Do Methodological Differences in Experiments with Stream Shredders Imply Variability in Outputs? A Microcosm Approach.

Experiments are useful scientific tools for testing hypotheses by manipulating variables of interest while controlling for other factors that can bias or confuse the results and their interpretation. To ensures accuracy and reproducibility, experiments must have transparent and repeatable methodologies. Due to the importance of shredder invertebrates in organic matter processing, carbon cycling, and nutrient cycling, we tested experimentally the effect of different methodological approaches in microcosm experiments on the consumption and survival of shredders. We found that the shredder species, the presence or absence of the case, and the use or non-use of air-pumps in the microcosms did not affect shredder performance (i.e., consumption and survival). Furthermore, the type of water (stream or bottled) did not affect shredder performance. On the other hand, the amount of light had a negative effect on shredder performance, with constant light (i.e., 24 h) reducing shredder consumption and survival. Our results demonstrate that the use of different methodologies does not always result in changes in outcomes, thus ensuring comparability. However, luminosity is a critical factor that deserves attention when conducting microcosm experiments. Our findings provide valuable insights that can assist researchers in designing experiments with shredders from neotropical streams and conducting systematic reviews and meta-analyses.

Morphological variations and demographic responses of the Mediterranean pond turtle Mauremys leprosa to heterogeneous aquatic habitats.

Human activities affect terrestrial and aquatic habitats leading to changes at both individual and population levels in wild animal species. In this study, we investigated the phenotype and demographics of the Mediterranean pond turtle Mauremys leprosa (Schweigger, 1812) in contrasted environments of Southern France: two peri-urban rivers receiving effluents from wastewater treatment plants (WWTP), and another one without sewage treatment plant. Our findings revealed the presence of pesticides and pharmaceuticals in the three rivers of investigation, the highest diversities and concentrations of pollutants being found in the river subsections impacted by WWTP effluents. Principal component analysis and hierarchical clustering identified three levels of habitat quality, with different pollutant concentrations, thermal conditions, nutrient, and organic matter levels. The highest turtle densities, growth rates, and body sizes were estimated in the most disturbed habitats, suggesting potential adult benefits derived from harsh environmental conditions induced by pollution and eutrophication. Conversely, juveniles were the most abundant in the least polluted habitats, suggesting adverse effects of pollution on juvenile survival or adult reproduction. This study suggests that turtles living in polluted habitats may benefit from enhanced growth and body size, at the expense of reproductive success.

Plastic Litter Emits the Foraging Infochemical Dimethyl Sulfide after Submersion in Freshwater Rivers.

Plastic pollution is widespread throughout aquatic environments globally, with many organisms known to interact with and ingest plastic. In marine environments, microbial biofilms that form on plastic surfaces can produce the odorous compound dimethyl sulfide (DMS), which is a known foraging cue. This has been shown to increase the ingestion of plastic by some invertebrates and therefore act as a biological factor which influences the risks of plastic to marine ecosystems. In freshwater however, the production of DMS has been largely overlooked, despite the known sensitivity of some freshwater species to this compound. To address this gap, the present study analyzed the production of DMS by biofilms which formed on low-density polyethylene and polylactic acid films after 3 and 6 weeks of submersion in either a rural or an urban United Kingdom river. Using gas chromatography-mass spectrometry, the production of DMS by these biofilms was consistently identified. The amount of DMS produced varied significantly across river locations and materials, with surfaces in the urban river generally producing a stronger signal and plastics producing up to seven times more DMS than glass control surfaces. Analysis of biofilm weight and photosynthetic pigment content indicated differences in biofilm composition across conditions and suggested that DMS production was largely driven by nonphotosynthetic taxa. For the first time this work has documented the production of DMS by plastic litter after submersion in freshwater rivers. Further work is now needed to determine if, as seen in marine systems, this production of DMS can encourage the interaction of freshwater organisms with plastic litter and therefore operate as a biological risk factor in the impacts of plastic on freshwater environments. Environ Toxicol Chem 2024;00:1-12. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Reservoir ecosystems support large pools of fish biomass.

Humans increasingly dominate Earth's natural freshwater ecosystems, but biomass production of modified ecosystems is rarely studied. We estimate potential fish total standing stock in USA reservoirs is 3.4 billion (B) kg, and approximate annual secondary production is 4.5 B kg y-1. We also observe varied and non-linear trends in reservoir fish biomass over time, thus previous assertions that reservoir fisheries decline over time are not universal. Reservoirs are globally relevant pools of freshwater fisheries, in part due to their immense limnetic footprint and spatial extent. This study further shows that reservoir ecosystems play major roles in food security and fisheries conservation. We encourage additional effort be expended to effectively manage reservoir environments for the good of humanity, biodiversity, and fish conservation.

Effects of biodiversity on functional stability of freshwater wetlands: a systematic review.

Freshwater wetlands are the wetland ecosystems surrounded by freshwater, which are at the interface of terrestrial and freshwater ecosystems, and are rich in ecological composition and function. Biodiversity in freshwater wetlands plays a key role in maintaining the stability of their habitat functions. Due to anthropogenic interference and global change, the biodiversity of freshwater wetlands decreases, which in turn destroys the habitat function of freshwater wetlands and leads to serious degradation of wetlands. An in-depth understanding of the effects of biodiversity on the stability of habitat function and its regulation in freshwater wetlands is crucial for wetland conservation. Therefore, this paper reviews the environmental drivers of habitat function stability in freshwater wetlands, explores the effects of plant diversity and microbial diversity on habitat function stability, reveals the impacts and mechanisms of habitat changes on biodiversity, and further proposes an outlook for freshwater wetland research. This paper provides an important reference for freshwater wetland conservation and its habitat function enhancement.

Distribution and Genetic Lineages of the Craspedacusta sowerbii Species Complex (Cnidaria, Olindiidae) in Italy.

Olindiid freshwater jellyfishes of the genus Craspedacusta Lankester, 1880 are native to eastern Asia; however, some species within the genus have been introduced worldwide and are nowadays present in all continents except Antarctica. To date, there is no consensus regarding the taxonomy within the genus Craspedacusta due to the morphological plasticity of the medusa stages. The species Craspedacusta sowerbii Lankester, 1880 was first recorded in Italy in 1946, and until 2017, sightings of the jellyfish Craspedacusta were reported for 40 water bodies. Here, we shed new light on the presence of the freshwater jellyfishes belonging to the genus Craspedacusta across the Italian peninsula, Sardinia, and Sicily. First, we report 21 new observations of this non-native taxon, of which eighteen refer to medusae sightings, two to environmental DNA sequencing, and one to the finding of polyps. Then, we investigate the molecular diversity of collected Craspedacusta specimens, using a Bayesian analysis of sequences of the mitochondrial gene encoding for Cytochrome c Oxidase Subunit I (mtDNA COI). Our molecular analysis shows the presence of two distinctive genetic lineages: (i) a group that comprises sequences obtained from populations ranging from central to northern Italy; (ii) a group that comprises three populations from northern Italy-i.e., those from the Lake Levico, the Lake Santo of Monte Terlago, and the Lake Endine-and the single known Sicilian population. We also report for the first time a mtDNA COI sequence obtained from a Craspedacusta medusa collected in Spain.

The Emergence of the Family Scirtidae (Insecta: Coleoptera) in Lotic Karst Habitats: A Case Study over 15 Years.

Due to ongoing changes and a decline in biodiversity, science today should rely on long-term species-based ecological studies. We have conducted a long-term ecological dynamics study on the water beetle family Scirtidae, which, although it is very abundant in benthic communities, is still poorly studied. The main objective of this study was to investigate the population aspects (composition, diversity, sex ratio) and ecological aspects (emergence patterns, seasonal dynamics and preferences for environmental factors) of the family Scirtidae over 15 years in Plitvice Lakes NP, Croatia. The study was conducted at three sites and in five different substrate types. A total of three taxa with different distributions were recorded in the study area: Hydrocyphon novaki and H. deflexicollis on the tufa barriers and the Elodes sp. in the spring area. The sex ratio was in favour of males in spring and early summer, while it shifted in favour of females towards the end of autumn. The abundance and emergence of the family were primarily determined by the environmental parameters which showed the greatest fluctuations over a period of 15 years: water temperature, water discharge and oxygen saturation. Our results clearly show that Scirtidae can be used as indicators of stream zonation and habitat quality. Based on the methodology and the results of this study, we conclude that Scirtidae should be used in future monitoring and protection measures in karst freshwater habitats in southeastern Europe.

Using single-species and algal communities to determine long-term adverse effects of silver nanoparticles on freshwater phytoplankton.

The physical and chemical properties of silver nanoparticles (AgNPs) have led to their increasing use in various fields such as medicine, food, and industry. Evidence has proven that AgNPs cause adverse effects in aquatic ecosystems, especially when the release of Ag is prolonged in time. Several studies have shown short-term adverse effects of AgNPs on freshwater phytoplankton, but few studies have analysed the impact of long-term exposures on these populations. Our studies were carried out to assess the effects of AgNPs on growth rate, photosynthesis activity, and reactive oxygen species (ROS) generation on the freshwater green algae Scenedesmus armatus and the cyanobacteria Microcystis aeruginosa, and additionally on microcystin (MC-LR) generation from these cyanobacteria. The tests were conducted both in single-species cultures and in phytoplanktonic communities exposed to 1 ngL-1 AgNPs for 28 days. The results showed that cell growth rate of both single-species cultures decreased significantly at the beginning and progressively reached control-like values at 28 days post-exposure. This effect was similar for the community-cultured cyanobacteria, but not for the green algae, which maintained a sustained decrease in growth rate. While gross photosynthesis (Pg) increased in both strains exposed in single cultures, dark respiration (R) and net photosynthesis (Pn) decreased in S. armatus and M. aeruginosa, respectively. These effects were mitigated when both strains were exposed under community culture conditions. Similarly, the ROS generation shown by both strains exposed in single-species cultures was mitigated when exposure occurred in community cultures. MC-LR production and release were significantly decreased in both single-species and community exposures. These results can supply helpful information to further investigate the potential risks of AgNPs and ultimately help policymakers make better-informed decisions about their utilization for environmental restoration.

Informing the Exposure Landscape: The Fate of Microplastics in a Large Pelagic In-Lake Mesocosm Experiment.

Understanding microplastic exposure and effects is critical to understanding risk. Here, we used large, in-lake closed-bottom mesocosms to investigate exposure and effects on pelagic freshwater ecosystems. This article provides details about the experimental design and results on the transport of microplastics and exposure to pelagic organisms. Our experiment included three polymers of microplastics (PE, PS, and PET) ranging in density and size. Nominal concentrations ranged from 0 to 29,240 microplastics per liter on a log scale. Mesocosms enclosed natural microbial, phytoplankton, and zooplankton communities and yellow perch (Perca flavescens). We quantified and characterized microplastics in the water column and in components of the food web (biofilm on the walls, zooplankton, and fish). The microplastics in the water stratified vertically according to size and density. After 10 weeks, about 1% of the microplastics added were in the water column, 0.4% attached to biofilm on the walls, 0.01% within zooplankton, and 0.0001% in fish. Visual observations suggest the remaining >98% were in a surface slick and on the bottom. Our study suggests organisms that feed at the surface and in the benthos are likely most at risk, and demonstrates the value of measuring exposure and transport to inform experimental designs and achieve target concentrations in different matrices within toxicity tests.

Efficacy of essential oil from ginger (Zingiber officinale) for anesthesia and transport sedation of pacu (Piaractus mesopotamicus).

This study evaluated the anesthetic and sedative effects of the essential oil of Zingiber officinale (EOZO) on juvenile pacu (Piaractus mesopotamicus). Experiment 1 evaluated concentrations of 0, 50, 100, 200 and 400 mg L-1 EOZO for times of induction and recovery from anesthesia. Furthermore, hematological responses and residual components of EOZO in plasma were determined immediately after anesthesia. Experiment 2 evaluated the effect of 0, 10, 20 and 30 mg L-1 EOZO on water quality, blood variables and residual components of EOZO in plasma and tissues (muscle and liver) immediately after 2 h of transport. Survival was 100%. The three main compounds of EOZO [zingiberene (32.27%), ß-sesquiphellandrene (18.42%) and ß-bisabolene (13.93%)] were observed in animal plasma and tissues (muscle and liver) after anesthesia and transport, demonstrating a direct linear effect among the evaluated concentrations. The concentration of 200 mg L-1 EOZO promoted surgical anesthesia of pacu and prevented an increase in monocyte and neutrophil levels, yet did not alter other hematological parameters. The use of 30 mg L-1 EOZO has a sedative effect on juvenile pacu, thereby reducing oxygen consumption during transport. Furthermore, the use of 30 mg L-1 EOZO in transport water prevented an increase in hemoglobin and hematocrit, with minimal influences on other blood variables.

Environmental heterogeneity at two spatial scales affects litter diversity-decomposition relationships.

The effects of biodiversity on ecological processes have been experimentally evaluated mainly at the local scale under homogeneous conditions. To scale up experimentally based biodiversity-functioning relationships, there is an urgent need to understand how such relationships are affected by the environmental heterogeneity that characterizes larger spatial scales. Here, we tested the effects of an 800-m elevation gradient (a large-scale environmental factor) and forest habitat (a fine-scale factor) on litter diversity-decomposition relationships. To better understand local and landscape scale mechanisms, we partitioned net biodiversity effects into complementarity, selection, and insurance effects as applicable at each scale. We assembled different litter mixtures in aquatic microcosms that simulated natural tree holes, replicating mixtures across blocks nested within forest habitats (edge, interior) and elevations (low, mid, high). We found that net biodiversity and complementarity effects increased over the elevation gradient, with their strength modified by forest habitat and the identity of litter in mixtures. Complementarity effects at local and landscape scales were greatest for combinations of nutrient-rich and nutrient-poor litters, consistent with nutrient transfer mechanisms. By contrast, selection effects were consistently weak and negative at both scales. Selection effects at the landscape level were due mainly to nonrandom overyielding rather than spatial insurance effects. Our findings demonstrate that the mechanisms by which litter diversity affects decomposition are sensitive to environmental heterogeneity at multiple scales. This has implications for the scaling of biodiversity-ecosystem function relationships and suggests that future shifts in environmental conditions due to climate change or land use may impact the functioning of aquatic ecosystems.

Spatial versus spatio-temporal approaches for studying metacommunities: a multi-taxon analysis in Mediterranean and tropical temporary ponds.

Prior research on metacommunities has largely focused on snapshot surveys, often overlooking temporal dynamics. In this study, our aim was to compare the insights obtained from metacommunity analyses based on a spatial approach repeated over time, with a spatio-temporal approach that consolidates all data into a single model. We empirically assessed the influence of temporal variation in the environment and spatial connectivity on the structure of metacommunities in tropical and Mediterranean temporary ponds. Employing a standardized methodology across both regions, we surveyed multiple freshwater taxa in three time periods within the same hydrological year from multiple temporary ponds in each region. To evaluate how environmental, spatial and temporal influences vary between the two approaches, we used nonlinear variation partitioning analyses based on generalized additive models. Overall, this study underscores the importance of adopting spatio-temporal analytics to better understand the processes shaping metacommunities. While the spatial approach suggested that environmental factors had a greater influence, our spatio-temporal analysis revealed that spatial connectivity was the primary driver influencing metacommunity structure in both regions. Temporal effects were equally important as environmental effects, suggesting a significant role of ecological succession in metacommunity structure.

The first case of human invasion by Clinostomum complanatum in the European part of Russia.

The article presents a rare case of human invasion by the trematode Clinostomum complanatum in the European part of Russia. The diagnosis was established based on a parasitological study of flukes removed from the tonsils and pharynx of a 42-year-old woman, a resident of the Tambov region of Russia.

Climate warming shifts riverine macroinvertebrate communities to be more sensitive to chemical pollutants.

Freshwaters are highly threatened ecosystems that are vulnerable to chemical pollution and climate change. Freshwater taxa vary in their sensitivity to chemicals and changes in species composition can potentially affect the sensitivity of assemblages to chemical exposure. Here we explore the potential consequences of future climate change on the composition and sensitivity of freshwater macroinvertebrate assemblages to chemical stressors using the UK as a case study. Macroinvertebrate assemblages under end of century (2080-2100) and baseline (1980-2000) climate conditions were predicted for 608 UK sites for four climate scenarios corresponding to mean temperature changes of 1.28 to 3.78°C. Freshwater macroinvertebrate toxicity data were collated for 19 chemicals and the hierarchical species sensitivity distribution model was used to predict the sensitivity of untested taxa using relatedness within a Bayesian approach. All four future climate scenarios shifted assemblage compositions, increasing the prevalence of Mollusca, Crustacea and Oligochaeta species, and the insect taxa of Odonata, Chironomidae, and Baetidae species. Contrastingly, decreases were projected for Plecoptera, Ephemeroptera (except for Baetidae) and Coleoptera species. Shifts in taxonomic composition were associated with changes in the percentage of species at risk from chemical exposure. For the 3.78°C climate scenario, 76% of all assemblages became more sensitive to chemicals and for 18 of the 19 chemicals, the percentage of species at risk increased. Climate warming-induced increases in sensitivity were greatest for assemblages exposed to metals and were dependent on baseline assemblage composition, which varied spatially. Climate warming is predicted to result in changes in the use, environmental exposure and toxicity of chemicals. Here we show that, even in the absence of these climate-chemical interactions, shifts in species composition due to climate warming will increase chemical risk and that the impact of chemical pollution on freshwater macroinvertebrate biodiversity may double or quadruple by the end of the 21st century.

Occurrence of Pharmaceutical Micropollutants in Lake Nahuel Huapi, Argentine Patagonia.

Tourism is one of the most important activities for the economy of Nor Patagonia Argentina. In Bariloche City, located on the shores of Lake Nahuel Huapi, both the permanent and the temporary populations have increased significantly in recent decades, and this has not necessarily been accompanied by an improvement in sewage networks. Emerging micropollutants such as pharmaceutical compounds reach aquatic systems directly, in the absence of a domestic sewage network, or through effluents from wastewater treatment plants (WWTP), which do not efficiently remove these substances and represent a major threat to the environment. Therefore, the objective of our study was to monitor the presence of pharmaceutical compounds discharged both through wastewater effluents and diffusely from housing developments into Lake Nahuel Huapi. The results obtained demonstrate the presence of pharmaceuticals in Lake Nahuel Huapi with concentrations ranging from not detectable (ND) to 110.6 ng L-1 (caffeine). The highest pharmaceutical concentration recorded in WWTP influent corresponded to caffeine (41728 ng L-1), and the lowest concentration was paracetamol (18.8 ng L-1). The removal efficiency of pharmaceuticals in the WWTP was calculated, and ranged from 0% for carbamazepine to 66% for ciprofloxacin. This antibiotic showed the lowest % of attenuation (73%) in Lake Nahuel Huapi. These results on the occurrence of a wide variety of pharmaceuticals are the first generated in Patagonia, representing a regional baseline for this type of micropollutant and valuable information for the subsequent design of removal strategies for emerging pharmaceutical pollutants in surface water. Environ Toxicol Chem 2024;00:1-11. © 2024 SETAC.

Sensitivity of cholinesterases and carboxylesterases to pharmaceutical products in Tinca tinca.

Discharges to the aquatic environment of pharmaceuticals represent a hazard to the aquatic organisms. Subchronic assay with 17-alpha-ethinylestradiol (EE2) and in vitro essays with pharmaceuticals of environmental concern were conducted to examine the sensitivity of tissue acetylcholinesterase (AChE) and carboxylesterase (CbE) activities of Tinca tinca to them. Subchronic exposure to 17-alpha-EE2 caused significant effects on brain, liver, and muscle CbE, but no on AChE activities. Most of the pharmaceuticals tested in vitro were considered as weak inhibitors of tissular AChE activity. Depending on the tissues, some compounds were classified as moderate inhibitors of CbE activity while other were categorized as weak enzymatic inhibitors. An opposite trend was observed depending on the tissue, while brain and liver CbE activities were inhibited, the muscle CbE activity was induced. Changes experienced on enzymatic activities after exposure to pharmaceuticals might affect the physiological functions in which these enzymes are involved. In vitro exposure to 17-alpha-EE2 in tench could be an informative, but not a surrogate model to know the effect of this synthetic estrogen on AChE and CbE activities.

Native freshwater lake microbial community response to an in situ experimental dilbit spill.

With the increase in crude oil transport throughout Canada, the potential for spills into freshwater ecosystems has increased and additional research is needed in these sensitive environments. Large enclosures erected in a lake were used as mesocosms for this controlled experimental dilbit (diluted bitumen) spill under ambient environmental conditions. The microbial response to dilbit, the efficacy of standard remediation protocols on different shoreline types commonly found in Canadian freshwater lakes, including a testing of a shoreline washing agent were all evaluated. We found that the native microbial community did not undergo any significant shifts in composition after exposure to dilbit or the ensuing remediation treatments. Regardless of the treatment, sample type (soil, sediment, or water), or type of associated shoreline, the community remained relatively consistent over a 3-month monitoring period. Following this, metagenomic analysis of polycyclic aromatic and alkane hydrocarbon degradation mechanisms also showed that while many key genes identified in PAH and alkane biodegradation were present, their abundance did not change significantly over the course of the experiment. These results showed that the native microbial community present in a pristine freshwater lake has the prerequisite mechanisms for hydrocarbon degradation in place, and combined with standard remediation practices in use in Canada, has the genetic potential and resilience to potentially undertake bioremediation.

Influence of heavy Canadian crude oil on pristine freshwater boreal lake ecosystems in an experimental oil spill.

The overall impact of a crude oil spill into a pristine freshwater environment in Canada is largely unknown. To evaluate the impact on the native microbial community, a large-scale in situ model experimental spill was conducted to assess the potential role of the natural community to attenuate hydrocarbons. A small volume of conventional heavy crude oil (CHV) was introduced within contained mesocosm enclosures deployed on the shoreline of a freshwater lake. The oil was left to interact with the shoreline for 72 h and then free-floating oil was recovered using common oil spill response methods (i.e. freshwater flushing and capture on oleophilic absorptive media). Residual PAH concentrations returned to near pre-oiling concentrations within 2 months, while the microbial community composition across the water, soil, and sediment matrices of the enclosed oligotrophic freshwater ecosystems did not shift significantly over this period. Metagenomic analysis revealed key polycyclic aromatic and alkane degradation mechanisms also did not change in their relative abundance over the monitoring period. These trends suggest that for small spills (<2 L of oil per 15 m2 of surface freshwater), physical oil recovery reduces PAH concentrations to levels tolerated by the native microbial community. Additionally, the native microbial community present in the monitored pristine freshwater ecosystem possesses the appropriate hydrocarbon degradation mechanisms without prior challenge by hydrocarbon substrates. This study corroborated trends found previously (Kharey et al. 2024) toward freshwater hydrocarbon degradation in an environmentally relevant scale and conditions on the tolerance of residual hydrocarbons in situ.

Acute Toxicity Testing of Pink Salmon (Oncorhynchus gorbuscha) with the Tire Rubber-Derived Chemical 6PPD-Quinone.

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine-quinone (6PPD-quinone) is a widespread contaminant of emerging concern resulting from oxidation of 6PPD, which is an antidegradant substance added to tires. The recent identification of 6PPD-quinone as the cause of acute mortality in coho salmon has quickly motivated studies on 6PPD-quinone toxicity to other species. Subsequent findings have shown that 6PPD-quinone toxicity is highly species specific. Closely related species can differ widely in response to 6PPD-quinone from extremely sensitive to tolerant. Hence toxicity testing is currently the only way to establish whether a species exhibits 6PPD-quinone toxicity. We investigated the acute toxicity of 6PPD-quinone in pink salmon alevins (sac fry). This species has is the only Pacific salmon that so far has not been tested for 6PPD-quinone sensitivity. Fish were exposed in static water in eight treatments with initial concentrations ranging from 0.1 to 12.8 µg/L. Fish were observed for 48 h, and changes in concentrations of 6PPD-quinone were monitored throughout the experiment. No mortalities or substantial changes in behavior were recorded. Environ Toxicol Chem 2024;00:1-7. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

A Comparison of In Vitro Metabolic Clearance of Various Regulatory Fish Species Using Hepatic S9 Fractions.

Bioaccumulation predictions can be substantially improved by combining in vitro metabolic rate measurements derived from rainbow trout hepatocytes and/or hepatic S9 fractions with quantitative structure-activity relationship (QSAR) modeling approaches. Compared with in vivo testing guidelines Organisation for Economic Co-operation and Development (OECD) 305 and Office of Chemical Safety and Pollution Prevention (OCSPP; an office of the US Environmental Protection Agency) 850.1730, the recently adopted OECD test guidelines 319A and 319B are in vitro approaches that have the potential to provide a time- and cost-efficient, humane solution, reducing animal use while addressing uncertainties in bioaccumulation across species. The present study compares the hepatic clearance of the S9 subcellular fraction of rainbow trout, bluegill, common carp, fathead minnow, and largemouth bass, discerning potential differences in metabolism between different warm- and cold-water species. With refinements to the in vitro metabolic S9 assay for high-throughput analysis, we measured in vitro clearance rates of seven chemicals crossing multiple classes of chemistry and modes of action. We confirmed that data from rainbow trout liver S9 fraction metabolic rates can be utilized to predict rainbow trout bioconcentration factors using an in vitro to in vivo extrapolation model, as intended in the OECD 319B applicability domain per the bioaccumulation prediction. Also, we determined that OECD 319B can be applied to other species, modified according to their habitat, adaptations to feeding behavior, and environmental conditions (e.g., temperature). Once toxicokinetics for each species is better understood and appropriate models are developed, this method can be an excellent tool to determine hepatic clearance and potential bioaccumulation across species. The present study could be leveraged prior to or in place of initiating in vivo bioconcentration studies, thus optimizing selection of appropriate fish species. Environ Toxicol Chem 2024;00:1-16. © 2024 SETAC.