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335100.9922Quantification of the mobility potential of antibiotic resistance genes through multiplexed ddPCR linkage analysis. There is a clear need for global monitoring initiatives to evaluate the risks of antibiotic resistance genes (ARGs) towards human health. Therefore, not only ARG abundances within a given environment, but also their potential mobility, hence their ability to spread to human pathogenic bacteria needs to be quantified. We developed a novel, sequencing-independent method for assessing the linkage of an ARG to a mobile genetic element by statistical analysis of multiplexed droplet digital PCR (ddPCR) carried out on environmental DNA sheared into defined, short fragments. This allows quantifying the physical linkage between specific ARGs and mobile genetic elements, here demonstrated for the sulfonamide ARG sul1 and the Class 1 integron integrase gene intI1. The method's efficiency is demonstrated using mixtures of model DNA fragments with either linked and unlinked target genes: Linkage of the two target genes can be accurately quantified based on high correlation coefficients between observed and expected values (R2) as well as low mean absolute errors (MAE) for both target genes, sul1 (R2 = 0.9997, MAE = 0.71%, n = 24) and intI1 (R2 = 0.9991, MAE = 1.14%, n = 24). Furthermore, we demonstrate that adjusting the fragmentation length of DNA during shearing allows controlling rates of false positives and false negative detection of linkage. The presented method allows rapidly obtaining reliable results in a labor- and cost-efficient manner.202336941120
908310.9920ARGNet: using deep neural networks for robust identification and classification of antibiotic resistance genes from sequences. BACKGROUND: Emergence of antibiotic resistance in bacteria is an important threat to global health. Antibiotic resistance genes (ARGs) are some of the key components to define bacterial resistance and their spread in different environments. Identification of ARGs, particularly from high-throughput sequencing data of the specimens, is the state-of-the-art method for comprehensively monitoring their spread and evolution. Current computational methods to identify ARGs mainly rely on alignment-based sequence similarities with known ARGs. Such approaches are limited by choice of reference databases and may potentially miss novel ARGs. The similarity thresholds are usually simple and could not accommodate variations across different gene families and regions. It is also difficult to scale up when sequence data are increasing. RESULTS: In this study, we developed ARGNet, a deep neural network that incorporates an unsupervised learning autoencoder model to identify ARGs and a multiclass classification convolutional neural network to classify ARGs that do not depend on sequence alignment. This approach enables a more efficient discovery of both known and novel ARGs. ARGNet accepts both amino acid and nucleotide sequences of variable lengths, from partial (30-50 aa; 100-150 nt) sequences to full-length protein or genes, allowing its application in both target sequencing and metagenomic sequencing. Our performance evaluation showed that ARGNet outperformed other deep learning models including DeepARG and HMD-ARG in most of the application scenarios especially quasi-negative test and the analysis of prediction consistency with phylogenetic tree. ARGNet has a reduced inference runtime by up to 57% relative to DeepARG. CONCLUSIONS: ARGNet is flexible, efficient, and accurate at predicting a broad range of ARGs from the sequencing data. ARGNet is freely available at https://github.com/id-bioinfo/ARGNet , with an online service provided at https://ARGNet.hku.hk . Video Abstract.202438725076
780920.9919Mitigating Antibiotic Resistance Genes in Wastewater by Sequential Treatment with Novel Nanomaterials. Wastewater (WW) has been widely recognized as the major sink of a variety of emerging pathogens (EPs), antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs), which may disseminate and impact wider environments. Improving and maximizing WW treatment efficiency to remove these microbial hazards is fundamentally imperative. Despite a variety of physical, biological and chemical treatment technologies, the efficiency of ARG removal is still far from satisfactory. Within our recently accomplished M-ERA.NET project, novel functionalized nanomaterials, i.e., molecularly imprinted polymer (MIP) films and quaternary ammonium salt (QAS) modified kaolin microparticles, were developed and demonstrated to have significant EP removal effectiveness on both Gram-positive bacteria (GPB) and Gram-negative bacteria (GNB) from WW. As a continuation of this project, we took the further step of exploring their ARG mitigation potential. Strikingly, by applying MIP and QAS functionalized kaolin microparticles in tandem, the ARGs prevalent in wastewater treatment plants (WWTPs), e.g., blaCTXM, ermB and qnrS, can be drastically reduced by 2.7, 3.9 and 4.9 log (copies/100 mL), respectively, whereas sul1, tetO and mecA can be eliminated below their detection limits. In terms of class I integron-integrase I (intI1), a mobile genetic element (MGE) for horizontal gene transfer (HGT), 4.3 log (copies/100 mL) reduction was achieved. Overall, the novel nanomaterials exhibit outstanding performance on attenuating ARGs in WW, being superior to their control references. This finding provides additional merit to the application of developed nanomaterials for WW purification towards ARG elimination, in addition to the proven bactericidal effect.202134063382
784930.9917Efficient removal of antibiotic-resistant bacteria and intracellular antibiotic resistance genes by heterogeneous activation of peroxymonosulfate on hierarchical macro-mesoporous Co(3)O(4)-SiO(2) with enhanced photogenerated charges. Antibiotic resistance genes (ARGs) and their host antibiotic-resistant bacteria (ARB) are widely detected in the environment and pose a threat to human health. Traditional disinfection in water treatment plants cannot effectively remove ARGs and ARB. This study explored the potential of a heterogeneous photo-Fenton-like process utilizing a hierarchical macro-mesoporous Co(3)O(4)-SiO(2) (MM CS) catalyst for activation of peroxymonosulfate (PMS) to inactivate ARB and degrade the intracellular ARGs. A typical gram-negative antibiotic-resistant bacteria called Pseudomonas sp. HLS-6 was used as a model ARB. A completed inactivation of ARB at ∼10(7) CFU/mL was achieved in 30 s, and an efficient removal rate of more than 4.0 log for specific ARGs (sul1 and intI1) was achieved within 60 min by the MM CS-based heterogeneous photo-Fenton-like process under visible light and neutral pH conditions. Mechanism investigation revealed that •O(2)(-) and (1)O(2) were the vital reactive species for ARB inactivation and ARG degradation. The formation and transformation of the active species were proposed. Furthermore, the hierarchical macro-mesoporous structure of MM CS provided excellent optical and photoelectrochemical properties that promoted the cycle of Co(3+)/Co(2+) and the effective utilization of PMS. This process was validated to be effective in various water matrices, including deionized water, underground water, source water, and secondary effluent wastewater. Collectively, this work demonstrated that the MM CS-based heterogeneous photo-Fenton-like process is a promising technology for controlling the spread of antibiotic resistance in aquatic environments.202235149504
785640.9916Boosting Low-Dose Ferrate(VI) Activation by Layered FeOCl for the Efficient Removal of Antibiotic-Resistant Bacteria and Antibiotic Resistance Genes via Enhancing Fe(IV)/Fe(V) Generation. Antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) in aquatic environments pose threats to ecosystem safety and human health, which could not be efficiently removed by conventional disinfection techniques. Herein, layered FeOCl with coordinatively unsaturated Fe sites were fabricated and used to activate Fe(VI) for the efficient ARB/ARG removal in the present study. We found that highly reactive Fe(IV)/Fe(V) intermediates were generated in the FeOCl/Fe(VI) system, rapidly disinfecting 1 × 10(7) CFU mL(-1) ARB to below the limit of detection within only 6 min. Via the combination of in situ characterization and theoretical calculations, we revealed that Fe(VI) was preferentially adsorbed onto Fe sites on the (010) plane of FeOCl and subsequently activated to produce reactive Fe(IV)/Fe(V) through direct electron transfer. Meanwhile, O(2)(•-) generated from O(2) activation on the FeOCl surface enhanced Fe(VI) conversion to Fe(IV)/Fe(V). During the disinfection process, intracellular/extracellular ARGs and DNA bases were simultaneously degraded, inhibiting the potential horizontal gene transfer process. The FeOCl/Fe(VI) system could effectively disinfect ARB under complex water matrices and in real water samples including tap water, lake water, and groundwater. When integrated into a continuous-flow reactor, the FeOCl/Fe(VI) system with excellent stability successively disinfected ARB. Overall, the FeOCl/Fe(VI) system showed great promise for eliminating ARB/ARGs from water.202540739812
786850.9916A double-quenching paperclip ECL biosensing platform for ultrasensitive detection of antibiotic resistance genes (mecA) based on Ti(3)C(2) MXene-Au NPs as a coreactant accelerator. The global spread of environmental biological pollutants, such as antibiotic-resistant bacteria and their antibiotic resistance genes (ARGs), has emerged as a critical public health concern. It is imperative to address this pressing issue due to its potential implications for public health. Herein, a DNA paperclip probe with double-quenching function of target cyclic cleavage was proposed, and an electrochemiluminescence (ECL) biosensing platform was constructed using Ti(3)C(2) MXene in-situ reduction growth of Au NPs (TCM-Au) as a coreactant accelerator, and applied to the sensitive detection of ARGs. Thanks to the excellent catalytic performance, large surface area and Au-S affinity of TCM-Au, the ECL performance of CdS QDs have been significantly improved. By cleverly utilizing the negative charge of the paperclip nucleic acid probe and its modification group, double-quenching of the ECL signal was achieved. This innovative approach, combined with target cyclic amplification, facilitated specific and sensitive detection of the mecA gene. This biosensing platform manifested highly selective and sensitive determination of mecA genes in the range of 10 fM to 100 nM and a low detection limit of 2.7 fM. The credible detectability and anti-interference were demonstrated in Yangtze river and Aeration tank outlet, indicating its promising application toward pollution monitoring of ARGs.202337666010
780560.9916Elimination of antibiotic-resistance bacterium and its associated/dissociative bla(TEM-1) and aac(3)-II antibiotic-resistance genes in aqueous system via photoelectrocatalytic process. The ubiquity of antibiotic-resistance bacteria (ARB) and antibiotic-resistance genes (ARGs) in various environmental matrices is a potential threat to human and ecological health. Therefore, the inactivation of ARB E. coli S1-23 and the elimination of its associated ARGs, bla(TEM-1) and aac(3)-II, were investigated using the photoelectrocatalytic (PEC) process. Results indicate that the ARB E. coli S1-23 (1 × 10(8) cfu mL(-1)) and its ARGs (extracellular and intracellular) could be fully inactivated within 10 and 16 h PEC treatment, respectively. In contrast, photocatalytic (PC) and electrochemical (EC) treatments displayed no obvious effect; however, ARG-containing DNA extracted from E. coli S1-23, which was used as a model for dissociative naked ARGs, could be completely decomposed within a few minutes through these three treatments. Further analyses, including PCR, AFM and HPLC, proved that the structural integrity and surface topography of naked ARGs are damaged during treatment and can be completely eliminated. Furthermore, there is no generation of cytosine, guanine, adenine or thymine intermediates during the PEC, PC, and EC treatments. This study is the first report to propose the PEC treatment as a promising method for complete decomposition of ARB and ARGs in aqueous systems.201728863344
844870.9916Genome-Wide Association Analysis for Resistance to Coniothyrium glycines Causing Red Leaf Blotch Disease in Soybean. Soybean is a high oil and protein-rich legume with several production constraints. Globally, several fungi, viruses, nematodes, and bacteria cause significant yield losses in soybean. Coniothyrium glycines (CG), the causal pathogen for red leaf blotch disease, is the least researched and causes severe damage to soybean. The identification of resistant soybean genotypes and mapping of genomic regions associated with resistance to CG is critical for developing improved cultivars for sustainable soybean production. This study used single nucleotide polymorphism (SNP) markers generated from a Diversity Arrays Technology (DArT) platform to conduct a genome-wide association (GWAS) analysis of resistance to CG using 279 soybean genotypes grown in three environments. A total of 6395 SNPs was used to perform the GWAS applying a multilocus model Fixed and random model Circulating Probability Unification (FarmCPU) with correction of the population structure and a statistical test p-value threshold of 5%. A total of 19 significant marker-trait associations for resistance to CG were identified on chromosomes 1, 5, 6, 9, 10, 12, 13, 15, 16, 17, 19, and 20. Approximately 113 putative genes associated with significant markers for resistance to red leaf blotch disease were identified across soybean genome. Positional candidate genes associated with significant SNP loci-encoding proteins involved in plant defense responses and that could be associated with soybean defenses against CG infection were identified. The results of this study provide valuable insight for further dissection of the genetic architecture of resistance to CG in soybean. They also highlight SNP variants and genes useful for genomics-informed selection decisions in the breeding process for improving resistance traits in soybean.202337372451
781380.9916A framework predicting removal efficacy of antibiotic resistance genes during disinfection processes with machine learning. Disinfection has been applied widely for the removal of antibiotic resistance genes (ARGs) to curb the spread of antibiotic resistance. Quantitative polymerase chain reaction (qPCR) is the most used method to quantify the damage of DNA thus calculating the ARG degradation during disinfection but suffers the deviation due to the limitation of amplicon length. In contrast, transformation assay more accurately measures ARG deactivation based on expression of disinfected ARG in the receiving bacteria but is typically laborious and material-intensive. This work applied machine learning (ML) to develop a framework by using qPCR results as a proxy to estimate the transformation assay measurements during disinfection with chlorine (FAC), ultraviolet (UV(254)), ozone (O(3)), and hydrogen peroxide/ultraviolet (UV/H(2)O(2)) for multiple kinds of ARGs. ARG degradation rates and deactivation rates were well predicted with the optimal correlation coefficient (R(2)) of all test sets > 0.926 and > 0.871, respectively. Besides, by concatenating the ARG degradation and deactivation predictive models, ARG removal efficiency under given disinfection conditions was directly predicted as the loss of transformation activity with R(2) > 0.828. Furthermore, an online platform was built to provide users with access to the developed ML models for rapid and accurate evaluation of ARG removal efficiency.202540179779
973790.9916Zinc Finger Nuclease: A New Approach to Overcome Beta-Lactam Antibiotic Resistance. BACKGROUND: The evolution of antibiotic-resistant bacteria (ARB) and antibiotic-resistance genes (ARGs) has been accelerated recently by the indiscriminate application of antibiotics. Antibiotic resistance has challenged the success of medical interventions and therefore is considered a hazardous threat to human health. OBJECTIVES: The present study aimed to describe the use of zinc finger nuclease (ZFN) technology to target and disrupt a plasmid-encoded β-lactamase, which prevents horizontal gene transfer-mediated evolution of ARBs. MATERIALS AND METHODS: An engineered ZFN was designed to target a specific sequence in the ampicillin resistance gene (amp(R)) of the pTZ57R plasmid. The Escherichia coli bacteria already contained the pZFN kanamycin-resistant (kana(R)) plasmid as the case or the pP15A, kana(R) empty vector as the control, were transformed with the pTZ57R; the ability of the designed ZFN to disrupt the β-lactamase gene was evaluated with the subsequent disturbed ability of the bacteria to grow on ampicillin (amp) and ampicillin-kanamycin (amp-kana)-containing media. The effect of mild hypothermia on the ZFN gene targeting efficiency was also evaluated. RESULTS: The growth of bacteria in the case group on the amp and amp-kana-containing media was significantly lower compared with the control group at 37°C (P < 0.001). Despite being more efficient in hypothermic conditions at 30°C (P < 0.001), there were no significant associations between the incubation temperature and the ZFN gene targeting efficiency. CONCLUSIONS: Our findings revealed that the ZFN technology could be employed to overcome ampicillin resistance by the targeted disruption of the ampicillin resistance gene, which leads to inactivation of β-lactam synthesis. Therefore, ZFN technology could be engaged to decrease the antibiotic resistance issue with the construction of a ZFN archive against different ARGs. To tackle the resistance issue at the environmental level, recombinant phages expressing ZFNs against different ARGs could be constructed and released into both hospital and urban wastewater systems.201627099691
7494100.9915DNA phosphorothioate modification facilitates the dissemination of mcr-1 and bla(NDM-1) in drinking water supply systems. The mechanism driving the dissemination of antibiotic resistance genes (ARGs) in drinking water supply systems (DWSSs) with multiple barriers remains poorly understood despite several recent efforts. Phosphorothioate (PT) modifications, governed by dndABCDE genes, occur naturally in various bacteria and involve the incorporation of sulfur into the DNA backbone. PT is regarded as a mild antioxidant in vivo and is known to provide protection against bacterial genomes. We combined quantitative polymerase chain reaction, metagenomic, and network analyses for the water treatment process and laboratory-scale experiments for chlorine treatment using model strains to determine if DNA PT modification occurred in DWSS and facilitated the dissemination of mobilized colistin resistance-1 (mcr-1) and New Delhi metallo-β-lactamase-1 (bla(NDM-1)) in DWSS. Our results indicated that the relative abundance of dndB increased in the effluent, compared with the influent, in the water treatment plants. Presence of dndB copies had a positive correlation with the concentration of chloramine disinfectant. Network analysis revealed Bdellovibrio as a potential host for MCR genes, NDM genes, and dndB in the DWSS. E. coli DH10B (Wild-type with the dndABCDE gene cluster and ΔdndB) model strains were used to investigate resistance to chlorine treatment at the concentration range of 0.5-3 mg/L. The resistance of the wild-type strain increased with increasing concentration of chlorine. DNA PT modification protected MCR- and NDM-carrying bacteria from chloramine disinfection during the water treatment process. The higher relative abundance of ARGs in the effluent of the water treatment plants may be due to the resistance of DNA PT modification to chloramine disinfection, thereby causing the enrichment of genera carrying MCR, NDM, and dndB. This study provides a new understanding on the mechanism of ARG dissemination in DWSS, which will help to improve the performance of drinking water treatment to control the risk associated with antibiotic-resistant bacteria.202133162214
4773110.9915Draft genome analysis for Enterobacter kobei, a promising lead bioremediation bacterium. Lead pollution of the environment poses a major global threat to the ecosystem. Bacterial bioremediation offers a promising alternative to traditional methods for removing these pollutants, that are often hindered by various limitations. Our research focused on isolating lead-resistant bacteria from industrial wastewater generated by heavily lead-containing industries. Eight lead-resistant strains were successfully isolated, and subsequently identified through molecular analysis. Among these, Enterobacter kobei FACU6 emerged as a particularly promising candidate, demonstrating an efficient lead removal rate of 83.4% and a remarkable lead absorption capacity of 571.9 mg/g dry weight. Furthermore, E. kobei FACU6 displayed a remarkable a maximum tolerance concentration (MTC) for lead reaching 3,000 mg/L. To further investigate the morphological changes in E. kobei FACU6 in response to lead exposure, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were employed. These analyses revealed significant lead adsorption and intracellular accumulation in treated bacteria in contrast to the control bacterium. Whole-genome sequencing was performed to gain deeper insights into E. kobei's lead resistance mechanisms. Structural annotation revealed a genome size of 4,856,454 bp, with a G + C content of 55.06%. The genome encodes 4,655 coding sequences (CDS), 75 tRNA genes, and 4 rRNA genes. Notably, genes associated with heavy metal resistance and their corresponding regulatory elements were identified within the genome. Furthermore, the expression levels of four specific heavy metal resistance genes were evaluated. Our findings revealed a statistically significant upregulation in gene expression under specific environmental conditions, including pH 7, temperature of 30°C, and high concentrations of heavy metals. The outstanding potential of E. kobei FACU6 as a source of diverse genes related to heavy metal resistance and plant growth promotion makes it a valuable candidate for developing safe and effective strategies for heavy metal disposal.202338260751
7710120.9915Reduced Antibiotic Resistance in the Rhizosphere of Lupinus albus in Mercury-Contaminated Soil Mediated by the Addition of PGPB. The emergence of antibiotic resistance (AR) poses a threat to the "One Health" approach. Likewise, mercury (Hg) pollution is a serious environmental and public health problem. Its ability to biomagnify through trophic levels induces numerous pathologies in humans. As well, it is known that Hg-resistance genes and AR genes are co-selected. The use of plant-growth-promoting bacteria (PGPB) can improve plant adaptation, decontamination of toxic compounds and control of AR dispersal. The cenoantibiogram, a technique that allows estimating the minimum inhibitory concentration (MIC) of a microbial community, has been postulated as a tool to effectively evaluate the evolution of a soil. The present study uses the metagenomics of 16S rRNA gene amplicons to understand the distribution of the microbial soil community prior to bacterial inoculation, and the cenoantibiogram technique to evaluate the ability of four PGPB and their consortia to minimize antibiotic resistance in the rhizosphere of Lupinus albus var. Orden Dorado grown in Hg-contaminated soils. Results showed that the addition of A1 strain (Brevibacterium frigoritolerans) and its consortia with A2, B1 and B2 strains reduced the edaphic community´s MIC against cephalosporins, ertapenem and tigecycline. The metagenomic study revealed that the high MIC of non-inoculated soils could be explained by the bacteria which belong to the detected taxa,. showing a high prevalence of Proteobacteria, Cyanobacteria and Actinobacteria.202337372086
7491130.9914Application of nematicide avermectin enriched antibiotic-resistant bacteria and antibiotic resistance genes in farmland soil. The extensive use of antibiotics in medicine and agriculture has resulted in the accumulation of antibiotic-resistant microorganisms and antibiotic resistance genes (ARGs) in environments, which threaten human health and contaminate environment. Nematicide avermectin is widely applied to control root-knot nematodes. The effect of five-years application of avermectin on rhizosphere microbiome and resistome of sick tobacco plants in farmland were investigated in present study. The environmental risks of avermectin was assessed adequately. Metagenomic method was used to analyze antibiotic-resistant bacteria and antibiotic resistance genes in the avermectin-treated soil. The abundance and distribution of antibiotic-resistant bacteria and their antibiotic resistance genes were affected by avermectin application. The antibiotic resistant Proteobacteria occupied the highest percentage (36%) in rhizosphere soil and carried 530 ARGs. Opportunistic human pathogens carrying antibiotic resistance genes were enriched in the avermectin-treated soil. Avermectin application increased the counts of many types of antibiotic resistance genes. The relative abundances of genes adeF, BahA, fusH, ileS, and tlrB in the avermectin-treated soil were significantly greater than in the untreated control soil. Different resistance mechanisms were revealed in the avermectin-treated soil. The efflux of antibiotic (670 ARGs), inactivation of antibiotic (475 ARGs), and alteration of antibiotic target (267 ARGs) were the main resistance mechanisms. Rigid control the avermectin dose and use frequency and other pesticides can decrease soil antibiotic resistance genes and protect agricultural products' safety and public health. Overall, application of nematicide avermectin enriched antibiotic-resistant bacteria and antibiotic resistance genes in farmland soil, which should be on the alert for environment protection.202337003554
7869140.9914Nano-CeO(2) activates physical and chemical defenses of garlic (Allium sativum L.) for reducing antibiotic resistance genes in plant endosphere. The transmission of manure- and wastewater-borne antibiotic-resistant bacteria (ARB) to plants contributes to the proliferation of antimicrobial resistance in agriculture, necessitating effective strategies for preventing the spread of antibiotic resistance genes (ARGs) from ARB in the environment to humans. Nanomaterials are potential candidates for efficiently controlling the dissemination of ARGs. The present study investigated the abundance of ARGs in hydroponically grown garlic (Allium sativum L.) following nano-CeO(2) (nCeO(2)) application. Specifically, root exposure to nCeO(2) (1, 2.5, 5, 10 mg L(-1), 18 days) reduced ARG abundance in the endosphere of bulbs and leaves. The accumulation of ARGs (cat, tet, and aph(3')-Ia) in garlic bulbs decreased by 24.2-32.5 % after nCeO(2) exposure at 10 mg L(-1). Notably, the lignification extent of garlic stem-disc was enhanced by 10 mg L(-1) nCeO(2), thereby accelerating the formation of an apoplastic barrier to impede the upward transfer of ARG-harboring bacteria to garlic bulbs. Besides, nCeO(2) upregulated the gene expression related to alliin biosynthesis and increased allicin content by 15.9-16.2 %, promoting a potent antimicrobial defense for reducing ARG-harboring bacteria. The potential exposure risks associated with ARGs and Ce were evaluated according to the estimated daily intake (EDI). The EDI of ARGs exhibited a decrease exceeding 95 %, while the EDI of Ce remained below the estimated oral reference dose. Consequently, through stimulating physical and chemical defenses, nCeO(2) contributed to a reduced EDI of ARGs and Ce, highlighting its potential for controlling ARGs in plant endosphere within the framework of nano-enabled agrotechnology.202438570269
7744150.9914Dynamics and removal mechanisms of antibiotic and antibiotic resistance genes during the fermentation process of spectinomycin mycelial dregs: An integrated meta-omics study. Antibiotic mycelial dregs (AMDs) have been listed as industrial hazardous wastes. With the aim of reducing the environmental risk, the integrated-omics and qPCR approaches were used to reveal the dynamics and removal mechanisms of antibiotic and antibiotic resistance genes (ARGs) during the fermentation of different spectinomycin mycelial dregs (SMDs). The results showed that the removal efficiency of antibiotic in the fermentation of high moisture SMDs reached up to 98%. The high abundance of aadA1 gene encoded by Streptomyces, Lactobacillus, and Pseudomonas was associated with the efficient degradation of spectinomycin, and the inactivating enzymes secreted by degradative bacteria were identified. Furthermore, the dominant microbiota was impacted by moisture content significantly under high temperature environments. In the fermentation of low moisture SMDs, Saccharopolyspora was the dominant microbiota which secreted S8 endopeptidase, M14, M15, S10, S13 carboxypeptidases, M1, M28, S15 aminopeptidases, and antioxidant enzymes, while in the fermentation of high moisture SMDs, Bacillus and Cerasibacillus were dominant genera which mainly secreted S8 endopeptidase and antioxidant enzymes. The abundance of ARGs and mobile genetic elements decreased significantly at thermophilic phase, with maximum drops of 93.7% and 99.9%, respectively. Maintaining moisture content below 30% at the end phase could prevent the transmission of ARGs effectively.202234396972
7319160.9914Comprehensive Study of Antibiotics and Antibiotic Resistance Genes in Wastewater and Impacted Mediterranean Water Environments. Background: The spread of antimicrobial resistance is a central public health problem. Wastewater treatment plants and impacted environments are well-known hotspots for antibiotic resistance. However, there is still limited knowledge regarding where antibiotic resistance genes (ARGs) acquire mobility. Method: In this study, we aimed to gather evidence on the seasonal patterns of ARG spread in two Mediterranean areas from NE and E of Spain (Ebro River and Ebro Delta, and Xúquer River and Albufera de València), correlating ARG presence, with special focus on the faecal bacteria Escherichia coli, with antibiotic residues and environmental conditions. The analytical methodology employed was based on a suspect screening approach, while a novel prioritisation approach for antibiotics was proposed to identify those areas more susceptible to the spread of ARG. Results: Our findings demonstrate that ARG levels in wastewater were similar across different seasons, although a greater diversity of ARGs was recorded in summer. We hypothesise that horizontal gene transfer among aquatic bacterial populations during the northeastern Mediterranean summer, when temperatures reach approximately 35~40 °C, could be a key driver of ARG dissemination. By contrast, the highest concentrations of antibiotics in winter samples, with temperatures around 5~10 °C, may promote the spread of microbial resistance. Conclusions: Our key findings highlight that water temperature and sunlight irradiation are crucial factors influencing antibiotic levels and microbial abundance, requiring further investigation in future studies.202540298490
3280170.9914Optimization of five qPCR protocols toward the detection and the quantification of antimicrobial resistance genes in environmental samples. Here, we describe the optimization and validation of five quantitative PCR (qPCR) assays by employing the SYBRGreen chemistry paired with melting curve analysis to detect and quantify clinically relevant antimicrobial resistance genes (ARGs) (i.e. ermB, bla(CTXM1-like), bla(CMY-2), qnrA and qnrS) from environmental samples (i.e. soil and manure). These five protocols accurately detected and quantified the aforementioned ARGs in complex environmental matrices and represent useful tools for both diagnostic and monitoring activities of resistant bacteria and ARGs into the environment.202134754761
9739180.9914Au-Fe(3)O(4) nanozyme coupled with CRISPR-Cas12a for sensitive and visual antibiotic resistance diagnosing. The accumulation and spread of antibiotic resistance bacteria (ARB) in the environment may accelerate the formation of superbugs and seriously threaten the health of all living beings. The timeliness and accurate diagnosing of antibiotic resistance is essential to controlling the propagation of superbugs in the environment and formulating effective public health management programs. Herein, we developed a speedy, sensitive, accurate, and user-friendly colorimetric assay for antibiotic resistance, via a synergistic combination of the peroxidase-like property of the Au-Fe(3)O(4) nanozyme and the specific gene identification capability of the CRISPR-Cas12a. Once the CRISPR-Cas12a system recognizes a target resistance gene, it activates its trans-cleavage activity and subsequently releases the Au-Fe(3)O(4) nanozymes, which oxidizes the 3,3,5,5-tetramethylbenzidine (TMB) with color change from transparent to blue. The diagnosing signals could be captured and analyzed by a smartphone. This method detected kanamycin-resistance genes, ampicillin-resistance genes, and chloramphenicol-resistance genes by simple operation steps with high sensitivity (<0.1 CFU μL(-1)) and speediness (<1 h). This approach may prove easy for the accurate and sensitive diagnosis of the ARGs or ARB in the field, thus surveilling and controlling the microbial water quality flexibly and efficiently.202336925313
7807190.9914Copper oxide/peroxydisulfate system for urban wastewater disinfection: Performances, reactive species, and antibiotic resistance genes removal. In this study, copper oxide (CuO) catalyzed peroxydisulfate (PDS) system was investigated for the inactivation of a broad range of pathogenic microorganisms from urban wastewater. Complete inactivation of Escherichia coli, Enterococcus, F-specific RNA bacteriophages from secondary treated wastewater was achieved after a short time (15-30 min) treatment with CuO (10 g/L)/PDS (1 mM) system, but spores of sulfite-reducing bacteria took 120 min. No bacterial regrowth occurred during storage after treatment. Significant reduction of the pathogens was explained by the generation of the highly selective Cu(III) oxidant, as the predominant reactive species, which could quickly oxidize guanine through a one-electron oxidation pathway. Additionally, the potential of the CuO (10 g/L)/PDS (1 mM) system to inactivate antibiotic-resistant bacteria and antibiotic resistance genes (ARB&Gs) was explored. Sulfamethoxazole-resistant E. coli was used as the model ARB and a 3.2 log of reduction was observed after 10 min of treatment. A considerable reduction (0.7-2.3 log) of selected ARGs including blaTEM, qnrS, emrB, sul1, and genes related to the dissemination of antibiotic resistance, including the Class 1 integron-integrase (intI1), and the insertion sequence (IS613) was achieved after 60 min treatment. All these findings indicated the promising applicability of the CuO/PDS system as a disinfection technology for wastewater reuse in agriculture.202234648831