Changes in egg composition of American kestrels exposed to dietary polychlorinated biphenyls

Changes in the quality of eggs of birds exposed to polychlorinated biphenyls (PCBs) have been described, but have never been directly attributed to PCBs. Polychlorinated biphenyl residues in eggs have been associated with reduced reproductive success and embryonic deformities in wild birds. Egg size and composition, specifically the amount of albumen, yolk, and water in an egg, also influence the growth and viability of embryos and hatchlings, and consequently the reproductive success of birds. To deter mine whether PCB exposure of adult birds affected the size and composition of their eggs, 25 pairs of captive American kestrels (Falco sparverius) were fed a mixture of PCB-spiked (1248:1254:1260) food to give an approximate exposure of 7 mg/kg body weight/d, beginning 1 mo prior to pairing, and continuing throughout the courtship, egg-laying, and incubation periods. This dietary level in the adult female kestrels resulted in mean total PCB residues in the eggs of 34.1 microg/g wet weight (geometric mean), which is environmentally relevant. PCB residues in eggs increased with the time of female exposure to the contaminated diet and laying date. Variation in egg size within PCB clutches was significantly greater than within control clutches, although absolute egg mass and volume did not differ markedly by treatment. Only infertile eggs and only one egg per clutch were used for egg composition analysis. Yolks in the PCB-contaminated eggs were heavier, with less wet and dry albumen relative to control eggs. Water content and eggshell thickness were not significantly affected by PCB exposure. These results suggest that eggs from the PCB treatment have relatively more lipid and less protein available for embryonic development. Changes in egg composition were not associated with egg size, lay date, ambient temperature, humidity, or precipitation, which are factors known to affect these variables in bird eggs. The PCB-induced changes in egg composition described here provide insight into possible mechanisms contributing to reduced reproductive performance in wild birds exposed to PCBs.     

Reproductive abnormalities, teratogenicity, and developmental problems in American kestrels (Falco sparverius) exposed to polychlorinated biphenyls

This study found abnormalities in multiple reproductive stages in captive American kestrels (Falco sparverius) when exposed to polychlorinated biphenyls (PCBs) through dietary and in ovo exposure. American kestrels laid eggs with environmentally relevant total PCB levels (34.1 micrograms/g whole egg wet weight) when consuming PCB-spiked (Aroclor 1248:1254:1260) food (5-7 micrograms/g body weight per day) for 100 d only in 1998. In 1999, the same adults laid eggs with estimated total PCBs of 23 micrograms/g. Effects of maternal (only female exposed) and paternal (only male exposed) in ovo PCB exposure were investigated. Maternal F1 eggs contained predicted total PCB concentrations of 0.34 microgram/g. Specific abnormalities occurred more frequently during dietary F0 exposure, particularly aggressive courtship interactions, clutch abandonment, occurrences of cracked eggs, and developmental effects. Multiple developmental effects were more pronounced during than after dietary PCB exposure of adults, and although these effects were limited, nevertheless they occurred in the F1 maternal and F1 paternal pairs. However, the incidence of multiple deformities throughout the breeding season increased dramatically from 1998 (13%) to 1999 (56%) in F0 PCB-exposed pairs. Developmental abnormalities were unlikely to be attributed to the extrinsic factors of disease, genetics, or nutritional (vitamin D3) deficiencies, but rather to adverse changes in parental behavior and intrinsic factors involving altered genetic material and PCB exposure. Readily cleared PCB congeners may induce specific types of behavioral and developmental abnormalities, but persistent congeners and metabolites are likely producing (1) odd laying patterns, (2) odd laying patterns, (2) developmental effects including embryonic underdevelopment and edema, and (3) increased incidences of multiple deformities within a clutch.     

Brood patches of American kestrels altered by experimental exposure to PCBs

Captive breeding (n = 25 pairs) and nonbreeding (n = 25) American kestrels were exposed to a mixture of polychlorinated biphenyls (PCBs) (Aroclor 1248:1254:1260) through their diet of day-old cockerels. Kestrels ingested approximately 7 mg/kg body weight each day of PCBs, and this dosage resulted in environmentally relevant total PCB residues in eggs (geometric mean of 34.1 microg/g). An equal number of unexposed birds served as controls. Bare areas of skin known as brood patches function during incubation to warm eggs; therefore, brood patch size could potentially influence hatching success, or patches may be a confounding factor in the relationship between poor incubation behavior and hatching failure observed in birds in toxicological studies. Exposure to PCBs altered the size of brood patches in American kestrels. PCB-exposed male and female nonbreeders had two of three brood patches that were larger than those of control nonbreeders. Breeding males exposed to PCBs had smaller patches than controls, whereas PCB-exposed female kestrels had one larger and one smaller patch than controls. Patch sizes were not related to total PCB residue levels in eggs of exposed birds. Brood patches were not related to various incubation behaviors or hatching success in either control or PCB-exposed kestrels.     

Association between PCBs and lower embryonic weight in black-crowned night herons in San Francisco Bay

Reproductive problems, including congenital malformations, reduced hatching success, and decreased survival of hatchlings, have been observed in colonial-nesting water birds at the San Francisco Bay National Wildlife Refuge (SFBNWR). Twenty-four black-crowned night heron (Nycticorax nycticorax) eggs were collected from SFBNWR in 1983. Twelve of these were collected from separate nests when late-stage embryos were pipping, and an additional egg was randomly collected from each nest for organochlorine analysis. Overt anomalies and skeletal defects were not apparent. Embryonic weights (with partially absorbed yolk sacs removed) were 15% lower (p less than 0.05) in SFBNWR embryos compared to control embryos from the Patuxent Wildlife Research Center (PWRC). Crown-rump length and femur length were shorter for SFBNWR embryos. The geometric mean polychlorinated biphenyl (PCB) concentration in SFBNWR eggs was 4.1 ppm wet weight, with a range of 0.8-52.0 ppm. A negative correlation (r = -0.61; p less than 0.05) existed between embryonic weight and log-transformed PCB residues in whole eggs collected from the same nest at SFBNWR, suggesting a possible impact of PCBs on embryonic growth. A correlation with embryonic weight did not occur for DDE [1,1-dichloro-2,2-bis(p-chlorophenyl) ethylene] residues. Liver microsomal aryl hydrocarbon hydroxylase activity was neither significantly elevated nor correlated with PCB, DDE, or PCB plus DDE log-transformed residues. It is unknown whether the apparent association between PCBs and lower weight is persistent through hatching.     

Effects of dietary PCB exposure on adrenocortical function in captive American kestrels (Falco sparverius)

We experimentally examined the effects of dietary exposure to polychlorinated biphenyls (PCBs) on adrenocortical function in American kestrels (Falco sparverius). Nine captive male American kestrels previously exposed to a PCB mixture (Aroclor1248:1254:1260; 1:1:1) in their diet were subjected to a standardized capture, handling and restraint protocol designed to produce an increase in circulating corticosterone. A similar protocol has been applied to a wide range of avian species and was used here to evaluate the response of PCB-exposed and control kestrels to a defined physical stressor. Both baseline and stress-induced corticosterone levels were significantly lower in PCB-exposed birds when compared with control birds of the same age. PCB-exposed birds exhibited significantly lower corticosterone levels during the corticosterone response when compared with control birds, independent of body condition. Furthermore, baseline corticosterone concentrations exhibited a hormetic response characterized by an inverted U-shaped dose response in relation to total PCB liver burden. These results support several recent studies which report decreased levels of circulating corticosterone in PCB-exposed wild birds. The results presented here provide the first evidence that exposure to an environmentally relevant level of PCBs (approximately 10 mg/kg body weight) can impair the corticosterone stress response in kestrels, potentially increasing the susceptibility of birds to environmental stressors such as severe weather and predatory and human disturbance.     

Polychlorinated biphenyls,dioxins, furans,and organochlorine pesticides in spotted sandpiper eggs from the upper Hudson River basin,New York

In 2004, spotted sandpipers (Actitis macularia) were studied on the Hudson River near Fort Edward south to New Baltimore, NY and on two river drainages that flow into the Hudson River. Concentrations of 28 organochlorine pesticides, 160 polychlorinated biphenyl (PCB) congeners, and 17 dioxin and furan (PCDD-F) congeners were quantified in eggs collected on and off the Hudson River. The pattern of organochlorine pesticides and PCDD-F congeners did not differ significantly between eggs collected on and off the Hudson River. In contrast, the pattern of PCB congeners differed significantly between the Hudson River and other rivers. Total PCBs were significantly greater in eggs from the Hudson River (geometric mean = 9.1 μg PCBs/g wet weight) than from the other two rivers (0.6 and 0.6 μg PCBs/g wet weight). Seven of 35 (20%) eggs exceeded 20 μg PCBs/g wet weight, the estimated threshold for reduced hatching in tree swallows (Tachycineta bicolor) and some raptor species; the maximum concentration was 72.3 μg PCBs/g wet weight. Models that predicted nest survival and egg success (the proportion of eggs hatching in a clutch if at least one egg hatched) as functions of contaminant levels were poorly distinguished from models that presumed no such associations. While small sample size could have contributed to the inability to distinguish among contaminant and no toxicant models, we cannot rule out the possibility that contaminant concentrations on the Hudson River were not sufficiently high to demonstrate a relationship between contaminant concentrations and reproductive success.     

Relationship between PCB accumulation and reproductive output in conditioned oysters Crassostrea virginica fed a contaminated algal diet

Because of their resistance to environmental degradation, polychlorinated biphenyls (PCBs) are among the most widespread environmental contaminants. PCBs have high bioaccumulation potential and may affect a number of biological/physiological processes including disruption of the endocrine system function, lipid metabolism and reproduction. The objective of this study was to test whether conditioning sexually immature oysters with PCB-contaminated algal diets affects their subsequent reproductive success. Sexually immature oysters were conditioned in individual containers and fed daily with 0.7 g algal paste containing 0, 0.35 or 3.5 microg PCBs for up to 76 days. The impact of sediment load on PCB accumulation in oysters was also tested by exposure of a subset of oysters to clay particles. Oysters in different treatments were sampled 56 days after conditioning with PCB-contaminated algal diets to determine uptake and distribution of PCBs in gonad, digestive gland, mantle, gill and muscle, and the presence of gametes. Tissues from oysters exposed to PCBs alone for 56 days were also analyzed for lipid and fatty acid composition. Following 61 and 76 days of PCB exposure, remaining oysters from all treatments were induced to spawn via thermal stimulation. Non-spawned oysters were stripped to determine if sexual products were present. Oysters exposed to PCBs alone and PCBs plus clay particles showed similar trends in PCB accumulation, but concentrations were generally lower in the latter. PCB accumulation in oysters increased with an increase in algal-associated PCB concentrations, varied with organ types and was correlated with lipid content. The highest PCB concentration was in the gonad and the lowest in gill and muscle. PCB-153, -138/158, -118, -90/101 and -149 were the dominant congeners in all tissue compartments, except the muscle where PCB-28/31 was the dominant congener pair. PCB exposure appeared to impair both lipid metabolism and reproductive success. Although PCB exposure produced only slight changes in the lipid class composition in the oysters, decreases in phospholipids were observed in gonad, muscle and mantle of oysters exposed to 3.5 microg PCBs daily for 56 days. After 56 days of conditioning with PCB-sorbed algal paste, no well-developed mature eggs were observed in any of the oysters examined for the presence of sexual products. No significant difference was noted in reproductive success (production of spawned females and males) between sediment-treated and non-treated groups after 76 days of PCB exposure compared to controls, PCB-exposed oysters produced fewer spawned females, but no dose-dependent relationship was observed.     

Effects of in ovo exposure to PCBs 126 and 77 on mortality, deformities and post-hatch immune function in chickens

In laboratory experiments, planar PCBs produce immune organ atrophy in chicken embryos. To study the immunotoxic effects of PCBs in birds, the coplanar congeners 3,3',4,4',5-pentachlorobiphenyl (PCB 126) and 3,3',4,4'-tetrachlorobiphenyl (PCB 77) were injected into the air cell of fertile white leghorn chicken eggs before incubation at doses of 0.25 and 0.5 ng/g egg PCB 126 and 0.64 ng/g egg PCB 77. Mortality and deformities were assessed during incubation of the eggs, and immune function was analyzed post-hatch using phytohemagglutinin (PHA) skin test for T-cell mediated immunity, antibody titers to sheep red blood cells (SRBC), mitogenesis of peripheral blood lymphocytes, and immune organ mass and cellularity. Exposure to 0.25 ng/g PCB 126 elevated mortality (61% and 69%) and deformities (31% and 32%), three or more times higher than controls. Two-fold suppression of antibody titers was observed in 28 day old chicks exposed to PCB 126 or PCB 77. No consistent alterations in PHA skin response or lymphocyte proliferation were observed. In 14 day old chicks in experiment two, PCB 126 decreased thymus and bursa cellularity by 33% and 35%, respectively. Immune organ atrophy was transient, recovering to control levels by 42 days of age. These experiments demonstrate that PCB 126 and 77 suppress antibody responses in juvenile chickens following an in ovo PCB exposure. Results reinforce the need for measuring multiple immune endpoints to detect immunotoxicity.     

Long-term food-exposure of zebrafish to PCB mixtures mimicking some environmental situations induces ovary pathology and impairs reproduction ability

Although the use of polychlorinated biphenyls (PCBs) has been banned for several decades, they are still present in the environment and are occasionally mechanically released from sediment or transferred through the trophic chain. Field analyses have established correlations between exposure to PCBs and alterations in fish physiology including reproductive function. Experimental exposures have been mainly performed using dioxin-like PCBs or other congeners at very high concentrations. However, these studies are often difficult to relate to real-life conditions. In the present study, we performed a life-cycle exposure using zebrafish model and mixtures representative of some environmental situations in terms of doses, composition and containing mainly non dioxin-like congeners. Exposure was performed through diet which is the main contamination route in the field. We demonstrated a bioaccumulation of PCBs in males and females as well as a maternal transfer to the eggs. Survival, growth and organ size were similar for all conditions. Several reproductive traits were altered after exposure to a PCB-contaminated diet, including a reduction in the number of fertilized eggs per spawn as well as an increase of the number of poorly fertilized spawns. This latter observation was found irrespective of the sex of contaminated fish. This is related to modifications of ovary histology revealing a decrease of maturing follicles and an increase of atretic follicles in the ovaries of females exposed to PCBs. These results indicate that exposure to PCBs mixtures mimicking some environmental situations, including mainly non dioxin-like congeners, can lead to a dramatic reduction in the number of offspring produced by a female over a lifetime. This is of great concern for wild species living under natural conditions.     

Avian transgenerational reproductive toxicity test with in ovo exposure

Ecological risk assessment of environmental pollutants requires effective laboratory assays and extrapolation of the resultant data to wild species. Because avian reproductive disorder and accumulation of persistent compounds in wild birds and their eggs have long been observed in polluted regions, we have developed an assay for investigating whether pollutants accumulated in eggs impair the reproduction of the exposed birds and the survival of the next generation using the Japanese quail. A typical estrogenic compound, diethylstilbestrol (DES), dissolved in olive oil was injected into the air-chamber of fertilized eggs on day 10 of incubation. After sexual maturation of hatched chicks, we made pairs of male and female quails following an observation period of egg production and colleted their eggs. The collected eggs were incubated and checked for the fertility and hatchability, and then the hatchlings were raised and observed in growth for 3 weeks. A dosage of 5 ng/g per egg of DES caused eggshell thinning in eggs laid by exposed females and reduction in eggshell strength. DES also induced shortening of the left oviduct and unexpected development of the right oviduct, while testis weight was reduced symmetrically. The ability of quail pairs to produce offspring was significantly diminished by exposure of females to DES independently of exposure of males, which mainly arose from production of abnormal and inviable eggs. Fertility of normal-shelled eggs and hatchability of fertilized eggs were unchanged regardless of treatments. External morphological abnormalities, which were mostly unopened toes of the foot, were frequently observed in hatchlings from exposed males independently of exposure of females. Additionally, we attempted to extrapolate the experimental results to the northern bobwhite and to predict population trends for quails in a polluted habitat using a population projection model composed of a combination of a Leslie matrix and the logistic equation. In the event of accumulation of an estrogenic compound equivalent to a dosage of 5 ng/g DES in quail eggs, the average population size was predicted to decrease by 20.2% after 1 year, to approximately half after 4 years, and to a fifth after 14 years. When observed weakening of individuals and the risk of egg breakage are taken into consideration, the decline in population was further accelerated. The proposed assay appears to be suitable not only for assessing adverse effects of chemicals on avian reproduction but for population projection of affected wild birds.     

Maternal transfer of trace elements in leatherback turtles (Dermochelys coriacea) of French Guiana

In sea turtles, parental investment is limited to the nutrients and energy invested in eggs that will support embryonic development. Leatherback females have the largest clutches with the biggest eggs of the sea turtles and the highest reproductive output in reptiles. The migration between foraging sites and nesting beaches also represents high energy expenditure. The toxicokinetic of pollutants in the tissues is thus expected to vary during those periods but there is a lack of information in reptiles. Concentrations of essential (Copper, Zinc, Selenium) and non-essentials elements (Cadmium, Lead, Mercury) were determined in blood (n=78) and eggs (n=76) of 46 free-ranging leatherback females collected in French Guiana. Maternal transfer to eggs and relationships between blood and eggs concentrations during the nesting season were investigated. All trace elements were detectable in both tissues. Levels of toxic metals were lower than essential elements likely due to the high pelagic nature of leatherbacks that seems to limit exposure to toxic elements. Significant relationships between blood and egg concentrations were observed for Se and Cd. Se could have an important role in embryonic development of leatherback turtles and Cd transfer could be linked to similar carrier proteins as Se. Finally, as multiple clutches were sampled from each female, trends in trace elements were investigated along the nesting season. No change was observed in eggs but changes were recorded in blood concentrations of Cu. Cu level decreased while blood Pb levels increased through the nesting season. The high demand on the body during the breeding season seems to affect blood Cu concentrations. Calcium requirement for egg production with concomitant Pb mobilization could explain the increase in blood Pb concentrations along the nesting season.     

Developmental Toxicity of PCB 126 (3,3',4,4',5-Pentachlorobiphenyl) in Nestling American Kestrels (Falco sparverius)

Planar PCB congeners are embryotoxic and teratogenic to birdsincluding American kestrels. The developmental toxicity of 3,3',4,4',5-pentachlorobiphenyl(PCB 126) was studied in the posthatching kestrel as a modelfor the eagle. Nestlings were dosed orally for 10 days with5 µl/g body weight of corn oil (controls) or the planarPCB 126 at concentrations of 50, 250, or 1000 ng/g body weight.Dosing with 50 ng/g of PCB 126 resulted in a hepatic concentrationof 156 ng/g wet weight, liver enlargement and mild coagulativenecrosis, over 10-fold increases in hepatic microsomal ethoxyresorufin-O-dealkylaseand benzyloxyresorufin-O-dealkylase, and approximately a 5-foldincrease in methoxyresorufin-O-dealkylase. At this dose, mildto moderate lymphoid depletion of the spleen was apparent, aswere decreased follicle size and content of the thyroid. At250 ng/g, concentration of PCB 126 in the liver was 380 ng/gwith increasing multifocal coagulative necrosis, decreased bonegrowth, decreased spleen weight with lymphocyte depletion ofthe spleen and bursa, and degenerative lesions of the thyroid.At 1000 ng/g, the liver concentration was 1098 ng/g, accompaniedby decreased bursa weight, decreased hepatic thiol concentration,and increased plasma enzyme activities (ALT, AST, and LDH-L)in addition to the previous effects. Highly significant positivecorrelations were noted between liver concentrations of PCB126 and the ratio of oxidized to reduced glutathone. These findingsindicate that nestling kestrels are more susceptible to PCB126 toxicity than adults, but less sensitive than embryos, andthat planar PCBs are of potential hazard to nestling birds.     

Changes in polychlorinated biphenyl (PCB) exposure in tree swallows (Tachycineta bicolor) nesting along the Sheboygan River,WI, USA

Exposure to polychlorinated biphenyls (PCBs) in tree swallow (Tachycineta bicolor) eggs on the Sheboygan River, Wisconsin in the 1990s was higher at sites downstream (geometric means = 3.33–8.69 μg/g wet wt.) of the putative PCB source in Sheboygan Falls, Wisconsin than it was above the source (1.24 μg/g) with the exposure declining as the distance downstream of the source increased. A similar pattern of declining exposure was present in the 2010s as well. Although exposure to PCBs in eggs along the Sheboygan River at sites downstream of Sheboygan Falls has declined by ~60 % since the mid-1990s (8.69 down to 3.27 μg/g) there still seems to be residual pockets of contamination that are exposing some individuals (~25 %) to PCB contamination, similar to exposure found in the 1990s. The exposure patterns in eggs and nestlings among sites, and the changes between the two decades, are further validated by accumulation rate information.     

BEHAVIOR OF FREE-RANGING AND CAPTIVE AMERICAN KESTRELS UNDER ELECTROMAGNETIC FIELDS

Wild birds, particularly raptors, commonly use electrical transmission structures for nesting, perching, hunting, and roosting. Consequently, birds are exposed to electromagnetic fields (EMFs). The amount of time that wild reproducing American kestrels (Falco sparverius) were exposed to EMFs was determined, and the effects of EMFs on the behavior of captive reproducing kestrels were examined. Wild kestrels were exposed to EMFs for 25% to 75% of the observed time. On a 24-h basis, estimated EMF exposure of wild kestrels ranged from 71% during courtship, to 90% during incubation, similar to that experienced by captive kestrels (88% of a 24-h period). Additionally, captive kestrels were exposed to EMF levels experienced by wild kestrels nesting under 735-kV power lines. Captive EMF females were more active, more alert, and perched on the pen roof more frequently than control females during courtship. EMF females preened and rested less often during brood rearing. EMF male kestrels were more active than control males during courtship, and more alert during incubation. Increased activity of kestrels during courtship may be linked to changes in corticosterone, but likely not melatonin. Observed behavioral changes were unlikely to directly result in the better growth of nestlings and fledging success, or poorer hatching success, of the EMF group, as previously reported. Behavioral changes of captive EMF kestrels may be observed in wild kestrels.     

Behavior of free-ranging and captive American kestrels under electromagnetic fields

Wild birds, particularly raptors, commonly use electrical transmission structures for nesting, perching, hunting, and roosting. Consequently, birds are exposed to electromagnetic fields (EMFs). The amount of time that wild reproducing American kestrels (Falco sparverius) were exposed to EMFs was determined, and the effects of EMFs on the behavior of captive reproducing kestrels were examined. Wild kestrels were exposed to EMFs for 25% to 75% of the observed time. On a 24-h basis, estimated EMF exposure of wild kestrels ranged from 71% during courtship, to 90% during incubation, similar to that experienced by captive kestrels (88% of a 24-h period). Additionally, captive kestrels were exposed to EMF levels experienced by wild kestrels nesting under 735-kV power lines. Captive EMF females were more active, more alert, and perched on the pen roof more frequently than control females during courtship. EMF females preened and rested less often during brood rearing. EMF male kestrels were more active than control males during courtship, and more alert during incubation. Increased activity of kestrels during courtship may be linked to changes in corticosterone, but likely not melatonin. Observed behavioral changes were unlikely to directly result in the better growth of nestlings and fledging success, or poorer hatching success, of the EMF group, as previously reported. Behavioral changes of captive EMF kestrels may be observed in wild kestrels.     

Use of epidemiology and clinical toxicology to determine human risk in regulating polychlorinated biphenyls in the food supply

Polychlorinated biphenyls (PCBs) became a national problem in 1971 when several accidental contaminations of foods were reported. Extensive efforts were successfully undertaken by FDA to reduce the residues of PCBs in food. However, the PCB levels in several species of freshwater fish have raised concern about PCB residues from environmental contamination. This concern prompted a reassessment of the human risk involved in consumption of such fish. The best evidence that a chemical may produce adverse health effects in humans is provided by adequate epidemiologic data confirmed or supplemented by data from valid animal tests. Traditionally, where the regulatory agencies have used results of animal toxicology experiments to evaluate hazard and predict hypothetical safety for humans, “safety factors” such as 1 to 10 or 1 to 100 have been used. The size of the safety factor and the potential exposure to a chemical are established by properly informed scientific judgment. More recent efforts have involved use of a combination of human and animal data and a variety of mathematical models to determine risk. The human epidemiology data and the animal toxicity data of PCB exposure are reviewed, as well as risk assessment in general. Specific examples of risk assessment are presented in which animal data are extrapolated to humans, based on several levels of human exposure to PCBs in fish.     

Effects of PCB 126 on primary immune organs and thymocyte apoptosis in chicken embryos

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and polychlorinated biphenyl (PCB) 126 produce thymic atrophy and immunosuppression. This study explored the hypothesis that the thymic atrophy produced by developmental exposure to PCB 126 is associated with an increase in apoptotic thymocytes at the end of incubation in chicken embryos. Eggs were injected via the air cell with PCB 126 (0.05, 0.13, 0.32, 0.64, and 0.80 ng/g egg) on d 0 of incubation, and tissues were collected on d 20. Controls included noninjected and vehicle-injected (sunflower oil) eggs. Thymocytes were cultured for 6 h and analyzed by flow cytometry for decreased DNA content (propidium iodide staining) and cell size (forward scatter), which indicate apoptosis. PCB 126 induced dose-dependent mortality with an LD50 of 1.01 ng/g and lowest-observed-effect concentration (LOEC) of 0.32 ng/g. Teratogenic effects commonly associated with TCDD and planar PCBs, including cranial and foot deformities and subcutaneous edema, tended to increase with dose of PCB 126. PCB 126 reduced thymus mass by approximately 20% at 0.64 and 0.8 ng/g, the number of viable thymocytes by approximately 20-24% at and above 0.13 ng/g, and the number of bursal lymphoid cells by 57% at 0.64 ng/g. The percentage of apoptotic thymocytes increased with dose, reaching levels 2 times greater than controls at 0.8 ng/g. Electrophoresis of low-molecular-weight DNA from thymocytes of all doses demonstrated fragments in multiples of 180 bp. This DNA laddering is a hallmark of apoptosis. At all doses, thymocytes exhibited caspase-3 activation, another indicator of apoptosis. The results of this experiment supported the hypothesis that the thymic atrophy produced by developmental exposure to PCB 126 in chicken embryos is associated with an increase in apoptotic thymocytes on embryonic d 20.     

Antibody-Mediated Immunotoxicity in American Kestrels (Falco sparverius) Exposed to Polychlorinated Biphenyls

Antibody-mediated immune function in adult and recently fledged (30 to 33 d old) American kestrels (Falco sparverius) was examined in birds exposed directly, or only in ovo, to polychlorinated biphenyls (PCBs). In 1998, 9 mature male and 9 female kestrels were fed PCBs, whereas 9 females and 10 males served as controls. A mixture of Aroclors 1248:1254:1260 suspended in safflower oil was injected into the kestrels' food items, while in control diets only the same volume of oil was added. The dosage of PCBs was approximately 7 mg/kg kestrel/d, beginning in March 1998 and continuing for 120 d. In 1998, the antibody-mediated immune response was stimulated by immunization and booster vaccinations of the kestrels using a nonpathogenic antigen, dinitrophenol-keyhole limpet hemocyanin (DNP-KLH). In 1999, offspring from three treatment groups based upon maternal exposure to PCBs were similarly tested for their antibody response. None of these mothers was vaccinated with DNP-KLH the previous year. The maternal groups were: (1) exposed to PCBs in 1998 for 120 d, (2) exposed in ovo in 1998 (i.e., mothers were produced by PCB-exposed parents), or (3) unexposed to PCBs. Serum antibody levels were determined using an enzyme-linked immunosorbent assay (ELISA). In 1998, PCB-exposed adult females had a significantly higher antibody response than did controls, whereas adult males exposed to PCBs had significantly suppressed antibody production. For the nestlings produced in 1999, maternal treatment significantly affected antibody response. Generally, the antibody response in the nestlings was much lower than that seen in adult kestrels. Yet both male and female offspring from mothers that had been fed PCBs the previous year had significantly higher postbooster anti-DNP-KLH titers than control and in ovo-exposed maternal groups, thus mimicking the response seen in the adult females the previous year. These sex-specific responses in PCB-exposed birds provide further evidence of the endocrine-disrupting behavior of PCBs. Both suppression and stimulation of the antibody response are undesirable because this indicates that the immune system is not able to respond normally to challenges by infectious or other disease-causing agents.     

PCDDs, PCDFs, PCBs, OC pesticides and mercury in fish and osprey eggs from Willamette River, Oregon (1993, 2001 and 2006) with calculated biomagnification factors

The osprey (Pandion haliaetus) population nesting along the main stem Willamette River and lower Santiam River was first studied to evaluate contaminants and reproductive rates in 1993 when 78 occupied nests were present. By 2001, the population increased to 234 occupied nests, a 13.7% annual rate of population increase. A sample egg was collected from each of a series of nests along the Upper River (river mile 55–187) in 1993, 2001 and 2006 to evaluate trends of persistent contaminants (organochlorine [OC] pesticides, polychlorinated biphenyls [PCBs], polychlorinated dibenzo-p-dioxins [PCDDs], and polychlorinated dibenzofurans [PCDFs]). Nearly all OC pesticide residues decreased significantly, e.g., p, p′-DDE (DDE) from 2,350 to 1,353 to 210 μg/kg wet weight (ww). PCBs followed a similar pattern over time, e.g., ΣPCBs 688 to 245 to 182 μg/kg ww, while PCDDs and PCDFs showed a more precipitous decline (often 85–95%) between 1993 and 2001, with no egg analyses warranted in 2006. During 2001–2002, sample osprey eggs were also collected from nests at three Headwater Reservoirs and two lower reaches (Newberg Pool and Tidal Portland) of the Willamette River, as well as the lower portion of the Santiam River to evaluate spatial residue patterns. Significant differences were seldom detected among the different sampling areas for OC pesticides (probably due to small sample sizes), although higher concentrations were often seen in the lower reaches, e.g., DDE 901 μg/kg ww (Headwater Reservoirs), 1,353 (Upper River), 1,384 (Newberg Pool) and 2,676 (Tidal Portland). PCB congener concentrations in eggs were usually higher in the Tidal Portland reach than at other locations and often significantly higher than at the Headwater Reservoirs or Upper River. Mercury (first analyzed in eggs in 2001), PCDDs and PCDFs were extremely low in 2001/2002 with no significant spatial patterns. Whole fish composite samples of largescale sucker (Catastomus macrocheilus) and northern pikeminnow (Ptychocheilus oregonensis), which account for about 90% of the biomass in the diet of this osprey population, were also collected from the Willamette River in 1993 and 2001 and analyzed for the same contaminants as osprey eggs. Contaminant residues in fish from the Upper River decreased between 1993 and 2001, paralleling findings for osprey eggs. Likewise, spatial patterns for fish residues paralleled findings for osprey eggs from the different reaches in 2001. A second empirical estimate of biomagnification factors (BMFs) from fish to osprey eggs for OC pesticides, PCBs, PCDDs and PCDFs (ww and lipid weight [lw] basis) was calculated based on residue data collected in 2001. The two independent BMF estimates (1993 and 2001) for each contaminant from the Upper River provide a measure of consistency, e.g., DDE (ww) 87 and 79, (lw) 103 and 112; ΣPCBs (ww) 11 and 8.4, (lw) 13 and 12. Mercury did not biomagnify from fish to osprey eggs (BMF = 0.60). Legacy contaminants investigated had limited (perhaps only DDE), if any, effects on reproductive success of the increasing osprey population nesting along the Willamette River by 2001.     

Antibody-mediated immunotoxicity in American kestrels (Falco sparverius) exposed to polychlorinated biphenyls

Antibody-mediated immune function in adult and recently fledged (30 to 33 d old) American kestrels (Falco sparverius) was examined in birds exposed directly, or only in ovo, to polychlorinated biphenyls (PCBs). In 1998, 9 mature male and 9 female kestrels were fed PCBs, whereas 9 females and 10 males served as controls. A mixture of Aroclors 1248:1254:1260 suspended in safflower oil was injected into the kestrels' food items, while in control diets only the same volume of oil was added. The dosage of PCBs was approximately 7 mg/kg kestrel/d, beginning in March 1998 and continuing for 120 d. In 1998, the antibody-mediated immune response was stimulated by immunization and booster vaccinations of the kestrels using a nonpathogenic antigen, dinitrophenol-keyhole limpet hemocyanin (DNP-KLH). In 1999, offspring from three treatment groups based upon maternal exposure to PCBs were similarly tested for their antibody response. None of these mothers was vaccinated with DNP-KLH the previous year. The maternal groups were: (1) exposed to PCBs in 1998 for 120 d, (2) exposed in ovo in 1998 (i.e., mothers were produced by PCB-exposed parents), or (3) unexposed to PCBs. Serum antibody levels were determined using an enzyme-linked immunosorbent assay (ELISA). In 1998, PCB-exposed adult females had a significantly higher antibody response than did controls, whereas adult males exposed to PCBs had significantly suppressed antibody production. For the nestlings produced in 1999, maternal treatment significantly affected antibody response. Generally, the antibody response in the nestlings was much lower than that seen in adult kestrels. Yet both male and female offspring from mothers that had been fed PCBs the previous year had significantly higher postbooster anti-DNP-KLH titers than control and in ovo-exposed maternal groups, thus mimicking the response seen in the adult females the previous year. These sex-specific responses in PCB-exposed birds provide further evidence of the endocrine-disrupting behavior of PCBs. Both suppression and stimulation of the antibody response are undesirable because this indicates that the immune system is not able to respond normally to challenges by infectious or other disease-causing agents.