Feeding preference assay

To assay feeding preferences, the food mixing protocol was performed as previously described. In brief, Erioglaucine (FD&C Blue #1) was mixed into the experimental (Fermented) or control (H₂O) food. Twenty mated females were allowed to feed for 4 h, after which they were frozen. Flies were then homogenized in 200 μl of PBS, and the homogenate was centrifuged at 8,000 g for 10 min. The absorbance values of the supernatant were measured at 625 nm and converted into the concentration of dye. Subsequently, feeding preferences was computed analogously to the oviposition preference index. For the capillary feeding assay, females were starved for 8 h, and then transferred to the vials with nylon plugs. Two capillary tubes supplying fresh or fermented grape juices were inserted through the plug. The volume of the juices consumed by flies was recorded. A feeding index (FI) was calculated: FI = (volume of fermented juice – volume of control juice)/total volume of juice.