RNA sequencing obtained 26,664 assembled transcripts, including 2,912 differentially expressed genes. Based on small RNA sequencing and secondary structure predictions, we identified 17 CyHV-2 encoded miRNAs, among which 14 were validated by stem-loop quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) and eight were validated by northern blotting. Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of miRNAs-mRNA pairs revealed diverse affected immune signalling pathways, including the RIG-I-like receptor and JAK-STAT pathways. Finally, four genes were presented to be involved in RIG-I-like pathways, including host gene IRF3, RBMX, PIN1, viral gene ORF4, which are negatively regulated by CyHV-2 encoded miRNA miR-C4.
This study is the first to provide a comprehensive overview of viral miRNA-mRNA co-regulation, which might have a key role in controlling post-transcriptomic regulation during CyHV-2 infection.
The level of CyHV-2 and the DEGs in the kidney of silver crucian carp.
Samples were collected at 0, 12, 24, 48, and 72 h after challenged with CyHV-2. (A) The accumulation curve of CyHV-2 in the kidney of silver crucian carp. Viruses were titrated using a real-time PCR approach. (B) The relative expression levels of MAPK7, MHC-I, and NF- κB; (C) IRF3, RBMX, and PIN1 in the kidney, β-actin served as the reference gene. Results are presented as mean ± SD of 3 independent experiments, *P < 0.05, **P < 0.01.