nce-miR-12220 is a vital regulator for microsporidian to infect honeybee via positive modulation of ATP-A and γ-tubulin genes

Abstract

Microsporidia rely extensively on host resources, yet how parasite-derived microRNAs coordinate infection remains poorly understood. Here, we investigated the function of nce-miR-12220, a miRNA identified in Nosema ceranae spores, during infection of Apis mellifera workers. Target prediction, dual-luciferase assays, and fluorescence in situ hybridization were combined with RNA interference and miRNA gain- and loss-of-function experiments. nce-miR-12220 interacted sequence-specifically with binding regions in ATP-A and γ-tubulin and was detected in infected honeybee midgut epithelial cells. Silencing either target gene reduced expression of the N. ceranae virulence-associated gene NcRBL and improved worker survival relative to the scramble control. In infected workers, nce-miR-12220 overexpression increased endogenous ATP-A and γ-tubulin transcript abundance, whereas inhibition produced the opposite effect. Overexpression also reduced expression of the Toll pathway-associated genes Cactus and dorsal and the antimicrobial peptide genes Defensin and Hymenoptaecin , increased N. ceranae spore load and sucrose consumption, and decreased midgut ATP content. Inhibition of nce-miR-12220 reversed these responses and reduced parasite burden. Survival after nce-miR-12220 manipulation changed in the predicted direction but did not reach statistical significance. Together, these findings identify nce-miR-12220 as a parasite-derived regulator that promotes N. ceranae proliferation while reshaping host immune and energetic responses, and suggest that this microRNA and its target network may provide candidates for controlling bee nosemosis.