An Assessment of the Toxicity of Pesticide Mixtures in Periphyton from Agricultural Streams to the Mayfly Neocloeon triangulifer.

Residual concentrations of pesticides are commonly found outside the intended area of application in Ontario's surface waters. Periphyton are a vital dietary component for grazing organisms in aquatic ecosystems but can also accumulate substantial levels of pesticides from the surrounding water. Consequently, grazing aquatic organisms are likely subjected to pesticide exposure through the consumption of pesticide-contaminated periphyton. The objectives of the present study were to determine if pesticides partition into periphyton in riverine environments across southern Ontario and, if so, to determine the toxicity of pesticides in periphyton when fed to the grazing mayfly Neocloeon triangulifer. Sites with low, medium, and high pesticide exposure based on historic water quality monitoring data were selected to incorporate a pesticide exposure gradient into the study design. Artificial substrate samplers were utilized to colonize periphyton in situ, which were then analyzed for the presence of approximately 500 pesticides. The results demonstrate that periphyton are capable of accumulating pesticides in agricultural streams. A novel 7-day toxicity test method was created to investigate the effects of pesticides partitioned into periphyton when fed to N. triangulifer. Periphyton collected from the field sites were fed to N. triangulifer and survival and biomass production recorded. Survival and biomass production significantly decreased when fed periphyton colonized in streams with catchments dominated by agricultural land use (p < 0.05). However, the relationship between pesticide concentration and survival or biomass production was not consistent. Using field-colonized periphyton allowed us to assess the dietary toxicity of environmentally relevant concentrations of pesticide mixtures; however, nutrition and taxonomic composition of the periphyton may vary between sites. Environ Toxicol Chem 2023;42:2143-2157. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Effects of Agricultural Stressors on Growth and an Immune Status Indicator in Wood Frog (Lithobates sylvaticus) Tadpoles and Metamorphs.

Like many amphibians, wood frog (Lithobates sylvaticus) populations have likely declined or experienced local extirpations as a result of habitat alterations. Despite this, wood frogs are still present and breeding in altered landscapes, like the agricultural Prairie Pothole Region of central Canada, and are exposed to a variety of anthropogenic impacts. As tadpoles, water contamination can have negative effects on growth, development, and immune systems. To investigate the potential effects of agricultural land use on tadpole growth and immune system stress, we used boosted regression trees to model body mass, body condition, and neutrophil to lymphocyte ratios, a measure of immune stress, against 32 variables including water quality, wetland habitat, and landscape-level measures. Developmental stage strongly influenced all 3 endpoints, and body mass was negatively influenced by higher levels of total dissolved solids (>600-700 mg/L) and at the first sign of pesticide detection (>0.01 proportion pesticides detected of those screened). While correlative, these data suggest that tadpoles developing in agricultural environments may experience survival and reproductive disadvantages if they metamorphose at smaller body sizes. Given the potential impacts this can have on adult frogs and frog populations, these results provide an impetus for further field-based investigation into the effects that pesticides, and especially total dissolved solids, may have on tadpoles. Environ Toxicol Chem 2021;40:2269-2281. © 2021 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Detecting Amphibians in Agricultural Landscapes Using Environmental DNA Reveals the Importance of Wetland Condition.

Amphibians are declining worldwide, in part because of large-scale degradation of habitat from agriculture and pervasive pathogens. Yet a common North American amphibian, the wood frog (Lithobates sylvaticus), ranges widely and persists in agricultural landscapes. Conventional survey techniques rely on visual encounters and dip-netting efforts, but detectability limits the ability to test for the effects of environmental variables on amphibian habitat suitability. We used environmental DNA to determine the presence of wood frogs and an amphibian pathogen (ranavirus) in Prairie Pothole wetlands and investigated the effects of 32 water quality, wetland habitat, and landscape-level variables on frog presence at sites representing different degrees of agricultural intensity. Several wetland variables influenced wood frog presence, the most influential being those associated with wetland productivity (i.e., nutrients), vegetation buffer width, and proportion of the surrounding landscape that is comprised of other water bodies. Wood frog presence was positively associated with higher dissolved phosphorus (>0.4 mg/L), moderate dissolved nitrogen (0.1-0.2 mg/L), lower chlorophyll a (≤15 µg/L), wider vegetation buffers (≥10 m), and more water on the landscape (≥0.25). These results highlight the effects of environmental factors at multiple scales on the presence of amphibians in this highly modified landscape-namely the importance of maintaining wetland water quality, vegetation buffers, and surrounding habitat heterogeneity. Environ Toxicol Chem 2019;38:2750-2763. © 2019 SETAC.

Developmental expression profiles and thyroidal regulation of cytokines during metamorphosis in the amphibian Xenopus laevis.

Early life-stages of amphibians rely on the innate immune system for defense against pathogens. While thyroid hormones (TH) are critical for metamorphosis and later development of the adaptive immune system, the role of TH in innate immune system development is less clear. An integral part of the innate immune response are pro-inflammatory cytokines - effector molecules that allow communication between components of the immune system. The objective of this study was to characterize the expression of key pro-inflammatory cytokines, tumor necrosis factor-α (TNFα), interleukin-1ß (IL-1ß) and interferon-γ (IFN-γ), throughout amphibian development and determine the impacts of thyroidal modulation on their expression. Xenopus laevis were sampled at various stages of development encompassing early embryogenesis to late prometamorphosis and cytokine expression was measured by real-time PCR. Expression of TNFα and IL-1ß were transient over development, increasing with developmental stage, while IFN-γ remained relatively stable. Functionally athyroid, premetamorphic tadpoles were exposed to thyroxine (0.5 and 2 µg/L) or sodium perchlorate (125 and 500 µg/L) for seven days. Tadpoles demonstrated characteristic responses of advanced development with thyroxine exposure and delayed development (although to a lesser extent) and increased thyroid gland area and follicular cell height with sodium perchlorate exposure. Exposure to thyroxine for two days resulted in decreased expression of IL-1ß in tadpole trunks. Sodium perchlorate had negligible effects on cytokine expression. Overall, these results demonstrate that cytokine transcript levels vary with stage of tadpole development but that their ontogenic regulation is not likely exclusively influenced by thyroid status. Understanding the direct and indirect effects of altered hormone status may provide insight into potential mechanisms of altered immune function during amphibian development.

Exposure to the pesticide linuron affects androgen-dependent gene expression in the three-spined stickleback (Gasterosteus aculeatus).

Previous research demonstrated that exposure to exogenous androgens and effluents with androgenic activity can induce spiggin mRNA production in the kidney of the three-spined stickleback (Gasterosteus aculeatus). In the present study, we determine whether a short-term exposure to a known antiandrogenic pesticide, linuron (LN), suppresses spiggin mRNA in male stickleback and in androgenized female stickleback. Primers were designed from previously characterized sequences for each androgen receptor (AR) isoform in stickleback, arα and arß, to assess whether these receptors are differentially regulated by androgen or antiandrogen exposure. Fish were exposed for 72 h to one of four treatments: control, LN (250 µg/L), 17α-methyltestosterone (MT, 500 ng/L), and an LN-MT mixture at those same concentrations. There was no effect of LN on spiggin and arß mRNA levels in male kidney, while levels of arα were significantly increased twofold. Exposure to LN significantly inhibited MT-induced spiggin RNA production in female kidney with no effect on expression of arα and arß. The present study is the first to demonstrate the antiandrogenic effect of LN at the transcript level and to examine androgenic/antiandrogenic responsiveness of the two ARs in the stickleback. From the present study, it was determined that measurement of spiggin RNA is a reliable and sensitive screening tool for the detection of both androgenic and antiandrogenic compounds.