Leveraging wastewater surveillance to detect viral diseases in livestock settings.

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.

The Importance of Fostering and Funding Scientific Research, and its Relevance to Environmental Toxicology and Chemistry.

What do environmental contaminants and climate change have in common with the virus SARS-CoV-2 and the disease COVID-19? We argue that one common element is the wealth of basic and applied scientific research that provides the knowledge and tools essential in developing effective programs for addressing threats to humans and social-ecological systems. Research on various chemicals, including dichlorodiphenyltrichloroethane and per- and polyfluoroalkyl substances, resulted in regulatory action to protect environmental and human health. Moreover, decades of research on coronaviruses, mRNA, and recently SARS-CoV-2 enabled the rapid development of vaccines to fight the COVID-19 pandemic. In the present study, we explore the common elements of basic and applied scientific research breakthroughs that link chemicals, climate change, and SARS-CoV-2/COVID-19 and describe how scientific information was applied for protecting human health and, more broadly, socio-ecological systems. We also offer a cautionary note on the misuse and mistrust of science that is not new in human history, but unfortunately is surging in modern times. Our goal was to illustrate the critical role of scientific research to society, and we argue that research must be intentionally fostered, better funded, and applied appropriately. To that end, we offer evidence that supports the importance of investing in scientific research and, where needed, ways to counter the spread of misinformation and disinformation that undermines legitimate discourse. Environ Toxicol Chem 2023;42:581-593. © 2022 SETAC.

Detection of the Omicron (B.1.1.529) variant of SARS-CoV-2 in aircraft wastewater.

On the 26th of November 2021, the World Health Organization (WHO) designated the newly detected B.1.1.529 lineage of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) the Omicron Variant of Concern (VOC). The genome of the Omicron VOC contains more than 50 mutations, many of which have been associated with increased transmissibility, differing disease severity, and potential to evade immune responses developed for previous VOCs such as Alpha and Delta. In the days since the designation of B.1.1.529 as a VOC, infections with the lineage have been reported in countries around the globe and many countries have implemented travel restrictions and increased border controls in response. We putatively detected the Omicron variant in an aircraft wastewater sample from a flight arriving to Darwin, Australia from Johannesburg, South Africa on the 25th of November 2021 via positive results on the CDC N1, CDC N2, and del(69-70) RT-qPCR assays per guidance from the WHO. The Australian Northern Territory Health Department detected one passenger onboard the flight who was infected with SARS-CoV-2, which was determined to be the Omicron VOC by sequencing of a nasopharyngeal swab sample. Subsequent sequencing of the aircraft wastewater sample using the ARTIC V3 protocol with Nanopore and ATOPlex confirmed the presence of the Omicron variant with a consensus genome that clustered with the B.1.1.529 BA.1 sub-lineage. Our detection and confirmation of a single onboard Omicron infection via aircraft wastewater further bolsters the important role that aircraft wastewater can play as an independent and unintrusive surveillance point for infectious diseases, particularly coronavirus disease 2019.

Wastewater surveillance demonstrates high predictive value for COVID-19 infection on board repatriation flights to Australia.

Controlling importation and transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from overseas travelers is essential for countries, such as Australia, New Zealand, and other island nations, that have adopted a suppression strategy to manage very low community transmission. Wastewater surveillance of SARS-CoV-2 RNA has emerged as a promising tool employed in public health response in many countries globally. This study aimed to establish whether the surveillance of aircraft wastewater can be used to provide an additional layer of information to augment individual clinical testing. Wastewater from 37 long-haul flights chartered to repatriate Australians was tested for the presence of SARS-CoV-2 RNA. Children 5 years or older on these flights tested negative for coronavirus disease 19 (COVID-19) (deep nasal and oropharyngeal reverse-transcription (RT)-PCR swab) 48 h before departure. All passengers underwent mandatory quarantine for 14-day post arrival in Howard Springs, NT, Australia. Wastewater from 24 (64.9 %) of the 37 flights tested positive for SARS-CoV-2 RNA. During the 14 day mandatory quarantine, clinical testing identified 112 cases of COVID-19. Surveillance for SARS-CoV-2 RNA in repatriation flight wastewater using pooled results from three RT-qPCR assays demonstrated a positive predictive value (PPV) of 87.5 %, a negative predictive value (NPV) of 76.9 % and 83.7% accuracy for COVID-19 cases during the post-arrival 14-day quarantine period. The study successfully demonstrates that the surveillance of wastewater from aircraft for SARS-CoV-2 can provide an additional and effective tool for informing the management of returning overseas travelers and for monitoring the importation of SARS CoV-2 and other clinically significant pathogens.

Minimizing errors in RT-PCR detection and quantification of SARS-CoV-2 RNA for wastewater surveillance.

Wastewater surveillance for pathogens using reverse transcription-polymerase chain reaction (RT-PCR) is an effective and resource-efficient tool for gathering community-level public health information, including the incidence of coronavirus disease-19 (COVID-19). Surveillance of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) in wastewater can potentially provide an early warning signal of COVID-19 infections in a community. The capacity of the world's environmental microbiology and virology laboratories for SARS-CoV-2 RNA characterization in wastewater is increasing rapidly. However, there are no standardized protocols or harmonized quality assurance and quality control (QA/QC) procedures for SARS-CoV-2 wastewater surveillance. This paper is a technical review of factors that can cause false-positive and false-negative errors in the surveillance of SARS-CoV-2 RNA in wastewater, culminating in recommended strategies that can be implemented to identify and mitigate some of these errors. Recommendations include stringent QA/QC measures, representative sampling approaches, effective virus concentration and efficient RNA extraction, PCR inhibition assessment, inclusion of sample processing controls, and considerations for RT-PCR assay selection and data interpretation. Clear data interpretation guidelines (e.g., determination of positive and negative samples) are critical, particularly when the incidence of SARS-CoV-2 in wastewater is low. Corrective and confirmatory actions must be in place for inconclusive results or results diverging from current trends (e.g., initial onset or reemergence of COVID-19 in a community). It is also prudent to perform interlaboratory comparisons to ensure results' reliability and interpretability for prospective and retrospective analyses. The strategies that are recommended in this review aim to improve SARS-CoV-2 characterization and detection for wastewater surveillance applications. A silver lining of the COVID-19 pandemic is that the efficacy of wastewater surveillance continues to be demonstrated during this global crisis. In the future, wastewater should also play an important role in the surveillance of a range of other communicable diseases.

SARS-CoV-2 RNA monitoring in wastewater as a potential early warning system for COVID-19 transmission in the community: A temporal case study.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus which causes coronavirus disease (COVID-19), has spread rapidly across the globe infecting millions of people and causing significant health and economic impacts. Authorities are exploring complimentary approaches to monitor this infectious disease at the community level. Wastewater-based epidemiology (WBE) approaches to detect SARS-CoV-2 RNA in municipal wastewater are being implemented worldwide as an environmental surveillance approach to inform health authority decision-making. Owing to the extended excretion of SARS-CoV-2 RNA in stool, WBE can surveil large populated areas with a longer detection window providing unique information on the presence of pre-symptomatic and asymptomatic cases that are unlikely to be screened by clinical testing. Herein, we analysed SARS-CoV-2 RNA in 24-h composite wastewater samples (n = 63) from three wastewater treatment plants (WWTPs) in Brisbane, Queensland, Australia from 24th of February to 1st of May 2020. A total of 21 samples were positive for SARS-CoV-2, ranging from 135 to 11,992 gene copies (GC)/100 mL of wastewater. Detections were made in a Southern Brisbane WWTP in late February 2020, up to three weeks before the first clininal case was reported there. Wastewater samples were generally positive during the period with highest caseload data. The positive SARS-CoV-2 RNA detection in wastewater while there were limited clinical reported cases demonstrates the potential of WBE as an early warning system to identify hotspots and target localised public health responses, such as increased individual testing and the provision of health warnings.

Intraday variability of indicator and pathogenic viruses in 1-h and 24-h composite wastewater samples: Implications for wastewater-based epidemiology.

We monitored the concentration of indicator viruses crAssphage and pepper mild mottle virus (PMMoV) and human pathogen adenovirus (HAdV) in influent from a wastewater treatment plant in Brisbane, Australia in 1-h and 24-h composite samples. Over three days of sampling, the mean concentration of crAssphage gene copies (GC)/mL in 24-h composite samples did not differ significantly (p = 0.72-0.92), while for PMMoV GC/mL (p value range: 0.0002-0.0321) and HAdV GC/mL (p value range: 0.0028-0.0068) significant differences in concentrations were observed on one day of sampling compared to the other two. For all three viruses, the variation observed in 1-h composite samples was greater than the variation observed in 24-h composite samples. For crAssphage, in 54.1% of 1-h composite samples, the concentration was less than that observed in 24-h composite samples; whereas for PMMoV and HAdV the concentration was less in 79.2 and 70.9% of 1-h composite samples, respectively, compared to the relevant 24-h composite samples. Similarly, the concentration of crAssphage in 1-h compared to 24-h composite samples did not differ (p = 0.1082) while the concentrations of PMMoV (p < 0.0001) and HAdV (p < 0.0001) in 1-h composite samples were significantly different from 24-h composite samples. These results suggest that 24-h composite samples offer increased analytical sensitivity and decreased variability compared to 1-h composite samples when monitoring wastewater, especially for pathogenic viruses with low infection rates within a community. Thus, for wastewater-based epidemiology applications, 24-h composite samples are less likely to produce false negative results and erroneous public health information.

Surveillance of SARS-CoV-2 RNA in wastewater: Methods optimisation and quality control are crucial for generating reliable public health information.

Monitoring for SARS-CoV-2 RNA in wastewater through the process of wastewater-based epidemiology (WBE) provides an additional surveillance tool, contributing to community-based screening and prevention efforts as these measurements have preceded disease cases in some instances. Numerous detections of SARS-CoV-2 RNA have been reported globally using various methods, demonstrating the technical feasibility of routine monitoring. However, in order to reliably interpret data produced from these efforts for informing public health interventions, additional quality control information and standardization in sampling design, sample processing, and data interpretation and reporting is needed. This review summarizes published studies of SARS-CoV-2 RNA detection in wastewater as well as available information regarding concentration, extraction, and detection methods. The review highlights areas for potential standardization including considerations related to sampling timing and frequency relative to peak fecal loading times; inclusion of appropriate information on sample volume collected; sample collection points; transport and storage conditions; sample concentration and processing; RNA extraction process and performance; effective volumes; PCR inhibition; process controls throughout sample collection and processing; PCR standard curve performance; and recovery efficiency testing. Researchers are recommended to follow the Minimum Information for Publication of Quantitative Real-Time PCR (MIQE) guidelines. Adhering to these recommendations will enable robust interpretation of wastewater monitoring results and improved inferences regarding the relationship between monitoring results and disease cases.

Comparison of virus concentration methods for the RT-qPCR-based recovery of murine hepatitis virus, a surrogate for SARS-CoV-2 from untreated wastewater.

There is currently a clear benefit for many countries to utilize wastewater-based epidemiology (WBE) as part of ongoing measures to manage the coronavirus disease 2019 (COVID-19) global pandemic. Since most wastewater virus concentration methods were developed and validated for nonenveloped viruses, it is imperative to determine the efficiency of the most commonly used methods for the enveloped severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Municipal wastewater seeded with a human coronavirus (CoV) surrogate, murine hepatitis virus (MHV), was used to test the efficiency of seven wastewater virus concentration methods: (A-C) adsorption-extraction with three different pre-treatment options, (D-E) centrifugal filter device methods with two different devices, (F) polyethylene glycol (PEG 8000) precipitation, and (G) ultracentrifugation. MHV was quantified by reverse-transcription quantitative polymerase chain reaction and the recovery efficiency was calculated for each method. The mean MHV recoveries ranged from 26.7 to 65.7%. The most efficient methods were adsorption-extraction methods with MgCl2 pre-treatment (Method C), and without pre-treatment (Method B). The third most efficient method used the Amicon® Ultra-15 centrifugal filter device (Method D) and its recovery efficiency was not statistically different from the most efficient methods. The methods with the worst recovery efficiency included the adsorption-extraction method with acidification (A), followed by PEG precipitation (F). Our results suggest that absorption-extraction methods with minimal or without pre-treatment can provide suitably rapid, cost-effective and relatively straightforward recovery of enveloped viruses in wastewater. The MHV is a promising process control for SARS-CoV-2 surveillance and can be used as a quality control measure to support community-level epidemic mitigation and risk assessment.

Silver Sulfide Nanoparticles Reduce Nitrous Oxide Emissions by Inhibiting Denitrification in the Earthworm Gut.

The accumulation of Ag2S in agricultural soil via application of Ag-containing sludge potentially affects the functioning of soil microorganisms and earthworms (EWs) due to the strong antimicrobial properties of Ag. This study examined the effects of Ag2S nanoparticles (Ag2S-NPs) on the EW-mediated (Eisenia fetida and Pontoscolex corethrurus) soil N cycle. We used 16S rRNA gene-based sequencing and quantitative polymerase chain reaction to examine the bacterial community and nitrification/denitrification-related gene abundance. The presence of either EWs or Ag significantly increased denitrification and N2O emissions. However, the addition of Ag2S to EW-inhabited soil reduced N2O emissions by 14-33%. Furthermore, Ag2S caused a low-dose stimulation but a high-dose inhibition to N2O flux from the EW gut itself. Accordingly, an increase in Ag in the EW gut caused a decrease in the relative abundance of denitrifiers in both the soil and the gut, especially for the dominant genus Bacillus. Ag2S also decreased the copy numbers of nitrification gene (nxrB) and denitrification genes (napA, nirS, and nosZ) in EW gut, leading to the observed decrease in N2O emissions. Collectively, applying Ag2S-containing sludge disturbs the denitrification function of the EW gut microbiota and the cycling of N in soil-based systems.

Preparing interdisciplinary leadership for a sustainable future.

Urgent sustainability challenges require effective leadership for inter- and trans-disciplinary (ITD) institutions. Based on the diverse experiences of 20 ITD institutional leaders and specific case studies, this article distills key lessons learned from multiple pathways to building successful programs. The lessons reflect both the successes and failures our group has experienced, to suggest how to cultivate appropriate and effective leadership, and generate the resources necessary for leading ITD programs. We present two contrasting pathways toward ITD organizations: one is to establish a new organization and the other is to merge existing organizations. We illustrate how both benefit from a real-world focus, with multiple examples of trajectories of ITD organizations. Our diverse international experiences demonstrate ways to cultivate appropriate leadership qualities and skills, especially the ability to create and foster vision beyond the status quo; collaborative leadership and partnerships; shared culture; communications to multiple audiences; appropriate monitoring and evaluation; and perseverance. We identified five kinds of resources for success: (1) intellectual resources; (2) institutional policies; (3) financial resources; (4) physical infrastructure; and (5) governing boards. We provide illustrations based on our extensive experience in supporting success and learning from failure, and provide a framework that articulates the major facets of leadership in inter- and trans-disciplinary organizations: learning, supporting, sharing, and training.

Decay of SARS-CoV-2 and surrogate murine hepatitis virus RNA in untreated wastewater to inform application in wastewater-based epidemiology.

Wastewater-based epidemiology (WBE) demonstrates potential for COVID-19 community transmission monitoring; however, data on the stability of SARS-CoV-2 RNA in wastewater are needed to interpret WBE results. The decay rates of RNA from SARS-CoV-2 and a potential surrogate, murine hepatitis virus (MHV), were investigated by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) in untreated wastewater, autoclaved wastewater, and dechlorinated tap water stored at 4, 15, 25, and 37 °C. Temperature, followed by matrix type, most greatly influenced SARS-CoV-2 RNA first-order decay rates (k). The average T90 (time required for 1-log10 reduction) of SARS-CoV-2 RNA ranged from 8.04 to 27.8 days in untreated wastewater, 5.71 to 43.2 days in autoclaved wastewater, and 9.40 to 58.6 days in tap water. The average T90 for RNA of MHV at 4 to 37 °C ranged from 7.44 to 56.6 days in untreated wastewater, 5.58-43.1 days in autoclaved wastewater, and 10.9 to 43.9 days in tap water. There was no statistically significant difference between RNA decay of SARS-CoV-2 and MHV; thus, MHV is suggested as a suitable persistence surrogate. Decay rate constants for all temperatures were comparable across all matrices for both viral RNAs, except in untreated wastewater for SARS-CoV-2, which showed less sensitivity to elevated temperatures. Therefore, SARS-CoV-2 RNA is likely to persist long enough in untreated wastewater to permit reliable detection for WBE application.

Detection of SARS-CoV-2 RNA in commercial passenger aircraft and cruise ship wastewater: a surveillance tool for assessing the presence of COVID-19 infected travellers.

BACKGROUND: Wastewater-based epidemiology (WBE) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can be an important source of information for coronavirus disease 2019 (COVID-19) management during and after the pandemic. Currently, governments and transportation industries around the world are developing strategies to minimize SARS-CoV-2 transmission associated with resuming activity. This study investigated the possible use of SARS-CoV-2 RNA wastewater surveillance from airline and cruise ship sanitation systems and its potential use as a COVID-19 public health management tool. METHODS: Aircraft and cruise ship wastewater samples (n = 21) were tested for SARS-CoV-2 using two virus concentration methods, adsorption-extraction by electronegative membrane (n = 13) and ultrafiltration by Amicon (n = 8), and five assays using reverse-transcription quantitative polymerase chain reaction (RT-qPCR) and RT-droplet digital PCR (RT-ddPCR). Representative qPCR amplicons from positive samples were sequenced to confirm assay specificity. RESULTS: SARS-CoV-2 RNA was detected in samples from both aircraft and cruise ship wastewater; however concentrations were near the assay limit of detection. The analysis of multiple replicate samples and use of multiple RT-qPCR and/or RT-ddPCR assays increased detection sensitivity and minimized false-negative results. Representative qPCR amplicons were confirmed for the correct PCR product by sequencing. However, differences in sensitivity were observed among molecular assays and concentration methods. CONCLUSIONS: The study indicates that surveillance of wastewater from large transport vessels with their own sanitation systems has potential as a complementary data source to prioritize clinical testing and contact tracing among disembarking passengers. Importantly, sampling methods and molecular assays must be further optimized to maximize detection sensitivity. The potential for false negatives by both wastewater testing and clinical swab testing suggests that the two strategies could be employed together to maximize the probability of detecting SARS-CoV-2 infections amongst passengers.

Application of sewage sludge containing environmentally-relevant silver sulfide nanoparticles increases emissions of nitrous oxide in saline soils.

Silver (Ag) is released from a range of products and accumulates in agricultural soils as silver sulfide (Ag2S) through the application of Ag-containing biosolids as a soil amendment. Although Ag2S is comparatively stable, its solubility increases with salinity, potentially altering its impacts on microbial communities due to the anti-microbial properties of Ag. In this study, we investigated the impacts of Ag on the microbially mediated N cycle in saline soils by examining the relationship between the (bio)availability of Ag2S and microbial functioning following the application of Ag2S-containing sludge. Synchrotron-based X-ray absorption spectroscopy (XAS) revealed that the Ag2S was stable within the soil, although extractable Ag concentrations increased up to 18-fold in soils with higher salinity. However, the extractable Ag accounted for <0.05% of the total Ag in all soils and had no impact on plant biomass or soil bacterial biomass. Interestingly, at high soil salinity, Ag2S significantly increased cumulative N2O emissions from 80.9 to 229.2 mg kg-1 dry soil (by 180%) compared to the corresponding control sludge treatment, which was ascribed to the increased abundance of nitrification and denitrification-related genes (amoA, nxrB, narG, napA, nirS, and nosZ) and increased relative abundance of denitrifiers (Rhodanobacter, Salinimicrobium, and Zunongwangia). Together, our findings show that the application of Ag2S-containing sludge to a saline soil can disrupt the N cycle and increase N2O emissions from agroecosystems.

Toxicogenomic responses of Caenorhabditis elegans to pristine and transformed zinc oxide nanoparticles.

Manufactured nanoparticles (MNPs) undergo transformation immediately after they enter wastewater treatment streams and during their partitioning to sewage sludge, which is applied to agricultural soils in form of biosolids. We examined toxicogenomic responses of the model nematode Caenorhabditis elegans to pristine and transformed ZnO-MNPs (phosphatized pZnO- and sulfidized sZnO-MNPs). To account for the toxicity due to dissolved Zn, a ZnSO4 treatment was included. Transformation of ZnO-MNPs reduced their toxicity by nearly ten-fold, while there was almost no difference in the toxicity of pristine ZnO-MNPs and ZnSO4. This combined with the fact that far more dissolved Zn was released from ZnO- compared to pZnO- or sZnO-MNPs, suggests that dissolution of pristine ZnO-MNPs is one of the main drivers of their toxicity. Transcriptomic responses at the EC30 for reproduction resulted in a total of 1161 differentially expressed genes. Fifty percent of the genes differentially expressed in the ZnSO4 treatment, including the three metal responsive genes (mtl-1, mtl-2 and numr-1), were shared among all treatments, suggesting that responses to all forms of Zn could be partially attributed to dissolved Zn. However, the toxicity and transcriptomic responses in all MNP treatments cannot be fully explained by dissolved Zn. Two of the biological pathways identified, one essential for protein biosynthesis (Aminoacyl-tRNA biosynthesis) and another associated with detoxification (ABC transporters), were shared among pristine and one or both transformed ZnO-MNPs, but not ZnSO4. When comparing pristine and transformed ZnO-MNPs, 66% and 40% of genes were shared between ZnO-MNPs and sZnO-MNPs or pZnO-MNPs, respectively. This suggests greater similarity in transcriptomic responses between ZnO-MNPs and sZnO-MNPs, while toxicity mechanisms are more distinct for pZnO-MNPs, where 13 unique biological pathways were identified. Based on these pathways, the toxicity of pZnO-MNPs is likely to be associated with their adverse effect on digestion and metabolism.

Nanotoxicity of engineered nanomaterials (ENMs) to environmentally relevant beneficial soil bacteria - a critical review.

Deposition of engineered nanomaterials (ENMs) in various environmental compartments is projected to continue rising exponentially. Terrestrial environments are expected to be the largest repository for environmentally released ENMs. Because ENMs are enriched in biosolids during wastewater treatment, agriculturally applied biosolids facilitate ENM exposure of key soil micro-organisms, such as plant growth-promoting rhizobacteria (PGPR). The ecological ramifications of increasing levels of ENM exposure of terrestrial micro-organisms are not clearly understood, but a growing body of research has investigated the toxicity of ENMs to various soil bacteria using a myriad of toxicity end-points and experimental procedures. This review explores what is known regarding ENM toxicity to important soil bacteria, with a focus on ENMs which are expected to accumulate in terrestrial ecosystems at the highest concentrations and pose the greatest potential threat to soil micro-organisms having potential indirect detrimental effects on plant growth. Knowledge gaps in the fundamental understanding of nanotoxicity to bacteria are identified, including the role of physicochemical properties of ENMs in toxicity responses, particularly in agriculturally relevant micro-organisms. Strategies for improving the impact of future research through the implementation of in-depth ENM characterization and use of necessary experimental controls are proposed. The future of nanotoxicological research employing microbial ecoreceptors is also explored, highlighting the need for continued research utilizing bacterial isolates while concurrently expanding efforts to study ENM-bacteria interactions in more complex environmentally relevant media, e.g. soil. Additionally, the particular importance of future work to extensively examine nanotoxicity in the context of bacterial ecosystem function, especially of plant growth-promoting agents, is proposed.

Colloidal nitrogen is an important and highly-mobile form of nitrogen discharging into the Great Barrier Reef lagoon.

Soil-borne colloids have been linked to long-distance transport of radionuclides, metal(loid)s and nutrients. Colloid-associated nitrogen (N) will have different mechanisms of biogeochemical cycling and potential for water-borne transport over longer distances compared to dissolved N. The role that colloids play in the supply and mobility of N within catchments discharging into the Great Barrier Reef (GBR) lagoon is unexplored. Here, we examine water-dispersible clay (WDC) from soil samples collected from gullies and agricultural drains within three different land uses (sugarcane, non-agricultural land and grazing) within the Townsville area. The proportion of soil N associated with WDC was inversely correlated with total soil N, with up to 45% of the total soil N being colloid-associated in low N gully soils. Within the <0.45 µm fraction of the WDC, only 17-25% of the N was truly dissolved (<3 kDa) at the gully sites compared to 58% in the sugarcane sites. Our results demonstrate the importance of colloidal N and the inaccuracy of assuming N < 0.45 µm is dissolved in the sampled areas, as well as providing an alternate explanation for the large amounts of what has previously been defined as dissolved inorganic N in runoff from non-fertilized grazing land. In particular, they describe why non-fertilized land uses can contribute significant N < 0.45 µm, and why catchment models of nutrient export based on soil N concentrations can over-estimate loads of particulate nitrogen derived from monitoring data (N > 0.45 µm). The findings suggest that managing soil erosion may also contribute to managing N < 0.45 µm.

Silver engineered nanomaterials and ions elicit species-specific O2 consumption responses in plant growth promoting rhizobacteria.

Metal containing engineered nanomaterials (ENMs) are now commonly used in various industrial and commercial applications. Many of these materials can be transformed during waste water treatment and ultimately enter terrestrial ecosystems via agriculturally applied biosolids. It is unclear how agriculturally important soil microbes will be affected by exposure to environmentally relevant, sublethal concentrations of ENMs and their transformation products (i.e., ions, aggregates, etc.). A method was developed, which puts O2 consumption responses in terms of viability, and tested by examining the toxic effects of Ag+, Zn2+, and Ni2+ ions on the plant growth promoting rhizobacterium (PGPR) Bacillus amyloliquefaciens GB03. The method was then used to examine the toxicity of Ag+, as-synthesized polyvinylpyrrolidone-coated silver ENM (PVP-AgENMs), and 100% sulfidized AgENM on B. amyloliquefaciens GB03, and two additional PGPRs Sinorhizobium meliloti 2011, and Pseudomonas putida UW4. S. meliloti was found to have the highest LC50 for Ag+ and PVP-AgENMs (6.6 and 207 µM, respectively), while B. amyloliquefaciens and P. putida exhibited LC50's for Ag+ and PVP-AgENMs roughly half those observed for S. meliloti. The authors observed species-specific O2 consumption responses to ENM and ion exposure. PVP-AgENMs were less toxic than ions on a molar basis, and abiotic dissolution likely explains a significant portion of the observed toxic responses. Our results suggest microbes may exhibit distinct metabolic responses to metal and ENM exposure, even when similar LC50's are observed. These findings together illustrate the importance of understanding species-specific toxic responses and the utility of examining O2 consumption for doing so.

Incorporating Transgenerational Epigenetic Inheritance into Ecological Risk Assessment Frameworks.

Chronic exposure to environmental contaminants can induce heritable "transgenerational" modifications to organisms, potentially affecting future ecosystem health and functionality. Incorporating transgenerational epigenetic heritability into risk assessment procedures has been previously suggested. However, a critical review of existing literature yielded numerous studies claiming transgenerational impacts, with little compelling evidence. Therefore, contaminant-induced epigenetic inheritance may be less common than is reported in the literature. We identified a need for multigeneration epigenetic studies that extend beyond what could be deemed "direct exposure" to F1 and F2 gametes and also include subsequent multiple nonexposed generations to adequately evaluate transgenerational recovery times. Also, increased experimental replication is required to account for the highly variable nature of epigenetic responses and apparent irreproducibility of current studies. Further, epigenetic end points need to be correlated with observable detrimental organism changes before a need for risk management can be properly determined. We suggest that epigenetic-based contaminant studies include concentrations lower than current "EC10-20" or "Lowest Observable Effect Concentrations" for the organism's most sensitive phenotypic end point, as higher concentrations are likely already regulated. Finally, we propose a regulatory framework and optimal experimental design that enables transgenerational epigenetic effects to be assessed and incorporated into conventional ecotoxicological testing.

Silver Nanoparticles Entering Soils via the Wastewater-Sludge-Soil Pathway Pose Low Risk to Plants but Elevated Cl Concentrations Increase Ag Bioavailability.

The widespread use of silver nanoparticles (Ag-NPs) results in their movement into wastewater treatment facilities and subsequently to agricultural soils via application of contaminated sludge. On-route, the chemical properties of Ag may change, and further alterations are possible upon entry to soil. In the present study, we examined the long-term stability and (bio)availability of Ag along the "wastewater-sludge-soil" pathway. Synchrotron-based X-ray absorption spectroscopy (XAS) revealed that ca. 99% of Ag added to the sludge reactors as either Ag-NPs or AgNO3 was retained in sludge, with ≥79% of this being transformed to Ag2S, with the majority (≥87%) remaining in this form even after introduction to soils at various pH values and Cl concentrations for up to 400 days. Diffusive gradients in thin films (DGT), chemical extraction, and plant uptake experiments indicated that the potential (bio)availability of Ag in soil was low but increased markedly in soils with elevated Cl, likely due to the formation of soluble AgClx complexes in the soil solution. Although high Cl concentrations increased the bioavailability of Ag markedly, plant growth was not reduced in any treatment. Our results indicate that Ag-NPs entering soils through the wastewater-sludge-soil pathway pose low risk to plants due to their conversion to Ag2S in the wastewater treatment process, although bioavailability may increase in saline soils or when irrigated with high-Cl water.