Differentially expressed microRNAs with potential regulatory roles in ileum of Holstein cows with subclinical Johne's disease

Abstract Paratuberculosis or Johne’s disease (JD), caused by Mycobacterium avium subsp. paratuberculosis (MAP), is a chronic inflammatory disorder of the intestinal tract of ruminants. The mechanisms of JD pathogenesis remain unclear but MAP can cause chronic inflammatory lesions in the terminal small intestine of cattle. Using microRNA (miRNA) transcriptome analysis, this study investigated the impact of MAP infection on miRNA expression and their potential regulatory roles in JD progression. Ileal tissues were collected from MAP infected Holstein cows (n = 5) and healthy cows (n = 5). Purified total RNA were used to construct libraries followed by next generation sequencing, bioinformatic processing using a standard pipeline, differential miRNA expression analysis with DESeq2 software, and functional enrichment with ClueGo app in Cytoscape. A total of 269 miRNAs were expressed in ileal tissue including three abundantly expressed miRNAs (Bta-miR-143; 54.05% of total read counts), bta-miR-145 (11.54%) and bta-miR-192 (4.31%)). Twenty miRNAs were significantly (P < 0.05) differently expressed (DE) when comparing ileal tissues from MAP infected and uninfected cows. Bta-miR-383 (fold change = -12.93) and bta-miR-1842 (fold-change = 8.17) were the most down- and up-regulated miRNAs, respectively. Amongst the DE miRNAs, bta-miR-146a, bta-miR-146b and bta-miR-393 are inflammation-related miRNAs and considered potential biomarkers of bovine mastitis and other diseases, while bta-miR-370 and bta-miR-383 are associated with lipid metabolism. The predicted target genes of DE miRNAs were enriched (P < 0.05) for gene ontology terms related to regulation of metabolic, cell and developmental processes, and immune system development. KEGG pathway analysis revealed potential involvement in immune regulation (e.g. T-cell receptor signaling and C-type lectin receptor signaling pathways) and pathways in cancer were also enriched (P < 0.05). Our results indicate that some DE miRNAs play important roles by regulating metabolism and immune processes during JD and provide DE miRNAs that might form targets for future development of intervention measures.