Diversity Of Resistance Genes Research Articles

Exploring Plasmid-Mediated Quinolone Resistance Gene Diversity among Ciprofloxacin-Resistant Proteus Species in a Tertiary Care Hospital of Bangladesh

Background: Increased resistance to fluoroquinolones has been observed in members of the Enterobacteriaceae, including Proteus species. The extensive use of fluoroquinolones and the presence of plasmid-mediated quinolone resistance (PMQR) genes are believed to contribute significantly to fluoroquinolone resistance. This study highlights the prevalence of ciprofloxacin-resistant Proteus species and the presence of specific PMQR genes among them, emphasizing their potential role in quinolone resistance. Method: The study included 30 ciprofloxacin-resistant Proteus isolates collected from wound swab, pus, urine, and blood samples. Identification of Proteus species was performed using culture and biochemical tests. Antibiotic susceptibility test was conducted using the Kirbey-Bauer disc-diffusion method. The minimum inhibitory concentration (MIC) of ciprofloxacin among ciprofloxacin-resistant Proteus species was determined using the agar dilution method. The study aimed to detect specific PMQR genes, namely aac(6´) lb-cr, qnrA, qnrB, qnrD, and qnrS, using polymerase chain reaction (PCR). Result: A total of 42 Proteus species were isolated from 310 various samples. Among the Proteus isolates, 71.43% were ciprofloxacin resistant. The study found that 30% of ciprofloxacin-resistant isolates were positive for aac(6´) lb-cr gene, 13.33% for qnrA and qnrS respectively and 10% for qnrD gene. In total, 60% of ciprofloxacin-resistant Proteus isolates were positive for at least one PMQR gene. The qnrB gene was not detected among ciprofloxacin-resistant Proteus species. The MIC of ciprofloxacin ranged from 8 μg/ml to 128 μg/ml among ciprofloxacin-resistant Proteus isolates. Conclusion: This study suggests aac(6´) lb-cr, qnrA, qnrS, and qnrD genes are emerging in Proteus species, potentially contributing to the development of quinolone resistance and has implications for clinical management and public health. J MEDICINE 2024; 25: 149-154

Whole-Genome Sequencing of Human and Porcine Escherichia coli Isolates on a Commercial Pig Farm in South Africa.

Escherichia coli is an indicator micro-organism in One Health antibiotic resistance surveillance programs. The purpose of the study was to describe and compare E. coli isolates obtained from pigs and human contacts from a commercial farm in South Africa using conventional methods and whole-genome sequencing (WGS). Porcine E. coli isolates were proportionally more resistant phenotypically and harbored a richer diversity of antibiotic resistance genes as compared to human E. coli isolates. Different pathovars, namely ExPEC (12.43%, 21/169), ETEC (4.14%, 7/169), EPEC (2.96%, 5/169), EAEC (2.96%, 5/169) and STEC (1.18%, 2/169), were detected at low frequencies. Sequence type complex (STc) 10 was the most prevalent (85.51%, 59/169) among human and porcine isolates. Six STcs (STc10, STc86, STc168, STc206, STc278 and STc469) were shared at the human-livestock interface according to multilocus sequence typing (MLST). Core-genome MLST and hierarchical clustering (HC) showed that human and porcine isolates were overall genetically diverse, but some clustering at HC2-HC200 was observed. In conclusion, even though the isolates shared a spatiotemporal relationship, there were still differences in the virulence potential, antibiotic resistance profiles and cgMLST and HC according to the source of isolation.

Azithromycin resistance in Escherichia coli and Salmonella from food-producing animals and meat in Europe.

To characterize the genetic basis of azithromycin resistance in Escherichia coli and Salmonella collected within the EU harmonized antimicrobial resistance (AMR) surveillance programme in 2014-18 and the Danish AMR surveillance programme in 2016-19. WGS data of 1007 E. coli [165 azithromycin resistant (MIC > 16 mg/L)] and 269 Salmonella [29 azithromycin resistant (MIC > 16 mg/L)] were screened for acquired macrolide resistance genes and mutations in rplDV, 23S rRNA and acrB genes using ResFinder v4.0, AMRFinder Plus and custom scripts. Genotype-phenotype concordance was determined for all isolates. Transferability of mef(C)-mph(G)-carrying plasmids was assessed by conjugation experiments. mph(A), mph(B), mef(B), erm(B) and mef(C)-mph(G) were detected in E. coli and Salmonella, whereas erm(C), erm(42), ere(A) and mph(E)-msr(E) were detected in E. coli only. The presence of macrolide resistance genes, alone or in combination, was concordant with the azithromycin-resistant phenotype in 69% of isolates. Distinct mph(A) operon structures were observed in azithromycin-susceptible (n = 50) and -resistant (n = 136) isolates. mef(C)-mph(G) were detected in porcine and bovine E. coli and in porcine Salmonella enterica serovar Derby and Salmonella enterica 1,4, [5],12:i:-, flanked downstream by ISCR2 or TnAs1 and associated with IncIγ and IncFII plasmids. Diverse azithromycin resistance genes were detected in E. coli and Salmonella from food-producing animals and meat in Europe. Azithromycin resistance genes mef(C)-mph(G) and erm(42) appear to be emerging primarily in porcine E. coli isolates. The identification of distinct mph(A) operon structures in susceptible and resistant isolates increases the predictive power of WGS-based methods for in silico detection of azithromycin resistance in Enterobacterales.

Genome-wide association analysis uncovers rice blast resistance alleles of Ptr and Pia

A critical step to maximize the usefulness of genome-wide association studies (GWAS) in plant breeding is the identification and validation of candidate genes underlying genetic associations. This is of particular importance in disease resistance breeding where allelic variants of resistance genes often confer resistance to distinct populations, or races, of a pathogen. Here, we perform a genome-wide association analysis of rice blast resistance in 500 genetically diverse rice accessions. To facilitate candidate gene identification, we produce de-novo genome assemblies of ten rice accessions with various rice blast resistance associations. These genome assemblies facilitate the identification and functional validation of novel alleles of the rice blast resistance genes Ptr and Pia. We uncover an allelic series for the unusual Ptr rice blast resistance gene, and additional alleles of the Pia resistance genes RGA4 and RGA5. By linking these associations to three thousand rice genomes we provide a useful tool to inform future rice blast breeding efforts. Our work shows that GWAS in combination with whole-genome sequencing is a powerful tool for gene cloning and to facilitate selection of specific resistance alleles for plant breeding.

Effects of different assembly strategies on gene annotation in activated sludge

Activated sludge comprises diverse bacteria, fungi, and other microorganisms, featuring a rich repertoire of genes involved in antibiotic resistance, pollutant degradation, and elemental cycling. In this regard, hybrid assembly technology can revolutionize metagenomics by detecting greater gene diversity in environmental samples. Nonetheless, the optimal utilization and comparability of genomic information between hybrid assembly and short- or long-read technology remain unclear. To address this gap, we compared the performance of the hybrid assembly, short- and long-read technologies, abundance and diversity of annotated genes, and taxonomic diversity by analysing 46, 161, and 45 activated sludge metagenomic datasets, respectively. The results revealed that hybrid assembly technology exhibited the best performance, generating the most contiguous and longest contigs but with a lower proportion of high-quality metagenome-assembled genomes than short-read technology. Compared with short- or long-read technologies, hybrid assembly technology can detect a greater diversity of microbiota and antibiotic resistance genes, as well as a wider range of potential hosts. However, this approach may yield lower gene abundance and pathogen detection. Our study revealed the specific advantages and disadvantages of hybrid assembly and short- and long-read applications in wastewater treatment plants, and our approach could serve as a blueprint to be extended to terrestrial environments.

A Study of Genotypic Characterization of ESBL and MBL Genes of β-Lactamase Producing Pseudomonas aeruginosa in Various Clinical Samples

Pseudomonas aeruginosa, a Gram-negative bacterium, presents a substantial challenge in healthcare due to its adaptability and resistance. This study delves into its genotypic characteristics, focusing on ESBL and MBL genes. The prevalence of P. aeruginosa in nosocomial infections, the research aims to decipher resistance mechanisms crucial for tailored interventions. The study includes 170 non-repetitive clinical samples with protocols. Antibiotic susceptibility testing reveals diverse resistance patterns. Molecular detection of ESBL and MBL genes involves DNA isolation, PCR amplification, and gel electrophoresis. The study examined 170 P. aeruginosa samples, revealing gender-specific variations with 65.91% male and 34.09% female isolates. Antimicrobial testing displayed resistance in Ceftazidime (59%) and Ciprofloxacin (48%), while Ticarcillin-clavulanic acid showed promising sensitivity (58%). Molecular identification unveiled diverse resistance genes across sample types, emphasizing genetic complexity. The study underscores the urgency for targeted therapeutic interventions and novel antimicrobial strategies against P. aeruginosa infections. As antimicrobial resistance complexities persist, this research guides efforts toward a profound understanding of clinical interventions and strategic antimicrobial management.

Whole genome sequencing analysis on antibiotic-resistant Escherichia coli isolated from pig farms in Banten Province, Indonesia.

The emergence and rapid increase in the incidence of multidrug-resistant (MDR) bacteria in pig farms has become a serious concern and reduced the choice of effective antibiotics. This study analyzed the phylogenetics and diversity of antibiotic resistance genes (ARGs) and molecularly identified the source of ARGs in antibiotic-resistant Escherichia coli isolated from pig farms in Banten Province, Indonesia. Forty-four antibiotic-resistant E. coli isolates from fecal samples from 44 pig farms in Banten Province, Indonesia, were used as samples. The samples were categorized into 14 clusters. Sequencing was performed using the Oxford Nanopore Technologies MinION platform, with barcoding before sequencing with Nanopore Rapid sequencing gDNA-barcoding (SQK-RBK110.96) according to manufacturing procedures. ARG detection was conducted using ResFinder, and the plasmid replicon was determined using PlasmidFinder. Three phylogenetic leaves of E. coli were identified in the pig farming cluster in Banten Province. The E. coli isolates exhibited potential resistance to nine classes of antibiotics. Fifty-one ARGs were identified across all isolates, with each cluster carrying a minimum of 10 ARGs. The ant(3'')-Ia and qnrS1 genes were present in all isolates. ARGs in the E. coli pig farming cluster originated mainly from plasmids, accounting for an average of 89.4%. The elevated potential for MDR events, coupled with the dominance of ARGs originating from plasmids, increases the risk of ARG spread among bacterial populations in animals, humans, and the environment.

Molecular Identification of invA gene from Salmonella species isolated from Human Sources in Southern Taraba, North-East Nigeria

Place and Duration of the Study: The study was conducted in Wukari, Donga, Ibi, and Takum Local Government Areas in Southern Taraba State, North-East Nigeria. These areas were chosen due to their high population of farmers and traders, making them suitable locations for investigating foodborne pathogens. The experimental research spanned a period of 7 months, during which 200 blood and stool samples were collected from food vendors in the selected areas. Methodology: Sample collection involved the collection of 200 blood and stool samples from food vendors in Wukari, Donga, Ibi, and Takum Local Government Areas. Isolation and confirmation of Salmonella species were carried out through cultural and biochemical analyses, with reference strains used for quality control. Deoxyribonucleic Acid (DNA) extraction was performed using the boiling technique, followed by Polymerase Chain Reaction (PCR) amplification of the invA gene. Electrophoresis on an agarose gel was used to visualize the presence of the invA gene in the isolates. Results: The results of the study revealed a high prevalence of Salmonella species among food vendors in Southern Taraba State, North-East Nigeria. Analysis of the invA gene showed its presence in all isolates, indicating the widespread distribution of virulent strains in the study area. The findings underscore the importance of PCR-based methods for the detection of Salmonella and suggest the need for comprehensive surveillance and control measures to prevent foodborne illnesses. Conclusion: The study demonstrates the significant prevalence of Salmonella species among food vendors in Southern Taraba State, North-East Nigeria. The detection of the invA gene in all isolates highlights the potential virulence of these strains and emphasizes the importance of effective surveillance and control strategies to mitigate the risk of foodborne diseases. Further research is warranted to elucidate the genetic diversity and antimicrobial resistance genes associated with Salmonella strains in the region, facilitating the development of targeted interventions for public health protection.

It’s a matter of microbes: a perspective on the microbiological aspects of micro- and nanoplastics in human health

Micro- and nanoplastics (MNPs) are everywhere: in the air we breathe, in our food and in virtually every type of water. Currently, it is unknown whether, and to what extent, these MNPs are hazardous to human health. Identifying risks of physical and chemical aspects of MNPs has gained a lot of attention over the last few years, and efforts have been made to quantify these risks. In our opinion, the momentum delivered by these efforts should be used to highlight the relevance of including another important aspect of MNPs: their associated microbes. Although more and more studies describe MNP-associated microbes, the interplay between physical, chemical and microbiological aspects are lacking. Hazard identification parameters describing this interplay are crucial to risk assessment strategies, yet the majority of effort has been directed towards optimizing human exposure parameters. Here, we address the importance of including microbiological aspects of MNPs in the risk assessment of MNPs. Physical and chemical aspects of MNPs impact the diversity and abundance of microbes and antibiotic resistance genes (ARGs). In turn, environmental factors (e.g., UV radiation, antibiotics) may impact the microbial composition directly, or indirectly by impacting physical or chemical MNP aspects. Future efforts should be directed towards investigating this interplay in order to determine the effects of these dynamic outcomes on human health. We believe that elucidating these pieces of the puzzle is needed to ultimately perform a more holistic risk assessment of MNPs on human health.

Genetic Diversification and Resistome of Coagulase-Negative Staphylococci from Nostrils of Healthy Dogs and Dog-Owners in La Rioja, Spain.

Coagulase-negative staphylococci (CoNS) species in healthy dogs and their owners could be transferred between these hosts and carry diverse antimicrobial resistance (AMR) genes of public health concern. This study determined the frequency, diversity, and AMR genes of nasal CoNS from healthy dogs and in-contact people as well as the rate of intra-household (between healthy dogs and dog-owners) transmission of CoNS. Nasal samples were collected and processed from 34 dogs and 41 humans from 27 households, and CoNS identification was done by MALDI-TOF-MS. The AMR determinants and genetic lineages were determined by PCR/sequencing. A total of 216 CoNS isolates were initially obtained and identified, and the AMR phenotypes were determined. From these, 130 non-repetitive CoNS were selected (one isolate of each species per sample or more than one if they presented different AMR phenotypes) and further characterized. The predominant species from dog carriers were S. epidermidis (26.5%), S. hominis (8.8%), and S. cohnii (8.8%), whereas in the human carriers, the predominant ones were S. epidermidis (80.4%), S. lugdunensis (9.8%), and S. hominis (9.8%). Intra-host species diversity (>one CoNS species) was detected in 37.5% of dogs and 21.6% of dog-owners. Conversely, 50% of dogs and 70.3% of dog-owners had intra-species AMR diversity (2-4 AMR-CoNS profiles). About 20% were susceptible to all antimicrobial agents tested, 31.5% displayed a multidrug resistance phenotype, and 17.4% were mecA-positive, located in SCCmec type V (24.2%), III (18.1%), IVc (12.1%), and II (6.1%). The other mec-A positive CoNS isolates (39.5%) had non-typeable SCCmec. The highest AMR rates were found against erythromycin (32.3%/mph(C), msr(A)) and mupirocin (20.8%/mupA), but the resistance rates for other antimicrobial agents were <10% each. Remarkably, one linezolid-resistant S. epidermidis-ST35 isolate was identified and mediated by four amino acid substitutions in L3 and one in L4 ribosomal proteins. Dogs and dog-owners as carriers of S. epidermidis with similar AMR patterns and genetic lineages (ST59, ST61, ST166 and ST278) were detected in four households (14.8%). Diverse CoNS carriage and moderate level of AMR were obtained from this study. The detection of CoNS carrying diverse SCCmec elements and intra-species AMR diversity highlights the roles of dog ownership in the potential transmission of antimicrobial-resistant CoNS in either direction.

16S rDNA-Based Amplicon Analysis Unveiled a Correlation Between the Bacterial Diversity and Antibiotic Resistance Genes of Bacteriome of Commercial Smokeless Tobacco Products.

The distribution of bacterial-derived antibiotic resistance genes (ARGs) in smokeless tobacco products is less explored and encourages understanding of the ARG profile of Indian smokeless tobacco products. Therefore, in the present investigation, ten commercial smokeless tobacco products were assessed for their bacterial diversity to understand the correlation between the inhabitant bacteria and predicted ARGs using a 16S rDNA-based metagenome analysis. Overall analysis showed the dominance of two phyla, i.e., Firmicutes (43.07%) and Proteobacteria (8.13%) among the samples, where Bacillus (9.76%), Terribacillus (8.06%), Lysinibacillus (5.8%), Alkalibacterium (5.6%), Oceanobacillus (3.52%), and Dickeya (3.1%) like genera were prevalent among these phyla. The phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt)-based analysis revealed 217 ARGs which were categorized into nine groups. Cationic antimicrobial polypeptides (CAMP, 33.8%), vancomycin (23.4%), penicillin-binding protein (13.8%), multidrug resistance MDR (10%), and β-lactam (9.3%) were among the top five contributors to ARGs. Staphylococcus, Dickeya, Bacillus, Aerococcus, and Alkalibacterium showed their strong and significant correlation (p value < 0.05) with various antibiotic resistance mechanisms. ARGs of different classes (blaTEM, blaSHV, blaCTX, tetX, vanA, aac3-II, mcr-1, intI-1, and intI2) were also successfully amplified in the metagenomes of SMT samples using their specific primers. The prevalence of ARGs in inhabitant bacteria of smokeless tobacco products suggests making steady policies to regulate the hygiene of commercial smokeless tobacco products.

Bacterial diversity and resistome analysis of drinking water stored in cisterns from two First Nations communities in Manitoba, Canada.

The microbiological content of water is an ongoing concern in First Nations communities in Canada. Many communities lack water treatment plants and continue to be under drinking water advisories. However, lack of access to treatment plants is only a part of the problem as poor water distribution systems also contribute to the failure to provide safe drinking water. Here, we studied the microbial diversity and antibiotic resistome from water stored in cisterns from two First Nations communities in Manitoba, Canada. We found that the cistern water contained a high number of bacteria and showed the presence of diverse antimicrobial resistance genes. Interestingly, the bacterial diversity and antimicrobial resistance genes varied considerably from that of the untreated source water, indicating that the origin of contamination in the cistern water came from within the treatment plant or along the delivery route to the homes. Our study highlights the importance of proper maintenance of the water distribution system in addition to access to water treatment facilities to ensure a supply of safe water to First Nations communities in Canada.IMPORTANCEThe work described addresses a critical issue in First Nations communities in Canada-the microbiological content of water. Many of these communities lack access to water treatment plants and frequently experience drinking water advisories. This study focused on the microbial diversity and antibiotic resistome in water stored in cisterns within two First Nations communities in Manitoba, Canada. These findings reveal that cistern water, a common source of drinking water in these communities, contains a high number of bacteria and a wide range of antimicrobial resistance genes. This highlights a serious health risk as exposure to such water can lead to the spread of drug-resistant infections, posing a threat to the well-being of the residents.

ESCHERICHIA COLI phenotypic characteristics and antagonistic activity in opisthorchiasis invasion

Opisthorchis felineus invasion in human causes inflammatory and dyskinetic disorders of the gastrointestinal tract accompanied by altered phenotypic characteristics in colon microbiota. The aim of research — study an impact of the Escherichia coli isolate phenotypic characteristics on Klebsiella spp. bacteria, isolated from colonic contents of patients with diagnosed opisthorchiasis as well as E. coli antagonistic activity. Materials and methods. The phenotypic properties of 54 E. coli isolates and 8 genus Klebsiella isolates obtained from colonic contents of patients with diagnosed opisthorchiasis were assessed. Identification of isolates and analysis of proteomic profiles were performed using Maldi BioTyper 3.0 software. 204 co-cultivation datasets were analyzed investigating antagonistic activity of E. coli isolates with varying properties on Klebsiella spp. E. coli and Klebsiella spp. isolates were examined by whole genome sequencing. Results. E. coli bacteria with typical phenotypic characteristics showed significantly more prominent antagonistic activity against Klebsiella spp. A significantly higher level of antagonistic activity against K. oxytoca bacteria vs K. pneumoniae strains. The proteomic bacterial strain profiles were divided into clusters depending on the level of antagonistic activity. E. coli molecular serotyping for O- and H-antigens revealed the genes of enterotoxigenic, enteroinvasive and extraintestinal pathogens in 60.0% of cases. Strains with the highest antagonistic activity index, which are carriers of the genes typical to enterotoxigenic E. coli sequence serotypes O6:H1 and O6:H5, were identified. The genome of such strains consisted of the largest number of virulence gene complexes: adhesins, invasins, toxins, bacteriocins. Multilocus sequence typing and sequence serotyping of E. coli and K. pneumoniae strains established their heterogeneity; K. oxytoca isolates were identified as ST242 and ST176. All strains were characterized by homology of antibiotic resistance markers (oqxA, oqxB, fosA) and a variety of beta-lactam resistance gene variants. Conclusion. It was found that E. coli isolates with typical phenotypic characteristics and carriers of virulence gene complexes exhibited significantly more pronounced antagonistic activity against Klebsiella spp. isolated from colonic contents of patients with diagnosed opisthorchiasis.

Minimal impact on the resistome of children in Botswana after azithromycin treatment for acute severe diarrhoeal disease.

Macrolide antibiotics, including azithromycin, can reduce under-five mortality rates and treat various infections in children in sub-Saharan Africa. These exposures, however, can select for antibiotic-resistant bacteria in the gut microbiota. Our previous randomized controlled trial (RCT) of a rapid-test-and-treat strategy for severe acute diarrhoeal disease in children in Botswana included an intervention (three-day azithromycin dose) group and a control group that received supportive treatment. In this prospective matched cohort study using stools collected at baseline and 60 days after treatment from RCT participants, the collection of antibiotic resistance genes or resistome was compared between groups. Certain macrolide resistance genes increased in prevalence by 13% to 55% at 60 days, without differences in gene presence between the intervention and control groups. These genes were linked to tetracycline resistance genes and mobile genetic elements. Azithromycin treatment for bacterial diarrhoea for young children in Botswana resulted in similar effects on the gut resistome as the supportive treatment and did not provide additional selective pressure for macrolide resistance gene maintenance. The gut microbiota of these children contains diverse macrolide resistance genes that may be transferred within the gut upon repeated exposures to azithromycin or co-selected by other antibiotics.

Distribution of bacterial community structures and spread of antibiotic resistome at industrially polluted sites of Mini River, Vadodara, Gujarat, India.

The influence of anthropogenic pollution on the distribution of bacterial diversity, antibiotic-resistant bacteria (ARBs), and antibiotic resistance genes (ARGs) was mapped at various geo-tagged sites of Mini River, Vadodara, Gujarat, India. The high-throughput 16S rRNA gene amplicon sequencing analysis revealed a higher relative abundance of Planctomycetota at the polluted sites, compared to the pristine site. Moreover, the relative abundance of Actinobacteriota increased, whereas Chloroflexi decreased in the water samples of polluted sites than the pristine site. The annotation of functional genes in the metagenome samples of Mini River sites indicated the presence of genes involved in the defence mechanisms against bacitracin, aminoglycosides, cephalosporins, chloramphenicol, streptogramin, streptomycin, methicillin, and colicin. The analysis of antibiotic resistome at the polluted sites of Mini River revealed the abundance of sulfonamide, beta-lactam, and aminoglycoside resistance. The presence of pathogens and ARB was significantly higher in water and sediment samples of polluted sites compared to the pristine site. The highest resistance of bacterial populations in the Mini River was recorded against sulfonamide (≥ 7.943 × 103CFU/mL) and ampicillin (≥ 8.128 × 103CFU/mL). The real-time PCR-based quantification of ARGs revealed the highest abundance of sulfonamide resistance genes sul1 and sul2 at the polluted sites of the Mini River. Additionally, the antimicrobial resistance genes aac(6')-Ib-Cr and blaTEM were also found abundantly at polluted sites of the Mini River. The findings provide insights into how anthropogenic pollution drives the ARG and ARB distribution in the riverine ecosystem, which may help with the development of antimicrobial resistance mitigation strategies.

Genomic landscape of NDM-1 producing multidrug-resistant Providencia stuartii causing burn wound infections in Bangladesh

The increasing antimicrobial resistance in Providencia stuartii (P. stuartii) worldwide, particularly concerning for immunocompromised and burn patients, has raised concern in Bangladesh, where the significance of this infectious opportunistic pathogen had been previously overlooked, prompting a need for investigation. The two strains of P. stuartii (P. stuartii SHNIBPS63 and P. stuartii SHNIBPS71) isolated from wound swab of two critically injured burn patients were found to be multidrug-resistant and P. stuartii SHNIBPS63 showed resistance to all the 22 antibiotics tested as well as revealed the co-existence of blaVEB-6 (Class A), blaNDM-1 (Class B), blaOXA-10 (Class D) beta lactamase genes. Complete resistance to carbapenems through the production of NDM-1, is indicative of an alarming situation as carbapenems are considered to be the last line antibiotic to combat this pathogen. Both isolates displayed strong biofilm-forming abilities and exhibited resistance to copper, zinc, and iron, in addition to carrying multiple genes associated with metal resistance and the formation of biofilms. The study also encompassed a pangenome analysis utilizing a dataset of eighty-six publicly available P. stuartii genomes (n = 86), revealing evidence of an open or expanding pangenome for P. stuartii. Also, an extensive genome-wide analysis of all the P. stuartii genomes revealed a concerning global prevalence of diverse antimicrobial resistance genes, with a particular alarm raised over the abundance of carbapenem resistance gene blaNDM-1. Additionally, this study highlighted the notable genetic diversity within P. stuartii, significant informations about phylogenomic relationships and ancestry, as well as potential for cross-species transmission, raising important implications for public health and microbial adaptation across different environments.

Airway 'Resistotypes' and Clinical Outcomes in Bronchiectasis.

Application of whole-genome shotgun metagenomics to the airway microbiome in bronchiectasis highlights a diverse pool of antimicrobial resistance genes: the 'resistome', the clinical significance of which remains unclear. Individuals with bronchiectasis were prospectively recruited into cross-sectional and longitudinal cohorts (n=280) including the international multicentre cross-sectional Cohort of Asian and Matched European Bronchiectasis 2 study (CAMEB 2; n=251) and two independent cohorts, one describing patients experiencing acute exacerbation and a further cohort of patients undergoing P. aeruginosa eradication treatment. Sputum was subjected to metagenomic sequencing and the bronchiectasis resistome evaluated in association with clinical outcomes and underlying host microbiomes. The bronchiectasis resistome features a unique resistance gene profile and elevated counts of aminoglycoside, bicyclomycin, phenicol, triclosan and multi-drug resistance genes. Longitudinally, it exhibits within-patient stability over time and during exacerbations despite between-patient heterogeneity. Proportional differences in baseline resistome profiles including increased macrolide and multi-drug resistance genes associate with shorter intervals to next exacerbation, while distinct resistome archetypes associate with frequent exacerbations, poorer lung function, geographic origin, and the host microbiome. Unsupervised analysis of resistome profiles identified two clinically relevant 'resistotypes' RT1 and RT2, the latter characterized by poor clinical outcomes, increased multi-drug resistance and P. aeruginosa. Successful targeted eradication in P. aeruginosa-colonized individuals mediated reversion from RT2 to RT1, a more clinically favourable resistome profile demonstrating reduced resistance gene diversity. The bronchiectasis resistome associates with clinical outcomes, geographic origin, and the underlying host microbiome. Bronchiectasis 'resistotypes' link to clinical disease and are modifiable through targeted antimicrobial therapy. This article is open access and distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives License 4.0 (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Diversity of antibiotic resistance gene variants at subsequent stages of the wastewater treatment process revealed by a metagenomic analysis of PCR amplicons.

Wastewater treatment plants have been recognised as point sources of various antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARG) which are considered recently emerging biological contaminants. So far, culture-based and molecular-based methods have been successfully applied to monitor antimicrobial resistance (AMR) in WWTPs. However, the methods applied do not permit the comprehensive identification of the true diversity of ARGs. In this study we applied next-generation sequencing for a metagenomic analysis of PCR amplicons of ARGs from the subsequent stages of the analysed WWTP. The presence of 14 genes conferring resistance to different antibiotic families was screened by PCR. In the next step, three genes were selected for detailed analysis of changes of the profile of ARG variants along the process. A relative abundance of 79 variants was analysed. The highest diversity was revealed in the ermF gene, with 52 variants. The relative abundance of some variants changed along the purification process, and some ARG variants might be present in novel hosts for which they were currently unassigned. Additionally, we identified a pool of novel ARG variants present in the studied WWTP. Overall, the results obtained indicated that the applied method is sufficient for analysing ARG variant diversity.

Prevalence of diverse antimicrobial resistance genes and bacteria in sewage treatment plant-derived sludge environment.

Antimicrobial resistance (AMR) contamination in the environment is one of the most significant worldwide threats of the 21st century. Since sludge is heavily exposed to diverse contaminants, including pharmaceuticals, the inhabitant bacterial population is expected to exhibit resistance to antimicrobial agents. In this study, sewage treatment plant (STP) sludge samples were analyzed to assess the antibiotic-resistant bacterial population, abundance of AMR genes (ermF, qnrS, Sul1, blaGES, blaCTX-M, and blaNDM), and mobile genetic elements (intl1 and IS26). Out of 16, six bacterial isolates exhibited resistance to 13 antibiotics with a high multiple antibiotic resistance index (MARI) (0.93) and high metal tolerance. Quantitative polymerase chain reaction showed the abundance of target genes ranging from 6.6×103 to 6.5×108 copies g-1 sludge. The overall outcome reveals that STP sludge comprised varied multidrug-resistant bacterial populations. It will give insights into the functions of heavy metals and biofilm development in the selection and spread of AMR genes and the associated bacteria. Therefore, the application of sludge needs proper screening for AMR and metal contamination prior to its countless applications. This study will contribute immensely to the risk analysis of STP effluents on environmental health, including control of AMR transmission.

High-throughput qPCR and Amplicon Sequencing as Complementary Methods for Profiling Antibiotic Resistance Genes in Urban Wetland Parks

Urban wetland parks are an important practice for urban wetland protection and utilization due to the vast ecosystem service value. As emerging contaminants, antibiotic resistance genes (ARGs) are great attractions for environmental research and public concerns. Based on high-throughput qPCR and high-throughput amplicon sequencing techniques, we investigated the occurrence, abundance, and distribution profiles of antibiotic resistance genes in the aquatic environment of Xiamen urban wetland parks (five sites). The influencing factors and driving mechanisms of antibiotic resistance genes were deciphered on the basis of microbial community structure and water quality. Diverse and abundant ARGs were observed and coexisted in urban wet parks. A total of 217 ARGs were detected in the water body of urban wetland parks, with an abundance up to 6.48×109 copies·L-1. Urban wetland parks were important hotspots and repositories of the antibiotic resistome. A total of nine bacterial genera, including Marivivens, NS5_marine_group, and Planktomarina, were identified as the potential carriers of diverse resistance genes (41 ARGs). The microbial communities could alone explain 51% of alterations in the antibiotic resistome in the aquatic environment of the urban wetland parks. Therefore, the microbial community was the key driving force for the occurrence and evolution of ARGs in urban wetland parks. Based on the results, with the presence of ARGs and antibiotic resistance bacteria, it is suggested that the water environments of urban wetland parks have potential risks of water ecological security and human health, and it is necessary to further enhance the research and control of microbial contaminants in the aquatic environment of urban wetland parks.