Infectious hematopoietic necrosis virus (IHNV) can cause widespread death of rainbow trout (Oncorhynchus mykiss), understanding the molecular mechanisms that occur in the rainbow trout in response to IHNV infection will be useful to decrease IHN-related morbidity and mortality in trout aquaculture. However, the molecular mechanisms of rainbow trout in response to IHNV are very limited. This study performed analysis of mRNAs and miRNAs based on RNA-seq technology on the intestine of rainbow trout infected with IHNV and control. There were 80 differentially expressed miRNAs that regulated 3355 target mRNAs, which overlapped with differentially expressed mRNAs obtained from RNA-seq. The expression patterns of DEGs and miRNAs differentially expressed were validated by qRT-PCR. GO enrichment and KEGG pathway analyses of the potential target genes of the DE miRNAs, revealed DEGs were mainly enriched in immune-related pathways such as Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway and IL-17 signaling pathway. These findings improve our understanding of the molecular mechanisms of IHNV infection. The study analyzed the immune regulatory target gene pairs and signal pathways of rainbow trout intestine against IHNV infection at the transcriptional level, and provided basic data for the study of rainbow trout against IHNV immune regulatory. Overall design: In this study, the target gene pairs and signaling pathways of immune regulation against IHNV infection in rainbow trout intestinal tract were analyzed at the transcriptional level.