A primary cell culture system was established for the first time from embryonic tissues of Asian honeybee, Apis cerana, and used to trace the early infection process of Chinese sacbrood virus (CSBV), an iflavirus in the family Iflaviridae. A monolayer of epithelium-like cells of A. cerana, approximately 8-10 μm in diameter, was grown in Kimura's insect medium at 28 °C within 3-4 days of setting up the cultures. Such cultured cells were inoculated with CSBV purified from infected larvae or pupae for 2 h. In electron and confocal micrographs, viral particles accumulated as filamentous or vesicular inclusions in the cytoplasm of infected cultured cells at 36 h post-inoculation (hpi). Real-time quantitative RT-PCR assay showed that the expression levels of four cistrons of CSBV in the cultured cells increased rapidly between 12 and 48 hpi. This newly established primary cell culture derived from A. cerana will be useful for further studies of infection caused by CSBV.