miRNA-Mediated Regulation of Meloidogyne arenaria Responses in Wild Arachis

MicroRNAs (miRNAs) are key post-transcriptional regulators of plant development and stress responses, with many being conserved across diverse plant lineages. In this study, we investigated the expression profiles of miRNAs and their corresponding target genes in <i>Arachis stenosperma</i>, a wild peanut relative that exhibits robust resistance to root-knot nematodes (RKN). Small RNA sequencing of nematode-infected roots identified 107 miRNA loci, of which 93 corresponded to conserved miRNA families and 14 represented novel candidates, designated as miRNOVO. Among these, 18 miRNAs belonging to 11 conserved families were identified as differentially expressed (DEMs). Notably, <i>miR399</i> and <i>miR319</i> showed the highest upregulation (logFC = 4.25 and 4.20), while <i>miR393</i> and <i>miR477</i> were the most downregulated (logFC = -0.83 and -0.79). Integrated analysis of miRNA and transcriptome data revealed several regulatory interactions involving key defense-related genes. These included NLR genes targeted by <i>miR393</i> and <i>miR477</i>, hormone signaling components such as the auxin response factor <i>ARF8</i> targeted by <i>miR167</i>, and the growth regulator <i>GRF2</i> targeted by <i>miR396</i>. Additionally, <i>miR408</i> was predicted to target <i>laccase3</i>, a gene involved in the oxidation of phenolic compounds, lignin biosynthesis, copper homeostasis and defense responses. Remarkably, four immune receptor genes belonging to the nucleotide-binding site leucine-rich repeat (NLR) family displayed inverse expression patterns relative to their regulatory miRNAs, suggesting miRNA-mediated post-transcriptional control during the early stages of nematode infection. These findings reveal both conserved and species-specific miRNA-mRNA modules associated with nematode resistance in <i>A. stenosperma</i>, highlighting promising targets for developing RKN-tolerant peanut cultivars through miRNA-based strategies.