Microparticles in Wild and Caged Biota, Sediments, and Water Relative to Large Municipal Wastewater Treatment Plant Discharges.

Anthropogenically modified microparticles including microplastics are present in municipal wastewater treatment plant (WWTP) effluents; however, it is unclear whether biotic exposures are elevated downstream of these outfalls. In the fall of 2019, the present study examined whether microparticle levels in resident fish, environmental samples, and caged organisms were elevated near the Waterloo and Kitchener WWTP outfalls along the Grand River, Ontario, Canada. Wild rainbow darters (Etheostoma caeruleum) were collected from a total of 10 sites upstream and downstream of both WWTPs, along with surface water and sediment samples to assess spatial patterns over an approximately 70-km river stretch. Amphipods (Hyalella azteca), fluted-shell mussels (Lasmigona costata), and rainbow trout (Oncorhynchus mykiss) were also caged upstream and downstream of one WWTP for 14 or 28 days. Whole amphipods, fish digestive tracts, and mussel tissues (hemolymph, digestive glands, gills) were digested with potassium hydroxide, whereas environmental samples were processed using filtration and density separation. Visual identification, measurement, and chemical confirmation (subset only) of microparticles were completed. Elevated abiotic microparticles were found at several upstream reference sites as well as at one or both wastewater-impacted sites. Microparticles in amphipods, all mussel tissues, and wild fish did not show patterns indicative of increased exposures downstream of effluent discharges. In contrast, elevated microparticle counts were found in trout caged directly downstream of the outfall. Across all samples, cellulose fibers (mainly blue and clear colors) were the most common. Overall, results suggest little influence of WWTP effluents on microparticles in biota but rather a ubiquitous presence across most sites that indicates the importance of other point and nonpoint sources to this system. Environ Toxicol Chem 2024;43:1047-1061. © 2024 His Majesty the King in Right of Canada and The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. Reproduced with the permission of the Minister of Environment and Climate Change Canada.

Environmental Risks of Pharmaceutical Mixtures in Aquatic Ecosystems: Reflections on a Decade of Research.

Pharmaceuticals and personal care products (PPCPs) occur as variable mixtures in surface waters receiving discharges of human and animal wastes. A key question identified a decade ago is how to assess the effects of long-term exposures of these PPCP mixtures on nontarget organisms. We review the recent progress made on assessing the aquatic ecotoxicity of PPCP mixtures-with a focus on active pharmaceutical ingredients-and the challenges and research needs that remain. New knowledge has arisen from the use of whole-mixture testing combined with component-based approaches, and these studies show that mixtures often result in responses that meet the concentration addition model. However, such studies have mainly been done on individual species over shorter time periods, and longer-term, multispecies assessments remain limited. The recent use of targeted and nontargeted gene analyses has improved our understanding of the diverse pathways that are impacted, and there are promising new "read-across" methods that use mammalian data to predict toxicity in wildlife. Risk assessments remain challenging given the paucity of ecotoxicological and exposure data on PPCP mixtures. As such, the assessment of PPCP mixtures in aquatic environments should remain a priority given the potential for additive-as well as nontarget-effects in nontarget organisms. In addition, we need to improve our understanding of which species, life stages, and relevant endpoints are most sensitive to which types of PPCP mixtures and to expand our knowledge of environmental PPCP levels in regions of the globe that have been poorly studied to date. We recommend an increased use of new approach methodologies, in particular "omics," to advance our understanding of the molecular mechanics of mixture effects. Finally, we call for systematic research on the role of PPCP mixtures in the development of antimicrobial resistance. Environ Toxicol Chem 2024;43:549-558. © 2023 SETAC.

Spatial patterns of sediment contamination and their influence on benthic infaunal communities in a highly tidal and industrial estuary in Atlantic Canada.

Sediment contamination can be elevated in ports, harbours, and estuaries with legacies of exploitation, negatively impacting infaunal invertebrate communities. Saint John Harbour (45.25° N, 66.05° W), New Brunswick, Canada, is an active harbour with strong tides and a long history of human activity. To examine spatial patterns of sediment contamination, samples were collected between 2011 and 2021 from subtidal sites near potential contaminant sources. Invertebrate data from the same samples were used to investigate potential effects on biological communities. Contaminant concentrations in the inner parts of the harbour were elevated compared to reference sites, but generally did not reach levels comparable to other highly contaminated harbours in the region. Effects on invertebrates were detectable, particularly at sites with higher contamination, although physical factors (depth, sediment grain size) were more important. Dynamic tidal conditions in the harbour may reduce the accumulation of contaminants in subtidal sediments and their impacts on infaunal communities.

Elements and omega-3 fatty acids in fishes along a large, dammed river.

Damming of a river can trap and elevate levels of sediment-bound elements and alter food web dynamics in created reservoirs. It follows that dams may alter how elements and other nutrients, like the beneficial omega-3 fatty acids (n-3 FAs) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), are accumulated in fish and thus the chemical composition of species above and below this barrier to migration. This study examined the spatial and species differences in contaminants and nutrients in fish from the Wolastoq | Saint John River (New Brunswick, Canada) in association with a large hydroelectric dam (Mactaquac Generating Station; MQGS), a river which supports both recreational fisheries and subsistence fishing by Indigenous communities. In 2020 and 2021, Smallmouth Bass, Yellow Perch, American Eel, and Striped Bass were collected from locations upstream (reservoir and river) and downstream of the MQGS and analyzed for mercury (Hg) and 30 other trace elements, n-3 FAs, δ15N, and δ13C. Fish from the reservoir were highest in the beneficial elements P, S, and K, while fish from upstream of the reservoir had lower levels of toxic elements, including Hg. The dam appeared to alter food web dynamics, as fish from the reservoir and immediately downstream of the dam had higher δ15N and reservoir fish were depleted in δ13C. DHA and Hg were positively corelated with δ15N, and EPA in Smallmouth Bass was higher in sites where fish had higher δ13C. Overall, this study suggests that the dam altered food web dynamics and the uptake of contaminants and nutrients by fish, and that location and species are important factors when examining the risks and benefits of consuming wild fish from a system impacted by a large dam.

Temporal and Spatial Trends in Benthic Infauna and Potential Drivers, in a Highly Tidal Estuary in Atlantic Canada.

Infaunal invertebrate communities of coastal marine sediments are often impacted by human activities, particularly in harbours and estuaries. However, while many studies have attempted to identify the key factors affecting benthic infauna, few have done so for highly energetic tidal estuaries. Samples were collected over a decade (2011-2020) from a series of reference sites in Saint John Harbour (45.25° N, 66.05° W), a highly tidal estuary in the Bay of Fundy, Canada. These data were used to examine spatial and temporal trends in infaunal invertebrate communities and sediment properties and to determine the extent to which the biological patterns were driven by measured physical and chemical variables. There were substantial differences among sites in infaunal invertebrate abundance (median ranging from 688 to 13,700 individuals per square meter), infaunal species richness (median ranging from 8 to 22), and Shannon diversity (median ranging from 1.26 to 2.34); multivariate analysis also revealed variation in species composition among sites. Sediment contaminant concentrations also varied among sites, but differences tended to be smaller (e.g. median chromium concentrations ranging from 21.6 to 27.6 mg/kg). Sample contaminant concentrations were all below probable effect levels, and almost all below threshold effect levels (Canadian interim sediment quality guidelines), but relationships with biological data were still detectable. However, physical variables (depth, sediment characteristics) were better predictors of biological variables and community composition. These results confirm the importance of physical factors in shaping infaunal communities in soft-sediment habitats in tidally influenced coastal waters. Supplementary Information: The online version contains supplementary material available at 10.1007/s12237-023-01222-w.

Human health risk assessment of metals and arsenic via consumption of commercial bivalves in the Gulf of California, Mexico.

Although various studies have reported the bioaccumulation of metals in bivalves of commercial interest from the Gulf of California (GC), the risk associated with their consumption remains poorly understood. In this study, our own and bibliographic concentrations of 14 elements in 16 species of bivalves from 23 locations were used to investigate (1) the species-specific and regional accumulation of metals and arsenic in bivalves, (2) the human health risks by age and sex, and (3) the maximum allowable consumption rates (CRlim). The assessments were done according to the US Environmental Protection Agency guidelines. The results indicate that the bioaccumulation of elements varies markedly between groups (oysters > mussels > clams) and localities (higher on Sinaloa due to intense anthropogenic activities). However, consuming bivalves from the GC remains safe for human health. To prevent health effects for residents or consumers on the GC, we recommend (1) following the CRlim proposed herein; (2) monitoring levels of Cd, Pb, and As (inorganic) in bivalves, as the elements of top concern, mainly when are consumed by children; (3) calculating CRlim for more species and locations, including at least: As, Al, Cd, Cu, Fe, Mn, Pb, and Zn; and (4) determine the regional consumption rates of bivalves.

Elevated Allochthony in Stream Food Webs as a Result of Longitudinal Cumulative Effects of Forest Management.

The river continuum concept (RCC) predicts a downstream shift in the reliance of aquatic consumers from terrestrial to aquatic carbon sources, but this concept has rarely been assessed with longitudinal studies. Similarly, there are no studies addressing how forestry related disturbances to the structure of headwater food webs manifest (accumulate/dissipate) downstream and/or whether forest management alters natural longitudinal trends predicted by the RCC. Using stable isotopes of carbon, nitrogen and hydrogen, we investigated how: 1) autochthony in macroinvertebrates and fish change from small streams to larger downstream sites within a basin with minimal forest management (New Brunswick, Canada); 2) longitudinal trends in autochthony and food web length compare among three basins with different forest management intensity [intensive (harvest and replanting), extensive (harvest only), minimal] to detect potential cumulative/dissipative effects; and 3) forest management intensity and other catchment variables are influencing food web dynamics. We showed that, as predicted, the reliance of some macroinvertebrate taxa (especially collector feeders) on algae increased from small streams to downstream waters in the minimally managed basin, but that autochthony in the smallest shaded stream was higher than expected based on the RCC (as high as 90% for some taxa). However, this longitudinal increase in autochthony was not observed within the extensively managed basin and was weaker within the intensively managed one, suggesting that forest management can alter food web dynamics along the river continuum. The dampening of downstream autochthony indicates that the increased allochthony observed in small streams in response to forest harvesting cumulates downstream through the river continuum. Supplementary Information: The online version contains supplementary material available at 10.1007/s10021-021-00717-6.

Using tissue cysteine to predict the trophic transfer of methylmercury and selenium in lake food webs.

The biomagnification of toxic methylmercury (MeHg) and selenium (Se) through aquatic food webs using nitrogen stable isotopes (δ15N) varies among ecosystems but underlying mechanisms are yet unexplained. Given the strong links between MeHg and thiol-containing amino acids and proteins containing selenocysteine, our hypothesis was that cysteine content is a better predictor of MeHg and Se transfer through lake food webs than δ15N. Food web samples were collected from six lakes in Kejimkujik National Park, Nova Scotia, Canada, and the regression slopes of log MeHg or Se versus protein-bound cysteine or bulk δ15N were compared. Across all six lakes, MeHg varied by a factor of 10 among taxa and was significantly and positively related to both cysteine (R2 = 0.65-0.80, p < 0.001) and δ15N (R2 = 0.88-0.94, p < 0.001), with no among-system differences in these slopes. In contrast, total Se concentrations varied by less than a factor of 2 among taxa in four lakes and were significantly related to cysteine in only two food webs (R2 = 0.20 & 0.37, p = 0.014 & < 0.001); however, δ15N was not a predictor of Se in any lake (p = 0.052-0.777). Overall, these novel results indicate that cysteine content predicts MeHg, and sometimes Se, across trophic levels, providing a potential mechanism for among-system differences in their biomagnification.

Effects of forest management on mercury bioaccumulation and biomagnification along the river continuum.

Forest management can alter the mobilization of mercury (Hg) into headwater streams and its conversion to methylmercury (MeHg), the form that bioaccumulates in aquatic biota and biomagnifies through food webs. As headwater streams are important sources of organic materials and nutrients to larger systems, this connectivity may also increase MeHg in downstream biota through direct or indirect effects of forestry on water quality or food web structure. In this study, we collected water, seston, food sources (biofilm, leaves, organic matter), five macroinvertebrate taxa and fish (slimy sculpin; Cottus cognata) at 6 sites representing different stream orders (1-5) within three river basins with different total disturbances from forestry (both harvesting and silviculture). Methylmercury levels were highest in water and some food sources from the basin with moderate disturbance (greater clearcutting but less silviculture). Water, leaves, stoneflies and fish increased in MeHg or total Hg along the river continuum in the least disturbed basin, and there were some dissipative effects of forest management on these spatial patterns. Trophic level (δ15N) was a significant predictor of MeHg (and total Hg in fish) within food webs across all 18 sites, and biomagnification slopes were significantly lower in the basin with moderate total disturbance but not different in the other two basins. The elevated MeHg in lower trophic levels but its reduced trophic transfer in the basin with moderate disturbance was likely due to greater inputs of sediments and of dissolved organic carbon that is more humic, as these factors are known to both increase transport of Hg to streams and its uptake in primary producers but to also decrease MeHg bioaccumulation in consumers. Overall, these results suggest that the type of disturbance from forestry affects MeHg bioaccumulation and trophic transfer in stream food webs and some longitudinal patterns along a river continuum.

Effects of municipal wastewater effluents on the digestive gland microbiome of wild freshwater mussels (Lasmigona costata).

Gut microbial communities are vital for maintaining host health, and are sensitive to diet, environment, and chemical exposures. Wastewater treatment plants (WWTPs) release effluents containing antimicrobials, pharmaceuticals, and other contaminants that may negatively affect the gut microbiome of downstream organisms. This study investigated changes in the diversity and composition of the digestive gland microbiome of flutedshell mussels (Lasmigona costata) from upstream and downstream of two large (service >100,000) WWTPs. Mussel digestive gland microbiome was analyzed following the extraction, PCR amplification, and sequencing of bacterial DNA using the V3-V4 hypervariable regions of the 16 S rRNA gene. Bacterial alpha diversity decreased at sites downstream of the second WWTP and these sites were dissimilar in beta diversity from sites upstream and downstream of the first upstream WWTP. The microbiomes of mussels collected downstream of the first WWTP had increased relative abundances of Actinobacteria, Bacteroidetes, and Firmicutes, with a decrease in Cyanobacteria, compared to upstream mussels. Meanwhile, those collected downstream of the second WWTP increased in Proteobacteria and decreased in Actinobacteria, Bacteroidetes, and Tenericutes. Increased Proteobacteria has been linked to adverse effects in mammals, but their functions in mussels is currently unknown. Finally, effluent-derived bacteria were found in the microbiome of mussels downstream of both WWTPs but not in those from upstream. Overall, results show that the digestive gland microbiome of mussels collected upstream and downstream of WWTPs differed, which has implications for altered host health and the transport of WWTP-derived bacteria through aquatic ecosystems.

Comparative Effects of Embryonic Metformin Exposure on Wild and Laboratory-Spawned Fathead Minnow (Pimephales promelas) Populations.

Metformin is routinely detected in aquatic ecosystems because of its widespread use as a treatment for Type 2 diabetes. Laboratory studies have shown that exposure to environmentally relevant concentrations of metformin can alter metabolic pathways and impact the growth of early life stage (ELS) fish; however, it is unknown whether these effects occur in wild populations. Herein, we evaluate whether findings from laboratory studies are representative and describe the relative sensitivities of both populations. Duplicate exposures (0, 5, or 50 µg/L metformin) were conducted using wild- and lab-spawned fathead minnow (Pimephales promelas) embryos. Apart from the water source, exposure conditions remained constant. Wild embryos were exposed to previously dosed lake water to account for changes in bioavailability, while reconstituted freshwater was used for the laboratory study. Developmental metformin exposure differentially impacted the growth and morphology of both cohorts, with energy dyshomeostasis and visual effects indicated. The fitness of wild-spawned larvae was impacted to a greater extent relative to lab-spawned fish. Moreover, baseline data reveal important morphological differences between wild- and lab-spawned ELS fatheads that may diminish representativeness of lab studies. Findings also confirm the bioavailability of metformin in naturally occurring systems and suggest current exposure scenarios may be sufficient to negatively impact developing fish.

Age, body size, growth and dietary habits: What are the key factors driving individual variability in mercury of lacustrine fishes in northern temperate lakes?

Fish total mercury concentration ([THg]) has been linked to various fish attributes, but the relative importance of these attributes in accounting for among-individual variation in [THg] has not been thoroughly assessed. We compared the contributions of ontogeny (age, length), growth (growth rate, body condition), and food web position (δ13C, δ15N) to among-individual variability in [THg] within populations of seven common fishes from 141 north temperate lakes. Ontogenetic factors accounted for most variation in [THg]; age was a stronger and less variable predictor than length for most species. Adding both indices of growth and food web position to these models increased explained variation (R2) in [THg] by 6-25% among species. Fish [THg] at age increased with growth rate, while fish [THg] at length decreased with growth rate, and the effect of body condition was consistently negative. Trophic elevation (inferred from δ15N) was a stronger predictor than primary production source (inferred from δ13C) for piscivores but not benthivores. Fish [THg] increased with δ15N in all species but showed a more variable relationship with δ13C. Among-individual variation in [THg] is primarily related to age or size in most temperate freshwater fishes, and effects of growth rate and food web position need to be considered in the context of these ontogenetic drivers.

PCB exposure is associated with reduction of endosymbionts in riparian spider microbiomes.

Microbial communities, including endosymbionts, play diverse and critical roles in host biology and reproduction, but contaminant exposure may cause an imbalance in the microbiome composition with subsequent impacts on host health. Here, we examined whether there was a significant alteration of the microbiome community within two taxa of riparian spiders (Tetragnathidae and Araneidae) from a site with historical polychlorinated biphenyl (PCB) contamination in southern Ontario, Canada. Riparian spiders specialize in the predation of adult aquatic insects and, as such, their contaminant levels closely track those of nearby aquatic ecosystems. DNA from whole spiders from sites with either low or high PCB contamination was extracted, and spider microbiota profiled by partial 16S rRNA gene amplicon sequencing. The most prevalent shift in microbial communities we observed was a large reduction in endosymbionts in spiders at the high PCB site. The abundance of endosymbionts at the high PCB site was 63 % and 98 % lower for tetragnathids and araneids, respectively, than at the low PCB site. Overall, this has potential implications for spider reproductive success and food webs, as riparian spiders are critical gatekeepers of energy and material fluxes at the land-water interface.

Are There Longitudinal Effects of Forest Harvesting on Carbon Quality and Flow and Methylmercury Bioaccumulation in Primary Consumers of Temperate Stream Networks?

Forest harvesting affects dissolved organic matter (DOM) and aqueous mercury inputs as well as the food web structure in small-headwater streams, but how these upstream changes manifest downstream is unclear. To address this uncertainty, we examined DOM quality, autochthony in the caddisfly Hydropsychidae (using δ2 H), and methylmercury (MeHg) concentrations in stream water and the caddisfly along a longitudinal gradient (first- to fourth-order streams, subcatchments of 50-1900 ha) in paired partially harvested and reference catchments in central Ontario, Canada. Although measures of DOM quality (specific ultraviolet absorbance at 254 nm 2.20-11.62) and autochthony in caddisflies (4.9%-34.0%) varied among sites, no upstream-to-downstream differences in these measures were observed between the paired harvested and reference catchments. In contrast, MeHg levels in stream water (0.06-0.35 ng/L) and caddisflies (29.7-192 µg/kg dry wt) were significantly higher in the upstream sites but not the farthest downstream sites in the harvested catchments compared to the reference catchments. This suggests that while current mitigation measures used by forestry companies did not prevent elevated MeHg in water and invertebrates at smaller spatial scales (subcatchments of 50-400 ha), these upstream impacts did not manifest at larger spatial scales (subcatchments of 800-1900 ha). The present study advances our understanding of spatially cumulative impacts within harvested catchments, which is critical to help forest managers maintain healthy forest streams and their provisioning of aquatic ecosystem services. Environ Toxicol Chem 2022;41:1490-1507. © 2022 SETAC.

Mercury concentrations and stable isotopes (δ15N and δ13C) in fish muscle indicate human impacts in tropical coastal lagoons.

Coastal lagoons are essential nursery habitats of many marine fishes, but they are often sites of intense human activities that impact water quality and potentially the fish using these habitats. We compared the variability in nutrient uptake (using δ15N and δ13C) and total mercury (THg) levels in juveniles and adults of three common species in two lagoons on the central Mexican coast of the Pacific Ocean during the wet and dry seasons. One of three species, Achirus mazatlanus had higher THg, δ15N, and δ13C levels in the lagoon with the greatest wastewater inputs (Barra de Navidad). Delta13C varied seasonally for all three species and THg was higher in the dry season for Lutjanus argentiventris and in males of A. mazatlanus. Our results demonstrate that mercury and stable isotopes can identify impacts of human activities on estuarine ichthyofauna and the importance of understanding seasonal and spatial variability of measures that could impact monitoring and predictions of impacts in these lagoons.

Impacts of wastewater treatment plants on benthic macroinvertebrate communities in summer and winter.

Treated effluent from municipal wastewater treatment plants (WWTPs) is a major source of contamination that can impact population size, community structure, and biodiversity of aquatic organisms. However, because the majority of field research occurs during warmer periods of the year, the impacts of wastewater effluent on aquatic communities during winter has largely been neglected. In this study, we assessed the impacts of wastewater effluent on aquatic benthic macroinvertebrate (benthos) communities along the effluent gradients of two WWTPs discharging into Hamilton Harbour, Canada, during summer and winter using artificial substrates incubated for 8 weeks. At the larger of the two plants, benthic macroinvertebrate abundance was higher and diversity was lower at sites downstream of the outfall compared to upstream sites in both seasons. Whereas at the smaller plant, the opposite was observed, abundance increased and diversity decreased with distance from the outfall in both seasons. While the impacts of wastewater on benthic communities were largely similar between seasons, we did detect several general seasonal trends - family diversity of macroinvertebrates was lower during winter at both WWTPs and total abundance was also lower during winter, but only significantly so at the smaller WWTP. Further, benthic macroinvertebrate community composition differed significantly along the effluent gradients, with sites closest and farthest from the outfall being the most dissimilar. Our contrasting results between the WWTPs demonstrate that plants, with different treatment capabilities and effluent-receiving environments (industrial/urban versus wetland), can dictate how wastewater effluent impacts benthic macroinvertebrate communities.

Trophodynamics of trace elements in marine organisms from cold and remote regions of southern hemisphere.

Trace metals bioaccumulate in aquatic organisms and some of them biomagnify through food webs, posing a threat to the organisms or their human consumers. Although the trophodynamics of many trace metals is well known in the northern hemisphere, much less is known about metals in aquatic food webs from cold and remote coastal zones of the southern hemisphere. To fill this gap, we investigated the trophodynamics of Al, Co, Cr, Li, Mo, Ni, Sr, and V, which were measured in marine macroinvertebrates and fishes from inshore and offshore locations in each of the Chilean Patagonia and the Antarctic Peninsula area. In Patagonia, there was biodilution of these metals across the whole food web, while biomagnification of Li and Ni was significantly found across the lower food web at the offshore site. In Antarctica, significant biodilution of Al, Li, Ni, Mo, Sr and V occurred through the whole food web for the inshore site, but no tendency (biodilution or biomagnification) was found (p > 0.05) across the organisms at lower trophic levels for the offshore site. Our data suggest that the geographic location and species influences the trophodynamics of these trace elements and expand our understanding of metal fate in remote locations of the southern hemisphere.

Altered microbiomes of aquatic macroinvertebrates and riparian spiders downstream of municipal wastewater effluents.

Municipal wastewater treatment plants (WWTPs) contain numerous contaminants, including antimicrobials, that could affect the composition of the beneficial bacterial communities associated with host aquatic organisms. There is also potential for these effects to transfer to terrestrial predators. Riparian spiders and five families of aquatic macroinvertebrates were collected from sites upstream and downstream of two WWTPs, Waterloo and Kitchener, discharging to the Grand River, Ontario, Canada. Whole-body microbiota were analyzed following the extraction, PCR amplification, and sequencing of bacterial DNA using the V3-V4 hypervariable regions of the 16S rRNA genetic barcode. Changes in the relative abundance of major microbiome phyla were observed in all downstream aquatic insects except Hydropsychidae caddisflies, which exhibited little site variation. Shannon alpha diversity differed among sites for Tetragnathidae spiders, Perlidae, Hydropsychidae, and Heptageniidae. Downstream of the Waterloo WWTP alpha diversity decreased in spiders, while downstream of the Kitchener WWTP this measure decreased in Perlidae and increased in spiders. Bray-Curtis beta diversity was dissimilar among sites in all invertebrate taxa; upstream sites differed from those downstream of Waterloo in spiders, Perlidae, and Hydropsychidae, and from those downstream of Kitchener in spiders, Perlidae, and Hydropsychidae. Finally, effluent-derived bacteria were found in the microbiomes of downstream spiders and aquatic insects and not upstream. Overall, results indicated that the microbiomes of invertebrates collected downstream differed from those collected upstream of WWTPs, which has implications for altered host health and transport of WWTP-derived bacteria through aquatic ecosystems.

Chronic Embryo-Larval Exposure of Fathead Minnows to the Pharmaceutical Drug Metformin: Survival, Growth, and Microbiome Responses.

Metformin is a glucose-lowering drug commonly found in municipal wastewater effluents (MWWEs). The present study investigated the chronic effects of metformin in early-life stages of the fathead minnow (Pimephales promelas). Endpoints assessed were growth, survival, and deformities. The larval gut microbiome was also examined using 16 S ribosomal RNA gene amplicon sequencing to determine microbial community composition and alpha and beta diversity. Eggs and larvae were exposed to metformin measured concentrations (mean [standard deviation]) of 0.020 (0.017) µg/L (for controls) and 3.44 (0.23), 33.6 (1.6), and 269 (11) µg/L in a daily static-renewal setup, with 20 embryos per beaker. The low and middle metformin exposure concentrations represent river and MWWE concentrations of metformin. To detect small changes in growth, we used 18 replicate beakers for controls and 9 replicates for each metformin treatment. Over the 21-d exposure (5 d as embryos and 16 d posthatch [dph]), metformin did not affect survival or growth of larval fish. Hatch success, time to hatch, deformities in hatched fry, and survival were similar across all treatments. Growth (wet wt, length, and condition factor) assessed at 9 and 16 dph was also unaffected by metformin. Assessment of the microbiome showed that the larvae microbiome was dominant in Proteobacteria and Firmicutes, with small increases in Proteobacteria and decreases in Firmicutes with increasing exposure to metformin. No treatment effects were found for microbiome diversity measures. Control fish euthanized with the anesthetic tricaine methane sulfonate had decreased alpha diversity compared to those sampled by spinal severance. This experiment demonstrates that metformin at environmentally relevant concentrations (3.44 and 33.6 µg/L) and at 10 times MWWE concentrations (269 µg/L) does not adversely affect larval growth or gut microbiome in this ubiquitous freshwater fish species. Environ Toxicol Chem 2022;41:635-647. © 2021 Her Majesty the Queen in Right of Canada. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. Reproduced with the permission of the Minister of Environment and Climate Change Canada.