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Assessing the population relevance of endocrine‐disrupting effects for nontarget vertebrates exposed to plant protection products

Assessing the population relevance of endocrine‐disrupting effects for nontarget vertebrates... Integrated Environmental Assessment and Management — Volume 15, Number 2—pp. 278–291 278 Received: 20 June 2018 Returned for Revision: 9 October 2018 Accepted: 21 November 2018 | | Environmental Policy & Regulation Assessing the Population Relevance of Endocrine-Disrupting Effects for Nontarget Vertebrates Exposed to Plant Protection Products Mark Crane,*y Nina Hallmark,z Laurent Lagadic,§ Katharina Ott,k Dan Pickford,# Thomas Preuss,§ Helen Thompson,# Pernille Thorbek,#yy Lennart Weltje,k and James R Wheelerzz yAG-HERA, Faringdon, United Kingdom zBayer SAS, Crop Science Division, Regulatory Toxicology, Sophia-Antipolis Cedex, France §Bayer AG, Crop Science Division, Environmental Safety, Monheim am Rhein, Germany kBASF SE, Crop Protection—Ecotoxicology, Limburgerhof, Germany #Syngenta, Jealott's Hill International Research Station, Bracknell, United Kingdom yyPresent address: BASF SE, APD/EE, Limburgerhof, Germany zzCorteva Agriscience, Agriculture Division of DowDuPont, Oxfordshire, United Kingdom ABSTRACT The European Commission intends to protect vertebrate wildlife populations by regulating plant protection product (PPP) active substances that have endocrine-disrupting properties with a hazard-based approach. In this paper we consider how the Commission’s hazard-based regulation and accompanying guidance can be operationalized to ensure that a technically robust process is used to distinguish between substances with adverse population-level effects and those for which it can be demonstrated that adverse effects observed (typically in the laboratory) do not translate into adverse effects at the population level. Our approach is to use population models within the adverse outcome pathway framework to link the nonlinear relationship between adverse effects at the individual and population levels in the following way: (1) use specific protection goals for focal wildlife populations within an ecosystem services framework; (2) model the effects of changes in population- related inputs on focal species populations with individual-based population models to determine thresholds between negligible and nonnegligible (i.e., adverse) population-level effects; (3) compare these thresholds with the relevant endpoints from laboratory toxicity tests to determine whether they are likely to be exceeded at hazard-based limits or the maximum tolerated dose/concentration from the experimental studies. If the population threshold is not exceeded, then the substance should not be classified as an endocrine disruptor with population-relevant adversity unless there are other lines of evidence within a weight-of-evidence approach to challenge this. We believe this approach is scientifically robust and still addresses the political and legal requirement for a hazard-based assessment. Integr Environ Assess Manag 2019;15:278–291. C 2018 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals, Inc. on behalf of Society of Environmental Toxicology & Chemistry (SETAC) Keywords: Endocrine disruption Plant protection product Vertebrate population European Union Hazard assessment INTRODUCTION populations of organisms (Munns et al. 2007); thus, The potential effects of endocrine-disrupting chemicals extrapolation from laboratory to field and from individual (EDCs) on wildlife have been a concern for many years organism to population is required. (Campbell and Hutchinson 1998; Kidd et al. 2007; Tyler et al. Some studies have demonstrated a strong link between 2008). Laboratory-based studies have shown that certain adverse endocrine effects found in the laboratory and chemicals can disrupt specific components of vertebrate adverse effects found in natural populations. For example, endocrine systems in individual organisms, but the regulatory Giesy et al. (2003) and Ottinger et al. (2011) review the effects goal for environmental risk assessment is usually to protect of endocrine disruptors in birds, including the effects of now- banned organochlorine substances on gull sex ratio and clutch “superabundance,” for which there is clear laboratory * Address correspondence to [email protected] and field-based evidence. However, other studies have Published 6 December 2018 on wileyonlinelibrary.com/journal/ieam. shown that many natural populations can compensate for This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in endocrine-mediated effects observed in the laboratory and any medium, provided the original work is properly cited. thereby maintain population numbers and biomass. For Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 279 example, a field study by Hamilton et al. (2014) supports the endocrine disruptor for regulatory purposes (WHO/IPCS conclusion that intersex and other signs of feminization in 2002; EC 2018). If this is not done, then useful products are individual fish observed in both the laboratory and the field likely to be banned on the basis of effects observed under do not necessarily result in adverse population-level effects. laboratory conditions that are unlikely to translate to the They sampled roach (Rutilus rutilus) from different southern population level in nature. It is worth noting that environ- English rivers and assessed the genetic structure of the mental risk assessments have been performed for decades to populations at each location. Despite widespread feminiza- help prevent adverse effects on vertebrate populations, tion of male roach in effluent-contaminated rivers, there was which, as shown by Matthiessen et al. (2018), may explain why no evidence of a correlation between effective population the evidence for endocrine disruption due to current-use size and predicted exposure to estrogens. In another study chemicals in wildlife populations is quite limited. with fish, Harris et al. (2011) examined the breeding ability of In an accompanying Learned Discourses paper (Crane intersex male roach when in competition with other males for et al. this issue), we discuss recent European Union (EU) regulation (EC 2018) and guidance (ECHA/EFSA 2018) females. They found that most intersex males were able to participate in spawning, and the reproductive success of intended to protect humans and vertebrate wildlife mildly intersex males was similar to nonintersex males, populations from adverse effects due to plant protection products (PPPs) containing an EDC. In this paper we although moderately and severely intersex males were less successful. describe possible approaches that complement this guid- Matthiessen and Weltje (2015) reviewed the effects of ance and would assist registrants and regulatory authorities in distinguishing between (i) substances with adverse azole compounds and their possible involvement in mascu- linization of wild fish populations to determine whether there endocrine-mediated population effects and (ii) substances is a biologically causal link between exposure to azoles and for which a potential endocrine disruption issue could be concluded from laboratory toxicity tests, but which would endocrine-disrupting effects in wild fish populations. They concluded that the available data on exposure and effects not translate into adverse effects on wildlife populations. provide reassurance that reported environmental concen- This activity is in response to, and complies with, the hazard-based approach that has been politically and legally trations of certain azoles are insufficient to cause adverse effects in fish by interference with their endocrine system, mandated by the European Commission (EC 2018). despite results from laboratory studies that show masculini- zation in fish under continuous exposure at significantly ASSESSMENT OF ADVERSE POPULATION EFFECTS higher concentrations. OF ENDOCRINE DISRUPTERS Recovery from endocrine-disrupting effects may also be The European Food Safety Authority (EFSA) benefits from reasonably rapid if the source of the endocrine-disrupting many expert opinions provided by their Scientific Committee substance is removed. For example, Kidd et al. (2007) (EFSA SC) and Panel on Plant Protection Products and their conducted a 7-year, whole-lake experiment at the Experi- Residues (EFSA PPR). We draw upon these opinions here to mental Lakes Area in Canada and showed that chronic assist in mapping out a coherent and conservative hazard- exposure of fathead minnow (Pimephales promelas) to low based approach (Figure 1) for demonstrating the population concentrations (5–6 ng  L ) of the potent estrogen 17a- relevance of endocrine disruptor effects observed in the ethynylestradiol (EE2) led to feminization and intersex in laboratory. males and altered oogenesis in females, leading to near The approach depends on clear identification of specific extinction of the population in the lake. However, in a follow- protection goals for focal wildlife populations within the up study, Blanchfield et al. (2015) quantified the physiologi- EFSA’s ecosystem services framework. The magnitude of cal, population, and genetic characteristics of the fathead relevant effects in toxicity tests performed at hazard-based minnow population for 7 years after EE2 additions ceased. thresholds (i.e., regulatory-defined limits or a maximum They found that 3 years after treatment, whole-body tolerated dose/concentration [MTD/C]) is compared to vitellogenin concentrations in male fathead minnow had population-relevant thresholds derived from population returned to baseline levels, there were no testicular modeling. The relevance of specific endpoints is taken abnormalities, and in the fourth year, adult size-frequency from an understanding of the adverse outcome pathway distribution and abundance had returned to pretreatment (AOP) concept (Ankley et al. 2010) focusing on adverse levels. effects caused by an endocrine disruptor at the individual Wild vertebrate populations may therefore be either level that may translate to the population level. If the resistant or resilient (for definitions, see Adverse Effects population threshold is not exceeded, then the substance section) to the effects of endocrine-disrupting substances, should not be classified as an endocrine disruptor with even when adverse effects of these substances are observed adverse population effects unless there are other lines of in laboratory tests. It is therefore important to establish a evidence within a weight-of-evidence approach to challenge biologically causal link between endocrine-disrupting effects this. This approach is analogous to that for human health in measured in laboratory toxicity tests and adverse population- which the relevance of animal model effects to humans can level effects on vertebrate wildlife in the field, because this is be challenged in the endocrine disruptor properties necessary to meet the requirements of the definition of an evaluation. Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 280 Integr Environ Assess Manag 15, 2019—M Crane et al. Figure 1. Evaluation scheme to determine the population relevance of laboratory-determined adverse endocrine-disrupting effects. Focal species EFSA (2009) distinguishes between “generic focal species” and “focal species” when assessing PPP risks to birds and It is not possible, desirable, or legally permissible to test mammals and defines them as follows: all vertebrate species in the laboratory, nor is it possible to perform risk assessments for every vertebrate species in an ecosystem. Therefore, the concept of “focal species” has A generic focal species is not a real species, however it is been adopted from conservation biology (Lambeck 1997) considered to be representative of all those species to focus assessments on those species of most relevance to potentially at risk, that is it is based on ecological perceived threats. For example, in the case of PPPs these knowledge of a range of species that could be at risk. It will be species that are present in specific agricultural has a high food intake rate and may consume a mixed landscapes and are thus most likely to be exposed to a diet rather than just one as for the indicator species. The PPP. Focal species may be keystone species (organisms diet is not real but is considered to be representative of with an effect on the ecosystem that is disproportionate to the species represented .... The “generic focal species” their numerical abundance), umbrella species (organisms is also considered to be a representative of the types of that cover a large geographical area in their daily or birds or mammals that occur across Member States. seasonal movements), flagship species (charismatic organ- isms with public appeal), or indicator species (organisms A focal species is a real species that actually occurs in the sensitive to change and therefore useful in monitoring crop when the PPP is being used. The aim of using a habitat quality). Focal species may fulfill more than one of “focal species” is to add realism to the risk assessment these criteria (e.g., they may be both an indicator and a insofar as the assessment is based on a real species that flagship species). uses the crop. It is essential that the species actually The concept of focal species has been adopted most fully occurs in the crop at a time when the PPP is being for birds and mammals in PPP assessment, although the applied. It is also essential that this species is considered concept has wider application for other vertebrate taxa and to be representative of all other species that may occur in has also been explored for fish (Ibrahim et al. 2013), the crop at that time. As a “focal species” needs to cover amphibians, and reptiles (EFSA PPR 2018). Mintram et al. all species present in the crop, it is possible that there (2017), in a review of modeling approaches for risk may be more than one “focal species” per crop. assessment of endocrine disruptors in fish, recommend that “the development of future models should include Focal bird and mammal species are identified for specific species representing a range of life-histories and.. .their exposure scenarios (i.e., a PPP application to a particular selection should be guided by the derivation of ecological crop type at a specific growth stage), ideally by use of scenarios which are relevant to major land use and water transect or field survey methods across a range of body types in which chemical exposures and effects are representative fields, although literature and census data predicted according to current risk assessments.” might also be useful (EFSA 2009). The survey data are then Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 281 We therefore also use the term “population” subsequently used to select focal species by considering their feeding strata, food intake rate, body weight, and diet to ensure in this paper as an operational description of any group of that species with the highest potential exposures are individuals from the same species that occurs in a particular space during a particular time, as defined by a regulatory considered. EFSA (2009) notes that a focal species is not automatically the species that was most frequently seen but authority. that it should represent the feeding guild(s) that raised Nontarget vertebrate wildlife populations provide a wide variety of ecosystem services including food (for human concern at earlier stages in the risk assessment and represent other species. consumption), genetic resources (biodiversity), education Selection of focal species when assessing the potential and inspiration, aesthetic values, pest and disease regulation (e.g., birds feeding on caterpillars), seed and propagule population relevance of endocrine disruptors should there- fore include the following considerations: dispersal, and recreation and ecotourism (e.g., bird-watch- ing, hunting, and fishing) (EFSA 2010). The EFSA SC (2016a) uses the concept of ecosystem services to derive specific 1) At least one relevant (generic) focal species should be selected for each regulatory exposure scenario (e.g., crop protection goals (SPGs) for service-providing units (SPUs). An and application scenario). Examples of generic focal bird SPU can be any ecological entity that provides an ecosystem service (provisioning, regulating, cultural, or supporting and mammal species are provided in EFSA (2009), and Ibrahim et al. (2013) have derived a list of focal fish species. services) to humans. EFSA SC (2016a) states that the An EFSA PPR (2018) opinion suggests that the following following need to be defined before setting an SPG: the ecological entity (e.g., individual, population, functional amphibian and reptile focal species should be considered: the great crested newt (Triturus cristatus), the natterjack group, or ecosystem), the attribute of that entity (e.g., toad (Epidalea calamita), the common treefrog (Hyla behavior, growth, abundance, biomass, or ecosystem processes), the magnitude of effects (i.e., negligible, small, arborea), the Hermann’s tortoise (Testudo hermanni), the sand lizard (Lacerta agilis), and the smooth snake medium, or large), the temporal scale of effect for the (Coronella austriaca). attribute (e.g., duration and frequency), and the spatial scales (e.g., in-field and off-field patches of landscapes). If the 2) Each of the selected focal species should, as a priority, be potentially ecologically susceptible to adverse effects ecological entity to protect is the population of a particular caused by exposure to endocrine disruptors (i.e., they species, then EFSA SC (2016a) suggests that in most cases the attribute to be protected will be population dynamics in should be indicator species). “Susceptibility” here does not mean that the species has a proven toxicological terms of abundance (e.g., numbers of individuals and their sensitivity to the substance of interest (most focal species fitness) or biomass. The ecosystem services approach was supported at a stakeholder workshop reported by EFSA will not have been tested in the laboratory, so we would not know). Instead it means that the ecology and (2010) and is now a more widely used concept in EU PPP demographic structure of the population make it vulnera- assessment (e.g., Topping and Luttik 2017). EFSA PPR (2014) makes the point that since the ecosystem ble to the adverse endocrine-mediated effects from the substance of interest. In this sense, the focal species acts services for SPGs for vertebrates derived in EFSA PPR (2010) as a surrogate for all other species in its taxonomic group are performed by populations or groups of populations, because it has life history and other ecological traits at the there needs to be development of appropriate population more susceptible end of the spectrum for that taxon. models for use in risk assessment. This view is supported by Similar thinking led EFSA (2010) to suggest that a species the EU guidance documents on bird and mammal risk is “vulnerable” if there is a combination of high exposure assessment (EFSA 2009) and aquatic risk assessment (EFSA potential, particular life history characteristics, sensitivity, PPR 2013). low dispersal ability, and low reproductive potential (i.e., a Adverse effects low potential for recovery). 3) The role of the selected species as a keystone, umbrella, or Table 1 sorts different types of effect measurements from flagship species is of secondary importance to its ability to mammalian, bird, fish, and amphibian test guidelines and act as an indicator. places them within the framework of the AOP concept (Ankley et al. 2010) and the revised OECD conceptual framework (OECD 2018a). The AOP concept is a robust Protection goals framework in which to organize such information on The PPP regulation (EC 2018) uses the term “(sub) endocrine potential and help support regulatory decision population,” which requires definition before it is possible making (Kramer et al. 2011; Becker et al. 2015; Wheeler and to set protection goals. This definition has usefully been Weltje 2015; Edwards et al. 2016). The concept reflects the provided in the guidance, which states that “The term (sub) definition of an endocrine disruptor: requiring an endocrine population is of predominant relevance with respect to mechanism (i.e., molecular initiating event[s]), causally linked humans, therefore for non-target organisms the term (via key events or key event relationships) to an adverse population is used throughout the document” (ECHA/ outcome (organism or, more relevant here, population EFSA [2018] guidance, footnote p 4). responses). The question that then needs to be addressed Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 282 Integr Environ Assess Manag 15, 2019—M Crane et al. � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Table 1. Toxicological endpoint and AOP classification table for endocrine disruptor effects in vertebrates Adverse outcome pathway Cellular Population Taxa Macromolecular interactions responses Organ responses Organism responses responses Amphibians Cf. OECD TGs Typically in vitro assessment in mammalian-based Vitellogenin Gross necropsy of Growth Potential 231 and 241 systems but considered qualitatively relevant to all endocrine organs Behavior effects on vertebrate taxa Cf. OECD TGs 455, 456, 458, and Histopathology, for Sex ratio population 493, US EPA OPPTS 890.1150 and 1200, and other example, thyroid, Hind-limb length Size relevant sources (e.g., EDSP21 and ToxCast assays) gonads, liver, kidney Developmental stage (abundance Liver Somatic Index Time to metamorphosis or biomass) Stability Birds Cf. OECD TG 206 Gross necropsy of Reproduction (fecundity, fertility) Recruitment endocrine organs Growth Behavior Hatching success Eggshell thickness Number of 14-day survivors Fishes Cf. OECD TGs 229, Vitellogenin Histopathology, for Reproduction (fecundity, fertility) 230, 234, and 240 Hormone levels example, thyroid, Time to maturity and modifications of US gonads, liver, kidney Growth EPA OPPTS 850.1500 Secondary sexual Behavior characteristics Sex ratio Gonadal Somatic Index Embryo time to hatch Hatching success Mammals Hormone levels Reproduction (fecundity, fertility) Steroidogenesis Gross necropsy of Time to maturity (gene or enzyme endocrine organs Growth changes) Histopathology, for Behavior Sperm example thyroid, Sex ratio morphology gonads, liver, kidney Dystocia Sperm motility Secondary sexual Fetal development Sperm numbers characteristics Gestation length Vaginal smears Sexual maturity Litter size landmarks, such as Litter viability –Age at first estrus Litter or pup weight –Estrus cyclicity Number of implantations, corpora lutea –Age at balanopreputial separation –Age at vaginal opening –Nipple development –Anogenital distance Keratinization and cornification of vagina Proliferation of endometrial epithelium (colloid area and follicular cell height) Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 283 to meet the requirements of the PPP regulation is whether measured effects at the organism or individual-animal level (i.e., apical effects measured in in vivo laboratory studies) translate into truly adverse effects at the population level. It is � � only endocrine disruptor–related effects on wildlife popula- tions that are relevant here, and not systemic or other types of toxicity. This focus is because the PPP regulation states that “Adverse effects that are non-specific secondary conse- quences of other toxic effects shall not be considered for the identification of the substance as an endocrine disruptor with respect to non-target organisms.” The concept and definition of an “adverse effect” has received considerable attention in toxicology (Kerlin et al. 2016). For example, Lewis et al. (2002) state that an effect is unlikely to be adverse if: 1) There is no alteration in the general function of the test organism or affected organ or tissue; 2) It is an adaptive response; 3) It is transient; 4) The severity is limited and below thresholds of concern; 5) The effect is isolated or independent; 6) The effect is not a precursor to adverse effects; 7) It is secondary to other adverse effect(s); or 8) It is a consequence of the experimental model. In addition, an effect is unlikely to be adverse if the response lies within historical control ranges (e.g., Valverde- Garcia et al. 2018). For an effect to be considered adverse in an environmental context it is normally assumed to be a measure that is evident at the whole-animal level, whereby the suborganism responses are integrated into an apical effect measure (e.g., growth, development, or reproduction). This integration is not intended to diminish the value of suborganism responses in an understanding of the overall toxicological response, especially in terms of establishing an endocrine mechanism and, potentially, as indicators for adversity. However, it is the organism responses (see Table 1, ‘“organism response” column) that provide the individual adverse response variables that will also serve as input variables for population modeling. This thinking aligns with the currently accepted testing paradigm in which the more definitive eco/toxicological assays sitting at level 5 of the OECD Conceptual Framework (OECD 2018a) are typically used to confirm whether endocrine activity identified at lower levels translates into endocrine-mediated adverse effects. EFSA SC (2013) identifies 3 aspects of a nontarget population that should be addressed when assessing the possible adverse effects of an endocrine disruptor: 1) Population recruitment; 2) Population size; and 3) Population stability. They also state that adverse consequences on reproduc- tion, growth and development, disease incidence, and Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors Data considerations (causal OECD CF level 1 þ 2 OECD CF level 3, OECD CF level 3, 4 þ 5 OECD CF level 4 þ 5 Population links must be established Mechanistic only, not indicative 4 þ 5 Predominately Adverse at the individual level. If responses between responses at of adversity Mechanistic mechanistic, may give an observed and confirmed in OECD CF unpredictable different biological levels) only, not indication for potential level 5 studies, in the absence of additional unless indicative of adversity data, assumed relevant at the population Simulated by adversity level, that is, assumed by default to be population relevant (cf. endocrine disruptor criteria). modeling, or Derived empirically from semifield or field studies 284 Integr Environ Assess Manag 15, 2019—M Crane et al. the stressor event took place) or to the level that is not survival in one or more species should be addressed “as these are the effects most likely to impact on population significantly different from that in control or reference recruitment and stability”, although we note that survival is systems. They note that a system that has been subject to an adaptive response or to recovery might not necessarily not an endpoint of relevance to endocrine disruption and, to the best of our knowledge, disease incidence is rarely return to the same state that it exhibited before the considered explicitly in chemical risk assessment. disturbance. Individual organism recovery from transient effects is a recognized concept under the PPP regulation. The EFSA (2009) guidance document on bird and mammal risk assessment mentions that although the magnitude of an However, “recovery” is unlikely to be an acceptable effect in an exposed group could be statistically significantly regulatory approach to endocrine disruptor effects on vertebrate populations (EFSA PPR 2013; EFSA SC 2016b), different to that in the controls, it might not be biologically relevant, and states: so we do not explore it further in this paper. Instead we use the related concepts of resistance and resilience, which are of considerable value because they are key components of In order to determine the biological relevance of an effect it should be considered whether the effect could population stability and allow discrimination between rele- lead to a functional deficit later on in the study, e.g. if a vant and negligible effects at the population level. EFSA SC (2016b) defines population resistance as the magnitude of reduction in the weight of pups at birth leads to a decrease in level of survival. If not, then the effect may environmental perturbation a population can tolerate with- not be biologically relevant, however if there is a carry- out being pushed out of its normal operating range (i.e., a population’s capacity to remain unaffected). Population over of effects into the number of survivors, it can be considered biologically relevant. resilience is related to the return time to equilibrium following a perturbation (EFSA SC 2016b and references therein). For example, in 2-generation mammalian reproduction Figure 2 illustrates both these concepts. EFSA SC (2016b) studies, a body-weight reduction in pups would not be specifies that population resilience depends on the ecologi- considered relevant if normal development was observed in cal context and is related to the degree to which induced the F1 generation, especially if F1 fertility and reproduction fluctuations in the population density are buffered by density- were comparable to the control (EFSA 2009). Similarly, a dependent feedback mechanisms and competition with reduction in sperm production in rodents with multiple other species (e.g., Knillmann et al. 2012). matings may not adversely affect reproduction. This definition is related to the EFSA SC (2017b) view that The EFSA SC opinion on the hazard assessment of between individual effect reversibility and population recov- endocrine disruptors (EFSA SC 2013) argues that the concept ery, there is also the concept of what they describe as of “biological relevance” is based on the assumption that a “population relevance.” For example, small effects on “normal” biological state can be defined. In turn, “normality” fecundity in density-regulated systems (e.g., a slightly is linked to the adversity of an effect observed during toxicity reduced number of eggs for fish that produce many more testing or in epidemiological studies. The SC states that the eggs than can possibly develop into juvenile fish) will not point at which endocrine modulation becomes an adverse translate adversely to the population level. EFSA SC (2017b) effect cannot be based on an absolute response value but concludes that as, under these circumstances, there is no only on a relative response (compared to the control or effect at the population level, there is no need for recovery background response). (e.g., Hamilton et al. 2014). In other words, the population EFSA SC (2017a) suggests that models can be used for has shown resistance. setting a critical effect level (i.e., a benchmark response Population modeling [BMR]). They envisage that models We propose, in agreement with EFSA SC (2017a), that of focal species could be used to determine endpoints population models should be used to determine what corresponding to cut-off values set by specific protection percentage effect observed in a laboratory toxicity test goals (SPG). These models can be used for calculating exceeds a threshold percentage that translates into a critical effect levels for certain types of effect, for instance nonnegligible reduction in population recruitment, size, or for egg cracking, number of surviving chicks or the size of stability across a representative “worst-case” landscape. We litters, above which the population of the focal species define a negligible effect as one that is buffered by population will be negatively affected to such an extent that the resistance so that population recruitment, size, and stability population will decline over time. remain within their normal operating ranges (Figure 2). These normal operating ranges may be determined from historical EFSA SC (2016b) also suggests that the concept of data, model simulations, or a combination of the 2. “recovery” may be useful when considering biological There are established methods to demonstrate the effects. Recovery is the return of the perturbed (ecological) relevance of effects with population models to extrapolate endpoint (e.g., species composition, population density) to from the laboratory to the field (e.g., Wang 2013; Liu et al the “window” of natural variability as observed in the 2013; Schmitt et al. 2016; Topping and Elmeros 2016; undisturbed state of the ecosystem of concern (e.g., before Topping and Weyman 2018). Mintram et al. (2017) reviewed Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 285 Figure 2. Illustration of population resistance (upper panel) and population resilience (lower panel). The gray area represents the normal operating range of the population parameter. modeling approaches potentially applicable to the environ- chronic (1-year) exposure regimes. Acute exposures to the mental risk assessment of endocrine-active substances in fish test substances had little effect on population-level end- and concluded that individual-based models (IBMs) are a points at any of the concentrations tested. Chronic exposures particularly useful model type because they can account for decreased population abundance at higher concentrations species-specific traits and behaviors (e.g., breeding behav- for both substances and most strongly with DHT, owing to the iors) and simulate interorganism interactions and organism– sex ratio shift to fewer females, which lowered population environment interactions (including responses to chemical reproductive output. However, these concentrations consid- exposure). IBMs therefore capture both the direct and erably exceeded environmentally realistic levels. The model indirect population-level effects of chemical exposures. predicted that a substance-induced change in sex ratio from Mintram et al. (2017) identify the main challenge as striking 50:50 to approximately 40:60 in favor of males (DHT) or a balance between site-specific versus generic applicability, females (4-tOP) would have limited effects on population because of the often complex and environmentally plastic life abundance after chronic exposure but that a large reduction histories of fish. in abundance would occur if the ratio changed to approxi- Hazlerigg et al. (2014) developed an IBM for zebrafish mately 20:80 in favor of either sex. This study suggests that an based on empirical data (e.g., growth, reproduction, and SPG for fish population abundance would be met at a mortality) derived from a combination of laboratory and field benchmark concentration that did not skew sex ratios to more experiments, literature values, and ecological theory. The than approximately 40:60. IBM was validated against size distributions for wild Hamilton et al. (2016) consider further examples for fish in a populations of zebrafish sampled in Bangladesh. Sensitivity critical review of population-level consequences for wild fish analysis showed that population abundance was most exposed to sublethal concentrations of chemicals. They sensitive to changes in density-dependent survival and the conclude that IBMs such as those developed by Hazlerigg availability of refugia for juveniles. The model was then used et al. (2014) include many more of the observed effects of to determine the population-level relevance of changes in chemical exposure on individuals than is the case for matrix sex ratio caused by androgenic (dihydrotestosterone [DHT]) population models and that they are useful for exploring and estrogenic (4-tert-octylphenol [4-tOP]) substances. Both which effects on individuals are likely to have the greatest substances were investigated under acute (10-day) and effect on populations. Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 286 Integr Environ Assess Manag 15, 2019—M Crane et al. individuals may be desirable to control for high natural Uncertainty analysis reported with the results of population modeling helps identify and prioritize the uncertainties variability in biomarker levels, although this approach associated with the assessment inputs and methodology must be balanced against the risk that repeated capture will alter the behavior of the animals and used (EFSA SC 2018). However, the use of additional ecological knowledge via population modeling of focal hence will bias the results. species can only reduce assessment uncertainty from current – Visual observations to monitor populations and activity of birds and large mammals. Interpretation levels. The ECHA/EFSA (2018) guidance is unclear as to how an applicant should report the results of population models or of results is difficult if the animals are not individually lines of evidence other than the results from laboratory toxicity marked. – Monitoring of reproductive performance of birds. test endpoints. This ambiguity is because the reporting template associated with the guidance does not include any endpoint types other than those from laboratory tests. EFSA (2009) guidance suggests that a well-performed However, EFSA PPR (2014) provides advice on good modeling practice specific to ecological models, including a template for field study should carry considerable weight when the model summary documents and a checklist for risk assessors. population relevance of endocrine disruptor effects on individual vertebrates is assessed. Such evidence should These suggestions are clearly applicable when establishing the population relevance of any endocrine disruptor effects via therefore be a powerful line of evidence, which may be modeling and would be suitable for submission with the used either to confirm or refute laboratory evidence of endocrine disruptor effects. This suggestion would be more endocrine disruptor reporting template. Raimondo et al. (2018) also provide useful advice on population models from consistent with the PPP regulation mandate that “Ade- the perspective of regulatory authorities. quate, reliable and representative field or monitoring data and/or results from population models shall as well be Field studies considered where available.” EFSA/ECHA (2018) guidance states that field studies In support of this, EFSA SC (2013) states that the use of field data is valuable in providing greater confidence about cannot be used to override or dismiss evidence of adversity found in laboratory studies. However, this contrasts with whether an adverse endocrine-mediated effect is likely to previous EFSA (2009) guidance, which provides the following have consequences at the population level. Indeed, they go further than this: main points on the use of field studies: Field studies of mortality and reproductive effects are In the absence of such data (i.e., field study data), neither simple nor inexpensive, but they have some regulators must be confident of being able to extrapo- important advantages over other study types: late from laboratory data on endpoints such as growth – They focus on the direct measurement of the effects of and reproduction to potential effects on populations, concern under realistic field conditions, so can take ideally but not necessarily through the use of population account of all routes of exposure and all relevant modeling. sources of variation. – They avoid uncertainties associated with extrapolation And, from models or laboratory studies to the field. – They reduce uncertainties associated with extrapolat- ing sensitivity (toxicity) from studied species to those Field monitoring is probably the most powerful tool in exposed in the field. establishing impacts at the population-level (e.g., Design of a study of appropriate power requires population declines). knowledge of the levels of effects that are considered acceptable and the degree of certainty that is required to We agree with the EFSA SC opinion that, when prevent the acceptable limit being exceeded. available, the use of field studies, including field monitor- “Extensive” approaches across a large number of sites to ing, should form an important line of evidence for cover a broad spectrum of use conditions are preferable assessing the population relevance of an endocrine to “intensive” studies across a small number of sites, disruptor, especially when used in combination with because they account for natural variability in exposure population modeling. Field studies can also provide and effects. invaluable inputs to modeling either for parameterization The choice of methods should be driven by the study or validation, andmodelscan addtofieldstudies by objectives and might include: extrapolating to untested conditions. If adequate, reliable, – Capture-mark-release-recapture studies to monitor and representative population models and field studies population changes, which include changes in age show that there is no evidence for an adverse population- structure, especially in small mammals. level effect from an endocrine disruptor then this should – Monitoring of sublethal effects with biomarkers (e.g., override laboratory data, which is a position consistent enzyme inhibition). Repeated sampling from the same with the PPP regulation. Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 287 ECHA/EFSA (2018) guidance recommends that vertebrates Linking laboratory effects to wildlife populations should be tested at concentrations or doses that do not Results from laboratory tests can currently be linked to exceed an MTD/C so that systemic toxicities that may potential adverse effects at the population level either potentially confound endocrine responses are avoided. quantitatively by use of a population model, or qualitatively Although well established in mammalian toxicology, the by use of expert judgment to bring together different lines of MTD/C concept is relatively new to ecotoxicology (Hutch- evidence (e.g., laboratory and field data) in an overall weight- inson et al. 2009; Wheeler et al. 2013). Test levels may also be of-evidence approach, as described by EFSA SC (2017b). limited by the practical limit of solubility in aquatic test media The population modeling for focal species described because tests should not be conducted above this level earlier, potentially complemented by data from field studies, (OECD 2000). If the MTD/C and water solubility are not would provide a set of simulations that show what percentage limiting, tests should be performed at regulatory limit effect on an input parameter leads to a nonnegligible effect concentrations (e.g., chronic aquatic toxicity at 10 mg/L on population recruitment, size, or stability. These values [OECD 2012] and bird toxicity at 1000 ppm diet [OECD from population models then provide benchmarks against 1984]). Limit test levels for certain taxa are intrinsically linked which an assessor can compare the results for specific to hazard assessment because they are levels specified as laboratory test endpoints such as eggshell cracking in a bird classification categories for hazardous to the environment toxicity test. This is likely to be a doubly conservative (e.g., UN 2011). This approach is illustrated in Figure 3. approach if the focal species chosen for modeling is ecologically sensitive and the laboratory species chosen for CASE STUDY testing over a prolonged chronic duration is toxicologically A study reported recently by Topping and Luttik (2017) sensitive. illustrates the value of the approach described above. They A wide range of potentially endocrine-related endpoints are routinely measured in laboratory tests to support the registra- simulated the effects of PPP exposure on populations of skylark (Alauda arvensis) across Danish agricultural land- tion of PPPs (Day et al. 2018). Marty et al. (2017) summarize the scapes in a study explicitly designed to show how the results main endpoints measured in laboratory toxicity tests designed to assess the effects of endocrine-active substances. They of population modeling might be used to establish SPGs for point out that most of these are “apical” endpoints such as this focal species. An IBM for skylarks was used to examine growth, development, sex ratio, and reproduction, which can several scenarios that represented the range of agricultural potentially be linked to adverse population effects. Increas- landscapes across Denmark. In this section we describe the ingly, as in mammalian toxicology, historical control data are way in which the information in Topping and Luttik (2017) can also available to allow a more thorough understanding of any be used to select focal species and protection goals, define statistically significant differences in these apical endpoints adverse effects, perform population modeling, and link (e.g., Valverde-Garcia et al. 2018). laboratory effects to wildlife populations. ECHA/EFSA (2018) guidance requires that an assessment Focal species for endocrine-disrupting properties is performed first for human health with use of the mammalian toxicology data set. The skylark is a small ground-nesting passerine that The guidance suggests that this requirement is primarily typically feeds on insects and breeds in fields during periods based on the results from tests on rats, such as the extended of PPP application. It is also one of the farmland bird species one-generation reproductive toxicity study (EOGRT; OECD in western Europe that has experienced a large population 2018b, with cohort 1a/1b, including the mating of cohort 1b decline, and it is the subject of an EU management plan to produce the F2 generation) (although this study is not (Topping and Luttik 2017). The skylark therefore fulfills EFSA currently a core data requirement for PPP registration in the (2009) criteria for a focal species because it occurs in the crop EU) or the two-generation reproduction toxicity test (OECD when PPPs are being used and it represents other bird 2001). If the criteria for establishing a substance as an species that may also occur in the crop at that time. endocrine disruptor are not met for human health, then the Protection goals assessment should move on to consider fish and amphibians, which may require further testing (the mammalian data EFSA SC (2016a) guidance states that the following need to package could be used to assess population relevance for be defined before setting a protection goal: the ecological wild mammals, but nothing is stated about this in the entity (e.g., individual, population, functional group, or guidance). The guidance suggests that this testing might ecosystem), the attribute of that entity (e.g., behavior, primarily include the medaka extended one-generation test growth, abundance, biomass, or ecosystem processes), the (MEOGRT; OECD 2015a) and the larval amphibian growth magnitude of effects (i.e., negligible, small, medium, or and development assay (LAGDA; OECD 2015b), although large), the temporal scale of effect for the attribute (e.g., again these are not currently data requirements for PPP duration and frequency), and the spatial scales (e.g., in-field registration in the EU. Any existing data on birds and reptiles and off-field patches of landscapes). If the ecological entity to should also be considered, although the guidance does not protect is the population of a particular species then EFSA SC identify specific tests for these taxa, and regulatory tests do (2016a) suggests that in most cases the attribute to be not currently exist for reptiles. Whichever test is used, the protected will be population dynamics in terms of abundance Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 288 Integr Environ Assess Manag 15, 2019—M Crane et al. Figure 3. The MTD/C concept and upper test level setting that can be applied to endocrine test guideline studies (adapted after Wheeler et al. 2013). Limit test 1 1 1 concentrations are 10 mg  L for fish and aquatic amphibians, 1000 ppm diet for birds (according to OECD TG 206), and 1000 mg  kg body weight  day for mammals (according to OECD TG 416). (e.g., numbers of individuals and their fitness) or biomass. constraints both as a limit to foraging success and as Topping and Luttik (2017) examined the effects of PPP increased energetic costs for cold weather. Initiation of application on total skylark population size and relative breeding depended upon a bird finding a suitable territory abundance occupancy, a measure that compares changes in and vegetation suitable for nesting. Breeding success spatial coverage (occupancy) and density (abundance) with or depended on the habitat being able to fulfil the energetic without PPP application. requirements of the birds during the breeding period and the Although the specific protection goal for effects on survival of eggs and nestlings, which was determined by food population abundance could be set at any level, it is highly resource quantity and availability (a function of management, likely that regulators in the EU would insist that there should weather, and skylark behavior). be negligible long-term adverse effects on abundance across Ten model landscapes were selected to be used in the the most ecologically sensitive landscapes. simulation runs, representing the range of agricultural practices in Denmark, from intensive to extensive. A wide Adverse effects range of different PPP application timing scenarios and levels EFSA SC (2013) identifies population recruitment, size, and of effect on different life stages were simulated. stability as the 3 aspects of a nontarget population that These simulations showed that in the most sensitive of the should be addressed when assessing the possible adverse 10 modeled landscapes a reduction in skylark eggshell effects of an endocrine disruptor. The skylark model thickness, which was associated with a 10% or less probability described by Topping and Luttik (2017) simulates population of clutch loss, led to negligible effects on population size over a 30-year period, so population stability and recruitment, size, and stability after a period of 20-30 years. recruitment are also addressed within the modeling process. Field studies Population modeling The model used by Topping and Luttik (2017) has been Topping and Luttik (2017) used the ALMaSS skylark model, extensively tested and reproduces a range of real-world which is an IBM within the open source Animal Landscape skylark population and individual behaviors. These attributes and Man Simulation System (https://gitlab.com/ include the mean and variation around time to hatch and nest ChrisTopping/ALMaSS_all). Individual birds were catego- leaving, densities of skylarks per farm, and within-season rized into 5 life stages: clutch, nestlings, prefledglings, males, phenology under different field conditions. and females. Available insect food biomass was determined Linking laboratory effects to wildlife populations by vegetation structure in the landscape and by its availability to birds foraging within a home range. Insect biomass was Bird eggs begin to crack in the laboratory when shell updated daily in the model and was affected by vegetation thickness is reduced by 18% (EFSA 2009). Topping and Luttik growth and human management, such as PPP application. (2017) show that a cracking rate of 10% or less did not lead to During the breeding period the model considered the adverse population effects for skylarks in even the most energetic balance of the adults, food requirements for sensitive simulated scenario that they investigated. A shell maintenance, requirements of the young, and weather thinning rate above 18% but below a value corresponding to Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 289 regulatory authorities and to adapt available models for the 10% cracking could therefore be set as a benchmark response. specific questions in the endocrine disruptor hazard assess- The results for eggshell thinning and cracking from ment. It may also be possible to use model outputs to develop look-up tables for adverse population effects, which laboratory-based avian reproduction tests can be directly linked to effects on skylark population recruitment, size, and could standardize and simplify the process, but this would stability, with this benchmark response indicating an inflec- need to be discussed with regulatory authorities. (iii) Definition of MTD/C for all vertebrate test methods so that tion point below which no adverse population effects would be expected. If avian testing is performed at the hazard- agreed hazard thresholds are available to minimize animal based dietary limit of 1000 ppm and the benchmark response testing and reduce the likelihood of observing systemic rather than endocrine disruptor-related toxicity in laboratory is not exceeded, then the tested substance is unlikely to cause adverse population effects in birds. tests. (iv) Perform detailed case studies to pilot the approach. The identification of endocrine disruptors in PPPs under the regulation is primarily a hazard-based and not a risk-based CONCLUSIONS In this paper we have suggested how to implement the new methodology, which is an approach to regulation that has endocrine disruptor criteria in the PPP regulation in a way that been strongly criticized in the past (Crane et al. 2019). The approach to identify adverse endocrine-disrupting popula- builds upon ECHA/EFSA (2018) guidance and is consistent with the PPP regulation legal text that mandates a hazard- tion-level effects that we recommend in this paper is hazard based approach. Our recommended approach is based to comply with the PPP regulation, but uncertainty would be significantly reduced and assessment relevance 1) Use SPGs for focal wildlife populations within EFSA’s improved if exposure was also explicitly considered during ecosystem services framework. The SPGs for vertebrate the process. For example, models are available at the organism level to predict whether an endocrine-disrupting wildlife can be summarized as (i) ecological entity: populations of vertebrate focal species; (ii) attribute: effect would be transient under realistic patterns of exposure population recruitment, size, and stability; (iii) effect (e.g., toxicokinetic and toxicodynamic models and dynamic energy budget models). Use of such models might negate magnitude: negligible; (iv) temporal scale: species life cycle, life span, or breeding period and an exposure the need to move along an AOP to the population level by frequency or duration agreed with regulatory authorities; showing that an adverse endocrine-disrupting effect had not occurred at the organism level. However, use of exposure (v) spatial scale: likely to be a watershed or landscape, but this element needs to be agreed upon with regulators; and duration and profiles information in these models implies a (vi) degree of certainty: high (e.g., 95%). risk-based approach, which is not currently allowed under the PPP regulation. We believe that further discussion of this 2) Model the effects of changes in population-related inputs on focal species populations with IBMs to determine issue between all stakeholders would help to reduce thresholds between negligible population effects and regulatory uncertainty and provide a more robust and science-based solution to the problem of detecting and nonnegligible (i.e., adverse) population effects. 3) Compare these thresholds with the relevant endpoints preventing adverse endocrine disruptor effects in the from laboratory toxicity tests to determine whether they environment. Regardless, all substances shown to be non– endocrine disrupting and potentially endocrine disrupting, are likely to be exceeded at the hazard-based limits or the MTD/C from the experimental studies. but without population-relevant adverse effects, will still 4) If the population threshold is not exceeded, then the undergo risk assessment, thus ensuring a high level of protection for the environment. substance should not be classified as an endocrine disruptor with adverse population effects unless there Acknowledgment—The authors thank Peter Day of the are other lines of evidence within a weight-of-evidence European Crop Protection Association for his support and approach to challenge this. comments and 2 anonymous reviewers for their interesting comments. If the proposals in this paper are accepted by stakeholders, Disclaimer—Mark Crane received funding from the Euro- then it is likely that future technical work will need to pean Crop Protection Association (ECPA) to research and concentrate on 4 main areas: (i) Selection of a small number of write this paper. All other co-authors work for agrochemical ecologically sensitive focal species for each taxonomic companies as indicated by their affiliations. group. This task has already been performed by EFSA for Data Accessibility—Please contact the corresponding birds and mammals (EFSA 2009), and Ibrahim et al. (2013) author, Mark Crane ([email protected]), for any and EFSA PPR (2018) have begun the process for fish, data used in this study. amphibians, and reptiles (although there are currently no laboratory test designs for the latter group). (ii) Development REFERENCES of suitable IBMs for these focal species. Models are already Ankley GT, Bennett RS, Erickson RJ, Hoff DJ, Hornung MW, Johnson RD, available for some bird and mammal focal species and for Mount DR, Nichols JW, Russom CL, Schmieder PK. 2010. Adverse some fish species. However, additional effort will be required outcome pathways: A conceptual framework to support ecotoxicology research and risk assessment. Environ Toxicol Chem 29:730–741. to develop models for additional focal species identified by Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 290 Integr Environ Assess Manag 15, 2019—M Crane et al. [EFSA SC] European Food Safety Authority Scientific Committee. 2017b. Becker RA, Ankley GT, Edwards SW, Kennedy SW, Linkov I, Meek B, Sachana M, Segner H, Van Der Burg B, Villeneuve DL, et al. 2015. Increasing Guidance on the use of the weight of evidence approach in scientific scientific confidence in adverse outcome pathways: Application of tailored assessments. EFSA J 15:4971. [EFSA SC] European Food Safety Authority Scientific Committee. 2018. Bradford-Hill considerations for evaluating weight of evidence. Regul Toxicol Pharmacol 72:514–537. Guidance on uncertainty analysis in scientific assessments. EFSA J Blanchfield PJ, Kidd KA, Docker MF, Palace VP, Park BJ, Postma LD. 2015. 16:5123. Recovery of a wild fish population from whole-lake additions of a synthetic Giesy JP, Feyk LA, Jones PD, Kannan K, Sanderson T. 2003. Review of the estrogen. Environ Sci Technol 49:3136–3144. effects of endocrine-disrupting chemicals in birds. Pure Appl Chem Campbell PM, Hutchinson TH. 1998. Wildlife and endocrine disrupters: 75:2287–2303. Requirements for hazard identification. Environ Toxicol Chem 17: Hamilton PB, Nicol E, De-Bastos ESR, Williams RJ, Sumpter JP, Jobling S, 127–135. Stevens JR, Tyler CR. 2014. Populations of a cyprinid fish are self-sustaining Crane M, Hallmark N, Lagadic L, Ott K, Pickford D, Preuss T, Thompson H, despite widespread feminization of males. BMC Biol 12:1. Thorbek P, Weltje L, Wheeler JR. 2019. Establishing the relevance of Hamilton PB, Cowx IG, Oleksiak MF, Griffiths AM, Grahn M, Stevens JR, endocrine disrupting effects for nontarget vertebrate populations. Integr Carvalho GR, Nicol E, Tyler CR. 2016. Population-level consequences for Environ Assess Manag 15:299–301. wild fish exposed to sublethal concentrations of chemicals—A critical Day P, Green RM, Gross M, Weltje L, Wheeler JR. 2018. Endocrine disruption: review. Fish Fish 17:545–566. Current approaches for regulatory testing and assessment of plant Harris CA, Hamilton PB, Runnalls TJ, Vinciotti V, Henshaw A, Hodgson D, Coe protection products are fit for purpose. Toxicol Lett 296:10–22. TS, Jobling S, Tyler CR, Sumpter JP. 2011. The consequences of [EC] European Commission. 2018. Commission Regulation (EU) 2018/605 of feminization in breeding groups of wild fish. Environ Health Perspect 19 April 2018 amending Annex II to Regulation (EC) No 1107/2009 by 119:306–311. setting out scientific criteria for the determination of endocrine disrupting Hazlerigg CRE, Tyler CR, Lorenzen K, Wheeler JR, Thorbek P. 2014. Population relevance of toxicant mediated changes in sex ratio in fish: An assessment properties. Off J Eur Union L 101:33–36. [ECHA/EFSA] European Chemicals Agency, European Food Safety Authority, using an individual-based zebrafish (Danio rerio) model. Ecol Modell Joint Research Centre. 2018. Guidance for the identification of endocrine 280:76–88. disruptors in the context of Regulations (EU) No 528/2012 and (EC) No Hutchinson TH, Bogi € C, Winter MJ, Owens JW. 2009. Benefits of the maximum 1107/2009. EFSA J 16:5311. tolerated dose (MTD) and maximum tolerated concentration (MTC) Edwards SW, Tan YM, Villeneuve DL, Meek ME, McQueen CA. 2016. Adverse concept in aquatic toxicology. Aquat Toxicol 91:197–202. outcome pathways: Organizing toxicological information to improve Ibrahim L, Preuss TG, Ratte HT, Hommen U. 2013. A list of fish species that are decision making. J Pharmacol Exp Ther 356:170–181. potentially exposed to pesticides in edge-of-field water bodies in the [EFSA] European Food Safety Authority. 2009. Guidance document on risk European Union—A first step towards identifying vulnerable representa- assessment for birds and mammals. EFSA J 7:1438. tives for risk assessment. Environ Sci Pollut Res Int 20:2679–2687. [EFSA] European Food Safety Authority. 2010. Report on the PPR stakeholder Kerlin R, Bolon B, Burkhardt J, Francke S, Greaves P, Meador V, Popp J. 2016. workshop protection goals for environmental risk assessment of pesti- Scientific and Regulatory Policy Committee: Recommended (“best”) cides: What and where to protect? EFSA J 8:1672. practices for determining, communicating, and using adverse effect data [EFSA PPR] European Food Safety Authority Panel on Plant Protection Products from nonclinical studies. Toxicol Pathol 44:147–162. and Their Residues. 2010. Scientific opinion on the development of Kidd KA, Blanchfield PJ, Mills KH, Palace VP, Evans RE, Lazorchak JM, Flick RW. specific protection goal options for environmental risk assessment of 2007. Collapse of a fish population after exposure to a synthetic estrogen. pesticides, in particular in relation to the revision of the Guidance Proc Natl Acad Sci USA 104:8897–8901. Documents on Aquatic and Terrestrial Ecotoxicology (SANCO/3268/2001 Knillmann S, Stampfli NC, Noskov YA, Beketov MA, Liess M. 2012. Interspecific and SANCO/10329/2002). EFSA J 8:1821. competition delays recovery of Daphnia spp. populations from pesticide [EFSA PPR] European Food Safety Authority Panel on Plant Protection Products stress. Ecotoxicology 21:1039–1049. and Their Residues. 2013. Guidance on tiered risk assessment for plant Kramer VJ, Etterson MA, Hecker M, Murphy CA, Roesijadi G, Spade DJ, protection products for aquatic organisms in edge-of-field surface waters. Spromberg JA, Wang M, Ankley GT. 2011. Adverse outcome pathways EFSA J 11:3290. and ecological risk assessment: Bridging to population-level effects. [EFSA PPR] European Food Safety Authority Panel on Plant Protection Products Environ Toxicol Chem 30:64–76. and Their Residues. 2014. Scientific opinion on good modelling practice in Lambeck RJ. 1997. Focal species: A multi-species umbrella for nature the context of mechanistic effect models for risk assessment of plant conservation. Conserv Biol 11:849–856. protection products. EFSA J 12:3589. Lewis RW, Billington R, Debryune E, Gamer A, Lang B, Carpanini F. 2002. [EFSA PPR] European Food Safety Authority Scientific Committee. 2018. Recognition of adverse and nonadverse effects in toxicity studies. Toxicol Scientific opinion on the state of the science on pesticide risk assessment Pathol 30:66–74. for amphibians and reptiles. EFSA J 16:5125. Liu C, Sibly RM, Grimm V, Thorbek P. 2013. Linking pesticide exposure and [EFSA SC] European Food Safety Authority Scientific Committee. 2013. spatial dynamics: An individual-based model of wood mouse (Apodemus Scientific opinion on the hazard assessment of endocrine disruptors: sylvaticus) populations in agricultural landscapes. Ecol Modell 248:92–102. Scientific criteria for identification of endocrine disruptors and appro- Marty MS, Blankinship A, Chambers J, Constantine L, Kloas W, Kumar A, priateness of existing test methods for assessing effects mediated by Lagadic L, Meador J, Pickford D, Schwarz T, et al. 2017. Population- these substances on human health and the environment. EFSA J relevant endpoints in the evaluation of endocrine-active substances (EAS) 11:3132. for ecotoxicological hazard and risk assessment. Integrated Environ Assess [EFSA SC] European Food Safety Authority Scientific Committee. 2016a. Manag 13:317–330. Guidance to develop specific protection goals options for environmental Matthiessen P, Weltje L. 2015. A review of the effects of azole compounds in risk assessment at EFSA, in relation to biodiversity and ecosystem services. fish and their possible involvement in masculinization of wild fish EFSA J 14:4499. populations. Crit Rev Toxicol 45:453–467. [EFSA SC] European Food Safety Authority Scientific Committee. 2016b. Matthiessen P, Wheeler JR, Weltje L. 2018. A review of the evidence for Scientific opinion on recovery in environmental risk assessments at EFSA. endocrine disrupting effects of current-use chemicals on wildlife pop- EFSA J 14:4313. ulations. Crit Rev Toxicol 48:195–216. [EFSA SC] European Food Safety Authority Scientific Committee. 2017a. Mintram KS, Brown AR, Maynard SK, Thorbek P, Tyler CR. 2017. Capturing Scientific opinion on guidance on the assessment of the biological ecology in modeling approaches applied to environmental risk assessment relevance of data in scientific assessments. EFSA J 15:4970. of endocrine active chemicals in fish. Crit Rev Toxicol 48:109–120. Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 291 Munns WR Jr, Barnthouse LW, Sorensen MT. 2007. Issues and recommenda- Raimondo S, Etterson M, Pollesch N, Garber K, Kanarek A, Lehmann W, tions. In: Barnthouse LW, Munns WR Jr, Sorensen MT, editors. Population- Awkerman J. 2018. A framework for linking population model develop- level ecological risk assessment. Boca Raton (FL): CRC Press. p 239–246. ment with ecological risk assessment objectives. Integrated Environ Assess Manag 14:369–380. [OECD] Organization for Economic Cooperation and Development. 1984. Avian reproduction test. Guideline for Testing of Chemicals No 206. Paris Schmitt W, Auteri D, Bastiansen F, Ebeling M, Liu C, Luttik R, Mastitsky S, Nacci (FR): OECD Publishing. D, Topping C, Wang M. 2016. An example of population-level risk [OECD] Organization for Economic Cooperation and Development. 2000. assessments for small mammals using individual-based population Guidance document on aquatic toxicity testing of difficult substances and models. Integr Environ Assess Manag 12:46–57. Topping CJ, Elmeros M. 2016. Modeling exposure of mammalian predators to mixtures. Environmental Health and Safety Publications Series on Testing and Assessment No 23. Paris (FR): OECD Publishing. anticoagulant rodenticides. Front Environ Sci 4:80. [OECD] Organization for Economic Cooperation and Development. 2001. Topping CJ, Luttik R. 2017. Simulation to aid in interpreting biological Two-generation reproduction toxicity study. Guideline for Testing of relevance and setting of population-level protection goals for risk Chemicals No 416. Paris (FR): OECD Publishing. assessment of pesticides. Regul Toxicol Pharmacol 89:40–49. [OECD] Organization for Economic Cooperation and Development. 2012. Fish Topping CJ, Weyman GS. 2018. Rabbit population landscape-scale simulation toxicity testing framework. OECD Environment, Health and Safety to investigate the relevance of using rabbits in regulatory environmental Publications Series on Testing and Assessment No 171. Paris (FR): risk assessment. Environ Model Assess 23:415–457. OECD Publishing. Tyler CR, Jobling S, Sumpter JP. 2008. Endocrine disruption in wildlife: A [OECD] Organization for Economic Cooperation and Development. 2015a. critical review of the evidence. Crit Rev Toxicol 28:319–361. Medaka extended one generation reproduction test (MEOGRT). Guideline [UN] United Nations. 2011. Globally harmonized system of classification for the Testing of Chemicals No 240. Paris (FR): OECD Publishing. and labelling of chemicals (GHS). 4th ed. New York (NY): United [OECD] Organization for Economic Cooperation and Development. 2015b. Nations. 561 p. The larval amphibian growth and development assay (LAGDA). Guideline Valverde-Garcia P, Springer T, Kramer V, Foudoulakis M, Wheeler JR. 2018. An for the Testing of Chemicals No 241. Paris (FR): OECD Publishing. avian reproduction study historical control database: A tool for data [OECD] Organization for Economic Cooperation and Development. 2018a. interpretation. Regul Toxicol Pharmacol 92:295–302. Revised guidance document 150 on standardised test guidelines for Wang M. 2013. From home range dynamics to population cycles: Validation evaluating chemicals for endocrine disruption. OECD Series on Testing and realism of a common vole population model for pesticide risk assessment. Integr Environ Assess Manag 9:294–307. and Assessment. Paris (FR): OECD Publishing. https://doi.org/10.1787/ 9789264304741-en Wheeler JR, Panter GH, Weltje L, Thorpe KL. 2013. Test concentration setting [OECD] Organization for Economic Cooperation and Development. 2018b. for fish in vivo endocrine screening assays. Chemosphere 92:1067–1076. Extended one-generation reproductive toxicity study. Guideline for the Wheeler JR, Weltje L. 2015. Adverse outcome pathways—An industry Testing of Chemicals No 443. Paris (FR): OECD Publishing. perspective. Environ Toxicol Chem 34:1937–1938. [WHO/IPCS] World Health Organization/International Programme on Chemical Ottinger MA, Dean K, McKernan M, Quinn MJ Jr. 2011. Endocrine disruption of reproduction in birds. In: Norris DO, Lopez KH, editors. Hormones and Safety. 2002. Global Assessment of the state-of-the-science of endocrine reproduction of vertebrates. Vol 4. London (UK): Academic Press. p disruptors. Geneva (CH): World Health Organization. WHO/PCS/EDC/02.2. 239–260. 180 p. Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Integrated Environmental Assessment and Management Oxford University Press

Assessing the population relevance of endocrine‐disrupting effects for nontarget vertebrates exposed to plant protection products

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Integrated Environmental Assessment and Management — Volume 15, Number 2—pp. 278–291 278 Received: 20 June 2018 Returned for Revision: 9 October 2018 Accepted: 21 November 2018 | | Environmental Policy & Regulation Assessing the Population Relevance of Endocrine-Disrupting Effects for Nontarget Vertebrates Exposed to Plant Protection Products Mark Crane,*y Nina Hallmark,z Laurent Lagadic,§ Katharina Ott,k Dan Pickford,# Thomas Preuss,§ Helen Thompson,# Pernille Thorbek,#yy Lennart Weltje,k and James R Wheelerzz yAG-HERA, Faringdon, United Kingdom zBayer SAS, Crop Science Division, Regulatory Toxicology, Sophia-Antipolis Cedex, France §Bayer AG, Crop Science Division, Environmental Safety, Monheim am Rhein, Germany kBASF SE, Crop Protection—Ecotoxicology, Limburgerhof, Germany #Syngenta, Jealott's Hill International Research Station, Bracknell, United Kingdom yyPresent address: BASF SE, APD/EE, Limburgerhof, Germany zzCorteva Agriscience, Agriculture Division of DowDuPont, Oxfordshire, United Kingdom ABSTRACT The European Commission intends to protect vertebrate wildlife populations by regulating plant protection product (PPP) active substances that have endocrine-disrupting properties with a hazard-based approach. In this paper we consider how the Commission’s hazard-based regulation and accompanying guidance can be operationalized to ensure that a technically robust process is used to distinguish between substances with adverse population-level effects and those for which it can be demonstrated that adverse effects observed (typically in the laboratory) do not translate into adverse effects at the population level. Our approach is to use population models within the adverse outcome pathway framework to link the nonlinear relationship between adverse effects at the individual and population levels in the following way: (1) use specific protection goals for focal wildlife populations within an ecosystem services framework; (2) model the effects of changes in population- related inputs on focal species populations with individual-based population models to determine thresholds between negligible and nonnegligible (i.e., adverse) population-level effects; (3) compare these thresholds with the relevant endpoints from laboratory toxicity tests to determine whether they are likely to be exceeded at hazard-based limits or the maximum tolerated dose/concentration from the experimental studies. If the population threshold is not exceeded, then the substance should not be classified as an endocrine disruptor with population-relevant adversity unless there are other lines of evidence within a weight-of-evidence approach to challenge this. We believe this approach is scientifically robust and still addresses the political and legal requirement for a hazard-based assessment. Integr Environ Assess Manag 2019;15:278–291. C 2018 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals, Inc. on behalf of Society of Environmental Toxicology & Chemistry (SETAC) Keywords: Endocrine disruption Plant protection product Vertebrate population European Union Hazard assessment INTRODUCTION populations of organisms (Munns et al. 2007); thus, The potential effects of endocrine-disrupting chemicals extrapolation from laboratory to field and from individual (EDCs) on wildlife have been a concern for many years organism to population is required. (Campbell and Hutchinson 1998; Kidd et al. 2007; Tyler et al. Some studies have demonstrated a strong link between 2008). Laboratory-based studies have shown that certain adverse endocrine effects found in the laboratory and chemicals can disrupt specific components of vertebrate adverse effects found in natural populations. For example, endocrine systems in individual organisms, but the regulatory Giesy et al. (2003) and Ottinger et al. (2011) review the effects goal for environmental risk assessment is usually to protect of endocrine disruptors in birds, including the effects of now- banned organochlorine substances on gull sex ratio and clutch “superabundance,” for which there is clear laboratory * Address correspondence to [email protected] and field-based evidence. However, other studies have Published 6 December 2018 on wileyonlinelibrary.com/journal/ieam. shown that many natural populations can compensate for This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in endocrine-mediated effects observed in the laboratory and any medium, provided the original work is properly cited. thereby maintain population numbers and biomass. For Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 279 example, a field study by Hamilton et al. (2014) supports the endocrine disruptor for regulatory purposes (WHO/IPCS conclusion that intersex and other signs of feminization in 2002; EC 2018). If this is not done, then useful products are individual fish observed in both the laboratory and the field likely to be banned on the basis of effects observed under do not necessarily result in adverse population-level effects. laboratory conditions that are unlikely to translate to the They sampled roach (Rutilus rutilus) from different southern population level in nature. It is worth noting that environ- English rivers and assessed the genetic structure of the mental risk assessments have been performed for decades to populations at each location. Despite widespread feminiza- help prevent adverse effects on vertebrate populations, tion of male roach in effluent-contaminated rivers, there was which, as shown by Matthiessen et al. (2018), may explain why no evidence of a correlation between effective population the evidence for endocrine disruption due to current-use size and predicted exposure to estrogens. In another study chemicals in wildlife populations is quite limited. with fish, Harris et al. (2011) examined the breeding ability of In an accompanying Learned Discourses paper (Crane intersex male roach when in competition with other males for et al. this issue), we discuss recent European Union (EU) regulation (EC 2018) and guidance (ECHA/EFSA 2018) females. They found that most intersex males were able to participate in spawning, and the reproductive success of intended to protect humans and vertebrate wildlife mildly intersex males was similar to nonintersex males, populations from adverse effects due to plant protection products (PPPs) containing an EDC. In this paper we although moderately and severely intersex males were less successful. describe possible approaches that complement this guid- Matthiessen and Weltje (2015) reviewed the effects of ance and would assist registrants and regulatory authorities in distinguishing between (i) substances with adverse azole compounds and their possible involvement in mascu- linization of wild fish populations to determine whether there endocrine-mediated population effects and (ii) substances is a biologically causal link between exposure to azoles and for which a potential endocrine disruption issue could be concluded from laboratory toxicity tests, but which would endocrine-disrupting effects in wild fish populations. They concluded that the available data on exposure and effects not translate into adverse effects on wildlife populations. provide reassurance that reported environmental concen- This activity is in response to, and complies with, the hazard-based approach that has been politically and legally trations of certain azoles are insufficient to cause adverse effects in fish by interference with their endocrine system, mandated by the European Commission (EC 2018). despite results from laboratory studies that show masculini- zation in fish under continuous exposure at significantly ASSESSMENT OF ADVERSE POPULATION EFFECTS higher concentrations. OF ENDOCRINE DISRUPTERS Recovery from endocrine-disrupting effects may also be The European Food Safety Authority (EFSA) benefits from reasonably rapid if the source of the endocrine-disrupting many expert opinions provided by their Scientific Committee substance is removed. For example, Kidd et al. (2007) (EFSA SC) and Panel on Plant Protection Products and their conducted a 7-year, whole-lake experiment at the Experi- Residues (EFSA PPR). We draw upon these opinions here to mental Lakes Area in Canada and showed that chronic assist in mapping out a coherent and conservative hazard- exposure of fathead minnow (Pimephales promelas) to low based approach (Figure 1) for demonstrating the population concentrations (5–6 ng  L ) of the potent estrogen 17a- relevance of endocrine disruptor effects observed in the ethynylestradiol (EE2) led to feminization and intersex in laboratory. males and altered oogenesis in females, leading to near The approach depends on clear identification of specific extinction of the population in the lake. However, in a follow- protection goals for focal wildlife populations within the up study, Blanchfield et al. (2015) quantified the physiologi- EFSA’s ecosystem services framework. The magnitude of cal, population, and genetic characteristics of the fathead relevant effects in toxicity tests performed at hazard-based minnow population for 7 years after EE2 additions ceased. thresholds (i.e., regulatory-defined limits or a maximum They found that 3 years after treatment, whole-body tolerated dose/concentration [MTD/C]) is compared to vitellogenin concentrations in male fathead minnow had population-relevant thresholds derived from population returned to baseline levels, there were no testicular modeling. The relevance of specific endpoints is taken abnormalities, and in the fourth year, adult size-frequency from an understanding of the adverse outcome pathway distribution and abundance had returned to pretreatment (AOP) concept (Ankley et al. 2010) focusing on adverse levels. effects caused by an endocrine disruptor at the individual Wild vertebrate populations may therefore be either level that may translate to the population level. If the resistant or resilient (for definitions, see Adverse Effects population threshold is not exceeded, then the substance section) to the effects of endocrine-disrupting substances, should not be classified as an endocrine disruptor with even when adverse effects of these substances are observed adverse population effects unless there are other lines of in laboratory tests. It is therefore important to establish a evidence within a weight-of-evidence approach to challenge biologically causal link between endocrine-disrupting effects this. This approach is analogous to that for human health in measured in laboratory toxicity tests and adverse population- which the relevance of animal model effects to humans can level effects on vertebrate wildlife in the field, because this is be challenged in the endocrine disruptor properties necessary to meet the requirements of the definition of an evaluation. Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 280 Integr Environ Assess Manag 15, 2019—M Crane et al. Figure 1. Evaluation scheme to determine the population relevance of laboratory-determined adverse endocrine-disrupting effects. Focal species EFSA (2009) distinguishes between “generic focal species” and “focal species” when assessing PPP risks to birds and It is not possible, desirable, or legally permissible to test mammals and defines them as follows: all vertebrate species in the laboratory, nor is it possible to perform risk assessments for every vertebrate species in an ecosystem. Therefore, the concept of “focal species” has A generic focal species is not a real species, however it is been adopted from conservation biology (Lambeck 1997) considered to be representative of all those species to focus assessments on those species of most relevance to potentially at risk, that is it is based on ecological perceived threats. For example, in the case of PPPs these knowledge of a range of species that could be at risk. It will be species that are present in specific agricultural has a high food intake rate and may consume a mixed landscapes and are thus most likely to be exposed to a diet rather than just one as for the indicator species. The PPP. Focal species may be keystone species (organisms diet is not real but is considered to be representative of with an effect on the ecosystem that is disproportionate to the species represented .... The “generic focal species” their numerical abundance), umbrella species (organisms is also considered to be a representative of the types of that cover a large geographical area in their daily or birds or mammals that occur across Member States. seasonal movements), flagship species (charismatic organ- isms with public appeal), or indicator species (organisms A focal species is a real species that actually occurs in the sensitive to change and therefore useful in monitoring crop when the PPP is being used. The aim of using a habitat quality). Focal species may fulfill more than one of “focal species” is to add realism to the risk assessment these criteria (e.g., they may be both an indicator and a insofar as the assessment is based on a real species that flagship species). uses the crop. It is essential that the species actually The concept of focal species has been adopted most fully occurs in the crop at a time when the PPP is being for birds and mammals in PPP assessment, although the applied. It is also essential that this species is considered concept has wider application for other vertebrate taxa and to be representative of all other species that may occur in has also been explored for fish (Ibrahim et al. 2013), the crop at that time. As a “focal species” needs to cover amphibians, and reptiles (EFSA PPR 2018). Mintram et al. all species present in the crop, it is possible that there (2017), in a review of modeling approaches for risk may be more than one “focal species” per crop. assessment of endocrine disruptors in fish, recommend that “the development of future models should include Focal bird and mammal species are identified for specific species representing a range of life-histories and.. .their exposure scenarios (i.e., a PPP application to a particular selection should be guided by the derivation of ecological crop type at a specific growth stage), ideally by use of scenarios which are relevant to major land use and water transect or field survey methods across a range of body types in which chemical exposures and effects are representative fields, although literature and census data predicted according to current risk assessments.” might also be useful (EFSA 2009). The survey data are then Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 281 We therefore also use the term “population” subsequently used to select focal species by considering their feeding strata, food intake rate, body weight, and diet to ensure in this paper as an operational description of any group of that species with the highest potential exposures are individuals from the same species that occurs in a particular space during a particular time, as defined by a regulatory considered. EFSA (2009) notes that a focal species is not automatically the species that was most frequently seen but authority. that it should represent the feeding guild(s) that raised Nontarget vertebrate wildlife populations provide a wide variety of ecosystem services including food (for human concern at earlier stages in the risk assessment and represent other species. consumption), genetic resources (biodiversity), education Selection of focal species when assessing the potential and inspiration, aesthetic values, pest and disease regulation (e.g., birds feeding on caterpillars), seed and propagule population relevance of endocrine disruptors should there- fore include the following considerations: dispersal, and recreation and ecotourism (e.g., bird-watch- ing, hunting, and fishing) (EFSA 2010). The EFSA SC (2016a) uses the concept of ecosystem services to derive specific 1) At least one relevant (generic) focal species should be selected for each regulatory exposure scenario (e.g., crop protection goals (SPGs) for service-providing units (SPUs). An and application scenario). Examples of generic focal bird SPU can be any ecological entity that provides an ecosystem service (provisioning, regulating, cultural, or supporting and mammal species are provided in EFSA (2009), and Ibrahim et al. (2013) have derived a list of focal fish species. services) to humans. EFSA SC (2016a) states that the An EFSA PPR (2018) opinion suggests that the following following need to be defined before setting an SPG: the ecological entity (e.g., individual, population, functional amphibian and reptile focal species should be considered: the great crested newt (Triturus cristatus), the natterjack group, or ecosystem), the attribute of that entity (e.g., toad (Epidalea calamita), the common treefrog (Hyla behavior, growth, abundance, biomass, or ecosystem processes), the magnitude of effects (i.e., negligible, small, arborea), the Hermann’s tortoise (Testudo hermanni), the sand lizard (Lacerta agilis), and the smooth snake medium, or large), the temporal scale of effect for the (Coronella austriaca). attribute (e.g., duration and frequency), and the spatial scales (e.g., in-field and off-field patches of landscapes). If the 2) Each of the selected focal species should, as a priority, be potentially ecologically susceptible to adverse effects ecological entity to protect is the population of a particular caused by exposure to endocrine disruptors (i.e., they species, then EFSA SC (2016a) suggests that in most cases the attribute to be protected will be population dynamics in should be indicator species). “Susceptibility” here does not mean that the species has a proven toxicological terms of abundance (e.g., numbers of individuals and their sensitivity to the substance of interest (most focal species fitness) or biomass. The ecosystem services approach was supported at a stakeholder workshop reported by EFSA will not have been tested in the laboratory, so we would not know). Instead it means that the ecology and (2010) and is now a more widely used concept in EU PPP demographic structure of the population make it vulnera- assessment (e.g., Topping and Luttik 2017). EFSA PPR (2014) makes the point that since the ecosystem ble to the adverse endocrine-mediated effects from the substance of interest. In this sense, the focal species acts services for SPGs for vertebrates derived in EFSA PPR (2010) as a surrogate for all other species in its taxonomic group are performed by populations or groups of populations, because it has life history and other ecological traits at the there needs to be development of appropriate population more susceptible end of the spectrum for that taxon. models for use in risk assessment. This view is supported by Similar thinking led EFSA (2010) to suggest that a species the EU guidance documents on bird and mammal risk is “vulnerable” if there is a combination of high exposure assessment (EFSA 2009) and aquatic risk assessment (EFSA potential, particular life history characteristics, sensitivity, PPR 2013). low dispersal ability, and low reproductive potential (i.e., a Adverse effects low potential for recovery). 3) The role of the selected species as a keystone, umbrella, or Table 1 sorts different types of effect measurements from flagship species is of secondary importance to its ability to mammalian, bird, fish, and amphibian test guidelines and act as an indicator. places them within the framework of the AOP concept (Ankley et al. 2010) and the revised OECD conceptual framework (OECD 2018a). The AOP concept is a robust Protection goals framework in which to organize such information on The PPP regulation (EC 2018) uses the term “(sub) endocrine potential and help support regulatory decision population,” which requires definition before it is possible making (Kramer et al. 2011; Becker et al. 2015; Wheeler and to set protection goals. This definition has usefully been Weltje 2015; Edwards et al. 2016). The concept reflects the provided in the guidance, which states that “The term (sub) definition of an endocrine disruptor: requiring an endocrine population is of predominant relevance with respect to mechanism (i.e., molecular initiating event[s]), causally linked humans, therefore for non-target organisms the term (via key events or key event relationships) to an adverse population is used throughout the document” (ECHA/ outcome (organism or, more relevant here, population EFSA [2018] guidance, footnote p 4). responses). The question that then needs to be addressed Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 282 Integr Environ Assess Manag 15, 2019—M Crane et al. � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � � Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Table 1. Toxicological endpoint and AOP classification table for endocrine disruptor effects in vertebrates Adverse outcome pathway Cellular Population Taxa Macromolecular interactions responses Organ responses Organism responses responses Amphibians Cf. OECD TGs Typically in vitro assessment in mammalian-based Vitellogenin Gross necropsy of Growth Potential 231 and 241 systems but considered qualitatively relevant to all endocrine organs Behavior effects on vertebrate taxa Cf. OECD TGs 455, 456, 458, and Histopathology, for Sex ratio population 493, US EPA OPPTS 890.1150 and 1200, and other example, thyroid, Hind-limb length Size relevant sources (e.g., EDSP21 and ToxCast assays) gonads, liver, kidney Developmental stage (abundance Liver Somatic Index Time to metamorphosis or biomass) Stability Birds Cf. OECD TG 206 Gross necropsy of Reproduction (fecundity, fertility) Recruitment endocrine organs Growth Behavior Hatching success Eggshell thickness Number of 14-day survivors Fishes Cf. OECD TGs 229, Vitellogenin Histopathology, for Reproduction (fecundity, fertility) 230, 234, and 240 Hormone levels example, thyroid, Time to maturity and modifications of US gonads, liver, kidney Growth EPA OPPTS 850.1500 Secondary sexual Behavior characteristics Sex ratio Gonadal Somatic Index Embryo time to hatch Hatching success Mammals Hormone levels Reproduction (fecundity, fertility) Steroidogenesis Gross necropsy of Time to maturity (gene or enzyme endocrine organs Growth changes) Histopathology, for Behavior Sperm example thyroid, Sex ratio morphology gonads, liver, kidney Dystocia Sperm motility Secondary sexual Fetal development Sperm numbers characteristics Gestation length Vaginal smears Sexual maturity Litter size landmarks, such as Litter viability –Age at first estrus Litter or pup weight –Estrus cyclicity Number of implantations, corpora lutea –Age at balanopreputial separation –Age at vaginal opening –Nipple development –Anogenital distance Keratinization and cornification of vagina Proliferation of endometrial epithelium (colloid area and follicular cell height) Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 283 to meet the requirements of the PPP regulation is whether measured effects at the organism or individual-animal level (i.e., apical effects measured in in vivo laboratory studies) translate into truly adverse effects at the population level. It is � � only endocrine disruptor–related effects on wildlife popula- tions that are relevant here, and not systemic or other types of toxicity. This focus is because the PPP regulation states that “Adverse effects that are non-specific secondary conse- quences of other toxic effects shall not be considered for the identification of the substance as an endocrine disruptor with respect to non-target organisms.” The concept and definition of an “adverse effect” has received considerable attention in toxicology (Kerlin et al. 2016). For example, Lewis et al. (2002) state that an effect is unlikely to be adverse if: 1) There is no alteration in the general function of the test organism or affected organ or tissue; 2) It is an adaptive response; 3) It is transient; 4) The severity is limited and below thresholds of concern; 5) The effect is isolated or independent; 6) The effect is not a precursor to adverse effects; 7) It is secondary to other adverse effect(s); or 8) It is a consequence of the experimental model. In addition, an effect is unlikely to be adverse if the response lies within historical control ranges (e.g., Valverde- Garcia et al. 2018). For an effect to be considered adverse in an environmental context it is normally assumed to be a measure that is evident at the whole-animal level, whereby the suborganism responses are integrated into an apical effect measure (e.g., growth, development, or reproduction). This integration is not intended to diminish the value of suborganism responses in an understanding of the overall toxicological response, especially in terms of establishing an endocrine mechanism and, potentially, as indicators for adversity. However, it is the organism responses (see Table 1, ‘“organism response” column) that provide the individual adverse response variables that will also serve as input variables for population modeling. This thinking aligns with the currently accepted testing paradigm in which the more definitive eco/toxicological assays sitting at level 5 of the OECD Conceptual Framework (OECD 2018a) are typically used to confirm whether endocrine activity identified at lower levels translates into endocrine-mediated adverse effects. EFSA SC (2013) identifies 3 aspects of a nontarget population that should be addressed when assessing the possible adverse effects of an endocrine disruptor: 1) Population recruitment; 2) Population size; and 3) Population stability. They also state that adverse consequences on reproduc- tion, growth and development, disease incidence, and Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors Data considerations (causal OECD CF level 1 þ 2 OECD CF level 3, OECD CF level 3, 4 þ 5 OECD CF level 4 þ 5 Population links must be established Mechanistic only, not indicative 4 þ 5 Predominately Adverse at the individual level. If responses between responses at of adversity Mechanistic mechanistic, may give an observed and confirmed in OECD CF unpredictable different biological levels) only, not indication for potential level 5 studies, in the absence of additional unless indicative of adversity data, assumed relevant at the population Simulated by adversity level, that is, assumed by default to be population relevant (cf. endocrine disruptor criteria). modeling, or Derived empirically from semifield or field studies 284 Integr Environ Assess Manag 15, 2019—M Crane et al. the stressor event took place) or to the level that is not survival in one or more species should be addressed “as these are the effects most likely to impact on population significantly different from that in control or reference recruitment and stability”, although we note that survival is systems. They note that a system that has been subject to an adaptive response or to recovery might not necessarily not an endpoint of relevance to endocrine disruption and, to the best of our knowledge, disease incidence is rarely return to the same state that it exhibited before the considered explicitly in chemical risk assessment. disturbance. Individual organism recovery from transient effects is a recognized concept under the PPP regulation. The EFSA (2009) guidance document on bird and mammal risk assessment mentions that although the magnitude of an However, “recovery” is unlikely to be an acceptable effect in an exposed group could be statistically significantly regulatory approach to endocrine disruptor effects on vertebrate populations (EFSA PPR 2013; EFSA SC 2016b), different to that in the controls, it might not be biologically relevant, and states: so we do not explore it further in this paper. Instead we use the related concepts of resistance and resilience, which are of considerable value because they are key components of In order to determine the biological relevance of an effect it should be considered whether the effect could population stability and allow discrimination between rele- lead to a functional deficit later on in the study, e.g. if a vant and negligible effects at the population level. EFSA SC (2016b) defines population resistance as the magnitude of reduction in the weight of pups at birth leads to a decrease in level of survival. If not, then the effect may environmental perturbation a population can tolerate with- not be biologically relevant, however if there is a carry- out being pushed out of its normal operating range (i.e., a population’s capacity to remain unaffected). Population over of effects into the number of survivors, it can be considered biologically relevant. resilience is related to the return time to equilibrium following a perturbation (EFSA SC 2016b and references therein). For example, in 2-generation mammalian reproduction Figure 2 illustrates both these concepts. EFSA SC (2016b) studies, a body-weight reduction in pups would not be specifies that population resilience depends on the ecologi- considered relevant if normal development was observed in cal context and is related to the degree to which induced the F1 generation, especially if F1 fertility and reproduction fluctuations in the population density are buffered by density- were comparable to the control (EFSA 2009). Similarly, a dependent feedback mechanisms and competition with reduction in sperm production in rodents with multiple other species (e.g., Knillmann et al. 2012). matings may not adversely affect reproduction. This definition is related to the EFSA SC (2017b) view that The EFSA SC opinion on the hazard assessment of between individual effect reversibility and population recov- endocrine disruptors (EFSA SC 2013) argues that the concept ery, there is also the concept of what they describe as of “biological relevance” is based on the assumption that a “population relevance.” For example, small effects on “normal” biological state can be defined. In turn, “normality” fecundity in density-regulated systems (e.g., a slightly is linked to the adversity of an effect observed during toxicity reduced number of eggs for fish that produce many more testing or in epidemiological studies. The SC states that the eggs than can possibly develop into juvenile fish) will not point at which endocrine modulation becomes an adverse translate adversely to the population level. EFSA SC (2017b) effect cannot be based on an absolute response value but concludes that as, under these circumstances, there is no only on a relative response (compared to the control or effect at the population level, there is no need for recovery background response). (e.g., Hamilton et al. 2014). In other words, the population EFSA SC (2017a) suggests that models can be used for has shown resistance. setting a critical effect level (i.e., a benchmark response Population modeling [BMR]). They envisage that models We propose, in agreement with EFSA SC (2017a), that of focal species could be used to determine endpoints population models should be used to determine what corresponding to cut-off values set by specific protection percentage effect observed in a laboratory toxicity test goals (SPG). These models can be used for calculating exceeds a threshold percentage that translates into a critical effect levels for certain types of effect, for instance nonnegligible reduction in population recruitment, size, or for egg cracking, number of surviving chicks or the size of stability across a representative “worst-case” landscape. We litters, above which the population of the focal species define a negligible effect as one that is buffered by population will be negatively affected to such an extent that the resistance so that population recruitment, size, and stability population will decline over time. remain within their normal operating ranges (Figure 2). These normal operating ranges may be determined from historical EFSA SC (2016b) also suggests that the concept of data, model simulations, or a combination of the 2. “recovery” may be useful when considering biological There are established methods to demonstrate the effects. Recovery is the return of the perturbed (ecological) relevance of effects with population models to extrapolate endpoint (e.g., species composition, population density) to from the laboratory to the field (e.g., Wang 2013; Liu et al the “window” of natural variability as observed in the 2013; Schmitt et al. 2016; Topping and Elmeros 2016; undisturbed state of the ecosystem of concern (e.g., before Topping and Weyman 2018). Mintram et al. (2017) reviewed Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 285 Figure 2. Illustration of population resistance (upper panel) and population resilience (lower panel). The gray area represents the normal operating range of the population parameter. modeling approaches potentially applicable to the environ- chronic (1-year) exposure regimes. Acute exposures to the mental risk assessment of endocrine-active substances in fish test substances had little effect on population-level end- and concluded that individual-based models (IBMs) are a points at any of the concentrations tested. Chronic exposures particularly useful model type because they can account for decreased population abundance at higher concentrations species-specific traits and behaviors (e.g., breeding behav- for both substances and most strongly with DHT, owing to the iors) and simulate interorganism interactions and organism– sex ratio shift to fewer females, which lowered population environment interactions (including responses to chemical reproductive output. However, these concentrations consid- exposure). IBMs therefore capture both the direct and erably exceeded environmentally realistic levels. The model indirect population-level effects of chemical exposures. predicted that a substance-induced change in sex ratio from Mintram et al. (2017) identify the main challenge as striking 50:50 to approximately 40:60 in favor of males (DHT) or a balance between site-specific versus generic applicability, females (4-tOP) would have limited effects on population because of the often complex and environmentally plastic life abundance after chronic exposure but that a large reduction histories of fish. in abundance would occur if the ratio changed to approxi- Hazlerigg et al. (2014) developed an IBM for zebrafish mately 20:80 in favor of either sex. This study suggests that an based on empirical data (e.g., growth, reproduction, and SPG for fish population abundance would be met at a mortality) derived from a combination of laboratory and field benchmark concentration that did not skew sex ratios to more experiments, literature values, and ecological theory. The than approximately 40:60. IBM was validated against size distributions for wild Hamilton et al. (2016) consider further examples for fish in a populations of zebrafish sampled in Bangladesh. Sensitivity critical review of population-level consequences for wild fish analysis showed that population abundance was most exposed to sublethal concentrations of chemicals. They sensitive to changes in density-dependent survival and the conclude that IBMs such as those developed by Hazlerigg availability of refugia for juveniles. The model was then used et al. (2014) include many more of the observed effects of to determine the population-level relevance of changes in chemical exposure on individuals than is the case for matrix sex ratio caused by androgenic (dihydrotestosterone [DHT]) population models and that they are useful for exploring and estrogenic (4-tert-octylphenol [4-tOP]) substances. Both which effects on individuals are likely to have the greatest substances were investigated under acute (10-day) and effect on populations. Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 286 Integr Environ Assess Manag 15, 2019—M Crane et al. individuals may be desirable to control for high natural Uncertainty analysis reported with the results of population modeling helps identify and prioritize the uncertainties variability in biomarker levels, although this approach associated with the assessment inputs and methodology must be balanced against the risk that repeated capture will alter the behavior of the animals and used (EFSA SC 2018). However, the use of additional ecological knowledge via population modeling of focal hence will bias the results. species can only reduce assessment uncertainty from current – Visual observations to monitor populations and activity of birds and large mammals. Interpretation levels. The ECHA/EFSA (2018) guidance is unclear as to how an applicant should report the results of population models or of results is difficult if the animals are not individually lines of evidence other than the results from laboratory toxicity marked. – Monitoring of reproductive performance of birds. test endpoints. This ambiguity is because the reporting template associated with the guidance does not include any endpoint types other than those from laboratory tests. EFSA (2009) guidance suggests that a well-performed However, EFSA PPR (2014) provides advice on good modeling practice specific to ecological models, including a template for field study should carry considerable weight when the model summary documents and a checklist for risk assessors. population relevance of endocrine disruptor effects on individual vertebrates is assessed. Such evidence should These suggestions are clearly applicable when establishing the population relevance of any endocrine disruptor effects via therefore be a powerful line of evidence, which may be modeling and would be suitable for submission with the used either to confirm or refute laboratory evidence of endocrine disruptor effects. This suggestion would be more endocrine disruptor reporting template. Raimondo et al. (2018) also provide useful advice on population models from consistent with the PPP regulation mandate that “Ade- the perspective of regulatory authorities. quate, reliable and representative field or monitoring data and/or results from population models shall as well be Field studies considered where available.” EFSA/ECHA (2018) guidance states that field studies In support of this, EFSA SC (2013) states that the use of field data is valuable in providing greater confidence about cannot be used to override or dismiss evidence of adversity found in laboratory studies. However, this contrasts with whether an adverse endocrine-mediated effect is likely to previous EFSA (2009) guidance, which provides the following have consequences at the population level. Indeed, they go further than this: main points on the use of field studies: Field studies of mortality and reproductive effects are In the absence of such data (i.e., field study data), neither simple nor inexpensive, but they have some regulators must be confident of being able to extrapo- important advantages over other study types: late from laboratory data on endpoints such as growth – They focus on the direct measurement of the effects of and reproduction to potential effects on populations, concern under realistic field conditions, so can take ideally but not necessarily through the use of population account of all routes of exposure and all relevant modeling. sources of variation. – They avoid uncertainties associated with extrapolation And, from models or laboratory studies to the field. – They reduce uncertainties associated with extrapolat- ing sensitivity (toxicity) from studied species to those Field monitoring is probably the most powerful tool in exposed in the field. establishing impacts at the population-level (e.g., Design of a study of appropriate power requires population declines). knowledge of the levels of effects that are considered acceptable and the degree of certainty that is required to We agree with the EFSA SC opinion that, when prevent the acceptable limit being exceeded. available, the use of field studies, including field monitor- “Extensive” approaches across a large number of sites to ing, should form an important line of evidence for cover a broad spectrum of use conditions are preferable assessing the population relevance of an endocrine to “intensive” studies across a small number of sites, disruptor, especially when used in combination with because they account for natural variability in exposure population modeling. Field studies can also provide and effects. invaluable inputs to modeling either for parameterization The choice of methods should be driven by the study or validation, andmodelscan addtofieldstudies by objectives and might include: extrapolating to untested conditions. If adequate, reliable, – Capture-mark-release-recapture studies to monitor and representative population models and field studies population changes, which include changes in age show that there is no evidence for an adverse population- structure, especially in small mammals. level effect from an endocrine disruptor then this should – Monitoring of sublethal effects with biomarkers (e.g., override laboratory data, which is a position consistent enzyme inhibition). Repeated sampling from the same with the PPP regulation. Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 287 ECHA/EFSA (2018) guidance recommends that vertebrates Linking laboratory effects to wildlife populations should be tested at concentrations or doses that do not Results from laboratory tests can currently be linked to exceed an MTD/C so that systemic toxicities that may potential adverse effects at the population level either potentially confound endocrine responses are avoided. quantitatively by use of a population model, or qualitatively Although well established in mammalian toxicology, the by use of expert judgment to bring together different lines of MTD/C concept is relatively new to ecotoxicology (Hutch- evidence (e.g., laboratory and field data) in an overall weight- inson et al. 2009; Wheeler et al. 2013). Test levels may also be of-evidence approach, as described by EFSA SC (2017b). limited by the practical limit of solubility in aquatic test media The population modeling for focal species described because tests should not be conducted above this level earlier, potentially complemented by data from field studies, (OECD 2000). If the MTD/C and water solubility are not would provide a set of simulations that show what percentage limiting, tests should be performed at regulatory limit effect on an input parameter leads to a nonnegligible effect concentrations (e.g., chronic aquatic toxicity at 10 mg/L on population recruitment, size, or stability. These values [OECD 2012] and bird toxicity at 1000 ppm diet [OECD from population models then provide benchmarks against 1984]). Limit test levels for certain taxa are intrinsically linked which an assessor can compare the results for specific to hazard assessment because they are levels specified as laboratory test endpoints such as eggshell cracking in a bird classification categories for hazardous to the environment toxicity test. This is likely to be a doubly conservative (e.g., UN 2011). This approach is illustrated in Figure 3. approach if the focal species chosen for modeling is ecologically sensitive and the laboratory species chosen for CASE STUDY testing over a prolonged chronic duration is toxicologically A study reported recently by Topping and Luttik (2017) sensitive. illustrates the value of the approach described above. They A wide range of potentially endocrine-related endpoints are routinely measured in laboratory tests to support the registra- simulated the effects of PPP exposure on populations of skylark (Alauda arvensis) across Danish agricultural land- tion of PPPs (Day et al. 2018). Marty et al. (2017) summarize the scapes in a study explicitly designed to show how the results main endpoints measured in laboratory toxicity tests designed to assess the effects of endocrine-active substances. They of population modeling might be used to establish SPGs for point out that most of these are “apical” endpoints such as this focal species. An IBM for skylarks was used to examine growth, development, sex ratio, and reproduction, which can several scenarios that represented the range of agricultural potentially be linked to adverse population effects. Increas- landscapes across Denmark. In this section we describe the ingly, as in mammalian toxicology, historical control data are way in which the information in Topping and Luttik (2017) can also available to allow a more thorough understanding of any be used to select focal species and protection goals, define statistically significant differences in these apical endpoints adverse effects, perform population modeling, and link (e.g., Valverde-Garcia et al. 2018). laboratory effects to wildlife populations. ECHA/EFSA (2018) guidance requires that an assessment Focal species for endocrine-disrupting properties is performed first for human health with use of the mammalian toxicology data set. The skylark is a small ground-nesting passerine that The guidance suggests that this requirement is primarily typically feeds on insects and breeds in fields during periods based on the results from tests on rats, such as the extended of PPP application. It is also one of the farmland bird species one-generation reproductive toxicity study (EOGRT; OECD in western Europe that has experienced a large population 2018b, with cohort 1a/1b, including the mating of cohort 1b decline, and it is the subject of an EU management plan to produce the F2 generation) (although this study is not (Topping and Luttik 2017). The skylark therefore fulfills EFSA currently a core data requirement for PPP registration in the (2009) criteria for a focal species because it occurs in the crop EU) or the two-generation reproduction toxicity test (OECD when PPPs are being used and it represents other bird 2001). If the criteria for establishing a substance as an species that may also occur in the crop at that time. endocrine disruptor are not met for human health, then the Protection goals assessment should move on to consider fish and amphibians, which may require further testing (the mammalian data EFSA SC (2016a) guidance states that the following need to package could be used to assess population relevance for be defined before setting a protection goal: the ecological wild mammals, but nothing is stated about this in the entity (e.g., individual, population, functional group, or guidance). The guidance suggests that this testing might ecosystem), the attribute of that entity (e.g., behavior, primarily include the medaka extended one-generation test growth, abundance, biomass, or ecosystem processes), the (MEOGRT; OECD 2015a) and the larval amphibian growth magnitude of effects (i.e., negligible, small, medium, or and development assay (LAGDA; OECD 2015b), although large), the temporal scale of effect for the attribute (e.g., again these are not currently data requirements for PPP duration and frequency), and the spatial scales (e.g., in-field registration in the EU. Any existing data on birds and reptiles and off-field patches of landscapes). If the ecological entity to should also be considered, although the guidance does not protect is the population of a particular species then EFSA SC identify specific tests for these taxa, and regulatory tests do (2016a) suggests that in most cases the attribute to be not currently exist for reptiles. Whichever test is used, the protected will be population dynamics in terms of abundance Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 288 Integr Environ Assess Manag 15, 2019—M Crane et al. Figure 3. The MTD/C concept and upper test level setting that can be applied to endocrine test guideline studies (adapted after Wheeler et al. 2013). Limit test 1 1 1 concentrations are 10 mg  L for fish and aquatic amphibians, 1000 ppm diet for birds (according to OECD TG 206), and 1000 mg  kg body weight  day for mammals (according to OECD TG 416). (e.g., numbers of individuals and their fitness) or biomass. constraints both as a limit to foraging success and as Topping and Luttik (2017) examined the effects of PPP increased energetic costs for cold weather. Initiation of application on total skylark population size and relative breeding depended upon a bird finding a suitable territory abundance occupancy, a measure that compares changes in and vegetation suitable for nesting. Breeding success spatial coverage (occupancy) and density (abundance) with or depended on the habitat being able to fulfil the energetic without PPP application. requirements of the birds during the breeding period and the Although the specific protection goal for effects on survival of eggs and nestlings, which was determined by food population abundance could be set at any level, it is highly resource quantity and availability (a function of management, likely that regulators in the EU would insist that there should weather, and skylark behavior). be negligible long-term adverse effects on abundance across Ten model landscapes were selected to be used in the the most ecologically sensitive landscapes. simulation runs, representing the range of agricultural practices in Denmark, from intensive to extensive. A wide Adverse effects range of different PPP application timing scenarios and levels EFSA SC (2013) identifies population recruitment, size, and of effect on different life stages were simulated. stability as the 3 aspects of a nontarget population that These simulations showed that in the most sensitive of the should be addressed when assessing the possible adverse 10 modeled landscapes a reduction in skylark eggshell effects of an endocrine disruptor. The skylark model thickness, which was associated with a 10% or less probability described by Topping and Luttik (2017) simulates population of clutch loss, led to negligible effects on population size over a 30-year period, so population stability and recruitment, size, and stability after a period of 20-30 years. recruitment are also addressed within the modeling process. Field studies Population modeling The model used by Topping and Luttik (2017) has been Topping and Luttik (2017) used the ALMaSS skylark model, extensively tested and reproduces a range of real-world which is an IBM within the open source Animal Landscape skylark population and individual behaviors. These attributes and Man Simulation System (https://gitlab.com/ include the mean and variation around time to hatch and nest ChrisTopping/ALMaSS_all). Individual birds were catego- leaving, densities of skylarks per farm, and within-season rized into 5 life stages: clutch, nestlings, prefledglings, males, phenology under different field conditions. and females. Available insect food biomass was determined Linking laboratory effects to wildlife populations by vegetation structure in the landscape and by its availability to birds foraging within a home range. Insect biomass was Bird eggs begin to crack in the laboratory when shell updated daily in the model and was affected by vegetation thickness is reduced by 18% (EFSA 2009). Topping and Luttik growth and human management, such as PPP application. (2017) show that a cracking rate of 10% or less did not lead to During the breeding period the model considered the adverse population effects for skylarks in even the most energetic balance of the adults, food requirements for sensitive simulated scenario that they investigated. A shell maintenance, requirements of the young, and weather thinning rate above 18% but below a value corresponding to Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 289 regulatory authorities and to adapt available models for the 10% cracking could therefore be set as a benchmark response. specific questions in the endocrine disruptor hazard assess- The results for eggshell thinning and cracking from ment. It may also be possible to use model outputs to develop look-up tables for adverse population effects, which laboratory-based avian reproduction tests can be directly linked to effects on skylark population recruitment, size, and could standardize and simplify the process, but this would stability, with this benchmark response indicating an inflec- need to be discussed with regulatory authorities. (iii) Definition of MTD/C for all vertebrate test methods so that tion point below which no adverse population effects would be expected. If avian testing is performed at the hazard- agreed hazard thresholds are available to minimize animal based dietary limit of 1000 ppm and the benchmark response testing and reduce the likelihood of observing systemic rather than endocrine disruptor-related toxicity in laboratory is not exceeded, then the tested substance is unlikely to cause adverse population effects in birds. tests. (iv) Perform detailed case studies to pilot the approach. The identification of endocrine disruptors in PPPs under the regulation is primarily a hazard-based and not a risk-based CONCLUSIONS In this paper we have suggested how to implement the new methodology, which is an approach to regulation that has endocrine disruptor criteria in the PPP regulation in a way that been strongly criticized in the past (Crane et al. 2019). The approach to identify adverse endocrine-disrupting popula- builds upon ECHA/EFSA (2018) guidance and is consistent with the PPP regulation legal text that mandates a hazard- tion-level effects that we recommend in this paper is hazard based approach. Our recommended approach is based to comply with the PPP regulation, but uncertainty would be significantly reduced and assessment relevance 1) Use SPGs for focal wildlife populations within EFSA’s improved if exposure was also explicitly considered during ecosystem services framework. The SPGs for vertebrate the process. For example, models are available at the organism level to predict whether an endocrine-disrupting wildlife can be summarized as (i) ecological entity: populations of vertebrate focal species; (ii) attribute: effect would be transient under realistic patterns of exposure population recruitment, size, and stability; (iii) effect (e.g., toxicokinetic and toxicodynamic models and dynamic energy budget models). Use of such models might negate magnitude: negligible; (iv) temporal scale: species life cycle, life span, or breeding period and an exposure the need to move along an AOP to the population level by frequency or duration agreed with regulatory authorities; showing that an adverse endocrine-disrupting effect had not occurred at the organism level. However, use of exposure (v) spatial scale: likely to be a watershed or landscape, but this element needs to be agreed upon with regulators; and duration and profiles information in these models implies a (vi) degree of certainty: high (e.g., 95%). risk-based approach, which is not currently allowed under the PPP regulation. We believe that further discussion of this 2) Model the effects of changes in population-related inputs on focal species populations with IBMs to determine issue between all stakeholders would help to reduce thresholds between negligible population effects and regulatory uncertainty and provide a more robust and science-based solution to the problem of detecting and nonnegligible (i.e., adverse) population effects. 3) Compare these thresholds with the relevant endpoints preventing adverse endocrine disruptor effects in the from laboratory toxicity tests to determine whether they environment. Regardless, all substances shown to be non– endocrine disrupting and potentially endocrine disrupting, are likely to be exceeded at the hazard-based limits or the MTD/C from the experimental studies. but without population-relevant adverse effects, will still 4) If the population threshold is not exceeded, then the undergo risk assessment, thus ensuring a high level of protection for the environment. substance should not be classified as an endocrine disruptor with adverse population effects unless there Acknowledgment—The authors thank Peter Day of the are other lines of evidence within a weight-of-evidence European Crop Protection Association for his support and approach to challenge this. comments and 2 anonymous reviewers for their interesting comments. If the proposals in this paper are accepted by stakeholders, Disclaimer—Mark Crane received funding from the Euro- then it is likely that future technical work will need to pean Crop Protection Association (ECPA) to research and concentrate on 4 main areas: (i) Selection of a small number of write this paper. All other co-authors work for agrochemical ecologically sensitive focal species for each taxonomic companies as indicated by their affiliations. group. This task has already been performed by EFSA for Data Accessibility—Please contact the corresponding birds and mammals (EFSA 2009), and Ibrahim et al. (2013) author, Mark Crane ([email protected]), for any and EFSA PPR (2018) have begun the process for fish, data used in this study. amphibians, and reptiles (although there are currently no laboratory test designs for the latter group). (ii) Development REFERENCES of suitable IBMs for these focal species. Models are already Ankley GT, Bennett RS, Erickson RJ, Hoff DJ, Hornung MW, Johnson RD, available for some bird and mammal focal species and for Mount DR, Nichols JW, Russom CL, Schmieder PK. 2010. Adverse some fish species. However, additional effort will be required outcome pathways: A conceptual framework to support ecotoxicology research and risk assessment. Environ Toxicol Chem 29:730–741. to develop models for additional focal species identified by Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors 290 Integr Environ Assess Manag 15, 2019—M Crane et al. [EFSA SC] European Food Safety Authority Scientific Committee. 2017b. Becker RA, Ankley GT, Edwards SW, Kennedy SW, Linkov I, Meek B, Sachana M, Segner H, Van Der Burg B, Villeneuve DL, et al. 2015. Increasing Guidance on the use of the weight of evidence approach in scientific scientific confidence in adverse outcome pathways: Application of tailored assessments. EFSA J 15:4971. [EFSA SC] European Food Safety Authority Scientific Committee. 2018. Bradford-Hill considerations for evaluating weight of evidence. Regul Toxicol Pharmacol 72:514–537. Guidance on uncertainty analysis in scientific assessments. EFSA J Blanchfield PJ, Kidd KA, Docker MF, Palace VP, Park BJ, Postma LD. 2015. 16:5123. Recovery of a wild fish population from whole-lake additions of a synthetic Giesy JP, Feyk LA, Jones PD, Kannan K, Sanderson T. 2003. Review of the estrogen. Environ Sci Technol 49:3136–3144. effects of endocrine-disrupting chemicals in birds. Pure Appl Chem Campbell PM, Hutchinson TH. 1998. Wildlife and endocrine disrupters: 75:2287–2303. Requirements for hazard identification. Environ Toxicol Chem 17: Hamilton PB, Nicol E, De-Bastos ESR, Williams RJ, Sumpter JP, Jobling S, 127–135. Stevens JR, Tyler CR. 2014. Populations of a cyprinid fish are self-sustaining Crane M, Hallmark N, Lagadic L, Ott K, Pickford D, Preuss T, Thompson H, despite widespread feminization of males. BMC Biol 12:1. Thorbek P, Weltje L, Wheeler JR. 2019. Establishing the relevance of Hamilton PB, Cowx IG, Oleksiak MF, Griffiths AM, Grahn M, Stevens JR, endocrine disrupting effects for nontarget vertebrate populations. Integr Carvalho GR, Nicol E, Tyler CR. 2016. Population-level consequences for Environ Assess Manag 15:299–301. wild fish exposed to sublethal concentrations of chemicals—A critical Day P, Green RM, Gross M, Weltje L, Wheeler JR. 2018. Endocrine disruption: review. Fish Fish 17:545–566. Current approaches for regulatory testing and assessment of plant Harris CA, Hamilton PB, Runnalls TJ, Vinciotti V, Henshaw A, Hodgson D, Coe protection products are fit for purpose. Toxicol Lett 296:10–22. TS, Jobling S, Tyler CR, Sumpter JP. 2011. The consequences of [EC] European Commission. 2018. Commission Regulation (EU) 2018/605 of feminization in breeding groups of wild fish. Environ Health Perspect 19 April 2018 amending Annex II to Regulation (EC) No 1107/2009 by 119:306–311. setting out scientific criteria for the determination of endocrine disrupting Hazlerigg CRE, Tyler CR, Lorenzen K, Wheeler JR, Thorbek P. 2014. Population relevance of toxicant mediated changes in sex ratio in fish: An assessment properties. Off J Eur Union L 101:33–36. [ECHA/EFSA] European Chemicals Agency, European Food Safety Authority, using an individual-based zebrafish (Danio rerio) model. Ecol Modell Joint Research Centre. 2018. Guidance for the identification of endocrine 280:76–88. disruptors in the context of Regulations (EU) No 528/2012 and (EC) No Hutchinson TH, Bogi € C, Winter MJ, Owens JW. 2009. Benefits of the maximum 1107/2009. EFSA J 16:5311. tolerated dose (MTD) and maximum tolerated concentration (MTC) Edwards SW, Tan YM, Villeneuve DL, Meek ME, McQueen CA. 2016. Adverse concept in aquatic toxicology. Aquat Toxicol 91:197–202. outcome pathways: Organizing toxicological information to improve Ibrahim L, Preuss TG, Ratte HT, Hommen U. 2013. A list of fish species that are decision making. J Pharmacol Exp Ther 356:170–181. potentially exposed to pesticides in edge-of-field water bodies in the [EFSA] European Food Safety Authority. 2009. Guidance document on risk European Union—A first step towards identifying vulnerable representa- assessment for birds and mammals. EFSA J 7:1438. tives for risk assessment. Environ Sci Pollut Res Int 20:2679–2687. [EFSA] European Food Safety Authority. 2010. Report on the PPR stakeholder Kerlin R, Bolon B, Burkhardt J, Francke S, Greaves P, Meador V, Popp J. 2016. workshop protection goals for environmental risk assessment of pesti- Scientific and Regulatory Policy Committee: Recommended (“best”) cides: What and where to protect? EFSA J 8:1672. practices for determining, communicating, and using adverse effect data [EFSA PPR] European Food Safety Authority Panel on Plant Protection Products from nonclinical studies. Toxicol Pathol 44:147–162. and Their Residues. 2010. Scientific opinion on the development of Kidd KA, Blanchfield PJ, Mills KH, Palace VP, Evans RE, Lazorchak JM, Flick RW. specific protection goal options for environmental risk assessment of 2007. Collapse of a fish population after exposure to a synthetic estrogen. pesticides, in particular in relation to the revision of the Guidance Proc Natl Acad Sci USA 104:8897–8901. Documents on Aquatic and Terrestrial Ecotoxicology (SANCO/3268/2001 Knillmann S, Stampfli NC, Noskov YA, Beketov MA, Liess M. 2012. Interspecific and SANCO/10329/2002). EFSA J 8:1821. competition delays recovery of Daphnia spp. populations from pesticide [EFSA PPR] European Food Safety Authority Panel on Plant Protection Products stress. Ecotoxicology 21:1039–1049. and Their Residues. 2013. Guidance on tiered risk assessment for plant Kramer VJ, Etterson MA, Hecker M, Murphy CA, Roesijadi G, Spade DJ, protection products for aquatic organisms in edge-of-field surface waters. Spromberg JA, Wang M, Ankley GT. 2011. Adverse outcome pathways EFSA J 11:3290. and ecological risk assessment: Bridging to population-level effects. [EFSA PPR] European Food Safety Authority Panel on Plant Protection Products Environ Toxicol Chem 30:64–76. and Their Residues. 2014. Scientific opinion on good modelling practice in Lambeck RJ. 1997. Focal species: A multi-species umbrella for nature the context of mechanistic effect models for risk assessment of plant conservation. Conserv Biol 11:849–856. protection products. EFSA J 12:3589. Lewis RW, Billington R, Debryune E, Gamer A, Lang B, Carpanini F. 2002. [EFSA PPR] European Food Safety Authority Scientific Committee. 2018. Recognition of adverse and nonadverse effects in toxicity studies. Toxicol Scientific opinion on the state of the science on pesticide risk assessment Pathol 30:66–74. for amphibians and reptiles. EFSA J 16:5125. Liu C, Sibly RM, Grimm V, Thorbek P. 2013. Linking pesticide exposure and [EFSA SC] European Food Safety Authority Scientific Committee. 2013. spatial dynamics: An individual-based model of wood mouse (Apodemus Scientific opinion on the hazard assessment of endocrine disruptors: sylvaticus) populations in agricultural landscapes. Ecol Modell 248:92–102. Scientific criteria for identification of endocrine disruptors and appro- Marty MS, Blankinship A, Chambers J, Constantine L, Kloas W, Kumar A, priateness of existing test methods for assessing effects mediated by Lagadic L, Meador J, Pickford D, Schwarz T, et al. 2017. Population- these substances on human health and the environment. EFSA J relevant endpoints in the evaluation of endocrine-active substances (EAS) 11:3132. for ecotoxicological hazard and risk assessment. Integrated Environ Assess [EFSA SC] European Food Safety Authority Scientific Committee. 2016a. Manag 13:317–330. Guidance to develop specific protection goals options for environmental Matthiessen P, Weltje L. 2015. A review of the effects of azole compounds in risk assessment at EFSA, in relation to biodiversity and ecosystem services. fish and their possible involvement in masculinization of wild fish EFSA J 14:4499. populations. Crit Rev Toxicol 45:453–467. [EFSA SC] European Food Safety Authority Scientific Committee. 2016b. Matthiessen P, Wheeler JR, Weltje L. 2018. A review of the evidence for Scientific opinion on recovery in environmental risk assessments at EFSA. endocrine disrupting effects of current-use chemicals on wildlife pop- EFSA J 14:4313. ulations. Crit Rev Toxicol 48:195–216. [EFSA SC] European Food Safety Authority Scientific Committee. 2017a. Mintram KS, Brown AR, Maynard SK, Thorbek P, Tyler CR. 2017. Capturing Scientific opinion on guidance on the assessment of the biological ecology in modeling approaches applied to environmental risk assessment relevance of data in scientific assessments. EFSA J 15:4970. of endocrine active chemicals in fish. Crit Rev Toxicol 48:109–120. Integr Environ Assess Manag 2019:278–291 DOI: 10.1002/ieam.4113 2018 The Authors Endocrine Effects and Nontarget Vertebrates—Integr Environ Assess Manag 15, 2019 291 Munns WR Jr, Barnthouse LW, Sorensen MT. 2007. Issues and recommenda- Raimondo S, Etterson M, Pollesch N, Garber K, Kanarek A, Lehmann W, tions. In: Barnthouse LW, Munns WR Jr, Sorensen MT, editors. Population- Awkerman J. 2018. A framework for linking population model develop- level ecological risk assessment. Boca Raton (FL): CRC Press. p 239–246. ment with ecological risk assessment objectives. Integrated Environ Assess Manag 14:369–380. [OECD] Organization for Economic Cooperation and Development. 1984. Avian reproduction test. Guideline for Testing of Chemicals No 206. Paris Schmitt W, Auteri D, Bastiansen F, Ebeling M, Liu C, Luttik R, Mastitsky S, Nacci (FR): OECD Publishing. D, Topping C, Wang M. 2016. An example of population-level risk [OECD] Organization for Economic Cooperation and Development. 2000. assessments for small mammals using individual-based population Guidance document on aquatic toxicity testing of difficult substances and models. Integr Environ Assess Manag 12:46–57. Topping CJ, Elmeros M. 2016. Modeling exposure of mammalian predators to mixtures. Environmental Health and Safety Publications Series on Testing and Assessment No 23. Paris (FR): OECD Publishing. anticoagulant rodenticides. Front Environ Sci 4:80. [OECD] Organization for Economic Cooperation and Development. 2001. Topping CJ, Luttik R. 2017. Simulation to aid in interpreting biological Two-generation reproduction toxicity study. Guideline for Testing of relevance and setting of population-level protection goals for risk Chemicals No 416. Paris (FR): OECD Publishing. assessment of pesticides. Regul Toxicol Pharmacol 89:40–49. [OECD] Organization for Economic Cooperation and Development. 2012. Fish Topping CJ, Weyman GS. 2018. Rabbit population landscape-scale simulation toxicity testing framework. OECD Environment, Health and Safety to investigate the relevance of using rabbits in regulatory environmental Publications Series on Testing and Assessment No 171. Paris (FR): risk assessment. Environ Model Assess 23:415–457. OECD Publishing. Tyler CR, Jobling S, Sumpter JP. 2008. Endocrine disruption in wildlife: A [OECD] Organization for Economic Cooperation and Development. 2015a. critical review of the evidence. Crit Rev Toxicol 28:319–361. Medaka extended one generation reproduction test (MEOGRT). Guideline [UN] United Nations. 2011. Globally harmonized system of classification for the Testing of Chemicals No 240. Paris (FR): OECD Publishing. and labelling of chemicals (GHS). 4th ed. New York (NY): United [OECD] Organization for Economic Cooperation and Development. 2015b. Nations. 561 p. The larval amphibian growth and development assay (LAGDA). Guideline Valverde-Garcia P, Springer T, Kramer V, Foudoulakis M, Wheeler JR. 2018. An for the Testing of Chemicals No 241. Paris (FR): OECD Publishing. avian reproduction study historical control database: A tool for data [OECD] Organization for Economic Cooperation and Development. 2018a. interpretation. Regul Toxicol Pharmacol 92:295–302. Revised guidance document 150 on standardised test guidelines for Wang M. 2013. From home range dynamics to population cycles: Validation evaluating chemicals for endocrine disruption. OECD Series on Testing and realism of a common vole population model for pesticide risk assessment. Integr Environ Assess Manag 9:294–307. and Assessment. Paris (FR): OECD Publishing. https://doi.org/10.1787/ 9789264304741-en Wheeler JR, Panter GH, Weltje L, Thorpe KL. 2013. Test concentration setting [OECD] Organization for Economic Cooperation and Development. 2018b. for fish in vivo endocrine screening assays. Chemosphere 92:1067–1076. Extended one-generation reproductive toxicity study. Guideline for the Wheeler JR, Weltje L. 2015. Adverse outcome pathways—An industry Testing of Chemicals No 443. Paris (FR): OECD Publishing. perspective. Environ Toxicol Chem 34:1937–1938. [WHO/IPCS] World Health Organization/International Programme on Chemical Ottinger MA, Dean K, McKernan M, Quinn MJ Jr. 2011. Endocrine disruption of reproduction in birds. In: Norris DO, Lopez KH, editors. Hormones and Safety. 2002. Global Assessment of the state-of-the-science of endocrine reproduction of vertebrates. Vol 4. London (UK): Academic Press. p disruptors. Geneva (CH): World Health Organization. WHO/PCS/EDC/02.2. 239–260. 180 p. Integr Environ Assess Manag 2019:278–291 wileyonlinelibrary.com/journal/ieam 2018 The Authors

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