosis transcriptional mechanism of feline infectious peritonitis virus infected cells

“…CCL8, CXCL10, and CCL17), genes related to innate immune responses (e.g. PHF11 and IRF1), and some genes like MX1 also showed significant up regulation ( Supplementary Table 1), confirming the findings from previous study by Harun and his colleagues [22].…”

“…Transcriptome analysis of FIPV infected cells showed that the virus probably increase the inflammation through the chemokine and cytokine signaling pathways, Wnt signaling, Cadherin signaling, to name but a few (Tables 3, 4, 5). After 9 hpi, this virus could significant deregulate 4526 genes, which is much higher than 96 genes at 3 hpi according to a previous study by Harun et al [22]. In addition, the expression of some of the ISG genes (e.g.…”

Inflammatory Mediators in the Mesenteric Lymph Nodes, Site of a Possible Intermediate Phase in the Immune Response to Feline Coronavirus and the Pathogenesis of Feline Infectious Peritonitis?

“…In the present study, gene expression for all three pyrogenic cytokines (IL-1b, IL-6 and TNF-a) was upregulated in the MLNs in FIP, as well as that for IL-15 (a stimulator of lymphocyte proliferation). We also found significantly higher transcription levels for the monocyte-recruiting chemokines CXCL10 and CCL8, which have both been found to be upregulated in CrandelleRees feline kidney cells after in vitro FCoV infection (Harun et al, 2013), indicating a mechanism of monocyte recruitment as a direct viral effect. Our results confirm their relevance in vivo, with recruitment of monocytes as the infected cell type being a potential amplifying step that is worthy of further investigation.…”

An update on feline infectious peritonitis: Virology and immunopathogenesis

“…One of the newest approaches to studying immune responses involves transcriptional profiling, wherein the various cellular mRNAs triggered by a viral infection can be monitored for up-or downregulation. Harun et al (2013) demonstrated the feasibility of such an approach using CRfk cells exposed to FIPV WSU-79-1146 for 3 h. They were able to detect 18,899 of the 19,046 annotated feline genes. Based on results from their culture studies and Kal's Z test, 44 genes were upregulated, 61 genes were not affected by virus infection and the remaining genes were down-regulated.…”