## 2 Standard Methods
### Field Trial  
This field trial involves standardizing the number of adult SHBs in hives, administering treatments (in this study: in modified CD cases for a set time), and then counting the number of living adult SHBs remaining in the hive and dead adult SHBs in traps to calculate treatment efficacy. To accomplish this, hives are made SHB-free (to standardize SHB populations) and then a known number of adult SHBs is added to each hive. The treatments and concentrations used are selected from data generated from the acute toxicity bioassay. 1. Prepare honey bee hives. 1.1 Ensure all hives have approximately the same colony strength (adult bees and brood) and size/equipment configuration (i.e., number of boxes and frames). 1.2 Standardize adult SHBs in each hive (t = 0 d). 1.2.1 Use the \"bounce method\" described in Section 3.2.3.1 of [PERSON] et al. [17] to remove all adult SHBs from hives. Find and set aside or cage the queen before using this method. 1.2.2 Use an electrically powered aspirator to collect SHBs quickly. 1.2.3 If needed, create an electrically powered aspirator by connecting the tubing of an insect aspirator (Table 1L) to a compact vacuum (Table 1M). 1.3 Number all hives and assign them evenly/randomly into treatments. 1.3.1 Include a minimum of three treatments: solvent control, positive control, and experimental compound of interest. 2. Prepare traps. * 2.1 Cut three entrances into the closed CD case (Table 1N) using a hot knife (Table 1O) inside a fume hood. The openings should be 3.5 cm long and 0.5 cm wide (Figure 2). Note: Ensure the openings lead directly into the center of the case without any obstructions. Center the openings along each of the three unhinged sides to avoid the ring of plastic inside the case. 2.2 Cover the holes on the center of the top and bottom of the CD case with tape (Figure 1B, Arrow 1). This will ensure the treated pollen remains inside the trap. 2.3 Prepare one trap for every hive box in the field trial. For example, a hive consisting of a deep brood box and medium super would receive two traps. Note: Clearly label each trap with the date that treated pollen is inserted, the treatment ID, the treatment concentration, hive number, and trap position in the hive. 3. Prepare treated pollen and place into traps. 3.1 Weigh pollen balls. 3.1.1 Weigh 25 \(\pm\) 0.1 g of bee-collected pollen or commercial pollen supplement (Table 1A) on a calibrated scale (Table 1B). Roll 25 \(\pm\) 0.1 g portions of pollen into balls with gloved hands. Prepare one pollen ball for each trap needed. Note: If the pollen mixture is sticky, place it in the freezer (\(-20\pm 1\)\({}^{\circ}\)C) to harden before handling. 3.1.2 Wrap individual pollen balls in plastic wrap (Table 1C). Store in refrigerator (+4 \(\pm\) 1\({}^{\circ}\)C) for up to two weeks before discarding. 3.2 Prepare solutions. 3.2.1 Select one concentration for each compound. The concentration is chosen based on the results of the laboratory bioassays. At a minimum, include a solvent (i.e., acetone) control, positive control, and one experimental compound. Note: Be sure to select a concentration that kills 100% of the adult SHBs for the positive control. We used fipronil and thiamethoxam as positive controls, but any compound with proven 100% efficacy can be used. 3.2.2 Clearly label amber vials (Table 1I) with the date, compound, and concentration of compound that will be added. Figure 2: Trap design used in the field trial. The CD cases were modified by cutting a 3.5 cm entrance on 3 sides (denoted by *) to allow small hive beetles access to treated pollen inside the cage. 3.2.3 Prepare 1375 \(\mu\)L of solution for each trap. This follows the 50 \(\mu\)L:1 g pollen ratio used in laboratory bioassays (25 g pollen \(\times\) 50 \(\mu\)L = 1250 \(\mu\)L) and includes 10% of the original volume excess to account for evaporation (1250 \(\mu\)L \(\times\) 0.10 = 125 \(\mu\)L).