Shrinking Body Size and Physiology Contribute to Geographic Variation and the Higher Toxicity of Pesticides in a Warming World

Abstract

S1 contains detailed information about the study populations and the associated thermal regimes during the experiment. S2 contains detailed information on measured chlorpyrifos concentrations and water quality. S3 contains detailed protocols of the physiological assays. S4 contains detailed material and methods, and results of the independent action model for mortality. S5 contains extended results. S6 contains extended discussion about general effects of the pesticide. The supporting Information contains eleven figures (S1-S11), eight tables (S1-S10), and is in total 45 pages. S2 Appendix S1. Details about the study populations and the associated thermal regimes during the experiment Study populations We studied three populations in Northern Europe (high latitude) and three populations in Southern Europe (low latitude). The populations at the high latitude came from southern Sweden: Hovgardsdammarna (57°14'24.3"N, 12°08'28.2"E) and Kalmar Dämme (56°40'04.6"N; 16°17'46.5"E), and from Denmark: Roskilde (55°39'09.8"N; 12°08'01.7"E). The populations at the low latitude came from southern France: Bassin de Réaltor (43°28'11.1"N; 05°19'44.1"E), La Durance (43°43'52.5"N; 05°44'53.0"E) and St. Martin de Crau (43°37'57.8"N; 04°46'55.1"E). About 30 mated females were caught per population. They were immediately placed individually in plastic cups (7.5 cm height, 3.5 cm diameter) and provided with wet filter paper for oviposition. Filter papers containing eggs were transferred to the laboratory (KU Leuven, Belgium) where the eggs were kept at a water temperature of 22 °C and a 14h:10h light-dark cycle. Hatchlings were daily fed Artemia nauplii ad libitum. Ten days after the eggs hatched, larvae were allocated individually to plastic cups (5 cm height, 6 cm diameter) filled with 90 mL dechlorinated tap water. From this day onwards, larvae were fed Artemia nauplii ad libitum during weekdays (Monday-Friday). Thermal regimes The thermal regimes were based on the current mean water temperature and the current maximum DTF during the summer period in shallow (< 1m) freshwater ponds inhabited by the study species at both latitudes. At the high-latitude sites there is a mean summer water temperature of 20 °C with a maximum DTF of 5 °C (temperatures fluctuate daily between 17.5 °C and 22.5 °C). At the low-latitude sites the mean summer water temperature is 24 °C with a S4 Figure S1. Data of measured water temperatures over a period of 24 hours are given for the six different thermal regimes: 3 levels of daily temperature fluctuations (constant = 0 °C, low = 5 °C and high = 10 °C) around a mean temperature of 20 °C (upper figure) and 24 °C (lower figure). Confidence intervals (95%) around the means of ca. 30 days are given by the grey area around the black lines. S5 Appendix S2: Information on measured chlorpyrifos concentrations and water quality Details about the pesticide solutions To obtain the CPF concentrations, a CPF stock solution of 1 mg/mL was made by dissolving CPF powder (Sigma-Aldrich, purity > 99%) in absolute ethanol (...

Negative bioenergetic responses to pesticides in damselfly larvae are more likely when it is hotter and when temperatures fluctuate

“…A key finding was that the net energy budget (CEA) of the damselfly larvae was lowered by high chlorpyrifos, and this mainly at 24 °C when combined with high DTF. This matches the higher chlorpyrifos-induced mortality under the combination of the high mean temperature and the presence of DTFs in the companion study (Verheyen and Stoks, 2019c). Notably, we could show the expected pattern that treatment groups with a lower CEA showed a higher mortality and a lower growth rate (cfr Goodchild et al, 2019).…”

“…Concentrations below 0.75 µg/L cause minor effects on growth rate while concentrations above 2.25 µg/L cause 50% mortality at 20 °C and are therefore less ideal to address our objectives (Op de Beeck et al, 2017, personal observations). This was confirmed in a companion study where at constant 20 °C the low chlorpyrifos concentration (0.75 µg/L) caused ~5.5% mortality, the medium chlorpyrifos (1.50 µg/L) caused ~15% mortality, and the high chlorpyrifos (2.25 µg/L) caused ~21 % mortality (Verheyen and Stoks, 2019c). Larvae of the study species may be continuously exposed to high chlorpyrifos concentrations (> 2.25 µg/L) for more than a week in edge-to-field water bodies.…”

“…Warm-adapted low-latitude and cold-adapted high-latitude populations (Debecker and Stoks, 2019) were used to test for spatial differences in the effects of the stressors. In a companion study we have shown that both increases in mean temperature and in DTF, and especially their combination made chlorpyrifos more toxic in terms of growth and mortality (Verheyen and Stoks, 2019c). To determine the cellular energy allocation (CEA), we quantified the energy availability by measuring the key energy reserve molecules (proteins, sugars and fat), and the energy consumption by measuring the electron transport system (ETS) activity at the mitochondrial level (following De Coen and Janssen, 2003).…”

“…To explore relationships between CEA and life history (mortality and growth rate) at the treatment group level (n = 48 treatment combinations), we correlated treatment group means for CEA obtained in current study with those for life history obtained in the companion study (Verheyen and Stoks, 2019c). We did so using Pearson's product moment correlation.…”

“…Janssens et al, 2014;Scheil et al, 2010), and to upregulate levels of the detoxification enzyme cytochrome P450 monooxygenase (CytP450; e.g. Verheyen and Stoks, 2019a;Verheyen and Stoks, 2019c). Moreover, the food intake may have been lower under chlorpyrifos exposure (e.g.…”