Ammonium sulfate [(NH₄)₂SO₄] deposition and elevated ozone (O₃) concentrations may negatively affect plants and trophic interactions. This study aimed to evaluate for the first time the interactive effects of high (NH₄)₂SO₄ load and elevated O₃ levels on cauliflower (Brassica oleracea L.) under field conditions. Cauliflower seedlings were treated with 0 (AS0) or 50 (AS50) kg ha⁻¹ (NH₄)₂SO₄ and exposed to ambient (AOZ, ≈20 ppb) or elevated (EOZ, ≈55 ppb) O₃ for about one month, in a Free Air O₃ Concentration Enrichment (FACE) system. The oligophagous diamondback moth (Plutella xylostella Linnaeus, 1758) showed a clear preference towards the seedlings treated with AS50, which intensively grazed. Plant-herbivore interactions were driven by (NH₄)₂SO₄ availability, rather than O₃, via increased nitrogen content in the leaves. Further laboratory bioassays were followed to confirm the validity of these observations using polyphagous Eri silkmoth larvae (Samia ricini) as a biological model in a standardized experimental setup. Choice assays, where larvae could select leaves among leaf samples from the different experimental conditions, and no-choice assays, where larvae could graze leaves from just one experimental condition, were conducted. In the choice assay, the larvae preferred AS50-treated leaves, in agreement with the field observations with diamondback moth. In the no-choice assay, larval body mass growth was inhibited when fed with leaves treated with EOZ and/or AS50. Larvae fed with AS50-treated leaves displayed increased mortality. These observations coincide with higher NO₃ and Zn content in AS50-treated leaves. This study shows that plant-herbivore interactions can be driven by (NH₄)₂SO₄ availability, independently of O₃, and suggests that high N deposition may have severe health implications in animals consuming such plant tissues.

Key message: Plant-herbivore interactions are driven by high (NH₄)₂SO₄ availability, independently of O₃.

The United States Environmental Protection Agency (US EPA) has recently proposed changes to strengthen the transparency of its pivotal regulatory science policy and procedures. In this context, the US EPA aims to enhance the transparency of dose-response data and models, proposing to consider for the first time non-linear biphasic dose-response models. While the proposed changes have the potential to lead to markedly improved ecological risk assessment compared to past and current approaches, we believe there remain open issues for improving the quality of ecological risk assessment, such as the consideration of adaptive, dynamic and interactive effects. Improved risk assessment including adaptive and dynamic non-linear models (beyond classic threshold models) can enhance the quality of regulatory decisions and the protection of ecological health. We suggest that other countries consider adopting a similar scientific-regulatory posture with respect to dose-response modeling via the inclusion of non-linear biphasic models, that incorporate the dynamic potential of biological systems to adapt (i.e., enhancing positive biological endpoints) or maladapt to low levels of stressor agents.

Evaluations of ozone effects on vegetation across the globe over the last seven decades have mostly incorporated exposure levels that were multi-fold the preindustrial concentrations. As such, global risk assessments and derivation of critical levels for protecting plants and food supplies were based on extrapolation from high to low exposure levels. These were developed in an era when it was thought that stress biology is framed around a linear dose-response. However, it has recently emerged that stress biology commonly displays non-linear, hormetic processes. The current biological understanding highlights that the strategy of extrapolating from high to low exposure levels may lead to biased estimates. Here, we analyzed a diverse sample of published empirical data of approximately 500 stimulatory, hormetic-like dose-responses induced by ozone in plants. The median value of the maximum stimulatory responses induced by elevated ozone was 124%, and commonly <150%, of the background response (control), independently of species and response variable. The maximum stimulatory response to ozone was similar among types of response variables and major plant species. It was also similar among clades, between herbaceous and woody plants, between deciduous and evergreen trees, and between annual and perennial herbaceous plants. There were modest differences in the stimulatory response between genera and between families which may reflect different experimental designs and conditions among studies. The responses varied significantly upon type of exposure system, with open-top chambers (OTCs) underestimating the maximum stimulatory response compared to free-air ozone-concentration enrichment (FACE) systems. These findings suggest that plants show a generalized hormetic stimulation by ozone which is constrained within certain limits of biological plasticity, being highly generalizable, evolutionarily based, and maintained over ecological scales. They further highlight that non-linear responses should be taken into account when assessing the ozone effects on plants.

Ground-level ozone (O₃) levels are nowadays elevated in wide regions of the Earth, causing significant effects on plants that finally lead to suppressed productivity and yield losses. Ethylenediurea (EDU) is a chemical compound which is widely used in research projects as phytoprotectant against O₃ injury. The EDU mode of action remains still unclear, while there are indications that EDU may contribute to plants with nitrogen (N) when the soil is poor in N and the plants have relatively small leaf area. To reveal whether the N content of EDU acts as a fertilizer to plants when the soil is not poor in N and the plants have relatively large total plant leaf area, willow plants (Salix sachalinensis Fr. Schm) were exposed to low ambient O₃ levels and treated ten times (9-day interval) with 200 mL soil drench containing 0, 800 or 1600 mg EDU L⁻¹. Fertilizer was added to a nutrient-poor soil, and the plants had an average plant leaf area of 9.1 m² at the beginning of EDU treatments. Indications for EDU-induced hormesis in maximum electron transport rate (J_max) and ratio of intercellular to ambient CO₂ concentration (C_i:C_a) were observed at the end of the experiment. No other EDU-induced effects on leaf greenness and N content, maximum quantum yield of photosystem II (F_v/F_m), gas exchange, growth and matter production suggest that EDU did not act as N fertilizer and did not cause toxicity under these experimental conditions.

Ground-level ozone (O₃) concentrations have been elevating in the last century. While there has been a notable progress in understanding O₃ effects on vegetation, O₃ effects on ecological stoichiometry remain unclear, especially early in the oxidative stress. Ethyelenediurea (EDU) is a chemical compound widely applied in research projects as protectant of plants against O₃ injury, however its mode of action remains unclear. To investigate O₃ and EDU effects early in the stress, we sprayed willow (Salix sachalinensis) plants with 0, 200 or 400 mg EDU L⁻¹, and exposed them to either low ambient O₃ (AOZ) or elevated O₃ (EOZ) levels during the daytime, for about one month, in a free air O₃ controlled exposure (FACE); EDU treatment was repeated every nine days. We collected samples for analyses from basal, top, and shed leaves, before leaves develop visible O₃ symptoms. We found that O₃ altered the ecological stoichiometry, including impacts in nutrient resorption efficiency, early in the stress. The relation between P content and Fe content seemed to have a critical role in maintaining homeostasis in an effort to prevent O₃-induced damage. Photosynthetic pigments and P content appeared to play an important role in EDU mode of action. This study provides novel insights on the stress biology which are of ecological and toxicological importance.