All RT-QPCR data generated during the infection of Galleria mellonella larvae with TBS buffer (TBS), Staphylococcus aureus HG003 (SA), EVs produced by Staphylococcus aureus HG003 (EV) and non infected. At 2, 24, and 72 h post-injection (hpi), two living larvae were randomly selected in each group for monitoring key larvae immune system actors. Experiment was conducted in biological triplicates. Total RNA was directly extracted from the hemolymph (HL) and the fat body (FB) larval content using the phenol-chloroform method. Dataset comprises raw, filtered, normalized and analyzed RT-qPCR data.