Microplastics may induce food dilution and endocrine disrupting effects in fathead minnows (Pimephales promelas), and decrease offspring quality.

Microplastics are a complex environmental contaminant that have been reported to cause a variety of impacts, although the mechanism of these impacts remains unclear. Many studies have investigated either sub-organismal or apical endpoints, while very few have attempted to integrate and link endpoints seen at multiple levels of organization. Here, we exposed fathead minnows to microplastics for their entire lifecycle, from the egg stage through to reproduction, and raised a subset of the offspring in clean water. We show that both preconsumer and environmentally sourced microplastics impact adult growth, lipid storage, and external colouration, suggesting a potential food dilution effect. Environmentally sourced microplastics, but not preconsumer microplastics, had further endocrine disrupting impacts on the parental generation and their offspring in the low concentration treatments such that egg production began later, eggs were less viable, and the offspring had higher rates of malformation. Low dose effects are a typical dose-response for endocrine disrupting contaminants. These results suggest that microplastic exposure, at concentrations relevant to what is being found in the environment, has potential implications for forage fish populations. Our findings also highlight the importance of using an integrative approach to understanding the mechanisms behind how and why microplastics impact organisms.

Impacts to Larval Fathead Minnows Vary between Preconsumer and Environmental Microplastics.

Microplastics are a complex suite of contaminants varying in size, shape, polymer, and associated chemicals and are sometimes referred to as a "multiple stressor." Still, the majority of studies testing hypotheses about their effects use commercially bought microplastics of a uniform size, shape, and type. We investigated the effects of polyethylene and polypropylene microplastics purchased as preproduction pellets (referred to as "preconsumer") and a mixture of polyethylene and polypropylene collected from the environment (environmental microplastic). Embryo-stage fathead minnows were exposed to either the physical plastic particles and their leachates or the chemical leachates alone at an environmentally relevant (280 particles/L) or high (2800 particles/L) concentration for 14 d. The effects of microplastics differed by polymer type and presence of environmental contaminants, and effects can be driven by the physical particles and/or the chemical leachates alone. Larvae exposed to preconsumer polyethylene experienced a decrease in survival, length, and weight, whereas preconsumer polypropylene caused an increase in weight. Environmental microplastics caused a more drastic increase in length and weight and almost 6 times more deformities as the preconsumer microplastics. Although preconsumer microplastics caused effects only when organisms were exposed to both the particles and the chemical leachates, the environmental microplastics caused effects when organisms were exposed to the chemical leachates alone, suggesting that the mechanism of effects are context-dependent. The present study provides further support for treating microplastics as a multiple stressor and suggests that testing for effects with pristine microplastics may underestimate the true effects of microplastics in the environment. Environ Toxicol Chem 2022;41:858-868. © 2021 SETAC.

Cyanotoxins within and Outside of Microcystis aeruginosa Cause Adverse Effects in Rainbow Trout (Oncorhynchus mykiss).

The global expansion of toxic Microcystis blooms, and production of cyanotoxins including microcystins, are an increasing risk to freshwater fish. Differentiating intracellular and extracellular microcystin toxicity pathways (i.e., within and outside of cyanobacterial cells) in fish is necessary to assess the severity of risks to populations that encounter harmful algal blooms in pre-to-postsenescent stages. To address this, adult and juvenile Rainbow Trout (Oncorhynchus mykiss) were, respectively, exposed for 96 h to intracellular and extracellular microcystins (0, 20, and 100 µg L-1) produced by Microcystis aeruginosa. Fish were dissected at 24 h intervals for histopathology, targeted microcystin quantification, and nontargeted proteomics. Rainbow Trout accumulated intracellular and extracellular microcystins in all tissues within 24 h, with greater accumulation in the extracellular state. Proteomics revealed intracellular and extracellular microcystins caused sublethal toxicity by significantly dysregulating proteins linked to the cytoskeletal structure, stress responses, and DNA repair in all tissues. Pyruvate metabolism in livers, anion binding in kidneys, and myopathy in muscles were also significantly impacted. Histopathology corroborated these findings with evidence of necrosis, apoptosis, and hemorrhage at similar severity in both microcystin treatments. We demonstrate that sublethal concentrations of intracellular and extracellular microcystins cause adverse effects in Rainbow Trout after short-term exposure.

Impact of salt-contaminated freshwater on osmoregulation and tracheal gill function in nymphs of the mayfly Hexagenia rigida.

The impact of freshwater (FW) salinization on osmoregulation as well as tracheal gill morphology and function was examined in nymphs of the mayfly Hexagenia rigida following exposure to salt contaminated water (SCW, 7.25 g/l NaCl) for a 7-day period. Ionoregulatory homeostasis was perturbed in SCW exposed H. rigida nymphs as indicated by increased hemolymph Na+, K+ and Cl- levels as well as hemolymph pH and water content. Despite this, SCW did not alter gill Na+-K+-ATPase (NKA) or V-type H+-ATPase (VA) activity. In addition, NKA and VA immunolocalization in gill ionocytes did not show alterations in enzyme location or changes in ionocyte abundance. The latter observation was confirmed using scanning electron microscopy (SEM) to examine exposed tracheal gill ionocyte numbers. Ionocyte surface morphometrics also revealed that SCW did not change individual ionocyte surface area or ionocyte fractional surface area. Nevertheless, analysis of Na+ movement across the tracheal gill of mayfly nymphs using scanning ion-selective electrode technique indicated that FW nymphs acquired Na+ from surrounding water, while tracheal gills of SCW nymphs had the capacity to secrete Na+. Because Na+ secretion across the gill of SCW-exposed animals occurred in the absence of any change in (1) NKA and VA activity or (2) ionocyte numbers/surface exposure, it was reasoned that Na+ movement across the gill of SCW animals may be occurring, at least in part, through the paracellular pathway. The ultrastructure of tracheal gill septate junctions (SJs) supported this idea as they exhibited morphological alterations indicative of a leakier pathway. Data provide a first look at alterations in osmoregulatory mechanisms that allow H. rigida nymphs to tolerate sub-lethal salinization of their surroundings.

Strategies of ionoregulation in the freshwater nymph of the mayfly Hexagenia rigida.

This study investigated ionoregulatory strategies used by freshwater (FW) nymphs of the mayfly Hexagenia rigida Like other FW organisms, H. rigida nymphs maintain hemolymph ion levels (in mmol l-1: Na+ ∼102; Cl- ∼84; K+ ∼6; pH ∼7.35) far in excess of their surroundings. This appears to be accomplished by the combined actions of the alimentary canal, Malpighian tubules (MTs) and tracheal gills. The alimentary canal contributes in a region-specific manner, a view supported by: (1) spatial differences in the activity of basolateral Na+/K+-ATPase (NKA) and apical V-type H+-ATPase (VA) and (2) region-specific Na+ and K+ flux rates. Both indicate a prominent role for the hindgut (rectum) in K+ reabsorption. MTs also exhibit region-specific differences in Na+ and K+ flux rates that are coupled with an organized but tortuous architecture. NKA and VA activities were highest in MTs versus all other organs examined. Tracheal gills were found to be sites of Na+ uptake, but no difference in Na+ uptake was found between gills taken from different regions of the abdomen or spatially along individual gills. This is likely because each gill exhibited a dense population of NKA and/or VA immunoreactive cells (putative ionocytes). Data provide new insight into how FW mayfly nymphs regulate salt and water balance using the alimentary canal, MTs and tracheal gills as well as the first direct evidence that tracheal gills acquire ions from FW.

Using a modified dredging elutriate testing approach to evaluate potential aquatic impacts associated with dredging a large freshwater industrial harbor.

Potential adverse impacts to the aquatic environment should be minimized whenever possible during an environmental dredging project by selecting realistic and technically feasible environmental targets. These targets need to balance short term impacts with the longer term benefit of removing contaminated sediments from the environment. Environmental dredging is part of the planned remediation of Randle Reef (a 60 hectare zone of mostly PAH-contaminated sediments) in Hamilton, Ontario, Canada. In this study, we describe the results of dredging elutriate toxicity testing (DETE) to assess the potential risks from dredging this PAH contaminated site. A modified elutriate preparation method intended as an alternative measure of conditions within the dredging plume was assessed with both standard water column species (Daphnia magna and fathead minnow [Pimephales promelas]) and alternative benthic and epibenthic test organisms (Chironomus dilutus and Hyalella azteca). The standard DETE test was also conducted with H. azteca to compare with the modified DETE results. The greatest toxic response was seen in the alternative test species; however, the modified DETE method resulted in less toxicity than the standard protocol. The relationship between toxicity results and chemical and/or physical characteristics of the samples was examined, but differences in toxicity could only be explained by differences in the total suspended solids concentrations in the elutriate samples. Challenges associated with DETE assessment of PAH-contaminated sediments and the implications for establishing dredging benchmarks are discussed. Integr Environ Assess Manag 2017;13:155-166. © 2016 SETAC.

Interlaboratory evaluation of the assessment of arsenic bioaccumulation from field collected sediments using Hexagenia spp.

Standardized bioaccumulation testing of aquatic organisms is essential to understanding the impact of historical contamination on the quality of water and sediment. A standardized 28-d laboratory bioaccumulation method with a freshwater burrowing mayfly, Hexagenia spp., has been developed and internally validated by the Ontario Ministry of the Environment and Climate Change (MOECC). An interlaboratory comparison was conducted to assess the precision of this method. Field-collected sediment contaminated with arsenic was chosen for the present study. Control and test sediments were subsampled and sent to 6 laboratories to perform the bioaccumulation test. One laboratory failed to meet the control survival criterion of ≥80%. When results of this laboratory are removed from the arsenic accumulation assessment, the mean interlaboratory variability (expressed as coefficient of variation) of the arsenic whole-body concentration is reduced from 44% to 24% in the test sediment-exposed Hexagenia spp. There was no significant interlaboratory difference between the Hexagenia spp. arsenic accumulations. While improved culturing and organism holding guidance may increase laboratory success, the MOECC Hexagenia spp. bioaccumulation test method has tight biological method precision when the control survival criterion is met. Environ Toxicol Chem 2016;35:2448-2455. © 2016 SETAC.

The evaluation of 3 diets for rearing Hyalella azteca and the influence of diet on acute ammonia toxicity.

Three Hyalella azteca cultures were reared on different diets since birth, reflecting the recommended diets of various investigators. The 3 diets consisted of fish flakes (FF), a mixture of fish flakes supplemented with the diatom Thalassiosira weissflogii (FF-D), and a mixture of fish flakes supplemented with yeast, cereal grass media, and trout chow (FF-YCT). The 3 diets were evaluated by comparing 20 wk of culturing data, along with the organism's response to standard 96-h toxicity testing with ammonium chloride over a range of pH and temperature. Hyalella azteca fed the FF-D diet had the highest overall survival rate (96.6%, standard deviation [SD] 4.3%) compared to those fed the FF diet (92.0%, SD 12.7%), or the FF-YCT diet (91.1%, SD 14.8%), although difference in survival was not statistically significant. Organisms fed the FF-D diet produced a higher number of young per week per adult (6.1, SD 2.8) than the FF diet (5.1, SD 2.2), or the FF-YCT diet (4.0, SD 1.2), although differences were not statistically significant. Of the diets evaluated, H. azteca reared on the FF-D diet were often significantly more resistant to total and un-ionized ammonia toxicity in acute 96-h testing than those reared on the other 2 diets across the 2 temperatures and 5 pHs tested, suggesting this may be the most optimal diet for this species. Environ Toxicol Chem 2016;35:2416-2424. © 2016 SETAC.

Complementary nontargeted and targeted mass spectrometry techniques to determine bioaccumulation of halogenated contaminants in freshwater species.

Assessing the toxicological significance of complex environmental mixtures is challenging due to the large number of unidentified contaminants. Nontargeted analytical techniques may serve to identify bioaccumulative contaminants within complex contaminant mixtures without the use of analytical standards. This study exposed three freshwater organisms (Lumbriculus variegatus, Hexagenia spp., and Pimephales promelas) to a highly contaminated soil collected from a recycling plant fire site. Biota extracts were analyzed by Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) and mass defect filtering to identify bioaccumulative halogenated contaminants. Specific bioaccumulative isomers were identified by comprehensive two-dimensional gas chromatography high-resolution time-of-flight mass spectrometry (GCxGC-HRToF). Targeted analysis of mixed brominated/chlorinated dibenzo-p-dioxins and dibenzofurans (PXDD/PXDFs, X = Br and Cl) was performed by atmospheric pressure gas chromatography tandem mass spectrometry (APGC-MS/MS). Relative sediment and biota instrument responses were used to estimate biota-sediment accumulation factors (BSAFs). Bioaccumulating contaminants varied among species and included polychlorinated naphthalenes (PCNs), polychlorinated dibenzofurans (PCDFs), chlorinated and mixed brominated/chlorinated anthracenes/phenanthrenes, and pyrenes/fluoranthenes (Cl-PAHs and X-PAHs, X = Br and Cl), as well as PXDD/PXDFs. Bioaccumulation potential among isomers also varied. This study demonstrates how complementary high-resolution mass spectrometry techniques identify persistent and bioaccumulative contaminants (and specific isomers) of environmental concern.