Zika virus exists in enterocytes and enteroendocrine cells of the midgut.

The midgut is crucial for blood digestion, nutrition, reproduction, and pathogen interaction. Using single-cell RNA sequencing, we explored virus infection and transcriptomic changes at the cellular level. We identified 12 distinct cell clusters in the midgut post-Zika virus infection, including intestinal stem cells, enteroblasts, enteroendocrine cells (EE), and enterocytes (ECs). The virus was found mainly in specific subsets of ECs and EE. Infection altered transcriptional profiles related to metabolism, signaling, and immune responses. Functional studies highlighted three significantly differentially expressed genes in infected cells. Notably, silencing apolipophorin III reduced virus RNA copy number in the midgut, emphasizing the role of specific genes in viral infection. These findings enhance our understanding of mosquito midgut cell processes during Zika virus infection and suggest potential targets for vector control.