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The co-circulation of mosquito-borne Japanese encephalitis virus (JEV), Murray Valley encephalitis virus (MVEV), and West Nile virus (WNV) has impacted human and animal health in multiple countries worldwide. To facilitate early warnings and surveillance of the presence of these viral infectious agents in the environment, a triplex reverse transcription-quantitative PCR (RT-qPCR) was developed for simultaneous quantification of JEV, MVEV, and WNV in potential hotspots such as piggery and urban wastewater and environmental water samples. The performance of the developed triplex RT-qPCR assay was compared with that of simplex counterparts, all using the same primer and probe sequences. The quantifiable results showed a concordance rate of 93.9%-100% (Cohen's kappa) between the triplex and simplex assays. The mean concentrations of exogenous JEV, MVEV, and WNV using the triplex and simplex RT-qPCR assays were remarkably similar in piggery/urban wastewater and environmental water samples. However, the impacts of the matrix effects (i.e., sample composition and PCR inhibition) of environmental water samples on the accurate quantification of these viruses need to be considered. Taken together, this newly developed triplex RT-qPCR assay of JEV, MVEV, and WNV will allow for a more rapid and cost-efficient sample analysis and data interpretation. The application of the triplex assay for environmental surveillance may be a valuable tool to complement the existing disease and mosquito surveillance approaches used to safeguard the health of both humans and animals.IMPORTANCEThe co-circulation of mosquito-borne Japanese encephalitis virus (JEV), Murray Valley encephalitis virus (MVEV), and West Nile virus (WNV) poses significant threats to human and animal health globally. In this study, a triplex RT-qPCR assay was developed for simultaneous quantification of these viruses in wastewater and environmental water samples. Results demonstrated high concordance and sensitivity of the newly developed triplex RT-qPCR assay compared to simplex assays, indicating its efficacy for environmental surveillance. This cost-effective and rapid assay offers a vital tool for timely monitoring of mosquito-borne viruses in environmental samples, enhancing our ability to mitigate potential outbreaks and safeguard public health.
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Barium (Ba) dissolution and mobilization in groundwater are predominantly controlled by sulfate because of the low solubility of barium sulfate (BaSO4) minerals. Naturally present at low concentrations in groundwater, elevated concentrations of Ba can occur as a result of anthropogenic activities, including use of barite in drill operations, and geogenic sources such as leaching from geological formations. No toxicity data exist for Ba with groundwater organisms (stygofauna) to assess the risk of elevated Ba concentrations. The present study measured Ba toxicity to two stygobiont Cyclopoida species: one collected from Wellington and the other from Somersby, New South Wales, Australia. Toxicity was measured as cyclopoid survival over 2, 4, 7, 14, 21, and 28 days in waters of varying sulfate concentration (<1-100 mg SO4/L). When sulfate was present, dissolved Ba concentrations decreased rapidly in toxicity test solutions forming a BaSO4 precipitate until dissolved sulfate was depleted. Barium in excess of sulfate remained in the dissolved form. The toxicity of Ba to cyclopoids was clearly attributed to dissolved Ba. Precipitated Ba was not toxic to the Wellington cyclopoid species. Toxicity values for dissolved Ba for the Wellington and Somersby cyclopoid species included a (21-day) no-effect concentration of 3.3 mg/L and an effective concentration to cause 5% mortality of 4.8 mg/L (at 21 days). Elevated dissolved Ba concentrations due to anthropogenic and/or biogeochemical processes may pose a risk to groundwater organisms. Further toxicity testing with other stygobiont species is recommended to increase the data available to derive a guideline value for Ba that can be used in contaminant risk assessments for groundwaters. Environ Toxicol Chem 2024;00:1-14. © 2024 The Author(s). Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.
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Diverse enteric pathogens, transmitted through human and animal feces, can cause gastroenteritis. Enteric viruses, such as human Aichi virus, specifically A (AiV-A), are emerging pathogens that cause illnesses even at low doses and are spreading globally. This research developed a reverse transcription quantitative polymerase chain reaction (RT-qPCR) assay targeting the 3CD junction and a reverse transcription colorimetric loop-mediated isothermal amplification (RT-cLAMP) duplex assay targeting junctions 2BC and 3CD of the AiV-A genome for rapid and sensitive detection of AiV-A in metropolitan and regional wastewater samples. The performance of these assays was evaluated using control materials and by analyzing wastewater samples. In serially diluted control materials, RT-qPCR provided quantifiable data (mean 1.51 log10 GC/2⯵L of nucleic acid) down to a dilution of 1â¯×â¯10-5â¯pg/µL. In comparison, the duplex RT-cLAMP assay detected down to 1â¯×â¯10-4â¯pg/µL, indicating that its sensitivity was one order of magnitude less than that of RT-qPCR. Of the 38 wastewater samples from 38 metropolitan and regional wastewater treatment plants (WWTPs) in Queensland, Australia, 21 (55.3â¯%) tested positive by RT-qPCR with concentrations in ranging from 3.60 to 6.23 log10 GC/L. In contrast, only 15 (39.5â¯%) of 38 wastewater samples were positive using the duplex RT-cLAMP assay. The methods demonstrated substantial qualitative agreement (κâ¯=â¯0.730), with a concordance of 86.5â¯%, demonstrating the reliability of RT-cLAMP for detecting AiV-A in wastewater samples. The duplex RT-cLAMP assay, despite demonstrating reduced detection sensitivity, has proven effective and holds promise as a supplementary approach, especially in settings with limited resources where rapid and affordable testing is crucial.
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Surface waters are vulnerable to contamination by human and animal feces, posing risks to human health due to potential exposure to enteric pathogens. This research developed a colorimetric loop-mediated isothermal amplification (cLAMP) assay to detect sewage associated Bacteroides dorei HF183/BacR287 (HF183) marker in wastewater and environmental water samples. The host sensitivity and host specificity of the assay were evaluated, and their performance was compared to the Bacteroides HF183 qPCR assay using control materials (gBlocks), environmental water samples seeded with untreated sewage, and ambient environmental water samples. In serial dilutions of control materials, qPCR produced quantifiable data across all dilutions, while cLAMP detected the marker down to 0.001 pg/µL of control materials, which was two orders of magnitude less sensitive than qPCR. All untreated sewage samples (n = 12) tested positive for HF183 by both the qPCR and cLAMP assays, demonstrating a host sensitivity value of 1.00 (maximum value of 1.00). The host specificity by analysing 70 non-human fecal nucleic acid samples revealed cLAMP's specificity value of 0.81 compared to qPCR's 0.64. When testing sewage-seeded environmental water samples, both methods detected HF183 for the lowest amount of sewage, indicating similar detection sensitivity. The application of cLAMP for tracking sewage pollution in environmental waters showed promising results, with moderate agreement between cLAMP and qPCR (κ = 0.510). However, cLAMP occasionally missed detections compared to qPCR, particularly in low-concentration samples. Overall, the cLAMP HF183 assay demonstrated promising potential as a rapid and sensitive method for detecting sewage pollution, offering a viable alternative to qPCR in certain environmental monitoring scenarios.
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Bacteroides , Aguas del Alcantarillado , Aguas del Alcantarillado/microbiología , Bacteroides/genética , Colorimetría/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , Monitoreo del Ambiente/métodos , Heces/microbiología , Humanos , Contaminación del Agua , Técnicas de Diagnóstico MolecularRESUMEN
This study evaluated ten nucleic acid extraction protocols (EP1 to EP10) for measuring five endogenous antibiotic resistance genes (ARGs) in four aircraft wastewater samples (AWW1 to AWW4). The targeted ARGs, including blaCTX-M, blaNDM-1, ermB, qnrS, and tetA, encompassed highly and minimally abundant ARGs. TetA and ermB were consistently detected across four aircraft wastewater samples using the DNeasy Blood and Tissue Kit and the AllPrep PowerViral DNA/RNA kit. QnrS displayed high detection rates with specific extraction protocols and aliquot volumes. Concentrations of ARGs varied across aircraft wastewater samples, with differing extraction protocols influencing quantitative results. The concentrations of tetA, ermB, and qnrS in AWW1 were distinct, while AWW2 to AWW4 exhibited a broader range for tetA, ermB, qnrS, blaCTX-M, and blaNDM-1. EP1 consistently produced the highest concentrations for several ARGs. Collective data analysis revealed varying ARG concentrations across the ten extraction protocols, suggesting the importance of careful extraction protocol selection in ARG monitoring in aircraft wastewater samples. Based on the results, we suggest that a small sample volume (as low as 0.2 mL) may be sufficient for ARG characterization in aircraft wastewater samples. The findings also emphasize the need for considering toilet paper removal without compromising nucleic acid extraction efficiency. The study highlights promising prospects for aircraft wastewater monitoring of ARGs, calling for further investigation into the import and spread of unique ARGs through transport hubs.
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Aeronaves , Aguas Residuales , Aguas Residuales/microbiología , Genes Bacterianos , Farmacorresistencia Microbiana/genética , Humanos , Ácidos Nucleicos/genética , Ácidos Nucleicos/aislamiento & purificación , Farmacorresistencia Bacteriana/genética , AntibacterianosRESUMEN
This study establishes site-specific risk-based threshold (RBT) concentrations for sewage-associated markers, including Bacteroides HF183 (HF183), Lachnospiraceae Lachno3 (Lachno3), cross-assembly phage (CrAssphage), and pepper mild mottle virus (PMMoV), utilizing quantitative microbial risk assessment (QMRA) for recreational estuarine waters (EW). The QMRA model calculates a RBT concentration corresponding to a selected target illness risk for ingestion of EW contaminated with untreated sewage. RBT concentrations were estimated considering site-specific decay rates and concentrations of markers and reference pathogen (human norovirus; HNoV), aiding in the identification of high-risk days during the swimming season. Results indicated varying RBT concentrations for fresh (Day 0) and aged (Days 1 to 10) sewage contamination scenarios over 10 days. HF183 exhibited the highest RBT concentration (26,600 gene copis (GC)/100 mL) initially but decreased rapidly with aging (2570 to 3120 GC/100 mL on Day 10) depending on the decay rates, while Lachno3 and CrAssphage remained relatively stable. PMMoV, despite lower initial RBT (3920 GC/100 mL), exhibited increased RBT (4700 to 6440 GC/100 mL) with aging due to its slower decay rate compared to HNoV. Sensitivity analysis revealed HNoV concentrations as the most influential parameter. Comparison of marker concentrations in estuarine locations with RBT concentrations showed instances of marker exceedance, suggesting days of potential higher risks. The observed discrepancies between bacterial and viral marker concentrations in EW highlight the need for optimized sample concentration method and simultaneous measurement of multiple markers for enhanced risk predictions. Future research will explore the utility of multiple markers in risk management. Overall, this study contributes to better understanding human health risks in recreational waters, aiding regulators, and water quality managers in effective decision-making for risk prioritization and mitigation strategies.
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Monitoreo del Ambiente , Estuarios , Aguas del Alcantarillado , Medición de Riesgo , Monitoreo del Ambiente/métodos , Microbiología del Agua , Tobamovirus , Natación , Biomarcadores/análisisRESUMEN
Agricultural run-off in Australia's Mackay-Whitsunday region is a major source of nutrient and pesticide pollution to coastal and inshore ecosystems of the Great Barrier Reef. While the effects of run-off are well documented for the region's coral and seagrass habitats, the ecological impacts on estuaries, the direct recipients of run-off, are less known. This is particularly true for fish communities, which are shaped by the physico-chemical properties of coastal waterways that vary greatly in tropical regions. To address this knowledge gap, we used environmental DNA (eDNA) metabarcoding to examine fish assemblages at four locations (three estuaries and a harbour) subjected to varying levels of agricultural run-off during a wet and dry season. Pesticide and nutrient concentrations were markedly elevated during the sampled wet season with the influx of freshwater and agricultural run-off. Fish taxa richness significantly decreased in all three estuaries (F = 164.73, P = <0.001), along with pronounced changes in community composition (F = 46.68, P = 0.001) associated with environmental variables (largely salinity: 27.48% contribution to total variance). In contrast, the nearby Mackay Harbour exhibited a far more stable community structure, with no marked changes in fish assemblages observed between the sampled seasons. Among the four sampled locations, variation in fish community composition was more pronounced within the wet season (F = 2.5, P = 0.001). Notably, variation in the wet season was significantly correlated with agricultural contaminants (phosphorus: 6.25%, pesticides: 5.22%) alongside environmental variables (salinity: 5.61%, DOC: 5.57%). Historically contaminated and relatively unimpacted estuaries each demonstrated distinct fish communities, reflecting their associated catchment use. Our findings emphasise that while seasonal effects play a key role in shaping the community structure of fish in this region, agricultural contaminants are also important contributors in estuarine systems.
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Agricultura , Arrecifes de Coral , ADN Ambiental , Monitoreo del Ambiente , Peces , Salinidad , Contaminantes Químicos del Agua , Animales , Contaminantes Químicos del Agua/análisis , Australia , Plaguicidas , Estuarios , EcosistemaRESUMEN
Wastewater surveillance has evolved into a powerful tool for monitoring public health-relevant analytes. Recent applications in tracking severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection highlight its potential. Beyond humans, it can be extended to livestock settings where there is increasing demand for livestock products, posing risks of disease emergence. Wastewater surveillance may offer non-invasive, cost-effective means to detect potential outbreaks among animals. This approach aligns with the "One Health" paradigm, emphasizing the interconnectedness of animal, human, and ecosystem health. By monitoring viruses in livestock wastewater, early detection, prevention, and control strategies can be employed, safeguarding both animal and human health, economic stability, and international trade. This integrated "One Health" approach enhances collaboration and a comprehensive understanding of disease dynamics, supporting proactive measures in the Anthropocene era where animal and human diseases are on the rise.
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Ganado , Aguas Residuales , Animales , Aguas Residuales/virología , COVID-19/prevención & control , Virosis/veterinaria , Virosis/diagnóstico , SARS-CoV-2 , Humanos , Monitoreo del Ambiente/métodos , Salud ÚnicaRESUMEN
Sediments are an integral component of aquatic systems, linking multiple water uses, functions, and services. Contamination of sediments by chemicals is a worldwide problem, with many jurisdictions trying to prevent future pollution (prospective) and manage existing contamination (retrospective). The present review assesses the implementation of sediment toxicity testing in environmental regulations globally. Currently, the incorporation of sediment toxicity testing in regulations is most common in the European Union (EU), North America, and Australasian regions, with some expansion in Asia and non-EU Europe. Employing sediment toxicity testing in prospective assessments (i.e., before chemicals are allowed on the market) is most advanced and harmonized with pesticides. In the retrospective assessment of environmental risks (i.e., chemicals already contaminating sediments), regulatory sediment toxicity testing practices are applied inconsistently on the global scale. International harmonization of sediment toxicity tests is considered an asset and has been successful through the widespread adoption and deployment of Organisation for Economic Co-operation and Development guidelines. On the other hand, retrospective sediment assessments benefit from incorporating regional species and protocols. Currently used toxicity testing species are diverse, with temperate species being applied most often, whereas test protocols are insufficiently flexible to appropriately address the range of environmental contaminants, including nanomaterials, highly hydrophobic contaminants, and ionized chemicals. The ever-increasing and -changing pressures placed on aquatic resources are a challenge for protection and management efforts, calling for continuous sediment toxicity test method improvement to insure effective use in regulatory frameworks. Future developments should focus on including more subtle and specific toxicity endpoints (e.g., incorporating bioavailability-based in vitro tests) and genomic techniques, extending sediment toxicity testing from single to multispecies approaches, and providing a better link with ecological protection goals. Environ Toxicol Chem 2024;43:1697-1716. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.
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Sedimentos Geológicos , Pruebas de Toxicidad , Contaminantes Químicos del Agua , Sedimentos Geológicos/química , Contaminantes Químicos del Agua/toxicidad , Contaminantes Químicos del Agua/análisis , Monitoreo del Ambiente/métodos , Política Ambiental , AnimalesRESUMEN
The lack of standardized methods and large differences in virus concentration and extraction workflows have hampered Severe Acute Respiratory Syndrome (SARS-CoV-2) wastewater surveillance and data reporting practices. Numerous studies have shown that adsorption-extraction (AE) method holds promise, yet several uncertainties remain regarding the optimal AE workflow. Several procedural components may influence the recovered concentrations of target nucleic acid, including membrane types, homogenization instruments, speed and duration, and lysis buffer. In this study, 42 different AE workflows that varied these components were compared to determine the optimal workflow by quantifying endogenous SARS-CoV-2, human adenovirus 40/41 (HAdV 40/41), and a bacterial marker gene of fecal contamination (Bacteroides HF183). Our findings suggest that the workflow chosen had a significant impact on SARS-CoV-2 concentrations, whereas it had minimal impact on HF183 and no effect on HAdV 40/41 concentrations. When comparing individual components in a workflow, such as membrane type (MF-Millipore™ 0.45 µm MCE vs. Isopore™ 0.40 µm), we found that they had no impact on SARS-CoV-2, HAdV 40/41, and HF183 concentrations. This suggests that at least some consumables and equipment are interchangeable. Buffer PM1 + TRIzol-based workflows yielded higher concentrations of SARS-CoV-2 than other workflows. HF183 concentrations were higher in workflows without chloroform. Similarly, higher homogenization speeds (5000-10,000 rpm) led to increased concentrations of SARS-CoV-2 and HF183 but had no effect on HAdV 40/41. Our findings indicate that minor enhancements to the AE workflow can improve the recovery of viruses and bacteria from the wastewater, leading to improved outcomes from wastewater surveillance efforts.
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Adenovirus Humanos , Ácidos Nucleicos , Aguas Residuales , Humanos , Adsorción , Monitoreo Epidemiológico Basado en Aguas Residuales , Flujo de Trabajo , SARS-CoV-2RESUMEN
The Fundão mine tailings dam rupture of 2015, in the Rio Doce basin, Brazil, resulted in the deposition of tailings downstream of the dam. It has yet to be determined if metals associated with the tailings have contributed toxicity to organisms, burying a time bomb that could be ticking. Currently the data on toxicity to benthic and aquatic organisms have not been assessed sufficiently to allow an informed assessment using an approach based on weight-of-evidence. This study was conducted to ascertain if sediments at "hot spots" that received Fundão tailings reflected elevated concentrations of metals and if these concentrations were sufficient to result in toxicity to freshwater organisms. The lines-of-evidence considered included assessing metals concentrations in relation to sediment quality criteria, establishing biogeochemical characterizations, completing an evaluation of potential metal release upon resuspension to provide information on bioavailability, and identifying acute and chronic toxicity effects using sensitive native species for waters (water flea, Daphnia similis) and sediments (burrowing midge larvae, Chironomus sancticaroli). Only porewater concentrations of iron and manganese exceeded Brazilian surface water criteria, whereas most trace elements exhibited no enrichment or elevated environmental indexes. The concentrations of bioavailable metals were assessed to be low, and metal concentrations did not increase in the overlying water upon resuspension; rather, they decreased through time. Toxicity testing in resuspended waters and bulk sediments resulted in no acute or chronic toxicity to either benthic or aquatic species. The low metal bioavailability and absence of toxicity of the tailings-enriched sediments was attributed to the strong binding and rapid removal of potentially toxic metal ions caused by oxyhydroxides and particles in the presence of iron-rich particulates. The findings of these sediment hot-spot studies indicate the Fundão dam release of tailings more than six years ago is not causing the current release of toxic concentrations of metals into the freshwaters of the Rio Doce. Integr Environ Assess Manag 2024;20:148-158. © 2023 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).
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Oligoelementos , Contaminantes Químicos del Agua , Monitoreo del Ambiente , Contaminantes Químicos del Agua/toxicidad , Contaminantes Químicos del Agua/análisis , Metales/toxicidad , Metales/análisis , Hierro , BrasilRESUMEN
Quantifying metal bioaccumulation in a sedimentary environment is a valuable line of evidence when evaluating the ecological risks associated with metal-contaminated sediments. However, the precision of bioaccumulation predictions has been hindered by the challenges in accurately modeling metal influx processes. This study focuses on nickel bioaccumulation from sediment and introduces an innovative approach using the isotopically modified bioassay to directly measure nickel assimilation rates in sediment. Tested in sediments spiked with two distinct nickel concentrations, the measured Ni assimilation rates ranged from 35 to 78 ng g-1 h-1 in the Low-Ni treatment and from 96 to 320 ng g-1 h-1 in the High-Ni treatment. Integrating these rates into a biodynamic model yielded predictions of nickel bioaccumulation closely matching the measured results, demonstrating high accuracy with predictions within a factor of 3 for the Low-Ni treatment and within a factor of 1 for the High-Ni treatment. By eliminating the need to model metal uptake from various sources, this streamlined approach provides a reliable method for predicting nickel bioaccumulation in contaminated sediments. This advancement holds promise for linking bioaccumulation with metal toxicity risks in sedimentary environments, enhancing our understanding of metal-contaminated sediment risks and providing valuable insights to support informed decision-making in ecological risk assessment and management.
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Níquel , Contaminantes Químicos del Agua , Sedimentos Geológicos , Bioacumulación , Metales , Bioensayo , Contaminantes Químicos del Agua/análisisRESUMEN
For the purpose of sediment quality assessment, the prediction of toxicity risk-levels for aquatic organisms based on simple environmental measurements is desirable. One commonly used approach is the comparison of total contaminant concentrations with corresponding water and sediment quality guideline values, serving as a Line of Evidence (LoE) based on chemistry-toxicity effects relationships. However, the accuracy of toxicity predictions can be improved by considering the factors that modify contaminant bioavailability. In this study we used paired chemistry-ecotoxicity data sets for sediments to evaluate the improvement in toxicity risk predictions using bioavailability-modified guidelines. The sediments were predominantly contaminated with metals, and measurements of sediment particle size, total organic carbon (TOC) and acid volatile sulfide (AVS) were used to modify hazard quotients (HQ). To further assess the predictive efficacy of the bioavailability-modified guideline models, sediments with differing contamination levels were tested for toxicity to a benthic amphipod's reproduction. To account for differences between laboratory exposure and field exposure scenarios, where the latter creates greater dilution, both static-renewal and flow-through test procedures were employed, and flow-through resulted in lower dissolved metal concentrations in the overlying waters. We also investigated how lower AVS concentration by oxidation modified the toxicity. This study reaffirmed that consideration of factors that influence contaminant bioavailability improves toxicity risk predictions, however the improvements may be modest. The sediment particle size data had the greatest influence on the modified HQ, indicating that higher percentage of fine particle size (<63 µm) contributed most to a lower predicted toxicity. The comparison of the static-renewal and flow-through test results continue to raise important questions about the relevance of static or static-renewal toxicity test results for risk assessment decisions, as both these test designs may cause unrealistically high contributions of dissolved metals in overlying waters to toxicity. Overall, this study underscores the value of incorporating outcomes from simple and routine sediment analysis (e.g., particle size, TOC, and consideration of AVS) to enhance the predictive efficacy of toxicity risk assessments in the context of sediment quality risk assessment.
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Anfípodos , Contaminantes Químicos del Agua , Animales , Sedimentos Geológicos/análisis , Contaminantes Químicos del Agua/toxicidad , Contaminantes Químicos del Agua/análisis , Metales/toxicidad , Metales/análisis , Agua/análisis , Sulfuros/análisisRESUMEN
Quantitative polymerase chain reaction (qPCR) measurement of antibiotic resistance genes (ARGs) in untreated municipal wastewater may prove useful in combating the antimicrobial resistance crisis. However, harmonizing and optimizing qPCR-based workflows is essential to facilitate comparisons across studies, and includes achieving highly-effective ARG capture through efficient concentration and extraction procedures. In the current study, combinations of sample volume, membrane types and DNA extraction kits within filtration and centrifugation-based workflows were used to quantify 16S ribosomal RNA (16S rRNA), class 1 integron-integrase gene (intI1) and an ARG encoding resistance to vancomycin (vanA) in untreated wastewater sampled from three wastewater treatment plants (WWTPs). Highly abundant 16S rRNA and intI1 were detected in 100 % of samples from all three WWTPs using both 2 and 20 mL sample volumes, while lower prevalence vanA was only detected when using the 20 mL volume. When filtering 2 mL of wastewater, workflows with 0.20-/0.40-µm polycarbonate (PC) membranes generally yielded greater concentrations of the three targets than workflows with 0.22-/0.45-µm mixed cellulose ester (MCE) membranes. The improved performance was diminished when the sample volume was increased to 20 mL. Consistently greater concentrations of 16S rRNA, intI1 and vanA were yielded by filtration-based workflows using PC membranes combined with a DNeasy PowerWater (DPW) Kit, regardless of the sample volume used, and centrifugation-based workflows with DNeasy Blood & Tissue Kit for 2-mL wastewater extractions. Within the filtration-based workflows, the DPW kit yielded more detection and quantifiable results for less abundant vanA than the DNeasy PowerSoil Pro Kit and FastDNA™ SPIN Kit for Soil. These findings indicate that the performance of qPCR-based workflows for surveillance of ARGs in wastewater varies across targets, sample volumes, concentration methods and extraction kits. Workflows must be carefully considered and validated considering the target ARGs to be monitored.
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As oil and gas infrastructure comes to the end of its working life, a decommissioning decision must be made: should the infrastructure be abandoned in situ, repurposed, partially removed, or fully removed? Environmental contaminants around oil and gas infrastructure could influence these decisions because contaminants in sediments could degrade the value of the infrastructure as habitat, enter the seafood supply if the area is re-opened for commercial and/or recreational fishing, or be made biologically available as sediment is resuspended when the structures are moved. An initial risk hypothesis, however, may postulate that these concerns are only relevant if contaminant concentrations are above screening values that predict the possibility of environmental harm or contaminant bioaccumulation. To determine whether a substantive contaminants-based risk assessment is needed for infrastructure in the Gippsland Basin (South-eastern Australia), we measured the concentration of metals and polycyclic aromatic hydrocarbons (PAHs) in benthic sediments collected around eight platforms earmarked for decommissioning. The measurements were compared to preset screening values and to background contaminant concentrations in reference sites. Lead (Pb), zinc (Zn), PAHs and other contaminants were occasionally measured at concentrations that exceeded reference values, most often within 150 m of the platforms. The exceedance of a few screening values by contaminants at some platforms indicates that these platforms require further analysis to determine the contaminant risks associated with any decommissioning option.
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Petróleo , Hidrocarburos Policíclicos Aromáticos , Contaminantes Químicos del Agua , Petróleo/análisis , Monitoreo del Ambiente , Sedimentos Geológicos/química , Metales/análisis , Australia , Hidrocarburos Policíclicos Aromáticos/análisis , Contaminantes Químicos del Agua/análisisRESUMEN
The effective detection of viruses in aircraft wastewater is crucial to establish surveillance programs for monitoring virus spread via aircraft passengers. This study aimed to compare the performance of two virus concentration workflows, adsorption-extraction (AE) and Nanotrap® Microbiome A Particles (NMAP), in detecting the prevalence and concentrations of 15 endogenous viruses including ssDNA, dsDNA, ssRNA in 24 aircraft lavatory wastewater samples. The viruses tested included two indicator viruses, four enteric viruses, and nine respiratory viruses. The results showed that cross-assembly phage (crAssphage), human polyomavirus (HPyV), rhinovirus A (RhV A), and rhinovirus B (RhV B) were detected in all wastewater samples using both workflows. However, enterovirus (EV), human norovirus GII (HNoV GII), human adenovirus (HAdV), bocavirus (BoV), parechovirus (PeV), epstein-barr virus (EBV). Influenza A virus (IAV), and respiratory syncytial virus B (RsV B) were infrequently detected by both workflows, and hepatitis A virus (HAV), influenza B virus (IBV), and respiratory syncytial virus B (RsV A) were not detected in any samples. The NMAP workflow had greater detection rates of RNA viruses (EV, PeV, and RsV B) than the AE workflow, while the AE workflow had greater detection rates of DNA viruses (HAdV, BoV, and EBV) than the NMAP workflow. The concentration of each virus was also analyzed, and the results showed that crAssphage had the highest mean concentration (6.76 log10 GC/12.5 mL) followed by HPyV (5.46 log10 GC/12.5 mL using the AE workflow, while the mean concentrations of enteric and respiratory viruses ranged from 2.48 to 3.63 log10 GC/12.5 mL. Using the NMAP workflow, the mean concentration of crAssphage was 5.18 log10 GC/12.5 mL and the mean concentration of HPyV was 4.20 log10 GC/12.5 mL, while mean concentrations of enteric and respiratory viruses ranged from 2.55 to 3.74 log10 GC/12.5 mL. Significantly higher (p < 0.05) mean concentrations of crAssphage and HPyV were observed when employing the AE workflow in comparison to the NMAP workflow. Conversely, the NMAP workflow yielded significantly greater (p < 0.05) concentrations of RhV A, and RhV B compared to the AE workflow. The findings of this study can aid in the selection of an appropriate concentration workflow for virus surveillance studies and contribute to the development of efficient virus detection methods.
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Adenovirus Humanos , Bacteriófagos , Infecciones por Virus de Epstein-Barr , Microbiota , Poliomavirus , Humanos , Aguas Residuales , Flujo de Trabajo , Adsorción , Cuartos de Baño , Herpesvirus Humano 4RESUMEN
The ongoing COVID-19 pandemic has emphasized the significance of wastewater surveillance in monitoring and tracking the spread of infectious diseases, including SARS-CoV-2. The wastewater surveillance approach detects genetic fragments from viruses in wastewater, which could provide an early warning of outbreaks in communities. In this study, we determined the concentrations of four types of endogenous viruses, including non-enveloped DNA (crAssphage and human adenovirus 40/41), non-enveloped RNA (enterovirus), and enveloped RNA (SARS-CoV-2) viruses, from wastewater samples using the adsorption-extraction (AE) method with electronegative HA membranes of different pore sizes (0.22, 0.45, and 0.80 µm). Our findings showed that the membrane with a pore size of 0.80 µm performed comparably to the membrane with a pore size of 0.45 µm for virus detection/quantitation (repeated measurement one-way ANOVA; p > 0.05). We also determined the recovery efficiencies of indigenous crAssphage and pepper mild mottle virus, which showed recovery efficiencies ranging from 50% to 94% and from 20% to 62%, respectively. Our results suggest that the use of larger pore size membranes may be beneficial for processing larger sample volumes, particularly for environmental waters containing low concentrations of viruses. This study offers valuable insights into the application of the AE method for virus recovery from wastewater, which is essential for monitoring and tracking infectious diseases in communities.
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COVID-19 , Virus , Humanos , Aguas Residuales , SARS-CoV-2/genética , Pandemias , Adsorción , Monitoreo Epidemiológico Basado en Aguas Residuales , ARN , ARN ViralRESUMEN
In this study, two virus concentration methods, namely Adsorption-Extraction (AE) and Nanotrap® Magnetic Virus Particles (NMVP) along with commercially available extraction kits were used to quantify endogenous pepper mild mottle virus (PMMoV) and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in nucleic acid extracted from 48 wastewater samples collected over six events from eight wastewater treatment plants (WWTPs). The main aim was to determine which workflow (i.e., concentration and extraction methods) produces greater concentrations of endogenous PMMoV and SARS-CoV-2 gene copies (GC) in comparison with each other. Turbidity and total suspended solids (TSS) of wastewater samples within and among the eight WWTPs were highly variable (41-385 NTU and 77-668 mg/L TSS). In 58 % of individual wastewater samples, the log10 GC concentrations of PMMoV were greater by NMVP workflow compared to AE workflow. Paired measurements of PMMoV GC/10 mL from AE and NMVP across all 48 wastewater samples were weakly correlated (r = 0.455, p = 0.001) and demonstrated a poor linear relationship (r2 = 0.207). The log10 GC concentrations of SARS-CoV-2 in 69 % of individual samples were greater by AE workflow compared to NMVP workflow. In contrast to PMMoV, the AE and NMVP derived SARS-CoV-2 GC counts were strongly correlated (r = 0.859, p < 0.001) and demonstrated a strong linear relationship (r2 = 0.738). In general, the PMMoV GC achieved by the NMVP workflow decreased with increasing turbidity, but the PMMoV GC by the AE workflow did not appear to be as sensitive to either turbidity or TSS levels. These findings suggest that wastewater sample turbidity or suspended solids concentration, and the intended target for analysis should be considered when validating an optimal workflow for wastewater surveillance of viruses.
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COVID-19 , Virus , Humanos , Aguas Residuales , SARS-CoV-2 , Heces , Monitoreo Epidemiológico Basado en Aguas Residuales , Virión , Fenómenos MagnéticosRESUMEN
Novel combinations of climatic conditions due to climate change and prolonged fire seasons have contributed to an increased occurrence of "megafires". Such large-scale fires pose an unknown threat to biodiversity due to the increased extent and severity of burn. Assessments of wildfires often focus on terrestrial ecosystems and effects on aquatic habitats are less documented, particularly in coastal environments. In a novel application of eDNA techniques, we assessed the impacts of the 2019-2020 Australian wildfires on the diversity of estuarine benthic sediment communities in six estuaries in NSW, Australia, before and after the fires. Estuaries differed in area of catchment burnt (0-92%) and amount of vegetative buffer that remained post-fire between burnt areas and waterways. We found greater dissimilarities in the composition and abundance of eukaryotic and bacterial sediment communities in estuaries from burnt catchments with no buffer compared to those with an intact buffer or from unburnt catchments. Shifts in composition in highly burnt catchments were associated with increased concentrations of nutrients, carbon, including fire-derived pyrogenic carbon, and copper, which was representative of multiple highly correlated trace metals. Changes in the relative abundances of certain taxonomic groups, such as sulfate-reducing and nitrifying bacterial groups, in the most impacted estuaries indicate potential consequences for the functioning of sediment communities. These results provide a unique demonstration of the use of eDNA to identify wildfire impacts on ecological communities and emphasize the importance of vegetative buffers in limiting wildfire-associated impacts.
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Incendios , Incendios Forestales , Ecosistema , Australia , Carbono , BosquesRESUMEN
The early warning and tracking of COVID-19 prevalence in the community provided by wastewater surveillance has highlighted its potential for much broader viral disease surveillance. In this proof-of-concept study, 46 wastewater samples from four wastewater treatment plants (WWTPs) in Queensland, Australia, were analyzed for the presence and abundance of 13 respiratory viruses, and the results were compared with reported clinical cases. The viruses were concentrated using the adsorption-extraction (AE) method, and extracted nucleic acids were analyzed using qPCR and RT-qPCR. Among the viruses tested, bocavirus (BoV), parechovirus (PeV), rhinovirus A (RhV A) and rhinovirus B (RhV B) were detected in all wastewater samples. All the tested viruses except influenza B virus (IBV) were detected in wastewater sample from at least one WWTP. BoV was detected with the greatest concentration (4.96-7.22 log10 GC/L), followed by Epstein-Barr virus (EBV) (4.08-6.46 log10 GC/L), RhV A (3.95-5.63 log10 GC/L), RhV B (3.74-5.61 log10 GC/L), and PeV (3.17-5.32 log10 GC/L). Influenza viruses and respiratory syncytial virus (RSV) are notifiable conditions in Queensland, allowing the gene copy (GC) concentrations to be compared with reported clinical cases. Significant correlations (ρ = 0.60, p < 0.01 for IAV and ρ = 0.53, p < 0.01 for RSV) were observed when pooled wastewater influenza A virus (IAV) and RSV log10 GC/L concentrations were compared to log10 clinical cases among the four WWTP catchments. The positive predictive value for the presence of IAV and RSV in wastewater was 97 % for both IAV and RSV clinical cases within the four WWTP catchments. The overall accuracy of wastewater analysis for predicting clinical cases of IAV and RSV was 97 and 90 %, respectively. This paper lends credibility to the application of wastewater surveillance to monitor respiratory viruses of various genomic characteristics, with potential uses for increased surveillance capabilities and as a tool in understanding the dynamics of disease circulation in the communities.