Cross-taxa sublethal impacts of plant protection products on honeybee in-hive and zebrafish swimming behaviours at environmentally relevant concentrations.

Single and mixture exposure to plant protection products (PPPs) can affect non-target organisms at sublethal concentrations, yet the ecological relevance of behavioural effects remains underexplored. Behavioural disruptions can compromise survival and fitness, with exposure occurring across terrestrial and aquatic ecosystems. Here, we assess the behavioural impact of environmentally relevant PPP concentrations on two ecologically and toxicologically important model species: honeybees (Apis mellifera) and zebrafish (Danio rerio). These organisms represent distinct exposure pathways: pollinator-specific routes such as oral uptake of contaminated nectar and pollen or contact during overspray and foraging, and freshwater contamination via runoff. In honeybees, in-hive behaviours were monitored using a snapshot method from days three to ten after exposure, while zebrafish behavioural endpoints were quantified using a 26-endpoint visual and acoustic motor response (VAMR) assay. Sublethal PPP exposure (1.99-7.81 ng/µL in honeybee hives, 0.0253-27.5 ng/µL in zebrafish assay) caused significant, substance-specific behavioural alterations. In honeybees, flupyradifurone (SIVANTO® prime) significantly decreased foraging and nectar processing, while boscalid (Cantus®) and terbuthylazine (ClickPro®) minimised brood-tending behaviours. Insecticides and fungicides affected honeybees most, while zebrafish embryos were especially sensitive to the herbicide terbuthylazine. They exhibited concentration-dependent neurotoxic phenotypes, with behavioural profiles of the PPPs mixture (consistent with concentrations in German streams: 41.54 % boscalid, 0.013 % flupyradifurone, 58.45 % terbuthylazine) shifting along a terbuthylazine-boscalid gradient. These findings show that PPPs can elicit pronounced behavioural changes in non-target species, even at low environmental concentrations. These results support incorporating in-hive and early-life stage behavioural assays into pesticide risk assessments and warrant mechanistic studies on PPP-induced neurotoxicity.