Extracellular Antibiotic Resistance Genes Research Articles

Deciphering the reduction of antibiotic resistance genes (ARGs) during medium-chain fatty acids production from waste activated sludge: Driven by inhibition of ARGs transmission and shift of microbial community

Medium-chain fatty acids (MCFAs) production from waste activated sludge (WAS) by chain extension (CE) is a promising technology. However, the effects and mechanisms of CE process on the fate of antibiotic resistance genes (ARGs) remain unclear. In this study, the results showed that the removal efficiency of ARGs was 81.15 % in CE process, suggesting its efficacy in reducing environmental risks. Further, the observed decrease in mobile genetic elements (MGEs) indicated that CE process restricted the horizontal gene transfer (HGT). Complementing this, the increase in soluble organic matters and extracellular 16 S rDNA confirmed that MCFAs production caused bacterial damage. Decreased intracellular ARGs and increased extracellular ARGs further revealed that MCFAs production impaired ARGs hosts, thereby limiting the vertical gene transfer (VGT) of ARGs. Shift of microbial community combined with co-occurrence network analysis demonstrated that functional bacteria without host potential for ARGs were enriched, but potential ARGs and MGEs hosts decreased, showing the role of functional bacterial phylogeny and selection pressure of MCFAs in reducing ARGs. Finally, partial least squares path model was used to systematic verify the mechanism of ARGs removal in CE process, which was attributed to the inhibition of ARGs transmission (HGT and VGT) and shift of microbial community.

Removal of Intracellular and Extracellular Antibiotic Resistance Genes and Virulence Factor Genes Using Electricity-Intensified Constructed Wetlands

Constructed wetland (CW) is considered a promising technology for the removal of emerging contaminants. However, its removal performance for antibiotic resistance genes (ARGs) is not efficient and influence of virulence factor genes (VFGs) have not been elucidated. Here, removal of intracellular and extracellular ARGs as well as VFGs by electricity-intensified CWs was comprehensively evaluated. The two electrolysis-intensified CWs can improve the removal of intracellular ARGs and MGEs to 0.96- and 0.85-logs, respectively. But cell-free extracellular ARGs (CF-eARGs) were significantly enriched with 1.8-logs in the electrolysis-intensified CW. Interestingly, adding Fe-C microelectrolysis to the electrolysis-intensified CW is conducive to the reduction of CF-eARGs. However, the detected number and relative abundances of intracellular and extracellular VFGs were increased in all of the three CWs. The biofilms attached onto the substrates and rhizosphere are also hotspots of both intracellular and particle-associated extracellular ARGs and VFGs. Structural equation models and correlation analysis indicated that ARGs and VFGs were significantly cooccurred, suggesting that VFGs may affect the dynamics of ARGs. The phenotypes of VFGs, such as biofilm, may act as protective matrix for ARGs, hindering the removal of resistance genes. Our results provide novel insights into the ecological remediation technologies to enhance the removal of ARGs. Environmental ImplicationConstructed wetland (CW) has been considered a promising technology for the removal of emerging contaminants. But its removal performance for antibiotic resistance genes (ARGs) is controversial. Here, we proposed electrolysis-intensified CW to enhance the removal of intracellular and extracellular ARGs. This strategy significantly promotes the removal of resistance genes. In addition, we evaluated the correlations between virulence factor genes (VFGs) and ARGs, suggesting that VFGs may affected the removal of ARGs using biofilm- or substrate-based technologies. These results lead significant guidance for the enhancement removal of ARGs using CWs, and highlight the need to take VFGs into account.

Enhanced sensitivity of extracellular antibiotic resistance genes (ARGs) to environmental concentrations of antibiotic

As emerging contaminants, antibiotics are frequently present in various environments, particularly rivers, albeit often at sublethal concentrations (ng/L∼μg/L). Assessing the risk associated with these low levels, which are far below the lethal threshold for most organisms, remains challenging. In this study, using microcosms containing planktonic bacteria and biofilm, we examined how antibiotic resistance genes (ARGs) in different physical states, including intracellular ARGs (iARGs) and extracellular ARGs (eARGs) responded to these low-level antibiotics. Our findings reveal a positive correlation between sub-lethal antibiotic exposure (ranging from 0.1 to 10 μg/L) and increased prevalence (measured as ARG copies/16s rDNA) of both iARGs and eARGs in planktonic bacteria. Notably, eARGs demonstrated greater sensitivity to antibiotic exposure compared to iARGs, with a lower threshold (0.1 μg/L for eARGs versus 1 μg/L for iARGs) for abundance increase. Moreover, ARGs in biofilms demonstrates higher sensitivity to antibiotic exposure compared to planktonic bacteria. To elucidate the underlying mechanisms, we established an integrated population dynamics-pharmacokinetics-pharmacodynamics (PD-PP) model. This model indicates that the enhanced sensitivity of eARGs is primarily driven by an increased potential for plasmid release from cells under low antibiotic concentrations. Furthermore, the accumulation of antibiotic in biofilms induces a greater sensitivity of ARG compared to the planktonic bacteria. This study provides a fresh perspective on the development of antibiotic resistance and offers an innovative approach for assessing the risk of sublethal antibiotic in the environment.

Dissolved oxygen facilitates efficiency of chlorine disinfection for antibiotic resistance

Controlling the dissemination of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) is a global concern. While commonly used chlorine disinfectants can damage or even kill ARB, dissolved oxygen (DO) may affect the formation of reactive chlorine species. This leads to the hypothesis that DO may play roles in mediating the effectiveness of chlorine disinfection for antibiotic resistance. To this end, this study investigated the impacts of DO on the efficiency of chlorine disinfection for antibiotic resistance. The results revealed that DO could increase the inactivation efficiency of ARB under chloramine and free chlorine exposure at practically relevant concentrations. Reactive species induced by DO, including H2O2, O2−, and OH, inactivated ARB strains by triggering oxidative stress response and cell membrane damage. In addition, the removal efficiency of extracellular ARGs (i.e. tetA and blaTEM) was enhanced with increasing dosage of free chlorine or chloramine under aerobic conditions. DO facilitated the fragmentation of plasmids, contributing to the degradation of extracellular ARGs under exposure to chlorine disinfectants. The findings suggested that DO facilitates disinfection efficiency for antibiotic resistance in water treatment systems.

Assessing intracellular and extracellular distribution of antibiotic resistance genes in the commercial organic fertilizers

Compost-based organic fertilizers often contain high levels of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs). Previous studies focused on quantification of total ARGs and MGEs. For a more accurate risk assessment of the dissemination risk of antibiotic resistance, it is necessary to quantify the intracellular and extracellular distribution of ARGs and MGEs. In the present study, extracellular ARGs and MGEs (eARGs and eMGEs) and intracellular ARGs and MGEs (iARGs and iMGEs) were separately analyzed in 51 commercial composts derived from different raw materials by quantitative polymerase chain reaction (qPCR) and metagenomic sequencing. Results showed that eARGs and eMGEs accounted for 11–56% and 4–45% of the total absolute abundance of ARGs and MGEs, respectively. Comparable diversity, host composition and association with MGEs were observed between eARGs and iARGs. Contents of high-risk ARGs were similar between eARGs and iARGs, with high-risk ARGs in the two forms accounting for 6.7% and 8.2% of the total abundances, respectively. Twenty-four percent of the overall ARGs were present in plasmids, while 56.7% of potentially mobile ARGs were found to be associated with plasmids. Variation partitioning analysis, null model and neutral community model indicated that the compositions of both eARGs and iARGs were largely driven by deterministic mechanisms. These results provide important insights into the cellular distribution of ARGs in manure composts that should be paid with specific attention in risk assessment and management.

Phthalates Boost Natural Transformation of Extracellular Antibiotic Resistance Genes through Enhancing Bacterial Motility and DNA Environmental Persistence.

The environmental dissemination of extracellular antibiotic resistance genes (eARGs) in wastewater and natural water bodies has aroused growing ecological concerns. The coexisting chemical pollutants in water are known to markedly affect the eARGs transfer behaviors of the environmental microbial community, but the detailed interactions and specific impacts remain elusive so far. Here, we revealed a concentration-dependent impact of dimethyl phthalate (DMP) and several other types of phthalate esters (common water pollutants released from plastics) on the natural transformation of eARGs. The DMP exposure at an environmentally relevant concentration (10 μg/L) resulted in a 4.8-times raised transformation frequency of Acinetobacter baylyi but severely suppressed the transformation at a high concentration (1000 μg/L). The promotion by low-concentration DMP was attributed to multiple mechanisms, including increased bacterial mobility and membrane permeability to facilitate eARGs uptake and improved resistance of the DMP-bounded eARGs (via noncovalent interaction) to enzymatic degradation (with suppressed DNase activity). Similar promoting effects of DMP on the eARGs transformation were also found in real wastewater and biofilm systems. In contrast, higher-concentration DMP suppressed the eARGs transformation by disrupting the DNA structure. Our findings highlight a potentially underestimated eARGs spreading in aquatic environments due to the impacts of coexisting chemical pollutants and deepen our understanding of the risks of biological-chemical combined pollution in wastewater and environmental water bodies.

Simultaneous elimination of antibiotic-resistant bacteria and antibiotic resistance genes by different Fe-N co-doped biochars activating peroxymonosulfate: The key role of pyridine-N and Fe-N sites

The coexistence of antibiotic resistance genes (ARGs) and antibiotic-resistant bacteria (ARB) in the environment poses a potential threat to public health. In our study, we have developed a novel advanced oxidation process for simultaneously removing ARGs and ARB by two types of iron and nitrogen-doped biochar derived from rice straw (FeN-RBC) and sludge (FeN-SBC). All viable ARB (approximately 108 CFU mL−1) was inactivated in the FeN-RBC/ peroxymonosulfate (PMS) system within 40 min and did not regrow after 48 h even in real water samples. Flow cytometry identified 96.7 % of dead cells in the FeN-RBC/PMS system, which verified the complete inactivation of ARB. Thorough disinfection of ARB was associated with the disruption of cell membranes and intracellular enzymes related to the antioxidant system. Whereas live bacteria (approximately 200 CFU mL−1) remained after FeN-SBC/PMS treatment. Intracellular and extracellular ARGs (tetA and tetB) were efficiently degraded in the FeN-RBC/PMS system. The production of active species, primarily •OH, SO4•- and Fe (IV), as well as electron transfer, were essential to the effective disinfection of FeN-RBC/PMS. In comparison with FeN-SBC, the better catalytic performance of FeN-RBC was mainly ascribed to its higher amount of pyridine-N and Fe0, and more reactive active sites (such as CO group and Fe-N sites). Density functional theory calculations indicated the greater adsorption energy and Bader charge, more stable Fe-O bond, more easily broken OO bond in FeN-RBC/PMS, which demonstrated the stronger electron transfer capacity between FeN-RBC and PMS. To encapsulate, our study provided an efficient and dependable method for the simultaneous elimination of ARGs and ARB in water.

Using transcriptome analysis to reveal the effects of bromoacetamide at drinking water ambient concentrations on the transformation of eDNA carrying ARGs

Antibiotic resistance genes (ARGs) are frequently detected in drinking water. Transformation of extracellular antibiotic resistance genes (eARGs) is an important pathway for horizontal gene transfer, which facilitates the spread of ARGs among different species. Although most studies have pointed out that the presence of some common carbon-containing disinfection by-products, such as trihalomethanes, can increase the frequency of transformation by increasing the content of reactive oxygen species (ROS), nitrogen-containing disinfection by-products (N-DBPs), such as bromoacetamide (BAcAm), can also induce bacterial resistance to antibiotics by stimulating ROS. However, it is unclear whether such N-DBPs can similarly promote the transformation of ARGs in bacteria. In this study, the effect of BAcAm on the transformation of eDNA carrying ARGs was investigated, and found that DNA integration and repair (ruv, rec), efflux pumps (emr), and genes related to membrane proteins (bam, opm) were up-regulated in E. coli after addition of BAcAm. Furthermore, in addition to analyzing common functional genes, in terms of metabolic gene expression, which is usually overlooked, the genes related to amino acid biosynthesis (pot, arg, lys) were up-regulated while genes related to sulfur metabolism, which has a higher metabolic cost (thi, sssu) were down-regulated, which suggests that E. coli is able to effectively defend itself against the threat posed by BAcAm to its survival. In conclusion, the factors affecting the conversion of eDNA carrying ARGs into E. coli cells by BAcAm were suggested and elaborated that E. coli carrying ARGs can adapt to various unfavorable environments by regulating relevant metabolic mechanisms and survive in drinking water environment for a long period of time.

Different mobile counter ion types of magnetic ion-exchange resin coupling with ozonation treatment of the organic matter, antibiotic resistance genes, and pathogens in secondary effluent

Advanced treatment and reuse of secondary effluents (SE) from wastewater treatment plants is considered an effective method of alleviating the conflict between water supply and demand. However, the effective removal of organic matter, antibiotic resistance genes (ARGs), and pathogens in SE is key to ensure the safety of reclaimed water. In this study, the effectiveness of three types of magnetic ion-exchange resins (R-X, where X refers to –OH, –HCO3, and -Cl) coupled with ozonation (R + O) in reducing different types of carbon, pathogens, intracellular and extracellular antibiotic resistance genes (iARGs and eARGs), and microbial communities in SE was investigated. The results indicated that R-Cl was the best pre-treatment resin for ozonation. R-Cl + O (8 mL/L R-Cl adsorption for 30 min, followed by 15 min of 8 mg/L ozonation) achieved removal efficiencies of 49.7 %, 86.6 %, 99.9 %, 87.1 %, and 99.9 % for dissolved organic carbon, UV254, ΣiARGs, ΣeARGs, and Σpathogens, respectively. Ozonation was more effective than resin adsorption in the removal of ARGs, and R-Cl + O had the lowest rebounding potential for iARGs and pathogens. In addition, R-OH had the lowest removal efficiency of organic matter, iARGs, and pathogens compared to other types of resins owing to the electrostatic repulsion mechanism. Molecular dynamics simulations and average reduced density gradient analysis revealed that the quaternary amine sites of the resin had a positive surface electrostatic potential, which could firmly adsorb double-stranded deoxyribonucleic acid by electrostatic action. Ozonation effectively reduced the risk of ARGs and pathogens in the effluent of resin. The R + O treatment reduced organic matter, ARGs, and pathogens more effectively than single treatment, despite the different anionic groups of the resin.

Gamma-ray irradiation as an effective method for mitigating antibiotic resistant bacteria and antibiotic resistance genes in aquatic environments

The prevalence of antibiotic resistance genes (ARGs) in municipal wastewater treatment plants (MWTPs) has emerged as a significant environmental concern. Despite advanced treatment processes, high levels of ARGs persist in the secondary effluent from MWTPs, posing ongoing environmental risks. This study explores the potential of gamma-ray irradiation as a novel approach for sterilizing antibiotic-resistant bacteria (ARB) and reducing ARGs in MWTP secondary effluent. Our findings reveal that gamma-ray irradiation at an absorbed dose of 1.6 kGy effectively deactivates all culturable bacteria, with no subsequent revival observed after exposure to 6.4 kGy and a 96-h incubation in darkness at room temperature. The removal efficiencies for a range of ARGs, including tetO, tetA, blaTEM-1, sulI, sulII, and tetW, were up to 90.5% with a 25.6 kGy absorbed dose. No resurgence of ARGs was detected after irradiation. Additionally, this study demonstrates a considerable reduction in the abundances of extracellular ARGs, with the transformation efficiencies of extracellular tetracycline and sulfadiazine resistance genes decreasing by 56.3–81.8% after 25.6 kGy irradiation. These results highlight the effectiveness of gamma-ray irradiation as an advanced and promising method for ARB sterilization and ARG reduction in the secondary effluent of MWTPs, offering a potential pathway to mitigate environmental risks associated with antibiotic resistance.

Mechanistic Aspects of Photodegradation of Deoxynucleosides Induced by Triplet State of Effluent Organic Matter.

Excited triplet states of wastewater effluent organic matter (3EfOM*) are known as important photo-oxidants in the degradation of extracellular antibiotic resistance genes (eArGs) in sunlit waters. In this work, we further found that 3EfOM* showed highly selective reactivity toward 2'-deoxyguanosine (dG) sites within eArGs in irradiated EfOM solutions at pH 7.0, while it showed no photosensitizing capacity toward 2'-deoxyadenosine, 2'-deoxythymidine, and 2'-deoxycytidine (the basic structures of eArGs). The 3EfOM* contributed to the photooxidation of dG primarily via one-electron transfer mechanism, with second-order reaction rate constants of (1.58-1.74) × 108 M-1 s-1, forming the oxidation intermediates of dG (dG(-H)•). The formed dG(-H)• could play a significant role in hole hopping and damage throughout eArGs. Using the four deoxynucleosides as probes, the upper limit for the reduction potential of 3EfOM* is estimated to be between 1.47 and 1.94 VNHE. Compared to EfOM, the role of the triplet state of terrestrially natural organic matter (3NOM*) in dG photooxidation was minor (∼15%) mainly due to the rapid reverse reactions of dG(-H)• by the antioxidant moieties of NOM. This study advances our understanding of the difference in the photosensitizing capacity and electron donating capacity between NOM and EfOM and the photodegradation mechanism of eArGs induced by 3EfOM*.

Impact of uranium on antibiotic resistance in activated sludge

The emergence of antibiotic resistance genes (ARGs) and antibiotic-resistant bacteria (ARB) in the environment is well established as a human health crisis. The impact of radioactive heavy metals on ecosystems and ultimately on human health has become a global issue, especially for the regions suffering various nuclear activities or accidents. However, whether the radionuclides can affect the fate of antibiotic resistance in bacteria remains poorly understood. Here, the dynamics of ARB, three forms of ARGs—intracellular ARGs (iARGs), adsorbed extracellular ARGs (aeARGs), and free extracellular ARGs (feARGs)—and microbial communities were investigated following exposure to uranium (U), a representative radioactive heavy metal. The results showed that 90-d of U exposure at environmentally relevant concentrations of 0.05 mg/L or 5 mg/L significantly increased the ARB concentration in activated sludge (p < 0.05). Furthermore, 90-d of U exposure slightly elevated the absolute abundance of aeARGs (except tetO) and sulfonamide iARGs, but decreased tetracycline iARGs. Regarding feARGs, the abundance of tetC, tetO, and sul1 decreased after 90-d of U stress, whereas sul2 showed the opposite trend. Partial least-squares path model analysis revealed that the abundance of aeARGs and iARGs under U stress was predominantly driven by increased cell membrane permeability/intI1 abundance and cell membrane permeability/reactive oxygen species concentration, respectively. Conversely, the changes in feARGs abundance depended on the composition of the microbial community and the expression of efflux pumps. Our findings shed light on the variations of ARGs and ARB in activated sludge under U exposure, providing a more comprehensive understanding of antibiotic resistance risks aggravated by radioactive heavy metal-containing wastewater.

Effective attenuation of extracellular antibiotic resistance gene risks in wastewater by capacitive deionization

Despite the effective inactivation on antibiotic resistant bacteria, current water and wastewater treatment technologies are often ineffective in removing antibiotic resistance genes (ARGs), which remain active outside cells, known as extracellular ARGs (eARGs). eARGs can persist for a long time and facilitate bacterial transformation in receiving environments, thereby, posing a threat to public health. This study applied capacitive deionization (CDI), as a promising desalination technology, for eARGs removal from wastewater. Results showed that CDI using carbon electrodes was significantly more effective than activated carbon adsorption in removing eARGs, with the removal efficiency and rate constant that are 2.1–11.2 times (p < 0.05) and 2.5–13 times (p < 0.05) higher. In the flow-by mode, CDI achieved the highest removal of eARGs (3.5 log) in just 15 min. Furthermore, CDI treatment significantly decreased the frequency of horizontal gene transfer (HGT) and increased potential for natural degradation, thereby reducing the risk of eARGs propagation. The high electric field (>1000 V/m) applied in CDI induces DNA conformational damage and DNA sequence mutations, which are the main mechanisms for removing eARG. This study will provide technical guidance and theoretical support for the removal of eARG in wastewater.

Biochar induces different responses of intracellular and extracellular antibiotic resistance genes and suppresses horizontal transfer during lincomycin fermentation dregs composting

This study aims to indicate the influence of biochar on extracellular and intracellular ARGs (e/iARGs) variation and proliferation during lincomycin fermentation dregs (LFDs) compost. Biochar addition made iARGs keep reducing but eARGs increase to the maximum at the middle thermophilic phase and reduce at the end of the compost. Compared to control 3.15-log and 5.42-log reduction of iARGs and eARGs were observed, respectively. Biochar addition, bacterial community, and MGEs were the major contributors to iARGs and eARGs removal, with the contribution percentages of 38.4%, 31.0%, 23.7%, and 27.2%, 29.1%, and 34.9%, respectively. Moreover, biochar significantly inhibited eARGs transformation and RP4 plasmid conjugative transfer among E. coli DH5α and Pseudomonas aeruginosa HLS-6. The underlying mechanism involved in broken cell membranes of bacteria, and altered expression of oxidative stress genes and save our souls (SOS) response-related genes. The results indicated that biochar addition in composting could limit the dissemination of ARGs.

Antibiotics and antibiotic resistance change bacterial community compositions in marine sediments

Emerging contaminants, including antibiotics, antibiotic-resistant bacteria (ARB), and extracellular antibiotic resistance genes (eARGs), have been detected in large numbers in the aquatic environment. The effects of emerging contaminants on bacterial communities in marine sediments are not well studied. In this study, the effects of emerging contaminants (antibiotics, ARB, and eARGs) on the variations of bacterial populations in marine sediments of the Bohai Sea, Yellow Sea, East China Sea, and South China Sea were investigated. The results showed that the abundance of the host bacterial phylum Probacteria in the marine sediments of the Bohai Sea was the lowest among the four seas after exposure to different antibiotics, ARB, and eARGs. The inputs of exogenous antibiotics and resistance genes significantly affected the community function, resulting in significant differences in community abundance at the genus level. The abundance of Halomonas, Sulfitobacter, and Alcanivorax in the four sea areas displayed noteworthy differences in response to the addition of exogenous antibiotics and eARGs. These findings contribute to a more comprehensive understanding of the intricate interplay between emerging contaminants and the dynamics of bacterial communities in natural ecosystems.

Facet-Specific Photocatalytic Degradation of Extracellular Antibiotic Resistance Genes by Hematite Nanoparticles in Aquatic Environments.

The persistence of extracellular antibiotic resistance genes (ARGs) in aquatic environments has attracted increasing attention due to their potential threat to public health and the environment. However, the fate of extracellular ARGs in receiving water remains largely unknown. This study investigated the influence of hematite nanoparticles, a widespread natural mineral, on the photodegradation of extracellular ARGs in river water. Results showed that under exposure to visible light, hematite nanoparticles, at environmental concentrations, resulted in a 3-5 orders of magnitude reduction in extracellular ARGs. This photodegradation of extracellular ARGs is shown to be facet-dependent; the (001) facet of hematite demonstrates a higher removal rate than that of the (100) facet, which is ascribed to its enhanced adsorption capability and higher hydroxyl radical (•OH) production. Density functional theory (DFT) calculations corroborate this finding, indicating elevated iron density, larger adsorption energy, and lower energy barrier of •OH formation on the (001) facet, providing more active sites and •OH generation for extracellular ARG interaction. Gel electrophoresis and atomic force microscopy analyses further confirm that the (001) facet causes more substantial damage to extracellular ARGs than the (100) facet. These findings pave the way for predicting the photodegradation efficiency of hematite nanoparticles with varied facets, thereby shedding light on the inherent self-purification capacity for extracellular ARGs in both natural and engineered aquatic environments.

Tracking the extracellular and intracellular antibiotic resistance genes across whole year in wastewater of intensive dairy farm

Monitoring the annual variation of antibiotic resistance genes (ARGs) in livestock wastewater is important for determining the high-risk period of transfer and spread of animal-derived antibiotic resistance into the environment. However, the knowledge regarding the variation patterns of ARGs, especially intracellular ARGs (iARGs) and extracellular ARGs (eARGs), over time in livestock wastewater is still unclear. Herein, we conducted a year-round study to trace the profiles of ARGs at a Chinese-intensive dairy farm, focusing on the shifts observed in different months. The results showed significant differences in the composition and variation between iARGs and eARGs. Tetracycline, sulfonamide, and macrolide resistance genes were the major types of iARGs, while cfr was the major type of eARG. The environmental adaptations of the host bacteria determine whether ARGs appear as intracellular or extracellular forms. The total abundance of ARGs was higher from April to September, which can be attributed to the favorable climatic conditions for bacterial colonization and increased antibiotic administration during this period. Integron was found to be highly correlated with most iARGs, potentially playing a role in the presence of these genes within cells and their similar transmission patterns in wastewater. The intracellular and extracellular bacterial communities were significantly different, primarily because of variations in bacterial adaptability to the high salt and anaerobic environment. The intracellular co-occurrence network indicated that some dominant genera in wastewater, such as Turicibacter, Clostridium IV, Cloacibacillus, Subdivision5_genera_incertae_sedis, Saccharibacteria_genera_incertae_sedis and Halomonas, were potential hosts for many ARGs. To the best of our knowledge, this study demonstrates, for the first time, the annual variation of ARGs at critical points in the reuse of dairy farm wastewater. It also offers valuable insights into the prevention and control of ARGs derived from animals.

Effects of oxytetracycline on variation in intracellular and extracellular antibiotic resistance genes during swine manure composting

This research aimed to investigate the alterations in extracellular (eARGs) and intracellular (iARGs) antibiotic resistance genes in response to oxytetracycline (OTC), and unravel the dissemination mechanism of ARGs during composting. The findings revealed both low (L-OTC) and high contents (H-OTC) of OTC significantly enhanced absolute abundance (AA) of iARGs (p < 0.05), compared to CK (no OTC). Composting proved to be a proficient strategy for removing eARGs, while AA of eARGs was significantly enhanced in H-OTC (p < 0.05). OTC resulted in an increase in AA of mobile genetic elements (MGEs), ATP levels, antioxidant and DNA repair enzymes in bacteria in compost product. Structural equation model further demonstrated that OTC promoted bacterial DNA repair and antioxidant enzyme activities, altered bacterial community and enhanced MGEs abundance, thereby facilitating iARGs dissemination. This study highlights OTC can increase eARGs and iARGs abundance, underscoring the need for appropriate countermeasures to mitigate potential hazards.

Degradation of chromosomal and plasmid encoded extracellular and intracellular resistance genes across different types and sizes in water matrices during ozone-based oxidation process

Antimicrobial resistance is a major and growing threat to public health and imposes significant environmental and economic burden worldwide. This study evaluates the degradation kinetics of resistance genes from ciprofloxacin (gyrAR), levofloxacin (qnrSR), tetracycline (tetAR), and mobile genetic elements (intl-1, plasmids) in water matrices by O3 treatment. For extracellular antibiotic resistance genes (e-ARGs) in deionized water, 3.6–5.2 logs removal was obtained, whereas intracellular ARGs (i-ARGs) in phosphate buffer saline, exhibited a removal of 3.5–4.5 logs at an O3 dosage of 3.7 × 10−2 M.s. While there was no significant impact of the wastewater matrix on e-gyrAR and e-intl-1 degradation as compared to deionized water (p > 0.05), the e-qnrSR reduction was higher (4.8–5.2 logs) at an ozone dosage of 3.7 × 10−2 M.s. For i-plasmid and e-plasmid encoded ARGs in wastewater, 4.0–5.5 logs removal was observed for the same dosage. Electrical Energy per Order (EEO, kWh/m3) was estimated, and the energy consumption on the degradation of the ARGs in the wastewater ranged between 0.68 and 1.1 kWh/m3. The findings of this study provide quantitative data for evaluating the fate and degradation of different ARG forms/types during ozone treatment of wastewater to mitigate the risk of AMR dissemination in the environment.

Efficient Removal of Antibiotic Resistance Genes through 4f‐2p‐3d Gradient Orbital Coupling Mediated Fenton‐Like Redox Processes

AbstractPeroxymonosulfate (PMS) mediated radical and nonradical active substances can synergistically achieve the efficient elimination of antibiotic resistance genes (ARGs). However, enhancing interface electron cycling and optimizing the coupling of the oxygen‐containing intermediates to improve PMS activation kinetics remains a major challenge. Here, Co doped CeVO4 catalyst (Co−CVO) with asymmetric sites was constructed based on Ce 4f−O 2p−Co 3d gradient orbital coupling. The catalyst achieved approximately 2.51×105 copies/mL of extracellular ARGs (eARGs) removal within 15 minutes, exhibited ultrahigh degradation rate (k=1.24 min−1). The effective gradient 4f‐2p‐3d orbital coupling precisely regulates the electron distribution of Ce−O−Co active center microenvironment, while optimizing the electronic structure of Co 3d states (especially the occupancy of eg), promoting the adsorption of oxygen‐containing intermediates. The generated radical and nonradical generated by interfacial electron cycling enhanced by the reduction reaction of PMS at the Ce site and the oxidation reaction at the Co site achieved a significant mineralization rate of ARGs (83.4 %). The efficient removal of ARGs by a continuous flow reactor for 10 hours significantly reduces the ecological risk of ARGs in actual wastewater treatment.