Metagenomics insights into microbiome and antibiotic resistance genes from free living amoeba in chlorinated wastewater effluents. - Related Documents




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773501.0000Metagenomics insights into microbiome and antibiotic resistance genes from free living amoeba in chlorinated wastewater effluents. Free living amoeba (FLA) are among the organisms commonly found in wastewater and are well-established hosts for diverse microbial communities. Despite its clinical significance, there is little knowledge on the FLA microbiome and resistome, with previous studies relying mostly on conventional approaches. In this study we comprehensively analyzed the microbiome, antibiotic resistome and virulence factors (VFs) within FLA isolated from final treated effluents of two wastewater treatment plants (WWTPs) using shotgun metagenomics. Acanthamoeba has been identified as the most common FLA, followed by Entamoeba. The bacterial diversity showed no significant difference (p > 0.05) in FLA microbiomes obtained from the two WWTPs. At phylum level, the most dominant taxa were Proteobacteria, followed by Firmicutes and Actinobacteria. The most abundant genera identified were Enterobacter followed by Citrobacter, Paenibacillus, and Cupriavidus. The latter three genera are reported here for the first time in Acanthamoeba. In total, we identified 43 types of ARG conferring resistance to cephalosporins, phenicol, streptomycin, trimethoprim, quinolones, cephalosporins, tigecycline, rifamycin, and kanamycin. Similarly, a variety of VFs in FLA metagenomes were detected which included flagellar proteins, Type IV pili twitching motility proteins (pilH and rpoN), alginate biosynthesis genes AlgI, AlgG, AlgD and AlgW and Type VI secretion system proteins and general secretion pathway proteins (tssM, tssA, tssL, tssK, tssJ, fha, tssG, tssF, tssC and tssB, gspC, gspE, gspD, gspF, gspG, gspH, gspI, gspJ, gspK, and gspM). To the best of our knowledge, this is the first study of its kind to examine both the microbiomes and resistome in FLA, as well as their potential pathogenicity in treated effluents. Additionally, this study showed that FLA can host a variety of potentially pathogenic bacteria including Paenibacillus, and Cupriavidus that had not previously been reported, indicating that their relationship may play a role in the spread and persistence of antibiotic resistant bacteria (ARBs) and antibiotic resistance genes (ARGs) as well as the evolution of novel pathogens.202438471337
327010.9989High-Throughput Shotgun Metagenomics of Microbial Footprints Uncovers a Cocktail of Noxious Antibiotic Resistance Genes in the Winam Gulf of Lake Victoria, Kenya. Background: A diverse range of pollutants, including heavy metals, agrochemicals, pharmaceutical residues, illicit drugs, personal care products, and other anthropogenic contaminants, pose a significant threat to aquatic ecosystems. The Winam Gulf of Lake Victoria, heavily impacted by surrounding human activities, faces potential contamination from these pollutants. However, studies exploring the presence of antibiotic resistance genes (ARGs) in the lake remain limited. In the current study, a shotgun metagenomics approach was employed to identify ARGs and related pathways. Genomic DNA was extracted from water and sediment samples and sequenced using the high-throughput Illumina NovaSeq platform. Additionally, phenotypic antibiotic resistance was assessed using the disk diffusion method with commonly used antibiotics. Results: The analysis of metagenomes sequences from the Gulf ecosystem and Comprehensive Antibiotic Resistance Database (CARD) revealed worrying levels of ARGs in the lake. The study reported nine ARGs from the 37 high-risk resistant gene families previously documented by the World Health Organization (WHO). Proteobacteria had the highest relative abundance of antibiotic resistance (53%), Bacteriodes (4%), Verrucomicrobia (2%), Planctomycetes Chloroflexi, Firmicutes (2%), and other unclassified bacteria (39%). Genes that target protection, replacement, change, and antibiotic-resistant efflux were listed in order of dominance. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed antibiotic resistance to beta-lactamase and vancomycin. Phenotypic resistance to vancomycin, tetracycline, sulfamethoxazole, erythromycin, trimethoprim, tetracycline, and penicillin was reported through the zone of inhibition. Conclusions: This study highlights that the Winam Gulf of Lake Victoria in Kenya harbors a diverse array of antibiotic-resistant genes, including those conferring multidrug resistance. These findings suggest that the Gulf could be serving as a reservoir for more antibiotic-resistant genes, posing potential risks to both human health and aquatic biodiversity. The insights gained from this research can guide policy development for managing antibiotic resistance in Kenya.202439741524
368920.9989Virome-associated antibiotic-resistance genes in an experimental aquaculture facility. We report the comprehensive characterization of viral and microbial communities within an aquaculture wastewater sample, by a shotgun sequencing and 16S rRNA gene profiling metagenomic approach. Caudovirales had the largest representation within the sample, with over 50% of the total taxonomic abundance, whereas approximately 30% of the total open reading frames (ORFs) identified were from eukaryotic viruses (Mimiviridae and Phycodnaviridae). Antibiotic resistance genes (ARGs) within the virome accounted for 0.85% of the total viral ORFs and showed a similar distribution both in virome and in microbiome. Among the ARGs, those encoding proteins involved in the modulation of antibiotic efflux pumps were the most abundant. Interestingly, the taxonomy of the bacterial ORFs identified in the viral metagenome did not reflect the microbial taxonomy as deduced by 16S rRNA gene profiling and shotgun metagenomic analysis. A limited number of ARGs appeared to be mobilized from bacteria to phages or vice versa, together with other bacterial genes encoding products involved in general metabolic functions, even in the absence of any antibiotic treatment within the aquaculture plant. Thus, these results confirm the presence of a complex phage-bacterial network in the aquaculture environment.201626738553
323330.9989A Metagenomic Investigation of Potential Health Risks and Element Cycling Functions of Bacteria and Viruses in Wastewater Treatment Plants. The concentration of viruses in sewage sludge is significantly higher (10-1000-fold) than that found in natural environments, posing a potential risk for human and animal health. However, the composition of these viruses and their role in the transfer of pathogenic factors, as well as their role in the carbon, nitrogen, and phosphorus cycles remain poorly understood. In this study, we employed a shotgun metagenomic approach to investigate the pathogenic bacteria and viral composition and function in two wastewater treatment plants located on a campus. Our analysis revealed the presence of 1334 amplicon sequence variants (ASVs) across six sludge samples, with 242 ASVs (41.22% of total reads) identified as pathogenic bacteria. Arcobacter was found to be the most dominant pathogen accounting for 6.79% of total reads. The virome analysis identified 613 viral genera with Aorunvirus being the most abundant genus at 41.85%. Approximately 0.66% of these viruses were associated with human and animal diseases. More than 60% of the virome consisted of lytic phages. Host prediction analysis revealed that the phages primarily infected Lactobacillus (37.11%), Streptococcus (21.11%), and Staphylococcus (7.11%). Furthermore, our investigation revealed an abundance of auxiliary metabolic genes (AMGs) involved in carbon, nitrogen, and phosphorus cycling within the virome. We also detected a total of 113 antibiotic resistance genes (ARGs), covering major classes of antibiotics across all samples analyzed. Additionally, our findings indicated the presence of virulence factors including the clpP gene accounting for approximately 4.78%, along with toxin genes such as the RecT gene representing approximately 73.48% of all detected virulence factors and toxin genes among all samples analyzed. This study expands our understanding regarding both pathogenic bacteria and viruses present within sewage sludge while providing valuable insights into their ecological functions.202438675877
323240.9988Metagenome-Based Analysis of the Microbial Community Structure and Drug-Resistance Characteristics of Livestock Feces in Anhui Province, China. We analyzed metagenome data of feces from sows at different physiological periods reared on large-scale farms in Anhui Province, China, to provide a better understanding of the microbial diversity of the sow intestinal microbiome and the structure of antibiotic-resistance genes (ARGs) and virulence genes it carries. Species annotation of the metagenome showed that in the porcine intestinal microbiome, bacteria were dominant, representing >97% of the microorganisms at each physiological period. Firmicutes and Proteobacteria dominated the bacterial community. In the porcine gut microbiome, the viral component accounted for an average of 0.65%, and the species annotation results indicated that most viruses were phages. In addition, we analyzed the microbiome for ARGs and virulence genes. Multidrug-like, MLS-like, and tetracycline-like ARGs were most abundant in all samples. Evaluation of the resistance mechanisms indicated that antibiotic inactivation was the main mechanism of action in the samples. It is noteworthy that there was a significant positive correlation between ARGs and the total microbiome. Moreover, comparative analysis with the Virulence Factor Database showed that adhesion virulence factors were most abundant.202438393105
309250.9988Potential reservoirs of antimicrobial resistance in livestock waste and treated wastewater that can be disseminated to agricultural land. Livestock manure, dairy lagoon effluent, and treated wastewater are known reservoirs of antibiotic resistance genes (ARGs), antibiotic-resistant bacteria (ARB), and virulence factor genes (VFGs), and their application to agricultural farmland could be a serious public health threat. However, their dissemination to agricultural lands and impact on important geochemical pathways such as the nitrogen (N) cycle have not been jointly explored. In this study, shotgun metagenomic sequencing and analyses were performed to examine the diversity and composition of microbial communities, ARGs, VFGs, and N cycling genes in different livestock manure/lagoon and treated wastewater collected from concentrated animal feeding operations (CAFOs) and a municipal wastewater treatment plant along the west coast of the United States. Multivariate analysis showed that diversity indices of bacterial taxa from the different microbiomes were not significantly different based on InvSimpson (P = 0.05), but differences in ARG mechanisms were observed between swine manure and other microbiome sources. Comparative resistome profiling showed that ARGs in microbiome samples belonged to four core resistance classes: aminoglycosides (40-55 %), tetracyclines (30-45 %), beta-lactam-resistance (20-35 %), macrolides (18-30 %), and >50 % of the VFGs that the 24 microbiomes harbored were phyletically affiliated with two bacteria, Bacteroidetes fragilis and Enterobacter aerogenes. Network analysis based on Spearman correlation showed co-occurrence patterns between several genes such as transporter-gene and regulator, efflux pump and involved-in-polymyxin- resistance, aminoglycoside, beta-lactam, and macrolide with VFGs and bacterial taxa such as Firmicutes, Candidatus Themoplasmatota, Actinobacteria, and Bacteroidetes. Metabolic reconstruction of metagenome-assembled genome (MAGs) analysis showed that the most prevalent drug resistance mechanisms were associated with carbapenem resistance, multidrug resistance (MDR), and efflux pump. Bacteroidales was the main taxa involved in dissimilatory nitrate reduction (DNRA) in dairy lagoon effluent. This study demonstrates that the dissemination of waste from these sources can increase the spread of ARGs, ARB, and VFGs into agricultural lands, negatively impacting both soil and human health.202336781130
327160.9988Metagenomic characterization of bacterial community and antibiotic resistance genes found in the mass transit system in Seoul, South Korea. Mass transit systems, including subways and buses, are useful environments for studying the urban microbiome, as the vast majority of populations in urban areas use public transportation. Microbial communities in urban environments include both human- and environment-associated bacteria that play roles in health and pathogen transmission. In this study, we used shotgun metagenomic sequencing to profile microbial communities sampled from various surfaces found in subway stations and bus stops within the Seoul mass transit system. The metagenomic approach and network analysis were used to investigate broad-spectrum antibiotic resistance genes (ARGs) and their co-occurrence patterns. We uncovered 598 bacterial species in 76 samples collected from various surfaces within the Seoul mass transit system. All samples were dominated by the potential human pathogen Salmonella enterica (40 %) and the human skin bacterium Cutibacterium acnes (19 %). Significantly abundant biomarkers detected in subway station samples were associated with bacteria typically found in the human oral cavity and respiratory tract, whereas biomarkers detected in bus stop samples were associated with bacteria commonly found in soil, water, and plants. Temperature and location had significant effects on microbial community structure and diversity. In total, 41 unique ARG subtypes were identified, associated with single-drug or multidrug resistance to clinically important and extensively used antibiotics, including aminoglycosides, carbapenem, glycopeptide, and sulfonamides. We revealed that Seoul subway stations and bus stops possess unique microbiomes containing potential human pathogens and ARGs. These findings provide insights for refining location-specific responses to reduce exposure to potentially causative agents of infectious diseases, improving public health.202236257123
768970.9988Discovery of Novel Antibiotic Resistance Determinants in Forest and Grassland Soil Metagenomes. Soil represents a significant reservoir of antibiotic resistance genes (ARGs), which can potentially spread across distinct ecosystems and be acquired by pathogens threatening human as well as animal health. Currently, information on the identity and diversity of these genes, enabling anticipation of possible future resistance development in clinical environments and the livestock sector, is lacking. In this study, we applied functional metagenomics to discover novel sulfonamide as well as tetracycline resistance genes in soils derived from forest and grassland. Screening of soil metagenomic libraries revealed a total of eight so far unknown ARGs. The recovered genes originate from phylogenetically diverse soil bacteria (e.g., Actinobacteria, Chloroflexi, or Proteobacteria) and encode proteins with a minimum identity of 46% to other antibiotic resistance determinants. In particular forest soil ecosystems have so far been neglected in studies focusing on antibiotic resistance. Here, we detected for the first time non-mobile dihydropteroate synthase (DHPS) genes conferring resistance to sulfonamides in forest soil with no history of exposure to these synthetic drugs. In total, three sulfonamide resistant DHPSs, differing in taxonomic origin, were discovered in beech or pine forest soil. This indicates that sulfonamide resistance naturally occurs in forest-resident soil bacterial communities. Besides forest soil-derived sulfonamide resistance proteins, we also identified a DHPS affiliated to Chloroflexi in grassland soil. This enzyme and the other recovered DHPSs confer reduced susceptibility toward sulfamethazine, which is widely used in food animal production. With respect to tetracycline resistance, four efflux proteins affiliated to the major facilitator superfamily (MFS) were identified. Noteworthy, one of these proteins also conferred reduced susceptibility toward lincomycin.201930899254
771880.9988Shotgun metagenomics offers novel insights into taxonomic compositions, metabolic pathways and antibiotic resistance genes in fish gut microbiome. Gut microbiota of freshwater carp (Labeo rohita) was investigated by shotgun metagenomics to understand its taxonomic composition and functional capabilities. With the presence of 36 phyla, 326 families and 985 genera, the fish gut microbiota was found to be quite diverse in nature. However, at the phylum level, more than three-fourths of gut microbes belonged to Proteobacteria. Very low prevalence of commonly used probiotic bacteria (Bacillus, Lactobacillus, Streptococcus, and Lactococcus) in fish gut suggested the need to search for alternative probiotics for aquaculture use. Biosynthesis pathways were found to be the most dominant (51%) followed by degradation (39%), energy metabolism (4%) and fermentation (2%). In conformity with herbivorous feeding habit of L. rohita, gut microbiome also had pathways for the degradation of cellulose, hemicellulose, chitin, pectin, starch, and other complex carbohydrates. High prevalence of Actinobacteria and antibiotic biosynthesis pathways in the fish gut microbiome indicated its potential for bioprospecting of potentially novel natural antibiotics. Fifty-one different types of antibiotic resistance genes (ARGs) belonging to 15 antimicrobial resistance (AMR) gene families and conferring resistance against 24 antibiotic types were detected in fish gut. Some of the ARGs for multi-drug resistance were also found to be located on sequences of plasmid origin. The presence of pathogenic bacteria and ARGs on plasmid sequences suggested the potential risk due to horizontal gene transfer in the confined gut environment. The role of ARGs in fish gut microbiome needs further investigations.201930604012
774190.9988Microbial diversity of a full-scale UASB reactor applied to poultry slaughterhouse wastewater treatment: integration of 16S rRNA gene amplicon and shotgun metagenomic sequencing. The 16S rRNA gene amplicon and whole-genome shotgun metagenomic (WGSM) sequencing approaches were used to investigate wide-spectrum profiles of microbial composition and metabolic diversity from a full-scale UASB reactor applied to poultry slaughterhouse wastewater treatment. The data were generated by using MiSeq 2 × 250 bp and HiSeq 2 × 150 bp Illumina sequencing platforms for 16S amplicon and WGSM sequencing, respectively. Each approach revealed a distinct microbial community profile, with Pseudomonas and Psychrobacter as predominant genus for the WGSM dataset and Clostridium and Methanosaeta for the 16S rRNA gene amplicon dataset. The virome characterization revealed the presence of two viral families with Bacteria and Archaea as host, Myoviridae, and Siphoviridae. A wide functional diversity was found with predominance of genes involved in the metabolism of acetone, butanol, and ethanol synthesis; and one-carbon metabolism (e.g., methanogenesis). Genes related to the acetotrophic methanogenesis pathways were more abundant than methylotrophic and hydrogenotrophic, corroborating the taxonomic results that showed the prevalence of the acetotrophic genus Methanosaeta. Moreover, the dataset indicated a variety of metabolic genes involved in sulfur, nitrogen, iron, and phosphorus cycles, with many genera able to act in all cycles. BLAST analysis against Antibiotic Resistance Genes Database (ARDB) revealed that microbial community contained 43 different types of antibiotic resistance genes, some of them were associated with growth chicken promotion (e.g., bacitracin, tetracycline, and polymyxin).201728229558
3184100.9988Comparative assessment of microbiome and resistome of influent and effluent of sewage treatment plant and common effluent treatment plant located in Delhi, India using shotgun approach. Antimicrobial resistance (AMR) is a significant threat that demands surveillance to identify and analyze trends of the emerging antibiotic resistance genes (ARGs) and potential microbial carriers. The influent of the wastewater treatment plants (WWTPs) reflects the microbes derived from the population and effluent being the source of dissemination of potential pathogenic microbes and AMR. The present study aimed to monitor microbial communities and antibiotic resistance genes in WWTPs employing a whole metagenome shotgun sequencing approach. The samples were collected from a sewage treatment plant (STP) and a common effluent treatment plant (CETP) in Delhi, India. The results showed the influent of STP to be rich in Bifidobacterium, Bacteroides, Escherichia, Arcobacter, and Pseudomonas residents of gut microbiota and known to cause diseases in humans and animals; whereas the CETP sample was abundant in Aeromonas, Escherichia, and Shewanella known to be involved in the degradation of different compounds. Interestingly, the effluent samples from both STPs and CETP were rich in microbial diversity, comprising organic and xenobiotic compound degrading and disease-causing bacteria, indicating the effluent being the source of dissemination of concerning bacteria to the environment. The functional profile at both sites displayed similarity with an abundance of housekeeping function genes as analyzed by Clusters of Orthologous Genes (COG), KEGG Orthology (KO), and subsystem databases. Resistome profiling by MEGARes showed the dominance of ARGs corresponding to beta-lactams having relative abundance ranging from 16% to 34% in all the metagenome datasets, followed by tetracycline (8%-16%), aminoglycosides (7%-9%), multi-drug (5%-9%), and rifampin (3%-9%). Also, AMR genes oxa, ant3-DPRIME, and rpoB, which are of clinical importance were predominantly and most prevalently present in all the samples. The presence of AMR in effluents from both types of treatment plants indicates that wastewater from both sources contributes to the spread of pathogenic bacteria and resistance genes, increasing the environmental AMR burden and therefore requires tertiary treatment before discharge. This work will facilitate further research towards the identification of suitable biomarkers for monitoring antibiotic resistance.202439232318
3269110.9988Exploring antibiotic resistance genes, mobile gene elements, and virulence gene factors in an urban freshwater samples using metagenomic analysis. Antibiotic resistance genes (ARGs) and antimicrobial resistance elements (AMR) are novel environmental contaminants that pose a significant risk to human health globally. Freshwater contains a variety of microorganisms that might affect human health; its quality must be assessed before use. However, the dynamics of mobile genetic elements (MGEs) and ARG propagation in freshwater have rarely been studied in Singapore. Therefore, this study used metagenomics to compare diversity, virulence factor composition, and ARG and MGE co-occurrence with bacterial communities in paired (n = 8) environmental freshwater samples. KneadData, FMAP, and Kraken2 were used for bioinformatics analysis and R (v4.1.1) for statistical analysis. Sequence reads with a total of 9043 species were taxonomically classified into 66 phyla, 130 classes, 261 orders, 584 families, and 2477 genera. Proteobacteria, Bacteroidetes, Actinobacteria, and Firmicutes were found the Phyla in all samples. Analysis of QIIME output by PICRUSt and ß-diversity showed unique clusters and functional microbial community structures. A total of 2961 ARGs were found that conferred resistance to multidrug, aminoglycosides, tetracyclines, elfamycins, and more. The classified ARG mechanism revealed significant distribution of virulence factors in bacterial cells. Transposes and transposon were highly correlated to ARG gene transfer. Co-occurrence network analysis showed several MGEs appear to use the same ARGs (intI and rho) and were dominant in all samples. Furthermore, ARGs are also highly correlated with bacteria like Campylobacter and Escherichia. This study enhances the understanding of antibiotic risk assessment and provides a new perspective on bacterial assembly contamination and the functional prevalence of ARGs and MGEs with antibiotic resistance bacteria. Moreover, it raises public awareness because these contaminants put people's lives at risk of acquiring bacterial infections. In addition, it can also help propose hybrid water treatment approaches.202335939194
3688120.9988Functional metagenomic characterization of antibiotic resistance genes in agricultural soils from China. Soil has been regarded as a rich source of antibiotic resistance genes (ARGs) due to the complex microbial community and diverse antibiotic-producing microbes in soil, however, little is known about the ARGs in unculturable bacteria. To investigate the diversity and distribution of ARGs in soil and assess the impact of agricultural practice on the ARGs, we screened soil metagenomic library constructed using DNA from four different agricultural soil for ARGs. We identified 45 clones conferring resistance to minocycline, tetracycline, streptomycin, gentamicin, kanamycin, amikacin, chloramphenicol and rifampicin. The similarity of identified ARGs with the closest protein in GenBank ranged from 26% to 92%, with more than 60% of identified ARGs had low similarity less than 60% at amino acid level. The identified ARGs include aminoglycoside acetyltransferase, aminoglycoside 6-adenyltransferase, ADP-ribosyl transferase, ribosome protection protein, transporters and other antibiotic resistant determinants. The identified ARGs from the soil with manure application account for approximately 70% of the total ARGs in this study, implying that manure amendment may increase the diversity of antibiotic resistance genes in soil bacteria. These results suggest that antibiotic resistance in soil remains unexplored and functional metagenomic approach is powerful in discovering novel ARGs and resistant mechanisms.201424412260
7714130.9987Functional traits and health implications of the global household drinking-water microbiome retrieved using an integrative genome-centric approach. The biological safety of drinking water plays a crucial role in public health protection. However, research on the drinking water microbiome remains in its infancy, especially little is known about the potentially pathogenic bacteria in and functional characteristics of the microbiome in household tap water that people are directly exposed to. In this study, we used a genomic-centric approach to construct a genetic catalogue of the drinking water microbiome by analysing 116 metagenomic datasets of household tap water worldwide, spanning nine countries/regions on five continents. We reconstructed 859 high-quality metagenome-assembled genomes (MAGs) spanning 27 bacterial and 2 archaeal phyla, and found that the core MAGs belonging to the phylum Proteobacteria encoded the highest metabolic functional diversity of the 33 key complete metabolic modules. In particular, we found that two core MAGs of Brevibacillus and Methylomona encoded genes for methane metabolism, which may support the growth of heterotrophic organisms observed in the oligotrophic ecosystem. Four MAGs of complete ammonia oxidation (comammox) Nitrospira were identified and functional metabolic analysis suggested these may enable mixotrophic growth and encode genes for reactive oxygen stress defence and arsenite reduction that could aid survival in the environment of oligotrophic drinking water systems. Four MAGs were annotated as potentially pathogenic bacteria (PPB) and thus represented a possible public health concern. They belonged to the genera Acinetobacter (n = 3) and Mycobacterium (n = 1), with a total relative abundance of 1.06 % in all samples. The genomes of PPB A. junii and A. ursingii were discovered to contain antibiotic resistance genes and mobile genetic elements that could contribute to antimicrobial dissemination in drinking water. Further network analysis suggested that symbiotic microbes which support the growth of pathogenic bacteria can be targets for future surveillance and removal.202438183799
4643140.9987Longitudinal metatranscriptomic sequencing of Southern California wastewater representing 16 million people from August 2020-21 reveals widespread transcription of antibiotic resistance genes. Municipal wastewater provides a representative sample of human fecal waste across a catchment area and contains a wide diversity of microbes. Sequencing wastewater samples provides information about human-associated and medically important microbial populations, and may be useful to assay disease prevalence and antimicrobial resistance (AMR). Here, we present a study in which we used untargeted metatranscriptomic sequencing on RNA extracted from 275 sewage influent samples obtained from eight wastewater treatment plants (WTPs) representing approximately 16 million people in Southern California between August 2020 - August 2021. We characterized bacterial and viral transcripts, assessed metabolic pathway activity, and identified over 2,000 AMR genes/variants across all samples. Because we did not deplete ribosomal RNA, we have a unique window into AMR carried as ribosomal mutants. We show that AMR diversity varied between WTPs (as measured through PERMANOVA, P < 0.001) and that the relative abundance of many individual AMR genes/variants increased over time (as measured with MaAsLin2, P(adj) < 0.05). Similarly, we detected transcripts mapping to human pathogenic bacteria and viruses suggesting RNA sequencing is a powerful tool for wastewater-based epidemiology and that there are geographical signatures to microbial transcription. We captured the transcription of gene pathways common to bacterial cell processes, including central carbon metabolism, nucleotide synthesis/salvage, and amino acid biosynthesis. We also posit that due to the ubiquity of many viruses and bacteria in wastewater, new biological targets for microbial water quality assessment can be developed. To the best of our knowledge, our study provides the most complete longitudinal metatranscriptomic analysis of a large population's wastewater to date and demonstrates our ability to monitor the presence and activity of microbes in complex samples. By sequencing RNA, we can track the relative abundance of expressed AMR genes/variants and metabolic pathways, increasing our understanding of AMR activity across large human populations and sewer sheds.202336455460
4555150.9987Characterization of known and novel clinically important antibiotic resistance genes and novel microbes from wastewater-impacted high Arctic fjord sediments. Arctic microbiota is enigmatic and highly underexplored. With the aim of understanding the resistome and microbiota of high-Arctic fjord sediments and the effect of wastewater discharge on sediment microbiota, we analyzed sediments from Advent fjord in Svalbard using metagenomics. We show the presence of 888 clinically relevant antibiotic resistance genes including genes coding resistance against last-resort antibiotics such as carbapenems, colistin, vancomycin, linezolid and tigecycline in the sediment microbiota. Using computational models, 478 novel β-lactamases belonging to 217 novel β-lactamase families were revealed in the sediment microbiota. Further, we identified hosts for 69 novel families and showed that these genes are widespread in the Arctic environment. We assembled 644 metagenome-assembled genomes (MAGs) from sediment metagenomes. Of these >97 % belonged to novel taxa with 89 bacterial MAGs representing seven putative novel phyla. These MAGs encoded important functions like nutrient cycling and methane metabolism etc. Our study demonstrated mixing of human associated bacteria and Arctic sediment microbiota. It provides the first comprehensive dataset of the distribution and diversity of novel microbes and β-lactamases in the wastewater-impacted high Arctic fjord sediments.202540424901
7716160.9987Metagenomic analysis fecal microbiota of dysentery-like diarrhoea in a pig farm using next-generation sequencing. Porcine enteric diseases including swine dysentery involves a wide range of possible aetiologies and seriously damages the intestine of pigs of all ages. Metagenomic next-generation sequencing is commonly used in research for detecting and analyzing pathogens. In this study, the feces of pigs from a commercial swine farm with dysentery-like diarrhea was collected and used for microbiota analysis by next-generation sequencing. While Brachyspira spp. was not detected in diarrheal pig fecal samples, indicating that the disease was not swine dysentery. The quantity of microbial population was extremely lowered, and the bacterial composition was altered with a reduction in the relative abundance of the probiotics organisms, Firmicutes and Bacteroidetes, with an increase in pathogens like Fusobacterium and Proteobacteria, in which the specific bacteria were identified at species-level. Viral pathogens, porcine circovirus type 2, porcine lymphotropic herpesviruses 1, and porcine mastadenovirus A were also detected at pretty low levels. Carbohydrate-active enzymes (CAZy) analysis indicated that the constitute of Firmicutes and Bacteroidete were also changed. Further, the Kyoto Encyclopedia of Genes and Genomes (KEGG) alignment analysis indicated that the microbiota of diarrheal pigs had a lower ability in utilizing energy sources but were enriched in multi-drug resistance pathways. Comprehensive Antibiotic Resistance Database (CARD) and Virulence Factors of Pathogenic Bacteria (VFDB) analysis indicated that genes for elfamycin and sulfonamide resistance and the iron uptake system were enriched in diarrheal pigs. This revealed potential bacterial infection and can guide antibiotic selection for treating dysentery. Overall, our data suggested that alterations in both the population and functional attributes of microbiota in diarrheal pigs with decreased probiotic and increased pathogenic microorganisms. These results will help elucidate the mechanism of dysentery-like diarrhea and the development of approaches to control the disease.202337915946
6891170.9987Feedstock-dependent antibiotic resistance gene patterns and expression profiles in industrial scale biogas plants revealed by meta-omics technology. This study investigated antimicrobial resistance in the anaerobic digesters of two industrial-scale biogas plants processing agricultural biomass and municipal wastewater sludge. A combination of deep sequencing and genome-centric workflow was implemented for metagenomic and metatranscriptomics data analysis to comprehensively examine potential antimicrobial resistance in microbial communities. Anaerobic microbes were found to harbour numerous antibiotic resistance genes (ARGs), with 58.85% of the metagenome-assembled genomes (MAGs) harbouring antibiotic resistance. A moderately positive correlation was observed between the abundance and expression of ARGs. ARGs were located primarily on bacterial chromosomes. A higher expression of resistance genes was observed on plasmids than on chromosomes. Risk index assessment suggests that most ARGs identified posed a significant risk to human health. However, potentially pathogenic bacteria showed lower ARG expression than non-pathogenic ones, indicating that anaerobic treatment is effective against pathogenic microbes. Resistomes at the gene category level were associated with various antibiotic resistance categories, including multidrug resistance, beta-lactams, glycopeptides, peptides, and macrolide-lincosamide-streptogramin (MLS). Differential expression analysis revealed specific genes associated with potential pathogenicity, emphasizing the importance of active gene expression in assessing the risks associated with ARGs.202539461216
3093180.9987Prevalence of antibiotic resistance and virulence genes in the biofilms from an aquifer recharged with stormwater. An improved understanding of the diversity and composition of microbial communities carrying antibiotic resistance genes (ARGs) and virulence genes (VGs) in aquifers recharged with stormwater is essential to comprehend potential human health risks from water reuse. A high-throughput functional gene array was used to study the prevalence of ARGs and VGs in aquifer biofilms (n = 27) taken from three boreholes over three months. Bacterial genera annotated as opportunistic pathogens such as Aeromonas, Burkholderia, Pseudomonas, Shewanella, and Vibrio were ubiquitous and abundant in all biofilms. Bacteria from clinically relevant genera, Campylobacter, Enterobacter, Klebsiella, Mycobacterium, Mycoplasma, and Salmonella were detected in biofilms. The mean travel time of stormwater from the injection well to P1 and P3 boreholes was 260 and 360 days respectively. The presence of ARGs and VGs in the biofilms from these boreholes suggest a high spatial movement of ARGs and VGs in the aquifer. The ARGs with the highest abundance were small multidrug resistance efflux pumps (SMR) and multidrug efflux (Mex) followed by β-lactamase C genes. β- lactamase C encoding genes were primarily detected in Enterobacteriaceae, Pseudomonadaceae, Bacillaceae, and Rhodobacteraceae families. The VGs encoding siderophores, including aerobactin (iro and iuc genes), followed by pilin, hemolysin, and type III secretion were ubiquitous. Canonical correspondence analysis suggested that Total Organic Carbon (TOC), Dissolved Organic Carbon (DOC), turbidity, and Fe concentration has a significant impact on the microbial community structure of bacteria carrying ARGs and VGs. Post abstraction treatment of groundwater may be prudent to improve water security and reduce potential health risks.202032798893
4539190.9987Dataset of 569 metagenome-assembled genomes from the caeca of multiple chicken breeds from commercial and backyard farming setups of Pakistan. This article focuses the recovery of prokaryotic organisms including bacteria and archaea from 9 different groups of chicken raised in different farm setups in Pakistan. The groups comprise of three different breeds (Broilers, White Layers, and Black Australorp) of chicken raised in different farming setups that include antibiotic-free control, commercial (open and controlled shed), and backyard farms. We have recovered 569 Metagenomics-Assembled Genomes (MAGs) with a completeness of ≥50 % and contamination of ≤10 %. For each MAG, functional annotations were obtained that include KEGG modules, carbohydrate active enzymes (CAZymes), peptidases, geochemical cycles, antibiotic resistance genes, stress genes, and virulence genes. Furthermore, two different sets of Single Copy Genes (SCGs) were used to construct the phylogenetic trees. Based on the reconstructed phylogeny, phylogenetic gain of each MAG is calculated to give an account of novelty.202438882194