miR‐701 modulates the humoral immune response of <i>Coptotermes formosanus</i> against <i>Metarhizium anisopliae</i>

Termites are not only social insects but also significant global insect pests. Investigating the molecular mechanisms regulating immune defense response in termites is beneficial for developing novel approaches to termite management. Currently, research mainly focuses on coding RNAs in termite immunity, with limited exploration of non-coding RNAs. Here, we identified miR-701, a markedly downregulated microRNA (miRNA) in the globally significant termite pest Coptotermes formosanus after Metarhizium anisopliae infection, which targets the immune gene Toll4. Transcriptome analysis of termites injected with miR-701 agomir revealed that miR-701 affects the immune-related response, growth, and development of termites. Treatment with miR-701 agomir, either through injection or ingestion, resulted in a notably reduced survival rate of termites infected with M. anisopliae compared to the control group infected with M. anisopliae alone. Additionally, termites injected with miR-701 agomir exhibited a significant decrease in the expression of antimicrobial peptide genes termicin and lysozyme, alongside a notable increase in the colony-forming units of M. anisopliae in the infected termites. Subsequent investigations demonstrated that miR-701 suppressed the expression of the target gene Toll4, consequently inhibiting the Toll signaling pathway and diminishing the expression of antimicrobial peptides. These findings suggest that termites can combat M. anisopliae by downregulating miR-701 expression to activate the Toll signaling pathway and enhance antimicrobial peptides synthesis. This discovery improves our comprehension of the role of miRNAs in termites' immune responses and the mechanism of termites managing miRNAs to boost their pathogen resistance. Additionally, it reveals a new molecular target for termite biological control.