Acaricide, fungicide and drug interactions in honey bees (Apis mellifera)

Abstract

Background: Chemical analysis shows that honey bees (Apis mellifera) and hive products contain many pesticides derived from various sources. The most abundant pesticides are acaricides applied by beekeepers to control Varroa destructor. Beekeepers also apply antimicrobial drugs to control bacterial and microsporidial diseases. Fungicides may enter the hive when applied to nearby flowering crops. Acaricides, antimicrobial drugs and fungicides are not highly toxic to bees alone, but in combination there is potential for heightened toxicity due to interactive effects.

Methodology/principal findings: Laboratory bioassays based on mortality rates in adult worker bees demonstrated interactive effects among acaricides, as well as between acaricides and antimicrobial drugs and between acaricides and fungicides. Toxicity of the acaricide tau-fluvalinate increased in combination with other acaricides and most other compounds tested (15 of 17) while amitraz toxicity was mostly unchanged (1 of 15). The sterol biosynthesis inhibiting (SBI) fungicide prochloraz elevated the toxicity of the acaricides tau-fluvalinate, coumaphos and fenpyroximate, likely through inhibition of detoxicative cytochrome P450 monooxygenase activity. Four other SBI fungicides increased the toxicity of tau-fluvalinate in a dose-dependent manner, although possible evidence of P450 induction was observed at the lowest fungicide doses. Non-transitive interactions between some acaricides were observed. Sublethal amitraz pre-treatment increased the toxicity of the three P450-detoxified acaricides, but amitraz toxicity was not changed by sublethal treatment with the same three acaricides. A two-fold change in the toxicity of tau-fluvalinate was observed between years, suggesting a possible change in the genetic composition of the bees tested.

Conclusions/significance: Interactions with acaricides in honey bees are similar to drug interactions in other animals in that P450-mediated detoxication appears to play an important role. Evidence of non-transivity, year-to-year variation and induction of detoxication enzymes indicates that pesticide interactions in bees may be as complex as drug interactions in mammals.

Figure 1. Median lethal dose (LD₅₀) of acaricides to honey bees in 2009 following sublethal treatment with acaricides, fungicides, or detoxicative enzyme inhibitors.

Confidence intervals (95%) are indicated below the LD₅₀ values. Significant differences compared to the control treatment are indicated with a superscript letter: a = significant pre-treatment effect, b = significant pre-treatment*acaricide dose effect (Table S1). LD₅₀ values taken from previous work: † = , ‡ = . Names for classical enzyme inhibitors are abbreviated as follows DEM = diethyl maleate, DEF = S,S,S-tributylphosphorotrithioate, PBO = piperonyl butoxide. A dash “−” indicates an LD₅₀ that could not be calculated because of insufficient data.

Figure 2. Median lethal doses (LD₅₀) of acaricides to honey bees in 2010 fed antimicrobial drugs used in beekeeping.

Confidence intervals (95%) are indicated below the LD₅₀ values. Significant differences compared to the respective treatment are indicated with a superscript letter a = significant pre-treatment effect, b = significant pre-treatment*acaricide dose effect (Table S2).

Figure 3. Plot of log-transformed dose and probit-transformed honey bee mortality data for tau-fluvalinate toxicity after oxalic acid or control pre-treatments.

Symbols represent raw mortality and solid lines indicate lines fitted using the full model. Dotted lines represent 95% confidence intervals for each line fit with the full model. Dashed green lines were fitted using the same slope for both pre-treatments. The dashed blue line was fitted with combined data from both pre-treatments. Likelihood ratio tests comparing the full model and the reduced models were used to determine pre-treatment effects and pre-treatment * dose effects.

Figure 4. Median lethal doses (LD₅₀) for tau-fluvalinate in honey bees pre-treated with piperonyl butoxide (PBO) or a sterol biosynthesis inhibiting (SBI) fungicide at three dose levels.

Significant differences compared to the control treatment are indicated with a superscript letter a = significant pre-treatment effect, b = significant pre-treatment*acaricide dose effect (Table S3).