Ecotoxicology and Environmental Safety

Previously, we have demonstrated that certain ferric salts common in Archean waters, such as iron trichloride and ferric ammonium citrate, can protect liposomes from destruction by short-wavelength UVC light. In this study, we investigate the propagation of 254 nm UV radiation through aqueous FeCl3 solutions and its interactions with liposomes. We then consider these findings in the context of early Earth UV environment, discuss their implications for our hypothesis of the Darwinian evolution of liposomes, and integrate them with our previous experimental results.

Wildfire smoke has increasingly affected air quality across North America, raising concerns about the health effects of fine particulate matter (PM2.5) exposure, including potential impacts on brain health. However, relatively few studies have characterized personal PM2.5 exposure during these events using wearable monitoring. We examined daily personal PM2.5 concentrations during wildfire smoke episodes in southeast Michigan alongside neighborhood outdoor PM2.5 estimates. Four participants (one adolescent and three adults) wore AirBeam3 personal monitors during ongoing studies. Neighborhood outdoor PM2.5 was estimated using the average of three nearest PurpleAir outdoor air quality sensors, and wildfire smoke days were identified using state air quality advisories. Group-level descriptive statistics summarized personal and neighborhood outdoor PM2.5 and self-reported time spent outdoors. Exploratory within-participant analyses quantified associations between neighborhood outdoor and personal PM2.5 concentrations on smoke and non-smoke days. Neighborhood outdoor daily PM2.5 concentrations were higher during wildfire smoke days than non-smoke days (87.6 + 80.2 vs. 12.3 + 7.0 {micro}g/m3). Personal PM2.5 concentrations were more than five times higher during wildfire smoke days (28.2 + 20.0 vs. 5.1 + 4.7 {micro}g/m3) than non-smoke days. Within participants, every 10 {micro}g/m3 increase in neighborhood PM2.5 was associated with 2.3 {micro}g/m3 increase in personal PM2.5 concentrations. Wearable PM2.5 monitoring captured elevated personal exposures while providing individual-level exposure information beyond neighborhood outdoor air quality estimates. These findings demonstrate that wearable monitoring complements neighborhood air quality measurements by capturing individual-level exposure, providing a more comprehensive assessment of real-world wildfire smoke exposure for future studies examining the effects on brain health.

High concentrations of steroidal hormone compounds are a growing source of concern for environmental pollution in aquatic ecosystems. In this study, we examine the effects of two estrogenic compounds (estriol and 17-ethinylestradiol) on fitness traits in the aquatic microcrustacean, Daphnia magna, a key bioindicator species for toxicology studies. The impacts were compared of two forms representing a natural and synthetic estrogenic compound. Growth and reproduction traits were assayed by exposing Daphnia to each estrogen type at four concentrations reflecting potential environmental exposure conditions up to acute toxicity levels (ranging from 0.1 - 50 {micro}g/L). Assaying the effects at a variety of concentrations is important given that it is known that hormone exposures can often result in non-monotonic responses. Both forms of estrogen impact a subset of the traits assessed, in some cases leading to beneficial changes and others causing harm. Estriol, the naturally-occurring estrogen, and EE2, the synthetic version, at high doses shift fitness traits in opposite directions such as adult growth rate as do at low doses for fecundity. In conclusion, our results support the need to assay a wide array of traits using multiple forms of steroidal hormones at a range of doses in order to assess non-monotonic patterns and their impact on an organismal fitness. In particular, assays that extend beyond the conventional measurements of lethality during acute exposure windows will be essential for understanding the impact of increased levels of hormone pollution on aquatic organisms and ecosystem health.

Concerns raised by ubiquitate plastic contamination are urging to develop alternative materials. In the recent years, bio-sourced polymers also coined as "bioplastics" have been proposed to mitigate plastic pollution while meeting industrial and commercial expectations. Like petro-sourced plastics, they are expected to break-down in the environment into fragments down to sub-micron size. However, only scarce data are available on the impacts of such biosourced nanoplastics once released into the environment. This study examines the effects on aquatic species of model nanoplastics made from several bio-sourced polymers (Bio-NPs) that are either already on market (PHA, PLA, PA11) or still under development (NIPU, PCAR). We exposed three species of micro-algae (at 10, 100, and 1000 {micro}g/L, for 24 and 48 hours, and one week) to test the effects of Bio-NPs on algal growth. We also exposed freshwater bivalves C. fluminea (at 1, 10 and 100 {micro}g/L, for one week) to test the filtration activity and gene expressions in response to Bio-NPs exposure. All five Bio-NPs tested generated growth inhibitions in at least one of the three algae tested. PLA and PA11 were the most deleterious ones for algal growth among the five tested Bio-NPs. The highest growth inhibitions were observed on the fresh water species D. subspicatus. Each Bio-NP tested resulted in significant decreases of the filtration rates of C. fluminea. PHA impaired filtration at the lowest concentrations tested (1 {micro}g/L), whereas PCAR, PA11 and NIPU led to significant effects only at higher concentrations (10 and 100 {micro}g/L). The results from gene expressions in C. fluminea showed strong inductions of all gene functions tested for all the five bio-NPs tested. These Bio-NPs triggered endocytosis and detoxification mechanisms. They impaired the mitochondrial metabolism and triggered oxidative stress and immune responses. PA11, NIPU and PHA exposures resulted in the strongest gene regulations. The present study brings brand new findings about a kind of nanoplastics that may be released into the environment in a near future as the use of bioplastics is growing fast. It will help better understanding the impacts of such fragmented bioplastic NPs on aquatic species.

Per- and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants associated with developmental abnormalities and adverse health outcomes, yet it remains unclear whether PFAS exposure imposes novel transcriptional programs during development or perturbs endogenous developmental processes. Here, we continuously exposed African killifish (Nothobranchius furzeri) to an environmentally relevant concentration of perfluorooctanoic acid (PFOA) from egg laying through juvenile development to mimic prenatal-to-adolescent exposure and performed whole-transcriptome sequencing at two developmental stages. Despite four weeks of embryonic exposure, newly hatched juveniles, approximately equivalent to human infants, exhibited remarkably limited transcriptional responses, with only a few differentially expressed genes identified. In contrast, older juveniles, equivalent to human adolescents, exposed for eight weeks displayed a dramatic expansion of transcriptional perturbation, with approximately 30-fold more differentially expressed genes spanning pathways involved in cell-cycle regulation, endocrine signaling, immune function, oxidative stress, and lipid metabolism. Unexpectedly, more than half of the PFOA-induced genes were the same genes that normally increase during juvenile maturation, representing a highly significant enrichment of the endogenous developmental program. These findings indicate that the embryonic transcriptome is largely buffered against chronic PFOA exposure, whereas post-hatch stages exhibit heightened vulnerability. Rather than inducing a distinct toxicological state, PFOA predominantly acted by amplifying existing developmental gene regulatory programs. Our results identify the juvenile stage following hatching, equivalent to human neonatal and adolescent developmental stages, as a critical window of PFAS susceptibility and suggest that environmental contaminants may exert their effects by exaggerating normal developmental trajectories, with potential consequences for growth, maturation, and long-term health.

Air pollution is a growing public health concern. The developing fetus is particularly vulnerable, with exposure during pregnancy linked to negative developmental health outcomes. Teratogenic studies rely on the use of model organisms, such as the chick embryo, a well- established model of human development. However, existing protocols for the exposure of chick embryos to gaseous and aerosol pollutants have financial and technical limitations. Here, we present a novel, non-invasive method for the long-term exposure of chicken embryos to a gaseous toxin, carbon monoxide (CO). Exposure is performed inside airtight incubation boxes, which can be used in a standard laboratory incubator. We demonstrate reliable dosing of precise internal CO concentrations up to 200ppm, using a simple volumetric approach. Following optimization of key incubation parameters, temperature and turning frequency, we determined the impact of the system on embryo viability and development. Closed box incubation caused minor developmental delay but had no effect on chick embryo viability. Internal oxygen concentrations remained above hypoxic levels. No significant effects of exposure up to 200ppm CO were observed on embryo viability, weight or developmental stage. In conclusion, we present a non-invasive, affordable, accessible and technically straightforward exposure method for air pollutant teratogenicity studies. This method can be applied to other model systems and organisms beyond the chick embryo as well as to other gaseous and aerosol toxins. Thus, the system offers a suitable platform for future research on teratogenic doses, mechanisms and effects of air pollutants.

Polycyclic aromatic hydrocarbons (PAHs) are hazardous organic contaminants for which microbial bioaugmentation is a promising remediation strategy, but poor persistence of introduced microorganisms can limit efficacy. Encapsulation may improve persistence, yet the influence of capsule design, microbial species, and environmental conditions on performance remains poorly understood. We evaluated alginate encapsulation of the PAH-degrading bacteria Pseudomonas putida and Novosphingobium aromaticivorans across nutrient conditions and capsule formulations. Encapsulation effects varied by species and medium, influencing growth rate, maximum cell density, overall growth, and lag time; notably, encapsulation shortened lag time of N. aromaticivorans in sRB15 medium (36.9 h to 3.9-5.3 h). Enumeration methods also affected apparent cell recovery. After 8 weeks, encapsulation had no significant effect on P. putida but resulted in increased concentrations of N. aromaticivorans relative to planktonic cultures (1.22 x 10; vs. 2.05 x 10; CFU/mL). Capsule composition further influenced cell retention: increasing alginate approximately doubled capsule-associated cell concentrations, while chitosan coatings reduced cell concentrations within capsules without affecting external concentrations. These findings demonstrate that the benefits of encapsulation are species- and environment-dependent and that capsule formulation can be tuned to influence bacterial persistence and release, informing the design of encapsulated inoculants for bioaugmentation applications.

Alternaria toxins are naturally occurring food contaminants with limited and often inconsistent genotoxicity and mutagenicity data. Within the European Partnership for the Assessment of Risks from Chemicals (PARC), an OECD-aligned in vitro testing strategy was applied to fill existing data gaps and to characterize the genotoxic potential of major Alternaria toxins using high-purity test materials. Mutagenicity was assessed using bacterial reverse mutation test (OECD TG 471) and SOS/umu assay, while chromosomal damage was assessed using the in vitro micronucleus (MN) assay (OECD TG 487) in TK6 and HepG2 cells, complemented by fluorescence in situ hybridization (FISH) and {gamma}H2AX assay in HepaRG cells. Alternariol (AOH), alternariol monomethyl ether (AME), and altertoxin-I (ATX-I) showed clear mutagenicity in bacteria, whereas altenuene (ALT), tenuazonic acid (TeA), and tentoxin (TEN) were negative under the tested conditions. In mammalian cells, AOH, AME, and ATX-I induced MN formation in TK6 cells at concentrations [≥]5.5 {micro}M, [≥]2.5 {micro}M, and [≥]0.21 {micro}M, respectively, with FISH analysis supporting a clastogenic mode of action. In HepG2 cells, all tested toxins induced chromosomal damage, with effect threshold ranging from [≥]6.25 {micro}M (AOH) to [≥]50 {micro}M (TeA). {gamma}H2AX induction confirmed DNA damage for AOH and ATX-I, and at higher concentrations for TeA (1000 {micro}M). Overall, the data indicate clear in vitro genotoxic potential for AOH, AME, and ATX-I and provide evidence of chromosomal damage for ALT, TEN, and TeA, thereby reducing critical data gaps for hazard assessment.

Metals and per- and polyfluoroalkyl substances (PFAS) represent a significant environmental concern, yet their association with epigenetic age acceleration (EAA) remain largely understudied in marine mammals. Here, associations between EAA in common dolphins (Delphinus delphis) and life history (sex and sexual maturity), trace metals, and PFAS were investigated. EAA was calculated as the residual in the regression of epigenetic age vs chronological age, hence providing a direct measure of the deviation of the epigenetic age of an organism (positive or negative) by comparison with expectation, given their actual chronological age. Sixteen trace elements were quantified in hepatic and renal tissues (n = 53). In addition, 28 PFAS were quantified in hepatic tissue (n = 58). Associations between EAA and explanatory variables were assessed using regression-based and multivariate modelling approaches (linear models and canonical analysis of principal coordinates). No effect of sex was observed, although sexual maturity did significantly increase EAA. Exposure to metals was significantly associated with EAA, explaining 55.4% of the variation, with hepatic metals (Se, Zn, Cu, Al, Mn) driving this relationship. Although EAA was not significantly related to the total PFAS exposure overall, a subset of PFAS variables (PFBA, PFDA, PFHxS-B, PFNA) showed significant association with EAA after adjusting for sex and sexual maturity. Together, these subsets of metal and PFAS variables, in addition to the selenium-to-mercury (Se:Hg) molar ratio, explained 66.7% of the variation in EAA. Our results identify sexual maturity and specific contaminant mixtures as key potential drivers of EAA in common dolphins, highlighting the possible use of EAA as a biomarker of environmental and physiological stress in marine mammals.

Background: Exercise training is a cornerstone intervention for cardiovascular disease, yet large cohort studies have reported attenuation of physical activity benefits at elevated air pollution concentrations, creating uncertainty around exercise prescription in polluted settings where cardiovascular disease burden is greatest. Objectives: To determine whether ambient PM2.5 concentration modifies the cardiovascular benefits of structured exercise training, using a global sample of trials spanning a >100-fold pollution gradient. Methods: We conducted a systematic review and multilevel meta-analysis of exercise training interventions reporting pre-post changes in systolic blood pressure (SBP), diastolic blood pressure (DBP), peak oxygen uptake (VO2Max), or resting heart rate (HR) in adults. Annual ambient PM2.5 concentrations (3.5-283 g/m3) were assigned to each study location from CAMS ERA5 reanalysis data. Three-level random-effects models with cluster-robust variance estimation accounted for arms nested within studies. PM2.5 meta-regression was conducted unadjusted and adjusted for world region, exercise mode, trial duration, and health condition, with subgroup analyses by exercise mode and hypertension status. Results: Across 465 studies (27,629 participants), exercise training produced clinically meaningful benefits for all outcomes (SBP - mmHg, DBP - mmHg, VO2Max +3.2 ml/kg/min, HR - bpm; all p < 0.001), with benefits consistently larger in higher-pollution settings. Hypertensive participants showed the greatest improvements, particularly from aerobic exercise (SBP standardised mean difference 0.396 in the Low vs 1.020 in the High PM2.5 stratum). Although aerobic and resistance training participants experience similar chronic ambient PM2.5 exposure, only aerobic exercise showed a stratum gradient (interaction p = 0.074). Discussion: Exercise training delivers clinically meaningful cardiovascular benefits at every pollution level tested. The larger benefits observed in higher-pollution settings reflect the greater cardiovascular risk burden of those populations, and hypertensive patients stand to gain the most, particularly from aerobic exercise.

Depleted uranium (DU) is an environmental contaminant with a 30 g/L (ppb; parts per billion) EPA maximum contaminant level (MCL) for drinking water. The mining of uranium and use of DU in modern weapons underly human exposure that disproportionally impacts military and tribal communities in the United States. Uranium's radiotoxic characteristics are understood, but its chemical hazards much less so. In zebrafish (Danio rerio) and human cell cultures we test the hypothesis that exposure to DU negatively impacts cellular function and development through disruption of mitochondrial metabolism. Using a novel shrapnel model with TEM/SEM+EDS, we showed uranium microparticles caused proximity-dependent mitochondrial disruption. In waterborne exposure paradigms, larval movement was reduced and hatching delayed as a result of reduced movement and not enzyme deficiencies in response to 18 ppb DU, below the MCL. Increased DNA damage accumulation was detected in exposed larva and cells. DNA-damage quantitative PCR of DU-exposed larvae showed increased damage in the ahr1 locus (nuclear gene) and decreased mitochondrial DNA (mtDNA) copy number, but mtDNA damage levels varied across experiments. Mitochondrial function was assessed using a resazurin-based assay in the presence and absence of antioxidants and showed diminished cytoplasmic reductive capacity. DU exposure alone did not enrich antioxidant gene expression, contrasting with arsenic exposure, a known ROS-inducer and Nrf2-activator. Sulforaphane (SFN), a potent Nrf2-activator, did not blunt the effects of DU exposure, despite activation of antioxidant response element (ARE) genes (gstp and gss), but did blunt the effects of arsenic exposure. The most enriched transcript in DU-exposed larvae coded for slingshot protein phosphatase (ssh), further exploration revealed ssh1b as the zebrafish-specific ortholog activated in response to DU, and inhibition using an identified SSH1 inhibitor, Sennoside A, partially rescued the metabolic and hatching defects observed. Our data points to a cytotoxic mechanism in which DU disrupts mitochondrial function through ssh1b enrichment that impairs normal mitophagy, leading to decreased cellular reductive potential independent of either ROS production or ARE-activation. Our results suggest that health impacts from DU exposure may be directly linked to impaired mitochondrial functions.

Food security is one of the major ongoing global anthropic challenges, driving the intensification of agricultural systems notably through the widespread use of phyto-pharmaceutical products (PPPs). Although effective for maximizing production yields through pest control, PPP contamination is pervasive across the entire agroecosystem, raising concerns about human and non-target species exposure and health effects. Organic farming has therefore been promoted as a sustainable alternative to ensure human food security and mitigate these impacts. Yet, the pathways of PPP non-target organisms contamination are insufficiently understood. While most research has focused on ingestion as the primary route of exposure, increasing evidence suggests more complex contamination dynamics. In this study, we investigated the role of food intake in shaping PPP contamination profiles in semi-captive grey partridges (Perdix perdix). To do so, 80 birds were housed in open aviaries within an agricultural landscape and fed either conventional or organic grains over a five-month period. After drawing their PPP blood profiles at the end of the exposure period, we assessed their loads and compared the pesticide prevalence and abundance between food treatments. We found similar quantity and diversity of PPP residues in blood from conventionally- and organically-fed partridges, with few differences in specific PPP compounds. Overall, these findings suggest that food intake is not necessarily the only source of contamination for non-target organisms, with alternative pathways (e.g., inhalation, dermal contact, soil and water exposure) likely playing a substantial role. Integrating multi-pathway contamination monitoring across trophic levels within a One Health framework would now be essential to better understand and mitigate PPP contamination risks. Submitted to PCI: Ecotoxicology and Environmental Chemistry

Endocrine-disrupting chemicals can target ovaries and interfere with key milestones of reproduction. Previously, we found that mixtures of the chemical exposome measured in follicular fluid (FF) were cumulatively associated with lower oocyte yield. Because ovaries age faster than many other organs, our current aim was to evaluate associations of FF chemical mixtures with epigenetic age acceleration and epigenetic pathways in FF cells. FF was collected during oocyte retrieval from 76 patients undergoing assisted reproduction in Atlanta. The exposome was measured using untargeted high-resolution mass spectrometry with gas (GC) and liquid (LC) chromatography. Weighted quantile sum (WQS-RS) indices were constructed for three mixtures of chemicals in association with oocyte yield, separated by instrument configuration (GC, LC-HILIC, LC-C18). DNA methylation was measured from the cellular component of FF using Illumina MethylationEPIC BeadChip, with age acceleration based on the GrimAge clock. Regression and pathway enrichment analyses elucidated relationships between chemical exposures or mixture indices and epigenetic markers, adjusted for age and technical covariates. All three chemical mixture indices were associated with epigenetic age acceleration in FF (p<0.05). For example, a standard-deviation increase in the GC-detected mixture was associated with 0.23 standard-deviations higher accelerated aging (95% CI: 0.0058-0.45; p=0.048). Twenty-seven frequently detected chemicals, including benzo[a]pyrene, plasticizers, flame retardants, forever chemicals, and pesticides, were associated with epigenetic pathways related to ovarian follicle growth and hormone signaling (p<0.05; six under false discovery rate<5%). In summary, environmental chemicals may accumulate in ovaries, contribute to accelerated epigenetic aging of ovarian somatic cells, and potentially affect follicle development.

Tenuazonic acid (TeA) is an emerging Alternaria mycotoxin frequently detected in food and feed commodities, raising concerns about its toxicological relevance. Chronic oral exposure to TeA has been reported to induce dysplastic alterations in the esophageal mucosa of mice, while human biomonitoring data indicate an association between TeA exposure and esophageal cancer, although a causal relationship has not yet been established. At a mechanistic level, the effects of TeA in esophageal cells remain poorly characterized. Therefore, this study investigated the impact of TeA on cytotoxicity, oxidative stress, DNA damage, mitochondrial homeostasis, cell-cycle distribution and transcriptomic stress responses in human esophageal KYSE-510 cells. TeA induced a concentration-dependent reduction in metabolic activity and total protein content after 24 h exposure to 0.1-100 M. Significant cytotoxicity was measured starting from 20 M. At sub-cytotoxic concentrations, TeA triggered rapid ROS formation within 5-30 min exposure and induced formamidopyrimidine-DNA glycosylase (FPG) sensitive DNA damage after 1 h exposure (5-7.5 M), indicating oxidative DNA lesions. In addition, TeA altered mitochondrial morphology after 4 h exposure at 7.5 M, manifested by shrinkage of the mitochondrial network area and perinuclear redistribution, while mitochondrial respiration showed only a non-significant tendency towards reduced respiratory capacity. RNA sequencing after 6 h exposure to 10 M TeA revealed oxidative stress-associated transcriptional changes, impaired antioxidant and stress-adaptive responses, and p53-associated stress signaling. Furthermore, TeA induced significant G2/M phase accumulation after 24 h exposure to 1-10 M.

Fine particulate matter (PM2.5) was found to be associated with elevated blood lipids, but fewer studies have examined the associations with specific constituents of PM2.5. We studied the associations between exposure to annual PM2.5 and its 14 constituents, and repeated blood lipid measurements among general responders enrolled in the World Trade Center Health Program between 2003 and 2019 (n = 44,876). We used generalized additive mixed effect models to investigate the single-pollutant associations with repeated measures of blood total cholesterol (TC), high and low-density lipoprotein (HDL-C and LDL-C) levels. We then used linear generalized weighted quantile sum regression with a random intercept for participant ID to account for the clustering of repeated measures and evaluate the combined associations with the component mixture. A decile increase in the mixture of 14 PM2.5 chemical components was associated with 0.375 mg/dL increase in TC levels (95% confidence Interval (CI): 0.174-0.577) and 0.302 mg/dL increase in LDL-C (95% CI: 0.063, 0.540). Lead, organic carbon, and iron were major drivers of both associations. Component-specific models also show higher TC and LDL levels associated with interquartile range increases in organic carbon (0.472, 95% CI [0.027, 0.918] and 0.648 95% CI [0.136, 1.160]) and iron exposure (1.081, 95% CI [0.630, 1.532] and 0.748, 95% CI [0.318, 1.178]). In conclusion, we found PM2.5 exposure to be associated with elevated lipid levels. The associations differed by PM2.5 composition, highlighting organic carbon, lead, and iron and major drivers. These findings are highly significant for a population exposed to extreme air pollution event and susceptible to lipid alterations that might trigger cardiovascular events.

Here, oyster larval developmental abnormalities within a New England hatchery were linked to a common UV sterilization technique that has been used for over 20 years. Because of the known link between phytoplankton oxylipins and egg mortality in copepods, we hypothesized that UV pretreatment of seawater results in the production of oxylipins that inhibit larval digestion of microalgae. We used lipidomics to observe changes in the organic compounds dissolved in estuarine seawater when filtered and when filtered and pretreated with UV. UV treatment resulted in an increase in the relative abundance of oxylipins associated with cyanobacteria, fungi, and macroalgae in 2020, whereas oxylipins typically produced by diatoms were more abundant in the UV treatments from 2021. Oxylipin concentrations were higher in 2020, when the hatchery reported the most severe problems with larval development. Removing the UV step allowed continued larval production in both years. However, the lack of UV sterilization led to an unidentified bacterial pathogen in 2021, which nearly decimated the overall seasonal production of oyster seed. To follow up in a more controlled environment, the larvae were exposed to exogenous oxidized lipids, which resulted in the same digestive syndrome and histological symptoms as the endogenous suite of compounds produced by UV. Further investigation of the lipidomes revealed that oxylipins were only one class of potentially harmful compounds linked to UV sterilization, and the dissolved concentrations of secondary metabolites associated with higher plants, a wide range of pharmaceuticals, and anthropogenic organic pollutants also increased under UV light. Future efforts will explore the sources of these compounds, the mechanisms by which they inhibit oysters, and whether this is an emerging environmental problem for other ecosystems and shellfish hatcheries. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=137 SRC="FIGDIR/small/738347v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@43d06corg.highwire.dtl.DTLVardef@2894daorg.highwire.dtl.DTLVardef@449e59org.highwire.dtl.DTLVardef@f81983_HPS_FORMAT_FIGEXP M_FIG C_FIG

BackgroundZebrafish (Danio rerio) are a powerful vertebrate model for developmental toxicology and chemical safety assessment, yet large-scale transcriptomics in zebrafish remains limited by cost and data heterogeneity. Targeted transcriptomics offers a cost-effective alternative, but gene extrapolation methods tailored to zebrafish have not been systematically developed or evaluated. ObjectivesWhile the S1500+ platform is widely used for toxicogenomics research with rat, mouse, and human cell lines as model systems, its use in zebrafish has been limited due to data scarcity and lack of suitable bioinformatics approaches for analysis of such data. To that end, we sought to (i) curate a large zebrafish transcriptomic training data resource, and (ii) evaluate multiple machine learning strategies for reconstructing unmeasured transcriptome-wide expression profiles for data originating from the zebrafish-specific reduced representation gene set ("Zf S1500+"). MethodsWe assembled 14,924 zebrafish RNA-Seq samples covering 21,930 genes across 1,246 studies. Using the Zf S1500+ gene subset (3,062 genes), we trained and tested three extrapolation approaches: principal components regression (PCR), a locally weighted extension of PCR (PCR+), and a neural network mixture-of-experts model (NN-MoE). Model performance was assessed using mean absolute error (MAE), mean squared regression error (MSRE), and weighted variants of these metrics. ResultsExtrapolation performance using the baseline approach was strongly influenced by tissue and developmental context, with within-tissue models outperforming cross-tissue models. Errors were lowest when training and testing were conducted within the same tissue or between developmentally related tissues. Both PCR+ and NN-MoE improved upon the baseline PCR approach, with NN-MoE reducing average MAE by [~]20% and MSRE by [~]17%. Importantly, extrapolation remained reliable for the majority of genes, even when limiting output to high-confidence predictions using an empirical MAE threshold. ConclusionsWe demonstrate that targeted transcriptomics can be effectively extended to zebrafish, enabling robust transcriptome-wide extrapolation at reduced cost. The NN-MoE method provided the most substantial gains, highlighting the value of non-linear and ensemble modeling in heterogeneous datasets. These results establish a scalable framework for zebrafish toxicogenomics and suggest that accuracy will continue to improve with larger, better-annotated datasets, paving the way for broader application in chemical safety assessments.

Several studies have been conducted on human exposure to ultrafine particulate matter (UFPM), Black carbon (BC), and polystyrene nanoplastics (PS-NPs). However, it remains unclear whether different chemical types of environmental nanoparticles induce a similar mitochondrial stress response or a unique particle-specific response. In the present study, we examined the molecular mechanisms underlying nanoparticle-induced mitochondrial stress response and immunotoxicity using human peripheral blood mononuclear cells exposed to UFPM, BC, and PS-NPs under similar experimental conditions. Oxidative stress, mitochondrial adaptation, respiratory chain integrity, mitochondrial integrated stress response, inflammatory signaling, and systems-level interactions between molecules were analyzed through the evaluation of the expression of NRF2, HIF-1, PGC-1, TFAM, OMA1, DELE1, mitochondrial ND1, Complex I-V, NF-{kappa}B, TNF-, and NLRP3 and the use of principal component analysis, hierarchical clustering, and correlation networks. All three nanoparticles caused oxidative stress and mitochondrial dysfunction with different kinetics and mechanisms. UFPM mostly induced an acute antioxidant response and mitochondrial adaptation; BC led to chronic mitochondrial dysfunction, chronic activation of the OMA1-DELE1-mediated mitochondrial ISR pathway, and inflammation; while PS-NPs induced low but chronic mitochondrial adaptation along with mitochondrial biogenesis and stress responses. Our systems-level analysis showed that oxidative stress, mitochondrial adaptation, mitochondrial ISR, and inflammation represent a highly connected molecular network regardless of the physicochemical nature of the nanoparticles, with the OMA1- DELE1 axis being a key regulatory node connecting mitochondrial stress response and inflammation. Overall, we have found that mitochondrial stress response is a common mechanism underlying the toxicity of chemically different nanoparticles and have also revealed particle-specific stress-response dynamics responsible for the degree and persistence of cellular damage. The current work presents novel insights into the molecular mechanisms of nanoparticle-induced immunotoxicity and suggests OMA1, DELE1, NRF2, PGC-1, TFAM, ND1, and Complex I-V as potential biomarkers.

Chemical mixtures and potential cocktail effects in aquatic ecosystems are recognised as a threat for river basins world-wide. Probabilistic risk approaches are becoming more common in environmental risk assessment, and offer new opportunities for metodological challenges such as of mixture risk characterisation. The Concentration Addition (CA) concept is commonly used in lower-tier risk assessment (e.g., sum of risk quotients), as a pragmatic and protective method. However, the alternative Independent Action (IA) concept can easily be implemented in probabilistic risk calculation (e.g., joint probability of threshold exceedances). We have developed a multi-level probabilistic model for integrating these two concepts, formulated as an object-oriented Bayesian network (BN). First, probabilistic risk quotients (RQ) are calculated for individual substances, as probability distributions of environmental concentrations divided by a threshold environmental value. Next, the CA concept is applied within groups of substances by summing the RQ distributions. Finally, the IA concept is applied across the different substance groups, assuming independent modes of action, to combine RQ distributions by joint probability calculation ("OR" expressions). Predicted exposure concentrations were obtained from the ENCORE fate model, a process-based model for simulation of chemicals in river basins across Europe. Here we present a pilot study focusing on a subset of the substances (15 pesticides) and river basins (in Belgium), as a proof-of-concept. The purpose of this pilot study was to demonstrate a novel probabilistic approach to mixture risk characterisation, by combining the CA and IA concepts in a multi-level BN. The results were consistent across scenarios as well as with literature, with CA-based risk characterisations being slightly higher the IA-based. The combined CA+IA-based risk represents a reasonable compromise. Sensitivity analysis of the BN can provide an effective ranking of the risk-driving substances and groups, to support chemical prioritisation and risk managment. Key pointsO_LIA multi-level Bayesian network (BN) was developed for probabilistic calculation of environmental risk from chemical mixtures, based on risk quotients (RQ) calculated for individual substances, with a selection of 15 pesticides in Belgium as a pilot study. C_LIO_LIPredicted environmental concentrations (PEC) are obtained from the ENCORE exposure model; a process-based model which can simulate transport, fate and concentration of >1000 substances (pesticides, pharmaceuticals, etc.) in rivers subcatchments across Europe based on chemical use and emission. C_LIO_LIThe BNs risk calculation uses substance groups (Fungicides, Herbicides and Insecticides) to combine two classical mixture concepts: (1) Concentration Addition: sum of RQs within groups, followed by (2) Independent Action: joint probability of threshold exceedance for one or more groups. C_LIO_LIThe resulting rankings of risk-driving substances by the BN for this pilot study are robust across scenarios, suggesting a potential for expanding this generic BN approach to different mixtures with higher numbers of substances and groups, and to larger regions of Europe. C_LI

Background: Population-based biomonitoring of contemporary-use pesticides remains limited in the United States, particularly in rural agricultural regions, and few studies have repeated measurements within the same individuals over time. Methods: We analyzed 28 urinary pesticide-related biomarkers among 600 adults from the population-based Survey of the Health of Wisconsin with archived urine collected during 2008-2016; 296 participants provided repeat urine and updated exposure information in 2025. Detection frequencies, co-detection, and within-person detection patterns were characterized. Generalized estimating equations were used for stacked, repeated-measures analyses of factors associated with detection of aminomethylphosphonic acid (AMPA), glyphosate, 2,4-dichlorophenoxyacetic acid (2,4-D), and any of these three. Prospective-only analyses evaluated more detailed agricultural and recent exposure measures. Results: Glyphosate, AMPA, and 2,4-D were detected in 7.7%, 6.2%, and 4.3% of retrospective specimens and 5.4%, 3.1%, and 4.1% of prospective specimens, respectively. Co-detection and persistent detection across the 9 to 17-year interval was rare. In repeated-measures models, greater fruit and vegetable intake, older age, and male sex were associated with higher 2,4-D detection. Lower household income was associated with lower AMPA detection, while afternoon/evening collection was associated with higher AMPA detection. In prospective analyses, working on field-crop agricultural land showed the strongest agricultural associations, particularly for 2,4-D and detection of any of the three pesticides. Associations were not seen with self-reported conventional versus organic produce consumption. Conclusions: Urinary pesticide detections were generally infrequent in this Wisconsin population. Diet and direct agricultural activities may be more informative exposure pathways than residing near cropland or private well drinking-water characteristics.