Polyclonal Spread of Fosfomycin Resistance among Carbapenemase-Producing Members of the Enterobacterales in the Czech Republic. - Related Documents




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158001.0000Polyclonal Spread of Fosfomycin Resistance among Carbapenemase-Producing Members of the Enterobacterales in the Czech Republic. Fosfomycin (FOS) has been recently reintroduced into clinical practice, but its effectiveness against multidrug-resistant (MDR) Enterobacterales is reduced due to the emergence of FOS resistance. The copresence of carbapenemases and FOS resistance could drastically limit antibiotic treatment. The aims of this study were (i) to investigate fosfomycin susceptibility profiles among carbapenem-resistant Enterobacterales (CRE) in the Czech Republic, (ii) to characterize the genetic environment of fosA genes among the collection, and (iii) to evaluate the presence of amino acid mutations in proteins involved in FOS resistance mechanisms. During the period from December 2018 to February 2022, 293 CRE isolates were collected from different hospitals in the Czech Republic. FOS MICs were assessed by the agar dilution method (ADM), FosA and FosC2 production was detected by the sodium phosphonoformate (PPF) test, and the presence of fosA-like genes was confirmed by PCR. Whole-genome sequencing was conducted with an Illumina NovaSeq 6000 system on selected strains, and the effect of point mutations in the FOS pathway was predicted using PROVEAN. Of these strains, 29% showed low susceptibility to fosfomycin (MIC, ≥16 μg/mL) by ADM. An NDM-producing Escherichia coli sequence type 648 (ST648) strain harbored a fosA10 gene on an IncK plasmid, while a VIM-producing Citrobacter freundii ST673 strain harbored a new fosA7 variant, designated fosA7.9. Analysis of mutations in the FOS pathway revealed several deleterious mutations occurring in GlpT, UhpT, UhpC, CyaA, and GlpR. Results regarding single substitutions in amino acid sequences highlighted a relationship between ST and specific mutations and an enhanced predisposition for certain STs to develop resistance. This study highlights the occurrence of several FOS resistance mechanisms in different clones spreading in the Czech Republic. IMPORTANCE Antimicrobial resistance (AMR) currently represents a concern for human health, and the reintroduction of antibiotics such as fosfomycin into clinical practice can provide further option in treatment of multidrug-resistant (MDR) bacterial infections. However, there is a global increase of fosfomycin-resistant bacteria, reducing its effectiveness. Considering this increase, it is crucial to monitor the spread of fosfomycin resistance in MDR bacteria in clinical settings and to investigate the resistance mechanism at the molecular level. Our study reports a large variety of fosfomycin resistance mechanisms among carbapenemase-producing Enterobacterales (CRE) in the Czech Republic. Our study summarizes the main achievements of our research on the use of molecular technologies, such as next-generation sequencing (NGS), to describe the heterogeneous mechanisms that reduce fosfomycin effectiveness in CRE. The results suggest that a program for widespread monitoring of fosfomycin resistance and epidemiology fosfomycin-resistant organisms can aide timely implementation of countermeasures to maintain the effectiveness of fosfomycin.202337098942
158210.9997Integrated Genomic and Phenotypic Characterization of an Mcr-10.1-Harboring Multidrug Resistant Escherichia coli Strain From Migratory Birds in China. Background: The global rise in antibiotic resistance among multidrug resistant (MDR) Gram-negative (GN) bacteria has posed significant health challenges, leading to the resurgence of colistin as a key defense against these bacteria. However, the widespread use of colistin has resulted in the rapid emergence of colistin resistance on a global scale. Ten members of the (mobile colistin resistance) mcr gene family, mcr-1 through mcr-10, have been reported and documented. Currently, bacteria reported to carry the mcr-10.1 gene are sensitive to colistin, but the mechanism underlying the low-level resistance phenomenon mediated by mcr-10.1 remains unclear. Methods: In this study, antimicrobial susceptibility testing (AST) was conducted on Escherichia coli (E.coli) isolated from Chinese migratory birds, resulting in the selection of 87 strains exhibiting MDR phenotypes. Whole-genome sequencing (draft) was performed on these 87 MDR E. coli strains, and for one of the E. coli strains carrying the mcr-10.1 gene, whole-genome sequencing, phenotypic characterization, AST and conjugation experiments were conducted to identify its resistance phenotypes and genetic characteristics. Results: Whole-genome sequencing (draft) of 87 MDR E. coli isolates revealed a diverse array of resistance genes, predominantly including aminoglycoside, β-lactam, tetracycline, and sulfonamide resistance genes. Remarkably, one isolate, despite being sensitive to colistin, harbored the mcr-10.1 gene. Further sequencing showed that mcr-10.1 was located in the conserved region of xerC-mcr-10.1, a hotspot for movable elements with various insertion sequences (ISs) or transposons nearby. Phenotypic characterization indicated that the MDR plasmid pGN25-mcr10.1 had no significant effect on the growth of GN25 and its derivatives but reduced the number of bacterial flagella. Conclusions: It is particularly important to note that bacteria harboring the mcr-10.1 gene may exhibit low minimum inhibitory concentration (MIC) values, but that the MIC values under colistin selective pressure can become progressively higher and exacerbate the difficulty of treating infections caused by mcr-10.1-associated bacteria. Therefore, vigilance for such "silent transmission" is warranted, and continuous monitoring of the spread of mcr-10.1 is necessary in the future.202540343190
189220.9997Colistin Resistance Mediated by Mcr-3-Related Phosphoethanolamine Transferase Genes in Aeromonas Species Isolated from Aquatic Environments in Avaga and Pakro Communities in the Eastern Region of Ghana. PURPOSE: Colistin is classified by the World Health Organization (WHO) as a critically important and last-resort antibiotic for the treatment of infections caused by carbapenem-resistant bacteria. However, colistin resistance mediated by chromosomal mutations or plasmid-linked mobilized colistin resistance (mcr) genes has emerged. METHODS: Thirteen mcr-positive Aeromonas species isolated from water samples collected in Eastern Ghana were analyzed using whole-genome sequencing (WGS). Antimicrobial susceptibility was tested using the broth microdilution method. Resistome analysis was performed in silico using a web-based platform. RESULTS: The minimum inhibitory concentration (MIC) of colistin for all except three isolates was >4 µg/mL. Nine new sequence types were identified and whole-genome analysis revealed that the isolates harbored genes (mcr-3-related genes) that code for Lipid A phosphoethanolamine transferases on their chromosomes. BLAST analysis indicated that the amino acid sequences of the mcr-3-related genes detected varied from those previously reported and shared 79.04-99.86% nucleotide sequence identity with publicly available mcr-3 variants and mcr-3-related phosphoethanolamine transferases. Analysis of the genetic context of mcr-3-related genes revealed that the genetic environment surrounding mcr-3-related genes was diverse among the different species of Aeromonas but conserved among isolates of the same species. Mcr-3-related-gene-IS-mcr-3-related-gene segment was identified in three Aeromonas caviae strains. CONCLUSION: The presence of mcr-3-related genes close to insertion elements is important for continuous monitoring to better understand how to control the mobilization and dissemination of antibiotic resistance genes.202439050833
189730.9997Plasmid-based replicon typing: Useful tool in demonstrating the silent pandemic of plasmid-mediated multi-drug resistance in Enterobacterales. BACKGROUND: Multi-drug resistant Enterobacterales increasingly isolated in hospital settings have a significant impact on therapy and overall treatment costs. Conjugative plasmids carrying relevant resistance genes have been described as the most frequent mechanism of acquisition and spread of resistance. PCR-based replicon typing (PBRT) is a method for plasmid identification and incompatibility typing which helps detect the presence of plasmid families in these bacteria. This study was undertaken to provide an insight into the prevalence of resistance plasmids in MDR Enterobacterales in our tertiary care setting. METHODS: A selection of one hundred multi-drug resistant Enterobacterale isolates sourced from clinical samples were subjected to PCR-based replicon typing. RESULTS: A total of 21 plasmid replicon types were detected from 85% of the isolates out of the 28 families described in literature. Majority of the isolates (54%) showed three or more replicons. IncF was the most frequent plasmid family detected with FIA being the most common replicon type (43%) followed by FII (29%) and FIB (28%) replicons. Among the IncX plasmid family, X3 replicon was the commonest (14%). IncF and IncX plasmid families are known to carry a large spectrum of resistance genes. CONCLUSION: The presence of these plasmids engenders emergent steps to be taken for prevention of their transmission in the form of strict infection control measures in the hospital and adoption of novel methods of plasmid curing to eliminate the plasmids from these organisms rendering them susceptible to the currently used antimicrobials.202540463599
168340.9997Colonization of a hand washing sink in a veterinary hospital by an Enterobacter hormaechei strain carrying multiple resistances to high importance antimicrobials. BACKGROUND: Hospital intensive care units (ICUs) are known reservoirs of multidrug resistant nosocomial bacteria. Targeted environmental monitoring of these organisms in health care facilities can strengthen infection control procedures. A routine surveillance of extended spectrum beta-lactamase (ESBL) producers in a large Australian veterinary teaching hospital detected the opportunistic pathogen Enterobacter hormaechei in a hand washing sink of the ICU. The organism persisted for several weeks, despite two disinfection attempts. Four isolates were characterized in this study. METHODS: Brilliance-ESBL selective plates were inoculated from environmental swabs collected throughout the hospital. Presumptive identification was done by conventional biochemistry. Genomes of multidrug resistant Enterobacter were entirely sequenced with Illumina and Nanopore platforms. Phylogenetic markers, mobile genetic elements and antimicrobial resistance genes were identified in silico. Antibiograms of isolates and transconjugants were established with Sensititre microdilution plates. RESULTS: The isolates possessed a chromosomal Tn7-associated silver/copper resistance locus and a large IncH12 conjugative plasmid encoding resistance against tellurium, arsenic, mercury and nine classes of antimicrobials. Clusters of antimicrobial resistance genes were associated with class 1 integrons and IS26, IS903 and ISCR transposable elements. The blaSHV-12, qnrB2 and mcr-9.1 genes, respectively conferring resistance to cephalosporins, quinolones and colistin, were present in a locus flanked by two IS903 copies. ESBL production and enrofloxacin resistance were confirmed phenotypically. The isolates appeared susceptible to colistin, possibly reflecting the inducible nature of mcr-9.1. CONCLUSIONS: The persistence of this strain in the veterinary hospital represented a risk of further accumulation and dissemination of antimicrobial resistance, prompting a thorough disinfection of the ICU. The organism was not recovered from subsequent environmental swabs, and nosocomial Enterobacter infections were not observed in the hospital during that period. This study shows that targeted routine environmental surveillance programs to track organisms with major resistance phenotypes, coupled with disinfection procedures and follow-up microbiological cultures are useful to control these risks in sensitive areas of large veterinary hospitals.202033087168
168950.9997Occurrence and Characteristics of Mcrs among Gram-Negative Bacteria Causing Bloodstream Infections of Infant Inpatients between 2006 and 2019 in China. The aim of this study was to determine the occurrence of mobilized colistin resistance (mcr) genes in Gram-negative bacteria causing bloodstream infections of child inpatients in China. Bacteria were collected between 2006 and 2019 in a maternal and child health hospital, and mcr genes were screened by PCR. Five of 252 isolates were mcr-positive, including one mcr-1-positive colistin-resistant Escherichia coli isolate, two mcr-9-positive colistin-susceptible Salmonella enterica isolates, and two mcr-9-positive colistin-susceptible Enterobacter hormaechei isolates. These were obtained from two neonate and three infant patients admitted between 2009 and 2018. The E. coli isolate was obtained from a neonate aged 20 min, suggestive of a possible mother-to-neonate transmission. The five mcr-positive isolates were multidrug resistant, and two S. enterica and one E. hormaechei isolate showed a hypervirulent phenotype compared to a hypervirulent Klebsiella pneumoniae type strain in a Galleria mellonella infection model. The mcr-1 gene was carried by an IncX4-type pA1-like epidemic plasmid, and the mcr-9 gene was detected on IncHI2/2A-type novel plasmids co-carrying multiple resistance genes. The four IncHI2/2A-type plasmids shared a backbone and a high similarity (≥77% coverage and ≥ 90% nucleotide identity), suggesting that they were derived from a common ancestor with cross-species transmission and have circulated locally over a long period. The conjugation assay showed that the mcr-1-encoding plasmid and one mcr-9-encoding plasmid were self-transmissible to E. coli with high conjugation frequencies. Our findings demonstrate that mcr genes have disseminated in the community and/or hospitals, mediated by epidemic/endemic plasmids over a long period. The study shows that continuous monitoring of mcr genes is imperative for understanding and tackling their dissemination. IMPORTANCE Antimicrobial resistance, especially the spread of carbapenemase-producing Enterobacteriaceae (CPE), represents one of the largest challenges to One Health coverage of environmental, animal, and human sectors. Colistin is one of the last-line antibiotics for clinical treatment of CPE. However, the emergence of the mobilized colistin resistance (mcr) gene largely threatens the usage of colistin in the clinical setting. In this study, we investigated the existence of mcr genes in 252 Gram-negative bacteria collected between 2006 and 2019 which caused bloodstream infections of child inpatients in China. We found a high prevalence of mcr carriage among children inpatients in the absence of professional exposure, and mcr might have widely disseminated in the community via different routes. This study emphasizes the importance of rational use of colistin in the One Health frame, and highlights both the urgent need for understanding the prevalence and dissemination of mcr genes in different populations and the importance of effective measures to control their spread.202235138190
192060.9997Exploring the resistome, virulome, and mobilome of multidrug-resistant Klebsiella pneumoniae isolates: deciphering the molecular basis of carbapenem resistance. BACKGROUND: Klebsiella pneumoniae, a notorious pathogen for causing nosocomial infections has become a major cause of neonatal septicemia, leading to high morbidity and mortality worldwide. This opportunistic bacterium has become highly resistant to antibiotics due to the widespread acquisition of genes encoding a variety of enzymes such as extended-spectrum beta-lactamases (ESBLs) and carbapenemases. We collected Klebsiella pneumoniae isolates from a local tertiary care hospital from February 2019-February 2021. To gain molecular insight into the resistome, virulome, and genetic environment of significant genes of multidrug-resistant K. pneumoniae isolates, we performed the short-read whole-genome sequencing of 10 K. pneumoniae isolates recovered from adult patients, neonates, and hospital tap water samples. RESULTS: The draft genomes of the isolates varied in size, ranging from 5.48 to 5.96 Mbp suggesting the genome plasticity of this pathogen. Various genes conferring resistance to different classes of antibiotics e.g., aminoglycosides, quinolones, sulfonamides, tetracycline, and trimethoprim were identified in all sequenced isolates. The highest resistance was observed towards carbapenems, which has been putatively linked to the presence of both class B and class D carbapenemases, bla(NDM,) and bla(OXA), respectively. Moreover, the biocide resistance gene qacEdelta1 was found in 6/10 of the sequenced strains. The sequenced isolates exhibited a broad range of sequence types and capsular types. The significant antibiotic resistance genes (ARGs) were bracketed by a variety of mobile genetic elements (MGEs). Various spontaneous mutations in genes other than the acquired antibiotic-resistance genes were observed, which play an indirect role in making these bugs resistant to antibiotics. Loss or deficiency of outer membrane porins, combined with ESBL production, played a significant role in carbapenem resistance in our sequenced isolates. Phylogenetic analysis revealed that the study isolates exhibited evolutionary relationships with strains from China, India, and the USA suggesting a shared evolutionary history and potential dissemination of similar genes amongst the isolates of different origins. CONCLUSIONS: This study provides valuable insight into the presence of multiple mechanisms of carbapenem resistance in K. pneumoniae strains including the acquisition of multiple antibiotic-resistance genes through mobile genetic elements. Identification of rich mobilome yielded insightful information regarding the crucial role of insertion sequences, transposons, and integrons in shaping the genome of bacteria for the transmission of various resistance-associated genes. Multi-drug resistant isolates that had the fewest resistance genes exhibited a significant number of mutations. K. pneumoniae isolate from water source displayed comparable antibiotic resistance determinants to clinical isolates and the highest number of virulence-associated genes suggesting the possible interplay of ARGs amongst bacteria from different sources.202438664636
158370.9997Identification of Novel Mobilized Colistin Resistance Gene mcr-9 in a Multidrug-Resistant, Colistin-Susceptible Salmonella enterica Serotype Typhimurium Isolate. Mobilized colistin resistance (mcr) genes are plasmid-borne genes that confer resistance to colistin, an antibiotic used to treat severe bacterial infections. To date, eight known mcr homologues have been described (mcr-1 to -8). Here, we describe mcr-9, a novel mcr homologue detected during routine in silico screening of sequenced Salmonella genomes for antimicrobial resistance genes. The amino acid sequence of mcr-9, detected in a multidrug-resistant (MDR) Salmonella enterica serotype Typhimurium (S Typhimurium) strain isolated from a human patient in Washington State in 2010, most closely resembled mcr-3, aligning with 64.5% amino acid identity and 99.5% coverage using Translated Nucleotide BLAST (tblastn). The S. Typhimurium strain was tested for phenotypic resistance to colistin and was found to be sensitive at the 2-mg/liter European Committee on Antimicrobial Susceptibility Testing breakpoint under the tested conditions. mcr-9 was cloned in colistin-susceptible Escherichia coli NEB5α under an IPTG (isopropyl-β-d-thiogalactopyranoside)-induced promoter to determine whether it was capable of conferring resistance to colistin when expressed in a heterologous host. Expression of mcr-9 conferred resistance to colistin in E. coli NEB5α at 1, 2, and 2.5 mg/liter colistin, albeit at a lower level than mcr-3 Pairwise comparisons of the predicted protein structures associated with all nine mcr homologues (Mcr-1 to -9) revealed that Mcr-9, Mcr-3, Mcr-4, and Mcr-7 share a high degree of similarity at the structural level. Our results indicate that mcr-9 is capable of conferring phenotypic resistance to colistin in Enterobacteriaceae and should be immediately considered when monitoring plasmid-mediated colistin resistance.IMPORTANCE Colistin is a last-resort antibiotic that is used to treat severe infections caused by MDR and extensively drug-resistant (XDR) bacteria. The World Health Organization (WHO) has designated colistin as a "highest priority critically important antimicrobial for human medicine" (WHO, Critically Important Antimicrobials for Human Medicine, 5th revision, 2017, https://www.who.int/foodsafety/publications/antimicrobials-fifth/en/), as it is often one of the only therapies available for treating serious bacterial infections in critically ill patients. Plasmid-borne mcr genes that confer resistance to colistin pose a threat to public health at an international scale, as they can be transmitted via horizontal gene transfer and have the potential to spread globally. Therefore, the establishment of a complete reference of mcr genes that can be used to screen for plasmid-mediated colistin resistance is essential for developing effective control strategies.201931064835
189480.9997Phenotypic and Genotypic Characterization of Multidrug-Resistant Enterobacter hormaechei Carrying qnrS Gene Isolated from Chicken Feed in China. Multidrug resistance (MDR) in Enterobacteriaceae including resistance to quinolones is rising worldwide. The plasmid-mediated quinolone resistance (PMQR) gene qnrS is prevalent in Enterobacteriaceae. However, the qnrS gene is rarely found in Enterobacter hormaechei (E. hormaechei). Here, we reported one multidrug resistant E. hormaechei strain M1 carrying the qnrS1 and bla(TEM-1) genes. This study was to analyze the characteristics of MDR E. hormaechei strain M1. The E. hormaechei strain M1 was identified as Enterobacter cloacae complex by biochemical assay and 16S rRNA sequencing. The whole genome was sequenced by the Oxford Nanopore method. Taxonomy of the E. hormaechei was based on multilocus sequence typing (MLST). The qnrS with the other antibiotic resistance genes were coexisted on IncF plasmid (pM1). Besides, the virulence factors associated with pathogenicity were also located on pM1. The qnrS1 gene was located between insertion element IS2A (upstream) and transposition element ISKra4 (downstream). The comparison result of IncF plasmids revealed that they had a common plasmid backbone. Susceptibility experiment revealed that the E. hormaechei M1 showed extensive resistance to the clinical antimicrobials. The conjugation transfer was performed by filter membrane incubation method. The competition and plasmid stability assays suggested the host bacteria carrying qnrS had an energy burden. As far as we know, this is the first report that E. hormaechei carrying qnrS was isolated from chicken feed. The chicken feed and poultry products could serve as a vehicle for these MDR bacteria, which could transfer between animals and humans through the food chain. We need to pay close attention to the epidemiology of E. hormaechei and prevent their further dissemination. IMPORTANCE Enterobacter hormaechei is an opportunistic pathogen. It can cause infections in humans and animals. Plasmid-mediated quinolone resistance (PMQR) gene qnrS can be transferred intergenus, which is leading to increase the quinolone resistance levels in Enterobacteriaceae. Chicken feed could serve as a vehicle for the MDR E. hormaechei. Therefore, antibiotic-resistance genes (ARGs) might be transferred to the intestinal flora after entering the gastrointestinal tract with the feed. Furthermore, antibiotic-resistant bacteria (ARB) were also excreted into environment with feces, posing a huge threat to public health. This requires us to monitor the ARB and antibiotic-resistant plasmids in the feed. Here, we demonstrated the characteristics of one MDR E. hormaechei isolate from chicken feed. The plasmid carrying the qnrS gene is a conjugative plasmid with transferability. The presence of plasmid carrying antibiotic-resistance genes requires the maintenance of antibiotic pressure. In addition, the E. hormaechei M1 belonged to new sequence type (ST). These data show the MDR E. hormaechei M1 is a novel strain that requires our further research.202235467399
190290.9997Large-scale analysis of putative plasmids in clinical multidrug-resistant Escherichia coli isolates from Vietnamese patients. INTRODUCTION: In the past decades, extended-spectrum beta-lactamase (ESBL)-producing and carbapenem-resistant (CR) Escherichia coli isolates have been detected in Vietnamese hospitals. The transfer of antimicrobial resistance (AMR) genes carried on plasmids is mainly responsible for the emergence of multidrug-resistant E. coli strains and the spread of AMR genes through horizontal gene transfer. Therefore, it is important to thoroughly study the characteristics of AMR gene-harboring plasmids in clinical multidrug-resistant bacterial isolates. METHODS: The profiles of plasmid assemblies were determined by analyzing previously published whole-genome sequencing data of 751 multidrug-resistant E. coli isolates from Vietnamese hospitals in order to identify the risk of AMR gene horizontal transfer and dissemination. RESULTS: The number of putative plasmids in isolates was independent of the sequencing coverage. These putative plasmids originated from various bacterial species, but mostly from the Escherichia genus, particularly E. coli species. Many different AMR genes were detected in plasmid contigs of the studied isolates, and their number was higher in CR isolates than in ESBL-producing isolates. Similarly, the bla(KPC-2), bla(NDM-5), bla(OXA-1), bla(OXA-48), and bla(OXA-181) β-lactamase genes, associated with resistance to carbapenems, were more frequent in CR strains. Sequence similarity network and genome annotation analyses revealed high conservation of the β-lactamase gene clusters in plasmid contigs that carried the same AMR genes. DISCUSSION: Our study provides evidence of horizontal gene transfer in multidrug-resistant E. coli isolates via conjugative plasmids, thus rapidly accelerating the emergence of resistant bacteria. Besides reducing antibiotic misuse, prevention of plasmid transmission also is essential to limit antibiotic resistance.202337323902
1573100.9996Genomic Analysis of a Pan-Resistant Isolate of Klebsiella pneumoniae, United States 2016. Antimicrobial resistance is a threat to public health globally and leads to an estimated 23,000 deaths annually in the United States alone. Here, we report the genomic characterization of an unusual Klebsiella pneumoniae, nonsusceptible to all 26 antibiotics tested, that was isolated from a U.S. PATIENT: The isolate harbored four known beta-lactamase genes, including plasmid-mediated bla(NDM-1) and bla(CMY-6), as well as chromosomal bla(CTX-M-15) and bla(SHV-28), which accounted for resistance to all beta-lactams tested. In addition, sequence analysis identified mechanisms that could explain all other reported nonsusceptibility results, including nonsusceptibility to colistin, tigecycline, and chloramphenicol. Two plasmids, IncA/C2 and IncFIB, were closely related to mobile elements described previously and isolated from Gram-negative bacteria from China, Nepal, India, the United States, and Kenya, suggesting possible origins of the isolate and plasmids. This is one of the first K. pneumoniae isolates in the United States to have been reported to the Centers for Disease Control and Prevention (CDC) as nonsusceptible to all drugs tested, including all beta-lactams, colistin, and tigecycline.IMPORTANCE Antimicrobial resistance is a major public health threat worldwide. Bacteria that are nonsusceptible or resistant to all antimicrobials available are of major concern to patients and the public because of lack of treatment options and potential for spread. A Klebsiella pneumoniae strain that was nonsusceptible to all tested antibiotics was isolated from a U.S. PATIENT: Mechanisms that could explain all observed phenotypic antimicrobial resistance phenotypes, including resistance to colistin and beta-lactams, were identified through whole-genome sequencing. The large variety of resistance determinants identified demonstrates the usefulness of whole-genome sequencing for detecting these genes in an outbreak response. Sequencing of isolates with rare and unusual phenotypes can provide information on how these extremely resistant isolates develop, including whether resistance is acquired on mobile elements or accumulated through chromosomal mutations. Moreover, this provides further insight into not only detecting these highly resistant organisms but also preventing their spread.201829615503
1627110.9996Screening of colistin-resistant bacteria in livestock animals from France. Colistin is frequently used as a growth factor or treatment against infectious bacterial diseases in animals. The Veterinary Division of the European Medicines Agency (EMA) restricted colistin use as a second-line treatment to reduce colistin resistance. In 2020, 282 faecal samples were collected from chickens, cattle, sheep, goats, and pigs in the south of France. In order to track the emergence of mobilized colistin resistant (mcr) genes in pigs, 111 samples were re-collected in 2021 and included pig faeces, food, and water from the same location. All samples were cultured in a selective Lucie Bardet Jean-Marc Rolain (LBJMR) medium and colonies were identified using MALDI-TOF mass spectrometry and then antibiotic susceptibility tests were performed. PCR and Sanger sequencing were performed to screen for the presence of mcr genes. The selective culture revealed the presence of 397 bacteria corresponding to 35 different bacterial species including Gram-negative and Gram-positive. Pigs had the highest prevalence of colistin-resistant bacteria with an abundance of intrinsically colistin-resistant bacteria and from these samples one strain harbouring both mcr-1 and mcr-3 has been isolated. The second collection allowed us to identify 304 bacteria and revealed the spread of mcr-1 and mcr-3 in pigs. In the other samples, naturally, colistin-resistant bacteria were more frequent, nevertheless the mcr-1 variant was the most abundant gene found in chicken, sheep, and goat samples and one cattle sample was positive for the mcr-3 gene. Animals are potential reservoir of colistin-resistant bacteria which varies from one animal to another. Interventions and alternative options are required to reduce the emergence of colistin resistance and to avoid zoonotic transmissions.202236414994
1684120.9996Plasmid-encoded gene duplications of extended-spectrum β-lactamases in clinical bacterial isolates. INTRODUCTION: The emergence of extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae is an urgent and alarming One Health problem. This study aimed to investigate duplications of plasmid-encoded ESBL genes and their impact on antimicrobial resistance (AMR) phenotypes in clinical and screening isolates. METHODS: Multi-drug-resistant bacteria from hospitalized patients were collected during routine clinical surveillance from January 2022 to June 2023, and their antimicrobial susceptibility patterns were determined. Genotypes were extracted from long-read whole-genome sequencing data. Furthermore, plasmids and other mobile genetic elements associated with ESBL genes were characterized, and the ESBL genes were correlated to ceftazidime minimal inhibitory concentration (MIC). RESULTS: In total, we identified four cases of plasmid-encoded ESBL gene duplications that match four genetically similar plasmids during the 18-month surveillance period: five Escherichia coli and three Klebsiella pneumoniae isolates. As the ESBL genes were part of transposable elements, the surrounding sequence regions were duplicated as well. In-depth analysis revealed insertion sequence (IS)-mediated transposition mechanisms. Isolates with duplicated ESBL genes exhibited a higher MIC for ceftazidime in comparison to isolates with a single gene copy (3-256 vs. 1.5-32 mg/L, respectively). CONCLUSION: ESBL gene duplications led to an increased phenotypic resistance against ceftazidime. Our data suggest that ESBL gene duplications by an IS-mediated transposition are a relevant mechanism for how AMR develops in the clinical setting and is part of the microevolution of plasmids.202438469349
1686130.9996Resistome of carbapenem- and colistin-resistant Klebsiella pneumoniae clinical isolates. The emergence and dissemination of carbapenemases, bacterial enzymes able to inactivate most β-lactam antibiotics, in Enterobacteriaceae is of increasing concern. The concurrent spread of resistance against colistin, an antibiotic of last resort, further compounds this challenge further. Whole-genome sequencing (WGS) can play a significant role in the rapid and accurate detection/characterization of existing and emergent resistance determinants, an essential aspect of public health surveillance and response activities to combat the spread of antimicrobial resistant bacteria. In the current study, WGS data was used to characterize the genomic content of antimicrobial resistance genes, including those encoding carbapenemases, in 10 multidrug-resistant Klebsiella pneumoniae isolates from Pakistan. These clinical isolates represented five sequence types: ST11 (n = 3 isolates), ST14 (n = 3), ST15 (n = 1), ST101 (n = 2), and ST307 (n = 1). Resistance profiles against 25 clinically-relevant antimicrobials were determined by broth microdilution; resistant phenotypes were observed for at least 15 of the 25 antibiotics tested in all isolates except one. Specifically, 8/10 isolates were carbapenem-resistant and 7/10 isolates were colistin-resistant. The blaNDM-1 and blaOXA-48 carbapenemase genes were present in 7/10 and 5/10 isolates, respectively; including 2 isolates carrying both genes. No plasmid-mediated determinants for colistin resistance (e.g. mcr) were detected, but disruptions and mutations in chromosomal loci (i.e. mgrB and pmrB) previously reported to confer colistin resistance were observed. A blaOXA-48-carrying IncL/M-type plasmid was found in all blaOXA-48-positive isolates. The application of WGS to molecular epidemiology and surveillance studies, as exemplified here, will provide both a more complete understanding of the global distribution of MDR isolates and a robust surveillance tool useful for detecting emerging threats to public health.201829883490
1896140.9996Difference analysis and characteristics of incompatibility group plasmid replicons in gram-negative bacteria with different antimicrobial phenotypes in Henan, China. BACKGROUND: Multi-drug-resistant organisms (MDROs) in gram-negative bacteria have caused a global epidemic, especially the bacterial resistance to carbapenem agents. Plasmid is the common vehicle for carrying antimicrobial resistance genes (ARGs), and the transmission of plasmids is also one of the important reasons for the emergence of MDROs. Different incompatibility group plasmid replicons are highly correlated with the acquisition, dissemination, and evolution of resistance genes. Based on this, the study aims to identify relevant characteristics of various plasmids and provide a theoretical foundation for clinical anti-infection treatment. METHODS: 330 gram-negative strains with different antimicrobial phenotypes from a tertiary hospital in Henan Province were included in this study to clarify the difference in incompatibility group plasmid replicons. Additionally, we combined the information from the PLSDB database to elaborate on the potential association between different plasmid replicons and ARGs. The VITEK mass spectrometer was used for species identification, and the VITEK-compact 2 automatic microbial system was used for the antimicrobial susceptibility test (AST). PCR-based replicon typing (PBRT) detected the plasmid profiles, and thirty-three different plasmid replicons were determined. All the carbapenem-resistant organisms (CROs) were tested for the carbapenemase genes. RESULTS: 21 plasmid replicon types were detected in this experiment, with the highest prevalence of IncFII, IncFIB, IncR, and IncFIA. Notably, the detection rate of IncX3 plasmids in CROs is higher, which is different in strains with other antimicrobial phenotypes. The number of plasmid replicons they carried increased with the strain resistance increase. Enterobacterales took a higher number of plasmid replicons than other gram-negative bacteria. The same strain tends to have more than one plasmid replicon type. IncF-type plasmids tend to be associated with MDROs. Combined with PLSDB database analysis, IncFII and IncX3 are critical platforms for taking bla(KPC-2) and bla(NDM). CONCLUSIONS: MDROs tend to carry more complex plasmid replicons compared with non-MDROs. The plasmid replicons that are predominantly prevalent and associated with ARGs differ in various species. The wide distribution of IncF-type plasmids and their close association with MDROs should deserve our attention. Further investigation into the critical role of plasmids in the carriage, evolution, and transmission of ARGs is needed.202438373913
2536150.9996Global Burden of Colistin-Resistant Bacteria: Mobilized Colistin Resistance Genes Study (1980-2018). Colistin is considered to be an antimicrobial of last-resort for the treatment of multidrug-resistant Gram-negative bacterial infections. The recent global dissemination of mobilized colistin resistance (mcr) genes is an urgent public health threat. An accurate estimate of the global prevalence of mcr genes, their reservoirs and the potential pathways for human transmission are required to implement control and prevention strategies, yet such data are lacking. Publications from four English (PubMed, Scopus, the Cochrane Database of Systematic Reviews and Web of Science) and two Chinese (CNKI and WANFANG) databases published between 18 November 2015 and 30 December 2018 were identified. In this systematic review and meta-analysis, the prevalence of mcr genes in bacteria isolated from humans, animals, the environment and food products were investigated. A total of 974 publications were identified. 202 observational studies were included in the systematic review and 71 in the meta-analysis. mcr genes were reported from 47 countries across six continents and the overall average prevalence was 4.7% (0.1-9.3%). China reported the highest number of mcr-positive strains. Pathogenic Escherichia coli (54%), isolated from animals (52%) and harboring an IncI2 plasmid (34%) were the bacteria with highest prevalence of mcr genes. The estimated prevalence of mcr-1 pathogenic E. coli was higher in food-animals than in humans and food products, which suggests a role for foodborne transmission. This study provides a comprehensive assessment of prevalence of the mcr gene by source, organism, genotype and type of plasmid.201931623244
1898160.9996Multiple-Replicon Resistance Plasmids of Klebsiella Mediate Extensive Dissemination of Antimicrobial Genes. Multiple-replicon resistance plasmids have become important carriers of resistance genes in Gram-negative bacteria, and the evolution of multiple-replicon plasmids is still not clear. Here, 56 isolates of Klebsiella isolated from different wild animals and environments between 2018 and 2020 were identified by phenotyping via the micro-broth dilution method and were sequenced and analyzed for bacterial genome-wide association study. Our results revealed that the isolates from non-human sources showed more extensive drug resistance and especially strong resistance to ampicillin (up to 80.36%). The isolates from Malayan pangolin were particularly highly resistant to cephalosporins, chloramphenicol, levofloxacin, and sulfamethoxazole. Genomic analysis showed that the resistance plasmids in these isolates carried many antibiotic resistance genes. Further analysis of 69 plasmids demonstrated that 28 plasmids were multiple-replicon plasmids, mainly carrying beta-lactamase genes such as bla (CTX-M-) (15), bla (CTX-M-) (14), bla (CTX-M-) (55), bla (OXA-) (1), and bla (TEM-) (1). The analysis of plasmids carried by different isolates showed that Klebsiella pneumoniae might be an important multiple-replicon plasmid host. Plasmid skeleton and structure analyses showed that a multiple-replicon plasmid was formed by the fusion of two or more single plasmids, conferring strong adaptability to the antibiotic environment and continuously increasing the ability of drug-resistant isolates to spread around the world. In conclusion, multiple-replicon plasmids are better able to carry resistance genes than non-multiple-replicon plasmids, which may be an important mechanism underlying bacterial responses to environments with high-antibiotic pressure. This phenomenon will be highly significant for exploring bacterial resistance gene transmission and diffusion mechanisms in the future.202134777312
1988170.9996Different fosA genes were found on mobile genetic elements in Escherichia coli from wastewaters of hospitals and municipals in Turkey. AIMS: The increasing number of globally established fosfomycin-resistant (Fos(R)) Gram-negative bacteria inspired us to investigate the occurrence of Fos(R)Enterobacterales populations (esp. E. coli) in samples of city wastewater treatment plants (WWTPs) and hospital sewage in Hatay, Turkey. Fos(R) target bacteria were further characterized for their clonal relatedness, resistomes and mobile genetic elements (MGEs) to evaluate their impact on fosfomycin resistance dissemination. METHODS: A total of 44 samples from raw and treated waters of WWTPs as well as of two hospitals in the Hatay province were subjected to selective cultivation for recovering Fos(R)Enterobacterales. The presence of fosA was verified by PCR and Sanger amplicon sequencing. Detected E. coli were further evaluated against antimicrobial susceptibility-testing, macrorestriction profiling (PFGE) and whole-genome sequencing (WGS). Bioinformatics analysis was performed for genome subtyping (i.e., MLST, serotype), resistome/virulome determination and dissection of the genetic determinants of plasmidic fosA3/4 resistances. RESULTS: Besides ten non-E. coli Enterobacterales, 29 E. coli were collected within this study. In silico-based subtyping revealed that E. coli isolates were assigned to six different serovars and 14 sequence types (ST), while O8:H21 and ST410 represented the major prevalent types, respectively. Fosfomycin resistance in the isolates was found to be mediated by the fosA4 (n = 18), fosA3 (n = 10) and fosA (n = 1), which are frequently associated with transmissible MGEs. Reconstruction of plasmid-associated fosA gene context revealed a linkage between the resistance cassette and IS6 (IS26 family) transposases, which might represent a major driver for the distribution of the genes and the generation of novel fosA-carrying plasmids. CONCLUSIONS: The occurrence of plasmid-mediated, transmissible Fos(R) in E. coli from wastewater pose a foreseeable threat to "One-Health". To minimize further spread of the resistances in bacterial populations associated with environmental, animal and human health further resistance monitoring and management strategies must be developed.202235182630
1895180.9996Comparative Genome Analysis of Livestock and Human Colistin-Resistant Escherichia coli Isolates from the Same Household. BACKGROUND: Emergence and dissemination of colistin-resistant bacteria that harbor mobile colistin resistance (mcr) genes pose a dire challenge for the treatment of intractable infections caused by multidrug-resistant bacteria. Current findings on colistin-resistant bacteria in both humans and livestock of the same households highlight the need to identify the dissemination mechanisms of colistin-resistant bacteria. METHODS: In this study, a comparative genome analysis of colistin-resistant Escherichia coli isolates from livestock and humans of the same household was performed to clarify the possible dissemination mechanism of mcr genes among bacteria. Pulsed-field gel electrophoresis and whole-genome sequencing followed by sequence typing of the isolates were performed for assessment of the samples. RESULTS: The study revealed that two colistin-resistant E. coli isolates, one each from a pig and a chicken, were phylogenetically similar but not identical to the human isolates obtained from the same household. The comparative genome analysis revealed that the chicken isolate and a human isolate shared the same IncHl2 plasmid harboring the mcr transposon (mcr-1-PAP2). The pig isolate and the other human isolate retained the mcr-1 transposon on the chromosome, with the pig isolate carrying the complete mcr transposon (ISApl1-mcr-1-PAP2-ISApl1) and the human isolate carrying the incomplete mcr transposon (ISApl1-mcr-1-PAP2). CONCLUSION: The results of the study confirm the distribution of colistin-resistant bacteria and subsequent transmission of the resistance gene-carrying transposon between livestock and humans of the same household. To the best of our knowledge, this is the first report on genomic analysis of colistin-resistant E. coli isolates obtained from livestock and residents of the same household.202133688219
1907190.9996Nationwide surveillance of carbapenem-resistant Gram-negative pathogens in the Lebanese environment. Gram-negative ESKAPE pathogens with carbapenem resistance pose a significant health threat. Despite extensive research on the spread of these pathogens within Lebanese hospital settings, their emergence in environmental settings remains understudied. This study aimed to explore the environmental spread of carbapenem resistance among Gram-negative bacteria isolated from environmental samples in nine districts across Lebanon. A total of 250 samples were collected from wild animals, sewage, water, and soil between June 2022 and September 2023. Samples were streaked on MacConkey agar plates supplemented with 2 mg/L meropenem. Bacterial species were identified primarily using API20E. Antimicrobial susceptibility profiles were determined by the disk diffusion method and the Vitek 2 compact system. Meropenem-resistant Gram-negative bacteria were further characterized by whole-genome sequencing, and each of the bacterial species, sequence types, resistance genes, and plasmids was detected by sequence data analysis. We successfully isolated 130 carbapenem-resistant isolates from various samples, 67 of which belonged to the ESKAPE pathogens list and showed a multidrug-resistant (MDR) profile. The distribution of the latter was as follows: Escherichia coli (65.67%), Acinetobacter baumannii (16.42%), Pseudomonas aeruginosa (11.94%), and Klebsiella pneumoniae (5.97%). Several carbapenem resistance genes were detected, with a prevalence of blaNDM-5 in Escherichia coli and Klebsiella pneumoniae, blaIMP-1 and mexAB-OprM efflux pumps in Pseudomonas aeruginosa, and blaOXA-23 in Acinetobacter baumannii. Our findings revealed a widespread distribution of carbapenem-resistant ESKAPE bacteria in Lebanon, underscoring the significant public health risk posed by these pathogens. This highlights the urgent need to address the dissemination of antibiotic resistance in Lebanese environmental settings. IMPORTANCE: The emergence of antimicrobial resistance (AMR) extremely burdens public health and increases morbid and mortal threats in Lebanon. While the majority of the studies in our country target antimicrobial resistance in clinical settings, fewer studies focus on antimicrobial resistance dissemination in the environment. The significance of our research is that it sheds light on the environment as a less explored yet equally crucial sector in the spread of AMR. Here, we isolated carbapenemase-producing bacteria (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii) that were categorized as multidrug resistant (MDR) from diverse environmental sources in multiple provinces across Lebanon. The finding of carbapenem-resistant bacteria carrying plasmids represents a potential risk due to the possible spread of resistance genes via horizontal gene transfer across the environment and hospital settings. This highly recommends the implementation of regular surveillance to monitor the spread of antimicrobial resistance among environmental bacteria, which consequently leads to its spread within communities and thus poses a great threat to human health.202540492734