Stage-specific effects of a fungicide and global warming on copper accumulation and development in a major vineyard insect pest

Bordeaux mixture (20% copper content, BB Caffaro WG) was incorporated into the larval diet at three concentrations: 25 mg.kg-1, 50 mg.kg-1, and 100 mg.kg-1. A control was also included (0 mg.kg-1). To assess Lobesia botrana responses under current and future climatic conditions, individuals exposed to different copper concentrations were placed in controlled environment chambers (Memmert HPP260eco, Schwabach, Germany).
A total of 3112 newly hatched larvae (<12 hours old) were used in all experiments. Larvae were randomly assigned to one of eight experimental groups, representing a full factorial design with four copper concentrations and two climatic conditions. Lobesia botrana undergoes five larval stages, which can be identified by the size of the head capsule. This anatomical feature grows in discontinuous stages and remains stable within each instar (Delbac et al., 2010). To avoid potential misclassification caused by environmental stressors such as copper or temperature, the first experiment (experiment 1, “Data_Development;xlsx”) focused on measuring head-capsule width across all larval instars. This experiment also included developmental metrics (i.e., body mass and development time), based on a total sample of 931 larvae and 471 adults. The second experiment (experiment 2, “Data_Copper_content.xlsx”) aimed to quantify copper content in the body tissues of L. botrana at the 3rd and 5th larval instars, as well as in newly emerged males and females (<12 h post-emergence). Since head-capsule width was characterized in experiment 1 across all larval instars and treatment conditions, it was subsequently used in experiment 2 to ensure accurate identification of the 3rd and 5th larval instars prior to copper content analysis. In total, 644 individuals were sampled for copper quantification, including 3rd and 5th instar larvae and adults. Analyses were performed at the PEA²t platform of the Chrono-environment laboratory (Université Marie et Louis Pasteur, France).