Transcriptomic and RNAi-mediated antiviral response to Kamiti River virus infection in Aedes albopictus U4.4 cells.

Mosquitoes transmit arboviruses that represent major global public health challenges. Increasing insecticide resistance and absence of effective antiviral therapies underscore the need for novel vector control strategies. Insect-specific viruses have emerged as candidates for biological control, however, the cellular mechanisms underlying their interactions with mosquito hosts remain poorly understood. Here, we examined the immune response of Aedes albopictus U4.4 cells to Kamiti River virus (KRV) infection, an insect-specific flavivirus. Cells were infected with KRV, and transcriptomic and small RNA profiles were analyzed at 24, 48 and 72 h post-infection. KRV infection induced production of virus-derived small interfering RNAs (vsiRNAs) and virus-derived PIWI-interacting RNA (vpiRNAs) from 24 to 72 h. The vsiRNAs predominantly mapped to the 3' untranslated region of the KRV genome, whereas vpiRNAs formed distinct hotspots in regions encoding the NS1, NS3, NS4A/B and NS5 proteins. Transcriptomic analysis revealed upregulation of genes associated with the humoral immune response, including defensin, cecropin, and glutathione S-transferase, and downregulation of Toll-like receptors and ecdysone-induced transcripts at later stages of infection. These gene expression patterns suggest an early activation followed by suppression of key immune signaling pathways. Collectively, the findings indicate that KRV leads to coordinated modulation of antiviral RNAi and host transcriptional responses, consistent with a balanced, commensal-like interaction in mosquito cells.