Developing an Approach for Integrating Chemical Analysis and Transcriptional Changes to Assess Contaminants in Water, Sediment, and Fish.

Pharmaceuticals in aquatic environments pose threats to aquatic organisms because of their continuous release and potential accumulation. Monitoring methods for these contaminants are inadequate, with targeted analyses falling short in assessing water quality's impact on biota. The present study advocates for integrated strategies combining suspect and targeted chemical analyses with molecular biomarker approaches to better understand the risks posed by complex chemical mixtures to nontarget organisms. The research aimed to integrate chemical analysis and transcriptome changes in fathead minnows to prioritize contaminants, assess their effects, and apply this strategy in Wascana Creek, Canada. Analysis revealed higher pharmaceutical concentrations downstream of a wastewater-treatment plant, with clozapine being the most abundant in fathead minnows, showing notable bioavailability from water and sediment sources. Considering the importance of bioaccumulation factor and biota-sediment accumulation factor in risk assessment, these coefficients were calculated based on field data collected during spring, summer, and fall seasons in 2021. Bioaccumulation was classified as very bioaccumulative with values >5000 L kg-1, suggesting the ability of pharmaceuticals to accumulate in aquatic organisms. The study highlighted the intricate relationship between nutrient availability, water quality, and key pathways affected by pharmaceuticals, personal care products, and rubber components. Prioritization of these chemicals was done through suspect analysis, supported by identifying perturbed pathways (specifically signaling and cellular processes) using transcriptomic analysis in exposed fish. This strategy not only aids in environmental risk assessment but also serves as a practical model for other watersheds, streamlining risk-assessment processes to identify environmental hazards and work toward reducing risks from contaminants of emerging concern. Environ Toxicol Chem 2024;00:1-22. © 2024 SETAC.

Mixtures of legacy and replacement perfluorosulphonic acids (PFSAs) demonstrate ratio-, concentration- and endpoint-dependent synergistic interactions in vitro.

The extensive use of poly- and per-fluoroalkyl substances (PFASs) has les to their widespread presence in the environment, raising concerns about potential toxicity. While certain PFASs of concern have been phased-out or banned, new PFASs continue to be produced. Two such substances are perfluoroethylcyclohexane sulphonate (PFECHS) and perfluorobutane sulphamide (FBSA), replacements of perfluoroctanesulphonic acid (PFOS) that have recently been detected in multiple environmental media around the globe. Despite PFASs generally occurring in the environment as mixtures, few data are available outlining the effects of PFAS mixtures. Therefore, this research investigated the interaction potential of binary and ternary mixtures of emerging and legacy PFASs. The immortalized rainbow trout gill cell line (RTgill-W1) was chosen as the experimental model to investigate two apical endpoints: cytotoxicity and phospholipidosis. RTgill-W1 cells were exposed for 24 h to each compound to obtain endpoint-specific effect concentrations (LCx; ECx). These values were then applied to formulate mixture predictions following the Loewes Additivity and Steel and Peckham methods. Based on cytotoxicity, relative potencies of individual compounds were: PFOS > PFECHS > FSBA. PFOS and PFECHS had nearly identical effects on phospholipidosis, while FSBA did not have any effects. Most mixtures had a synergistic effect on cytotoxicity, but the effect was both dose- and ratio-dependent. PFOS and PFECHS were additive at lower concentrations (LC10) and synergistic at higher concentrations (LC50; 3:1, 1:1, and 1:3). PFECHS and FSBA mixtures were synergistic at all doses and ratios (3:1, 1:1, 1:3), while FBSA and PFOS were mainly synergistic at higher concentrations and at ratios favouring PFOS (1:1, 1:3). Tertiary combinations were mainly synergistic. For phospholipidosis, mixtures were strictly additive. These results are strongly suggestive of synergism between emerging PFAS replacements and highlight that independent apical mechanisms of different PFASs could combine to induce unexpected toxicity. Considering that emerging replacements are continuing to increase in concentration in the environment, such mixture scenarios are also likely to continue to increase in probability.

Solid-liquid distribution of SARS-CoV-2 in primary effluent of a wastewater treatment plant.

Distributions of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and fecal viral biomarkers between solid and liquid phases of wastewater are largely unknown. Herein, distributions of SARS-CoV-2, Pepper Mild Mottle Virus (PMMoV), and F-RNA bacteriophage group II (FRNAPH-II) were determined by viral RNA RT-qPCR. Comparison of viral recovery using three conventional fractionation methods included membrane filtration, a combination of mid-speed centrifugation and membrane filtration, and high-speed centrifugation. SARS-CoV-2 partitioned to the solids fraction in greater abundance compared to liquid fractions in a combination of mid-speed centrifugation and membrane filtration and high-speed centrifugation, but not in membrane filtration method in a particular assay, while fecal biomarkers (PMMoV and FRNAPH-II) exhibited the reciprocal relationship. The wastewater fractionation method had minimal effects on the solids-liquids distribution for all viral and phage markers tested; however, viral RNA load was significantly greater in solid-liquid fractions viral RNA loads compared with the than whole-wastewater PEG precipitation. A RNeasy PowerWater Kit with PCR inhibitor removal resulted in greater viral RNA loads and lesser PCR inhibition compared to a QIAamp Viral RNA Mini Kit without PCR inhibitor removal. These results support the development of improved methods and interpretation of WBE of SARS-CoV-2. •Distribution of SARS-CoV-2 to liquid and solid portions was addressed.•Addressing PCR inhibition is important in wastewater-based epidemiology.•Fraction methods have minimal effect.

Emergence and spread of SARS-CoV-2 variants of concern in Canada: a retrospective analysis from clinical and wastewater data.

BACKGROUND: The spread of SARS-CoV-2 has been studied at unprecedented levels worldwide. In jurisdictions where molecular analysis was performed on large scales, the emergence and competition of numerous SARS-CoV-2lineages have been observed in near real-time. Lineage identification, traditionally performed from clinical samples, can also be determined by sampling wastewater from sewersheds serving populations of interest. Variants of concern (VOCs) and SARS-CoV-2 lineages associated with increased transmissibility and/or severity are of particular interest. METHOD: Here, we consider clinical and wastewater data sources to assess the emergence and spread of VOCs in Canada retrospectively. RESULTS: We show that, overall, wastewater-based VOC identification provides similar insights to the surveillance based on clinical samples. Based on clinical data, we observed synchrony in VOC introduction as well as similar emergence speeds across most Canadian provinces despite the large geographical size of the country and differences in provincial public health measures. CONCLUSION: In particular, it took approximately four months for VOC Alpha and Delta to contribute to half of the incidence. In contrast, VOC Omicron achieved the same contribution in less than one month. This study provides significant benchmarks to enhance planning for future VOCs, and to some extent for future pandemics caused by other pathogens, by quantifying the rate of SARS-CoV-2 VOCs invasion in Canada.

Advancing exposure assessment approaches to improve wildlife risk assessment.

The exposure assessment component of a Wildlife Ecological Risk Assessment aims to estimate the magnitude, frequency, and duration of exposure to a chemical or environmental contaminant, along with characteristics of the exposed population. This can be challenging in wildlife as there is often high uncertainty and error caused by broad-based, interspecific extrapolation and assumptions often because of a lack of data. Both the US Environmental Protection Agency (USEPA) and European Food Safety Authority (EFSA) have broadly directed exposure assessments to include estimates of the quantity (dose or concentration), frequency, and duration of exposure to a contaminant of interest while considering "all relevant factors." This ambiguity in the inclusion or exclusion of specific factors (e.g., individual and species-specific biology, diet, or proportion time in treated or contaminated area) can significantly influence the overall risk characterization. In this review, we identify four discrete categories of complexity that should be considered in an exposure assessment-chemical, environmental, organismal, and ecological. These may require more data, but a degree of inclusion at all stages of the risk assessment is critical to moving beyond screening-level methods that have a high degree of uncertainty and suffer from conservatism and a lack of realism. We demonstrate that there are many existing and emerging scientific tools and cross-cutting solutions for tackling exposure complexity. To foster greater application of these methods in wildlife exposure assessments, we present a new framework for risk assessors to construct an "exposure matrix." Using three case studies, we illustrate how the matrix can better inform, integrate, and more transparently communicate the important elements of complexity and realism in exposure assessments for wildlife. Modernizing wildlife exposure assessments is long overdue and will require improved collaboration, data sharing, application of standardized exposure scenarios, better communication of assumptions and uncertainty, and postregulatory tracking. Integr Environ Assess Manag 2024;20:674-698. © 2023 SETAC.

Cross-species apical microinjected selenomethionine toxicity in embryo-larval fishes.

Selenium (Se) is an essential micronutrient that becomes toxic when exposures minimally exceed those that are physiologically required. Studies on Se contaminated aquatic environments have identified that embryo-larval fishes are at particular risk of Se toxicity, primarily due to maternal Se transfer to developing eggs during oogenesis. This study emulated these exposures in embryo-larval fathead minnow (FHM), rainbow trout (RBT), white sucker (WSu), and white sturgeon (WSt) using embryonic selenomethionine (SeMet) microinjections. Adverse Se-outcomes observed across these species included spinal and edematous deformities, total individuals deformed, and reduced survival. Spinal deformity was the most sensitive sublethal endpoint and developed at the lowest concentrations in WSt (10 % effects concentration (EC10) = 12.42 µg (total) Se/g dry weight (d.w.)) followed by WSu (EC10 = 14.49 µg Se/g d.w.) and FHM (EC10 = 18.10 µg Se/g d.w.). High mortality was observed in RBT, but SeMet influences were confounded by the species' innate sensitivity to the microinjections themselves. 5 % hazardous concentrations derived across exposure type data subsets were ∼49 % higher when derived from within-species maternal transfer exclusive data as opposed to all, or within-species microinjection exclusive, data. These results support the current exclusion of SeMet microinjections during regulatory guideline derivation and their inclusion when studying mechanistic Se toxicity across phylogenetically distant fishes.

Interspecies Differences in 6PPD-Quinone Toxicity Across Seven Fish Species: Metabolite Identification and Semiquantification.

N-(1,3-Dimethylbutyl)-N'-phenyl-p-phenylenediamine-quinone (6PPD-Q) is a recently identified contaminant that originates from the oxidation of the tire antidegradant 6PPD. 6PPD-Q is acutely toxic to select salmonids at environmentally relevant concentrations, while other fish species display tolerance to concentrations that surpass those measured in the environment. The reasons for these marked differences in sensitivity are presently unknown. The objective of this research was to explore potential toxicokinetic drivers of species sensitivity by characterizing biliary metabolites of 6PPD-Q in sensitive and tolerant fishes. For the first time, we identified an O-glucuronide metabolite of 6PPD-Q using high-resolution mass spectrometry. The semiquantified levels of this metabolite in tolerant species or life stages, including white sturgeon (Acipenser transmontanus), chinook salmon (Oncorhynchus tshawytscha), westslope cutthroat trout (Oncorhynchus clarkii lewisi), and nonfry life stages of Atlantic salmon (Salmo salar), were greater than those in sensitive species, including coho salmon (Oncorhynchus kisutch), brook trout (Salvelinus fontinalis), and rainbow trout (Oncorhynchus mykiss), suggesting that tolerant species might detoxify 6PPD-Q more effectively. Thus, we hypothesize that differences in species sensitivity are a result of differences in basal expression of biotransformation enzyme across various fish species. Moreover, the semiquantification of 6PPD-Q metabolites in bile extracted from wild-caught fish might be a useful biomarker of exposure to 6PPD-Q, thereby being valuable to environmental monitoring and risk assessment.

Assessment of rapid wastewater surveillance for determination of communicable disease spread in municipalities.

Wastewater surveillance (WS) helps to improve the understanding of the spread of communicable diseases in communities. WS can assist public health decision-makers in the design and implementation of timely mitigation measures. There is an increased need to use reliable, cost-effective, simple, and rapid WS systems, given traditional analytical (or 'gold-standard') programs are instrument/time-intensive, and dependent on highly skilled personnel. This study investigated the application of the portable GeneXpert platform for WS of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus (IAV), influenza B virus (IBV), and respiratory syncytial virus (RSV). The GeneXpert system with the Xpert Xpress-SARS-CoV-2/Flu/RSV test kit uses reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to analyze wastewater samples. From September 2022 through January 2023, wastewater samples were collected from the influents of municipal wastewater treatment plants (MWTPs) of Saskatoon, Prince Albert, and North Battleford in the province of Saskatchewan, Canada. Both raw and concentrated wastewater samples were subjected to the GeneXpert analysis. Results showed that the Saskatoon wastewater viral loads were significantly correlated to Saskatchewan's influenza and COVID-19 clinical cases, with a lead time of 10 days for IAV and a lag time of 4 days for SARS-CoV-2. Additionally, the GeneXpert analysis of the three cities' wastewater samples showed that the raw WS could capture the dynamics of SARS-CoV-2 and IAV due to their correlation with concentrated WS. Interestingly, IBV loads were not detected in any wastewater samples, while the Saskatoon and Prince Albert wastewater samples collected following the 2023 holiday season (end of December and beginning of January) were positive for RSV. This study indicates that the GeneXpert has excellent potential for use in the development of an early warning system for transmissible disease in municipalities and limited-resource communities while simultaneously providing stakeholders with an efficient WS methodology.

Levels and composition of microplastics and microfibers in the South Saskatchewan River and stormwater retention ponds in the City of Saskatoon, Canada.

In recent decades, contamination of the environment with microplastics and microfibers has been recognized as a pervasive and ubiquitous issue of global concern. While much research in this field has been undertaken in marine environments, more recent studies have identified rivers as major conveyors of plastic pollution from terrestrial into marine systems. However, reports on the levels and composition of microplastic and microfiber contamination in rivers of the Canadian prairie region, specifically the South Saskatchewan River (SSR), are scarce, which leaves this vital source of water for societies and ecosystems in a vulnerable state. To fill this gap, we obtained samples from seven sites along the Saskatchewan portion of the SSR, as well as three stormwater retention ponds (SRP) in the city of Saskatoon during the spring, summer, and fall of 2020. We used optical and Raman microscopy to enumerate and characterize particles in these samples. Total levels of particles and fibers in all samples ranged from 32 to 116 particles m-3. Most particles (approximately 77%) were natural fibers, while polymers accounted for the remaining 33%. Average microplastic levels were lower (3.18 ± 3 particles m-3) downstream of Lake Diefenbaker, a large reservoir on the SSR, compared to upstream (12.0 ± 9 particles m-3). Retention of microplastics in the reservoir could explain the lower mean microplastic concentration of 4.43 ± 3 particles m-3 recorded in the SSR compared to mean concentrations of 26.2 ± 18 particles m-3 reported in the North Saskatchewan River, which is not dammed. This study is among the first to describe microplastic and microfiber levels in the SSR and thereby helps improve our understanding of this pervasive environmental contaminant on the Canadian prairies.

Apical and mechanistic effects of 6PPD-quinone on different life-stages of the fathead minnow (Pimephales promelas).

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine-quinone (6PPD-quinone) is an emerging contaminant of concern that is generated through the environmental oxidation of the rubber tire anti-degradant 6PPD. Since the initial report of 6PPD-quinone being the cause of urban runoff mortality syndrome of Coho salmon, numerous species have been identified as either sensitive or insensitive to acute lethality caused by 6PPD-quinone. In sensitive species, acute lethality might be caused by uncoupling of mitochondrial respiration in gills. However, little is known about effects of 6PPD-quinone on insensitive species. Here we demonstrate that embryos of fathead minnows (Pimephales promelas) are insensitive to exposure to concentrations as great as 39.97 µg/L for 168 h, and adult fathead minnows are insensitive to exposure to concentrations as great as 9.4 µg/L for 96 h. A multi-omics approach using a targeted transcriptomics array, (EcoToxChips), and proton nuclear magnetic resonance (1H NMR) was used to assess responses of the transcriptomes and metabolomes of gills and livers from adult fathead minnows exposed to 6PPD-quinone for 96 h to begin to identify sublethal effects of 6PPD-quinone. There was little agreement between results of the EcoToxChip and metabolomics analyses, likely because genes present on the EcoToxChip were not representative of pathways suggested to be perturbed by metabolomic analysis. Changes in abundances of transcripts and metabolites in livers and gills suggest that disruption of one­carbon metabolism and induction of oxidative stress might be occurring in gills and livers, but that tissues differ in their sensitivity or responsiveness to 6PPD-quinone. Overall, several pathways impacted by 6PPD-quinone were identified as candidates for future studies of potential sublethal effects of this chemical.

Toxicity and mutagenicity studies of 6PPD-quinone in a marine invertebrate species and bacteria.

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine-quinone (6PPD-quinone), an oxidation product of the tire additive, 6PPD, has been associated with high mortality of salmonids (0.1 µg/L). The objective of this study was to determine the acute toxicity using neonates and mutagenicity (micronuclei in hemolymph of exposed adults) of 6PPD-quinone in the marine amphipod Parhyale hawaiensis. Also, we studied its mutagenicity in the Salmonella/microsome assay using five strains of Salmonella with and without metabolic system (rat liver S9, 5%). 6PPD-quinone did not present acute toxicity to P. hawaiensis from 31.25 to 500 µg/L. Micronuclei frequency increased after 96 h-exposure to 6PPD-quinone (250 and 500 µg/L) when compared to the negative control. 6PPD-quinone also showed a weak mutagenic effect for TA100 only in the presence of S9. We conclude that 6PPD-quinone is mutagenic to P. hawaiensis and weakly mutagenic to bacteria. Our work provides information for future risk assessment of the presence of 6PPD-quinone in the aquatic environment.

Contrast-enhanced ultrasound imaging for assessment of intestinal inflammation in rainbow trout.

The intestine is essential for nutritional uptake and as a barrier to pathogens. Inflammation of the intestine can result from chemical contaminants, dietary irritants, or disease and may lead to serious health consequences, including reduced growth rates or increased pathogen susceptibility. Traditionally, intestinal inflammation in fish has been detected through histology completed post-mortem through excision and processing of the affected tissue. However, in human clinical settings, tools have been developed to assess intestinal inflammation non-invasively. Contrast-enhanced ultrasound (CEUS) imaging is an important tool for measuring inflammation in patients because it is cost-effective and minimally invasive. Specifically, CEUS allows real-time visualization and quantification of vascular perfusion. Changes in blood flow are typical in areas of inflamed or diseased tissue, and by measuring these changes, it is possible to assess the degree of inflammation. We demonstrate that standard CEUS protocols used for small mammals can be used to quantify vascular perfusion in the intestines of rainbow trout. Our resolution was sufficient to measure a significant difference in perfusion between control and TNBS-inflamed trout intestines, with inflamed intestines showing decreased perfusion. The presence of inflammation in the TNBS-treated intestines was verified ex vivo histologically and was characterized by the thickening of intestinal folds. The minimally invasive nature of CEUS imaging presents the opportunity to evaluate intestinal health in novel ways by allowing for longitudinal observations and avoiding mortality in valuable or at-risk specimens. Recent developments of highly portable, cost-effective CEUS systems will allow broad application of this tool, from industry to research.

Predicting the Kinetics of Resupply of Organic Pollutants from Sediments Using Diffusive Gradients in Thin Film Samplers and their Bioavailability to Aquatic Invertebrates.

The present study used diffusive gradients in thin film (DGT) samplers deployed in situ at a wastewater-impacted site (Clarkboro Ferry) for 20 days to develop a predictive model between time-weighted mean concentrations of seven selected antipsychotic compounds in water and those in resident benthic invertebrates, specifically crayfish (Faxonius virilis). The model was further combined with a model of desorption of antipsychotic compounds to predict kinetics at the sediment-water interface. Antipsychotic compounds were mostly detected in adult crayfish and internal concentrations were similar among targeted compounds, except for lesser concentrations of duloxetine. The model, based on the mass balance of organic chemicals, to predict uptake by organisms exhibited good agreement with measured values (R2 = 0.53-0.88), except for venlafaxine (R2 = 0.35). At the sediment-water interface, positive fluxes were observed for antipsychotic compounds and the results from DGT-induced fluxes in sediments (DIFS) coupled with equilibrium hydroxyl-ß-cyclodextrin extraction further indicated partial resupply of antipsychotic compounds from sediments to the aqueous phase, despite the labile pool being relatively limited. The results of the present study affirm that DGT techniques can be used as a predictive tool for contamination in benthic invertebrates and can simulate the ability of contaminant resupply from sediments. Environ Toxicol Chem 2023;42:1696-1708. © 2023 SETAC.

Understanding common population markers for SARS-CoV-2 RNA normalization in wastewater - A review.

Wastewater monitoring and epidemiology have seen renewed interest during the recent COVID-19 pandemic. As a result, there is an increasing need to normalize wastewater-derived viral loads in local populations. Chemical tracers, both exogenous and endogenous compounds, have proven to be more stable and reliable for normalization than biological indicators. However, differing instrumentation and extraction methods can make it difficult to compare results. This review examines current extraction and quantification methods for ten common population indicators: creatinine, coprostanol, nicotine, cotinine, sucralose, acesulfame, androstenedione 5-hydroindoleacetic acid (5-HIAA), caffeine, and 1,7-dimethyluric acid. Some wastewater parameters such as ammonia, total nitrogen, total phosphorus, and daily flowrate were also evaluated. The analytical methods included direct injection, dilute and shoot, liquid/liquid, and solid phase extraction (SPE). Creatine, acesulfame, nicotine, 5-HIAA and androstenedione have been analysed by direct injection into LC-MS; however, most authors prefer to include SPE steps to avoid matrix effects. Both LC-MS and GC-MS have been successfully used to quantify coprostanol in wastewater, and the other selected indicators have been quantified successfully with LC-MS. Acidification to stabilize the sample before freezing to maintain the integrity of samples has been reported to be beneficial. However, there are arguments both for and against working at acidic pHs. Wastewater parameters mentioned earlier are quick and easy to quantify, but the data does not always represent the human population effectively. A preference for population indicators originating solely from humans is apparent. This review summarises methods employed for chemical indicators in wastewater, provides a basis for choosing an appropriate extraction and analysis method, and highlights the utility of accurate chemical tracer data for wastewater-based epidemiology.

Wastewater discharges alter microbial community composition in surface waters of the canadian prairies.

Microbial communities are an important component of freshwater biodiversity that is threatened by anthropogenic impacts. Wastewater discharges pose a particular concern by being major sources of anthropogenic contaminants and microorganisms that may influence the composition of natural microbial communities. Nevertheless, the effects of wastewater treatment plant (WWTP) effluents on microbial communities remain largely unexplored. In this study, the effects of wastewater discharges on microbial communities from five different WWTPs in Southern Saskatchewan were investigated using rRNA gene metabarcoding. In parallel, nutrient levels and the presence of environmentally relevant organic pollutants were analyzed. Higher nutrient loads and pollutant concentrations resulted in significant changes in microbial community composition. The greatest changes were observed in Wascana Creek (Regina), which was found to be heavily polluted by wastewater discharges. Several taxa occurred in greater relative abundance in the wastewater-influenced stream segments, indicating anthropogenic pollution and eutrophication, especially taxa belonging to Proteobacteria, Bacteroidota, and Chlorophyta. Strong decreases were measured within the taxa Ciliphora, Diatomea, Dinoflagellata, Nematozoa, Ochrophyta, Protalveolata, and Rotifera. Across all sample types, a significant decline in sulfur bacteria was measured, implying changes in functional biodiversity. In addition, downstream of the Regina WWTP, an increase in cyanotoxins was detected which was correlated with a significant change in cyanobacterial community composition. Overall, these data suggest a causal relationship between anthropogenic pollution and changes in microbial communities, possibly reflecting an impairment of ecosystem health.

A wastewater-based risk index for SARS-CoV-2 infections among three cities on the Canadian Prairie.

Wastewater surveillance (WWS) is useful to better understand the spreading of coronavirus disease 2019 (COVID-19) in communities, which can help design and implement suitable mitigation measures. The main objective of this study was to develop the Wastewater Viral Load Risk Index (WWVLRI) for three Saskatchewan cities to offer a simple metric to interpret WWS. The index was developed by considering relationships between reproduction number, clinical data, daily per capita concentrations of virus particles in wastewater, and weekly viral load change rate. Trends of daily per capita concentrations of SARS-CoV-2 in wastewater for Saskatoon, Prince Albert, and North Battleford were similar during the pandemic, suggesting that per capita viral load can be useful to quantitatively compare wastewater signals among cities and develop an effective and comprehensible WWVLRI. The effective reproduction number (Rt) and the daily per capita efficiency adjusted viral load thresholds of 85 × 106 and 200 × 106 N2 gene counts (gc)/population day (pd) were determined. These values with rates of change were used to categorize the potential for COVID-19 outbreaks and subsequent declines. The weekly average was considered 'low risk' when the per capita viral load was 85 × 106 N2 gc/pd. A 'medium risk' occurs when the per capita copies were between 85 × 106 and 200 × 106 N2 gc/pd. with a rate of change <100 %. The start of an outbreak is indicated by a 'medium-high' risk classification when the week-over-week rate of change was >100 %, and the absolute magnitude of concentrations of viral particles was >85 × 106 N2 gc/pd. Lastly, a 'high risk' occurs when the viral load exceeds 200 × 106 N2 gc/pd. This methodology provides a valuable resource for decision-makers and health authorities, specifically given the limitation of COVID-19 surveillance based on clinical data.

Acute Exposure of Zebrafish (Danio rerio) to the Next-Generation Perfluoroalkyl Substance, Perfluoroethylcyclohexanesulfonate, Shows Similar Effects as Legacy Substances.

Perfluoroethylcyclohexanesulfonate (PFECHS) is an emerging perfluoroalkyl substance (PFAS) that has been considered a potential replacement for perfluorooctanesulfonic acid (PFOS). However, there is little information characterizing the toxic potency of PFECHS to zebrafish embryos and its potential for effects in aquatic environments. This study assessed toxic potency of PFECHS in vivo during both acute (96-hour postfertilization) and chronic (21-day posthatch) exposures and tested concentrations of PFECHS from 500 ng/L to 2 mg/L. PFECHS was less likely to cause mortalities than PFOS for both the acute and chronic experiments based on previously published values for PFOS exposure, but exposure resulted in a similar incidence of deformities. Exposure to PFECHS also resulted in significantly increased abundance of transcripts of peroxisome proliferator activated receptor alpha (pparα), cytochrome p450 1a1 (cyp1a1), and apolipoprotein IV (apoaIV) at concentrations nearing those of environmental relevance. Overall, these results provide further insight into the safety of an emerging PFAS alternative in the aquatic environment and raise awareness that previously considered "safer" alternatives may show similar effects as legacy PFASs.

Perfluoroethylcyclohexane sulphonate, an emerging perfluoroalkyl substance, disrupts mitochondrial membranes and the expression of key molecular targets in vitro.

Perfluoroethylcyclohexane sulphonate (PFECHS) is an emerging, replacement perfluoroalkyl substance (PFAS) with little information available on the toxic effects or potencies with which to characterize its potential impacts on aquatic environments. This study aimed to characterize effects of PFECHS using in vitro systems, including rainbow trout liver cells (RTL-W1 cell line) and lymphocytes separated from whole blood. It was determined that exposure to PFECHS caused minor acute toxic effects for most endpoints and that little PFECHS was concentrated into cells with a mean in vitro bioconcentration factor of 81 ± 25 L/kg. However, PFECHS was observed to affect the mitochondrial membrane and key molecular receptors, such as the peroxisome proliferator receptor, cytochrome p450-dependent monooxygenases, and receptors involved in oxidative stress. Also, glutathione-S-transferase was significantly down-regulated at a near environmentally relevant exposure concentration of 400 ng/L. These results are the first to report bioconcentration of PFECHS, as well as its effects on the peroxisome proliferator and glutathione-S-transferase receptors, suggesting that even with little bioconcentration, PFECHS has potential to cause adverse effects.