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When a Tritrophic Interaction Goes Wrong to the Third Level: Xanthoxylin From Trees Causes the Honeybee Larval Mortality in Colonies Affected by the River Disease

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The “River Disease” (RD), a disorder impacting honeybee colonies located close to waterways with abundant riparian vegetation (including Sebastiania schottiana, Euphorbiaceae), kills newly hatched larvae. Forager bees from RD-affected colonies collect honeydew excretions from Epormenis cestri (Hemiptera: Flatidae), a planthopper feeding on trees of S. schottiana. First-instar honeybee larvae fed with this honeydew died. Thus, we postulated that the nectars of RD-affected colonies had a natural toxin coming from either E. cestri or S. schottiana. An untargeted metabolomics characterization of fresh nectars extracts from colonies with and without RD allowed to pinpoint xanthoxylin as one of the chemicals present in higher amounts in nectar from RD-affected colonies than in nectars from healthy colonies. Besides, xanthoxylin was also found in the aerial parts of S. schottiana and the honeydew excreted by E. cestri feeding on this tree. A larva feeding assay where xanthoxylin-enriched diets were offered to 1st instar larvae showed that larvae died in the same proportion as larvae did when offered enriched diets with nectars from RD-colonies. These findings demonstrate that a xenobiotic can mimic the RD syndrome in honeybee larvae and provide evidence of an interspecific flow of xanthoxylin among three trophic levels. Further, our results give information that can be considered when implementing measures to control this honeybee disease.

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We would like to thank Horacio Pezaroglo, Andrés López and Guillermo Moyna for technical assistance in the NMR analyses.


Financial support was received from grants from INIA (Instituto Nacional de Investigación Agropecuaria, FPTA 2012–322) and CSIC (Comisión Sectorial de Investigación Científica, Universidad de la República, Uruguay. Programa Grupos, CSIC-Grupos 2004).

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All authors contribute to the different experiments: CR, LA and ARB conceived and performed chemical analyses; DA, KA and CR performed larval assay. CI, ES and KA conceived the presented idea. ES and CI performed field sampling. CR wrote the manuscript with input from DA, KA, ES and CI. CR and CI were responsible of the funding grants.

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Correspondence to Carmen Rossini.

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Rossini, C., Almeida, L., Arredondo, D. et al. When a Tritrophic Interaction Goes Wrong to the Third Level: Xanthoxylin From Trees Causes the Honeybee Larval Mortality in Colonies Affected by the River Disease. J Chem Ecol 47, 777–787 (2021).

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