Genetically engineered eucalyptus expressing pesticidal proteins from Bacillus thuringiensis for insect resistance: a risk assessment evaluation perspective.

Eucalyptus covers approximately 7.5 million hectares in Brazil and serves as the primary woody species cultivated for commercial purposes. However, native insects and invasive pests pose a significant threat to eucalyptus trees, resulting in substantial economic losses and reduced forest productivity. One of the primary lepidopteran pests affecting eucalyptus is Thyrinteina arnobia (Stoll, 1782) (Lepidoptera: Geometridae), commonly referred to as the brown looper caterpillar. To address this issue, FuturaGene, the biotech division of Suzano S.A., has developed an insect-resistant (IR) eucalyptus variety, which expresses Cry pesticidal proteins (Cry1Ab, Cry1Bb, and Cry2Aa), derived from Bacillus thuringiensis (Bt). Following extensive safety assessments, including field trials across various biomes in Brazil, the Brazilian National Technical Commission of Biosafety (CTNBio) recently approved the commercialization of IR eucalyptus. The biosafety assessments involved the analysis of molecular genomics, digestibility, thermostability, non-target organism exposure, degradability in the field, and effects on soil microbial communities and arthropod communities. In addition, in silico studies were conducted to evaluate allergenicity and toxicity. Results from both laboratory and field studies indicated that Bt eucalyptus is as safe as the conventional eucalyptus clone for humans, animals, and the environment, ensuring the secure use of this insect-resistant trait in wood production.

Corals nitrogen and carbon isotopic signatures alters under Artificial Light at Night (ALAN).

This study examines the impact of Artificial Light at Night (ALAN) on two coral species, Acropora eurystoma and Pocillopora damicornis, in the Gulf of Aqaba/Eilat Red Sea, assessing their natural isotopic responses to highlight changes in energy and nutrient sourcing due to sensory light pollution. Our findings indicate significant disturbances in photosynthetic processes in Acropora eurystoma, as evidenced by shifts in δ13C values under ALAN, pointing to alterations in carbon distribution or utilization. In Pocillopora damicornis, similar trends were observed, with changes in δ13C and δ15N values suggesting a disruption in its nitrogen cycle and feeding strategies. The study also uncovers species-specific variations in heterotrophic feeding, a crucial factor in coral resilience under environmental stress, contributing to the corals' fixed carbon budget. Light measurements across the Gulf demonstrated a gradient of light pollution which possess the potential of affecting marine biology in the region. ALAN was found to disrupt natural diurnal tentacle behaviors in both coral species, crucial for prey capture and nutrient acquisition, thereby impacting their isotopic composition and health. Echoing previous research, our study underscores the need to consider each species' ecological and physiological contexts when assessing the impacts of anthropogenic changes. The findings offer important insights into the complexities of marine ecosystems under environmental stress and highlight the urgency of developing effective mitigation strategies.

Five-years post commercial approval monitoring of eucalyptus H421.

Eucalyptus comprises the largest planted area of cultivated production forest in Brazil. Genetic modification of eucalyptus can provide additional characteristics for increasing productivity, protecting plant yield, and potentially altering fiber for various industrial uses. With this objective, a transgenic eucalyptus variety, event H421, received regulatory approval for commercial release after 6 years of approved risk assessment studies by the Brazilian National Technical Biosafety Commission (CTNBio) in 2015, becoming the first approved genetically modified (GM) eucalyptus in the world. GM event H421 enables increased plant biomass accumulation through overexpression of the Arabidopsis 1,4-ß-endoglucanase Cel1, which remodels the xyloglucan-cellulose matrix of the cell wall during development to promote cell expansion and growth. As required, in that time, by the current normative from CTNBio, a post-commercial release monitoring plan for H421 was submitted, incorporating general surveillance for five consecutive years with the submission of annual reports. The monitoring plan was conducted on fields of H421 progenies, with conventional clones as comparators, cultivated in representative regions where eucalyptus is cultivated in the states of São Paulo, Bahia, and Maranhão, representing Southeast, Northeast, and Northern Brazil. Over the course of the five-year general surveillance monitoring plan for the approved GM eucalyptus H421, no adverse effect that could impact the biosafety of the commercially approved event was identified. Additionally, the GM eucalyptus exhibited behavior highly consistent with that of conventional commercial clones. Therefore, there was no need for an extra risk assessment study of a case-specific monitoring plan. The results show the importance of continuously updating the regulation norms of governmental agencies to align with scientific advances.

Safety Assessment of the CP4 EPSPS and NPTII Proteins in Eucalyptus.

Glyphosate herbicide treatment is essential to sustainable Eucalyptus plantation management in Brazil. Eucalyptus is highly sensitive to glyphosate, and Suzano/FuturaGene has genetically modified eucalyptus to tolerate glyphosate, with the aim of both protecting eucalyptus trees from glyphosate application damage and improving weed management. This study presents the biosafety results of the glyphosate-tolerant eucalyptus event 751K032, which expresses the selection marker neomycin phosphotransferase II (NPTII) enzyme and CP4-EPSPS, a glyphosate-tolerant variant of plant 5-enolpyruvyl-shikimate-3-phosphate synthase enzyme. The transgenic genetically modified (GM) event 751K032 behaved in the plantations like conventional non-transgenic eucalyptus clone, FGN-K, and had no effects on arthropods and soil microorganisms. The engineered NPTII and CP4 EPSPS proteins were heat-labile, readily digestible, and according to the bioinformatics analyses, unlikely to cause an allergenic or toxic reaction in humans or animals. This assessment of the biosafety of the glyphosate-tolerant eucalyptus event 751K032 concludes that it is safe to be used for wood production.

Impact of industrial air pollution on the quality of atmospheric water production.

The atmospheric water generator (AWG) is a commercially available device that produces water from the air in large volumes over short times. This method can be applied in most regions of the world to solve chronic and acute drinking water scarcity. However, knowledge of the effects of air chemical composition on AWG-produced water quality is still very limited. In this study, a comprehensive survey of AWG-produced water quality was conducted in a heavily polluted industrial environment; 83 AWG water samples were analyzed for 99 different quality parameters, including organic, inorganic, and microbial contamination. Two parameters-nickel (15 samples) and dichloromethane (2 samples)-exceeded sporadically their drinking water standards of EPA, EU and IL. Ammonia was the only parameter consistently above standard limits of 0.5 mg/L (61% of samples, relevant to 47 countries) and even higher than 1.5 mg/L. Comparison to real air concentrations of volatile pollutants in the same environment did not reveal any significant correlations; while some pollutants were found at high concentrations in the air, this was not reflected by their presence in the produced water. The findings show that even in areas that are considered excessively polluted relative to the natural environment, the water produced from the air by AWG could be considered suitable for drinking, with careful attention to very specific contaminants.

Pharmaceutical Transformation Products Formed by Ozonation-Does Degradation Occur?

The efficiency of an advanced oxidation process (AOP) using direct and indirect ozonation for the removal of pharmaceutical residues from deliberately spiked deionized water was examined. Both direct and indirect ozonation demonstrated 34% to 100% removal of the parent compounds. However, based on the products' chemical structure and toxicity, we suggest that despite using accepted and affordable ozone and radical concentrations, the six parent compounds were not fully degraded, but merely transformed into 25 new intermediate products. The transformation products (TPs) differed slightly in structure but were mostly similar to their parent compounds in their persistence, stability and toxicity; a few of the TPs were found to be even more toxic than their parent compounds. Therefore, an additional treatment is required to improve and upgrade the traditional AOP toward degradation and removal of both parent compounds and their TPs for safer release into the environment.

Metal peroxides as potential photocatalysts for environmental remediation.

Inorganic oxide materials such as TiO2 and ZnO have been extensively studied for environmental remediation, that operates through photo generated Reactive Oxygen Species (ROS) such as H2O2, ·OH and O2 - to decontaminate waste water. However, inorganic solid oxidants such as metal peroxides capable of generating ROS in aqueous solutions have not been studied for environmental remediation. Towards this objective, we have synthesized peroxides of Zn, Mg, and Ba and characterized these by powder X-ray diffraction, Transmission Electron Microscopy, UV-visible spectroscopy, and X-ray photoelectron spectroscopy. The photocatalytic activity of these wide band gap semiconductors has also been investigated. The novelty of the work is in the use of these peroxides as chemical sources of ROS in aqueous suspensions in addition to their photochemical generation. Hence, these peroxides, in particular Ba, exhibit high photocatalytic activity, better than the well-known ZnO. The mechanisms of ROS generation and subsequent dye degradation are elucidated. ROS has been estimated and is correlated to the photocatalytic activity. This work reports for the first time BaO2 as potential photocatalyst.

Removal of carbamazepine, venlafaxine and iohexol from wastewater effluent using coupled microalgal-bacterial biofilm.

We evaluated the removal capacity of a coupled microalgal-bacterial biofilm (CMBB) to eliminate three recalcitrant pharmaceuticals. The CMBB's efficiency, operating at different biofilm concentrations, with or without light, was compared and analyzed to correlate these parameters to pharmaceutical removal and their effect on the microorganism community. Removal rates changed with changing pharmaceutical and biofilm concentrations: higher biofilm concentrations presented higher removal. Removal of 82-94% venlafaxine and 18-51% carbamazepine was obtained with 5 days of CMBB treatment. No iohexol removal was observed. Light, microorganism composition, and dissolved oxygen concentration are essential parameters governing the removal of pharmaceuticals and ammonia. Chlorophyll concentration increased with time, even in the dark. Three bacterial phyla were dominant: Proteobacteria, Bacteroidetes and Firmicutes. The dominant eukaryotic supergroups were Archaeplastida, Excavata and SAR. A study of the microorganisms' community indicated that not only do the species in the biofilm play an important role; environment, concentration and interactions among them are also important. CMBB has the potential to provide low-cost and sustainable treatment for wastewater and recalcitrant pharmaceutical removal. The microenvironments on the biofilm created by the microalgae and bacteria improved treatment efficiency.

Nano-enabled sensing of per-/poly-fluoroalkyl substances (PFAS) from aqueous systems - A review.

Per-/poly-fluoroalkyl substances (PFAS) are an emerging class of environmental contaminants used as an additive across various commodity and fire-retardant products, for their unique thermo-chemical stability, and to alter their surface properties towards selective liquid repellence. These properties also make PFAS highly persistent and mobile across various environmental compartments, leading to bioaccumulation, and causing acute ecotoxicity at all trophic levels particularly to human populations, thus increasing the need for monitoring at their repositories or usage sites. In this review, current nano-enabled methods towards PFAS sensing and its monitoring in wastewater are critically discussed and benchmarked against conventional detection methods. The discussion correlates the materials' properties to the sensitivity, responsiveness, and reproducibility of the sensing performance for nano-enabled sensors in currently explored electrochemical, spectrophotometric, colorimetric, optical, fluorometric, and biochemical with limits of detection of 1.02 × 10-6 µg/L, 2.8 µg/L, 1 µg/L, 0.13 µg/L, 6.0 × 10-5 µg/L, and 4.141 × 10-7 µg/L respectively. The cost-effectiveness of sensing platforms plays an important role in the on-site analysis success and upscalability of nano-enabled sensors. Environmental monitoring of PFAS is a step closer to PFAS remediation. Electrochemical and biosensing methods have proven to be the most reliable tools for future PFAS sensing endeavors with very promising detection limits in an aqueous matrix, short detection times, and ease of fabrication.

UV-LED Combined with Small Bioreactor Platform (SBP) for Degradation of 17α-Ethynylestradiol (EE2) at Very Short Hydraulic Retention Time.

Degradation of 17α-ethynylestradiol (EE2) and estrogenicity were examined in a novel oxidative bioreactor (OBR) that combines small bioreactor platform (SBP) capsules and UV-LED (ultraviolet light emission diode) simultaneously, using enriched water and secondary effluent. Preliminary experiments examined three UV-LED wavelengths-267, 279, and 286 nm, with (indirect photolysis) and without (direct photolysis) H2O2. The major degradation wavelength for both direct and indirect photolysis was 279 nm, while the major removal gap for direct vs. indirect degradation was at 267 nm. Reduction of EE2 was observed together with reduction of estrogenicity and mineralization, indicating that the EE2 degradation products are not estrogens. Furthermore, slight mineralization occurred with direct photolysis and more significant mineralization with the indirect process. The physical-biological OBR process showed major improvement over other processes studied here, at a very short hydraulic retention time. The OBR can feasibly replace the advanced oxidation process of UV-LED radiation with catalyst in secondary sedimentation tanks with respect to reduction ratio, and with no residual H2O2. Further research into this OBR system is warranted, not only for EE2 degradation, but also to determine its capabilities for degrading mixtures of pharmaceuticals and pesticides, both of which have a significant impact on the environment and public health.

The Endosymbiotic Coral Algae Symbiodiniaceae Are Sensitive to a Sensory Pollutant: Artificial Light at Night, ALAN.

Artificial Light at Night, ALAN, is a major emerging issue in biodiversity conservation, which can negatively impact both terrestrial and marine environments. Therefore, it should be taken into serious consideration in strategic planning for urban development. While the lion's share of research has dealt with terrestrial organisms, only a handful of studies have focused on the marine milieu. To determine if ALAN impacts the coral reef symbiotic algae, that are fundamental for sustainable coral reefs, we conducted a short experiment over a period of one-month by illuminating isolated Symbiodiniaceae cell cultures from the genera Cladocopium (formerly Clade C) and Durusdinium (formerly Clade D) with LED light. Cell cultures were exposed nightly to ALAN levels of 0.15 µmol quanta m-2 s-1 (∼4-5 lux) with three light spectra: blue, yellow and white. Our findings showed that even in very low levels of light at night, the photo-physiology of the algae's Electron Transport Rate (ETR), Non-Photochemical Quenching, (NPQ), total chlorophyll, and meiotic index presented significantly lower values under ALAN, primarily, but not exclusively, in Cladocopium cell cultures. The findings also showed that diverse Symbiodiniaceae types have different photo-physiology and photosynthesis performances under ALAN. We believe that our results sound an alarm for the probable detrimental effects of an increasing sensory pollutant, ALAN, on the eco-physiology of symbiotic corals. The results of this study point to the potential effects of ALAN on other organisms in marine ecosystem such as fish, zooplankton, and phytoplankton in which their biorhythms is entrained by natural light and dark cycles.

Arsenate reducing bacteria isolated from the marine sponge Theonella swinhoei: Bioremediation potential.

Arsenic (As) contamination of freshwater resources constitutes a major environmental issue affecting over 200 million people worldwide. Although the use of microorganisms for the bioremediation of As has been well studied, only very few candidates have been identified to date. Here, we investigated bacteria associated with the Red Sea sponge Theonella swinhoei and their potential to reduce As in a low-salinity liquid medium. This Indo-Pacific common sponge has been shown to hyper-accumulate As, at an average concentration of 8600 mg/g-1 in an environment uncontaminated by arsenic or barium. Four isolated strains of bacteria exhibited arsenic reduction potential by transforming inorganic As in the form of arsenate (iAsV) to arsenite (iAsIII). Two of these isolates were identified as Alteromonas macleodii and Pseudovibrio ascidisceicola, and the other two isolates, both belonging to the same species, were identified as Pseudovibrio denitrificans. The four isolates were then cultured in a low-salinity iAsV-rich medium (5 mM) and As concentration was measured over time using a specifically designed high-performance liquid chromatograph coupled to a mass spectrometer (HPLC-MS). Out of the four isolates, A. macleodii and P. ascidisceicola grew successfully in a low-salinity liquid medium and reduced AsV to AsIII at an average rate of 0.094 and 0.083 mM/h, respectively, thereby demonstrating great potential for the bioremediation of As-contaminated groundwater.

Air-water interactions: The signature of meteorological and air-quality parameters on the chemical characteristics of water produced from the atmosphere.

Atmospheric water is considered an alternative sustainable solution for global water scarcity. We analyzed the effects of meteorological and air-quality parameters on the chemical characteristics of atmospheric water. First, we measured the chemical characteristics of water produced by a unique atmospheric water generator (AWG) apparatus in Tel Aviv, Israel. To examine the complex air-water relationships, we obtained atmospheric data from several sources: adjacent air-quality-monitoring stations, aerosol robotic network (AERONET), aerosol pollution profile using PollyXT lidar, and air back-trajectory simulation (HYSPLIT). We found a strong impact of different pollution sources on the water quality. The integration between HYSPLIT, AERONET and lidar analyses shows that the pathway crossed by the air parcel three days before arrival at the site affected the chemical properties of the produced water. Nearby sea salt aerosols from the Mediterranean were persistently observed in the water (medians: sodium 69 µg/L, chloride ions 120 µg/L), corresponding to lidar identification of a sea-breeze layer (30-50 sr lidar ratio in lower elevation). Seasonal variability in climatic conditions affected the concentration of dust-related elements in the water. During dust-storm events, calcium was the most dominant element (median 900 µg/L). Thus, the chemical characteristics of the water can be considered a "footprint" of both regional, local, and phenological composition of the atmosphere.

Detection of N-nitrosodimethylamine (NDMA) and its formation potential in hospital wastewater.

Hospital wastewaters contain high concentrations of pharmaceutical residues and other chemicals, and may present an important source for NDMA (N-nitrosodimethylamine) and its precursors in the aquatic environment. The present study evaluates the contribution of hospital wastewater to NDMA environmental load and identifies important sources within the hospital itself. For this purpose, wastewaters from five large hospitals in Israel were analyzed, and concentrations of NDMA were found in the range of 20.7-56.7 ng/L, which are similar to NDMA concentrations typically detected in domestic wastewater. The relative contribution of day surgery, oncology, laboratories, and central kitchen (in Sheba hospital) to the daily load of NDMA was calculated as 20.2%, 8.2%, 10%, and 43.2%, respectively. In addition, NDMA concentration in Sheba's mixed wastewater stream, measured throughout a complete working day, was highest at 14:00. This suggests the possible impact of lunchtime on NDMA concentration, and emphasizes the dominant contribution of central kitchen waste. Finally, formation potential of NDMA in the mixed stream was 7300 ng/L, in the upper range of domestic wastewater, but could be decreased by 70% during subsequent aerobic biological wastewater treatment.

Solid phase extraction based on trimethylsilyloxy silica aerogel.

Solid-phase extraction (SPE) based on trimethylsilyloxy-modified silica aerogel was developed for extraction of chemotherapeutic drugs from water. The developed method is easy and affordable, can be performed in separating funnel and does not require a vacuum and SPE manifold. The extraction and recovery of cyclophosphamide (CYP), dexamethasone (DEX), and paclitaxel (TAX) by the aerogel from water were investigated. The factors governing the extraction efficiency such as sample pH, sample volume, volume of eluent and concentration of analytes were studied. The LOD and LOQ of the developed method were calculated and linearity was found in the range of 4-100 µg L-1. The extraction efficiency of the aerogel was compared to that of other SPE cartridges, Oasis HLB, Strata-X-C, C18 and polymeric reversed phase, and the aerogel showed similar or better performance than the other commercial cartridges available on the market. The developed method was also used to extract chemotherapeutic drugs spiked in hospital wastewater.

Coral Gametogenesis Collapse under Artificial Light Pollution.

Artificial light at night (ALAN) can have negative impacts on the health of humans and ecosystems.1-4 Marine organisms, including coral reefs in particular, rely on the natural light cycles of sunlight and moonlight to regulate various physiological, biological, and behavioral processes.5-8 Here, we demonstrate that light pollution caused delayed gametogenesis and unsynchronized gamete release in two coral species, Acropora millepora and Acropora digitifera, from the Indo-Pacific Ocean. Given the urbanization along major coasts, light pollution could thus further threaten coral communities' populations, which are already under severe degradation. A worldwide-modeled light pollution impact assessment is provided, which can help incorporate an important variable in coral reef conservation planning.

Assessing pharmaceutical contamination along the Mediterranean and Red Sea coasts of Israel: Ascidians (Chordata, Ascidiacea) as bioindicators.

Global increase in the use of pharmaceutically-active compounds (PhACs), and their insufficient removal in wastewater treatment plants, have resulted in their continuous release into the marine environment. We investigated the use of the solitary ascidians Herdmania momus, Microcosmus exasperatus, and Styela plicata as bioindicators of three common PhACs in the Israeli coastal waters: Bezafibrate, carbamazepine and diclofenac. Both the Mediterranean and the Red-Sea coasts were found contaminated with PhACs, detected at all 11 sampling sites, with four sites contaminated with all three studied PhACs. Diclofenac was most frequent, present in nine of the 11 sites with concentrations reaching 51.9 ng/g of dry weight sample (dw). Bezafibrate and carbamazepine reached concentrations of 47.8 ng/g dw and 14.3 ng/g dw, respectively. The alarming detection of such high concentrations of PhACs in ascidians along Israel's coasts demonstrates both the extent of PhACs contamination in the region, and the potential of ascidians as bioindicators, and emphasizes the urgent need for additional research into PhAC contamination sources and effects.

Rapid visible-light degradation of EE2 and its estrogenicity in hospital wastewater by crystalline promoted g-C3N4.

Metal-free, chemically activated crystalline graphitic carbon nitride (g-C3N4) nanorods with enhanced visible-light photoactivity demonstrated rapid photodegradation of 17α-ethinylestradiol (EE2) in water and real hospital wastewater. Pure g-C3N4 and another three crystalline promoted g-C3N4 photocatalysts developed by hydrothermal method were characterized by, High-Resolution Transmission Electron Microscopy (HRTEM), X-ray diffraction (XRD), Fourier-Transform Infrared spectroscopy (FTIR), Brunauer-Emmett-Teller (BET), Photoluminescence (PL), Electron spin resonance (ESR), X-ray Photoelectron Spectroscopy (XPS) and Diffuse Reflectance Spectroscopy (DRS). Hydrothermal-based chemical activation did not alter the crystal structure, functional group or surface morphology, but it enhanced the specific surface area of activated g-C3N4 due to intralayer delamination and depolymerization of g-C3N4. Compared to pure g-C3N4, the activated g-C3N4-3 demonstrated efficient degradation of EE2 (<30 min, 3 mg/l) by visible wavelengths of the solar spectrum. This work provides advanced insight into the construction of heterojunction visible-light photocatalysts and production of O2- via reduction of O2 with photogenerated electrons. Proposed and derived mechanism for photodegradation of EE2 by g-C3N4-3 using gas chromatography-mass spectrometry (GCMS). Yeast Estrogen Screen (YES) was performed to evaluate the estrogenicity of treated water samples. Efficient removal of EE2 estrogenic activity (<45 min, 3 mg/l) was achieved using the visible light-activated g-C3N4. Estrogenicity removal rate corresponded well with EE2-degradation rate.

Trace Organic Compound Removal from Wastewater Reverse-Osmosis Concentrate by Advanced Oxidation Processes with UV/O3/H2O2.

Advanced technologies, such as reverse osmosis (RO), allow the reuse of treated wastewater for direct or indirect potable use. However, even highly efficient RO systems produce ~10-15% highly contaminated concentrate as a byproduct. This wastewater RO concentrate (WWROC) is very rich in metal ions, nutrients, and hard-to-degrade trace organic compounds (TOrCs), such as pharmaceuticals, plasticizers, flame retardants, and detergents, which must be treated before disposal. WWROC could be up to 10 times more concentrated than secondary effluent. We examined the efficiency of several advanced oxidation processes (AOPs) on TOrC removal from a two-stage WWROC matrix in a pilot wastewater-treatment facility. WWROC ozonation or UV irradiation, with H2O2 addition, demonstrated efficient removal of TOrCs, varying between 21% and over 99% degradation, and indicating that radical oxidation (by HO·) is the dominant mechanism. However, AOPs are not sufficient to fully treat the WWROC, and thus, additional procedures are required to decrease metal ion and nutrient concentrations. Further biological treatment post-AOP is also highly important, to eliminate the degradable organic molecules obtained from the AOP.

Microbial and Viral Communities and Their Antibiotic Resistance Genes Throughout a Hospital Wastewater Treatment System.

Antibiotic resistance poses a serious threat to global public health, and antibiotic resistance determinants can enter natural aquatic systems through discharge of wastewater effluents. Hospital wastewater in particular is expected to contain high abundances of antibiotic resistance genes (ARGs) compared to municipal wastewater because it contains human enteric bacteria that may include antibiotic-resistant organisms originating from hospital patients, and can also have high concentrations of antibiotics and antimicrobials relative to municipal wastewater. Viruses also play an important role in wastewater treatment systems since they can influence the bacterial community composition through killing bacteria, facilitating transduction of genetic material between organisms, and modifying the chromosomal content of bacteria as prophages. However, little is known about the fate and connections between ARGs, viruses, and their associated bacteria in hospital wastewater systems. To address this knowledge gap, we characterized the composition and persistence of ARGs, dsDNA viruses, and bacteria from influent to effluent in a pilot-scale hospital wastewater treatment system in Israel using shotgun metagenomics. Results showed that ARGs, including genes conferring resistance to antibiotics of high clinical relevance, were detected in all sampling locations throughout the pilot-scale system, with only 16% overall depletion of ARGs per genome equivalent between influent and effluent. The most common classes of ARGs detected throughout the system conferred resistance to aminoglycoside, cephalosporin, macrolide, penam, and tetracycline antibiotics. A greater proportion of total ARGs were associated with plasmid-associated genes in effluent compared to in influent. No strong associations between viral sequences and ARGs were identified in viral metagenomes from the system, suggesting that phage may not be a significant vector for ARG transfer in this system. The majority of viruses in the pilot-scale system belonged to the families Myoviridae, Podoviridae, and Siphoviridae. Gammaproteobacteria was the dominant class of bacteria harboring ARGs and the most common putative viral host in all samples, followed by Bacilli and Betaproteobacteria. In the total bacterial community, the dominant class was Betaproteobacteria for each sample. Overall, we found that a variety of different types of ARGs and viruses were persistent throughout this hospital wastewater treatment system, which can be released to the environment through effluent discharge.