Comparing the three exposure matrices, soil had the greatest number of neonicotinoids (clothianidin, imidacloprid, and thiamethoxam) detected, whereas imidacloprid was the only neonicotinoid detected in pollen and nectar. The maximum concentration of imidacloprid detected in soil (41.6 ng/g) was substantially greater than in pollen (4.3 ng/g) or nectar (1.1 ng/g), because imidacloprid is applied directly to soil and can bind to soil particles31 (Table S3). Although thiamethoxam is more soluble than imidacloprid, it is quickly metabolized to clothianidin6,7,32, which is much less soluble than either imidacloprid or thiamethoxam (Table S4). Dively and Kamel10 reported higher imidacloprid residue concentrations (60.9 ng/g for pollen, 7.4 ng/g for nectar) and greater frequencies of detection (92% of pollen samples; 88% of nectar samples) in the pollen and nectar of Cucurbita crops. However, the interval between application and sampling was shorter in their study (5 weeks) than in our study (8 weeks). Stoner and Eitzer11 also reported higher concentrations of imidacloprid and thiamethoxam in Cucurbita pollen and nectar than found here, likely because they included anther and nectary tissue in their samples.

The analysis of the hazard of pesticides to hoary squash bees in Cucurbita crops is based on LD 50 /LC 50 for adult honey bees as this represents current regulatory standards for testing toxicity in bees33. It is possible that differences in sensitivity exist between these two species as not all bee species are equally sensitive34,35,36. Neonicotinoids are often more toxic to bees when exposure is via ingestion20,21,37,38, and solitary bees are more sensitive than either honey bees or bumble bees to oral exposure35,36,38, therefore using oral honey bee LD 50 values in this study may under-represent oral toxicity to hoary squash bees. Toxicity via contact exposure varies much less among species34,39, thus honey bee contact LC 50 values may adequately represent contact toxicity for adult female hoary squash bees. The evaluation of hazard to larval squash bees using adult honey bee LD 50 or LC 50 values in this study may underestimate hazard because of developmental stage differences in sensitivity to neonicotinoids6,20,21,40. Contact lethal doses are higher than oral lethal doses for the residues we detected (Table 1).

Thiamethoxam appears to pose minimal hazard because residues were only detected in a single sample at concentrations below the limit of quantification. However, an absence of thiamethoxam residues may be the result of rapid metabolization to clothianidin, which was detected in our soil samples6,7. Both clothianidin (HQ = 1.82) and imidacloprid (HQ = 2.76) pose a hazard in Cucurbita growing systems in Ontario because they are detected frequently, and their respective HQs exceed one when summed across all exposure matrices. Imidacloprid appears to be more hazardous in Cucurbita-crops than clothianidin based on this approach because it was found at higher concentrations in soil and was present in both nectar and pollen (Table S3). The combined hazard of all neonicotinoids in the system was also high (HQ combined neonicotinoid = 4.59), suggesting that hoary squash bee populations may be exposed to lethal concentrations of neonicotinoids in a worst-case scenario. Combined hazard from chlorantraniliprole in all matrices was low (HQ = 0.02), likely because it has a much higher LC 50 than neonicotinoids.

There is common agreement that bees can be exposed to neonicotinoids from nectar and pollen18,20,21. Although neither pollen nor nectar were deemed hazardous here, HQs were higher for adult hoary squash bees via Cucurbita nectar (HQ nectar = 0.45) than via pollen (HQ pollen = 0.03). This was because adult exposure to pollen was considered to be contact rather than oral (as adult consumption of pollen was not evaluated), whereas adult exposure to nectar was oral and also likely an overestimation based on honey bee nectar consumption values. Although no hazard from pollen or nectar was found for the lethal dose endpoint, sublethal effects are still possible at these low HQs.

Currently, there are no studies that evaluate the risk to ground-nesting bees from direct exposure to neonicotinoids in soil, although some studies assess effects on other soil fauna41. Because both imidacloprid and thiamethoxam (which metabolizes to clothianidin) are applied to the soil in Cucurbita crops, and may persist in the soil for longer than a single growing season in Canada13, it is unsurprising that hazard to ground-nesting hoary squash bees from neonicotinoids in soil (HQ soil = 4.32) is much higher than even the combined hazard from neonicotinoids in both pollen and nectar (HQ pollen+nectar = 0.27; Table 1). Therefore, soil appears to be the most important route of exposure to systemic pesticides for hoary squash bees.

The combined hazard from insecticides for adult female hoary squash bees from all exposure matrices (soil, pollen, and nectar) was high, with 93% of this hazard attributable to neonicotinoids in soil (Table 1). Hoary squash bees can construct more than one nest per season when environmental conditions (e.g. nectar and pollen resources, weather) permit17. However, female hoary squash bees in this study were already exposed to doses above lethal levels of both imidacloprid and clothianidin (HQs > 1) during the construction of a single nest, rendering it unlikely they could construct another. Indeed, under present soil conditions in Ontario Cucurbita crops, pesticide exposure may preclude the construction of even a single 5-cell nest in a season, even if all other conditions are favourable. Excluding exposure from nectar, which was not evaluated, hazard to larval stages was low (HQ larvae pollen = 0.06). It is reasonable to expect that hazard from nectar is also low for larval stages because their consumption of nectar is much less than for adult females as they do not undertake energy-expensive nest construction or foraging activities. For honey bees, adult pollen foragers consume 150% more nectar than larvae42. Here we have assumed that larval hoary squash bees are protected from direct exposure to neonicotinoids in soil by the waterproof nature of the nest cell lining17, an assumption that requires further critical investigation.

The percentage translocation of insecticide residues from soil to bees is currently unknown. Our initial assumption was translocation at 100%, but recognizing the uncertainty around translocation rates, we also present exceedances for four alternative scenarios with lower rates (Table S8). Using a probabilistic approach for the hoary squash bee in the acute exposure scenario (48 h, 2.23 g soil; Fig. 2, Table S6) at 100% translocation, the probability of exceedance of the mean honey bee LC 50 was below 5% for imidacloprid and clothianidin. However, for the solitary bee surrogate LC 50 endpoint residues of both imidacloprid (31.2%) and clothianidin (28.3%) in soil exceeded 5% (Fig. 2, Table S6). In the chronic exposure scenario, the amount of exposure to soil (33.5 g soil, 30 days) was much greater, and exceedance for imidacloprid and clothianidin was greater than 5% for all lethal endpoints (Fig. 3a,b, Table S6). It is likely that at least some of the clothianidin found in Cucurbita crop soil is from the application of thiamethoxam to seeds6. Hilton et al.7 found that ~3–46% of the residues recovered from soil sampled more than 60 days after thiamethoxam application were the metabolite clothianidin. Further work on the fate of thiamethoxam in Cucurbita crop fields is needed to determine whether seed-applied thiamethoxam poses a risk to hoary squash bees via its metabolite, clothianidin. Exceedance for chlorantraniliprole was not greater than 5% for any exposure endpoint in either exposure scenario, suggesting that it did not pose a risk in Cucurbita-crop soils in 2016 (Fig. 3c, Table S6).

There were at least three issues generating uncertainty during our assessment of risk to hoary squash bees posed by neonicotinoid residues in soil. The lack of information about insecticide toxicity for this, or indeed any other solitary ground-nesting species, is a large knowledge gap. As hoary squash bees are similar in size to honey bees, they may be well represented by the available toxicity data for honey bees, especially for contact exposure which tends to vary less among species than oral exposure39. Secondly, although soil-applied neonicotinoids are known to elicit negative effects on Lepidoptera pupae43, Carabid beetles44, Hexapoda, Collembola and Thysanoptera that live in soil45, we could find no information on the extent to which insecticide residues in soil can pass through the cuticle or into spiracles of ground-nesting bees. We have assumed a worst-case scenario in which all the soil residues are translocated during exposure, but this is unlikely, although neonicotinoids have relatively low organic carbon-water partition coefficients (Table S4)46. However, even at 10% translocation, exceedances were greater than 5% for all imidacloprid endpoints, and also for the lowest honey bee LC 50 and solitary bee surrogate LC 50 for clothianidin, for chronic exposure in Cucurbita crop soils. In field crops, exceedances were greater than 5% for all endpoints for clothianidin assuming 10% translocation in a chronic exposure scenario (Table S8). Lastly, lower residue detection limits for soil are needed to align them with lethal dose concentrations for some compounds (Table S3). Confidence intervals for the EEDs in Cucurbita-crop soil are large due to limited sample size and high LODs. Interestingly, even at the extreme high values for confidence intervals (where exceedance would be lowest), exceedance remains above 5% for the solitary bee surrogate LC 50 for clothianidin and imidacloprid in both acute and chronic scenarios (Figs 2 and 3). Despite its limitations, this study is significant because it is the first evaluation of risk from insecticide residues in soil for any ground-nesting bee species.

Because little information about exposure to soil for most ground-nesting bee species exists, we used hoary squash bee exposure to soil as a surrogate for other ground-nesting bees. We estimated soil exposure of 33.5 g for hoary squash bee females, which is comparable to data for some other ground-nesting bee species. For example, soil exposure for Andrena prunorum or Nomia melanderi females are estimated as 30.23 g47 and 26.3 ± 5.7 g48 respectively. The difference between the low-end estimate of soil excavated by N. melanderi (20.6 g) and hoary squash bees (33.5 g) could represent as much as 38.5% of the latter species’ exposure, highlighting the potential variability in exposure via soil across species and the limitations of the hoary squash bee model. Ground-nesting bee species vary greatly in size14, and many are much smaller than hoary squash bees. For ground-nesting solitary bees, tunnel diameter is related to bee size because bees excavate tunnels and nest cells large enough for themselves14. Although other solitary ground-nesting bee species may be appreciably larger or smaller than squash bees, the ratio of their body size to the volume of soil they excavate when building a nest may be similar, providing a possible future basis upon which to compare exposure for different solitary bee species. Smaller bees may have lower exposure to neonicotinoids in soil because they construct narrower tunnels and nest cells to fit their smaller bodies14, therefore contacting lower soil volumes overall. However smaller bee species may be more sensitive to insecticide exposure16,34,49. Bee body size is not necessarily correlated to the depth that vertical tunnels in nests are excavated in soil50. Solitary bees are physiologically limited in their reproductive capacity and generally build 1–8 brood cells per nest14. Until more information emerges, the hoary squash bee is the best model available to evaluate risk from exposure to pesticide residues in soil for ground-nesting bees in general and provides a starting point to understand risk from insecticides residues in soil for these bees.

Neonicotinoid residues detected in Ontario agricultural soils reflect variation in usage for different crops. For Cucurbita crops, imidacloprid and clothianidin were the most commonly detected neonicotinoids, with a single detection of thiamethoxam at an unquantifiable concentration. For field crops, clothianidin and thiamethoxam were more commonly detected. Exposure to clothianidin residues for bees nesting in field crop soils appeared to be ubiquitous: 96.34% of soil samples taken before spring planting contained clothianidin applied in the previous season. The probabilities of exceedance were high for all clothianidin exposure endpoints for both the acute (Fig. 4) and chronic (Fig. 5) scenarios. As clothianidin-treated seeds are planted in a new cropping cycle, releasing more residues into the soil, these exceedances would likely increase. About 70% of the solitary bee species in eastern Canada nest in the ground51, many of which are associated with agriculture, including species in the genera Agapostemon, Andrena, Anthophora, Colletes, Eucera, Halictus, Lasioglossum, Megachile and Melissodes52,53. Ground-nesting bees from 13 genera have been collected in corn and soybean fields in Iowa54,55. Although this does not prove that these species were nesting within fields, the small foraging ranges of solitary bees56 suggest that many likely were. Various bees are active at different times during the season51,57. Those species active in the early spring may be exposed to the minimum residue concentrations described here, but those active post-planting may be exposed to much higher concentrations in soil.

Taken together, this evidence suggests the risk to ground-nesting bees from exposure to clothianidin in field crop soil is high, necessitating action to mitigate such risks to preserve pollination services. If clothianidin residues found in soil are metabolites of applied thiamethoxam, then use of thiamethoxam should also be addressed.

Thiamethoxam is used as a seed treatment on both field corn and soybean crops in Ontario and was applied to 1.98 million acres (66% of treated acres) in the 2016 season58. For the chronic exposure scenario, the risk to ground-nesting bees from thiamethoxam was high for all exposure endpoints (Fig. 5b). For the acute exposure scenario, the risk from thiamethoxam was less than 5% for all exposure endpoints except the solitary bee surrogate LC 50 (25.56%: Fig. 4b). The apparently lower risk to ground-nesting bees from thiamethoxam may be the product of its tendency to break down quickly into clothianidin in soil6. Although the risk to ground-nesting bees from acute exposure to imidacloprid in field crop soil was below the 5% threshold for all exposure endpoints (Fig. S1), exceedance rose above the threshold for the solitary bee surrogate LC 50 (9.24%) under the chronic exposure scenario (Fig. S2). The lower risk associated with imidacloprid in field crop soil may be because it is used in only 11% of treated field crop acres58. One of the main concerns around neonicotinoid insecticide exposure for ground-nesting bees is their use in soil applications as treated seed. This has been partially mitigated in Ontario by increased regulation of neonicotinoid-treated corn and soybean seed59 but has not yet been addressed for other crops.

In conclusion, neonicotinoid residues in soil pose a high risk to female hoary squash bees as they construct their nests in Cucurbita-crop growing systems or in field crop soils. These demonstrable risks for hoary squash bees seem likely to be applicable to other species of ground-nesting bees nesting in agricultural soils. Further work is needed to determine the relative sensitivity of the hoary squash bee to neonicotinoid exposure compared to honey bees, and to explicitly determine the extent and impacts of larval exposure in soil. Advances in analytical techniques are also needed to achieve lower limits of detection in soil that mirror lethal endpoints for solitary bees. Recognition and mitigation of risks from exposure to neonicotinoids in agricultural soil are urgently needed to protect these important crop pollinators.