Identification of a novel mitovirus in Colocasia esculenta through high‑throughput sequencing and in-silico miRNA prediction for sequence-specific gene silencing.

Mitoviruses are simple RNA viruses typically associated with fungal mitochondria, although recent studies suggest their presence in plant transcriptomes. In this study, we report the identification and molecular characterization of a novel mitovirus from Colocasia esculenta using high-throughput sequencing (HTS). Analysis of symptomatic leaf tissues revealed a 2,644 nucleotide-long viral genome encoding a single open reading frame (ORF) for an RNA-dependent RNA polymerase (RdRp) of 544 amino acids. The viral sequence, tentatively named Colocasia esculenta associated mitovirus 2 (CeaMV2), showed 60.76% amino acid identity with known mitoviruses, supporting its classification as a novel species within the family Mitoviridae. Phylogenetic analysis showed that CeaMV2 clustered within the family Mitoviridae, grouped with other plant-associated mitoviruses, supporting its placement within the family Mitoviridae. The conserved domain searches confirmed the presence of the mitovirus-specific RdRp domain (Pfam: PF05919). RT-PCR and Sanger sequencing further validated the presence of CeaMV2 in C. esculenta leaf samples. The relatively short RdRp and its expression in plant transcriptome suggest possible association with plant mitochondria. Furthermore, in-silico analysis identified several plant-derived mature miRNAs, such as sbi-miR6231-3p, with high binding affinity to the CeaMV2 RdRp gene, suggesting a potential role for RNA interference in the host-virus interaction. This finding expands the known diversity of mitoviruses and highlights the utility of HTS in uncovering cryptic viral infections and potential RNAi-based defense mechanisms in economically important crops.