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Acute Copper Toxicity Displays a Nonmonotonic Relationship with Age Across the Medaka ( Oryzias latipes ) Life Span

The ability of an organism to cope with environmental stressors varies across the life span because of developmental stage–specific responses and age-related functional declines. In the present study, we examined the effect of age on acute copper toxicity in Japanese medaka (Oryzias latipes). We first determined the median lethal concentration (LC50) at 96 h for embryos, 7-day-old fry, and 6-month-old medaka. Embryos were exposed to 0, 15, 30, 60, 125, 250, and 500 ppb CuSO 4 through hatching. Fry were exposed to 0, 20, 50, 75, 100, 150, 250, and 500 ppb CuSO 4 for 96 h. Adult fish were exposed to 0, 100, 150, 200, 250, and 300 ppb CuSO 4 for 96 h. The 96-h LC50 was 804 ppb for embryos, 262 ppb for embryonically exposed larvae, 60.3 ppb for 7-day-old fry, and 226 ppb for adults. We then challenged cohorts of fish aged 2, 3, 5, 6, 7, 8, 9, 10, 11, 13, 14, 15, and 16 months with a 225-ppb CuSO 4 exposure to determine the acute toxicity across the life span. The fish exhibited a bimodal tolerance to copper, with tolerance peaking in 2- and 3-month-old fish and again at 10 and 11 months of age. Our data demonstrate that copper sensitivity is dynamic throughout the medaka life span and may be influenced by trade-offs with reproduction. Environ Toxicol Chem 2022;41:2999–3006. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

54 ENVIRONMENTAL SCIENCES↗

Transcriptomics-Based Points of Departure for Daphnia magna Exposed to 18 Per- and Polyfluoroalkyl Substances

Here, per- and polyfluoroalkyl substances (PFAS) represent a large group of contaminants of concern based on their widespread use, environmental persistence, and potential toxicity. Many traditional models for estimating toxicity, bioaccumulation, and other toxicological properties are not well suited for PFAS. Consequently, there is a need to generate hazard information for PFAS in an efficient and cost-effective manner. In the present study, Daphnia magna were exposed to multiple concentrations of 22 different PFAS for 24 h, in 96-well plate format. Following exposure, whole body RNA was extracted and extracts, each representing five exposed individuals, were subjected to RNA sequencing. Following analytical measurements to verify PFAS exposure concentrations, and quality control on processed cDNA libraries for sequencing, concentration-response modeling was applied to the data sets for 18 of the tested compounds, and the concentration at which a concerted molecular response occurred (transcriptomic point of departure; tPOD) was calculated. The tPODs, based on measured concentrations of PFAS, generally ranged from 0.03-0.58 µM (9.9-350 µg/L; interquartile range). In most cases, these concentrations were two orders of magnitude lower than similarly calculated tPODs for human cell lines exposed to PFAS. They were also lower than apical effect concentrations reported for seven PFAS for which some crustacean or invertebrate toxicity data were available, although there were a few exceptions. Despite being lower than most other available hazard benchmarks, Daphnia magna tPODs were, on average, four orders of magnitude greater than the maximum aqueous concentrations of PFAS measured in Great Lakes tributaries. Overall, this high throughput transcriptomics assay with Daphnia magna holds promise as a component of a tiered hazard evaluation strategy employing new approach methodologies.

59 BASIC BIOLOGICAL SCIENCES↗

Transient Copper Exposure During Embryogenesis and Temperature Affect Developmental Rate, Survival, and Fin Regeneration in Japanese Medaka ( Oryzias latipes )

Combined environmental stressors that an organism experiences can have both immediate and lasting consequences. In this paper we exposed medaka (Oryzias latipes) embryos to sublethal copper sulfate (CuSO 4 ) (0, 10, and 100 ppb) in combination with different rearing temperatures (27, 30, and 33°C) to assess acute and latent effects on development, growth, and regenerative capacity. Embryos exposed to CuSO 4 and/or higher temperatures hatched significantly earlier. At 4 months post-exposure, fish exposed to low levels of CuSO 4 during development had higher survival, while fish exposed to both 100 ppb CuSO 4 and 33°C temperatures had significantly lower survival. Additionally, a sex-specific effect of embryonic CuSO 4 exposure was observed as female mass decreased with increasing copper dose. We also assessed caudal fin regenerative capabilities in both embryo-exposed fish at 4 months of age and adult medaka that were exposed to 0, 10, and 100 ppb CuSO 4 at room temperature during a 14- day trial. Whereas fin regeneration was unaffected by adult exposure to copper, fish transiently exposed during embryogenesis displayed an initial increase in fin growth rate and an increased incidence of abnormal fin morphology following regrowth. Collectively, these data suggest that developmental copper exposure has the potential to exert long lasting impacts to organismal growth, survival, and function.

63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTAN↗

Comparing Transcriptomic Points of Departure to Apical Effect Concentrations For Larval Fathead Minnow Exposed to Chemicals with Four Different Modes Of Action

It is postulated that below a transcriptomic-based point of departure, adverse effects are unlikely to occur, thereby providing a chemical concentration to use in screening level hazard assessment. The present study extends previous work describing a high-throughput fathead minnow assay that can provide full transcriptomic data after exposure to a test chemical. One-day post-hatch fathead minnows were exposed to ten concentrations of three representatives of four chemical modes of action: organophosphates, ecdysone receptor agonists, plant photosystem II inhibitors, and estrogen receptor agonists for 24 h. Concentration response modeling was performed on whole body gene expression data from each exposure, using measured chemical concentrations when available. Transcriptomic points of departure in larval fathead minnow were lower than apical effect concentrations across fish species but not always lower than toxic effect concentrations in other aquatic taxa like crustaceans and insects. The point of departure was highly dependent on measured chemical concentration which were often lower than the nominal concentration. Differentially expressed genes between chemicals within modes of action were compared and often showed statistically significant overlap. In addition, reproducibility between identical exposures using a positive control chemical (CuSO 4 ) and variability associated with the transcriptomic point of departure using in silico sampling were considered. Results extend a transcriptomic-compatible fathead minnow high-throughput assay for possible use in ecological hazard screening.

59 BASIC BIOLOGICAL SCIENCES↗

Amphibian Dispersal Traits Not Impacted by Triclopyr Exposure during the Juvenile Stage

Exposure to agrochemicals can have lethal and sublethal effects on amphibians. Most toxicology studies only examine exposure during the aquatic larval stage. Survival of the juvenile stage is the most important for population persistence and it is critical to understand the potential impacts of exposure during this life stage. We investigated how short-term exposure to triclopyr, an herbicide commonly used in forestry management, might impact several juvenile traits. To determine if juveniles perceived exposure as an environmental stressor, we measured their release of corticosterone. We also examined dispersal traits by measuring foraging and hopping behavior. We found no evidence that exposure negatively impacted these traits or was a stressor. Our results provide a preliminary assessment of the potential impact of triclopyr on juvenile amphibians, but we recommend additional research on the effects of agrochemicals on juvenile amphibians.

59 BASIC BIOLOGICAL SCIENCES↗

Metal–Organic Frameworks for Per- and Polyfluoroalkyl Substances Treatment in Contaminated Water

Per- and polyfluoroalkyl substances (PFAS) are synthetic pollutants known for their chemical stability, environmental persistence, and toxicological risks. Their widespread use has led to extensive contamination, particularly in aquatic systems. Conventional treatment methods often face challenges such as high energy consumption and the production of secondary pollutants. Metal− organic frameworks (MOFs), with their high surface areas and tunable structures, have emerged as promising materials for PFAS remediation. This review summarizes recent progress in MOF-based PFAS adsorption and degradation, highlighting key frameworks such as MIL, UiO, and ZIF. Mechanistic insights into adsorption behavior and regeneration capabilities are discussed, along with the catalytic performance of MOF composites and postmodified systems in degradation pathways. The review concludes with design strategies for next-generation MOF materials aimed at efficient, sustainable PFAS removal under realistic conditions.

Adsorption↗

Yeast Deletomics to Uncover Gadolinium Toxicity Targets and Resistance Mechanisms

Among the rare earth elements (REEs), a crucial group of metals for high-technologies. Gadolinium (Gd) is the only REE intentionally injected to human patients. The use of Gd-based contrasting agents for magnetic resonance imaging (MRI) is the primary route for Gd direct exposure and accumulation in humans. Consequently, aquatic environments are increasingly exposed to Gd due to its excretion through the urinary tract of patients following an MRI examination. The increasing number of reports mentioning Gd toxicity, notably originating from medical applications of Gd, necessitates an improved risk–benefit assessment of Gd utilizations. To go beyond toxicological studies, unravelling the mechanistic impact of Gd on humans and the ecosystem requires the use of genome-wide approaches. We used functional deletomics, a robust method relying on the screening of a knock-out mutant library of Saccharomyces cerevisiae exposed to toxic concentrations of Gd. The analysis of Gd-resistant and -sensitive mutants highlighted the cell wall, endosomes and the vacuolar compartment as cellular hotspots involved in the Gd response. Furthermore, we identified endocytosis and vesicular trafficking pathways (ESCRT) as well as sphingolipids homeostasis as playing pivotal roles mediating Gd toxicity. Finally, tens of yeast genes with human orthologs linked to renal dysfunction were identified as Gd-responsive. Therefore, the molecular and cellular pathways involved in Gd toxicity and detoxification uncovered in this study underline the pleotropic consequences of the increasing exposure to this strategic metal.

59 BASIC BIOLOGICAL SCIENCES↗

Development of a New Aggregation Method to Remove Nanoplastics from the Ocean: Proof of Concept Using Mussel Exposure Tests

The overproduction and mismanagement of plastics has led to the accumulation of these materials in the environment, particularly in the marine ecosystem. Once in the environment, plastics break down and can acquire microscopic or even nanoscopic sizes. Given their sizes, microplastics (MPs) and nanoplastics (NPs) are hard to detect and remove from the aquatic environment, eventually interacting with marine organisms. This research mainly aimed to achieve the aggregation of micro- and nanoplastics (MNPs) to ease their removal from the marine environment. To this end, the size and stability of polystyrene (PS) MNPs were measured in synthetic seawater with the different components of the technology (ionic liquid and chitosan). The MPs were purchased in their plain form, while the NPs displayed amines on their surface (PS NP-NH2). The results showed that this technology promoted a significant aggregation of the PS NP-NH2, whereas, for the PS MPs, no conclusive results were found, indicating that the surface charge plays an essential role in the MNP aggregation process. Moreover, to investigate the toxicological potential of MNPs, a mussel species (M. galloprovincialis) was exposed to different concentrations of MPs and NPs, separately, with and without the technology. In this context, mussels were sampled after 7, 14, and 21 days of exposure, and the gills and digestive glands were collected for analysis of oxidative stress biomarkers and histological observations. In general, the results indicate that MNPs trigger the production of reactive oxygen species (ROS) in mussels and induce oxidative stress, making gills the most affected organ. Yet, when the technology was applied in moderate concentrations, NPs showed adverse effects in mussels. The histological analysis showed no evidence of MNPs in the gill’s tissues.

Cid-Samamed, Antonio (ORCID:0000000205073394)↗

Atmospheric emission of nanoplastics from sewer pipe repairs

Nanoplastic particles are inadequately characterized environmental pollutants with significant adverse effects on aquatic and atmospheric systems, causing drastic impacts on human health through inhalation, ingestion, and skin penetration. At present, it is explicitly assumed that environmental nanoplastics (EnvNP) are weathering fragments of microplastic or larger plastic debris discharged into terrestrial and aquatic environments, while atmospheric occurrence of EnvNP is solely attributed to aerosolization by wind and other mechanical forces. However, sources and emissions of unintended EnvNP are poorly understood and are therefore largely unaccounted for in various risk assessments. In this work, we show that large quantities of EnvNP may be directly emitted into the atmosphere as steam laden waste components discharged from a commonly used technology to repair sewer pipes in urban areas. Comprehensive chemical analysis of the discharged waste condensate reveals abundant presence of insoluble colloids, which after drying form solid organic particles with composition and viscosity consistent with EnvNP. We suggest that airborne emissions of EnvNP from these globally used sewer repair practices may be prevalent in highly populated urban areas. Airborne emissions of these EnvNP particles may have important implications on air quality and toxicological levels in large urban areas that need to be mitigated.

77 NANOSCIENCE AND NANOTECHNOLOGY↗

Spatiotemporal Changes in Trace Metal Bioavailability in the Sediment Pore water of a Constructed Wetland Using Passive Pore water Samplers

Sediments in aquatic systems often act as a major sink for contaminants. Diffusive gradient in thin films (DGTs) and in situ equilibrium dialysis samplers (peepers) are two major in situ pore water sampling devices that overcome the problems associated with conventional pore water sampling methods. In the present study, DGTs and peepers were used to study the spatial and seasonal effects (cool months, October–February; warm months, May–September) on metal bioavailability in the H‐02 constructed wetland and the sink versus source role of the sediments by calculating the metal resupply capacity. Data showed similar seasonal trends in metal concentrations using passive samplers, peepers, and DGTs. Pooled Cu and Zn concentrations measured using DGTs were lower in warm months (1.67 ± 1.50 and 2.62 ± 0.68 μg L –1 , respectively, p < 0.001) versus in cool months (2.12 ± 0.65 and 5.58 ± 1.33 μg L –1 , respectively, p < 0.001; mean ± 95% confidence interval). Sulfate (SO 4 2− ) concentrations were significantly (p = 0.0139) lower in warm months (averaged at 0.22 ± 0.05 mg L –1 ) compared to in cool months (0.16 ± 0.05 mg L –1 ). The increase in SO 4 2− concentration is an indicator of the lower activity of sulfate‐reducing bacteria, which need SO 4 2− during anaerobic respiration, in which SO 4 2− is reduced to sulfide (S 2– ) that forms insoluble salts with Cu and Zn, which could partially explain the higher bioavailability of these metals in the cool season. Metal resupply capacity of the sediments was mostly <0.2 for Cu and Zn. Taken together, the H0‐2 wetland sediments mostly acted as a sink to both Cu and Zn over the course of the present study. Environ Toxicol Chem 2023;42:2726–2736. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

54 ENVIRONMENTAL SCIENCES↗

MIBiG 4.0: advancing biosynthetic gene cluster curation through global collaboration

Specialized or secondary metabolites are small molecules of biological origin, often showing potent biological activities with applications in agriculture, engineering and medicine. Usually, the biosynthesis of these natural products is governed by sets of co-regulated and physically clustered genes known as biosynthetic gene clusters (BGCs). To share information about BGCs in a standardized and machine-readable way, the Minimum Information about a Biosynthetic Gene cluster (MIBiG) data standard and repository was initiated in 2015. Since its conception, MIBiG has been regularly updated to expand data coverage and remain up to date with innovations in natural product research. Here, we describe MIBiG version 4.0, an extensive update to the data repository and the underlying data standard. In a massive community annotation effort, 267 contributors performed 8304 edits, creating 557 new entries and modifying 590 existing entries, resulting in a new total of 3059 curated entries in MIBiG. Particular attention was paid to ensuring high data quality, with automated data validation using a newly developed custom submission portal prototype, paired with a novel peer-reviewing model. MIBiG 4.0 also takes steps towards a rolling release model and a broader involvement of the scientific community. MIBiG 4.0 is accessible online at https://mibig.secondarymetabolites.org/.

59 BASIC BIOLOGICAL SCIENCES↗

Mixture Effects of Per- and Polyfluoroalkyl Substances on Embryonic and Larval Sheepshead Minnows ( Cyprinodon variegatus )

Per- and polyfluoroalkyl substances (PFAS) are ubiquitous and persistent environmental contaminants originating from many everyday products. Perfluorooctane sulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) are two PFAS that are commonly found at high concentrations in aquatic environments. Both chemicals have previously been shown to be toxic to fish, as well as having complex and largely uncharacterized mixture effects. However, limited information is available on marine and estuarine species. In this study, embryonic and larval sheepshead minnows (Cyprinodon variegatus) were exposed to several PFAS mixtures to assess lethal and sublethal effects. PFOS alone was acutely toxic to larvae, with a 96 h LC50 of 1.97 mg/L (1.64–2.16). PFOS + PFOA resulted in a larval LC 50 of 3.10 (2.62–3.79) mg/L, suggesting an antagonistic effect. These observations were supported by significant reductions in malondialdehyde (105% ± 3.25) and increases in reduced glutathione concentrations (43.8% ± 1.78) in PFOS + PFOA exposures compared to PFOS-only treatments, indicating reduced oxidative stress. While PFOA reduced PFOS-induced mortality (97.0% ± 3.03), perfluorohexanoic acid (PFHxA) and perfluorobutanoic acid (PFBA) did not. PFOS alone did not affect expression of peroxisome proliferator-activated receptor alpha (pparα) but significantly upregulated apolipoprotein A4 (apoa4) (112.4% ± 17.8), a downstream product of pparα, while none of the other individually tested PFAS affected apoa4 expression. These findings suggest that there are antagonistic interactions between PFOA and PFOS that may reduce mixture toxicity in larval sheepshead minnows through reduced oxidative stress. Elucidating mechanisms of toxicity and interactions between PFAS will aid environmental regulation and management of these ubiquitous pollutants.

59 BASIC BIOLOGICAL SCIENCES↗

Accumulation of trace elements in soil and fauna within a site historically contaminated with coal combustion residues

Legacy contaminants tied to energy production are a worldwide concern. Coal combustion residues (CCRs) contain high concentrations of potentially toxic trace elements such as arsenic (As), mercury (Hg), and selenium (Se), which can persist for decades after initial contamination. Coal combustion residue disposal methods, including aquatic settling basins and landfills, can facilitate environmental exposure through intentional and accidental releases. Wildlife exposed to CCRs can experience numerous deleterious effects, such as on development, reproduction, and survival. In the current study, we quantified and compared concentrations of As, Hg, Se, and strontium (Sr) within soils and target fauna (three vertebrate and three invertebrate taxa) from a CCR-contaminated site and a reference site within the U.S. Department of Energy’s Savannah River Site, South Carolina, United States. Our objectives were to (1) compare current concentrations of tested elements in soil and resident fauna to levels from our reference site, (2) assess natural attenuation of elements in soils by comparing current concentrations to historic levels, and (3) evaluate the biomagnification potential of the elements measured via body burden and trophic position correlations among fauna. Here, element concentrations were higher in contaminated soils than reference soils; however, concentrations in 2022 were unchanged from concentrations measured in 2003, suggesting no natural attenuation of tested elements. Additionally, target fauna had elevated As, Se, and Sr levels in comparison to reference samples. A positive correlation was observed in southern toads between Sr concentrations and trophic position, as assessed by nitrogen stable isotope ratios, suggesting potential for biomagnification of Sr within our study system. Collectively, our results demonstrate that legacy contaminants are still present and bioaccumulate in a diversity of taxa in a CCR-contaminated site that has not received effluents in more than 50 years, suggesting monitoring programs in CCR-contaminated sites should be maintained long term in the absence of remediation.

63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTAN↗

Metallothionein expression in Eastern mosquitofish ( Gambusia holbrooki ) exposed to 137 Cs and lead: implications for using metallothionein as a metal exposure biomarker

Given the range of threats facing aquatic ecosystems, it is critical we have monitoring tools to quickly and efficiently identify stressors within ecosystems. Metallothionein’s (MTs) are a group of proteins widely used as biomarkers of metal exposure; however, their suitability has been questioned due to their influenceable nature and ability to be induced by various stressors. Through our study, we sought to (1) evaluate induction of MT in eastern mosquitofish (Gambusia holbrooki) exposed to a radionuclide ( 137 Cs) and metal (Pb), (2) assess the influence of in situ 137 Cs pre-exposure on Pb-induced MT expression, and (3) quantify changes in MT concentrations during Pb depuration. We conducted our experiment in three phases using mosquitofish as a model species: Phase I—pre-exposure to 137 Cs in the field, Phase II—exposure to Pb in the lab immediately following 137 Cs exposure, and Phase III—depuration in the laboratory. We quantified MT concentrations over time and assessed differences among treatments. We did not find in situ exposure of mosquitofish to 137 Cs to induce MT, nor did pre-exposure to 137 Cs appear to influence induction of MT following Pb exposure. Metallothionein was rapidly induced in mosquitofish following Pb exposure; however, MT induction was also observed in control fish. Additionally, there was no apparent relationship between MT and Pb body burdens. During depuration, we found MT concentrations to be highly variable among fish demonstrating no clear response. Furthermore, our study demonstrates the high variability exhibited by MT, the need for further validation of MT as a metal exposure biomarker, and the value in additional research on these proteins under varying exposure scenarios, particularly environmentally realistic concentrations.

54 ENVIRONMENTAL SCIENCES↗