Search

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.

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.

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 r 2  = 0.212, P = 0.001) than did the host species (t = 4.762, P < 0.001; r 2  = 0.070, P = 0.001) or their interaction (t = 3.449; r 2  = 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)., (Published by Elsevier Ltd.)

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., (Published by Elsevier Ltd.)

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., (Copyright © 2021 US Geological Survey, The Author(s). Published by Elsevier Ltd.. All rights reserved.)

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 17β-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 17β-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 17β-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., (© 2021 SETAC. This article has been contributed to by US Government employees and their work is in the public domain in the USA.)

Short chain chlorinated paraffins (SCCPs) are complex mixtures of polychlorinated n-alkanes, shown to bioaccumulate but with unknown effects in wild birds. The present study examined development-related effects of SCCPs on captive American kestrels (Falco sparverius) treated in ovo on embryonic day (ED) 5 by injection with technical Chloroparaffin® (C 10-13 , 55.5% Cl) at environmentally relevant nominal (measured) concentrations of 10 (10), 50 (29) or 100 (97) ng ΣSCCP/g egg ww, and artificially incubated until hatching (ED27-ED29). The SCCP concentrations measured in the yolk sacs of the hatchling kestrels bracketed concentrations reported in the eggs of wild birds. Uptake and deposition of these SCCPs differed between male and female hatchlings, with only males showing differences in SCCP concentrations, being highest in the high-dose males than each of the other male groups. Embryonic exposure to SCCPs suppressed glandular total thyroxine (TT4) (20-33%) and reduced circulating triiodothyronine (TT3) (37-40%) in male hatchlings only when compared to control males, but had no effect on glandular TT3 or circulating TT4 in male or female kestrels. Histological assessments of thyroid glands showed that both sexes experienced significant structural changes indicative of gland activation. These thyroid glandular changes and the variations in SCCP concentrations were related to circulating TT3 in female hatchlings. Hepatic deiodinase enzyme (D1, D2) activities were stable and no SCCP-related changes were observed in hatching success, hatchling size, or immune organ size. However, several of the thyroid function indicators were correlated with hatchling size and smaller bursas and spleens, possibly indirectly through SCCP-induced changes in thyroid function. Because changes in thyroid function were evident at concentrations measured in wild bird eggs, similar changes may occur in wild nestlings. The potential impact of these changes on thyroid-mediated growth and survival in wild birds requires further investigation., (Crown Copyright © 2020. Published by Elsevier Ltd. All rights reserved.)

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., (Crown Copyright © 2019. Published by Elsevier Inc. All rights reserved.)

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., (Crown Copyright © 2019. Published by Elsevier Inc. All rights reserved.)

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.

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., (© 2018 SETAC.)

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., (Crown Copyright © 2018. Published by Elsevier B.V. All rights reserved.)

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.

We investigated the effects of the androgenic growth promoter 17β-trenbolone (17βTB) 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 17βTB treated F0 and F1 males. In F0 and F1 females, testosterone was suppressed by 17βTB, 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 17βTB. 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 17βTB in Japanese quail but the response was dependent on sex, developmental stage at initiation of exposure, and dose., (Published by Oxford University Press on behalf of the Society of Toxicology 2017. This work is written by US Government employees and is in the public domain in the US.)

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, 17β-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)., (© 2017 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals, Inc. on behalf of Society of Environmental Toxicology & Chemistry (SETAC).)

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)., (© 2016 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals, Inc. on behalf of Society of Environmental Toxicology & Chemistry (SETAC).)

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., (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.)

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., (Published by Elsevier Ltd.)

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.

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.

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., (Published by Elsevier Ltd.)

Despite extensive studies of methylmercury (MeHg) toxicity in birds, molecular effects on birds are poorly characterized. To improve our understanding of toxicity pathways and identify novel indicators of avian exposure to Hg, the authors investigated genomic changes, glutathione status, and oxidative status indicators in liver from laughing gull (Larus atricilla) hatchlings that were exposed in ovo to MeHg (0.05-1.6 µg/g). Genes involved in the transsulfuration pathway, iron transport and storage, thyroid-hormone related processes, and cellular respiration were identified by suppression subtractive hybridization as differentially expressed. Quantitative polymerase chain reaction (qPCR) identified statistically significant effects of Hg on cytochrome C oxidase subunits I and II, transferrin, and methionine adenosyltransferase RNA expression. Glutathione-S-transferase activity and protein-bound sulfhydryl levels decreased, whereas glucose-6-phosphate dehydrogenase activity increased dose-dependently. Total sulfhydryl concentrations were significantly lower at 0.4 µg/g Hg than in controls. Together, these endpoints provided some evidence of compensatory effects, but little indication of oxidative damage at the tested doses, and suggest that sequestration of Hg through various pathways may be important for minimizing toxicity in laughing gulls. This is the first study to describe the genomic response of an avian species to Hg. Laughing gulls are among the less sensitive avian species with regard to Hg toxicity, and their ability to prevent hepatic oxidative stress may be important for surviving levels of MeHg exposures at which other species succumb., (Copyright © 2012 SETAC.)

We surveyed four Chesapeake Bay tributaries for skin and liver tumors in brown bullhead (Ameiurus nebulosus). We focused on the South River, where the highest skin tumor prevalence (53%) in the Bay watershed had been reported. The objectives were to 1) compare tumor prevalence with nearby rivers (Severn and Rhode) and a more remote river (Choptank); 2) investigate associations between tumor prevalence and polynuclear aromatic hydrocarbons (PAHs) and alkylating agents; and 3) statistically analyze Chesapeake Bay bullhead tumor data from 1992 through 2008. All four South River collections exhibited high skin tumor prevalence (19% to 58%), whereas skin tumor prevalence was 2%, 10%, and 52% in the three Severn collections; 0% and 2% in the Choptank collections; and 5.6% in the Rhode collection. Liver tumor prevalence was 0% to 6% in all but one South River collection (20%) and 0% to 6% in the three other rivers. In a subset of samples, PAH-like biliary metabolites and (32)P-DNA adducts were used as biomarkers of exposure and response to polycyclic aromatic compounds (PACs). Adducts from alkylating agents were detected as O6-methyl-2'-deoxyguanosine (O6Me-dG) and O6-ethyl-2'-deoxyguanosine (O6Et-dG) modified DNA. Bullheads from the contaminated Anacostia River were used as a positive control for DNA adducts. (32)P-DNA adduct concentrations were significantly higher in Anacostia bullhead livers compared with the other rivers. We identified alkyl DNA adducts in bullhead livers from the South and Anacostia, but not the Choptank. Neither the PAH-like bile metabolite data, sediment PAH data, nor the DNA adduct data suggest an association between liver or skin tumor prevalence and exposure to PACs or alkylating agents in the South, Choptank, Severn, or Rhode rivers. Logistic regression analysis of the Chesapeake Bay database revealed that sex and length were significant covariates for liver tumors and length was a significant covariate for skin tumors., (Published by Elsevier B.V.)

The health benefits of regular consumption of fish and seafood have been espoused for many years. However, fish are also a potential source of environmental contaminants that have well known adverse effects on human health. We investigated the consumption risks for largemouth bass (Micropterus salmoides; n = 104) and striped mullet (Mugil cephalus; n = 170), two commonly harvested and consumed fish species inhabiting fresh and estuarine waters in northwest Florida. Skinless fillets were analyzed for total mercury, inorganic arsenic, polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDD/F), polychlorinated biphenyls (PCBs), and organochlorine pesticides. Contaminant levels were compared to screening values (SV) calculated using U.S. Environmental Protection Agency (EPA) recommendations for establishing consumption advisories. Largemouth bass were found to contain high levels of total mercury at all sampling locations (0.37-0.89 ug/g) and one location exhibited elevated total PCBs (39.4 ng/g). All of the samples exceeded Florida fish consumption advisory trigger levels for total mercury and one location exceeded the U.S. EPA SV for total PCBs. As a result of the high mercury levels, the non-cancer health risks (hazard index-HI) for bass were above 1 for all locations. Striped mullet from several locations with known point sources contained elevated levels of PCBs (overall range 3.4-59.3 ng/g). However, total mercury levels in mullet were low. Eight of the 16 mullet sampling locations exceeded the U.S. EPA SV for total PCBs and two locations exceeded an HI of 1 due to elevated PCBs. Despite the elevated levels of total PCBs in some samples, only two locations exceeded the acceptable cancer risk range and therefore cancer health risks from consumption of bass and mullet were determined to be low at most sampling locations., (Published by Elsevier Ltd.)

Polymerase chain reaction (PCR)-based methods to determine the sex of birds are well established and have seen few modifications since they were first introduced in the 1990s. Although these methods allowed for sex determination in species that were previously difficult to analyse, they were not conducive to high-throughput analysis because of the laboriousness of DNA extraction and gel electrophoresis. We developed a high-throughput real-time PCR-based method for analysis of sex in birds, which uses noninvasive sample collection and avoids DNA extraction and gel electrophoresis., (Published 2010. This article is a US Government Work and is in the public domain in the USA.)

In the present study, we identified and characterized an inducible heat shock protein 70 (HSP70) from the midge Chironomus dilutus and investigated the transcriptional profile of the gene under baseline and environmentally stressful conditions. Using real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR), we observed increased expression of CD-HSP70-1 in response to both heat shock and copper stress. We also investigated the expression of this gene during midge development. All C. dilutus developmental stages expressed CD-HSP70-1 under normal conditions, although at extremely low levels. Phylogenetic analysis of the amino acid sequence demonstrated distinct clustering of this gene with inducible HSP70s from other insect species.

The southeastern United States, and in particular the coastal areas along the Gulf of Mexico (Gulf Coast) in Florida, experience some of the highest levels of mercury deposition in the country. Although the State of Florida's coastal border is among the longest in the United States, and the State has issued fish consumption advisories due to mercury on multiple fish species, few data have been systematically collected to assess mercury levels in the human population of the state or to assess the efficacy of the consumption advisories. Because of the generally high rate of seafood consumption among coastal populations, the human population in the Florida Panhandle, near Pensacola, FL is potentially exposed to elevated levels of mercury. In the present study, we analyzed hair mercury levels in women of child-bearing age (16-49 years) who had resided near Pensacola, FL for at least 1 year. We also surveyed the fish consumption practices of the cohort and evaluated awareness of the Florida Fish Consumption Advisory. Hair mercury levels were significantly higher in women who consumed fish within the 30 days prior to sampling (p<0.05) and in those women who were unaware of the consumption advisory (p<0.05). Only 31% of the women reported knowledge of the consumption advisory and pregnant women exhibited lower awareness of the advisory than non-pregnant women. The data suggest that public health interventions such as education and fish advisories have not reached the majority of women in the counties surrounding Pensacola who are most at risk from consumption of fish with high levels of mercury.

The Escambia Wood Treating Company (ETC) Superfund site, Pensacola, FL, is contaminated with polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDD/F), benzo(a)pyrene, lead and arsenic from pentachlorophenol (PCP), creosote, and other compounds used to treat utility poles and foundation pilings. Although ETC's operations ceased in 1982, soils in the areas surrounding the facility continue to exhibit elevated levels of contaminants attributable to ETC operations. In July 2000, individuals who may have been affected by contamination from the ETC site, including current and former residents and former workers and their household members were invited to participate in a study, which included a health and exposure history and routine blood analysis. We also conducted a toxicological health evaluation of a subset of these eligible workers/residents by analyzing serum levels of 17 PCDD/F congeners. Members of the ETC cohort exhibited elevated serum PCDD/F relative to the general population, and congener profiles in members of the cohort reflected patterns commonly observed in persons exposed to PCP. Hypertension prevalence in the cohort was found to correlate with PCDD/F levels, although no other significant relationships were identified with monitored health indices.

We conducted a screening level assessment of contaminants in blue crabs (Callinectes sapidus) and oysters (Crassostrea virginica) from bays and bayous in the Pensacola, FL area. Tissue samples were analyzed for 17 dioxins/furans, 12 dioxin-like PCB (DL-PCBs) congeners, mercury, and various metals. Contaminant levels were compared to screening values (SV) calculated using U.S. EPA recommendations for establishing consumption advisories. All sampling locations exceeded the SV (0.098pgg(-1)) for dioxins/furans/DL-PCBs, based on a Florida-specific consumption rate (46gday(-1)). Arsenic (inorganic), mercury, cadmium, and zinc levels exceeded SVs in samples from select locations, and with the exception of mercury, these locations were generally downstream of known contaminated areas. We also assessed potential human health risks from consumption of these species. Risks to human health were greatest from consumption of crab hepatopancreas, suggesting that consumption of hepatopancreas, whether directly or indirectly, from crabs collected anywhere in the Pensacola Bay region should be avoided.

Two complementary deoxyribonucleic acid (cDNA) clones encoding 2 different 70-kDa heat shock proteins (HSPs) were isolated from the prawn Macrobrachium rosenbergii. The cDNA clones were 2448 and 2173 bp in length and contained 1950- and 1734-bp open reading frames (ORFs), respectively. The ORFs encoded 649- and 577-amino acid polypeptides, which were named Mar-HSC70 and Mar-HSP70, respectively, according to the sequence identities with other known HSC70s and HSP70s and based on their inducibility in response to heat shock stress (at 35 degrees C). Genomic DNA sequence analysis revealed no introns in either gene. The major structural differences between the 2 proteins were a 60-amino acid segment and a 14-amino acid segment present in the N-terminal and C-terminal, respectively, of Mar-HSC70 that were not found in Mar-HSP70. Northern blotting and semiquantitative reverse transcription-polymerase chain reaction analyses indicated that the Mar-HSP70 gene was expressed under heat shock (35 degrees C) stress in a non-tissue-specific manner. In contrast, Mar-HSC70 messenger ribonucleic acid was constitutively expressed in every tissue except muscle, and its expression in response to heat shock (at 35 degrees C) changed only in muscle.

In the present study, we investigated the induction of HSP70 proteins in Chironomus tentans in response to copper exposure. Larvae were exposed for 24 h to 0, 0.25, 0.5, 1.0 and 2.0 mgl(-1) copper, concentrations that were previously shown to cause growth reduction or mortality in 10 d bioassays. The induction of HSP70 was evaluated by Western blotting, and we observed a significant increase in HSP70 proteins over controls at all tested concentrations. The results suggest that short-term laboratory exposures to chronically toxic but acutely sublethal copper concentrations induce HSP70 proteins in C. tentans larvae.

In the present study we carried out the isolation and characterization of an HSC70 gene from two midges, Chironomus tentans and C. yoshimatsui. The HSC70 cDNAs are approximately 2424 (C. tentans) and 2464 bp (C. yoshimatsui) long, and contain 1950 and 1956 bp open reading frames, respectively. Analysis of genomic DNA revealed the presence of two introns in these genes. The 5' untranslated regions of the HSC70 genes are adenosine-rich, a feature found in inducible HSP70 genes. The nucleotide and amino acid sequences exhibit high identity with cytosolic HSC70s from other Dipterans. Northern hybridization indicated that HSC70 is expressed at all developmental stages, from embryo to adult, and Southern hybridization confirmed the presence of multiple HSP70 genes in Chironomus.

Stress proteins (heat shock proteins, HSPs) have been proposed as general biomarkers for environmental monitoring. In the present study, we evaluated the environmental stress-burden on the aquatic midge Chironomus yoshimatsui using hsp70 expression. Larvae collected from streams receiving polluted runoff (field strain) were resistant to the organophosphorus insecticide, fenitrothion (F), and the synthetic pyrethroid, ethofenprox (E), whereas a strain originally collected from an unpolluted area (susceptible strain) showed low resistance to insecticide exposure. To examine the expression of an HSP70 gene in C. yoshimatsui, an hsp70 cDNA probe was prepared using RNA obtained from the field strain larvae and used for Northern blot analyses. The expression of this HSP70 gene in larvae collected from two field sites in May about 1 week after insecticide spraying in the fields was 2.3 (p = 0.018) to 3.3 fold higher than that in the susceptible strain and was also 4.6 and 1.4 (p = 0.033) fold higher than those collected in November 3 months after the cessation of insecticide spraying. In order to identify potential inducers of the HSP70 gene of the field strain, larvae of the susceptible strain were exposed to F or E for 24 h and hsp70 mRNA levels determined. Exposures to F at 0.4 microg/L and E at 1.1 microg/L increased hsp70 mRNA levels 2.7 (p = 0.049) and 4.4 (p = 0.043) fold over controls, respectively. These results suggest that larvae collected from polluted areas are burdened by environmental stressors and the tested insecticides are potential inducers of HSP70. The results also support the suggestion that HSP70 gene expression is a sensitive indicator of low level (nonlethal) exposures to certain insecticides., (Copyright 2002 Wiley Periodicals, Inc. J Biochem Mol Toxicol 16:10–17, 2002; DOI 10.1002/jbt.10018)

Stormwater treatment ponds receive elevated levels of metals from urban runoff, but the effects of these pollutants on organisms residing in the ponds are unknown. We investigated the accumulation of Cu, Zn, and Pb by macroinvertebrates collected from stormwater treatment ponds in Maryland serving commercial, highway, residential and open-space watersheds, and determined whether watershed land-use classification influences metal concentrations in macroinvertebrates, sediments, and water. Three types of invertebrate samples were analyzed--molluscs, odonates, and composite. Zn concentrations in odonates from ponds draining watersheds with commercial development (mean = 113.82 micrograms g-1) were significantly higher than concentrations in the other land-use categories. Similarly, Cu levels in odonates from commercial ponds (mean = 27.12 micrograms g-1) were significantly higher than from highway (mean = 20.23 micrograms g-1) and open space (mean = 17.79 micrograms g-1) ponds. However, metal concentrations in sediments and water did not differ significantly among land-uses. The results suggest that despite the high variation in ambient metal concentrations within each land-use category, macroinvertebrates in ponds serving commercial watersheds accumulate higher levels of Cu and Zn. The levels of Cu, Zn, and Pb in invertebrates from all ponds were less than dietary concentrations considered toxic to fish.