Experiment One
Pesticide type explained 43 percent of the variation in Bi. glabrata biomass (Table S4), whereas most of the variation in Bu. truncatus biomass (35 percent) was explained by pesticide identity nested within class nested within type (Table S4). Likelihood ratio tests determined that the above pesticide effects were significant for both snail species (Table S5). Model selection favored using pesticide type and pesticide nested within class within type for Bi. glabrata and Bu. truncatus biomass, respectively, and the lowest AICc values included a term for zero-inflation and treating water and solvent as a single control group for both snail species (Tables S6-S7). Model selection (Table S6) indicated significant positive fertilizer effects (bottom-up) on Bi. glabrata biomass (Fig. 1a; Table 1). In contrast, bottom-up effects of herbicides for Bi. glabratabiomass were generally not significant in the presence of predators (Table 1; p > 0.05). Insecticides were associated with positive top-down effects on Bi. glabrata biomass (Fig. 1a; Table 1). Similar to Bi. glabrata , fertilizers had significant positive bottom-up effects for Bu. truncatus biomass, after model selection (Table S7), while none of the six herbicides were significantly associated with Bu. truncatus biomass in the presence of predators (Fig. 1b; Table 1). Similar to Bi. glabrata , insecticides were associated with positive top-down effects on Bu. truncatus biomass (Fig. 1b). Four insecticides led to significant increases in average Bu. truncatus biomass relative to the controls (chlorpyrifos: 55% increase, terbufos: 36%, esfenvalerate: 28%, and permethrin: 61%), while one insecticide from each insecticide class had no significant effects (malathion and λ-cyhalothrin both changed average biomass by < 10%).
Structural equation modeling (SEM) was consistent with the above relationships (Fig. 2). The final SEM was a good fit for Bu. truncatus (CFI = 0.99, χ2 = 15.25, df = 26, p = 0.95; Table S8) and Bi. glabrata (CFI = 0.99, χ2 = 28.04, df = 26, p = 0.36; Table S9). The SEM model for Bu. truncatus suggested positive and indirect top-down effects of insecticides on snail biomass by lowering 24 hr crayfish survival, which lowered final crayfish abundance (Fig. 2). Fertilizer increased both phytoplankton and periphyton chlorophyll a, the latter of which had positive but non-significant effects upon snail eggs and biomass (Table S8). Instead, significant positive bottom-up effects of fertilizer occurred via increasing submerged vegetation, which increased both snail eggs and biomass (Fig. 2). The sum of standardized coefficients of indirect pathways from fertilizer to snail eggs or biomass was over an order of magnitude greater via submerged vegetation than via periphyton chlorophyll a alone. Submerged vegetation was reduced by crayfish via consumption, which also formed a significant and negative indirect pathway from crayfish predators to snail eggs or biomass that was independent of crayfish predation on snails (Fig. 2). Thus, insecticide-induced reductions in crayfish were positively associated with snails both by reducing predation and increasing snail habitat. The significance and direction of pathways in the SEM model for Bi. glabrata were identical to Bu. truncatus except that both snail eggs and submerged vegetation pathways were negative for Bi. glabrata biomass (Table S9). Bi. glabrata biomass at the end of the experiment was also approximately an order of magnitude greater than for Bu. truncatus (Table S10).