The CRISPR-Cas toolkit for mosquito-borne virus surveillance: detection, tracing, and discovery.

Mosquito-borne virus surveillance increasingly requires rapid, distributed detection of co-circulating pathogens, serotypes, and lineages across clinical and vector-sampling sites. CRISPR-Cas platforms offer a programmable toolkit for this purpose, but their readiness differs substantially across surveillance functions. Here, we review CRISPR-Cas methods for mosquito-borne virus surveillance across detection, tracing, and discovery-supporting targeted screening. Detection is the most advanced application: selected Cas12- and Cas13-based assays for dengue, Zika, chikungunya, West Nile, Japanese encephalitis, and related mosquito-associated viruses report sub-hour workflows, portable readouts, and targeted serotype- or lineage-marker discrimination. However, performance remains assay-, target-, and sample-matrix-dependent, and validation in pooled mosquito samples and field settings is still limited. Tracing currently relies mainly on validated portable amplicon-sequencing workflows, whereas CRISPR-aided sample-preparation methods such as DASH, FLASH, RAPID-DASH, and Cas9-targeted enrichment remain transferable opportunities for host depletion or target enrichment rather than established mosquito-borne virus genomic-surveillance workflows. For discovery-oriented surveillance, multiplex CRISPR-Cas systems such as CARMEN can support targeted screening of known or near-neighbor viruses represented by predesigned crRNAs, while metagenomic next-generation sequencing remains necessary for divergent or previously unknown viruses. Across these functions, CRISPR-Cas programmability may accelerate parts of assay redesign, but practical retargeting still requires compatible amplification primers, effector-specific target constraints, cross-reactivity assessment, and analytical revalidation. Routine surveillance use will require integrated demonstrations with clinical and pooled-vector samples, comparison against established molecular and sequencing methods, cost validation, and regulatory evidence.