Metagenomic approach revealed the mobility and co-occurrence of antibiotic resistomes between non-intensive aquaculture environment and human.

BACKGROUND: Aquaculture is an important food source worldwide. The extensive use of antibiotics in intensive large-scale farms has resulted in resistance development. Non-intensive aquaculture is another aquatic feeding model that is conducive to ecological protection and closely related to the natural environment. However, the transmission of resistomes in non-intensive aquaculture has not been well characterized. Moreover, the influence of aquaculture resistomes on human health needs to be further understood. Here, metagenomic approach was employed to identify the mobility of aquaculture resistomes and estimate the potential risks to human health. RESULTS: The results demonstrated that antibiotic resistance genes (ARGs) were widely present in non-intensive aquaculture systems and the multidrug type was most abundant accounting for 34%. ARGs of non-intensive aquaculture environments were mainly shaped by microbial communities accounting for 51%. Seventy-seven genera and 36 mobile genetic elements (MGEs) were significantly associated with 23 ARG types (p < 0.05) according to network analysis. Six ARGs were defined as core ARGs (top 3% most abundant with occurrence frequency > 80%) which occupied 40% of ARG abundance in fish gut samples. Seventy-one ARG-carrying contigs were identified and 75% of them carried MGEs simultaneously. The qacEdelta1 and sul1 formed a stable combination and were detected simultaneously in aquaculture environments and humans. Additionally, 475 high-quality metagenomic-assembled genomes (MAGs) were recovered and 81 MAGs carried ARGs. The multidrug and bacitracin resistance genes were the most abundant ARG types carried by MAGs. Strikingly, Fusobacterium_A (opportunistic human pathogen) carrying ARGs and MGEs were identified in both the aquaculture system and human guts, which indicated the potential risks of ARG transfer. CONCLUSIONS: The mobility and pathogenicity of aquaculture resistomes were explored by a metagenomic approach. Given the observed co-occurrence of resistomes between the aquaculture environment and human, more stringent regulation of resistomes in non-intensive aquaculture systems may be required. Video Abstract.

Is ICE hot? A genomic comparative study reveals integrative and conjugative elements as "hot" vectors for the dissemination of antibiotic resistance genes.

IMPORTANCE: Different from other extensively studied mobile genetic elements (MGEs) whose discoveries were initiated decades ago (1950s-1980s), integrative and conjugative elements (ICEs), a diverse array of more recently identified elements that were formally termed in 2002, have aroused increasing concern for their crucial contribution to the dissemination of antibiotic resistance genes (ARGs). However, the comprehensive understanding on ICEs' ARG profile across the bacterial tree of life is still blurred. Through a genomic study by comparison with two key MGEs, we, for the first time, systematically investigated the ARG profile as well as the host range of ICEs and also explored the MGE-specific potential to facilitate ARG propagation across phylogenetic barriers. These findings could serve as a theoretical foundation for risk assessment of ARGs mediated by distinct MGEs and further to optimize therapeutic strategies aimed at restraining antibiotic resistance crises.

Plasmid permissiveness of wastewater microbiomes can be predicted from 16S rRNA sequences by machine learning.

MOTIVATION: Wastewater treatment plants (WWTPs) harbor a dense and diverse microbial community. They constantly receive antimicrobial residues and resistant strains, and therefore provide conditions for horizontal gene transfer (HGT) of antimicrobial resistance (AMR) determinants. This facilitates the transmission of clinically important genes between, e.g. enteric and environmental bacteria, and vice versa. Despite the clinical importance, tools for predicting HGT remain underdeveloped. RESULTS: In this study, we examined to which extent water cycle microbial community composition, as inferred by partial 16S rRNA gene sequences, can predict plasmid permissiveness, i.e. the ability of cells to receive a plasmid through conjugation, based on data from standardized filter mating assays using fluorescent bio-reporter plasmids. We leveraged a range of machine learning models for predicting the permissiveness for each taxon in the community, representing the range of hosts a plasmid is able to transfer to, for three broad host-range resistance IncP plasmids (pKJK5, pB10, and RP4). Our results indicate that the predicted permissiveness from the best performing model (random forest) showed a moderate-to-strong average correlation of 0.49 for pB10 [95% confidence interval (CI): 0.44-0.55], 0.43 for pKJK5 (0.95% CI: 0.41-0.49), and 0.53 for RP4 (0.95% CI: 0.48-0.57) with the experimental permissiveness in the unseen test dataset. Predictive phylogenetic signals occurred despite the broad host-range nature of these plasmids. Our results provide a framework that contributes to the assessment of the risk of AMR pollution in wastewater systems. AVAILABILITY AND IMPLEMENTATION: The predictive tool is available as an application at https://github.com/DaneshMoradigaravand/PlasmidPerm.

Plasmid-mediated antibiotic resistance gene transfer under environmental stresses: Insights from laboratory-based studies.

Although clinical settings play a major role in the current global dissemination of antibiotic resistance, once antibiotic resistance bacteria and genes are released into the environment, their fate will be subject to complex ecological processes. One of the processes prevalent in microbial communities - horizontal gene transfer - can largely facilitate the dissemination of antibiotic resistance genes (ARGs) across phylogenetic and ecological boundaries. Especially, plasmid transfer has aroused increasing concern as it has been proved a significant role in promoting ARG dissemination. As a multi-step process, plasmid transfer can be influenced by various factors, among which those stresses caused by environmental pollutants are important elements affecting the plasmid mediated ARG transfer in the environment. In fact, diverse traditional and emerging pollutants are continuously entering the environment nowadays, as evidenced by the global occurrence of pollutants like metals and pharmaceuticals in aquatic and terrestrial systems. It is therefore imperative to understand to what extent and in which way the plasmid mediated ARG dissemination can be influenced by these stresses. Over the past decades, numerous research endeavours have been made to understand the plasmid mediated ARG transfer under various environmental relevant pressures. In this review, progress and challenges of studies on environmental stress regulating plasmid mediated ARG dissemination will be discussed, with specific focus on emerging pollutants like antibiotics and non-antibiotic pharmaceuticals, metals and their nanoparticles, disinfectants and disinfection by-products, as well as the emerging particulate matter like microplastics. Despite the previous efforts, we are still lacking insights into the in situ plasmid transfer under environmental stresses, which can be addressed by future studies considering environmental relevant pollution status and multi-species microbial communities. We believe that future development of standardized high-throughput screening platforms will assist in rapidly identifying which pollutants enhance plasmid transfer and also which ones may block such gene transfer processes.

Polycyclic aromatic hydrocarbon (PAH) biodegradation capacity revealed by a genome-function relationship approach.

BACKGROUND: Polycyclic aromatic hydrocarbon (PAH) contamination has been a worldwide environmental issue because of its impact on ecosystems and human health. Biodegradation plays an important role in PAH removal in natural environments. To date, many PAH-degrading strains and degradation genes have been reported. However, a comprehensive PAH-degrading gene database is still lacking, hindering a deep understanding of PAH degraders in the era of big data. Furthermore, the relationships between the PAH-catabolic genotype and phenotype remain unclear. RESULTS: Here, we established a bacterial PAH-degrading gene database and explored PAH biodegradation capability via a genome-function relationship approach. The investigation of functional genes in the experimentally verified PAH degraders indicated that genes encoding hydratase-aldolase could serve as a biomarker for preliminarily identifying potential degraders. Additionally, a genome-centric interpretation of PAH-degrading genes was performed in the public genome database, demonstrating that they were ubiquitous in Proteobacteria and Actinobacteria. Meanwhile, the global phylogenetic distribution was generally consistent with the culture-based evidence. Notably, a few strains affiliated with the genera without any previously known PAH degraders (Hyphomonas, Hoeflea, Henriciella, Saccharomonospora, Sciscionella, Tepidiphilus, and Xenophilus) also bore a complete PAH-catabolic gene cluster, implying their potential of PAH biodegradation. Moreover, a random forest analysis was applied to predict the PAH-degrading trait in the complete genome database, revealing 28 newly predicted PAH degraders, of which nine strains encoded a complete PAH-catabolic pathway. CONCLUSIONS: Our results established a comprehensive PAH-degrading gene database and a genome-function relationship approach, which revealed several potential novel PAH-degrader lineages. Importantly, this genome-centric and function-oriented approach can overcome the bottleneck of conventional cultivation-based biodegradation research and substantially expand our current knowledge on the potential degraders of environmental pollutants.

Global environmental resistome: Distinction and connectivity across diverse habitats benchmarked by metagenomic analyses.

The widely distributed antibiotic resistance genes (ARGs) were unevenly proliferated in various habitats. Great endeavors are needed to resolve the resistome features that can differentiate or connect different habitats. This study retrieved a broad spectrum of resistome profiles from 1723 metagenomes categorized into 13 habitats, encompassing industrial, urban, agricultural, and natural environments, and spanning most continents and oceans. The resistome features (ARG types, subtypes, indicator ARGs, and emerging mobilizable ARGs: mcr and tet(X)) in these habitats were benchmarked via a standardized workflow. We found that wastewater and wastewater treatment works were characterized to be reservoirs of more diverse genotypes of ARGs than any other habitats including human and livestock fecal samples, while fecal samples were with higher ARG abundance. Bacterial taxonomy composition was significantly correlated with resistome composition across most habitats. Moreover, the source-sink connectivities were disentangled by developing the resistome-based microbial attribution prediction model. Environmental surveys with standardized bioinformatic workflow proposed in this study will help comprehensively understand the transfer of ARGs in the environment, thus prioritizing the critical environments with high risks for intervention to tackle the problem of ARGs.

Roadmap to tackle antibiotic resistance in the environment under the One Health framework.

Antibiotic resistance has been recognized as a major challenge worldwide for humans. "One Health" has been recognized as a key concept for containment of antibiotic resistance. Under the framework, the role of the environment in the development of antibiotic resistance genes (ARGs) has become increasingly obvious. Despite numerous efforts, response to antibiotic resistance is considered to be inadequate, which is probably due to the lack of a clear roadmap. Here, we propose a "One Health" roadmap to combat antibiotic resistance in the environment through (1) understanding environmental resistome. The environmental gene pool has long been recognized as the single largest reservoir of both known and novel ARGs. (2) Standardizing ARG quantification. Systematic joint efforts based on standardized quantification are urgently needed to understand the true tempospatial profiles of the environmental resistome. (3) Identifying mechanisms of resistome development. Horizontal gene transfer and co-selection have been recognized as the two main mechanisms contributing to the environmental resistome. (4) Establishing a risk-assessment framework. The first critical step for large-scale cost-effective targeted ARG management in the environment is the risk assessment to identify the priority ARGs for control. (5) Formulating regulatory standards. By correlating the environmental ARG profile with public health, we may identify the indicator ARGs that can be integrated into current environmental quality standards. (6) Developing control strategies. Systematic analysis of available control technologies is required to identify the most feasible ones to curtail the spread of ARGs in the environment. The proposed roadmap under the "One Health" framework provides a guide to tackle antibiotic resistance in the environment.

The Complex Interplay Between Antibiotic Resistance and Pharmaceutical and Personal Care Products in the Environment.

Antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) are important environmental contaminants. Nonetheless, what drives the evolution, spread, and transmission of antibiotic resistance dissemination is still poorly understood. The abundance of ARB and ARGs is often elevated in human-impacted areas, especially in environments receiving fecal wastes, or in the presence of complex mixtures of chemical contaminants, such as pharmaceuticals and personal care products. Self-replication, mutation, horizontal gene transfer, and adaptation to different environmental conditions contribute to the persistence and proliferation of ARB in habitats under strong anthropogenic influence. Our review discusses the interplay between chemical contaminants and ARB and their respective genes, specifically in reference to co-occurrence, potential biostimulation, and selective pressure effects, and gives an overview of mitigation by existing man-made and natural barriers. Evidence and strategies to improve the assessment of human health risks due to environmental antibiotic resistance are also discussed. Environ Toxicol Chem 2023;00:1-16. © 2022 SETAC.

Reliable and Scalable Identification and Prioritization of Putative Cellulolytic Anaerobes With Large Genome Data.

In the era of high-throughput sequencing, genetic information that is inherently whispering hints of the microbes' functional niches is becoming easily accessible; however, properly identifying and characterizing these genetic hints to infer the microbes' functional niches remains a challenge. Regarding genome-centric interpretation on the specific functional niche of cellulose hydrolysis for anaerobes, often encountered in practice is a lack of confidence in predicting the anaerobes' real cellulolytic competency based solely on abundances of the varying carbohydrate-active enzyme modules annotated or on their taxonomy affiliation. Recognition of the synergy machineries that include but not limited to the cellulosome gene clusters is equally important as the annotation of individual carbohydrate-active modules or genes. In the interpretation of complete genomes of 2,768 microbe strains whose phenotypes have been well documented, with the incorporation of an automatic recognition of synergy among the carbohydrate active elements annotated, an explicit genotype-phenotype correlation was evidenced to be feasible for cellulolytic anaerobes, and a bioinformatic pipeline was developed accordingly. This genome-centric pipeline would categorize putative cellulolytic anaerobes into six genotype groups based on differential cellulose-hydrolyzing capacity and varying synergy mechanisms. Suggested in this genotype-phenotype correlation analysis was a finer categorization of the cellulosome gene clusters: although cellulosome complexes, by their nature, could enable the assembly of a number of carbohydrate-active units, they do not certainly guarantee the formation of the cellulose-enzyme-microbe complex or the cellulose-hydrolyzing activity of the corresponding anaerobe strains, for example, the well-known Clostridium acetobutylicum strains. Also, recognized in this genotype-phenotype correlation analysis was the genetic foundation of a previously unrecognized machinery that may mediate the microbe-cellulose adhesion, to be specific, enzymes encoded by genes harboring both the surface layer homology and cellulose-binding CBM modules. Applicability of this pipeline on scalable annotation of large genome datasets was further tested with the annotation of 7,902 reference genomes downloaded from NCBI, from which 14 genomes of putative paradigm cellulose-hydrolyzing anaerobes were identified. We believe the pipeline developed in this study would be a good add as a bioinformatic tool for genome-centric interpretation of uncultivated anaerobes, specifically on their functional niche of cellulose hydrolysis.

Absolute quantification and genome-centric analyses elucidate the dynamics of microbial populations in anaerobic digesters.

Anaerobic digestion (AD) relies on myriads of functions performed by complex microbial communities in customized settings, thus, a comprehensive investigation on the AD microbiome is central to the fine-tuned control. Most current AD microbiome studies are based on relative abundance, which hinders the interpretation of microbes' dynamics and inter-sample comparisons. Here, we developed an absolute quantification (AQ) approach that integrated cellular spike-ins with metagenomic sequencing to elucidate microbial community variations and population dynamics in four anaerobic digesters. Using this method, 253 microbes were defined as decaying populations with decay rates ranging from -0.05 to -5.85 d-1, wherein, a population from Flavobacteriaceae family decayed at the highest rates of -3.87 to -5.85 d-1 in four digesters. Meanwhile, 25 microbes demonstrated the growing trend in the AD processes with growth rates ranging from 0.11 to 1.77 d-1, and genome-centric analysis assigned some of the populations to the functional niches of hydrolysis, short-chain fatty acids metabolism, and methane generation. Additionally, we observed that the specific activity of methanogens was lower in the prolonged digestion stage, and redundancy analysis revealed that the feedstock composition and the digestion duration were the two key parameters in governing the AD microbial compositions.

An assessment of resistome and mobilome in wastewater treatment plants through temporal and spatial metagenomic analysis.

Wastewater treatment plants (WWTPs) are regarded as critical points in disseminating antibiotic resistance genes (ARGs). In particular, the discharging effluents from WWTPs generally bring downstream catchment areas exogenous ARGs and resistant bacteria. However, there lacks a sufficient assessment of the resistome and mobilome in effluents. In this study, a consecutive monthly sampling was conducted over 13 months in three Hong Kong (HK) WWTPs for metagenomic sequencing. Prevalence information of ARGs and mobile genetic elements (MGEs) was compared with counterparts in effluents from cities of North America, South America, Europe, and Asia. Moreover, a publicly accessible platform integrating the exposure ranking scheme, which was based on the global archive of ARG abundance, and a readily implementable online pipeline was developed to benefit communication in academia and government consultancy. Results demonstrated HK WWTPs were featured high ARG removal efficiency of 2.34-2.43 log reduction rate, and effluents were ranked in moderate levels of Level 2 and Level 3 in the exposure prioritizing scheme based on total ARG abundance. Moreover, absolute quantification of temporal variations of effluent resistome disclosed distinct changes over time among varied ARG types which were associated with prevalently used antibiotics, including quinolone and sulfonamide. This reinforces the need for real-time management of WWTP systems. Notably, ARGs of anthropogenic prevalence, high mobility, and potential pathogenicity were found to be present in HK effluents, drawing attention to the necessity for improved risk management. In addition, source tracking of effluent resistome and structural equation model analysis was conducted to explore the disparity in ARG abundance and diversity in different samples. The discovery of this study and the recommendation of a comprehensive exposure assessment will facilitate decision-making in resistome management in WWTPs to reduce the ARG and antibiotic resistant bacteria (ARB) contamination in the receiving environments.

An omics-based framework for assessing the health risk of antimicrobial resistance genes.

Antibiotic resistance genes (ARGs) are widespread among bacteria. However, not all ARGs pose serious threats to public health, highlighting the importance of identifying those that are high-risk. Here, we developed an 'omics-based' framework to evaluate ARG risk considering human-associated-enrichment, gene mobility, and host pathogenicity. Our framework classifies human-associated, mobile ARGs (3.6% of all ARGs) as the highest risk, which we further differentiate as 'current threats' (Rank I; 3%) - already present among pathogens - and 'future threats' (Rank II; 0.6%) - novel resistance emerging from non-pathogens. Our framework identified 73 'current threat' ARG families. Of these, 35 were among the 37 high-risk ARGs proposed by the World Health Organization and other literature; the remaining 38 were significantly enriched in hospital plasmids. By evaluating all pathogen genomes released since framework construction, we confirmed that ARGs that recently transferred into pathogens were significantly enriched in Rank II ('future threats'). Lastly, we applied the framework to gut microbiome genomes from fecal microbiota transplantation donors. We found that although ARGs were widespread (73% of genomes), only 8.9% of genomes contained high-risk ARGs. Our framework provides an easy-to-implement approach to identify current and future antimicrobial resistance threats, with potential clinical applications including reducing risk of microbiome-based interventions.

Extended-Spectrum ß-Lactamase and Carbapenemase Genes are Substantially and Sequentially Reduced during Conveyance and Treatment of Urban Sewage.

Urban wastewater systems (UWSs) are a main receptacle of excreted antibiotic resistance genes (ARGs) and their host microorganisms. However, we lack integrated and quantitative observations of the occurrence of ARGs in the UWS to characterize the sources and identify processes that contribute to their fate. We sampled the UWSs from three medium-size cities in Denmark, Spain, and the United Kingdom and quantified 70 clinically important extended-spectrum ß-lactamase and carbapenemase genes along with the mobile genetic elements and microbial communities. Results from all three countries showed that sewage-especially from hospitals-carried substantial loads of ARGs (106-107 copies per person equivalent), but these loads progressively declined along sewers and through sewage treatment plants, resulting in minimal emissions (101-104 copies per person equivalent). Removal was primarily during sewage conveyance (65 ± 36%) rather than within sewage treatment (34 ± 23%). The extended-spectrum ß-lactamase and carbapenemase genes were clustered in groups based on their persistence in the UWS compartments. The less-persistent groups were associated to putative host taxa (especially Enterobacteriaceae and Moraxellaceae), while the more persistent groups appeared horizontally transferred and correlated significantly with total cell numbers and mobile genetic elements. This documentation of a substantial ARG reduction during sewage conveyance provides opportunities for antibiotic resistance management and a caution for sewage-based antibiotic resistance surveillance.

Source tracking of antibiotic resistance genes in the environment - Challenges, progress, and prospects.

Antibiotic resistance has become a global public health concern, rendering common infections untreatable. Given the widespread occurrence, increasing attention is being turned toward environmental pathways that potentially contribute to antibiotic resistance gene (ARG) dissemination outside the clinical realm. Studies during the past decade have clearly proved the increased ARG pollution trend along with gradient of anthropogenic interference, mainly through marker-ARG detection by PCR-based approaches. However, accurate source-tracking has been always confounded by various factors in previous studies, such as autochthonous ARG level, spatiotemporal variability and environmental resistome complexity, as well as inherent method limitation. The rapidly developed metagenomics profiles ARG occurrence within the sample-wide genomic context, opening a new avenue for source tracking of environmental ARG pollution. Coupling with machine-learning classification, it has been demonstrated the potential of metagenomic ARG profiles in unambiguously assigning source contribution. Through identifying indicator ARG and recovering ARG-host genomes, metagenomics-based analysis will further increase the resolution and accuracy of source tracking. In this review, challenges and progresses in source-tracking studies on environmental ARG pollution will be discussed, with specific focus on recent metagenomics-guide approaches. We propose an integrative metagenomics-based framework, in which coordinated efforts on experimental design and metagenomic analysis will assist in realizing the ultimate goal of robust source-tracking in environmental ARG pollution.

Coexistence of Antibiotic Resistance Genes and Virulence Factors Deciphered by Large-Scale Complete Genome Analysis.

Widespread use of antibiotics has enhanced the evolution of highly resilient pathogens and poses a severe risk to human health via coselection of antibiotic resistance genes (ARGs) and virulence factors (VFs). In this study, we rigorously evaluate the abundance relationship and physical linkage between ARGs and VFs by performing a comprehensive analysis of 9,070 bacterial genomes isolated from multiple species and hosts. The coexistence of ARGs and VFs was observed in bacteria across distinct phyla, pathogenicities, and habitats, especially among human-associated pathogens. The coexistence patterns of gene elements in different habitats and pathogenicity groups were similar, presumably due to frequent gene transfer. A shorter intergenic distance between mobile genetic elements and ARGs/VFs was detected in human/animal-associated bacteria, indicating a higher transfer potential. Increased accumulation of exogenous ARGs/VFs in human pathogens highlights the importance of gene acquisition in the evolution of human commensal bacteria. Overall, the findings provide insights into the genic features of combinations of ARG-VF and expand our understanding of ARG-VF coexistence in bacteria.IMPORTANCE Antibiotic resistance has become a serious global health concern. Despite numerous case studies, a comprehensive analysis of ARG and VF coexistence in bacteria is lacking. In this study, we explore the coexistence profiles of ARGs and VFs in diverse categories of bacteria by using a high-resolution bioinformatics approach. We also provide compelling evidence of unique ARG-VF gene pairs coexisting in specific bacterial genomes and reveal the potential risk associated with the coexistence of ARGs and VFs in organisms in both clinical settings and environments.

Online searching platform for the antibiotic resistome in bacterial tree of life and global habitats.

Metagenomic analysis reveals that antibiotic-resistance genes (ARGs) are widely distributed in both human-associated and non-human-associated habitats. However, it is difficult to equally compare ARGs between samples without a standard method. Here, we constructed a comprehensive profile of the distribution of potential ARGs in bacterial tree of life and global habitats by investigating ARGs in 55 000 bacterial genomes, 16 000 bacterial plasmid sequences, 3000 bacterial integron sequences and 850 metagenomes using a standard pipeline. We found that >80% of all known ARGs are not carried by any plasmid or integron sequences. Among potential mobile ARGs, tetracycline and beta-lactam resistance genes (such as tetA, tetM and class A beta-lactamase gene) distribute in multiple pathogens across bacterial phyla, indicating their clinical relevance and importance. We showed that class 1 integrases (intI1) display a poor linear relationship with total ARGs in both non-human-associated and human-associated environments. Furthermore, both total ARGs and intI1 genes show little correlation with the degree of anthropogenicity. These observations highlight the need to differentiate ARGs of high clinical relevance. This profile is published on an online platform (ARGs-OSP, http://args-osp.herokuapp.com/) as a valuable resource for the most challenging topics in this field, i.e. the risk, evolution and emergence of ARGs.

Abundance and diversity of microbial arsenic biotransformation genes in the sludge of full-scale anaerobic digesters from a municipal wastewater treatment plant.

Arsenic (As) is a potential contaminant in sewage sludge that may affect waste treatment and limit the use of these waste materials as soil amendments. Anaerobic digestion (AD) is an important and effective process for the treatment of sewage sludge and the chemical speciation of As is particularly important in sludge AD. However, the biotransformation genes of As in sludge during AD has not been fully explored. In this study, the influent and effluent sludge of anaerobic digester in a wastewater treatment plant (WWTP) was collected to investigate the species transformations of As, the abundance and diversity of As biotransformation genes was explored by real-time PCR (qPCR) and metagenomic sequencing, separately. The results showed that arsenite [As(III)] and arsenate [As(V)] were predominant in the influent sludge, whereas the relative abundance of monomethylarsenic acid (MMA) increased by 25.7% after digestion. As biotransformation genes were highly abundant, and the As(III) S-adenosylmethionine methyltransferase (arsM) gene was the predominant which significantly increased after AD by qPCR analysis. Metagenomic analysis indicated that the diversity of the arsM-like sequences also increased significantly after AD. Most of the arsM-like sequences in all the influent and effluent sludge samples were related to Bacteroidetes and Alphaproteobacteria. Furthermore, co-occurrence network analysis indicated a strong correlation between the microbial communities and As. This study provides a direct and reliable reference on As biotransformation genes and microbial community in the AD of sludge.

Plasmids persist in a microbial community by providing fitness benefit to multiple phylotypes.

The current epidemic of antibiotic resistance has been facilitated by the wide and rapid horizontal dissemination of antibiotic resistance genes (ARGs) in microbial communities. Indeed, ARGs are often located on plasmids, which can efficiently shuttle genes across diverse taxa. While the existence conditions of plasmids have been extensively studied in a few model bacterial populations, their fate in complex bacterial communities is poorly understood. Here, we coupled plasmid transfer assays with serial growth experiments to investigate the persistence of the broad-host-range IncP-1 plasmid pKJK5 in microbial communities derived from a sewage treatment plant. The cultivation conditions combined different nutrient and oxygen levels, and were non-selective and non-conducive for liquid-phase conjugal transfer. Following initial transfer, the plasmid persisted in almost all conditions during a 10-day serial growth experiment (equivalent to 60 generations), with a transient transconjugant incidence up to 30%. By combining cell enumeration and sorting with amplicon sequencing, we mapped plasmid fitness effects across taxa of the microbial community. Unexpected plasmid fitness benefits were observed in multiple phylotypes of Aeromonas, Enterobacteriaceae, and Pseudomonas, which resulted in community-level plasmid persistence. We demonstrate, for the first time, that plasmid fitness effects across community members can be estimated in high-throughput without prior isolation. By gaining a fitness benefit when carrying plasmids, members within complex microbial communities might have a hitherto unrecognised potential to maintain plasmids for long-term community-wide access.

Distribution of arsenic and its biotransformation genes in sediments from the East China Sea.

Microbial transformation of arsenic (As) plays a key role in As biogeochemical cycling and affects the mobility, bioavailability, and toxicity of As. This study aims to investigate the accumulation of As in marine sediments at different water depths in the East China Sea and reveal the abundance and diversity of the aioA, arrA, arsC, and arsM genes through quantitative real-time polymerase chain reaction (qPCR) and high-throughput sequencing. Results showed that the As content in sediments ranged from 5.53 mg kg-1 to 17.70 mg kg-1, which decreased with water depth. Abundant As biotransformation genes with low diversity were identified in these sediments, of which arsM and arrA were the most abundant. Significant positive correlation exists between the arsM and arrA gene abundance and between arsC and aioA, indicating the co-occurrence of the As biotransformation genes in microbes in marine sediments. Metagenomics analysis revealed that arsM gene was mainly distributed in Alphaproteobacteria, Solibacteres, Deltaproteobacteria, Clostridia, and Bacilli in these sediments. Among the sediment properties, total N, total S, C/N, and TOC were important factors that shaped the abundance profile of the genes involved in As transformation. This study provides a picture of As biotransformation genes in marine sediments from the East China Sea, which may affect As transformation and the ultimate fate of As in a marine environment.