Judit E.G. Smits

Assessing the exposure risk and impacts of pharmaceuticals in the environment on individuals and ecosystems

The use of human and veterinary pharmaceuticals is increasing. Over the past decade, there has been a proliferation of research into potential environmental impacts of pharmaceuticals in the environment. A Royal Society-supported seminar brought together experts from diverse scientific fields to discuss the risks posed by pharmaceuticals to wildlife. Recent analytical advances have revealed that pharmaceuticals are entering habitats via water, sewage, manure and animal carcases, and dispersing through food chains. Pharmaceuticals are designed to alter physiology at low doses and so can be particularly potent contaminants. The near extinction of Asian vultures following exposure to diclofenac is the key example where exposure to a pharmaceutical caused a population-level impact on non-target wildlife. However, more subtle changes to behaviour and physiology are rarely studied and poorly understood. Grand challenges for the future include developing more realistic exposure assessments for wildlife, assessing the impacts of mixtures of pharmaceuticals in combination with other environmental stressors and estimating the risks from pharmaceutical manufacturing and usage in developing countries. We concluded that an integration of diverse approaches is required to predict ‘unexpected’ risks; specifically, ecologically relevant, often long-term and non-lethal, consequences of pharmaceuticals in the environment for wildlife and ecosystems.

Compromised metamorphosis and thyroid hormone changes in wood frogs (Lithobates sylvaticus) raised on reclaimed wetlands on the Athabasca oil sands.

The wet landscape approach to oil sands tailings reclamation in the Athabasca Oil Sands region involves creating wetlands from fluid tailings in mined-out pits. We measured time to metamorphosis, thyroid hormone status, and detoxification enzyme (EROD) induction in Wood frog (Lithobates sylvaticus) tadpoles raised on reclaimed oil sands wetlands of different ages [young (≤ 7 yr) vs. old (> 7 yr)] and compared data with tadpoles raised on reference (control) wetlands. Metamorphosis was delayed or never occurred in tadpoles raised in young tailings; those exposed to older tailings developed similarly to those in reference wetlands. Thyroid hormone disruption likely played an important role in the metamorphosis delay as the T3:T4 ratio was lowest in tadpoles raised in young, tailings-affected wetlands. Our findings suggest tailings wetlands become less toxic with age, and that these amphibians will be able to complete their life cycle in tailing wetlands that have sufficiently detoxified with age.

Pathological, immunological and biochemical markers of subchronic arsenic toxicity in rats

Subchronic exposure to arsenic in rats was investigated to identify sensitive indicators of subclinical toxicity in rats. Immunological, pathological, and biochemical bioindicators were examined in rats exposed to arsenic in their drinking water. Juvenile male Wistar rats were allocated to four treatment groups receiving 0, 0.4, 4, and 40 ppm of arsenite in drinking water for 18 wks. Besides daily monitoring for clinical signs of adverse health effects, clinical biochemistry, B‐cell‐mediated and innate immune responses, plus gross, and histopathology were examined. In vitro tests of oxidative damage to basic cellular constituents, lipids, proteins, and nucleic acids, were measured using thiobarbituric acid reacting substances (TBARS) assays, protein carbonyl formation, and 8‐hydroxydeoxyguanosine (8‐OHdG), respectively. Clinical changes in the rats were limited to decreased feed and water intake in the high‐ (40 ppm) dose group (P < 0.05), however, growth rate was not affected. Serum biochemical changes occurred in blood urea nitrogen, K+, Cl−, and alanine aminotransferase (ALT) from arsenic exposure. Immunotoxicity was evident through a dose‐dependent suppression of the secondary antibody‐mediated response to a T‐cell‐dependent antigen, keyhole limpet hemocyanin (KLH). Histopathology of the liver revealed marked fatty infiltration and vacuolization particularly evident in periacinar hepatocytes. This pattern of toxicopathology in the high‐exposure group may be related to the significantly higher (P < 0.05) oxidative stress, demonstrated through lipid peroxidation (TBARS assay) in the rats exposed to 40 ppm arsenite. The present study revealed that young, growing rats exposed to arsenic for 18 wks tolerated exposures up to 4 ppm. At higher doses, there was evidence of hepatotoxicity, humoral immunity was compromised, and an adverse effect on hepatic organelle and cell membranes was evident through a dose dependent increased in oxidative stress.

Stress and Reproductive Hormones in Grizzly Bears Reflect Nutritional Benefits and Social Consequences of a Salmon Foraging Niche

Physiological indicators of social and nutritional stress can provide insight into the responses of species to changes in food availability. In coastal British Columbia, Canada, grizzly bears evolved with spawning salmon as an abundant but spatially and temporally constrained food source. Recent and dramatic declines in salmon might have negative consequences on bear health and ultimately fitness. To examine broadly the chronic endocrine effects of a salmon niche, we compared cortisol, progesterone, and testosterone levels in hair from salmon-eating bears from coastal BC (n = 75) with the levels in a reference population from interior BC lacking access to salmon (n = 42). As predicted, testosterone was higher in coastal bears of both sexes relative to interior bears, possibly reflecting higher social density on the coast mediated by salmon availability. We also investigated associations between the amount of salmon individual bears consumed (as measured by stable isotope analysis) and cortisol and testosterone in hair. Also as predicted, cortisol decreased with increasing dietary salmon and was higher after a year of low dietary salmon than after a year of high dietary salmon. These findings at two spatial scales suggest that coastal bears might experience nutritional or social stress in response to on-going salmon declines, providing novel insights into the effects of resource availability on fitness-related physiology.

Detection and drivers of exposure and effects of pharmaceuticals in higher vertebrates

Pharmaceuticals are highly bioactive compounds now known to be widespread environmental contaminants. However, research regarding exposure and possible effects in non-target higher vertebrate wildlife remains scarce. The fate and behaviour of most pharmaceuticals entering our environment via numerous pathways remain poorly characterized, and hence our conception and understanding of the risks posed to wild animals is equally constrained. The recent decimation of Asian vulture populations owing to a pharmaceutical (diclofenac) offers a notable example, because the exposure route (livestock carcasses) and the acute toxicity observed were completely unexpected. This case not only highlights the need for further research, but also the wider requirement for more considered and comprehensive ‘ecopharmacovigilance’. We discuss known and potential high risk sources and pathways in terrestrial and freshwater ecosystems where pharmaceutical exposure in higher vertebrate wildlife, principally birds and mammals, may occur. We examine whether approaches taken within existing surveillance schemes (that commonly target established classes of persistent or bioaccumulative contaminants) and the risk assessment approaches currently used for pesticides are relevant to pharmaceuticals, and we highlight where new approaches may be required to assess pharmaceutical-related risk.

Avian wildlife as sentinels of ecosystem health.

Birds have been widely used as sentinels of ecosystem health reflecting changes in habitat quality, increased incidence of disease, and exposure to and effects of chemical contaminants. Numerous studies addressing these issues focus on the breeding period, since hormonal, behavioural, reproductive, and developmental aspects of the health can be observed over a relatively short time-span. Many body systems within individuals are tightly integrated and interdependent, and can be affected by contaminant chemicals, disease, and habitat changes in complex ways. Animals higher in the food web will reflect cumulative effects of multiple stressors. Such features make birds ideal indicators for assessing environmental health in areas of environmental concern. Five case studies are presented, highlighting the use of different species which have provided insight into ecosystem sustainability, including (i) the consequences of anthropogenic disturbances of sagebrush habitat on the greater northern sage grouse Centrocercus urophasianus; (ii) the high prevalence of disease in very specific passerine species in the Canary Islands closely paralleling deterioration of formerly productive desert habitat and ensuing interspecific stressors; (iii) fractures, abnormal bone structure, and associated biochemical aberrations in nestling storks exposed to acidic tailings mud from a dyke rupture at an iron pyrite mine near Sevilla, Spain; (iv) newly presented data demonstrating biochemical changes in nestling peregrine falcons Falco peregrinus and associations with exposure to major chemical classes in the Great Lakes Basin of Canada; and (v) the variability in responses of tree swallows Tachycineta bicolor to contaminants, biological and meteorological challenges when breeding in the Athabasca oil sands.

Detoxification, endocrine, and immune responses of tree swallow nestlings naturally exposed to air contaminants from the Alberta oil sands

Changes in environmental and wildlife health from contaminants in tailings water on the Canadian oil sands have been well-studied; however, effects of air contaminants on wildlife health have not. A field study was conducted to assess biological costs of natural exposure to oil sands-related air emissions on birds. Nest boxes for tree swallows (Tachycineta bicolor) were erected at two sites; within 5 km of active oil sands mining and extraction, and ≥ 60 km south, at one reference site. Passive air monitors were deployed at the nest boxes to measure nitrogen dioxide, sulfur dioxide, ozone, volatile organic compounds, and polycyclic aromatic hydrocarbons (PAHs). Nestlings were examined at day 9 post hatching to assess T cell function and morphometry. At day 14 post hatching, a subset of nestlings was euthanized to measure detoxification enzymes, endocrine changes, and histological alterations of immune organs. Except for ozone, all air contaminants were higher at the two oil sands sites than the reference site (up to 5-fold). Adult birds had similar reproductive performance among sites (p>0.05). Nestlings from industrial sites showed higher hepatic ethoxyresorufin O-dealkylase (EROD) induction (p<0.0001) with lower relative hepatic mass (p=0.0001), a smaller T cell response to the phytohemagglutinin skin test (p=0.007), and smaller bursae of Fabricius (p<0.02); a low sample size for one site indicating lower body condition scores (p=0.01) at day 14 warrants cautious interpretation. There were no differences among nestlings for feather corticosterone (p>0.6), and no histological alterations in the spleen or bursa of Fabricius (p>0.05). This is the first report examining toxicological responses in wild birds exposed to air contaminants from industrial activity in the oil sands. It is also the first time that small, individual air contaminant monitors have been used to determine local contaminant levels in ambient air around nest boxes of wild birds.

Treating chronic arsenic toxicity with high selenium lentil diets

Arsenic (As) toxicity causes serious health problems in humans, especially in the Indo-Gangetic plains and mountainous areas of China. Selenium (Se), an essential micronutrient is a potential mitigator of As toxicity due to its antioxidant and antagonistic properties. Selenium is seriously deficient in soils world-wide but is present at high, yet non-toxic levels in the great plains of North America. We evaluate the potential of dietary Se in counteracting chronic As toxicity in rats through serum biochemistry, blood glutathione levels, immunotoxicity (antibody response), liver peroxidative stress, thyroid response and As levels in tissues and excreta. To achieve this, we compare diets based on high-Se Saskatchewan (SK) lentils versus low-Se lentils from United States. Rats drank control (0ppm As) or As (40ppm As) water while consuming SK lentils (0.3ppm Se) or northwestern USA lentils (<0.01ppm Se) diets for 14weeks. Rats on high Se diets had higher glutathione levels regardless of As exposure, recovered antibody responses in As-exposed group, higher fecal and urinary As excretion and lower renal As residues. Selenium deficiency caused greater hepatic peroxidative damage in the As exposed animals. Thyroid hormones, triiodothyronine (T3) and thyroxine (T4), were not different. After 14weeks of As exposure, health indicators in rats improved in response to the high Se lentil diets. Our results indicate that high Se lentils have a potential to mitigate As toxicity in laboratory mammals, which we hope will translate into benefits for As exposed humans.

The effects of oil sands wetlands on wood frogs (Rana sylvatica)

Extraction of crude oil from oil sand produces solid (sand) and liquid (water with suspended fine particles) tailings materials, called oil sands process-affected materials (OSPM). These waste materials are stored on the mine site due to a “zero discharge” policy and must be reclaimed when operations end. The liquid tailings materials are known to contain naphthenic acids and polycyclic aromatic hydrocarbons and have high pH and salinity. One method of reclamation is the “wet landscape” approach, which involves using oil sands tailings materials to form wetlands that would mimic natural wetland ecological function. This study investigated the effects of wetlands formed with oil sands tailings materials on the survival and growth of wood frog (Rana sylvatica) larvae. In spring 2007, in-situ caging studies were completed in 14 wetlands that were of four different classes; young or old, reference or reclaimed. Tadpole survival was different between types of wetlands, with young tailings-affected wetlands (≤7 years old) having 41.5%, 62.6%, and 54.7% higher tadpole mortality than old tailings-affected (>7 years old), young reference, and old reference wetlands, respectively. Since old wetlands created from OSPM showed effects on tadpoles similar to those of reference wetlands, which had markedly lower toxicity than young tailings-affected wetlands, we provide evidence that wetlands, at least 7 years old, can sustain amphibian life.

Heavily hunted wolves have higher stress and reproductive steroids than wolves with lower hunting pressure

Summary 1. Human-caused harassment and mortality (e.g. hunting) affects many aspects of wildlife population dynamics and social structure. Little is known, however, about the social and physiological effects of hunting, which might provide valuable insights into the mechanisms by which wildlife respond to human-caused mortality. 2. To investigate physiological consequences of hunting, we measured stress and reproductive hormones in hair, which reflect endocrine activity during hair growth. Applying this novel approach, we compared steroid hormone levels in hair of wolves (Canis lupus) living in Canada’s tundra–taiga (n = 103) that experience heavy rates of hunting with those in the northern boreal forest (n = 45) where hunting pressure is substantially lower. 3. The hair samples revealed that progesterone was higher in tundra–taiga wolves, possibly reflecting increased reproductive effort and social disruption in response to human-related mortality. Tundra–taiga wolves also had higher testosterone and cortisol levels, which may reflect social instability. 4. To control for habitat differences, we also measured cortisol in an out-group of boreal forest wolves (n = 30) that were killed as part of a control programme. Cortisol was higher in the boreal out-group than in our study population from the northern boreal forest. 5. Overall, our findings support the social and physiological consequences of human-caused mortality. Long-term implications of altered physiological responses should be considered in management and conservations strategies.