Genetic traits and transmission of antimicrobial resistance characteristics of cephalosporin resistant Escherichia coli in tropical aquatic environments.

This study investigates the genetic traits and transmission mechanisms of cephalosporin-resistant Escherichia coli in tropical aquatic environments in Singapore. From 2016 to 2020, monthly samples were collected from wastewater treatment plants, marine niches, community sewage, beaches, reservoirs, aquaculture farms, and hospitals, yielding 557 isolates that were analyzed for antimicrobial resistance genes (ARGs) and mobile genetic elements (MGEs) using genomic methods. Findings reveal significant genotypic similarities between environmental and hospital-derived strains, particularly the pandemic E. coli ST131. Environmental strains exhibited high levels of intrinsic resistance mechanisms, including mutations in porins and efflux pumps, with key ARGs such as CMY-2 and NDM-9 predominantly carried by MGEs, which facilitate horizontal gene transfer. Notably, pathogenic EPEC and EHEC strains were detected in community sewage and aquaculture farms, posing substantial public health risks. This underscores the critical role of these environments as reservoirs for multidrug-resistant pathogens and emphasizes the interconnectedness of human activities and environmental health.

A multi-pronged approach to assessing antimicrobial resistance risks in coastal waters and aquaculture systems.

Antimicrobial resistance (AMR) is a global challenge that has impacted aquaculture and surrounding marine environments. In this study, a year-long monitoring program was implemented to evaluate AMR in two different aquaculture settings (i.e., open cage farming, recirculating aquaculture system (RAS)) and surrounding marine environment within a tropical coastal region. The objectives of this study are to (i) investigate the prevalence and co-occurrence of antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARGs), antibiotics (AB) and various associated chemical compounds at these study sites; (ii) explore the contributing factors to development and propagation of AMR in the coastal environment; and (iii) assess the AMR risks from different perspectives based on the three AMR determinants (i.e., ARB, ARGs and AB). Key findings revealed a distinct pattern of AMR across the different aquaculture settings, notably a higher prevalence of antibiotic-resistant Vibrio at RAS outfalls, suggesting a potential accumulation of microorganisms within the treatment system. Despite the relative uniform distribution of ARGs across marine sites, specific genes such as qepA, blaCTX-M and bacA, were found to be abundant in fish samples, especially from the RAS. Variations in chemical contaminant prevalence across sites highlighted possible anthropogenic impacts. Moreover, environmental and seasonal variations were found to significantly influence the distribution of ARGs and chemical compounds in the coastal waters. Hierarchical cluster analysis that was based on ARGs, chemical compounds and environmental data, categorized the sites into three distinct clusters which reflected strong association with location, seasonality and aquaculture activities. The observed weak correlations between ARGs and chemical compounds imply that low environmental concentrations may be insufficient for resistance selection. A comprehensive risk assessment using methodologies such as the multiple antibiotic resistance (MAR) index, comparative AMR risk index (CAMRI) and Risk quotient (RQ) underscored the complexity of AMR risks. This research significantly contributes to the understanding of AMR dynamics in natural aquatic systems and provides valuable insights for managing and mitigating AMR risks in coastal environments.

Characterizing PFASs in aquatic ecosystems with 3D hydrodynamic and water quality models.

Understanding how per- and polyfluoroalkyl substances (PFASs) enter aquatic ecosystems is challenging due to the complex interplay of physical, chemical, and biological processes, as well as the influence of hydraulic and hydrological factors and pollution sources at the catchment scale. The spatiotemporal dynamics of PFASs across various media remain largely unknown. Here we show the fate and transport mechanisms of PFASs by integrating monitoring data from an estuarine reservoir in Singapore into a detailed 3D model. This model incorporates hydrological, hydrodynamic, and water quality processes to quantify the distributions of total PFASs, including the major components perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS), across water, particulate matter, and sediments within the reservoir. Our results, validated against four years of field measurements with most relative average deviations below 40%, demonstrate that this integrated approach effectively characterizes the occurrence, sources, sinks, and trends of PFASs. The majority of PFASs are found in the dissolved phase (>95%), followed by fractions sorbed to organic particles like detritus (1.0-3.5%) and phytoplankton (1-2%). We also assess the potential risks in both the water column and sediments of the reservoir. The risk quotients for PFOS and PFOA are <0.32 and < 0.00016, respectively, indicating an acceptable risk level for PFASs in this water body. The reservoir also exhibits substantial buffering capacity, even with a tenfold increase in external loading, particularly in managing the risks associated with PFOA compared to PFOS. This study not only enhances our understanding of the mechanisms influencing the fate and transport of surfactant contaminants but also establishes a framework for future research to explore how dominant environmental factors and processes can mitigate emerging contaminants in aquatic ecosystems.

Screening of the PA14NR Transposon Mutant Library Identifies Genes Involved in Resistance to Bacteriophage Infection in Pseudomomas aeruginosa.

Multidrug-resistant P. aeruginosa infections pose a serious public health threat due to the rise in antimicrobial resistance. Phage therapy has emerged as a promising alternative. However, P. aeruginosa has evolved various mechanisms to thwart phage attacks, making it crucial to decipher these resistance mechanisms to develop effective therapeutic strategies. In this study, we conducted a forward-genetic screen of the P. aeruginosa PA14 non-redundant transposon library (PA14NR) to identify dominant-negative mutants displaying phage-resistant phenotypes. Our screening process revealed 78 mutants capable of thriving in the presence of phages, with 23 of them carrying insertions in genes associated with membrane composition. Six mutants exhibited total resistance to phage infection. Transposon insertions were found in genes known to be linked to phage-resistance such as galU and a glycosyl transferase gene, as well as novel genes such as mexB, lasB, and two hypothetical proteins. Functional experiments demonstrated that these genes played pivotal roles in phage adsorption and biofilm formation, indicating that altering the bacterial membrane composition commonly leads to phage resistance in P. aeruginosa. Importantly, these mutants displayed phenotypic trade-offs, as their resistance to phages inversely affected antibiotic resistance and hindered biofilm formation, shedding light on the complex interplay between phage susceptibility and bacterial fitness. This study highlights the potential of transposon mutant libraries and forward-genetic screens in identifying key genes involved in phage-host interactions and resistance mechanisms. These findings support the development of innovative strategies for combating antibiotic-resistant pathogens.

A rapid approach with machine learning for quantifying the relative burden of antimicrobial resistance in natural aquatic environments.

The massive use and discharge of antibiotics have led to increasing concerns about antimicrobial resistance (AMR) in natural aquatic environments. Since the dose-response mechanisms of pathogens with AMR have not yet been fully understood, and the antibiotic resistance genes and bacteria-related data collection via field sampling and laboratory testing is time-consuming and expensive, designing a rapid approach to quantify the burden of AMR in the natural aquatic environment has become a challenge. To cope with such a challenge, a new approach involving an integrated machine-learning framework was developed by investigating the associations between the relative burden of AMR and easily accessible variables (i.e., relevant environmental variables and adjacent land-use patterns). The results, based on a real-world case analysis, demonstrate that the quantification speed has been reduced from 3-7 days, which is typical for traditional measurement procedures with field sampling and laboratory testing, to approximately 0.5 hours using the new approach. Moreover, all five metrics for AMR relative burden quantification exceed the threshold level of 85%, with F1-score surpassing 0.92. Compared to logistic regression, decision trees, and basic random forest, the adaptive random forest model within the framework significantly improves quantification accuracy without sacrificing model interpretability. Two environmental variables, dissolved oxygen and resistivity, along with the proportion of green areas were identified as three key feature variables for the rapid quantification. This study contributes to the enrichment of burden analyses and management practices for rapid quantification of the relative burden of AMR without dose-response information.

Picocyanobacterial-bacterial interactions sustain cyanobacterial blooms in nutrient-limited aquatic environments.

Cyanobacterial blooms (CBs) and concomitant water quality issues in oligotrophic/mesotrophic waters have been recently reported, challenging the conventional understanding that CBs are primarily caused by eutrophication. To elucidate the underlying mechanism of CBs in nutrition-deficient waters, the changes in Chlorophyll a (Chl-a), cyanobacterial-bacterial community composition, and certain microbial function in Qingcaosha Reservoir, the global largest tidal estuary storage reservoir, were analyzed systematically and comprehensively after its pilot run (2011-2019) in this study. Although the water quality was improved and stabilized, more frequent occurrences of bloom level of Chl-a (>20 µg L-1) in warm seasons were observed during recent years. The meteorological changes (CO2, sunshine duration, radiation, precipitation, evaporation, and relative humidity), water quality variations (pH, total organic carbon content, dissolved oxygen, and turbidity), accumulated sediments as an endogenous source, as well as unique estuarine conditions collectively facilitated picocyanobacterial-bacterial coexistence and community functional changes in this reservoir. A stable and tight co-occurrence pattern was established between dominant cyanobacteria (Synechococcus, Cyanobium, Planktothrix, Chroococcidiopsis, and Prochlorothrix) and certain heterotrophic bacteria (Proteobacteria, Actinobacteria, and Bacteroidetes), which contributed to the remineralization of organic matter for cyanobacteria utilization. The relative abundance of chemoorganoheterotrophs and bacteria related to nitrogen transformation (Paracoccus, Rhodoplanes, Nitrosomonas, and Zoogloea) increased, promoting the emergence of CBs in nutrient-limited conditions through enhanced nutrient recycling. In environments with limited nutrients, the interaction between photosynthetic autotrophic microorganisms and heterotrophic bacteria appears to be non-competitive. Instead, they adopt complementary roles within their ecological niche over long-term succession, mutually benefiting from this association. This long-term study confirmed that enhanced nutrient cycling, facilitated by cyanobacterial-bacterial symbiosis following long-term succession, could promote CBs in oligotrophic aquatic environments devoid of external nutrient inputs. This study advances understanding of the mechanisms that trigger and sustain CBs under nutritional constraints, contributing to developing more effective mitigation strategies, ensuring water safety, and maintaining ecological balance.

Usefulness of aircraft and airport wastewater for monitoring multiple pathogens including SARS-CoV-2 variants.

BACKGROUND: As global travel resumed in coronavirus disease 2019 (COVID-19) endemicity, the potential of aircraft wastewater monitoring to provide early warning of disease trends for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants and other infectious diseases, particularly at international air travel hubs, was recognized. We therefore assessed and compared the feasibility of testing wastewater from inbound aircraft and airport terminals for 18 pathogens including SARS-CoV-2 in Singapore, a popular travel hub in Asia. METHODS: Wastewater samples collected from inbound medium- and long-haul flights and airport terminals were tested for SARS-CoV-2. Next Generation Sequencing was carried out on positive samples to identify SARS-CoV-2 variants. Airport and aircraft samples were further tested for 17 other pathogens through quantitative reverse transcription polymerase chain reaction. RESULTS: The proportion of SARS-CoV-2-positive samples and the average virus load was higher for wastewater samples from aircraft as compared with airport terminals. Cross-correlation analyses indicated that viral load trends from airport wastewater led local COVID-19 case trends by 2-5 days. A total of 10 variants (44 sub-lineages) were successfully identified from aircraft wastewater and airport terminals, and four variants of interest and one variant under monitoring were detected in aircraft and airport wastewater 18-31 days prior to detection in local clinical cases. The detection of five respiratory and four enteric viruses in aircraft wastewater samples further underscores the potential to expand aircraft wastewater to monitoring pathogens beyond SARS-CoV-2. CONCLUSION: Our findings demonstrate the feasibility of aircraft wastewater testing for monitoring infectious diseases threats, potentially detecting signals before clinical cases are reported. The triangulation of similar datapoints from aircraft wastewater of international travel nodes could therefore serve as a useful early warning system for global health threats.

Size-dependent effects of microplastics on intestinal microbiome for Perna viridis.

Microplastics pollution threatens to marine organisms, particularly bivalves that actively ingest and accumulate microplastics of certain sizes, potentially disrupting intestinal homeostasis. This study investigated the microplastic abundance in wild and farmed mussels around Singapore, and examined the size-dependent effects of nano- to micro-scale polystyrene (0.5 µm/5 µm/50 µm) on the mussel intestinal microbiome in the laboratory. The field investigation revealed higher microplastic abundance in farmed mussels compared to wild ones. Experimentally, mussels exposed to 0.6 mg/L of microplastics for 7 days, followed by a 7-day depuration period, showed substantial impacts on Spirochaetes and Proteobacteria, facilitating the proliferation of pathogenic species and differentially affecting their pathogenic contributions. Metagenomics analysis revealed that microplastic exposure reduced Spirochaeta's contribution to virulence and pathogenicity loss, did not affect Vibrio and Oceanispirochaeta's pathogenicity, and increased Treponema and Oceanispirochaeta's contributions to pathogenicity loss. Moreover, microplastics increased transmembrane transporters and impacted oxidative phosphorylation enzymes, impairing energy metabolism. These effects persisted after depuration, indicating lack of resilience in the microbiome. Nano- and micro-scale plastics perturbed the mussel microbiome composition and functions in a size-dependent manner, with nano-plastics being the most disruptive. The increasing use and sale of aquaculture equipment of plastic may exacerbate the intestinal dysbiosis in bivalves, which threatens consumers' health.

Predicting Antibiotic Resistance and Assessing the Risk Burden from Antibiotics: A Holistic Modeling Framework in a Tropical Reservoir.

Predicting the hotspots of antimicrobial resistance (AMR) in aquatics is crucial for managing associated risks. We developed an integrated modeling framework toward predicting the spatiotemporal abundance of antibiotics, indicator bacteria, and their corresponding antibiotic-resistant bacteria (ARB), as well as assessing the potential AMR risks to the aquatic ecosystem in a tropical reservoir. Our focus was on two antibiotics, sulfamethoxazole (SMX) and trimethoprim (TMP), and on Escherichia coli (E. coli) and its variant resistant to sulfamethoxazole-trimethoprim (EC_SXT). We validated the predictive model using withheld data, with all Nash-Sutcliffe efficiency (NSE) values above 0.79, absolute relative difference (ARD) less than 25%, and coefficient of determination (R2) greater than 0.800 for the modeled targets. Predictions indicated concentrations of 1-15 ng/L for SMX, 0.5-5 ng/L for TMP, and 0 to 5 (log10 MPN/100 mL) for E. coli and -1.1 to 3.5 (log10 CFU/100 mL) for EC_SXT. Risk assessment suggested that the predicted TMP could pose a higher risk of AMR development than SMX, but SMX could possess a higher ecological risk. The study lays down a hybrid modeling framework for integrating a statistic model with a process-based model to predict AMR in a holistic manner, thus facilitating the development of a better risk management framework.

Virus surrogates throughout a full-scale advanced water reuse system.

Water reuse as an alternative water supply is increasing throughout the world due to water stress and scarcity; however, there are no standard practices for monitoring virus pathogens in such systems. This study aimed to identify suitable surrogates for virus fate, transport, and removal throughout a water reuse scheme. Various microbial targets (11 viruses, two phage, and three bacteria) were monitored using molecular and culture methods across all treatment stages in a wastewater reclamation facility and advanced water treatment facility. Criteria were established for identifying suitable surrogates, which included reliable detection, observable fate and transport, calculable log-reduction values (LRVs), correlations with other targets, and various morphological types. In total, five viruses (PMMoV, AiV, GII NoV, AdV, FRNA GII) met these stringent criteria and were suggested as potential virus surrogates. These surrogates enabled successful comparison of assigned versus actual LRVs throughout a water reuse scheme. Results suggest that virus pathogens are effectively removed throughout water reuse treatment and the suggested surrogates can be utilized for monitoring treatment performance and ensuring public health safety. This study provides a framework that water utilities across the world can reference for establishing virus monitoring practices.

Dissolved organic carbon spurs bacterial-algal competition and phosphorus-paucity adaptation: Boosting Microcystis' phosphorus uptake capacity.

Dissolved organic carbon (DOC) can alter the availability of background nutrients by affecting the proliferation of heterotrophic bacteria, which exerts a notable influence on algal growth and metabolism. However, the mechanism of how allochthonous DOC (aDOC) precipitates shifts in bacterial-algal interactions and modulates the occurrence of cyanobacteria blooms remains inadequately elucidated. Therefore, this study investigated the relationship between bacteria and algae under aDOC stimulation. We found that excess aDOC triggered the breakdown and reestablishment of the equilibrium between Microcystis and heterotrophic bacteria. The rapid proliferation of heterotrophic bacteria led to a dramatic decrease in soluble phosphorus and thereby resulted in the inhibition of the Microcystis growth. When the available DOC was depleted, the rapid death of heterotrophic bacteria released large amounts of dissolved phosphorus, which provided sufficient nutrients for the recovery of Microcystis. Notably, Microcystis rejuvenated and showed higher cell density compared to the carbon-absent group. This phenomenon can be ascribed that Microcystis regulated the compositions of extracellular polymeric substances (EPS) and the expression of relevant proteins to adapt to a nutrient-limited environment. Using time of flight secondary ion mass spectrometry (TOF-SIM) and proteomic analysis, we observed an enhancement of the signal of organic matter and metal ions associated with P complexation in EPS. Moreover, Microcystis upregulated proteins related to organic phosphorus transformation to increase the availability of phosphorus in various forms. In summary, this study emphasized the role of DOC in algal blooms, revealing the underestimated enhancement of Microcystis nutrient utilization through DOC-induced heterotrophic competition and providing valuable insights into eutrophication management and control.

Removal efficiency of antibiotic residues, antibiotic resistant bacteria, and genes across parallel secondary settling tank and membrane bioreactor treatment trains in a water reclamation plant.

Antimicrobial resistance is recognized as a potent threat to human health. Wastewater treatment facilities are viewed as hotspots for the spread of antimicrobial resistance. This study provides comprehensive data on the occurrences of 3 different antibiotic resistant opportunistic pathogens (with resistance to up to 5 antibiotics), 13 antibiotic resistant genes and intI1, and 22 different antimicrobial residues in a large water reclamation plant (176 million gallons per day) that runs a conventional Modified Ludzack-Ettinger (MLE) reactor followed by a secondary settling tank (SST) and membrane bioreactor (MBR) in parallel. All the antibiotic resistant bacteria and most of the antibiotic resistance genes were present in the raw influent, ranging from 2.5 × 102-3.7 × 106 CFU/mL and 1.2× 10-1-6.5 × 1010 GCN/mL, respectively. MBR outperformed the SST system in terms of ARB removal as the ARB targets were largely undetected in MBR effluent, with log removals ranging from 2.7 to 6.8, while SST only had log removals ranging from 0.27 to 4.6. Most of the ARG concentrations were found to have significantly higher in SST effluent than MBR permeate, and MBR had significantly higher removal efficiency for most targets (p < 0.05) except for sul1, sul2, blaOXA48, intI1 and 16S rRNA genes (p > 0.05). As for the antibiotic residues (AR), there was no significant removal from the start to the end of the treatment process, although MBR had higher removal efficiencies for azithromycin, chloramphenicol, erythromycin, erythromycin-H2O, lincomycin, sulfamethoxazole and triclosan, compared to the SST system. In conclusion, MBR outperformed SST in terms of ARB and ARGs removal. However low removal efficiencies of most AR targets were apparent.

Food waste compost and digestate as novel fertilizers: Impacts on antibiotic resistome and potential risks in a soil-vegetable system.

As a novel agricultural practice, the reuse of food waste compost and digestate as fertilizers leads to a circular economy, but inevitably introduces bio-contaminants such as antibiotic resistance genes (ARGs) into the agroecosystem. Moreover, heavy metal and antibiotic contamination in farmland soil may exert selective pressures on the evolution of ARGs, posing threats to human health. This study investigated the fate, influencing mechanisms and potential risks of ARGs in a soil-vegetable system under different food waste fertilization and remediation treatments and soil contamination conditions. Application of food waste fertilizers significantly promoted the pakchoi growth, but resulted in the spread of ARGs from fertilizers to pakchoi. A total of 56, 80, 84, 41, and 73 ARGs, mobile genetic elements (MGEs) and metal resistance genes (MRGs) were detected in the rhizosphere soil (RS), bulk soil (BS), control soil (CS), root endophytes (RE), and leaf endophytes (LE), respectively. Notably, 7 genes were shared in the above five subgroups, indicating a specific soil-root-endophytes transmission pathway. 36 genes were uniquely detected in the LE, which may originate from airborne ARGs. The combined application of biochar and fertilizers reduced the occurrence of ARGs and MGEs to some extent, showing the remediation effect of biochar. The average abundance of ARGs in the RS, BS and CS was 3.15 × 10-2, 1.31 × 10-2 and 2.35 × 10-1, respectively. Rhizosphere effects may reduce the abundance of ARGs in soil. The distribution pattern of ARGs was influenced by the types of soil, endophyte and contaminant. MGEs is the key driver shaping ARGs dynamics. Soil properties and pakchoi growth status may affect the bacterial composition, and consequently regulate ARGs fate, while endophytic ARGs were more impacted by biotic factors. Moreover, the average daily doses of ARGs from pakchoi consumption is 107-109 copies/d/kg, and its potential health risks should be emphasized.

Nano-scale and micron-scale plastics amplify the bioaccumulation of benzophenone-3 and ciprofloxacin, as well as their co-exposure effect on disturbing the antioxidant defense system in mussels, Perna viridis.

Plastics ranging from nano-scale to micron-scale are frequently ingested by many marine animals. These particles exhibit biotoxicity and additionally perform as vectors that convey and amass adsorbed chemicals within organisms. Meanwhile, the frequency of detection of the benzophenone-3 and ciprofloxacin can be adsorbed on plastic particles, then accumulated in bivalves, causing biotoxicity. To understand their unknown accumulative kinetics in vivo affected by different plastic sizes and toxic effect from co-exposure, several scenarios were set up in which the mode organism were exposed to 0.6 mg/L of polystyrene carrying benzophenone-3 and ciprofloxacin in three sizes (300 nm, 38 µm, and 0.6 mm). The live Asian green mussels were chosen as mode organism for exposure experiments, in which they were exposed to environments with plastics of different sizes laden with benzophenone-3 and ciprofloxacin, then depurated for 7 days. The bioaccumulation and depuration kinetics of benzophenone-3 and ciprofloxacin were measured using HPLC-MS/MS after one week of exposure and depuration. Meanwhile, their toxic effect were investigated by measuring the changes in six biomarkers (condition index, reactive oxygen species, catalase, glutathione, lipid peroxidation, cytochrome P450 and DNA damage). The bioconcentration factors in mussels under different exposure conditions were 41.48-111.75 for benzophenone-3 and 6.45 to 12.35 for ciprofloxacin. The results suggested that microplastics and nanoplastics can act as carriers to increase bioaccumulation and toxicity of adsorbates in mussels in a size-dependent manner. Overproduction of reactive oxygen species caused by microplastics and nanoplastics led to increased DNA damage, lipid peroxidation, and changes in antioxidant enzymes and non-enzymatic antioxidants during exposure. Marked disruption of antioxidant defenses and genotoxic effects in mussels during depuration indicated impaired recovery. Compared to micron-scale plastic with sizes over a hundred micrometers that had little effect on bivalve bioaccumulation and toxicity, nano-scale plastic greatly enhanced the biotoxicity effect.

Photochemical fate of quaternary ammonium compounds (QACs) and degradation pathways predication through computational analysis.

Quaternary ammonium compounds (QACs) are commonly used in many products, such as disinfectants, detergents and personal care products. However, their widespread use has led to their ubiquitous presence in the environment, posing a potential risk to human and environmental health. Several methods, including direct and indirect photodegradation, have been explored to remove QACs such as benzylalkyldimethyl ammonium compounds (BACs) and alkyltrimethyl ammonium compounds (ATMACs) from the environment. Hence, in this research, a systematic review of the literature was conducted using PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analysis) method to understand the fate of these QACs during direct and indirect photodegradation in UV/H2O2, UV/PS, UV/PS/Cu2+, UV/chlorine, VUV/UV/chlorine, O3/UV and UV/O3/TiO2 systems which produce highly reactive radicals that rapidly react with the QACs, leading to their degradation. As a result of photodegradation, several transformation products (TPs) of QACs are formed, which can pose a greater risk to the environment and human health than the parent QACs. Only limited research in this area has been conducted with fewer QACs. Hence, quantum mechanical calculations such as density functional theory (DFT)-based computational calculations using Gaussian09 software package were used here to explain better the photo-resistant nature of a specific type of QACs, such as BACs C12-18 and ATMACs C12, C14, C18, and their transformation pathways, providing insights into active sites participating in the phototransformation. Recognizing that different advanced oxidation processes (AOPs) come with pros and cons in the elimination of QACs, this review also highlighted the importance of implementing each AOP concerning the formation of toxic transformation products and electrical energy per order (EEO), especially when QACs coexist with other emerging contaminants (ECs).

Microcystis genotypes in a tropical freshwater lake: Discovery of novel MIB-producing Microcystis with potentially unique synthesis pathway.

Harmful algal blooms (HABs) are a threat to freshwater systems over the world due to the production of hepatotoxins like microcystin (MC), and nuisance taste and odour (T&O) compounds like 2-methylisoborneol (MIB). While MCs are known to cause detrimental effects to both water quality and human health, MIB is only reported to cause aesthetical problems. In this study, we investigated a tropical, urban lake that was experiencing persistent MC and MIB events. Although it was dominated by Microcystis blooms, analysis revealed that the toxigenic Microcystis were not the only species driving the MC concentrations. Additionally, there was also a lack of causative species for the MIB events. Through isolation, we have identified three toxigenic Microcystis found to produce four different variants of MCs, and two novel non-toxigenic Microcystis that were capable of producing MIB. The ability to produce MIB had never been previously reported for this species. Compared to other major producers such as Planktothricoides sp. and Streptomyces sp., the MIB synthase genes of our Microcystis sp. strains were partial, illustrating the possibility of unique synthesis pathways. The Microcystis sp. strains were found to produce about 2.77-5.22 fg MIB cell-1, with a majority of the contents (70-80 %) existing in the extracellular phase. Correlation analysis of field study indicated that phosphorus limitation may have an indirect effect on non-toxigenic Microcystis abundance and proportion by influencing the toxigenic genotype, suggesting that current measures to control HABs may favour the proliferation of the non-toxigenic Microcystis. The potential for Microcystis sp. to produce MIB through unique synthesis pathway, coupled with the potential dominance of non-toxigenic genotypes in Microcystis blooms, signals the possibility that non-toxigenic Microcystis should be monitored as well.

Assessing the additional health burden of antibiotic resistant Enterobacteriaceae in surface waters through an integrated QMRA and DALY approach.

Antibiotic resistant Enterobacteriaceae pose a significant threat to public health. However, limited studies have evaluated the health risks associated with exposure to antibiotic-resistant bacteria (ARB), especially in natural environments. While quantitative microbial risk assessment (QMRA) assesses microbial risks in terms of the probability of infection, it does not account for the severity of health outcomes. In this study, a QMRA-DALY model was developed to integrate QMRA with health burden (disability-adjusted life years (DALY)) from infections caused by ARB. The model considers uncertainties in probability of infection and health burden assessment using Monte Carlo simulations. The study collected antimicrobial resistance (AMR) surveillance data from surface waters with different land uses. Results revealed water bodies with agricultural land use to be the main AMR hotspots, with the highest additional health burden observed in infections caused by meropenem-resistant E. coli (∆DALY = 0.0105 DALY/event) compared to antibiotic-susceptible E. coli. The estimated ∆DALY for antibiotic-resistant K. pneumoniae was lower than for antibiotic-resistant E. coli (highest ∆DALY = 0.00048 DALY/event). The study highlights the need for better evaluation of AMR associated health burden, and effective measures to mitigate the risks associated with antibiotic-resistant bacteria in natural environments.

Size-dominated biotoxicity of microplastics laden with benzophenone-3 and ciprofloxacin: Enhanced integrated biomarker evaluation on mussels.

Microplastics (MPs) are emerging pollutants with diverse sizes in aquatic environments. This paper investigates the toxicity of micron- and nano-scale polystyrene (50 µm, 5 µm, 0.5 µm) loaded with 2-hydroxy-4-methoxy-benzophenone (BP-3) and ciprofloxacin (CIP) by eight biomarker responses in mussels, perna viridis. The mussels were exposed to MPs and chemicals for 7 days before 7 days of depuration. Eight biomarkers were measured to determine biotoxicity over time by using the weighted integrated biomarkers index evaluation (EIBR). Mussels exposed to MPs on a daily basis demonstrated a cumulative toxic effect. The toxicity of MPs for mussels was inversely related to the size at which they can be ingested. Then toxicity was reversed when exposure was halted. EIBR mold has shown a significant difference in the biotoxicity of each biological level under different exposure scenarios. In general, the mussel toxicity influenced by BP-3 and CIP exposure without an adsorbent was insignificant. MPs laden with them increased the toxicity of mussels. Under condition of lower concentration of ECs (Emerging contaminants), the presence of MPs as a component of a combined pollutant in water dominated the biotoxicity for mussels. The EIBR assessment further validated that the biotoxicity of mussels was size-dependent. Its application simplified the biomarkers' response index and enhanced the accuracy of evaluation by weighing on molecular, cellular and physiological level. Specifically, mussels were physiologically sensitive to nano-scale plastics, with nano-scale plastics causing a higher level of cellular immunity destruction and genotoxicity than micron-scale plastics. Enzymatic antioxidant systemswere upregulated based on size-differential plastics; however, the total antioxidant effect of non-enzymatic defenses appeared to be least affected by the size effect.

Low concentration quaternary ammonium compounds promoted antibiotic resistance gene transfer via plasmid conjugation.

During the pandemic of COVID-19, the amounts of quaternary ammonium compounds (QACs) used to inactivate the virus in public facilities, hospitals and households increased, which raised concerns about the evolution and transmission of antimicrobial resistance (AMR). Although QACs may play an important role in the propagation of antibiotic resistance gene (ARGs), the potential contribution and mechanism remains unclear. Here, the results showed that benzyl dodecyl dimethyl ammonium chloride (DDBAC) and didecyl dimethyl ammonium chloride (DDAC) significantly promoted plasmid RP4-mediated ARGs transfer within and across genera at environmental relevant concentrations (0.0004-0.4 mg/L). Low concentrations of QACs did not contribute to the permeability of the cell plasma membrane, but significantly increased the permeability of the cell outer membrane due to the decrease in content of lipopolysaccharides. QACs altered the composition and content of extracellular polymeric substances (EPS) and were positively correlated with the conjugation frequency. Furthermore, transcriptional expression levels of genes encode for mating pairing formation (trbB), DNA replication and translocation (trfA), and global regulators (korA, korB, trbA) are regulated by QACs. And we demonstrate for the first time that QACs decreased the concentration of extracellular AI-2 signals, which was verified to be involved in regulating conjugative transfer genes (trbB, trfA). Collectively, our findings underscore the risk of increased disinfectant concentrations of QACs on the ARGs transfer and provide new mechanisms of plasmid conjugation.

Per/polyfluoroalkyl substances modulate plasmid transfer of antibiotic resistance genes: A balance between oxidative stress and energy support.

Emerging contaminants can accelerate the transmission of antibiotic resistance genes (ARGs) from environmental bacteria to human pathogens via plasmid conjugation, posing a great challenge to the public health. Although the toxic effects of per/polyfluoroalkyl substances (PFAS) as persistent organic pollutants have been understood, it is still unclear whether and how PFAS modulate the transmission of ARGs. In this study, we for the first time reported that perfluorooctanoic acid (PFOA), perfluorododecanoic acid (PFDoA) and ammonium perfluoro (2-methyl-3-oxahexanoate) (GenX) at relatively low concentrations (0.01, 0.1 mg/L) promoted the conjugative transfer of plasmid RP4 within Escherichia coli, while the plasmid conjugation was inhibited by PFOA, PFDoA and GenX at relatively high concentrations (1, 10 mg/L). The non-unidirectional conjugation result was ascribed to the co-regulation of ROS overproduction, enhanced cell membrane permeability, shortage of energy support as well as l-arginine pool depletion. Taking the well-known PFOA as an example, it significantly enhanced the conjugation frequency by 1.4 and 3.4 times at relatively low concentrations (0.01, 0.1 mg/L), respectively. Exposure to PFOA resulted in enhanced cell membrane permeability and ROS overproduction in donor cells. At high concentrations of PFOA (1, 10 mg/L), although enhanced oxidative stress and cell membrane permeability still occurred, the ATP contents in E. coli decreased, which contributed to the inhibited conjugation. Transcriptome analysis further showed that the expression levels of genes related to arginine biosynthesis (argA, argC, argF, argG, argI) and transport (artJ, artM, artQ) pathways were significantly increased. Intracellular l-arginine concentration deficiency were observed at high concentrations of PFOA. With the supplementary exogenous arginine, it was demonstrated that arginine upregulated conjugation transfer- related genes (trfAp, trbBp) and restores the cell number of transconjugants in PFOA-treated group. Therefore, the inhibited conjugation at high concentrations PFOA were attributed to the shortage of ATP and the depletion of L-arginine pool. These findings provide important insights into the effect environmental concentrations of PFAS on the conjugative transfer of ARGs, and update the regulation mechanism of plasmid conjugation, which is critical for the management of antibiotic resistance in aquatic environments.