Characterizing the Movement of Per- and Polyfluoroalkyl Substances in an Avian Aquatic-Terrestrial Food Web.

The movement of per- and polyfluoroalkyl substances (PFAS) through linked aquatic-terrestrial food webs is not well understood. Tree swallows (Tachycineta bicolor) in such systems may be exposed to PFAS from multiple abiotic and/or biotic compartments. We show from fatty acid signatures and carbon stable isotopes that tree swallow nestlings in southwestern Ontario fed on both terrestrial and aquatic macroinvertebrates. The PFAS profiles of air, terrestrial invertebrates, and swallows were dominated by perfluorooctanesulfonic acid (PFOS). Short-chain perfluoroalkyl acids (PFAAs) were largely restricted to air, surface water, and sediment, and long-chain PFAAs were mainly found in aquatic invertebrates and tree swallows. PFOS, multiple long-chain perfluorocarboxylic acids [perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), perfluorotridecanoic acid (PFTrDA)] and perfluorooctane sulfonamide precursors were estimated to bioaccumulate from air to tree swallows. PFOS bioaccumulated from air to terrestrial invertebrates, and PFOS, PFDA, and perfluorooctane sulfonamidoacetic acids (FOSAAs) bioaccumulated from water to aquatic invertebrates. PFOS showed biomagnification from both terrestrial and aquatic invertebrates to tree swallows, and PFDA and FOSAAs were also biomagnified from aquatic invertebrates to tree swallows. The movement of PFAS through aquatic-terrestrial food webs appears congener- and compartment-specific, challenging the understanding of PFAS exposure routes for multiple species involved in these food webs.

Hepatic Gene Expression Profiling of American Kestrels (Falco sparverius) Exposed In Ovo to Three Alternative Brominated Flame Retardants.

A number of brominated flame retardants (BFRs) have been reported to interfere with the thyroid signaling pathway and cause oxidative stress in birds, yet the underlying shifts in gene expression associated with these effects remain poorly understood. In this study, we measured hepatic transcriptional responses of 31 genes in American kestrel (Falco sparverius) hatchlings following in ovo exposure to one of three high-volume alternative BFRs: 1,2-bis(2,4,6-tribromophenoxy) ethane (BTPBE), bis(2-ethylhexyl)-2,3,4,5-tetrabromophthalate (TBPH), or 2-ethylhexyl-2,3,4,5-tetrabromobenzoate (EHTBB). Hatchling kestrels exhibited shifts in the expression of genes related to oxidative stress (CYP, GSTA, SOD, and GPX1), thyroid hormone metabolism and transport (DIO1, DIO2, and TTR), lipid and protein metabolism (PPAR, HMGCR, FAB1, and LPL), and cytokine-mediated inflammation (TLR3, IL18, IRF7, STAT3, RACK1, and CEBPB). Male and female hatchlings differed in which genes were differentially expressed, as well as the direction of the effect (up- vs. downregulation). These results build upon our previous findings of increased oxidative stress and disrupted thyroid signaling pathway in the same hatchlings. Furthermore, our results indicate that inflammatory responses appear to occur in female hatchlings exposed to BTBPE and EHTBB in ovo. Gene expression analysis revealed multiple affected pathways, adding to the growing evidence that sublethal physiological effects are complex and are a concern for birds exposed to BTBPE, EHTBB, or TBPH in ovo.

Exposure to crop production alters cecal prokaryotic microbiota, inflates virulome and resistome in wild prairie grouse.

Chemically intensive crop production depletes wildlife food resources, hinders animal development, health, survival, and reproduction, and it suppresses wildlife immune systems, facilitating emergence of infectious diseases with excessive mortality rates. Gut microbiota is crucial for wildlife's response to environmental stressors. Its composition and functionality are sensitive to diet changes and environmental pollution associated with modern crop production. In this study we use shotgun metagenomics (median 8,326,092 sequences/sample) to demonstrate that exposure to modern crop production detrimentally affects cecal microbiota of sharp-tailed grouse (Tympanuchus phasianellus: 9 exposed, 18 unexposed and greater prairie chickens (T. cupido; 11, 11). Exposure to crop production had greater effect on microbiota richness (t = 6.675, P < 0.001) and composition (PERMANOVA r2 = 0.212, P = 0.001) than did the host species (t = 4.762, P < 0.001; r2 = 0.070, P = 0.001) or their interaction (t = 3.449; r2 = 0.072, both P = 0.001), whereas sex and age had no effect. Although microbiota richness was greater in exposed (T. cupido chao1 = 152.8 ± 20.5; T. phasianellus 115.3 ± 17.1) than in unexposed (102.9 ± 15.1 and 101.1 ± 17.2, respectively) birds, some beneficial bacteria dropped out of exposed birds' microbiota or declined and were replaced by potential pathogens. Exposed birds also had higher richness and load of virulome (mean ± standard deviation; T. cupido 24.8 ± 10.0 and 10.1 ± 5.5, respectively; T. phasianellus 13.4 ± 6.8/4.9 ± 2.8) and resistome (T. cupido 46.8 ± 11.7/28.9 ± 10.2, T. phasianellus 38.3 ± 16.7/18.9 ± 14.2) than unexposed birds (T. cupido virulome: 14.2 ± 13.5, 4.5 ± 4.2; T. cupido resistome: 31.6 ± 20.2 and 13.1 ± 12.0; T. phasianellus virulome: 5.2 ± 4.7 and 1.4 ± 1.5; T. phasianellus resistome: 13.7 ± 16.1 and 4.0 ± 6.4).

Establishment of baseline cytology metrics in nestling American kestrels (Falco sparverius): Immunomodulatory effects of the flame retardant isopropylated triarylphosphate isomers.

Avian populations must mount effective immune responses upon exposure to environmental stressors such as avian influenza and xenobiotics. Although multiple immune assays have been tested and applied to various avian species, antibody-mediated immune responses in non-model avian species are not commonly reported due to the lack of commercially available species-specific antibodies. The objectives of the present study were to advance methods for studying wild bird immune responses and to apply these to the evaluation of cytological responses after exposure of American kestrels, Falco sparverius, to a commercial flame retardant mixture containing isopropylated triarylphosphate isomers (ITP). Hatchlings were gavaged daily with safflower oil or 1.5 ug/g bw/day of ITP suspended in safflower oil, then bled on days 9, 17, and 21. The ITP treatment group (n = 18) and a subset of controls (Poly I:C treatment group; n = 10) were injected on days 9 and 15 with a synthetic analog of viral double-stranded RNA, polyinosinic:polycytidylic acid (Poly I:C), a toll-like receptor ligand and synthetic viral mimic, and responses compared to a sham injected control group (n = 8). The hypotheses tested whether kestrels showed immunological differences among treatment groups, genetic sex, and/or white blood cell (WBC) subpopulation type over time. A flow cytometry (FCM) gating strategy categorized heterophils (H), lymphocytes (L), and monocytes (M) and their proportions, and measured relative fluorescence in response to anti-chicken CD4 binding. Fluorescent cell surfaces and some granular/vacuolar inclusions were visualized by epifluorescence microscopy. A fourth subpopulation with higher levels of granularity than M but less than H became increasingly apparent with time and was gated along with the H subpopulation; its frequency of occurrence was lowest in the ITP group (P = 0.0023). The percentages of cells differed among treatment groups, days, and sexes (P = 0.0001). For both sexes, percentages of H and L were higher than M in control and Poly I:C. In the ITP group, L percentages were higher than H and M (P = 0.0457), and H and L were higher than M on days 9 and 21 (P = 0.0001). The ratios of H:L and H:WBC, indicators of robust immunity, were also higher on days 9 and 21 than on 17 (P = 0.0079). For each sex, the highest levels of activity measured by FCM geometric means (GEO) of fluorescence (indicative of antibody binding) were observed on day 9 (P = 0.0001 female, and P = 0.0011 male) in H over both L and M (P < 0.0001 for each). In males, GEO of the Poly I:C group was higher than that of the ITP group (P = 0.0374), with no difference observed among females over all days. By using a FCM algorithm for population comparisons of fluorescence to investigate binding within H, the T(x) scores indicated higher fluorescence in control and Poly I:C groups over ITP (P = 0.0001). Unlike chickens, Gallus gallus, which express CD4 primarily on L, kestrels bound the commercial antibody primarily within the gated H subpopulation, suggesting an immunophenotypic difference between taxa, despite a ~60% identity of Falco CD4 amino acid sequences with chicken CD4. The emergent cell subset within the gated H presented dendritic-like cell (DLC) morphological and functional properties, apparently serving as an effector cell. This study adds interpretive context to ecological investigations of infection and of potential immunomodulation by emerging compounds, whereby the early innate responses are mediated by the various cell subsets serving as useful quantitative markers of immunological condition. Data showed that dietary exposure to ITP was immunosuppressive for male and female kestrels over the course of the experiment, reducing DLC frequency compared to the Poly I:C controls. Heterophils and DLC were important in facilitating innate immunological responses.

Thyroid disruption and oxidative stress in American kestrels following embryonic exposure to the alternative flame retardants, EHTBB and TBPH.

Brominated flame retardant chemicals, such as 2-ethylhexyl-2,3,4,5-tetrabromobenzoate (EHTBB) (CAS #: 183658-27-7) and bis(2-ethylhexyl)-2,3,4,5-tetrabromophthalate (TBPH) (CAS #: 26040-51-7), have been detected in avian tissues and eggs from remote regions. Exposure to EHTBB and TBPH has been shown to cause oxidative stress and altered thyroid function in rodents and fish, yet no controlled studies have examined potential adverse effects of exposure in birds. Because flame retardants have been detected in wild raptors, we used American kestrels (Falco sparverius) as a model raptor to determine whether in ovo exposure to EHTBB or TBPH affected growth, hatching success, oxidative stress, or thyroid function. We exposed kestrel embryos to nominal concentrations (10, 50, or 100 ng g-1 egg weight) of EHTBB and TBPH via egg-injection on embryonic day 5. Embryonic exposure (~23 d) to EHTBB increased thyroid gland mass, reduced glandular colloid and total thyroxine (T4) in hatchling males and females, whereas deiodinase enzyme activity increased in males but decreased in females. Hatchlings exposed to TBPH in ovo exhibited reduced colloid and increased oxidative stress. Although exposure to EHTBB and TBPH caused several physiological effects (e.g., heart and brain mass), only exposure to 50 ng g-1 EHTBB appeared to reduce hatching success. Our results suggest these flame retardants may be hazardous for predatory birds. Future research should evaluate long-term survival and fitness consequences in birds exposed to these chemicals.

Sex- and Developmental Stage-Related Differences in the Hepatic Transcriptome of Japanese Quail (Coturnix japonica) Exposed to 17ß-Trenbolone.

Endocrine-disrupting chemicals can cause transcriptomic changes that may disrupt biological processes associated with reproductive function including metabolism, transport, and cell growth. We investigated effects from in ovo and dietary exposure to 17ß-trenbolone (at 0, 1, and 10 ppm) on the Japanese quail (Coturnix japonica) hepatic transcriptome. Our objectives were to identify differentially expressed hepatic genes, assess perturbations of biological pathways, and examine sex- and developmental stage-related differences. The number of significantly differentially expressed genes was higher in embryos than in adults. Male embryos exhibited greater differential gene expression than female embryos, whereas in adults, males and females exhibited similar numbers of differentially expressed genes (>2-fold). Vitellogenin and apovitellenin-1 were up-regulated in male adults exposed to 10 ppm 17ß-trenbolone, and these birds also exhibited indications of immunomodulation. Functional grouping of differentially expressed genes identified processes including metabolism and transport of biomolecules, enzyme activity, and extracellular matrix interactions. Pathway enrichment analyses identified as perturbed peroxisome proliferator-activated receptor pathway, cardiac muscle contraction, gluconeogenesis, growth factor signaling, focal adhesion, and bile acid biosynthesis. One of the primary uses of 17ß-trenbolone is that of a growth promoter, and these results identify effects on mechanistic pathways related to steroidogenesis, cell proliferation, differentiation, growth, and metabolism of lipids and proteins. Environ Toxicol Chem 2021;40:2559-2570. © 2021 SETAC. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

In ovo exposure to brominated flame retardants Part I: Assessment of effects of TBBPA-BDBPE on survival, morphometric and physiological endpoints in zebra finches.

Tetrabromobisphenol A bis(2,3-dibromopropyl) ether (TBBPA-BDBPE) is an additive flame retardant used in polyolefins and polymers. It has been detected in biota, including in avian eggs, yet little is known of its effects. We assessed the pattern of TBBPA-BDBPE concentrations in songbird eggs over the incubation period, and the effects of embryonic exposure to TBBPA-BDBPE in a model songbird species, the zebra finch (Taeniopygia guttata). To assess concentrations during embryo development, eggs were injected on the day they were laid with the vehicle control (safflower oil) or 100 ng TBBPA-BDBPE/g egg, and whole egg contents were collected throughout embryonic development on day 0 (unincubated), 5, 10 and 13. To evaluate effects of embryonic exposure to TBBPA-BDBPE, eggs were injected at Hamburger-Hamilton stage 18 (∼80 h after initiation of incubation) with safflower oil only, 10, 50 or 100 ng TBBPA-BDBPE/g egg (albumin injection volume 1 µl/g). Eggs were monitored for hatching success, and nestlings were monitored for growth and survival. At 15 days post-hatch, tissues were collected to assess physiological effects. TBBPA-BDBPE was incorporated into the egg as the embryo developed, and concentrations started declining in late incubation, suggesting biotransformation by the embryo. There were no effects on hatching success, nestling survival, growth, organ somatic indices, or thyroid hormone homeostasis; however, there was evidence that body condition declined in a dose-dependent manner towards the end of the rapid nestling growth phase. This decreased body condition could be a delayed effect of early developmental exposure, or it may be the result of increased exposure to biotransformation products of TBBPA-BDBPE produced over the nestling period, which are predicted to be more bioaccumulative and toxic than the parent compound.

In ovo exposure to brominated flame retardants Part II: Assessment of effects of TBBPA-BDBPE and BTBPE on hatching success, morphometric and physiological endpoints in American kestrels.

Tetrabromobisphenol A bis(2,3-dibromopropyl ether) (TBBPA-BDBPE) and 1,2-bis(2,4,6-tribromophenoxy)ethane (BTPBE) are both brominated flame retardants (BFRs) that have been detected in birds; however, their potential biological effects are largely unknown. We assessed the effects of embryonic exposure to TBBPA-BDBPE and BTBPE in a model avian predator, the American kestrel (Falco sparverius). Fertile eggs from a captive population of kestrels were injected on embryonic day 5 (ED5) with a vehicle control or one of three doses within the range of concentrations that have been detected in biota (nominal concentrations of 0, 10, 50 or 100 ng/g egg; measured concentrations 0, 3.0, 13.7 or 33.5 ng TBBPA-BDBPE/g egg and 0, 5.3, 26.8 or 58.1 ng BTBPE/g egg). Eggs were artificially incubated until hatching (ED28), at which point blood and tissues were collected to measure morphological and physiological endpoints, including organ somatic indices, circulating and glandular thyroid hormone concentrations, thyroid gland histology, hepatic deiodinase activity, and markers of oxidative stress. Neither compound had any effects on embryo survival through 90% of the incubation period or on hatching success, body mass, organ size, or oxidative stress of hatchlings. There was evidence of sex-specific effects in the thyroid system responses to the BTBPE exposures, with type 2 deiodinase (D2) activity decreasing at higher doses in female, but not in male hatchlings, suggesting that females may be more sensitive to BTBPE. However, there were no effects of TBBPA-BDBPE on the thyroid system in kestrels. For the BTPBE study, a subset of high-dose eggs was collected throughout the incubation period to measure changes in BTBPE concentrations. There was no decrease in BTBPE over the incubation period, suggesting that BTBPE is slowly metabolized by kestrel embryos throughout their ∼28-d development. These two compounds, therefore, do not appear to be particularly toxic to embryos of the American kestrel.

Toxicokinetics of Imidacloprid-Coated Wheat Seeds in Japanese Quail ( Coturnix japonica) and an Evaluation of Hazard.

Birds are potentially exposed to neonicotinoid insecticides by ingestion of coated seeds during crop planting. Adult male Japanese quail were orally dosed with wheat seeds coated with an imidacloprid (IMI) formulation at either 0.9 or 2.7 mg/kg body weight (BW) (∼3 and 9% of IMI LD50 for Japanese quail, respectively) for 1 or 10 days. Quail were euthanized between 1 and 24 h postexposure to assess toxicokinetics. Analysis revealed rapid absorption (1 h) into blood and distribution to the brain, muscle, kidney, and liver. Clearance to below detection limits occurred at both dose levels and exposure durations in all tissues within 24 h. Metabolism was extensive, with 5-OH-IMI and IMI-olefin detected at greater concentrations than IMI in tissues and fecal samples. There was no lethality or overt signs of toxicity at either dose level. Furthermore, no evidence of enhanced expression of mRNA genes associated with hepatic xenobiotic metabolism, oxidative DNA damage, or alterations in concentrations of corticosterone and thyroid hormones was observed. Application of the toxicokinetic data was used to predict IMI residue levels in the liver with reasonable results for some field exposure and avian mortality events. It would appear that some affected species of birds are either consuming larger quantities of seeds or exhibit differences in ADME or sensitivity than predicted by read-across from these data.

Sex-specific responses in neuroanatomy of hatchling American kestrels in response to embryonic exposure to the flame retardants bis(2-ethylhexyl)-2,3,4,5-tetrabromophthalate and 2-ethylhexyl-2,3,4,5-tetrabromobenzoate.

Bis(2-ethylhexyl)-2,3,4,5-tetrabromophthalate (BEH-TEBP) and 2-ethylhexyl-2,3,4,5-tetrabromobenzoate (EH-TBB), flame retardant components of FireMaster 550® and 600® have been detected in tissues of wild birds. To address the paucity of information regarding potential impacts of flame retardants on the brain, brain volume regions of hatchling American kestrels (Falco sparverius) were evaluated following in ovo injection at embryonic day 5 with safflower oil or to 1 of 3 doses of either BEH-TEBP (12, 60, or 107 ng/g egg) or EH-TBB (11, 55, or 137 ng/g egg). The doses for both chemicals reflected concentrations reported in wild birds. The volumes of the hippocampus and telencephalon and volumetric differences between left and right hemispheres were measured in hatchlings (embryonic day 28). A sex-specific effect of BEH-TEBP on relative hippocampus volume was evident: the hippocampus was significantly enlarged in high-dose females compared to control females but smaller in low-dose females than the other females. There was no significant effect of EH-TBB on hippocampus volume in female kestrel hatchlings or of either chemical in male hatchlings and no effects of these concentrations of EH-TBB or BEH-TEBP on telencephalon volume or the level of symmetry between the hemispheres of the brain. In sum, embryonic exposure of female kestrels to these BEH-TEBP concentrations altered hippocampus volume, having the potential to affect spatial memory relating to ecologically relevant behavior such as prey capture, predator avoidance, and migration. Environ Toxicol Chem 2018;37:3032-3040. © 2018 SETAC.

Female hatchling American kestrels have a larger hippocampus than males: A link with sexual size dimorphism?

The brain and underlying cognition may vary adaptively according to an organism's ecology. As with all raptor species, adult American kestrels (Falco sparverius) are sexually dimorphic with females being larger than males. Related to this sexual dimorphism, kestrels display sex differences in hunting and migration, with females ranging more widely than males, suggesting possible sex differences in spatial cognition. However, hippocampus volume, the brain region responsible for spatial cognition, has not been investigated in raptors. Here, we measured hippocampus and telencephalon volumes in American kestrel hatchlings. Female hatchlings had a significantly larger hippocampus relative to the telencephalon and brain weight than males (∼12% larger), although telencephalon volume relative to brain weight and body size was similar between the sexes. The magnitude of this hippocampal sex difference is similar to that reported between male and female polygynous Microtus voles and migratory and non-migratory subspecies of Zonotrichia sparrows. Future research should determine if this sex difference in relative hippocampus volume of hatchling kestrels persists into adulthood and if similar patterns exist in other raptor species, thus potentially linking sex differences in the brain to sex differences of space use of adults in the wild.

Biomarker responses of Peromyscus leucopus exposed to lead and cadmium in the Southeast Missouri Lead Mining District.

Biomarker responses and histopathological lesions have been documented in laboratory mammals exposed to elevated concentrations of lead and cadmium. The exposure of white-footed mice (Peromyscus leucopus) to these metals and the potential associated toxic effects were examined at three contaminated sites in the Southeast Missouri Lead Mining District and at a reference site in MO, USA. Mice from the contaminated sites showed evidence of oxidative stress and reduced activity of red blood cell δ-aminolevulinic acid dehydratase (ALAD). Histological examinations of the liver and kidney, cytologic examination of blood smears, and biomarkers of lipid peroxidation and DNA damage failed to show indications of toxic effects from lead. The biomagnification factor of cadmium (hepatic concentration/soil concentration) at a site with a strongly acid soil was 44 times the average of the biomagnification factors at two sites with slightly alkaline soils. The elevated concentrations of cadmium in the mice did not cause observable toxicity, but were associated with about a 50% decrease in expected tissue lead concentrations and greater ALAD activity compared to the activity at the reference site. Lead was associated with a decrease in concentrations of hepatic glutathione and thiols, whereas cadmium was associated with an increase. In addition, to support risk assessment efforts, we developed linear regression models relating both tissue lead dosages (based on a previously published a laboratory study) and tissue lead concentrations in Peromyscus to soil lead concentrations.

Effects on Circulating Steroid Hormones and Gene Expression along the Hypothalamus-Pituitary-Gonadal Axis in Adult Japanese Quail Exposed to 17ß-Trenbolone across Multiple Generations.

We investigated the effects of the androgenic growth promoter 17ß-trenbolone (17ßTB) on adult Japanese quail (Coturnix japonica) exposed across three generations. The F0 generation was exposed after sexual maturity to 0, 1, 5, 10, 20, and 40 ppm through feed. The F1 generation was exposed in ovo by maternal transfer and through feed at the same doses as their parents. The F2 generation was exposed in ovo only. Levels of plasma sex steroids, gonadal Cytochrome P450 aromatase (CYP19A1) mRNA and select brain neuroendocrine peptide mRNAs were measured. In males, testosterone levels did not differ in any generation from those in controls. Estradiol was significantly elevated in 17ßTB treated F0 and F1 males. In F0 and F1 females, testosterone was suppressed by 17ßTB, whereas estradiol was significantly higher at 40 ppm in F0 and at 10 ppm in F1 females. CYP19A1 expression in F1 males and females increased suggesting a compensatory response to the androgenic effects of 17ßTB. Few significant effects were observed in the F2 birds indicating that in ovo exposure had limited effects on the monitored endpoints. Overall, our results confirmed endocrine disrupting effects of dietary 17ßTB in Japanese quail but the response was dependent on sex, developmental stage at initiation of exposure, and dose.

Recommended approaches to the scientific evaluation of ecotoxicological hazards and risks of endocrine-active substances.

A SETAC Pellston Workshop® "Environmental Hazard and Risk Assessment Approaches for Endocrine-Active Substances (EHRA)" was held in February 2016 in Pensacola, Florida, USA. The primary objective of the workshop was to provide advice, based on current scientific understanding, to regulators and policy makers; the aim being to make considered, informed decisions on whether to select an ecotoxicological hazard- or a risk-based approach for regulating a given endocrine-disrupting substance (EDS) under review. The workshop additionally considered recent developments in the identification of EDS. Case studies were undertaken on 6 endocrine-active substances (EAS-not necessarily proven EDS, but substances known to interact directly with the endocrine system) that are representative of a range of perturbations of the endocrine system and considered to be data rich in relevant information at multiple biological levels of organization for 1 or more ecologically relevant taxa. The substances selected were 17α-ethinylestradiol, perchlorate, propiconazole, 17ß-trenbolone, tributyltin, and vinclozolin. The 6 case studies were not comprehensive safety evaluations but provided foundations for clarifying key issues and procedures that should be considered when assessing the ecotoxicological hazards and risks of EAS and EDS. The workshop also highlighted areas of scientific uncertainty, and made specific recommendations for research and methods-development to resolve some of the identified issues. The present paper provides broad guidance for scientists in regulatory authorities, industry, and academia on issues likely to arise during the ecotoxicological hazard and risk assessment of EAS and EDS. The primary conclusion of this paper, and of the SETAC Pellston Workshop on which it is based, is that if data on environmental exposure, effects on sensitive species and life-stages, delayed effects, and effects at low concentrations are robust, initiating environmental risk assessment of EDS is scientifically sound and sufficiently reliable and protective of the environment. In the absence of such data, assessment on the basis of hazard is scientifically justified until such time as relevant new information is available. Integr Environ Assess Manag 2017;13:267-279. © 2017 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals, Inc. on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Current limitations and recommendations to improve testing for the environmental assessment of endocrine active substances.

In the present study, existing regulatory frameworks and test systems for assessing potential endocrine active chemicals are described, and associated challenges are discussed, along with proposed approaches to address these challenges. Regulatory frameworks vary somewhat across geographies, but all basically evaluate whether a chemical possesses endocrine activity and whether this activity can result in adverse outcomes either to humans or to the environment. Current test systems include in silico, in vitro, and in vivo techniques focused on detecting potential endocrine activity, and in vivo tests that collect apical data to detect possible adverse effects. These test systems are currently designed to robustly assess endocrine activity and/or adverse effects in the estrogen, androgen, and thyroid hormone signaling pathways; however, there are some limitations of current test systems for evaluating endocrine hazard and risk. These limitations include a lack of certainty regarding: 1) adequately sensitive species and life stages; 2) mechanistic endpoints that are diagnostic for endocrine pathways of concern; and 3) the linkage between mechanistic responses and apical, adverse outcomes. Furthermore, some existing test methods are resource intensive with regard to time, cost, and use of animals. However, based on recent experiences, there are opportunities to improve approaches to and guidance for existing test methods and to reduce uncertainty. For example, in vitro high-throughput screening could be used to prioritize chemicals for testing and provide insights as to the most appropriate assays for characterizing hazard and risk. Other recommendations include adding endpoints for elucidating connections between mechanistic effects and adverse outcomes, identifying potentially sensitive taxa for which test methods currently do not exist, and addressing key endocrine pathways of possible concern in addition to those associated with estrogen, androgen, and thyroid signaling. Integr Environ Assess Manag 2017;13:302-316. © 2016 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals, Inc. on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Chesapeake Bay fish-osprey (Pandion haliaetus) food chain: Evaluation of contaminant exposure and genetic damage.

From 2011 to 2013, a large-scale ecotoxicological study was conducted in several Chesapeake Bay (USA) tributaries (Susquehanna River and flats, the Back, Baltimore Harbor/Patapsco Rivers, Anacostia/ middle Potomac, Elizabeth and James Rivers) and Poplar Island as a mid-Bay reference site. Osprey (Pandion haliaetus) diet and the transfer of contaminants from fish to osprey eggs were evaluated. The most bioaccumulative compounds (biomagnification factor > 5) included p,p'-dichlorodiphenyldichloroethylene (DDE), total polychlorinated biphenyls (PCBs), total polybrominated diphenyl ethers (PBDEs), and bromodiphenyl ether (BDE) congeners 47, 99, 100, and 154. This analysis suggested that alternative brominated flame retardants and other compounds (methoxytriclosan) are not appreciably biomagnifying. A multivariate analysis of similarity indicated that major differences in patterns among study sites were driven by PCB congeners 105, 128, 156, 170/190, and 189, and PBDE congeners 99 and 209. An integrative redundancy analysis showed that osprey eggs from Baltimore Harbor/Patapsco River and the Elizabeth River had high residues of PCBs and p,p'-DDE, with PBDEs making a substantial contribution to overall halogenated contamination on the Susquehanna and Anacostia/middle Potomac Rivers. The redundancy analysis also suggested a potential relation between PBDE residues in osprey eggs and oxidative DNA damage in nestling blood samples. The results also indicate that there is no longer a discernible relation between halogenated contaminants in osprey eggs and their reproductive success in Chesapeake Bay. Osprey populations are thriving in much of the Chesapeake, with productivity rates exceeding those required to sustain a stable population. Environ Toxicol Chem 2016;35:1560-1575. Published 2016 Wiley Periodicals Inc. on behalf of SETAC. This article is a US Government work and, as such, is in the public domain in the United States of America.

Decadal re-evaluation of contaminant exposure and productivity of ospreys (Pandion haliaetus) nesting in Chesapeake Bay Regions of Concern.

The last large-scale ecotoxicological study of ospreys (Pandion haliaetus) in Chesapeake Bay was conducted in 2000-2001 and focused on U.S. EPA-designated Regions of Concern (ROCs; Baltimore Harbor/Patapsco, Anacostia/middle Potomac, and Elizabeth Rivers). In 2011-2012, ROCs were re-evaluated to determine spatial and temporal trends in productivity and contaminants. Concentrations of p,p'-DDE were low in eggs and below the threshold associated with eggshell thinning. Eggs from the Anacostia/middle Potomac Rivers had lower total PCB concentrations in 2011 than in 2000; however, concentrations remained unchanged in Baltimore Harbor. Polybrominated diphenyl ether flame retardants declined by 40%, and five alternative brominated flame retardants were detected at low levels. Osprey productivity was adequate to sustain local populations, and there was no relation between productivity and halogenated contaminants. Our findings document continued recovery of the osprey population, declining levels of many persistent halogenated compounds, and modest evidence of genetic damage in nestlings from industrialized regions.

Investigating endocrine and physiological parameters of captive American kestrels exposed by diet to selected organophosphate flame retardants.

Organophosphate triesters are high production volume additive flame retardants (OPFRs) and plasticizers. Shown to accumulate in abiotic and biotic environmental compartments, little is known about the risks they pose. Captive adult male American kestrels (Falco sparverius) were fed the same dose (22 ng OPFR/g kestrel/d) daily (21 d) of tris(2-butoxyethyl) phosphate (TBOEP), tris(2-chloroethyl) phosphate (TCEP), tris(2-chloroisopropyl) phosphate (TCIPP), or tris(1,2-dichloro-2-propyl) phosphate (TDCIPP). Concentrations were undetected in tissues (renal, hepatic), suggesting rapid metabolism. There were no changes in glutathione status, indicators of hepatic oxidative status, or the cholinergic system (i.e., cerebrum, plasma cholinesterases; cerebrum muscarinic, nicotinic receptors). Modest changes occurred in hepatocyte integrity and function (clinical chemistry). Significant effects on plasma free triiodothyronine (FT3) concentrations occurred with exposure to TBOEP, TCEP, TCIPP, and TDCIPP; TBOEP and TCEP had additional overall effects on free thyroxine (FT4), whereas TDCIPP also influenced total thyroxine (TT4). Relative increases (32%-96%) in circulating FT3, TT3, FT4, and/or TT4 were variable with each OPFR at 7 d exposure, but limited thereafter, which was likely maintained through decreased thyroid gland activity and increased hepatic deiodinase activity. The observed physiological and endocrine effects occurred at environmentally relevant concentrations and suggest parent OPFRs or metabolites may have been present despite rapid degradation.

Assessment of mitochondrial DNA damage in little brown bats (Myotis lucifugus) collected near a mercury-contaminated river.

Historical discharges of Hg into the South River near the town of Waynesboro, VA, USA, have resulted in persistently elevated Hg concentrations in sediment, surface water, ground water, soil, and wildlife downstream of the discharge site. In the present study, we examined mercury (Hg) levels in in little brown bats (Myotis lucifugus) from this location and assessed the utility of a non-destructively collected tissue sample (wing punch) for determining mitochondrial DNA (mtDNA) damage in Hg exposed bats. Bats captured 1 and 3 km from the South River, exhibited significantly higher levels of total Hg (THg) in blood and fur than those from the reference location. We compared levels of mtDNA damage using real-time quantitative PCR (qPCR) analysis of two distinct regions of mtDNA. Genotoxicity is among the many known toxic effects of Hg, resulting from direct interactions with DNA or from oxidative damage. Because it lacks many of the protective protein structures and repair mechanisms associated with nuclear DNA, mtDNA is more sensitive to the effects of genotoxic chemicals and therefore may be a useful biomarker in chronically exposed organisms. Significantly higher levels of damage were observed in both regions of mtDNA in bats captured 3 km from the river than in controls. However, levels of mtDNA damage exhibited weak correlations with fur and blood THg levels, suggesting that other factors may play a role in the site-specific differences.

Comparative embryotoxicity of a pentabrominated diphenyl ether mixture to common terns (Sterna hirundo) and American kestrels (Falco sparverius).

Concentrations of polybrominated diphenyl ethers (PBDEs) in Forster's tern (Sterna forsteri) eggs from San Francisco Bay have been reported to range up to 63µgg(-1) lipid weight. This value exceeds the lowest-observed-adverse-effect level (1.8µgg(-1) egg wet weight; ∼32µg(-1) lipid weight) reported in an embryotoxicity study with American kestrels (Falco sparverius). As a surrogate for Forster's terns, common tern (Sterna hirundo) eggs were treated by air cell injection with corn oil vehicle (control) or a commercial penta-BDE formulation (DE-71) at nominal concentrations of 0.2, 2, and 20µgg(-1) egg. As a positive control, kestrel eggs received vehicle or 20µg DE-71g(-1) egg. In terns, there were no effects of DE-71 on embryonic survival, and pipping or hatching success; however, treated eggs hatched later (0.44d) than controls. Organ weights, organ-to-body weight ratios, and bone lengths did not differ, and histopathological observations were unremarkable. Several measures of hepatic oxidative stress in hatchling terns were not affected by DE-71, although there was some evidence of oxidative DNA damage (8-hydroxy-deoxyguanosine; 8-OH-dG). Although DE-71 did not impair pipping and hatching of kestrels, it did result in a delay in hatch, shorter humerus length, and reduced total thyroid weight. Concentrations of oxidized glutathione, reduced glutathione, thiobarbituric acid reactive substances, and 8-OH-dG in liver were greater in DE-71-treated kestrels compared to controls. Our findings suggest common tern embryos, and perhaps other tern species, are less sensitive to PBDEs than kestrel embryos.