Evaluating toxicity of varroa mite (Varroa destructor)-active dsRNA to monarch butterfly (Danaus plexippus) larvae

Authors: KRISHNAN, NIRANJANA - Iowa State University; HALL, MAURA - Iowa State University; Hellmich Ii, Richard; COATS, JOEL - Iowa State University; BRADBURY, STEVEN - Iowa State University

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/5/2021
Publication Date: 6/2/2021
Citation: Krishnan, N., Hall, M.J., Hellmich II, R.L., Coats, J.R., Bradbury, S.P. 2021. Evaluating toxicity of varroa mite (Varroa destructor)-active dsRNA to monarch butterfly (Danaus plexippus) larvae. PLoS ONE. 16(6). Article e0251884. https://doi.org/10.1371/journal.pone.0251884.
DOI: https://doi.org/10.1371/journal.pone.0251884

Interpretive Summary: Varroa mites are parasites that feed on developing honey bees (brood) that contribute to high colony losses. These mites are a major concern for beekeepers worldwide. A new approach to control this pest uses double-stranded RNA (dsRNA) to silence a gene vital to this mite called calmodulin. This study is part of an environmental risk assessment of this product. The 372 base pair sequence of this gene contains a 21 base pair match with the same gene of the monarch butterfly. Although unlikely, this raises the possibility that environmental exposure could have non-target effects on this popular butterfly. In this study, throughout the entire larval stage, larvae were exposed to milkweed leaves treated with one- and ten-fold estimated environmental concentrations of the Varroa-active dsRNA. Potassium arsenate and a previously designed monarch dsRNA with a 100% base pair match to the monarch V-ATPase gene were used as positive controls. The Varroa mite and monarch dsRNA compounds did not cause significant differences in larval mortality, larval or pupal development, pupal weights, or adult emergence rates when compared to negative controls. These results suggest this new approach to controlling Varroa mites would have no detrimental effects on monarch butterflies. This information is useful to scientists interested using dsRNA technology to control any type of pest, but it is especially useful to scientists and beekeepers interested in controlling Varroa mites.

Technical Abstract: Varroa mites (Varroa destructor) are parasitic mites that, combined with other factors, are contributing to high levels of honey bee (Aphis mellifera) colony losses. A Varroa-active dsRNA was recently developed to control Varroa mites within honey bee brood cells. This dsRNA has 372 base pairs that are homologous to a sequence region within the Varroa mite calmodulin gene (cam). The Varroa-active dsRNA also shares a 21 base pair match with monarch butterfly (Danaus plexippus) calmodulin mRNA, raising the possibility of non-target effects if there is environmental exposure. We chronically exposed the entire monarch larval stage to common (Asclepias syriaca) and tropical milkweed (Asclepias curassavica) leaves treated with one- and ten-fold estimated environmental concentrations of Varroa-active dsRNA. This corresponded to concentrations of 0.15-0.20 and 1.45-1.51 mg/g leaf, respectively. Potassium arsenate and a previously designed monarch dsRNA with a 100% base pair match to the monarch V-ATPase A mRNA (leaf concentration was 0.08-0.10 mg/g) were used as positive controls. The Varroa mite and monarch dsRNA compounds did not cause significant differences in larval mortality, larval or pupal development, pupal weights, or adult eclosion rates when compared to negative controls. Irrespective of control or dsRNA treatment, larvae that consumed approximately 7.5 to 10.5-gram milkweed leaf within 10 to 12 days had the highest pupal weights. The lack of mortality and sublethal effects following dietary exposure to dsRNA with 21 base pair and 100% base pair match to mRNAs that correspond to regulatory genes, suggest monarch mRNA may be refractory to silencing by dsRNA or monarch dsRNase activity may degrade dsRNA to a concentration that is insufficient to silence mRNA signaling.