A Beetle In Vitro: Establishment of a Short-Term Cell Culture from the Pest Popillia japonica.

The Japanese beetle (Popillia japonica Newman) is a highly invasive, polyphagous scarab causing significant agricultural and ecological damage across invaded regions. While molecular studies are gaining traction, the unavailability of P. japonica cell lines has constrained in vitro investigations. To overcome these limitations and provide a platform for controlled biological investigation, we developed the first cell culture derived from P. japonica larvae. Fat bodies from field-collected third-instar larvae were dissected and cultured. Cells initially formed floating spheroids before transitioning to adherent monolayers. Cultures remained stable over several splits, whereas a marked reduction in cell number was observed at the eighth split due to the onset of contamination. Fluorescence microscopy confirmed nuclear integrity, while transmission electron microscopy at split 5 revealed cytoplasmic features consistent with insect fat body cells, including lipid droplets. The cell culture predominantly contained trophocyte-like cells, consistent with the known cellular composition of insect fat bodies. Transcriptomic analyses comparing fresh fat bodies and cultured cells revealed moderate transcriptional divergence, with limited upregulation of genes associated with iron homeostasis and stress response, consistent with adaptive responses to in vitro conditions. While not immortalized, this cell culture offers a short-term model for studying P. japonica physiology, toxicology, host-pathogen interactions, and potential gene-targeting strategies under controlled conditions. This work represents a first step toward enabling molecular and cellular research in this economically important pest species.