Climate warming increases the proportions of specific antibiotic resistance genes in natural soil ecosystems. - Related Documents




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697801.0000Climate warming increases the proportions of specific antibiotic resistance genes in natural soil ecosystems. Understanding the future distribution of antibiotic resistance in natural soil ecosystems is important to forecast their impacts on ecosystem and human health under projected climate change scenarios. Therefore, it is critical and timely to decipher the links between climate warming and antibiotic resistance, two of Earth's most imminent problems. Here, we explored the role of five-year simulated climate warming (+ 4 °C) on the diversity and proportions of soil antibiotic resistance genes (ARGs) across three seasons in both plantation and natural forest ecosystems. We found that the positive effects of warming on the number and proportions of ARGs were dependent on the sampling seasons (summer, autumn and winter), and seasonality was a key factor driving the patterns of ARG compositions in forest soils. Fifteen ARGs, conferring resistance to common antibiotics including aminoglycoside, beta-lactam, macrolide-lincosamide-streptogramin B, multidrug, sulfonamide, and tetracycline, were significantly enriched in the warming treatment. We showed that changes in soil properties and community compositions of bacteria, fungi and protists can explain the changes in soil ARGs under climate warming. Taken together, these findings advance our understanding of environmental ARGs under the context of future climate change and suggest that elevated temperature may promote the abundance of specific soil ARGs, with important implications for ecosystem and human health.202235158246
697710.9999Tracking virulence genes and their interaction with antibiotic resistome during manure fertilization. Antibiotic resistance genes, collectively termed as antibiotic resistome, are regarded as emerging contaminants. Antibiotics resistome can be highly variable in different environments, imposing environmental safety concern and public health risk when it is in conjunction with pathogenic bacteria. However, it remains elusive how pathogenic bacteria interact with antibiotic resistome, making it challenging to assess microbial risk. Here, we examined the presence and relative abundance of bacterial virulence genes representing potential pathogens in swine manure, compost, compost-amended soil, and unamended agricultural soil in five suburban areas of Beijing, China. The absolute abundances of virulence genes were marginally significantly (p < 0.100) increased in compost-amended soils than unamended soil, revealing potential health risks in manure fertilization. The composition of potential pathogens differed by sample types and was linked to temperature, antibiotics, and heavy metals. As antibiotics can confer pathogens the resistance to clinic treatment, it was alarming to note that virulence genes tended to co-exist with antibiotic resistance genes, as shown by prevalently positive links among them. Collectively, our results demonstrate that manure fertilization in agriculture might give rise to the development of potentially antibiotic-resistant pathogens, unveiling an environmental health risk that has been frequently overlooked.202235810986
682620.9999Impact of land use on antibiotic resistance genes and bacterial communities in rivers. River ecosystems support essential ecosystem functions and services, including supplying water for domestic, agricultural, and industrial activities, provisioning of hydropower and fisheries, supporting navigation and recreational activities, and regulating water quality. In recent decades, the presence and spread of antibiotic resistance genes (ARGs) have emerged as a key threat to ecosystem health and human well-being. Rivers that are surrounded by human-modified landscapes serve as primary repositories and sources of ARGs. However, our understanding of the relationship between the diversity of ARGs and land use remain limited. We collected 30 sediment samples from five rivers in Ningbo, China, and then classified the sampling sites into two groups (i.e., group A with low levels of human impacts and group B with intense human impact) based on land use in their upstream areas. In total, we found 31 types of ARGs and 148 phyla of bacteria in the samples. ARGs abundance had a positive relationship with the levels of anthropogenic activities, and exhibited significant difference between the two groups. Co-occurrence networks showed that the interrelationship between bacteria and ARGs was more complex in group B than in group A. Moreover, Structural Equation Modeling (SEM) revealed that anthropogenic activity not only posed direct effect on ARGs but also indirectly affected ARGs through bacteria. Our results underscore the profound impacts of land-use changes on the diversity of ARGs, bacteria communities, and their relationships, which highlight the need for integrating ARGs in river assessments in regions with human-dominated land use.202540154785
697630.9999Unveiling the critical role of overlooked consumer protist-bacteria interactions in antibiotic resistance gene dissemination in urban sewage systems. Antibiotic resistance genes (ARGs) are emerging contaminants of significant concern due to their role in facilitating the spread of antibiotic resistance, especially high-risk ARGs, which are characterized by high human accessibility, gene mobility, pathogenicity, and clinical availability. Studies have shown that cross-domain interactions, such as those between consumer protists (consumers) and bacteria, can influence bacterial diversity, distribution, and function through top-down control. The consumers-bacteria interactions may also affect the occurrence and distribution of ARGs, yet this has been scarcely explored in field investigations. We conducted a city-scale investigation of ARGs, protists, and bacterial communities across each unit of the urban sewage system (USS), including 49 sewage pumping stations (SW), as well as influent (IF), activated sludge (AS), and effluent (EF) from seven wastewater treatment plants. Interestingly, consumers-bacteria interactions, as indicated by indices of bipartite relevance networks (i.e., connectedness and cohesion), increased from SW and IF to AS and EF. Structural equation modelling (SEM) revealed that consumers-bacteria interactions had a greater influence on the abundance of total ARGs and high-risk ARGs than seasonal or environmental factors. Notably, the total effects of consumers-bacteria interactions in SEM were significant (P < 0.05) and comparable in both IF and EF, even with the decrease in ARG abundance from IF to EF. This suggests a potential risk of ARG spread to the environment, facilitated by consumer protists in the EF. Additionally, the relevance network also demonstrated an increasing trend in the relationships between consumer protists and potential hosts of high-risk ARGs from raw sewage (SW and IF) to AS and EF. Overall, this study emphasizes the importance of integrating multitrophic microbial interactions to better understand and mitigate the dissemination of ARGs in sewage systems.202539662352
683140.9999Diversity of herbaceous plants and bacterial communities regulates soil resistome across forest biomes. Antibiotic resistance is ancient and prevalent in natural ecosystems and evolved long before the utilization of synthetic antibiotics started, but factors influencing the large-scale distribution patterns of natural antibiotic resistance genes (ARGs) remain largely unknown. Here, a large-scale investigation over 4000 km was performed to profile soil ARGs, plant communities and bacterial communities from 300 quadrats across five forest biomes with minimal human impact. We detected diverse and abundant ARGs in forests, including over 160 genes conferring resistance to eight major categories of antibiotics. The diversity of ARGs was strongly and positively correlated with the diversity of bacteria, herbaceous plants and mobile genetic elements (MGEs). The ARG composition was strongly correlated with the taxonomic structure of bacteria and herbs. Consistent with this strong correlation, structural equation modelling demonstrated that the positive effects of bacterial and herb communities on ARG patterns were maintained even when simultaneously accounting for multiple drivers (climate, spatial predictors and edaphic factors). These findings suggest a paradigm that the interactions between aboveground and belowground communities shape the large-scale distribution of soil resistomes, providing new knowledge for tackling the emerging environmental antibiotic resistance.201829687575
682750.9999Metagenomic profiles of planktonic bacteria and resistome along a salinity gradient in the Pearl River Estuary, South China. Estuarine ecosystems undergo pronounced and intricate changes due to the mixing of freshwater and saltwater. Additionally, urbanization and population growth in estuarine regions result in shifts in the planktonic bacterial community and the accumulation of antibiotic resistance genes (ARGs). The dynamic changes in bacterial communities, environmental factors, and carriage of ARGs from freshwater to seawater, as well as the complex interrelationships among these factors, have yet to be fully elucidated. Here, we conducted a comprehensive study based on metagenomic sequencing and full-length 16S rRNA sequencing, covering the entire Pearl River Estuary (PRE) in Guangdong, China. The abundance and distribution of the bacterial community, ARGs, mobile genetic elements (MGEs), and bacterial virulence factors (VFs) were analyzed on a site-by-site basis through sampling along the salinity gradient in PRE, from upstream to downstream. The structure of the planktonic bacterial community undergoes continuous changes in response to variations in estuarine salinity, with the phyla Proteobacteria and Cyanobacteria being dominant bacterial throughout the entire region. The diversity and abundance of ARGs and MGEs gradually decreased with the direction of water flow. A large number of ARGs were carried by potentially pathogenic bacteria, especially in Alpha-proteobacteria and Beta-proteobacteria. Multi-drug resistance genes have the highest abundance and subtypes in PRE. In addition, ARGs are more linked to some MGEs than to specific bacterial taxa and disseminate mainly by HGT and not by vertical transfer in the bacterial communities. Various environmental factors, such as salinity and nutrient concentrations, have a significantly impact on the community structure and distribution of bacteria. In conclusion, our results represent a valuable resource for further investigating the intricate interplay between environmental factors and anthropogenic disturbances on bacterial community dynamics. Moreover, they contribute to a better understanding of the relative impact of these factors on the dissemination of ARGs.202337211102
697460.9999Comparison of soil and grass microbiomes and resistomes reveals grass as a greater antimicrobial resistance reservoir than soil. Grasslands cover a large proportion of global agricultural landmass used to feed herbivores and ruminants and link the environment to the food chain via animals onto humans. However, most scientific studies of antimicrobial resistance and microbiomes at the environmental - animal nexus have focused on soil or vegetables rather than grasslands. Based on previous microbiome phyllosphere-soil studies we hypothesised that the microbiome and resistomes across soil and grass would have a core of shared taxa and antimicrobial resistance genes (ARGs), but that in addition each would also have a minority of unique signatures. Our data indicated grass contained a wider variety and higher relative abundance of ARGs and mobile genetic elements (MGEs) than soil with or without slurry amendments. The microbiomes of soil and grass were similar in content but varied in the composition proportionality. While there were commonalities across many of the ARGs present in soil and on grass their correlations with MGEs and bacteria differed, suggesting a source other than soil is also relevant for the resistome of grass. The variations in the relative abundances of ARGs in soil and on grass also indicated that either the MGEs or the bacteria carrying the ARGs comprised a higher relative abundance on grass than in soil. We conclude that while soil may be a source of some of these genes it cannot be the source for all ARGs and MGEs. Our data identifies grass as a more diverse and abundant reservoir of ARGs and MGEs in the environment than soil, which is significant to human and animal health when viewed in the context of grazing food animals.202336191722
683270.9999Historical trajectories of antibiotics resistance genes assessed through sedimentary DNA analysis of a subtropical eutrophic lake. Investigating the occurrence of antibiotic-resistance genes (ARGs) in sedimentary archives provides opportunities for reconstructing the distribution and dissemination of historical (i.e., non-anthropogenic origin) ARGs. Although ARGs in freshwater environments have attracted great attention, historical variations in the diversity and abundance of ARGs over centuries to millennia remain largely unknown. In this study, we investigated the vertical change patterns of bacterial communities, ARGs and mobile genetic elements (MGEs) found in sediments of Lake Chenghai spanning the past 600 years. Within resistome preserved in sediments, 177 ARGs subtypes were found with aminoglycosides and multidrug resistance being the most abundant. The ARG abundance in the upper sediment layers (equivalent to the post-antibiotic era since the 1940s) was lower than those during the pre-antibiotic era, whereas the ARG diversity was higher during the post-antibiotic era, possibly because human-induced lake eutrophication over the recent decades facilitated the spread and proliferation of drug-resistant bacteria. Statistical analysis suggested that MGEs abundance and the bacterial community structure were significantly correlated with the abundance and diversity of ARGs, suggesting that the occurrence and distribution of ARGs may be transferred between different bacteria by MGEs. Our results provide new perspectives on the natural history of ARGs in freshwater environments and are essential for understanding the temporal dynamics and dissemination of ARGs.202438621322
698280.9998Viral Communities Suppress the Earthworm Gut Antibiotic Resistome by Lysing Bacteria on a National Scale. Earthworms are critical in regulating soil processes and act as filters for antibiotic resistance genes (ARGs). Yet, the geographic patterns and main drivers of earthworm gut ARGs remain largely unknown. We collected 52 earthworm and soil samples from arable and forest ecosystems along a 3000 km transect across China, analyzing the diversity and abundance of ARGs using shotgun metagenomics. Earthworm guts harbored a lower diversity and abundance of ARGs compared to soil, resulting in a stronger distance-decay rate of ARGs in the gut. Greater deterministic assembly processes of ARGs were found in the gut than in soil. The earthworm gut had a lower frequency of co-occurrence patterns between ARGs and mobile genetic elements (MGEs) in forest than in arable systems. Viral diversity was higher in the gut compared to soil and was negatively correlated with bacterial diversity. Bacteria such as Streptomyces and Pseudomonas were potential hosts of both viruses and ARGs. Viruses had negative effects on the diversity and abundance of ARGs, likely due to the lysis on ARG-bearing bacteria. These findings provide new insights into the variations of ARGs in the earthworm gut and highlight the vital role of viruses in the regulation of ARGs in the soil ecosystem.202439037720
697990.9998Urbanization increases high-risk antibiotic resistance genes and pathogenic bacteria in soil and phyllosphere microbiomes. Rapid urban expansion has transformed agricultural and natural land into industrial, commercial, and residential areas, leading to substantial changes in land use and vegetation types, which exert a profound impact on microbial diversity. However, the responses of soil-plant multitrophic microbial communities to urbanization and its upshots on the profiles of antibiotic resistance genes (ARGs) and virulence factor genes (VFGs) remain poorly explored. This study explored the urban soil and grass phyllosphere microbiomes across five levels of urban gradient, defined by building densities. Our findings reveal the lowest α-diversity of bacteria, fungi, and protists in highly urbanized areas linked with a notable increase in ARGs, and VFGs. The highly populated areas mostly associated with the decrease of habitat green patch sizes that are exposed to the various anthropogenic stocks, and high-risk ARGs pose the utmost vulnerability risks for human well-being. The high-risk genes encoding resistance to multidrug (mdtG, emrD, and mepA), and glycopeptide (vanA) and their associated human potential pathogens were remarkably abundant in soil and grass phyllosphere. Our findings underscore the complex relationships between urbanization, microbial diversity, and resistome, ultimately, it is crucial to monitor the main source of clinical ARGs/VFGs for proper, effective sustainable urban management and public health interventions.202540555022
6981100.9998Decline in the Relative Abundance of Antibiotic Resistance Genes in Long-Term Fertilized Soil and Its Driving Factors. The changes in antibiotic resistance genes (ARGs) in long-term fertilized soil remain controversial. We aimed to analyze the variation characteristics of ARGs in long-term fertilized soil using metagenomic sequencing. The relative abundance of ARGs did not increase significantly after 7 years of fertilization. However, a clear decline in the relative abundance of ARGs was observed compared to the data from the 4th year. Microbial adaptation strategies in response to changes in the ARG abundance were associated with shifts in microbiome composition and function. Among these, bacterial abundance was the primary driving factor. Additionally, total heavy metal content might serve as the most significant co-selective pressure influencing ARG number. We believe that increasing the selective pressure from heavy metals and antibiotics might result in the loss of certain microbial species and a decrease in ARG abundance. This study provides novel insights into the variations of soil resistance genes under long-term fertilization.202540785530
7389110.9998Temporal changes of antibiotic-resistance genes and bacterial communities in two contrasting soils treated with cattle manure. The emerging environmental spread of antibiotic-resistance genes (ARGs) and their subsequent acquisition by clinically relevant microorganisms is a major threat to public health. Animal manure has been recognized as an important reservoir of ARGs; however, the dissemination of manure-derived ARGs and the impacts of manure application on the soil resistome remain obscure. Here, we conducted a microcosm study to assess the temporal succession of total bacteria and a broad spectrum of ARGs in two contrasting soils following manure application from cattle that had not been treated with antibiotics. High-capacity quantitative PCR detected 52 unique ARGs across all the samples, with β-lactamase as the most dominant ARG type. Several genes of soil indigenous bacteria conferring resistance to β-lactam, which could not be detected in manure, were found to be highly enriched in manure-treated soils, and the level of enrichment was maintained over the entire course of 140 days. The enriched β-lactam resistance genes had significantly positive relationships with the relative abundance of the integrase intI1 gene, suggesting an increasing mobility potential in manure-treated soils. The changes in ARG patterns were accompanied by a significant effect of cattle manure on the total bacterial community compositions. Our study indicates that even in the absence of selective pressure imposed by agricultural use of antibiotics, manure application could still strongly impact the abundance, diversity and mobility potential of a broad spectrum of soil ARGs. Our findings are important for reliable prediction of ARG behaviors in soil environment and development of appropriate strategies to minimize their dissemination.201626712351
6983120.9998Deciphering Potential Roles of Earthworms in Mitigation of Antibiotic Resistance in the Soils from Diverse Ecosystems. Earthworms are capable of redistributing bacteria and antibiotic resistance genes (ARGs) through soil profiles. However, our understanding of the earthworm gut microbiome and its interaction with the antibiotic resistome is still lacking. Here, we characterized the earthworm gut and soil microbiome and antibiotic resistome in natural and agricultural ecosystems at a national scale, and microcosm studies and field experiments were also employed to test the potential role of earthworms in dynamics of soil ARGs. The diversity and structure of bacterial communities were different between the earthworm gut and soil. A significant correlation between bacterial community dissimilarity and spatial distance between sites was identified in the earthworm gut. The earthworm gut consistently had lower ARGs than the surrounding soil. A significant reduction in the relative abundance of mobile genetic elements and dominant bacterial phylotypes that are the likely hosts of ARGs was observed in the earthworm gut compared to the surrounding soil, which might contribute to the decrease of ARGs in the earthworm gut. The microcosm studies and field experiments further confirmed that the presence of earthworms significantly reduced the number and abundance of ARGs in soils. Our study implies that earthworm-based bioremediation may be a method to reduce risks associated with the presence of ARGs in soils.202133977709
6955130.9998Soil antibiotic resistance genes accumulate at different rates over four decades of manure application. Manure can be a source of antibiotic resistance genes (ARGs) that enter the soil. However, previous studies assessing ARG persistence in soil have generally lacked continuity over sampling times, consistency of location, and assessing the impact of discontinuing manure application. We evaluated both short- and long-term ARG accumulation dynamics in soil with a 40-year known history of manure use. Manure application caused a greater abundance of tetracycline, macrolide, and sulfonamide ARGs in the soil. There was an initial spike in ARG abundance resulting from manure bacteria harboring ARGs being introduced to soil, followed by resident soil bacteria out-competing them, which led to ARG dissipation within a year. However, over four decades, annual manure application caused linear or exponential ARG accumulation, and bacteria associated with ARGs differed compared to those in the short term. Eleven years after discontinuing manure application, most soil ARG levels declined but remained elevated. We systematically explored the historical accumulation of ARGs in manured soil, and provide insight into factors that affect their persistence.202336444046
7506140.9998Risk assessment and dissemination mechanism of antibiotic resistance genes in compost. In recent years, the excessive of antibiotics in livestock and poultry husbandry, stemming from extensive industry experience, has resulted in the accumulation of residual antibiotics and antibiotic resistance genes (ARGs) in livestock manure. Composting, as a crucial approach for the utilization of manure resources, has the potential to reduce the levels of antibiotics and ARGs in manure, although complete elimination is challenging. Previous studies have primarily focused on the diversity and abundance of ARGs in compost or have solely examined the correlation between ARGs and their carriers, potentially leading to a misjudgment of the actual risk associated with ARGs in compost. To address this gap, this study investigated the transfer potential of ARGs in compost and their co-occurrence with opportunistic pathogenic bacteria by extensively analyzing metagenomic sequencing data of compost worldwide. The results demonstrated that the potential risk of ARGs in compost was significantly lower than in manure, suggesting that composting effectively reduces the risk of ARGs. Further analysis showed that the microbes shifted their life history strategy in manure and compost due to antibiotic pressure and formed metabolic interactions dominated by antibiotic-resistant microbes, increasing ARG dissemination frequency. Therefore, husbandry practice without antibiotic addition was recommended to control ARG evolution, dissemination, and abatement both at the source and throughout processing.202337562342
6816150.9998The impact of microplastics on antibiotic resistance genes, metal resistance genes, and bacterial community in aquaculture environment. Microplastics are emerging contaminants. However, their effects on antibiotic resistance genes (ARGs), metal resistance genes (MRGs), and the structure and abundance of bacterial communities, particularly pathogens, in aquaculture environments remains poorly understood. Therefore, this study investigated the effect of microplastics of different sizes on the abundance and distribution of ARGs, MRGs, and bacterial communities in aquaculture environments. The results revealed that, compared with pond water, large microplastics harbored significantly higher ARG abundances, particularly for multidrug-resistant genes; notably, level-I- and -II-risk ARGs were more prevalent on microplastics, highlighting the potential for horizontal gene transfer. Microplastics also exhibited a propensity to aggregate pathogenic bacteria such as Brucella and Pseudomonas, which could pose direct risks to aquatic product safety and public health. Network and differential network analyses revealed significant correlations between bacterial genera and ARG/MRG abundance, particularly on microplastics. Therefore, our findings suggest that microplastics act as vectors for the spread of ARGs, MRGs, and pathogens in aquaculture, potentially leading to the formation of complexes of these materials that threaten ecosystem health and human well-being. This study provides critical insights into the need for targeted management strategies to mitigate microplastic pollution in aquaculture settings.202539987738
6884160.9998The changes in antibiotic resistance genes during 86 years of the soil ripening process without anthropogenic activities. This study aimed to reveal the baseline of natural variations in antibiotic resistance genes (ARGs) in soil without anthropogenic activities over the decades. Nine soil samples with different time of soil formation were taken from the Yancheng Wetland National Nature Reserve, China. ARGs and mobile genetic elements (MGEs) were characterized using metagenomic analysis. A total of 196 and 192 subtypes of ARGs were detected in bulk soil and rhizosphere, respectively. The diversity and abundance of ARGs were stable during 69 years probably due to the alkaline pH soil environment but not due to antibiotics. Increases in ARGs after 86 years were probably attributed to more migrant birds inhabited compared with other sampling sites. Multidrug was the most abundant type, and largely shared by soil samples. It was further shown that soil samples could not be clearly distinguished, suggesting a slow process of succession of ARGs in the mudflat. The variation partitioning analysis revealed that the ARG profile was driven by the comprehensive effects exhibited by the bacterial community, MGEs, and environmental factors. Besides, pathogenic bacteria containing ARGs mediated by migrant birds in the area with 86 years of soil formation history nearing human settlements needed special attention. This study revealed the slow variations in ARGs in the soil ripening process without anthropogenic activities over decades, and it provided information for assessing the effect of human activities on the occurrence and dissemination of ARGs.202133228990
6980170.9998Effects of agricultural inputs on soil virome-associated antibiotic resistance and virulence: A focus on manure, microplastic and pesticide. Soil viruses are increasingly recognized as crucial mediators of horizontal gene transfer, yet their role in disseminating antibiotic resistance genes (ARGs) and virulence factor genes (VFGs) under agricultural disturbances remains poorly understood. Here, we characterized the viromes and associated ARGs and VFGs in agricultural soils treated with low- and high-dose manures, microplastics, and pesticides. Using metagenomic sequencing coupled with advanced viral identification tools, we found that manure fertilization markedly altered viral community composition and increased viral diversity. Manure also enhanced the abundance of ARGs and VFGs in viromes by 2.0-9.8-fold and 2.0-8.1-fold, respectively, while microplastics and pesticides had limited impacts. Additionally, gene pathways related to human diseases and environmental adaptation were enriched in soil viromes treated with manures and high-dose pesticides. Virus-host prediction revealed that Actinomycetia dominated bacterial hosts of both ARG- and VFG-carrying viruses, with some VFG-carrying viruses linked to potential human pathogens, e.g., Escherichia albertii and Klebsiella pneumoniae. Co-occurrence network analysis indicated that these disturbances strengthened connections between bacteria, viruses, and ARGs (or VFGs). Our study provides a comprehensive profile of viromes and associated risks in agricultural soil under three disturbances, highlighting the role of viruses in spread of antibiotic resistance and pathogenic risks in agricultural soil.202540752173
6815180.9998Bacterial community succession and the enrichment of antibiotic resistance genes on microplastics in an oyster farm. Microplastics can be colonized by microorganisms and form plastisphere. However, knowledge of bacterial community succession and the enrichment of antibiotic resistance genes (ARGs) and pathogens on microplastics in aquaculture environments is limited. Here, we conducted a 30-day continuous exposure experiment at an oyster farm. Results showed that the alpha-diversity of communities on most microplastics continuously increased and was higher than in planktonic communities after 14 days. Microplastics could selectively enrich certain bacteria from water which can live a sessile lifestyle and promote colonization by other bacteria. The composition and function of plastisphere communities were distinct from those in the surrounding water and influenced by polymer type and exposure time. Microplastics can enrich ARGs (sul1, qnrS and bla(TEM)) and harbor potential pathogens (e.g., Pseudomonas aeruginosa). Therefore, microplastic pollution may pose a critical threat to aquaculture ecosystems and human health. Our study provides further insight into the ecological risks of microplastics.202337611336
6878190.9998Reduction in antimicrobial resistance in a watershed after closure of livestock farms. Natural environments play a crucial role in transmission of antimicrobial resistance (AMR). Development of methods to manage antibiotic resistance genes (ARGs) in natural environments are usually limited to the laboratory or field scale, partially due to the complex dynamics of transmission between different environmental compartments. Here, we conducted a nine-year longitudinal profiling of ARGs at a watershed scale, and provide evidence that restrictions on livestock farms near water bodies significantly reduced riverine ARG abundance. Substantial reductions were revealed in the relative abundance of genes conferring resistance to aminoglycosides (42%), MLSB (36%), multidrug (55%), tetracyclines (53%), and other gene categories (59%). Additionally, improvements in water quality were observed, with distinct changes in concentrations of dissolved reactive phosphorus, ammonium, nitrite, pH, and dissolved oxygen. Antibiotic residues and other pharmaceuticals and personal care products (PPCPs) maintain at a similarly low level. Microbial source tracking demonstrates a significant decrease in swine fecal indicators, while human fecal pollution remains unchanged. These results suggest that the reduction in ARGs was due to a substantial reduction in input of antibiotic resistant bacteria and genes from animal excreta. Our findings highlight the watershed as a living laboratory for understanding the dynamics of AMR, and for evaluating the efficacy of environmental regulations, with implications for reducing environmental risks associated with AMR on a global scale.202438925006