Soil Pollution

We used six healthy honeybee colonies (Apis mellifera ligustica Spinola, 1806, located at the UCSD Biology Field Station apiary, La Jolla, USA), studied forager and in-hive honeybees, and followed standard collection and rearing methodologies [49 (link)]. To test the effect of season, we collected bees at two different colony developmental stages: early spring (February–March 2016) and summer (July 2016). We tested the synergistic and individual effects of FPF exposing bees to five acute oral doses of FPF or FPF + PRO. Based on current guidelines [50 ,51 (link)], we tested FPF doses (375 and 750 ng bee⁻¹) considered field-realistic, since bees can ingest higher FPF doses while foraging (see electronic supplementary material for the worst-case scenario estimations).
Following previous studies [22,28,52], we used a relatively high PRO dose that nonetheless, on its own, has no impact on bee survival (7000 ng bee⁻¹ [22 (link),52 (link)]). PRO is one of the most commonly used fungicides that contaminates bees and the environment [31 (link),32 (link)]. Bees can be simultaneously exposed to FPF and PRO (or another SBI fungicide with similar mode of action) because they are used on the same crops and ornamentals, including fruits (e.g. citrus), oilseeds (e.g. soya bean, peanuts), cereals (e.g. corn, sorghum) [10 ,12 (link),15 ,53 –55 ], although guidelines state that flupyradifurone should not be directly tank-mixed with azole fungicides when applied to flowering crops [10 ]. These pesticides can be used multiple times over a year in the same crop (and over different seasons) and applied in multiple ways (i.e. aerial, chemigation or ground application). In addition, bees can also be exposed to pesticides that drift from different crops (i.e. buffer zones) or are stored in the same hive [56 (link),57 (link)]. FPF and PRO are easily taken up by plants and thus contaminated soil and water may lead to unintended absorption. This can result in prolonged, multi-year contamination [56 (link),58 (link),59 (link)]. Bees can therefore be exposed to pesticide combinations that are contraindicated in tank mixes [6 (link)].
We tested a control dose (0 ng bee⁻¹), a total of six doses of FPF (375, 750, 1500, 3000, 6000, 12 000 ng bee⁻¹, respectively corresponding to 37.5, 75, 150, 300, 600, 1200 ppm), and five doses of the positive control dimethoate (DIM; 50, 100, 200, 400, 800 ng bee⁻¹, respectively corresponding to 5, 10, 20, 40, 80 ppm). In the combined FPF + PRO treatment, each FPF dose was tested in combination with a single sublethal dose of PRO (7000 ng bee⁻¹, corresponding to 700 ppm). We used technical grades of all active ingredients. The test solutions (sucrose 50% w/w, 100 µl cage⁻¹, 10 µl bee⁻¹,) were provided inside each cage using an Eppendorf cap [60 ], contained acetone as a solvent (0.7%) and were completely consumed 60 min after oral administration [60 ].
We measured the effects of treatment on bee survival (1–48 h) and the frequency of bees exhibiting abnormal behaviours (1–4 h, see below).
Detailed methods are reported in the electronic supplementary material.

The degree of soil HM pollution was assessed as follows: first soil pH values were categorized into three classes: <6.5, 6.5 ≤ pH ≤ 7.5, and >7.5; second, the pollution threshold for each soil HM was determined by land use (e.g., paddy fields) and pH class; third, the single pollution index (SPI) for each HM was determined (Equation (1)); finally, the Nemerow composite pollution index (NCPI) was calculated (Equation (2)).
$P_i=\frac{C_i}{S_i}$
where $C_i$ is the concentration of soil HM i, and S_i is the pollution threshold of $i$ [32 ].
$NCPI=\sqrt{\frac{(P_{imax}{)}^2+({\overline{P}}_i{)}^2}{2}}$
where $P_{\mathrm{imax}}$ is the maximum SPI value of each HM and $\overline{P}$ is the mean SPI of each HM [32 ].
As NCPI is a comprehensive index, it was used to classify the soils in terms of HM pollution. Table 1 shows the classifications of the single pollution index (SPI) and Table 2 shows the classifications of Nemerow composite pollution index (NCPI).

Black soldier fly larvae (BSFL; Hermetia illucens) were obtained from the commercial BSF producing company Bestico, (Berkel en Rodenrijs, The Netherlands). The following substrates were tested: chicken feed (CF; control diet), pig manure slurry mixed with roadside silage grass (PMLSG), the organic wet fraction of municipal household waste (OWF), secondary sludge from slaughter waste (SW), fast food waste (FFW), mushroom stems (MS) and pig manure solid (PMS). Chicken feed is a commercial broiler feed which was used as the control diet [33 (link),36 (link),37 (link)]. Pig manure slurry was a mixture of pig feces and urine, and it was mixed with roadside silage grass (1:1 w/w) to produce PMLSG. The organic wet fraction used in this experiment makes around 30–35% of the municipal household waste. It was contaminated by physical contaminants such as glass and plastic that were not removed. The solid phase of the secondary sludge from slaughter waste was also used as an experimental substrate. The fast food waste consisted of fries, vegetables, bread and meat products but not any non-food waste and was collected within maximum 4 days after disposal. The mushroom stems are a soft substrate and may have been contaminated by soil. The different substrates were selected based on the results of a prior study published by Veldkamp [33 (link)].
Substrates were obtained one week before the start of the rearing cycle and stored at 4 °C until use. Some of the substrates were pre-treated in a cutter to decrease particle size which included PMLSG (~2 cm), FFW (~1 cm) and MS substrates (~0.5 cm). All substrates are brought to 35% dry matter by adding water and/or cellulose/wood shavings to decrease or increase the DM in the substrate, respectively. Since the used substrates all have a different weight-to-volume ratio, different quantities of substrates and larvae were added to the containers to maintain a substrate layer of approximately 5 cm such that every BSFL gets 0.54 g of the wet substrate (Table 1). The containers were filled one day before starting the experiment, thus allowing them to adapt to the ambient temperature in the climate chamber without any external heating. On top of each substrate, 1850 starter BSFL (8 day old) per kilogram of wet substrate were incubated in 21 plastic containers (75 cm × 47 cm × 15 cm). Each substrate was tested in triplicate in a climate chamber (7 treatments × 3 replicates). The chamber temperature was set to 28 °C and the relative humidity (RH) was 70% from day 0 until day 5 and was 40–60% from day 6 until the end of the experiment. The rearing chamber was dark. The plastic containers were stacked in three columns each with seven containers (one container per repetition) arranged based on escaping probability, i.e., the containers with the highest moisture content were placed at the bottom to avoid escaping larvae falling into containers below them. Each column was placed in a non-escape box (cubic box; 120 cm × 100 cm × 60 cm). The experimental period was 7 to 8 days which was determined by visual checking of the substrate consumption or the presence of ~10% prepupae.

The oil-contaminated soil sample was collected from the Southwest University of Science and Technology, Mianyang, Sichuan Province (N 31.53791, E 104.69388). For the isolation and screening of lipase-producing strains, 5 g of the soil sample was weighed in 45 mL of physiological saline, incubated for 1 h at 30 °C while being shaken at 200 r/min, and then rested. A quantity of 1 mL of supernatant was added to 20 mL of enrichment medium (yeast paste 0.2 g/L, NaCl 0.5 g/L, Na₂HPO₄ 3.5 g/L, KH₂PO₄ 1.5 g/L, MgSO₄-7H₂O 0.5 g/L), and this was incubated for 24 h at 30 °C while being shaken at 200 r/min. After gradient dilution of the enrichment medium, 100 μL of 10⁻⁶~10⁻⁸ dilution was coated onto the oil assimilation plate containing rhodamine B (25 mL olive oil emulsion was mixed well with 175 mL LB medium, 200 μL 10% rhodamine B solution was added, and the mixture was shaken). After incubation at 30 °C for 2 d, a pure culture capable of producing degradation circles was obtained, named WCO-9. The single colony of strain WCO-9 was selected and inoculated in an LB liquid medium. Cells in the logarithmic growth phase were obtained by shaking at 200 r/min and incubating at 30 °C for 12 h and were then stored in glycerol tubes. Finally, the WCO-9 strain was identified and registered by the Guangdong Microbial Culture Collection Center with the serial number GDMCC No: 61851. The scanning electron microscopy observation, 16S rDNA analysis and whole genome sequencing were performed using cells from the logarithmic growth stage.