Environmental Science & Technology

Per- and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants associated with adverse female reproductive outcomes. However, most prior studies examined individual PFAS, despite that humans are exposed to complex PFAS mixtures. Here, we investigated the ovarian effects of a mixture of five commonly detected long-chain PFAS, including PFOS, PFOA, PFNA, PFDA, and PFHxS, using complementary in vivo mouse and in vitro 3D ovarian follicle culture models. Young adult female mice were exposed to a range of concentrations of the PFAS mixture (PFASmix) through daily drinking water for 8 weeks. PFASmix at high concentrations (PFASmix-high) disrupted mouse estrous cyclicity, decreased circulating estradiol, increased testosterone, and promoted follicular atresia. Antral follicles from exposed mice showed decreased Lhcgr and increased Amh expression. Although the number of ovulated oocytes was unchanged following superovulation, PFASmix-high reduced oocyte size and polar body extrusion and altered oocyte cytoskeletal characteristics. In the 3D follicle culture model, direct PFASmix exposure impaired follicle growth, ovulation, and oocyte maturation in a concentration-dependent manner. PFASmix-high suppressed the periovulatory expression of Areg, Ereg, Tnfaip6, and Adamts1 and impaired corpus luteal (CL) development and function, as evidenced by reduced CL spheroid growth and cell survival, decreased expression of Star, Cyp11a1, Hsd3b1, and Lhcgr, and reduced progesterone secretion. PFASmix-high exposure additionally altered the expression of multiple PPAR{gamma}-associated genes in cultured ovarian follicles. Benchmark dose modeling identified estrous cyclicity and follicle rupture as sensitive in vivo and in vitro endpoints, respectively. Together, these findings demonstrated that exposure to the mixture of long-chain PFAS disrupts multiple stages of ovarian function, from follicular development, oocyte maturation to ovulation and luteal development and function and provided mechanistic and quantitative evidence for the reproductive toxicity of PFAS mixtures.

Chemical exposures during pregnancy are linked to an increased risk of pregnancy complications that contribute significantly to maternal and infant morbidity and mortality and can lead to long term health consequences for both the mother and the offspring. The placenta, a central regulator of pregnancy health, is a direct target of environmental toxicants. Epidermal growth factor receptor (EGFR), highly expressed in the placenta, regulates proliferation, migration, invasion, fusion, and cellular bioenergetics. To identify compounds of environmental concern with potential for EGFR-disrupting activity, we optimized a high-throughput virtual screening protocol for the identification of EGFR inhibitors and achieved enrichment factors of EF1% = 10.09, EF5% = 3.86, and EF10% = 3.0 in a benchmarking dataset. We applied this protocol to screen the Collaborative Estrogen Receptor Activity Prediction Project database, finding that top-scoring compounds were enriched for aromatic and fused-ring chemical classes, including dyes. Kinase activity assays revealed that two out of thirteen selected compounds, Vat Red 32 and Reactive Red 136, inhibited EGFR kinase activity with micromolar IC50 values. Additionally, pose refinement with molecular dynamics simulations characterized the binding interactions of Reactive Red 136 within the EGFR kinase domain, and functional assays in HTR-8/SVneo placental trophoblast cells showed that Reactive Red 136, but not Vat Red 32, partially attenuated EGF-mediated cell migration despite both compounds inhibiting EGFR kinase activity. Together, this study has generated an enriched dataset of candidate environmental EGFR modulators, with experimental validation confirming enrichment for EGFR-disrupting activity among the selected compounds. These results provide a valuable resource for toxicological studies.

Large-scale chemical screens are important tools for hazard identification and chemical prioritization, but they less commonly progress from identifying a phenotype to identifying its mechanism. Here, we used zebrafish embryos to screen 4,657 chemicals from the U.S. EPA ToxCast Phase III library for disruption of embryonic development and advanced selected hits through sequential validation using original library stocks, independently sourced chemicals, concentration-response analysis, transcriptomics, and functional experiments. Of 61 primary hits subjected to repeat testing, 33 reproduced the original phenotype, and four of eight compounds subsequently tested using independently sourced chemicals exhibited reproducible concentration-dependent developmental toxicity. We then investigated purpurin, an understudied anthraquinone pigment that caused pericardial edema, circulation defects, and body-axis abnormalities. Transcriptomic analysis of purpurin-exposed embryos revealed coordinated suppression of pathways involved in calcium regulation, ion transport, and neuronal signaling. Increasing extracellular calcium produced a concentration-dependent rescue of purpurin-induced developmental abnormalities, whereas equivalent magnesium supplementation did not, supporting a role for calcium availability or homeostasis in purpurin developmental toxicity. These results demonstrate that large-scale in vivo toxicity screening can be integrated with independent chemical validation and functional follow-up to move beyond hazard identification toward mechanistic understanding of how environmental chemicals disrupt embryonic development.

Hormesis is an important consideration in chemical risk assessment across most aquatic environments. Evidence suggests that hormetic responses are shaped by organismal physiological processes. Accordingly, predictions of hormesis under environmentally realistic conditions should account for physiological traits. Yet the role of these traits remains largely unexplored. This study investigated how baseline growth rate modulates antibiotic responses in the harmful cyanobacterium Microcystis and the underlying mechanisms. Results showed that at an antibiotic concentration of 200 ng L-1 (representing environmental concentration levels), slow-growing Microcystis populations exhibited an obvious growth enhancement more than 4-fold greater than that of fast-growing populations, and this enhancement was positively correlated with cellular reserves. A baseline growth rate below 0.19 day-1 emerged as the threshold below which hormetic responses were consistently observed. This study provides pioneering insights into how organisms' physiological states influence pollutant-induced hormetic responses in aquatic environments. Our findings point to a central principle for extrapolation that laboratory-derived toxicological responses may not translate directly to the field unless key physiological traits of the organisms are explicitly considered. In this regard, baseline growth rate and energy reserves may be key physiological traits for reliably predicting toxic effects at the population level. This study also makes an initial attempt to conceptualize a physiology-anchored framework for strengthening risk assessment of low-level emerging contaminants.

Per- and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants with the potential to disrupt metabolic processes and contribute to carcinogenesis; however, molecular evidence linking alternative and emerging PFAS to ovarian cancer (OC) remains limited. We conducted a biomarker-based case-control study integrating exposureomics with multi-omics profiling. Plasma samples from 128 patients with OC and 128 controls were analyzed for 26 PFAS, untargeted metabolomics, lipidomics, and clinical phenotypes, while urine samples from an additional 100 patients and 100 controls underwent untargeted metabolomic and clinical profiling. Single-compound, burden-based, and mixture models were integrated with PFAS-omics association scans, cross-matrix metabolite prioritization, observational exposure-omics-clinical association-chain analyses, and repeated internal cross-validation. Higher PFAS burdens were consistently associated with OC case status, and DONA, GenX, PFNA, PFPeS, PFUdA, and PFTrDA constituted an OC-prioritized six-PFAS mixture. DONA and GenX exhibited the broadest lipid-related bridging signals, particularly across phospholipids, lysophospholipids, sphingolipids, neutral lipids, and acylcarnitines. Integrated plasma and urine analyses further identified five shared metabolites-elaidic acid, hippuric acid, betaine, corticosterone, and taurocholic acid-that provided incremental and complementary discrimination beyond reference models based on clinical variables and tumor markers in internal cross-validation. Collectively, these findings demonstrate that both alternative and emerging PFAS are associated with a distinct OC-related molecular phenotype involving coordinated lipidomic, metabolomic, and hematologic alterations, while highlighting DONA and GenX as the most prominent exposure-related signals.

Typical environmental exposures to the toxic class of chemicals known as polycyclic aromatic hydrocarbons (PAHs) involve complex mixtures; however, relatively few mechanistic toxicity studies have evaluated them as environmental mixtures, instead focusing on individual compounds or simple mixtures. In this study, we first derived Republic Sediment Extract (REPSE), a complex PAH mixture extracted from sediment at the Republic Creosoting site of the Elizabeth River in Norfolk, Virginia. After characterizing the PAH contents of REPSE, we evaluated its mechanisms of developmental neurotoxicity in three evolutionarily distinct taxa, leveraging the unique strengths of Atlantic killifish, zebrafish, and Caenorhabditis elegans as model species, with a focus on the Aryl hydrocarbon Receptor (AhR) pathway. Embryonic REPSE exposure caused induction of CYP1A in both fish species at sub-teratogenic concentrations, consistent with activation of the canonical AhR pathway. These sub-teratogenic exposures nevertheless induced neurotoxicity across both fish species, altering neurobehavioral phenotypes in fish, and induced dopaminergic neuronal damage in worms, again at non-teratogenic concentrations. To determine whether these effects were linked to canonical AhR response pathways, we examined killifish offspring from the pollution-adapted Republic Creosoting population, which exhibited characteristic recalcitrance to CYP1A induction, but remained susceptible to the neurobehavioral effects of REPSE. The induction of neuronal damage in worms provides orthogonal evidence for a non-AhR mechanism, because C. elegans AhR is not transcriptionally activated by PAHs as in vertebrates. Further probing of potential mechanisms underlying REPSE-induced neurotoxicity in worms revealed altered neuronal redox status (roGFP) and energy availability (ATP:ADP ratio). Collectively, our multispecies approach reveals conserved mechanisms of PAH mixture neurotoxicity, including effects that extend beyond canonical AhR signaling.

Hexafluoropropylene oxide dimer acid (GenX), a replacement for legacy per- and polyfluoroalkyl substances (PFAS), is increasingly detected in the environment, yet its chronic toxicity remains poorly characterized. Current safety assessments rely largely on short-term, high-dose studies that may not capture the biological consequences of long-term, low-dose exposure. To address this gap, we employed 3D human liver (HepG2/C3A) spheroids cultured in a continuously rotating bioreactor system (ClinoStar) to systematically evaluate dose- and time-dependent mRNA changes in response to GenX under environmentally relevant conditions. Spheroids were exposed to GenX (0.08-50 M, spanning environmentally relevant to mechanistically informative concentrations) for acute (4 days) and chronic (4 weeks) durations, followed by genome-wide TempO-Seq transcriptomic profiling and benchmark dose (BMD) modeling. GenX elicited pronounced non-monotonic mRNA changes in acute exposure conditions, with the greatest number of differentially expressed genes (DEGs) observed at an intermediate concentration (0.4 M). In contrast, chronic exposure exhibited a generally concentration-dependent increase in DEGs, except for the 10 M condition, indicating a more consistent dose-response relationship than acute exposure. Notably, acute and chronic exposures elicited qualitatively distinct mRNA changes with low concordance across matched concentrations, demonstrating that exposure duration was a major determinant of mRNA changes. Acute low-dose GenX exposure preferentially modulated mRNA encoding components of cell cycle-related pathways, whereas acute higher dose exposures suppress mRNA levels of the constituents of lipid metabolic pathways and increase expression of mRNA encoding proteins involved in stress- and toxicity-associated signaling. Chronic exposure revealed a different pattern of changes in mRNA expression not observed under acute exposure conditions, including suppression of cellular components involved in lipid-related pathways at the lowest concentration tested. At higher concentrations, mRNA levels of components of multiple metabolic pathways were altered. Benchmark dose modeling identified a significantly lower transcriptomic point of departure (tPOD) for chronic exposure as compared to acute exposure, suggesting increased cellular sensitivity to prolonged GenX exposure and supporting the relevance of chronic models for human exposure assessment. Collectively, these findings demonstrate that GenX elicits time-dependent and non-monotonic changes in mRNA levels of human liver (HepG2/C3A) spheroids, with distinct responses depending on the exposure duration and dose. This study, therefore, highlights the importance of incorporating chronic, human-relevant in vitro models and transcriptomic endpoints into PFAS risk assessment and suggests that conventional short-term assays may underestimate the biological impact of sustained low-dose exposure. Key message (Impact of the study)This study provides systematic comparisons of short term (4 day) versus longer term (4 weeks), environmentally relevant GenX exposure in human liver spheroids, revealing non-monotonic, time-dependent changes in mRNA levels encoding cellular components of lipid metabolism-related pathways with potential implications for appropriate dose and time exposure parameters for use in New Approach Methods to be applied in risk assessment.

Environmental exposure involves pollutant mixtures. Building on evidence that individual bisphenols alter zebrafish larval physiology and transcriptomic profiles at ng/L concentrations, we assessed the effects of specific mixtures of BPS, BPF, and BPAF without (M1) or with (M2) BPA. Experimentally, M1 and M2 induced 4.16% and 6.54% mortality in zebrafish larvae, despite the concentration addition model predicting no toxicity. Both mixtures reduced swimming-locomotor activities. These behaviors were accompanied by organ-level functional defects, as in vivo optical mapping revealed bradycardia and increased high-frequency (30-80 Hz) brain oscillations for both mixtures, while M2 additionally altered low-frequency (0.2-4 Hz) brain activity. The neurophysiological changes were accompanied by neurovascular remodeling, with in vivo two-photon microscopy revealing decreased vascular length and branching. Finally, transcriptomic analysis revealed common M1- and M2-induced alterations linked to pathways of vascular function, neuronal communication, and metabolism. M2 altered additional pathways of cardiac and neuronal signaling, consistent with the reported organ-physiology effects. We discuss the environmental relevance of mixture exposures and assess whether BPA intensifies mixture-induced responses.

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

Inhalation of airborne per- and polyfluoroalkyl substances (PFAS) represents an important yet understudied exposure route. While volatile organic compounds (VOCs) have been widely explored as biomarkers of toxicant exposure, their potential for early detection of PFAS-induced pulmonary toxicity remains largely unexplored. This study investigated VOC profiles in human alveolar (A549) and bronchial (BEAS-2B) epithelial cells following exposure to environmentally relevant PFAS concentrations (1, 5, and 10 ppb) for 12, 24, and 48 h. VOCs were analyzed using headspace solid-phase microextraction coupled with gas chromatography-Orbitrap mass spectrometry. We observed significant concentration- and time-dependent alterations in VOC profiles in both cell models. The principal VOCs associated with PFAS exposure include butanal, acetone, 2-butanone, 2-nonanone, (E)-2-nonenal, (E)-2-octenal, (E)-2-heptenal, 2,4-heptadienal, 2,4-nonadienal, 2,4-decadienal, 1-pentanol, and 1-hexanol. Cell type-specific regulations were observed, reflecting differential metabolic responses between transformed and non-transformed lung epithelial cells. Metabolite set enrichment analysis indicated perturbations in ketone body metabolism, consistent with altered cellular energy metabolism. Complementary qPCR analysis revealed altered expressions of CYP1A1, CYP2E1, ALDH1A1, and ALDH2, supporting a mechanistic link between PFAS exposure and oxidative stress, lipid peroxidation, and impaired aldehyde detoxification. These findings demonstrate that VOC profiling, combined with transcriptional analysis, can reveal PFAS-induced metabolic perturbations in pulmonary epithelial cells.

IntroductionGut microbiota are vulnerable to foreign chemicals (xenobiotics) including pharmaceuticals, environmental pollutants, and dietary contaminants such as aflatoxin B1 (AFB1) and fumonisin B1 (FB1). Assessing the effect of these xenobiotics in the laboratory requires their dissolution in a solvent vehicle, such as dimethyl sulfoxide (DMSO). While DMSO is typically used at low concentrations under the assumption of neutrality, its independent impact on microbial dynamics is a potential experimental confounder that has not been fully explored. MethodsHuman fecal microbiota were cultivated invitrofor 16 days, supplemented with 0, 10, 100, and 1000 ppb of the tested xenobiotics (AFB1 or FB1) in 0.05% DMSO (v/v), with a DMSO-free control included for comparison. Microbial community dynamics were characterized via full-length 16S rRNA gene sequencing, and metabolic activity was assessed by measuring production of short-chain fatty acids and gases. ResultsDMSO significantly altered microbial metabolism and drove the consistent enrichment of Desulfovibriodesulfuricans. This shift occurred across all AFB1 and FB1 treatment groups regardless of their concentrations, indicating that the biological impact of the DMSO vehicle overshadowed the specific effects of the xenobiotics. DiscussionThese findings demonstrate that DMSO can induce significant microbial shifts independent of the xenobiotics under study, potentially confounding biological interpretations. This highlights a critical need for rigorous vehicle validation and the identification of safe thresholds for solvents used in microbiota research.

Bauxite residue (BR), the haloalkaline byproduct of alumina refining, represents the largest and most costly environmental challenge facing the global aluminium industry, yet sustainable remediation has remained elusive because no rapid and field-feasible technology can overcome its recalcitrant alkalinity. Here, we establish a self-amplifying microbial-abiotic sulfur relay that drives rapid in situ acid generation and sustained dealkalization of BR across laboratory and glasshouse experiments and a field trial, where dealkalized residue subsequently supported spontaneous pioneer-plant colonization. Mechanistic assays and multi-omics analyses show that the relay is initiated by microbial reduction of elemental sulfur (S8) to HS- under oxygen-limited conditions. The resulting HS- abiotically attacks and solubilizes solid S_8, generating a mobile pool of polysulfides (Sx2-). In anoxic microsites, polysulfide reduction regenerates HS^-, which mobilizes additional S8 and amplifies sulfur turnover by increasing sulfur mobilization and bioavailability. In oxic microsites, Sx(2-) are abiotically converted to thiosulfate and reactive S0, which are subsequently microbially oxidized to sulfate and acidity. By coupling biotic reductive initiation and regeneration with abiotic sulfur mobilization and oxidation, followed by biotic terminal oxidation, this relay overcomes the low bioavailability of S8 and the constraints of extreme haloalkaline conditions, providing a low-cost, field-feasible strategy for efficient and sustained BR remediation.

Wastewater treatment is an important barrier to the environmental dissemination of antimicrobial resistance (AMR). However, its effectiveness is typically assessed using measures that do not directly capture resistance gene mobility or pathogen association. Here, we applied a configuration-based genomic risk framework (MetaCompare2.0) to 100 metagenomes (50 paired influent and effluent samples) collected from nine wastewater treatment works over 11 months, quantifying resistome risk based on associations between antimicrobial resistance genes (ARGs), mobile genetic elements, and host taxa. Influent samples exhibited highly variable ecological risk associated with mobile ARGs. Pathogen-associated human health risk was lower and more constrained. Wastewater treatment consistently reduced both ecological and human health risks, driving convergence towards uniformly lower risk resistome configurations. Risk attenuation was reproducible across treatment technologies and could not be explained by assembly quality or physicochemical variation. These findings indicate that wastewater treatment disrupts high-risk ARG configurations and highlight configuration-based genomic risk metrics as scalable tools for benchmarking environmental AMR mitigation.

Per- and polyfluoroalkyl substances (PFAS) such as GenX (HFPO-DA) are aerobically recalcitrant contaminants for which biological treatment options remain scarce; the best-characterized microbial degraders require strictly anaerobic conditions and external cofactors. We isolated Cladosporium halotolerans strain CsHGX-1 from activated sludge at a municipal wastewater treatment plant using GenX as the sole carbon source. Whole-genome sequencing (32.4 Mb; 11,201 genes) revealed a PFAS-degradation gene repertoire substantially expanded relative to congeneric Cladosporium species, including 26 dehalogenases (four type-II haloacid dehalogenases, HADs), 141 cytochrome P450s, and 558 esterases/hydrolases. Under aerobic conditions with GenX (50 mg L-1) as the sole carbon source, strain CsHGX-1 removed 47.6 {+/-} 1.7% of GenX within 48 h, accompanied by fluoride release (0.079 {+/-} 0.018 mM) that was absent in abiotic controls, confirming genuine C-F bond cleavage. Time-resolved RNA sequencing (0, 6, 24, 48 h; n = 6 biological replicates) revealed a phase-structured transcriptional program: oxidative genes, including cytochrome P450s, peaked first (6 h; up to 25.4-fold), hydroxylation and reactive-oxygen-species-management genes peaked next (24 h; up to 33.5-fold), and the three type-II HAD genes peaked last (48 h; up to 50.9-fold), coincident with fluoride accumulation. A parallel resazurin metabolic assay over 5 days confirmed sustained catabolic activity in GenX-exposed cultures relative to controls (1.37-1.52-fold; p [≤] 0.003). These findings identify strain CsHGX-1 as, to our knowledge, the first Ascomycete fungus for which genomic and transcriptomic evidence links oxidative activation to haloacid-dehalogenase-mediated defluorination of an aerobically recalcitrant PFAS, extending the known diversity of fungal PFAS degraders beyond Basidiomycota white-rot taxa. IMPORTANCEGenX is a PFAS "replacement" chemical that the U.S. Environmental Protection Agency added to its list of hazardous constituents in 2024, yet no aerobic biological treatment exists for it: every well-characterized microbial degrader requires oxygen-free conditions and added cofactors. We show that a fungus recovered from ordinary wastewater sludge breaks down GenX while using oxygen, the same conditions already used in conventional treatment plants, with no nutrient or reductant supplementation. Genome sequencing showed why this strain is unusual: it carries far more dehalogenase and cytochrome P450 genes than its close relatives. Time-course RNA sequencing showed these genes switch on in a defined order, oxidation first, then carbon-fluorine bond cleavage, matching the appearance of free fluoride in the culture. This links genome content to a functional outcome in an Ascomycete fungus, suggesting aerobic fungal defluorinators may already be present, unrecognized, in engineered wastewater systems.

Essential for nutrient growth, due to intensive farming practices nitrate has become a common water contaminant in agricultural states. US nitrate regulations are designed to protect the health of newborns and infants, but have never been revised to consider potential harms of prenatal exposure. Recent evidence, however, suggests that exposure below the 10 mg/L regulatory limit may be associated with adverse birth outcomes. This current study examines whether first-trimester exposure below the regulatory limit is associated with adverse birth outcomes in Iowa (an intensive agricultural state). Finished drinking-water nitrate measurements from Iowa public water systems were linked to birth records. Nitrate measures were aggregated to a balanced county-quarter panel (1983 to 1988). Each pregnancy was assigned an average nitrate exposure over a 98-day first-trimester window. All births with a first trimester county-quarter nitrate measure >= 10 mg/L were excluded. Linear probability models with county-by-year and year-by-conception-quarter fixed effects, with standard errors clustered by county, estimated both uncorrected and Sidak corrected inference. The final analytic sample included 113,328 singleton births. Relative to lowest exposure category (0.1 mg/L or below), exposure to 5 mg/L or more nitrate was associated with a 1.04 percentage-point increase in the probability of preterm birth (CI = 0.24 to 1.83), a 13.5% relative increase, and associated with a 0.53 percentage-point increase in very preterm birth (CI = 0.08 to 0.97), a 46.2% relative increase. The statistical significance of both estimates persisted after correction for multiple tests. Exposure to > 0.1 mg/L and < 5 mg/L was not associated with any outcome. These preterm and very preterm birth associations were consistent across several sensitivity checks. Because nitrate concentrations between 5 to <10 mg/L comply with current rules, these findings warrant reassessing approaches to protect the health of pregnant women and their children.

Wastewater is a key reservoir and transmission route for antibiotic resistance genes (ARGs), enabling their spread from influent to effluent and into receiving environments. However, how combined selective pressures (antibiotics, biocides, heavy metals, pharmaceuticals) influence resistant bacteria and ARG persistence over space and time remains poorly understood. Likewise, the role of the wastewater microbiome in ARG dynamics is still unclear, as few studies integrate microbiome shifts with chemical and environmental drivers. Here, we investigated how microbiome dynamics, chemical exposures, and environmental conditions shape clinically relevant ARG dynamics from sewage to receiving environments in Guadeloupe, French Caribbean. We analysed data collected from three wastewater continuums, (hospital-based, domestic, touristic) over four campaigns (September 2021-February 2023). We characterised ARG and microbiome composition spatiotemporal patterns and used a mixed-effect model to investigate ARG associations with potential drivers, including exposome factors, microbiome dissimilarity and environmental factors. Several ARGs were negatively associated with microbiome dissimilarity (Bray-Curtis distances) (aac(6)-Ib, aph(3)-III, blaSHV, blaTEM, intI1, qnrS, sul1 and tetM). Negative associations were also observed between upstream-downstream differences in anti-inflammatory drug concentrations and the abundance of aac(6)-Ib, aph(3)-III, blaCTX-M, ermB, intI1, and tetM. In contrast, ARG relative abundance was positively associated with upstream-downstream differences in antibiotic concentrations, suggesting selection along the continuum. These findings indicate that ARG dissemination along wastewater-to-coastal pathways is shaped by opposing processes, with microbiome turnover potentially limiting ARG persistence while chemical gradients promote specific gene enrichment. The outcome is ARG-specific, with implications for antimicrobial resistance risks associated with recreational waters, seafood consumption, and coastal ecosystem interactions.

The anaerobic biotransformation of lindane ({gamma}-hexachlorocyclohexane), a persistent legacy pesticide and challenging subsurface contaminant was modelled using an integrated Monod biokinetics framework. Calibrated against batch experimental data, this model accounted for the dissolution of solid phase lindane and the thermodynamics of reactions involved. The study focused on a methanogenic, lindane-degrading enrichment culture (GT2) that used ethanol as the electron donor, where ethanol fermentation supplied hydrogen to support reductive dechlorination of lindane. A Dehalobacter strain was identified as the lindane-degrading organism, while a member of bacterial order Geobacterales carried out ethanol fermentation. First-order decay constants for lindane-degrading Dehalobacter were estimated from experimental data and showed variation across different growth conditions. The biokinetics model captured interactions among key microbial processes, namely lindane dechlorination, ethanol fermentation, hydrogenotrophic methanogenesis, and acetoclastic methanogenesis. Thirteen biokinetics parameters including maximum substrate utilization rates (k), half-velocity constants (Ks), and decay constants for these microbial processes were estimated through a Bayesian optimization approach. Global sensitivity analysis showed that eight parameters, primarily k and Ks, had significant linear impact on the model output, whereas responses to decay constants were more variable and potentially non-linear. This study proposed an effective modelling approach for characterizing syntrophic microbial growth within enrichment cultures and provides valuable biokinetics parameters estimates for real-world applications of anaerobic lindane bioremediation.

The 2020 SARS-CoV-2 pandemic renewed global interest in wastewater-based epidemiology (WBE) as a tool for monitoring public health. Molecular analyses of wastewater are often limited by the small volumes of wastewater that can be processed, due to column clogging, handling constraints, and processing time. Additionally, inhibitors in the complex wastewater matrix reduce the sensitivity of downstream assays such as RT-PCR and sequencing. To address these limitations, we developed a novel column by incorporating a hydrophobic pre-filtration layer and sequential glass fiber filters. This enhanced column design, PureBioX Xpurify Column, enables processing of 1.58 times more wastewater (a 58% increase in throughput) while reducing RT-PCR inhibitors and maintaining compatibility with existing workflows. Despite a modest reduction in nucleic acid yield, the modified column consistently improved viral RNA detection sensitivity, including for SARS-CoV-2. This accessible, scalable upgrade strengthens the utility of direct capture methods in WBE-based public health surveillance.

Integrating high-throughput in vitro data into next-generation risk assessment (NGRA) workflows requires screening strategies that yield quantitative potency estimates and mechanistically interpretable biological signals. Transcriptomic and morphological profiling are increasingly adopted for early-stage hazard identification by enabling triage of substances for resource-intensive follow-up and prioritizing candidates most likely to present meaningful risk. In this study, we aimed to characterize biological concordance and uncertainty by quantifying how well high-throughput transcriptomics (HTTr) and Cell Painting PLUS (CPP) bioactivity profiles recover target-relevant biological signals in immortalized human renal proximal tubule epithelial RPTEC/TERT1 cells using 313 reference chemicals with high-confidence target annotations. Through quality control procedures and biological activity filters we yielded 142 reference chemicals spanning 66 different targets, which were systematically evaluated for biological concentration-responses by HTTr and CPP. HTTr was evaluated using TXG-MAPr-based qualitative and quantitative gene network activity analysis. HTTr showed the most prominent activity for targets that were highest expressed in RPTEC/TERT1 cells. Active chemical-pairs showed strong gene network activity correlation albeit with different potencies. Similarly, the highest transcriptomic concordance was observed for reference chemicals acting in the same pathway, such as EGFR/MEK or PI3K/AKT/mTOR. CPP often showed high sensitivity primarily at the organelle level providing limited statistical power for chemical grouping. Collectively, the results support HTTr and CPP as complementary early-tier assays within an in vitro weight-of-evidence safety testing framework. Although CPP is suitable as a cost-effective screening modality, HTTr offers higher mechanistic resolution for mode-of-action inference in high-throughput bioactivity screening and therefore remains necessary for high-confidence mechanistic interpretation.

Biodiversity is globally threatened by chemical pollution, yet toxicity data remain unavailable for millions of species and tens of thousands of chemicals, severely limiting our ability to assess ecological impacts. Here we present TRIDENT-2, a multimodal artificial intelligence model for predicting chemical toxicity across evolutionarily diverse eukaryotic species. Trained on 560,780 toxicity assays spanning 82,775 chemicals, 6,793 species, and multiple exposure scenarios, TRIDENT-2 accurately predicts toxicity across Eukaryota with an average median absolute error ranging from 1.76 to 3.80. By jointly learning from chemical, biological, and experimental information, it remains accurate across broad chemical and taxonomic distances, allowing for toxicity assessment for species and chemicals beyond the current experimental evidence. Our findings demonstrate that artificial intelligence can help overcome longstanding data limitations in ecotoxicology, paving the way for improved decision-making and reducing chemical impacts on biodiversity and ecosystems.