Innate immune responses are effective for insect survival to defend against entomopathogens including a fungal pathogen, Metarhizium rileyi , that infects a lepidopteran Spodoptera exigua . In particular, the fungal virulence was attenuated by cellular immune responses, in which the conidia were phagocytosed by hemocytes (insect blood cells) and hyphal growth was inhibited by hemocyte encapsulation. However, the chemokine signal to drive hemocytes to the infection foci was little understood. The hemocyte behaviors appeared to be guided by a Ca 2+ signal stimulating cell aggregation to the infection foci. The induction of the Ca 2+ signal was significantly inhibited by the cyclooxygenase (COX) inhibitor. Under the inhibitory condition, the addition of thromboxane A 2 or B 2 (TXA 2 or TXB 2 ) among COX products was the most effective to recover the Ca 2+ signal and hemocyte aggregation. TXB 2 alone induced a microaggregation behavior of hemocytes under in vitro conditions. Indeed, TXB 2 titer was significantly increased in the plasma of the infected larvae. The elevated TXB 2 level was further supported by the induction of phospholipase A 2 (PLA 2 ) activity in the hemocytes and subsequent up-regulation of COX-like peroxinectins ( SePOX-F and SePOX-H ) in response to the fungal infection. Finally, the expression of a thromboxane synthase ( Se-TXAS ) gene was highly expressed in the hemocytes. RNA interference (RNAi) of Se-TXAS expression inhibited the Ca 2+ signal and hemocyte aggregation around fungal hyphae, which were rescued by the addition of TXB 2 . Without any ortholog to mammalian thromboxane receptors, a prostaglandin receptor was essential to mediate TXB 2 signal to elevate the Ca 2+ signal and mediate hemocyte aggregation behavior. Specific inhibitor assays suggest that the downstream signal after binding TXB 2 to the receptor follows the Ca 2+ -induced Ca 2+ release pathway from the endoplasmic reticulum of the hemocytes. These results suggest that hemocyte aggregation induced by the fungal infection is triggered by TXB 2 via a Ca 2+ signal through a PG receptor.