A wastewater-based risk index for SARS-CoV-2 infections among three cities on the Canadian Prairie
Femi Francis Oloye,
Jonathan K. Challis,
Kerry N. McPhedran,
Chantel De Lange,
Mark R. Servos,
Paul D. Jones,
John P. Giesy,
Science of The Total Environment, Volume 876
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.
A comparative analysis of the partitioning behaviour of SARS-CoV-2 RNA in liquid and solid fractions of wastewater
Patrick R. Breadner,
Hadi A. Dhiyebi,
Marc G. Aucoin,
Scott Joseph Boegel,
Leslie M. Bragg,
Paul M. Craig,
John P. Giesy,
Mark R. Servos
Science of The Total Environment, Volume 895
As fragments of SARS-CoV-2 RNA can be quantified and measured temporally in wastewater, surveillance of concentrations of SARS-CoV-2 in wastewater has become a vital resource for tracking the spread of COVID-19 in and among communities. However, the absence of standardized methods has affected the interpretation of data for public health efforts. In particular, analyzing either the liquid or solid fraction has implications for the interpretation of how viral RNA is quantified. Characterizing how SARS-CoV-2 or its RNA fragments partition in wastewater is a central part of understanding fate and behaviour in wastewater. In this study, partitioning of SARS-CoV-2 was investigated by use of centrifugation with varied durations of spin and centrifugal force, polyethylene glycol (PEG) precipitation followed by centrifugation, and ultrafiltration of wastewater. Partitioning of the endogenous pepper mild mottled virus (PMMoV), used to normalize the SARS-CoV-2 signal for fecal load in trend analysis, was also examined. Additionally, two surrogates for coronavirus, human coronavirus 229E and murine hepatitis virus, were analyzed as process controls. Even though SARS-CoV-2 has an affinity for solids, the total RNA copies of SARS-CoV-2 per wastewater sample, after centrifugation (12,000 g, 1.5 h, no brake), were partitioned evenly between the liquid and solid fractions. Centrifugation at greater speeds for longer durations resulted in a shift in partitioning for all viruses toward the solid fraction except for PMMoV, which remained mostly in the liquid fraction. The surrogates more closely reflected the partitioning of SARS-CoV-2 under high centrifugation speed and duration while PMMoV did not. Interestingly, ultrafiltration devices were inconsistent in estimating RNA copies in wastewater, which can influence the interpretation of partitioning. Developing a better understanding of the fate of SARS-CoV-2 in wastewater and creating a foundation of best practices is the key to supporting the current pandemic response and preparing for future potential infectious diseases.
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.
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.
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.
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.
DNA metabarcoding can provide a high-throughput and rapid method for characterizing responses of communities to environmental stressors. However, within bulk samples, DNA metabarcoding hardly distinguishes live from the dead organisms. Here, both DNA and RNA metabarcoding were applied and compared in experimental freshwater mesocosms conducted for assessment of ecotoxicological responses of zooplankton communities to remediation treatment until 38 days post oil-spill. Furthermore, a novel indicator of normalized vitality (NV), sequence counts of RNA metabarcoding normalized by that of DNA metabarcoding, was developed for assessment of ecological responses. DNA and RNA metabarcoding detected similar taxa richness and rank of relative abundances. Both DNA and RNA metabarcoding demonstrated slight shifts in measured α-diversities in response to treatments. NV presented relatively greater magnitudes of differential responses of community compositions to treatments compared to DNA or RNA metabarcoding. NV declined from the start of the experiment (3 days pre-spill) to the end (38 days post-spill). NV also differed between Rotifer and Arthropoda, possibly due to differential life histories and sizes of organisms. NV could be a useful indicator for characterizing ecological responses to anthropogenic influence; however, the biology of target organisms and subsequent RNA production need to be considered. • RNA normalized by DNA metabarcoding functions as normalized vitality. • Normalized vitality reflected temporal dynamics of zooplankton communities. • Normalized vitality revealed greater community differences between treatments. • Rotifer had greatest normalized vitality compared to Arthropoda.
Absorption and elimination of per and poly-fluoroalkyl substances substitutes in salmonid species after pre-fertilization exposure
Paul D. Jones,
Jason C. Raine,
John P. Giesy
Science of The Total Environment, Volume 814
Due to their relatively large production and few restrictions on uses, novel substitutes for historically used per and poly-fluoroalkyl substances (PFAS) are being used and accumulating in the environment. However, due to a lack of information on their toxicological properties their hazards and risks are hard to estimate. Before fertilization, oocytes of two salmonid species, Arctic Char (Salvelinus alpinus) and Rainbow Trout (Oncorhynchus mykiss), were exposed to three PFAS substances used as substitutes for traditional PFAS, PFBA, PFBS or GenX or two archetypical, historically used, longer-chain PFAS, PFOA and PFOS. Exposed oocytes were subsequently fertilized, incubated and were sampled during several developmental stages, until swim-up. All five PFAS were accumulated into egg yolks with similar absorption rates, and their concentrations in egg yolks were less than respective concentrations in/on egg chorions. Rapid elimination of the five PFAS was observed during the first 3 days after fertilization. Thereafter, amounts of PFOS and PFOA were stable until swim-up, while PFBA, PFBS and GenX were further eliminated during development from one month after the fertilization to swim-up. In these two salmonid species, PFBA, PFBS and GenX were eliminated faster than were PFOS or PFOA.
The microbiome of the gut is vital for homeostasis of hosts with its ability to detoxify and activate toxicants, as well as signal to the immune and nervous systems. However, in the field of environmental toxicology, the gut microbiome has only recently been identified as a measurable indicator for exposure to environmental pollutants. Antidepressants found in effluents of wastewater treatment plants and surface waters have been shown to exhibit antibacterial-like properties in vitro, where some bacteria are known to express homologous proteins that bind antidepressants in vertebrates. Therefore, it has been hypothesized that exposure to antidepressant drugs might affect gut microbiota of aquatic organisms. In this study, the common antidepressant, fluoxetine, was investigated to determine whether it can modulate the gut microbiome of adult fathead minnows. A 28-day, sub-chronic, static renewal exposure was performed with nominal fluoxetine concentrations of 0.01, 10 or 100 μg/L. Using 16S rRNA amplicon sequencing, shifts among the gut-associated microbiota were observed in individuals exposed to the greatest concentration, with greater effects observed in females. These changes were associated with a decrease in relative proportions of commensal bacteria, which can be important for health of fish including bacteria essential for fatty acid oxidation, and an increase in relative proportions of pathogenic bacteria associated with inflammation. Results demonstrate, for the first time, how antidepressants found in some aquatic environments can influence gut microbiota of fishes.
RNA in Municipal Wastewater Reveals Magnitudes of COVID-19 Outbreaks across Four Waves Driven by SARS-CoV-2 Variants of Concern
Jonathan K. Challis,
Femi Francis Oloye,
Kerry N. McPhedran,
Natacha S. Hogan,
Paul D. Jones,
Chand S. Mangat,
Mark R. Servos,
John P. Giesy
ACS ES&T Water, Volume 2, Issue 11
There are no standardized protocols for quantifying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wastewater to date, especially for population normalization. Here, a pipeline was developed, applied, and assessed to quantify SARS-CoV-2 and key variants of concern (VOCs) RNA in wastewater at Saskatoon, Canada. Normalization approaches using recovery ratio and extraction efficiency, wastewater parameters, or population indicators were assessed by comparing to daily numbers of new cases. Viral load was positively correlated with daily new cases reported in the sewershed. Wastewater surveillance (WS) had a lead time of approximately 7 days, which indicated surges in the number of new cases. WS revealed the variant α and δ driving the third and fourth wave, respectively. The adjustment with the recovery ratio and extraction efficiency improved the correlation between viral load and daily new cases. Normalization of viral concentration to concentrations of the artificial sweetener acesulfame K improved the trend of viral load during the Christmas and New Year holidays when populations were dynamic and variable. Acesulfame K performed better than pepper mild mottle virus, creatinine, and ammonia for population normalization. Hence, quality controls to characterize recovery ratios and extraction efficiencies and population normalization with acesulfame are promising for precise WS programs supporting decision-making in public health.
Activities of gut microbiomes are often overlooked in assessments of ecotoxicological effects of environmental contaminants. Effects of the polycyclic aromatic hydrocarbon, benzo[a]pyrene (BaP) on active gut microbiomes of juvenile fathead minnows (Pimephales promelas) were investigated. Fish were exposed for two weeks, to concentrations of 0, 1, 10, 100, or 1000 μg BaP g-1 in the diet. The active gut microbiome was characterized using 16S rRNA metabarcoding to determine its response to dietary exposure of BaP. BaP reduced alpha-diversity at the greatest exposure concentrations. Additionally, exposure to BaP altered community composition of active microbiome and resulted in differential proportion of taxa associated with hydrocarbon degradation and fish health. Neighborhood selection networks of active microbiomes were not reduced with greater concentrations of BaP, which suggests ecological resistance and/or resilience of gut microbiota. The active gut microbiome had a similar overall biodiversity as that of the genomic gut microbiota, but had a distinct composition from that of the 16S rDNA profile. Responses of alpha- and beta-diversities of the active microbiome to BaP exposure were consistent with that of genomic microbiomes. Normalized activity of microbiome via the ratio of rRNA to rDNA abundance revealed rare taxa that became active or dormant due to exposure to BaP. These differences highlight the need to assess both 16S rDNA and rRNA metabarcoding to fully derive bacterial compositional changes resulting from exposure to contaminants.
Widespread application of poly- and per-fluoroalkyl substances (PFAS) has resulted in some substances being ubiquitous in environmental matrices. That and their resistance to degradation have allowed them to accumulate in wildlife and humans with potential for toxic effects. While specific substances of concern have been phased-out or banned, other PFAS that are emerging as alternative substances are still produced and are being released into the environment. This review focuses on describing three emerging, replacement PFAS: perfluoroethylcyclohexane sulphonate (PFECHS), 6:2 chlorinated polyfluoroalkyl ether sulfonate (6:2 Cl-PFAES), and hexafluoropropylene oxide dimer acid (HFPO-DA). By summarizing their physicochemical properties, environmental fate and transport, and toxic potencies in comparison to other PFAS compounds, this review offers insight into the viabilities of these chemicals as replacement substances. Using the chemical scoring and ranking assessment model, the relative hazards, uncertainties, and data gaps for each chemical were quantified and related to perfluorooctane sulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) based on their chemical and uncertainty scores. The substances were ranked PFOS > 6:2 Cl-PFAES > PFOA > HFPO-DA > PFECHS according to their potential toxicity and PFECHS > HFPO-DA > 6:2 Cl-PFAES > PFOS > PFOA according to their need for future research. Since future uses of PFAS remain uncertain in the face of governmental regulations and production bans, replacement PFAS will continue to emerge on the world market and in the environment, raising concerns about their general lack of information on mechanisms and toxic potencies.
Effects of in situ experimental selenium exposure on finescale dace (Phoxinus neogaeus) gut microbiome
Phillip J. Ankley,
Stephanie D. Graves,
David M. Janz,
John P. Giesy
Environmental Research, Volume 212
Selenium (Se) is an environmental contaminant of global concern that can cause adverse effects in fish at elevated levels. Fish gut microbiome play essential roles in gastrointestinal function and host health and can be perturbed by environmental contaminants, including metals and metalloids. Here, an in-situ Se exposure of female finescale dace (Phoxinus neogaeus) using mesocosms was conducted to determine the impacts of Se accumulation on the gut microbiome and morphometric endpoints. Prior to this study, the gut microbiome of finescale dace, a widespread Cyprinid throughout North America, had not been characterized. Exposure to Se caused a hormetic response of alpha diversity of the gut microbiome, with greater diversity at the lesser concentration of 1.6 μg Se/L, relative to that of fish exposed to the greater concentration of 5.6 μg Se/L. Select gut microbiome taxa of fish were differentially abundant between aqueous exposure concentrations and significantly correlated with liver-somatic index (LSI). The potential effects of gut microbiome dysbiosis on condition of wild fish might be a consideration when assessing adverse effects of Se in aquatic environments. More research regarding effects of Se on field-collected fish gut microbiome and the potential adverse effects or benefits on the host is warranted.
Rapid transition between SARS-CoV-2 variants of concern Delta and Omicron detected by monitoring municipal wastewater from three Canadian cities
Femi Francis Oloye,
Jonathan K. Challis,
Kerry N. McPhedran,
Mark P. Keller,
Paul D. Jones,
Chand S. Mangat,
Mark R. Servos,
John P. Giesy
Science of The Total Environment, Volume 841
Monitoring the communal incidence of COVID-19 is important for both government and residents of an area to make informed decisions. However, continuous reliance on one means of monitoring might not be accurate because of biases introduced by government policies or behaviours of residents. Wastewater surveillance was employed to monitor concentrations of SARS-CoV-2 RNA in raw influent wastewater from wastewater treatment plants serving three Canadian Prairie cities with different population sizes. Data obtained from wastewater are not directly influenced by government regulations or behaviours of individuals. The means of three weekly samples collected using 24 h composite auto-samplers were determined. Viral loads were determined by RT-qPCR, and whole-genome sequencing was used to charaterize variants of concern (VOC). The dominant VOCs in the three cities were the same but with different proportions of sub-lineages. Sub-lineages of Delta were AY.12, AY.25, AY.27 and AY.93 in 2021, while the major sub-lineage of Omicron was BA.1 in January 2022, and BA.2 subsequently became a trace-level sub-variant then the predominant VOC. When each VOC was first detected varied among cities; However, Saskatoon, with the largest population, was always the first to present new VOCs. Viral loads varied among cities, but there was no direct correlation with population size, possibly because of differences in flow regimes. Population is one of the factors that affects trends in onset and development of local outbreaks during the pandemic. This might be due to demography or the fact that larger populations had greater potential for inter- and intra-country migration. Hence, wastewater surveillance data from larger cities can typically be used to indicate what to expect in smaller communities.
Using zooplankton metabarcoding to assess the efficacy of different techniques to clean-up an oil-spill in a boreal lake
Phillip J. Ankley,
Tyler A. Black,
Mark L. Hanson,
Scott N. Higgins,
John P. Giesy,
Aquatic Toxicology, Volume 236
Abstract Regulators require adequate information to select best practices with less ecosystem impacts for remediation of freshwater ecosystems after oil spills. Zooplankton are valuable indicators of aquatic ecosystem health as they play pivotal roles in biochemical cycles while stabilizing food webs. Compared with morphological identification, metabarcoding holds promise for cost-effective, high-throughput, and benchmarkable biomonitoring of zooplankton communities. The objective of this study was to apply DNA and RNA metabarcoding of zooplankton for ecotoxicological assessment and compare it with traditional morphological identification in experimental shoreline enclosures in a boreal lake. These identification methods were also applied in context of assessing response of the zooplankton community exposed to simulated spills of diluted bitumen (dilbit), with experimental remediation practices (enhanced monitored natural recovery and shoreline cleaner application). Metabarcoding detected boreal zooplankton taxa up to the genus level, with a total of 24 shared genera, and while metabarcoding-based relative abundance served as an acceptable proxy for biomass inferred by morphological identification (ρ ≥ 0.52). Morphological identification determined zooplankton community composition changes due to treatments at 11 days post-spill (PERMANOVA, p = 0.0143) while metabarcoding methods indicated changes in zooplankton richness and communities at 38 days post-spill (T-test, p
Life Cycle Exposure to Environmentally Relevant Concentrations of Diphenyl Phosphate (DPhP) Inhibits Growth and Energy Metabolism of Zebrafish in a Sex-Specific Manner
Jonathan K. Challis,
John P. Giesy,
Environmental Science & Technology
Due to commercial uses and environmental degradation of aryl phosphate esters, diphenyl phosphate (DPhP) is frequently detected in environmental matrices and is thus of growing concern worldwide. However, information on potential adverse effects of chronic exposure to DPhP at environmentally realistic concentrations was lacking. Here, we investigated the effects of life cycle exposure to DPhP on zebrafish at environmentally relevant concentrations of 0.8, 3.9, or 35.6 μg/L and employed a dual-omics approach (metabolomics and transcriptomics) to characterize potential modes of action. Exposure to DPhP at 35.6 μg/L for 120 days resulted in significant reductions in body mass and length of male zebrafish, but did not cause those same effects to females. Predominant toxicological mechanisms, including inhibition of oxidative phosphorylation, down-regulation of fatty acid oxidation, and up-regulation of phosphatidylcholine degradation, were revealed by integrated dual-omics analysis and successfully linked to adverse outcomes. Activity of succinate dehydrogenase and protein content of carnitine O-palmitoyltransferase 1 were significantly decreased in livers of male fish exposed to DPhP, which further confirmed the proposed toxicological mechanisms. This study is the first to demonstrate that chronic, low-level exposure to DPhP can retard growth via inhibiting energy output in male zebrafish.
Comparison of approaches to quantify SARS-CoV-2 in wastewater using RT-qPCR: Results and implications from a collaborative inter-laboratory study in Canada
Alex H. S. Chik,
Melissa B. Glier,
Mark R. Servos,
Chand S. Mangat,
Patrick M. D’Aoust,
John P. Giesy,
R. Michael L. McKay,
Michael R. Mulvey,
Steve E. Hrudey
Journal of Environmental Sciences, Volume 107
Detection of SARS-CoV-2 RNA in wastewater is a promising tool for informing public health decisions during the COVID-19 pandemic. However, approaches for its analysis by use of reverse transcription quantitative polymerase chain reaction (RT-qPCR) are still far from standardized globally. To characterize inter- and intra-laboratory variability among results when using various methods deployed across Canada, aliquots from a real wastewater sample were spiked with surrogates of SARS-CoV-2 (gamma-radiation inactivated SARS-CoV-2 and human coronavirus strain 229E [HCoV-229E]) at low and high levels then provided "blind" to eight laboratories. Concentration estimates reported by individual laboratories were consistently within a 1.0-log10 range for aliquots of the same spiked condition. All laboratories distinguished between low- and high-spikes for both surrogates. As expected, greater variability was observed in the results amongst laboratories than within individual laboratories, but SARS-CoV-2 RNA concentration estimates for each spiked condition remained mostly within 1.0-log10 ranges. The no-spike wastewater aliquots provided yielded non-detects or trace levels (<20 gene copies/mL) of SARS-CoV-2 RNA. Detections appear linked to methods that included or focused on the solids fraction of the wastewater matrix and might represent in-situ SARS-CoV-2 to the wastewater sample. HCoV-229E RNA was not detected in the no-spike aliquots. Overall, all methods yielded comparable results at the conditions tested. Partitioning behavior of SARS-CoV-2 and spiked surrogates in wastewater should be considered to evaluate method effectiveness. A consistent method and laboratory to explore wastewater SARS-CoV-2 temporal trends for a given system, with appropriate quality control protocols and documented in adequate detail should succeed.
The microbiome has been described as an additional host “organ” with well-established beneficial roles. However, the effects of exposures to chemicals on both structure and function of the gut microbiome of fishes are understudied. To determine effects of benzo[ a ]pyrene (BaP), a model persistent organic pollutant, on structural shifts of gut microbiome in juvenile fathead minnows ( Pimephales promelas ), fish were exposed ad libitum in the diet to concentrations of 1, 10, 100, or 1000 μg BaP g −1 food, in addition to a vehicle control, for two weeks. To determine the link between exposure to BaP and changes in the microbial community, concentrations of metabolites of BaP were measured in fish bile and 16S rRNA amplicon sequencing was used to evaluate the microbiome. Exposure to BaP only reduced alpha-diversity at the greatest exposure concentrations. However, it did alter community composition assessed as differential abundance of taxa and reduced network complexity of the microbial community in all exposure groups. Results presented here illustrate that environmentally-relevant concentrations of BaP can alter the diversity of the gut microbiome and community network connectivity. Highlights • Dominant phyla of gut microbiome are consistent with those of other freshwater fishes. • BaP metabolites and exposure doses were consistent with those found in contaminated sites. • Dietary BaP exposure has significant, dose-dependent effects on the fish gut microbiome. • Dietary BaP exposure altered association networks of gut microbiome. Environmentally-relevant concentrations of BaP can alter the diversity of the gut microbiome and community network connectivity via dietary exposure route.
Remodeling of Arctic char ( <i>Salvelinus alpinus</i> ) lipidome under a stimulated scenario of Arctic warming
Jonathan K. Challis,
Jason C. Raine,
Paul D. Jones,
John P. Giesy
Global Change Biology, Volume 27, Issue 14
Arctic warming associated with global climate change poses a significant threat to populations of wildlife in the Arctic. Since lipids play a vital role in adaptation of organisms to variations in temperature, high-resolution mass-spectrometry-based lipidomics can provide insights into adaptive responses of organisms to a warmer environment in the Arctic and help to illustrate potential novel roles of lipids in the process of thermal adaption. In this study, we studied an ecologically and economically important species-Arctic char (Salvelinus alpinus)-with a detailed multi-tissue analysis of the lipidome in response to chronic shifts in temperature using a validated lipidomics workflow. In addition, dynamic alterations in the hepatic lipidome during the time course of shifts in temperature were also characterized. Our results showed that early life stages of Arctic char were more susceptible to variations in temperature. One-year-old Arctic char responded to chronic increases in temperature with coordinated regulation of lipids, including headgroup-specific remodeling of acyl chains in glycerophospholipids (GP) and extensive alterations in composition of lipids in membranes, such as less lyso-GPs, and more ether-GPs and sphingomyelin. Glycerolipids (e.g., triacylglycerol, TG) also participated in adaptive responses of the lipidome of Arctic char. Eight-week-old Arctic char exhibited rapid adaptive alterations of the hepatic lipidome to stepwise decreases in temperature while showing blunted responses to gradual increases in temperature, implying an inability to adapt rapidly to warmer environments. Three common phosphatidylethanolamines (PEs) (PE 36:6|PE 16:1_20:5, PE 38:7|PE 16:1_22:6, and PE 40:7|PE 18:1_22:6) were finally identified as candidate lipid biomarkers for temperature shifts via machine learning approach. Overall, this work provides additional information to a better understanding of underlying regulatory mechanisms of the lipidome of Arctic organisms in the face of near-future warming.
Environmental DNA of preservative ethanol performed better than water samples in detecting macroinvertebrate diversity using metabarcoding
John P. Giesy,
Diversity and Distributions, Volume 27, Issue 10
High‐throughput pipelines supported by eDNA metabarcoding have been applied in various freshwater ecosystems. Both eDNA in ethanol (EtOH) samples (ES‐eDNA) and in water samples (WS‐eDNA) can provide comprehensive classification lists with good taxonomic resolution and coverage for determining freshwater biodiversity and biomonitoring. But, the advantages of ES‐eDNA metabarcoding over WS‐eDNA metabarcoding remain unclear for routine assessments of diversity of benthic macroinvertebrates in streams.
In addition to aiding in digestion of food and uptake of nutrients, microbiota in guts of vertebrates are responsible for regulating several beneficial functions, including development of an organism and maintaining homeostasis. However, little is known about effects of exposures to chemicals on structure and function of gut microbiota of fishes. To assess effects of exposure to polycyclic aromatic hydrocarbons (PAHs) on gut microbiota, male and female fathead minnows ( Pimephales promelas ) were exposed to environmentally-relevant concentrations of the legacy PAH benzo[ a ]pyrene (BaP) in water. Measured concentrations of BaP ranged from 2.3 × 10 −3 to 1.3 μg L −1 . The community of microbiota in the gut were assessed by use of 16S rRNA metagenetics. Exposure to environmentally-relevant aqueous concentrations of BaP did not alter expression levels of mRNA for cyp1a1 , a “classic” biomarker of exposure to BaP, but resulted in shifts in relative compositions of gut microbiota in females rather than males. Results presented here illustrate that in addition to effects on more well-studied molecular endpoints, relative compositions of the microbiota in guts of fish can also quickly respond to exposure to chemicals, which can provide additional mechanisms for adverse effects on individuals. • Female and male fathead minnows exhibited significantly different gut microbiota. • Exposure to BaP altered structures in female gut microbiota, but not in males. • Exposure to BaP altered predicted functions in gut microbiota of fathead minnow. • Gut microbiome was more sensitive to a low dose BaP than host’s ahr1 and cyp1a1.
Integrated assessment of west coast of South Korea by use of benthic bacterial community structure as determined by eDNA, concentrations of contaminants, and in vitro bioassays
Aslan Hwanhwi Lee,
John P. Giesy,
Jong Seong Khim
Environment International, Volume 137
During the past few decades, contamination of sediments by persistent toxic substances (PTSs) has been observed in estuarine and coastal areas on the west coast of South Korea. The contaminants are suspected to cause toxicities in aquatic biota, but little is known about their ecological effects, particularly on benthic microbial communities. In this study, an eDNA-based assessment was applied along with classic assessments of exposure, such as chemistry and in vitro bioassays, to evaluate condition of benthic bacterial communities subjected to PTSs. Two strategies were adopted for the study. One was to conduct a comprehensive assessment in space (by comparing seawater and freshwater sites at five coastal regions) and in time (by following change over a 5-y period). Although we found that bacterial composition varied among and within years, some phyla, such as Proteobacteria (28.7%), Actinobacteria (13.1%), Firmicutes (12.7%), and Chloroflexi (12.5%) were consistently dominated across the study regions. Certain bacterial groups, such as Firmicutes and Verrucomicrobia have been linked to contamination at some sites in the study area and at specific points in time. Bacterial communities were not significantly correlated with salinity or AhR- and ER-mediated potencies, whereas concentrations of PAHs, APs, and certain metals (Cd and Hg) exhibited significant associations to the structure of bacterial communities at the phylum level. In fact, the relative abundance of microbes in the phylum Planctomycetes was significantly and negatively correlated with concentrations of PAHs and metals. Thus, the relative abundance of Planctomycetes could be used as an indicator of sedimentary contamination by PAHs and/or metals. Based on our correlation analyses, Cd and ER-mediated potencies were associated more with bacterial abundances at the class taxonomic level than were other PTSs and metals. Overall, the eDNA-based assessment was useful by augmenting more traditional measures of exposure and responses in a sediment triad approach and has potential as a more rapid screening tool.
• Concentrations of PAHs in muscle suggests continued exposure to the residual spilled oil. • Identity of the host species was the dominant driver in shaping the gut microbiome of fish. • Structures of gut microbiomes were correlated with concentrations of PAHs in muscle in walleye. In July 2016, a Husky Energy pipeline spilled 225,000 L of diluted heavy crude oil, with a portion of the oil entering the North Saskatchewan River near Maidstone, SK, Canada. This event provided a unique opportunity to assess potential effects of a crude oil constituent (namely polycyclic aromatic hydrocarbons, PAHs) on a possible sensitive indicator of freshwater ecosystem health, the gut microbiota of native fishes. In summer 2017, goldeye ( Hiodon alosoides ), walleye ( Sander vitreus ), northern pike ( Esox lucius ), and shorthead redhorse ( Moxostoma macrolepidotum ) were collected at six locations upstream and downstream of the spill. Muscle and bile were collected from individual fish for quantification of PAHs and intestinal contents were collected for characterization of the microbial community of the gut. Results suggested that host species is a significant determinant of gut microbiota, with significant differences among the species across sites. Concentrations of PAHs in dorsal muscle were significantly correlated with gut community compositions of walleye, but not of the other fishes. Concentrations of PAHs in muscle were also correlated with abundances of several families of bacteria among fishes. This study represents one of the first to investigate the response of the gut microbiome of wild fishes to chemical stressors.
Understanding the extent and directionality of the impact of human activities on ecosystems is directly related to their management and protection. However, the lack of historical data limits our understanding of ecosystem changes with long-term exposure to human activities. Recently, lake sedimentary DNA (sedDNA) has become a powerful tool for revealing changes in ecosystems at the century and millennium scales. Here, we used sedDNA to reveal the dynamic of the microbial community (including bacteria and micro-eukaryotes) in Lake Chao over the past 150 years, and further explored the effects of long-term nutrient and heavy metal loads on these communities. Our data show that nutrient and heavy metal loads in Lake Chao have increased by ca. 2 to 4-fold since the 1960s. In response, the community structure, diversity, and ecological network of bacteria and micro-eukaryotes changed significantly during the 1960s, the 1980s and the 2010s. Importantly, community structure was more sensitive to human activities than diversity. We also found that the relative abundance of some taxa associated with nitrification and algal blooms (e.g., taxa in Nitrospira sp., Peridinales) has increased ca. 100-fold since the 1960s. Nutrient could better explain the variation in the bacterial community (ca. twice as much as heavy metal), while heavy metal explained micro-eukaryotes better (ca. 3 or 5-fold as much as nutrient). In particular, based on parsimonious models from distance-based linear model (distLM), we further identified that Pb is the key factor affecting the bacterial and micro-eukaryotes community in Lake Chao in addition to nutrient. Our study reveals the impacts of long-term human activities on lake ecosystems from multiple perspectives of nutrient and heavy metal loads, community structure, diversity and ecological network, these findings will contribute to the management and conservation of lakes in the future.
Selenium (Se) enrichment has been demonstrated to vary by several orders of magnitude among species of planktonic algae. This is a substantial source of uncertainty when modelling Se biodynamics in aquatic systems. In addition, Se bioconcentration data are largely lacking for periphytic species of algae, and for multi-species periphyton biofilms, adding to the challenge of modelling Se transfer in periphyton-based food webs. To better predict Se dynamics in periphyton dominated, freshwater ecosystems, the goal of this study was to assess the relative influence of periphyton community composition on the uptake of waterborne Se oxyanions. Naturally grown freshwater periphyton communities, sampled from five different water bodies, were exposed to environmentally relevant concentrations of selenite [Se(IV)] or selenate [Se(VI)] (nominal concentrations of 5 and 25 μg Se L-1) under similar, controlled laboratory conditions for a period of 8 days. Unique periphyton assemblages were derived from the five different field sites, as confirmed by light microscopy and targeted DNA sequencing of the plastid 23S rRNA gene in algae. Selenium accumulation demonstrated a maximum of 23.6-fold difference for Se(IV) enrichment and 2.1-fold difference for Se(VI) enrichment across the periphyton/biofilm assemblages tested. The assemblage from one field site demonstrated both high accumulation of Se(IV) and iron, and was subjected to additional experimentation to elucidate the mechanism(s) of Se accumulation. Selenite accumulation (at nominal concentrations of 5 and 25 μg Se L-1 and mean pH of 7.5 across all treatment replicates) was assessed in both unaltered and heat-killed periphyton, and in periphyton from the same site grown without light to exclude phototrophic organisms. Following an exposure length of 8 days, all periphyton treatments showed similar levels of Se accumulation, indicating that much of the apparent uptake of Se(IV) was due to non-biological processes (i.e., surface adsorption). The results of this study will help reduce uncertainty in the prediction of Se dynamics and food-chain transfer in freshwater environments. Further exploration of the ecological consequences of extracellular adsorption of Se(IV) to periphyton, rather than intracellular absorption, is recommended to further refine predictions related to Se biodynamics in freshwater food webs.
Acid mine drainage affects the diversity and metal resistance gene profile of sediment bacterial community along a river
John P. Giesy,
Chemosphere, Volume 217
Acid mine drainage (AMD) is one of the most hazardous byproducts of some types of mining. However, research on how AMD affects the bacterial community structure of downstream riverine ecosystems and the distribution of metal resistance genes (MRGs) along pollution gradient is limited. Comprehensive geochemical and high-throughput next-generation sequencing analyses can be integrated to characterize spatial distributions and MRG profiles of sediment bacteria communities along the AMD-contaminated Hengshi River. We found that (1) diversities of bacterial communities significantly and gradually increased along the river with decreasing contamination, suggesting community composition reflected changes in geochemical conditions; (2) relative abundances of phyla Proteobacteria and genus Halomonas and Planococcaceae that function in metal reduction decreased along the AMD gradient; (3) low levels of sediment salinity, sulfate, aquatic lead (Pb), and cadmium (Cd) were negatively correlated with bacterial diversity despite pH was in a positive manner with diversity; and (4) arsenic (As) and copper (Cu) resistance genes corresponded to sediment concentrations of As and Cu, respectively. Altogether, our findings offer initial insight into the distribution patterns of sediment bacterial community structure, diversity and MRGs along a lotic ecosystem contaminated by AMD, and the factors that affect them.
Rivers are among the most threatened freshwater ecosystems, and anthropogenic activities are affecting both river structures and water quality. While assessing the organisms can provide a comprehensive measure of a river's ecological status, it is limited by the traditional morphotaxonomy-based biomonitoring. Recent advances in environmental DNA (eDNA) metabarcoding allow to identify prokaryotes and eukaryotes in one sequencing run, and could thus allow unprecedented resolution. Whether such eDNA-based data can be used directly to predict the pollution status of rivers as a complementation of environmental data remains unknown. Here we used eDNA metabarcoding to explore the main stressors of rivers along which community structure changes, and to identify the method's potential for predicting pollution status based on eDNA data. We showed that a broad range of taxa in bacterial, protistan, and metazoan communities could be profiled with eDNA. Nutrients were the main driving stressor affecting communities' structure, alpha diversity, and the ecological network. We specifically observed that the relative abundance of indicative OTUs was significantly correlated with nutrient levels. These OTUs data could be used to predict the nutrient status up to 79% accuracy on testing data sets. Thus, our study gives a novel approach to predicting the pollution status of rivers by eDNA data.
Conventional assessment and evaluation of sediment quality are based on laboratory-based ecotoxicological and chemical measurements with lack of concern for ecological relevance. Microbiotas in sediment are responsive to pollutants and can be used as alternative ecological indicators of sediment pollutants; however, the linkage between the microbial ecology and ecotoxicological endpoints in response to sediment contamination has been poorly evaluated. Here, in situ microbiotas from the Three Gorges Reservoir (TGR) area of the Yangtze River were characterized by DNA metabarcoding approaches, and then, changes of in situ microbiotas were compared with the ecotoxicological endpoint, aryl hydrocarbon receptor (AhR) mediated activity, and level of polycyclic aromatic hydrocarbons (PAHs) in sediments. PAHs and organic pollutant mixtures mediating AhR activity had different effects on the structures of microbiotas. Specifically, Shannon indices of protistan communities were negatively correlated with the levels of AhR mediated activity and PAHs. The sediment AhR activity was positively correlated with the relative abundance of prokaryotic Acetobacteraceae, but had a negative correlation with protistan Oxytrichidae. Furthermore, a quantitative classification model was built to predict the level of AhR activity based on the relative abundances of Acetobacteraceae and Oxytrichidae. These results suggested that in situ Protista communities could provide a useful tool for monitoring and assessing ecological stressors. The observed responses of microbial community provided supplementary evidence to support that the AhR-active pollutants, such as PAHs, were the primary stressors of the aquatic community in TGR area.
eDNA-based bioassessment of coastal sediments impacted by an oil spill
Seon Jin Kim,
John P. Giesy,
Un Hyuk Yim,
Won Joon Shim,
Jong Seong Khim
Environmental Pollution, Volume 238
Oil spills offshore can cause long-term ecological effects on coastal marine ecosystems. Despite their important ecological roles in the cycling of energy and nutrients in food webs, effects on bacteria, protists or arthropods are often neglected. Environmental DNA (eDNA) metabarcoding was applied to characterize changes in the structure of micro- and macro-biota communities of surface sediments over a 7-year period since the occurrence of Hebei Spirit oil spill on December 7, 2007. Alterations in diversities and structures of micro- and macro-biota were observed in the contaminated area where concentrations of polycyclic aromatic hydrocarbons were greater. Successions of bacterial, protists and metazoan communities revealed long-term ecological effects of residual oil. Residual oil dominated the largest cluster of the community-environment association network. Presence of bacterial families (Aerococcaceae and Carnobacteriaceae) and the protozoan family (Platyophryidae) might have conferred sensitivity of communities to oil pollution. Hydrocarbon-degrading bacterial families (Anaerolinaceae, Desulfobacteraceae, Helicobacteraceae and Piscirickettsiaceae) and algal family (Araphid pennate) were resistant to adverse effects of spilt oil. The protistan family (Subulatomonas) and arthropod families (Folsomia, Sarcophagidae Opomyzoidea, and Anomura) appeared to be positively associated with residual oil pollution. eDNA metabarcoding can provide a powerful tool for assessing effects of anthropogenic pollution, such as oil spills on sediment communities and its long-term trends in coastal marine environments.