Kerry Wautier

Dietary accumulation of hexabromocyclododecane diastereoisomers in juvenile rainbow trout (Oncorhynchus mykiss) I: bioaccumulation parameters and evidence of bioisomerization.

Juvenile rainbow trout (Oncorhynchus mykiss) were exposed to three diastereoisomers (alpha, beta, gamma) of hexabromocyclododecane (C12H18Br6) via their diet for 56 d followed by 112 d of untreated food to examine bioaccumulation parameters and test the hypothesis of in vivo bioisomerization. Four groups of 70 fish were used in the study. Three groups were exposed to food fortified with known concentrations of an individual diastereoisomer, while a fourth group were fed unfortified food. Bioaccumulation of the gamma-diastereoisomer was linear during the uptake phase, while the alpha- and beta-diastereoisomers were found to increase exponentially with respective doubling times of 8.2 and 17.1 d. Both the beta- and the gamma-diastereoisomers followed a first-order depuration kinetics with calculated half-lives of 157 +/- 71 and 144 +/- 60 d (+/-1 x standard error), respectively. The biomagnification factor (BMF) for the alpha-diastereoisomer (BMF = 9.2) was two times greater than the beta-diastereoisomer (BMF = 4.3); the large BMF for the beta-diastereoisomer is consistent with this diastereoisomer dominating higher-trophic-level organisms. Although the BMF of the beta-diastereoisomer suggests that it will biomagnify, it is rarely detected in environmental samples because it is present in small quantities in commercial mixtures. Results from these studies also provide evidence of bioisomerization of the beta- and gamma-diastereoisomers. Most importantly, the alpha-diastereoisomer that was recalcitrant to bioisomerization by juvenile rainbow trout in this study and known to be the dominant diastereosiomer in fish was bioformed from both the beta- and the gamma-diastereoisomers. To our knowledge, this is the first report of bioisomerization of a halogenated organic pollutant in biota.

Developmental effects of bioaccumulated selenium in eggs and larvae of two salmonid species

Elevated concentrations of Se have been detected in cold, flowing water habitats near uranium and coal mines in Canada. Fish from these systems have concentrations of Se in their tissues that exceed toxic effect thresholds that have been established for warm-water fishes. However, the applicability of toxic effect thresholds and guidelines to cold water, lotic habitats is a matter of contention in the literature since most cases of Se toxicosis have been documented in standing, warm-water systems. To examine the possibility of impaired reproduction in wild rainbow trout (Oncorhynchus mykiss) and brook trout (Salvelinusfontinalis) near coal mining activity in the northeastern slopes region of Alberta, Canada, spawn from both species were collected from exposure and reference sites. Gametes were fertilized in the laboratory, reared to the swim-up stage, and examined for deformities. A significant relationship was observed for rainbow trout between the amount of Se in eggs and the incidence of developmental abnormalities, specifically craniofacial defects, skeletal deformities, and edema. These associations approximate exponential functions with probabilities that 15% of the population would be affected occurring between 8.8 and 10.5 microg Se per gram of wet egg weight, based on probit analysis. These relationships are similar to those described for centrarchids inhabiting a seleniferous warm-water lake. No such relationships were established for brook trout.

Biochemical and histopathological effects in pearl dace (Margariscus margarita) chronically exposed to a synthetic estrogen in a whole lake experiment.

Potential effects of exposure to the synthetic estrogen 17alpha-ethynylestradiol (EE2) were examined in several species of fish from a lake experimentally treated with environmentally relevant concentrations of the contaminant. Ethynylestradiol was added to Lake 260, a small Precambrian shield lake at the Experimental Lakes Area in northwestern Ontario, Canada, from May to October of 2001, 2002, and 2003. Mean concentrations of EE2 in epilimnetic waters ranged between 4.5 and 8.1 ng/L during the three years, with overall means of 6.1 (+/- 2.8), 5.0 (+/- 1.8), and 4.8 (+/- 1.0) ng/L for the three years, respectively. Male and female pearl dace (Margariscus margarita) captured after EE2 additions began contained up to 4,000-fold higher concentrations of the egg yolk precursor vitellogenin than fish captured from the same lake before the EE2 additions or when compared to fish from reference lakes. Edema in the ovaries, inhibited development of testicular tissue, intersex, and histopathological kidney lesions were all evident in fish exposed to EE2. Some indications that EE2 exposure affected in vitro steroidogenic capacity of the ovaries and the testes existed, although results were not always consistent between years. Pearl dace abundance was similar in the lake treated with EE2 and the reference lake. A trend exists toward a reduced overall population of pearl dace from the treated and reference lakes, as do indications that young-of-the-year size classes are less abundant in the EE2-treated lake. Biochemical and histopathological impacts observed in fish exposed to EE2 in this study have not yet been linked to clear population level impacts in pearl dace. Monitoring of these populations is ongoing.

Waterborne ethynylestradiol induces vitellogenin and alters metallothionein expression in lake trout (Salvelinus namaycush)

Estrogenic contaminants isolated from waters receiving sewage treatment plant effluents are known to induce the egg yolk precursor vitellogenin (VTG) in male fish. Levels of the metal binding protein metallothionein (MT) have also been shown to be affected by estrogens in fish. It has been postulated that MT declines in estrogen exposed fish to facilitate transfer of the essential metal Zn to cellular components required for VTG synthesis. To examine the changes in MT and VTG concentrations in fish exposed to an estrogen contaminant, lake trout (Salvelinus namaycush) were exposed to waterborne ethynylestradiol at 0, 4, 40 or 400 ng/l(-1) for 21 days. Blood and tissues were collected after 21 days of exposure to measure circulating levels of VTG as well as MT concentrations in liver and kidney. VTG increased in male and female fish from all three exposure groups compared to control fish. MT in liver significantly decreased in males and females compared to the controls, in the two highest exposures. MT in kidney was significantly higher in both sexes of fish exposed to the two highest concentrations of ethynylestradiol. These data are supportive of a relationship between estrogen exposure and the regulation of MT. Further studies to examine the specific links between estrogen exposure, VTG induction and regulation of essential metals like Zn are required.

A modified mercury saturation assay for measuring metallothionein.

A thorough re-evaluation of assay procedures for a mercury displacement analytical method for metallothionein (MT) described in 1993, was conducted using an analytical standard of rabbit liver MT-II and MT determinations, in liver, kidney, gill and intestine from lake trout (Salvelinus namaycush). These studies indicated that: (1) loss of MT by adherence to polypropylene material occurs with the dilution series; (2) use of a calibration curve using a rabbit liver MT-II analytical standard, is superior to the dilution series for determining unknown MT concentrations; (3) Hg incubation time requirements are less in 20% TCA than in 10% TCA; and (4) employing both 20% TCA and heat at 95 degrees C for 5 min to denature non MT proteins results in the most accurate MT spike recoveries, or recoveries that did not differ significantly from those closest to 100%, in liver, kidney, gill and intestine. Calibration curves with an average r(2) of 0.9998 for 69 trials over different days, while passing through the origin, provided evidence of negligible non-specific binding of mercury. Use of water or saline for preparing tissue homogenates, produced slight, but significant, differences in MT estimates in the intestine and kidney. Comparisons between the original and the modified methods were made on liver, kidney and gill from control, sham-injected and Cd-treated lake trout. Differences in MT estimates obtained between the two methods depended upon the tissue. MT estimates in liver by the modified method were 9, 13, and 10% higher in control, sham-injected and Cd-treated groups, respectively, than those resulting from the original assay. MT estimates in gill by the modified method, were 3.2-fold, 4.0-fold and 69% higher in control, sham-injected, and Cd-treated groups, respectively. Corresponding values for kidney were 37, 75, and 57% higher. Because percent recoveries of MT-II spikes were virtually 100%, in all four tissues, using the modified method, MT estimates obtained using these modifications were more accurate than those obtained using the procedures of the original assay. Application of the modified assay to fish tissues in a Canadian biomonitoring program for metal mining, demonstrated that in 1 week an experienced technologist can complete 90 MT analyses of liver and kidney and 75 analyses of gill and intestine.

Thyroid axis disruption in juvenile brown trout (Salmo trutta) exposed to the flame retardant β-tetrabromoethylcyclohexane (β-TBECH) via the diet.

Tetrabromoethylcyclohexane (TBECH) is an additive brominated flame retardant used in domestic and industrial applications. It has been detected in wildlife, and there is early evidence that it is an endocrine disruptor. Whereas other brominated flame retardants with similar physicochemical properties have been shown to disrupt the thyroid axis, no such evaluation has been conducted for TBECH. To elucidate this, juvenile brown trout (Salmo trutta) were fed either a control diet or diets containing low, medium, or high doses of β-TBECH, the isomer most frequently detected in wildlife, for 56 days (uptake phase) followed by a control diet for an additional 77 days (depuration phase). Eight fish per treatment were lethally sampled on uptake days 7, 14, 21, 35, 49, and 56 and on depuration days 7, 21, 35, 49, and 77 to assess fish condition, circulating free and total triiodothyronine and thyroxine, and thyroid epithelial cell height. Although there was no effect on condition factor, there was a significant reduction in total plasma thyroxine in the high dose group and a significant increase in mean thyroid epithelial cell height in the low, medium, and high dose groups during the uptake phase, whereas there were no differences in the depuration phase. These results indicate that β-TBECH may modulate the thyroid axis in fish at environmentally relevant concentrations.

Effect of Different Levels of Fine-Sediment Loading on the Escapement Success of Rainbow Trout Fry from Artificial Redds

Abstract Increased sedimentation from logging operations can affect water flow over salmonid redds, potentially impairing embryonic development in incubating eggs and fry. Effects may result from reduced oxygen delivery or waste removal or from physical entrapment of fry by sediment cap formation. A system of artificial redds was used to examine the effects of varying loads of mixed fine sediments on the escapement success of rainbow trout Oncorhynchus mykiss fry. Replicates of 80 eyed rainbow trout eggs were seeded in redds loaded with low (11.8%), medium (21.2%), high (28.6%), or no (0%) additional fine sediments. Interstitial oxygen saturation was determined in each chamber throughout incubation and emergence. Total percent emergence, residual yolk sac remaining, prevalence of deformities, and condition factor were determined in emergent fry. Fry began to emerge at 507.6 degree-days postfertilization, and overall emergence was more than 70% in all treatment levels. Emergence rate patterns approximated ...

An Assessment of the Potential for Selenium to Impair Reproduction in Bull Trout, Salvelinus confluentus, from an Area of Active Coal Mining

Selenium (Se) is an essential nutrient, but in higher concentrations can reduce recruitment in fish populations by increasing rates of deformities during early development. Recent work has identified elevated levels of Se in water and biota collected downstream from coal mining activity in Albertas northeast slopes region. We also recently identified increased incidence of terata and edema in rainbow trout and brook trout with elevated tissue Se from this area. However, there is currently no information regarding the potential for Se to contribute to declining stocks of bull trout, a species of concern in the area. The present study provides an assessment of the potential for Se to contribute to low recruitment in bull trout downstream from coal mining activity. Non-destructive muscle biopsy sampling and a sensitive atomic fluorescence analysis technique are used to determine Se. Results indicate that most bull trout (>90%) captured immediately downstream from coal mining activity in the region have concentrations of Se that would be expected to impair recruitment. Additional work is required to determine the extent of Ses contribution to low recruitment in bull trout.