Insect diet bioassays

The diet bioassays were conducted as described in Huynh et al.²¹. All materials used in the diet assays were surface-treated via exposure to UV light for 10 min in a biological cabinet. Each formulation was tested in a 12-well row of a 96 well plate and values from the 12 sub-samples were averaged to yield a single replicate. Each formulation was replicated at least 4 times. Eight different formulations were randomly assigned on the plate. One larva (<24 h after hatching) was placed in each well of the diet plate using a fine paintbrush. A sealing film (TSS-RTQ-100, Excel Scientific, Inc., Victorville, CA) was used to cover the plate and one vent hole was made per well using an insect pin (size 0). The plates were stored at 25 °C in darkness for 10 days. Collections made at 10 days were used to determine differences in survival, weight and molting to the 2^nd instar. Larval survival, time of molt to 2^nd instar, and evidence of diet contamination were recorded daily. Live larvae were collected and pooled within a replication into 95% ethanol, dried in an oven (602752, Blue M Therm Dry Bacteriological Incubator) at 50 °C for 2 days, and then were weighed using a micro balance (MSU6.6S-000-DM, Sartorius Lab Instruments GmbH & Co. KG, Goettingen, Germany).