Search Results (208)

Background/Objectives: Soil microbiomes in agroecosystems are natural reservoirs of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs), creating conditions for horizontal gene transfer (HGT) to clinically relevant bacteria. Southern West Siberia—a globally significant grain-producing region—lacks metagenomic characterization of its soil resistome. This study aimed to establish the first baseline profile of resistome and mobilome composition for West Siberian agricultural soils. Methods: Twelve composite soil samples were collected from agroecosystems under seven crop types across diverse soil types in southern West Siberia (September 2022). Shotgun metagenomics was performed on an Illumina NovaSeq 6000 platform. Taxonomic profiling used Kraken2/Bracken; ARG annotation used Prokka/DeepARG (identity ≥ 70%, probability score ≥ 0.8); while MGE characterization used Platon, HMMER v3.3.2, and Prokka-based integrase annotation. Resistome load was normalized to the single-copy housekeeping gene rpoB; ARG–MGE associations were defined as co-localization within 10 kb on the same contig. Results: Microbial communities were dominated by Pseudomonadota and Bacillota, with a stable core of Streptomycetaceae, Nitrobacteraceae, and Sphingomonadaceae. Normalized resistome load (N/rpoB 2.30–5.37) indicated moderate anthropogenic pressure. Dominant ARGs included efflux pumps (emrA, drrA, tetA, bcr, fsr), target modification (lnrL), and lipid A modification (arnA) genes. Class 1 integron integrase (intI1/rpoB 0.64–1.59) was detected in all 12 samples, exceeding unity in 9 of 12. ARG–MGE co-localizations were found in 11 of 12 samples. In sample Mg_155, genes emrA–emrB and bcr (NODE_16) and arnA and lnrL (NODE_6) were each independently associated with distinct prophage IntA integrase copies within Pseudomonas contigs, documenting multiple parallel horizontal transfer events encompassing resistance to five antibiotic classes. Conclusions: This work establishes the first metagenomic baseline of resistome and mobilome for West Siberian agroecosystems. The obtained data indicate moderate anthropogenic pressure on soil microbiomes, consistent with temperate agricultural systems with limited organic fertilizer input. The detected ARG–MGE co-localizations and evidence of prophage-mediated transfer of resistance determinants beyond their natural hosts suggest that mobilization potential in the region warrants consideration in future AMR monitoring programs. Full article

The escalating crisis of antimicrobial resistance claims nearly 5 million lives annually. Resistant infections now account for 4.95 million deaths worldwide and economic losses projected to reach $300 billion by 2030. Despite this urgent threat, traditional antibiotic discovery has declined precipitously. New chemical entity approvals have fallen by over 50%, while existing therapeutics are rapidly rendered obsolete by sophisticated bacterial resistance mechanisms including extended-spectrum β-lactamases, carbapenemases, and multidrug efflux pumps. Bio-based products have historically provided humanity’s most transformative antibiotics, yet conventional discovery pipelines face insurmountable bottlenecks. A total of 99.9% of environmental microbes remain unculturable. Biosynthetic gene clusters are predominantly silent under laboratory conditions, and dereplication efforts achieve only 2 to 5% annotation rates. This review presents a comprehensive examination of how artificial intelligence (AI) is revolutionizing bio-based product-based antibacterial discovery. We analyze AI-driven genome mining tools that have identified over 170,000 biosynthetic gene clusters across bacterial genomes, deep learning architectures achieving 88.5% bioactivity prediction accuracy, and generative models delivering experimental hit rates exceeding 50%—representing 50- to 90-fold improvements over traditional screening. Through validated case studies spanning in silico prediction to in vivo efficacy, we demonstrate that AI integration is not merely accelerating discovery but fundamentally transforming our capacity to access nature’s previously inaccessible chemical diversity in the fight against antimicrobial resistance. Full article

Background/Objectives: Freshwater microalgae–bacteria consortia are increasingly utilized in wastewater treatment and biomass production. However, bacteria associated with the algal phycosphere may act as environmental reservoirs of multidrug-resistant (MDR) phenotypes and antibiotic resistance genes (ARGs), including resistance to last-resort antibiotics such as colistin. Methods: An axenic culture of the freshwater microalga Pectinodesmus pectinatus was established using a NaClO-based cleaning protocol. Three phycosphere-associated bacterial strains (Chryseobacterium sp., Pseudomonas monteilii, and Stenotrophomonas pavanii) were isolated and identified by 16S rRNA gene analysis. Antimicrobial susceptibility testing was performed using broth microdilution against 16 antibiotics. Whole-genome sequencing of the most resistant isolate, S. pavanii, was conducted using Oxford Nanopore technology, followed by genome annotation and in silico resistome analysis using CARD, AMRFinderPlus, and ResFinder. Results: Among the three isolates, S. pavanii exhibited the broadest resistance profile, including high minimum inhibitory concentrations (MICs) to multiple β-lactams and colistin (MIC ≥ 16 μg/mL). No plasmid-borne mcr genes were detected. Instead, the genome encoded multiple chromosomal determinants potentially associated with polymyxin non-susceptibility, including lipid A and lipopolysaccharide modification pathways (e.g., arn genes and eptA), outer-membrane maintenance and LPS transport systems, multidrug efflux pumps, and regulatory elements. Integration of genomic and phenotypic data suggested that the observed colistin non-susceptibility may be associated with intrinsic chromosomal determinants inferred from whole-genome analysis. Conclusions: This study demonstrates that the P. pectinatus phycosphere can harbor multidrug-resistant (MDR) bacteria, including strains exhibiting colistin non-susceptibility potentially associated with a repertoire of intrinsic chromosomal resistance mechanisms inferred from genomic analysis. Therefore, freshwater microalgae-based systems should be considered potential environmental reservoirs contributing to the dissemination of antimicrobial resistance. Full article

Pseudomonas aeruginosa is a versatile opportunistic pathogen that thrives in hostile environments by tightly regulating zinc (Zn) homeostasis. The CzcRS two-component system is pivotal for Zn resistance, primarily by activating the CzcCBA efflux pump, yet its basal activity and full regulon remain poorly defined. Here, we analyzed putative CzcR targets under zinc sufficiency (ZS) and zinc excess (ZE) conditions in P. aeruginosa PAO1 using ChIP-seq. Under ZE, we identified 32 CzcR binding sites, potentially regulating 39 genes, many of which are linked to virulence, antibiotic resistance, and stress response. Under ZS, 10 binding sites were detected, revealing distinct CzcR targets. Considering the presence of a CzcR binding motif close to the peaks summit and RNA-seq data, we identified seven and four novel CzcR-regulated genes under ZE and ZS conditions, respectively, mostly implicated in bacterial virulence. Our findings highlight that CzcR may exhibit different functionalities depending on Zn concentration: its basal activity maintains physiological robustness, while its activated form orchestrates Zn detoxification and virulence modulation. This study expands our understanding of how P. aeruginosa integrates metal sensing with clinically relevant phenotypes, highlighting CzcR as a key regulator at the intersection of metal homeostasis and pathogenicity. Full article

Background/Objectives: Glyphosate-based formulations are globally pervasive pollutants increasingly recognized as potential contributors to antimicrobial resistance (AMR) in environmental microbiomes. Although glyphosate is designed to inhibit plant 5-enolpyruvylshikimate-3-phosphate synthase, it also affects microbial metabolism, stress response, and genetic exchange. This review synthesizes the pathways through which glyphosate, its metabolite aminomethylphosphonic acid (AMPA), and commercial mixtures influence resistance-associated phenotypes and the dissemination of antibiotic resistance (ABR). Methods: A critical synthesis of the literature was conducted to evaluate the mechanistic and ecological interactions between glyphosate exposure and bacterial resistance in soil, aquatic, and host-associated microbiomes. Results: Experimental evidence showed that sublethal glyphosate exposure induced oxidative stress, altered membrane permeability, activated multidrug efflux pumps, and promoted tolerance phenotypes that could modify antibiotic susceptibility. It also enhances mutation rates and horizontal gene transfer processes associated with the emergence of resistance under controlled conditions. At the community level, glyphosate exposure is associated with microbiome restructuring and enrichment of resistance determinants, often without major shifts in overall diversity of the microbiome. These effects have been reported at environmentally relevant concentrations, although the evidence remains largely derived from laboratory and mesocosm studies. Conclusions: Glyphosate acts as both a biochemical modulator of resistance-related phenotypes and an environmental selective pressure that shapes microbial communities. Its widespread use and environmental persistence position it as a context-dependent contributor to the emergence and dissemination of AMR through interacting mechanistic and ecological pathways. Integrating AMR endpoints into pesticide risk assessments and surveillance frameworks is warranted, in addition to expanded field-based validation. Full article

Bacterial biofilms are structured microbial communities enclosed within a self-produced extracellular polymeric substance matrix composed of polysaccharides, proteins, extracellular DNA, and lipids. This matrix promotes adhesion, structural stability, and the development of heterogeneous microenvironments that restrict antimicrobial penetration and shield bacteria from host immune responses. As a result, biofilms are major contributors to chronic, recurrent, device-related, and difficult-to-treat infections, posing a major challenge for clinical management and antimicrobial stewardship. This review summarizes current understandings of biofilm biology, its clinical relevance, including the stages of biofilm development, the composition and protective roles of the matrix, and the physiological heterogeneity that arises during maturation. It also examines key mechanisms underlying biofilm tolerance and resistance, such as limited antibiotic diffusion, and sequestration, enzymatic inactivation, efflux pump upregulation, persister cell formation, and horizontal gene transfer. In addition, it highlights important clinical settings in which biofilms are implicated, including cystic fibrosis, chronic wounds, osteomyelitis, implant- or device-associated infections, and breast implant illness, in which persistent implant-associated biofilms and the resulting chronic inflammatory milieu have been hypothesized to contribute to local and systemic manifestations in a subset of patients. The review further discusses conventional and emerging approaches for biofilm detection alongwith real-time monitoring. Biofilm-associated infections remain difficult to eradicate because persistence is driven by multiple interconnected protective mechanisms. Effective management therefore requires integrated strategies that combine accurate detection with multifaceted therapies, including antibiotics alongside matrix-disrupting enzymes, quorum-sensing inhibitors, bacteriophages, metabolic reactivators, and nanotechnology-based delivery systems. Advances in multi-omics and system-level modeling will be essential for developing next-generation strategies to prevent, monitor, and treat biofilm-associated disease. Full article

Background: Reusable tourniquets are widely used across clinical settings, yet their role as reservoirs of microbial contamination and antimicrobial resistance remains poorly characterized. Methods: In this cross-sectional study, 53 polyester–elastane tourniquets were collected from an Emergency Department (SR) and Operating Theater (SBO) over a 28-day period to assess bacterial burden and resistome composition. A 180-target qPCR panel targeting antibiotic and disinfectant resistance determinants was used. qPCR analysis identified 112 distinct resistance genes across all samples, with SR tourniquets harboring significantly richer resistomes than SBO (median 34 vs. 15 genes; p < 0.001). Efflux pump- and disinfectant-associated genes were pervasive, and β-lactamase and quinolone-resistance determinants increased over time in SR samples. Results: Principal component analysis showed clear segregation of resistome profiles by clinical unit and progressive enrichment over time. These findings indicate that reusable, porous tourniquets can accumulate extensive resistance gene profiles under routine clinical use, particularly in high-contact environments. Conclusions: Enhanced decontamination strategies, development of new materials or transition to single-use alternatives may be necessary to mitigate their potential contribution to environmental antimicrobial resistance in hospitals. Full article

Background/Objectives: Municipal wastewater treatment plants (WWTPs) act as reservoirs for antibiotic-resistant bacteria, which pose a threat to global public health. In this study, we used whole-genome sequencing (WGS) to characterize antibiotic resistance genes (ARGs) and their association with mobile genetic elements (MGEs) in five multidrug-resistant (MDR) Escherichia coli isolates from dewatered sludge cake samples collected from a municipal WWTP in Cheongju, Republic of Korea. Methods: Susceptibility to nine antibiotics was evaluated via disk diffusion assay. Among the isolates exhibiting multidrug resistance (MDR) to three or more antibiotic classes, five isolates were randomly selected for whole-genome sequencing using the Illumina NovaSeqX platform. Additionally, we compared the genomic structures of five WWTP isolates with 35 environmental E. coli isolates from South Korea deposited in the NCBI pathogen database. ARGs and MGEs, including plasmids, integrons, and insertion sequences (ISs), were detected in the genome assemblies. Results: ARGs were differentially distributed between chromosomal and plasmid-derived contigs. Efflux pump-related genes were predominantly located on the chromosome across all isolates, whereas several beta-lactamase genes (e.g., bla_TEM-30 and bla_TEM-33), fluoroquinolone, and tetracycline resistance genes were localized on putative plasmid contigs. Furthermore, we characterized specific MGEs associated with these ARGs, including a class 1 integron gene cassette (dfrA17–aadA5–qacEΔ1–sul1) and an IS-mediated module (mph(A)–mrx–IS6100). Core-genome multilocus sequence typing (cgMLST) revealed that these MDR isolates represented diverse genetic lineages rather than a single clonal cluster. Conclusions: The results from this study highlight the necessity of enhanced post-treatment management of wastewater byproducts and WGS-based surveillance to mitigate the environmental spread of MDR bacteria. Full article

Rhodococcus equi is an opportunistic intracellular pathogen responsible for severe pneumonia in foals and has emerged as an important cause of infection in immunocompromised humans. The treatment of R. equi infections in foals relies mainly on the combination of macrolides and rifampin. However, the increasing incidence of multidrug-resistant (MDR) isolates has raised significant therapeutic challenges. The mechanisms underlying this resistance include mutations in target genes, activation of efflux pumps, and biofilm formation, which collectively compromise the efficacy of conventional antibiotics. Recently, growing concern over antibiotic failure has accelerated research into alternative and synergistic strategies to enhance antibacterial efficacy and reduce the development of resistance. Natural and synthetic compounds, as well as optimized antibiotic combinations, have shown promising synergistic effects by enhancing intracellular accumulation, disrupting redox homeostasis, or inhibiting efflux systems. Experimental models employing checkerboard and time-kill assays, as well as redox-sensitive biosensors, have demonstrated that certain antibiotic combinations can influence bacterial susceptibility to antibiotic exposure. Furthermore, integrating molecular tools provides valuable insight into bacterial responses to oxidative and antibiotic stress, paving the way for novel therapeutic designs. This review summarizes the current understanding of the molecular factors contributing to antimicrobial resistance in R. equi and assesses new therapeutic approaches aimed at overcoming these challenges. It highlights recent findings on strategies to improve treatment outcomes and manage antimicrobial resistance. Full article

Background/Objectives: Carbapenem-resistant Acinetobacter baumannii poses a critical threat due to its ability to acquire multiple resistance mechanisms and persist under antibiotic pressure. This study aimed to elucidate the molecular basis of imipenem resistance in clinical A. baumannii isolates by integrating phenotypic, molecular, transcriptional, and clonal analyses. Methods: Eleven A. baumannii isolates identified by MALDI-TOF MS (matrix-assisted laser desorption ionization time-of-flight mass spectrometry) were investigated. Antimicrobial susceptibility to imipenem and meropenem was assessed, followed by polymerase chain reaction (PCR) detection of Ade efflux pump, outer membrane porin, and OXA-type carbapenemase genes. Transcriptional responses to sub-inhibitory imipenem exposure were evaluated using quantitative real-time PCR, and clonal relatedness was assessed by arbitrarily primed PCR. Results: All isolates were carbapenem-resistant, with bla_OXA-23 detected in all isolates and bla_OXA-24 absent in one isolate. Transcriptional analysis revealed isolate-specific responses to imipenem exposure. Among Ade efflux pump components, only adeR exhibited expression changes, displaying either downregulation or upregulation depending on the isolate, whereas adeA, adeB, adeC, and adeS transcripts were not detected under the tested conditions. Outer membrane porin genes showed heterogeneous regulation, with ompA and carO downregulated, while some isolates showed increased expression. Expression of oprD varied among isolates, and omp33–36 transcripts were detected in a single isolate and were reduced after exposure. Clonal analysis identified nine distinct genotypes, indicating genetic diversity and the absence of clonal dominance. Conclusions: These findings highlight the multifactorial and heterogeneous nature of carbapenem resistance in A. baumannii, emphasizing the interplay between regulatory efflux mechanisms, porin modulation, and carbapenemase carriage. Full article

The expansion of tropical specialty livestock farming raises urgent concerns about airborne pathogen and antibiotic resistance dissemination. Ostrich farming, characterized by high-density stocking and feed exposure, yet their microbial ecology remain poorly characterized. This study analyzed 48 bioaerosols samples from an ostrich farm in Hainan, China, across dry and rainy seasons using 16S rRNA sequencing and metagenomics. The bacterial community were dominated by Firmicutes, Proteobacteria, and Actinobacteria, followed by Staphylococcus, Bacillus, and Acinetobacter as predominant genera, with particle size significantly shaping their structure. Large particles (>7.0 μm) carried higher species richness, while medium particles (2.1–3.3 μm) exhibited the highest diversity and evenness. Notably, small particles (0.65–1.1 μm), which can penetrate deep into the lungs, were enriched with Brevibacillus and Corynebacterium. Metagenomic analysis identified 638 antibiotic resistance genes (ARGs), dominated by efflux pump-associated determinants. The detection of clinically relevant ARGs (e.g., mcr-1 and bla_TEM) reflects the genetic potential of the airborne resistome, rather than confirmed resistance phenotypes or active horizontal gene transfer. Functional analysis revealed a strong potential for organic matter degradation, driven by abundant carbohydrate-active enzymes (CAZymes) and their corresponding CAZyme genes, as well as a nitrogen cycle dominated by assimilation and reduction pathways, while genes for nitrogen fixation and nitrification were absent. Our findings demonstrate that ostrich farming enhanced airborne microbial diversity and functional potential, facilitating the ARG dissemination and nitrogen transformation. This study provides critical insights into the ecological and health risks of bioaerosols in tropical livestock farms, informing environmental monitoring and risk management strategies. Full article

Background/Objectives: Multidrug resistance (MDR) remains a critical global public health concern, compromising the efficacy of currently available antibiotics. As the development of new antibiotics offers limited long-term solutions, alternative approaches such as efflux pump inhibition have gained attention. This study reports the development of nanostructured lipid carriers (NLCs) co-loaded with Norfloxacin (NOR) and the efflux pump inhibitor 2-amino-thiophen-6CN-Ethyl, to modulate NOR activity against resistant Staphylococcus aureus strains overexpressing efflux pump genes. Methods: NLCs were produced via the hot emulsion method followed by sonication. The formulations were characterized for encapsulation efficiency (EE%), particle size, polydispersity index (PDI), zeta potential, X-ray diffraction (XRD), infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), scanning electron microscopy (SEM), in vitro release kinetics, and stability. Antibacterial activity was evaluated against S. aureus 1199B and K2068 strains. Results: The NLC formulation containing norfloxacin and 6CN-Ethyl (NLC10NOR + 106CN) demonstrated high EE% for both compounds (99.50% for 6CN-Ethyl and 90.91% for NOR) and physicochemical stability over 60 days (particle size < 255 nm, PDI < 0.3, zeta potential < −20 mV). Structural analyses confirmed amorphization and effective encapsulation of the active constituents. Antibacterial assays showed that NLC10NOR + 106CN significantly increased NOR activity compared to the free drug and physical mixture; the effect in 1199B was notably superior to the NOR + CCCP (carbonyl cyanide m-chlorophenylhydrazone) combination. Conclusions: These findings highlight the potential of NLC-based co-delivery systems as innovative strategies to overcome bacterial resistance, particularly through efflux pump inhibition enhancing antibiotic efficacy. Full article

Background/Objectives: It is of the utmost importance to study environmental bacteria, as these microorganisms remain poorly characterized regarding their diversity, antimicrobial resistance, and impact on the global ecosystem. This knowledge gap is particularly pronounced for marine bacteria. In this study, we aimed to isolate bacteria from different marine samples and to gain insights into the environmental bacterial resistome, an aspect that remains largely neglected. Methods: Bacteria were isolated from several marine sources using two different culture media, and their identification was based on 16S rRNA gene analysis. Whole-genome sequencing was performed for selected isolates belonging to novel taxa. Antimicrobial susceptibility to seven antibiotics was evaluated using the disk diffusion method. Results: A total of 171 bacterial isolates belonging to the phyla Pseudomonadota, Bacteroidota, Planctomycetota, Actinomycetota, and Bacillota were obtained from diverse marine samples. The most abundant group belonged to the class Alphaproteobacteria. Thirty isolates represented novel taxa, comprising 16 new species and one new genus. Despite the challenges associated with determining antibiotic resistance profiles in environmental bacteria, only one isolate (1.8%) was pan-susceptible, whereas 54 (98.2%) showed resistance to at least one of the tested antibiotics. Moreover, 33 isolates exhibited a multidrug-resistant phenotype. Genome analysis of four novel taxa revealed the presence of an incomplete AdeFGH efflux pump. Conclusions: This study highlights the high bacterial diversity in marine environments, the striking prevalence of antibiotic resistance, and the major methodological challenges in studying environmental bacteria. Importantly, it emphasizes the relevance of culturomics-based approaches for uncovering hidden microbial diversity and characterizing environmental resistomes. Full article

Methicillin-resistant coagulase-negative staphylococci (MRCoNS) are a major cause of infectious diseases, owing to their ability to form biofilms and colonize community and hospital environments. MRCoNS strains were identified using biochemical tests, an MALDI-TOF MS analyzer, and PCR-based 16S rRNA gene confirmation. This study was designed to assess antibiotic resistance and biofilm-forming capacity and to determine the presence of the mecA, mecC, agrA, srtA, icaABCD, bap, fnbAB, and clfAB genes in MRCoNS isolates. From patients undergoing random screening during hospitalization in the Orthopedics Clinic in Slovakia, 28 strains of MRCoNS were identified: S. epidermidis (n = 10), S. hominis (n = 8), S. haemolyticus (n = 4), S. lugdunensis (n = 3), while S. simulans, S. pasteuri, and S. warneri were detected only once. The highest rates of resistance were observed for ampicillin, oxacillin, rifampicin, trimethoprim (100%), and erythromycin (62%). The mecA gene was detected in 12 analyzed isolates. In 12 isolates, MDR, strong efflux pump activity, and strong or moderate biofilm formation were simultaneously detected. Our findings highlight the problems posed by biofilm-forming, resistant CoNS in hospitalized patients and the importance of diagnostics, separation, rapid treatment, and proper hospital hygiene. Full article

Antimicrobial resistant (AMR) infections are a persistent public health issue causing excess death and economic impacts globally. Because AMR in clinical settings is often acquired from nonpathogenic bacteria that surround us, environmental surveillance must be better characterized. It has been well established that metals can co-select for bacterial AMR. Furthermore, recent studies have shown that compromised microbial community diversity may lead to community invasion by antibiotic resistance genes (ARGs). Widespread legacy mining has led to acid mine drainage and metal contamination of waterways and sediments throughout the western United States, potentially compromising microbial community diversity while simultaneously selecting for AMR bacteria. Our study objectives were to survey metal contaminated sediments from the Bonita Peak Mining District (BPMD) in southwestern Colorado, USA, compared to sites downstream in Durango, CO for bacterial and ARG diversity. Sediment bacteria were characterized using 16S rRNA Ilumina and metagenomic sequencing. We found that overall, bacterial diversity was lower in metal-contaminated, acidic sites (p = 0.04). Metagenomic sequencing revealed 31 different ARGs, with those encoding for efflux pumps (mex and spe gene families) substantially more prevalent in the BPMD sites, elucidating a specific AMR marker fingerprint from the high metal concentration sediments. Raising awareness and providing antimicrobial tracking techniques to resource limited communities could help provide information needed for better antibiotic use recommendations and environmental monitoring. Full article