Cyprinid herpesvirus 3 was not replicate in EPC cells but mediated cell cycle arrest in the S phase

EPC (Epithelioma Papulosum Cyprini) cell line is widely used for culturing various aquatic animal viruses. However, prior studies reported that EPC cells were non-permissive to CyHV-3 replication. In this study, CyHV-3 were inoculated to EPC cells and conducted cytopathic effect (CPE) observation, PCR and quantitative real-time PCR for viral gene detection and load quantification, electron microscopy to visualize virion formation, transcriptome sequencing (RNA-seq) to explore infection mechanisms, and flow cytometry to assess cell cycle alterations. Results demonstrated that CyHV-3-infected EPC cells exhibited no CPE at 24h, 48h, 72h, 96h, 7d, and 14d post-infection. While viral DNA and transcripts of immediate-early (IE), early (E), and late (L) genes were detected in cell pellets, neither viral genes nor cDNA were found in supernatant or cell-free fractions. RT-qPCR confirmed low viral loads at all time-points, and no virions were observed via transmission electron microscopy, collectively indicating the establishment of latent infection. Transcriptomic analysis of EPC cells at 96h post-infection identified 607 differentially expressed genes (DEGs). GO enrichment categorized these DEGs into biological processes (BP), molecular functions (MF), and cellular components (CC), primarily associated with cellular compartments, binding, metabolic processes, catalytic activity, and biological regulation. KEGG pathway analysis revealed enrichment in sterol biosynthesis, cell cycle, terpenoid backbone synthesis, and DNA replication. Flow cytometry showed that CyHV-3 infection induced S-phase arrest in EPC cells, with the arrested population increasing over time. These findings suggest that CyHV-3 may establish latency by modulating host cell cycle-related gene expression.