Chemical mixture immunotoxicity to rainbow trout

Abstract

Several studies were conducted, and their results compiled into this thesis to explore the utility of incorporating immunotoxicological endpoints into field studies for ecotoxicological risk assessment. In the following chapters, the hazard of chemical mixture exposure to rainbow trout was assessed using a panel of recently developed immune assays. Chapter one contains a review the risk assessment process, and discusses the relevance and placement of immunological endpoints in the framework for ecotoxicological risk assessment. Fish immunotoxicological endpoints that show the greatest potential as biomarkers for ecotoxicological risk assessment are emphasized in the discussion.

Chapter two contains results from an outdoor creosote microcosm field study. In this study, several immune parameters were evaluated in rainbow trout (Oncorhynchus mykiss) after they had been exposed for 28 days in microcosms dosed initially with liquid creosote concentrations of 0, 5, 9, 17, 31, 56 and 100 ul/l. The most noticeable changes were concentration-dependent reductions in pronephros leukocyte oxidative burst and the number of slg^+ peripheral blood leukocytes. Plasma lysozyme levels were reduced, while pronephros leukocyte phagocytic activity was enhanced marginally across creosote concentrations. Blastogenesis in response to lipopolysaccharide (LPS) was slightly impaired in head kidney leukocyte cultures prepared from creosote-exposed fish, whereas blastogenesis in response to phytohaemagglutinin (PHA) and concanavalin A (ConA) was unaffected. Overall the results suggest that creosote has the potential to alter some innate immune functions in rainbow trout. Polycyclic aromatic hydrocarbons (PAHs), a major constituent of liquid creosote, are the suspected immune altering agents. The LOEC or the immune responses measured in this study was 17 ul/l using nominal creosote concentrations, representing a total PAH concentration of 611.63 ng/l in the water.

Chapter three includes results from a second outdoor microcosm study, conducted in order to re-evaluate the immunotoxic effects of liquid creosote to rainbow trout (Oncorhynchus mykiss). During this study, two separate experiments were conducted. The first experiment was designed to monitor the kinetic effects of exposure to creosote; caged fish were sampled on days 7, 14, 21, and 28 from microcosms initially dosed with 0, 3, and 10 ul/l creosote. A second experiment was designed to monitor immune parameters after 37 d of exposure in microcosms initially dosed with 0.3, 1, 3, 10, and 30 ul/l creosote. Two replicate control microcosms were included in the 37 d exposure to evaluate variability across microcosms. Pronephros leukocytes were monitored in both experiments for phagocytic activity, oxidative burst, and surface immunoglobulin-positive (Slg^+) B bell counts. Serum lysozyme activity was also measured for the 28 d kinetic experiment. During the kinetic study, oxidative burst was progressively inhibited in fish exposed to 3 and 10 ul/l creosote, returning to control levels by day 28. Phagocytic activity was initially stimulated after seven days of exposure, returning to control levels by day 28. Although control Slg^+ B cell counts were quite variable across sampling days, Slg^+ B cell counts were also elevated after seven days of exposure in creosote exposed fish. These cell numbers decreased significantly during the remainder of the study. Lysozyme activity appeared to be unaffected by creosote exposure. After the 37 d of exposure, oxidative burst was suppressed in creosote exposed fish; phagocytic activity was enhanced. No detectable change was observed for Slg^+ B cell counts. The overall results confirm that creosote has the potential to alter certain measured immune parameters. Modulation of the measured immune parameters varied with the duration of exposure. Polycyclic aromatic hydrocarbons (PAHs), a major constituent of liquid creosote, are the suspected immune-altering agents.

The final chapter contains results from a field study, used to assess whether or not exposure to sites within the Hamilton Harbour, which are known to be highly contaminated with polycyclic aromatic hydrocarbons (PAHs), heavy metals, and sewage treatment plant (STP) effluent, could affect immune function in fish. Caged rainbow trout (Oncorhynchus mykiss) were sampled after 7, 14, and 21 days of exposure from five sites within the Harbour, and a Lake Ontario reference site. The Harbour sites were selected for their proximity to either highly contaminated sediments or industrial and municipal discharges. Pronephros leukocytes from fish at the six sites were evaluated for phagocytic activity, oxidative burst, and the number of surface immunoglobulin-positive B cells. Serum lysozyme activity was also measured. Fish from the Harbour sites showed a greater reduction in pronephros leukocyte phagocytic activity over the course of the study when compared to the Lake Ontario reference fish; pronephros leukocyte oxidative burst was also reduced at two of the Harbour sites. Although B cell counts did not change throughout the duration of the study, overall counts were lower at two of the Harbour sites when compared to the Lake Ontario site. Serum lysozyme activity increased at site 4 and at the Lake Ontario reference site over the 21 days of exposure. Lysozyme activity was elevated at sites 5 and 6, when compared to Lake Ontario fish lysozyme activity, and did not change throughout the course of the study. These results indicate that several immune parameters were altered in fish during exposure at various sites within the Harbour. The role of chemical and physical stressors at these sites in the observed immunomodulation is discussed.