scholarly journals Linezolid resistance in Enterococcus faecium and Enterococcus faecalis from hospitalized patients in Ireland: high prevalence of the MDR genes optrA and poxtA in isolates with diverse genetic backgrounds

2020 ◽  
Vol 75 (7) ◽  
pp. 1704-1711 ◽  
Author(s):  
Sarah A Egan ◽  
Anna C Shore ◽  
Brian O’Connell ◽  
Grainne I Brennan ◽  
David C Coleman
Keyword(s):  
Enterococcus Faecalis ◽  
Resistance Genes ◽  
Enterococcus Faecium ◽  
Illumina Miseq ◽  
Conjugative Plasmid ◽  
Sequence Coverage ◽  
Linezolid Resistance ◽  
Oxford Nanopore ◽  
High Prevalence ◽  
Mdr Genes

Abstract Objectives To investigate the prevalence of the optrA, poxtA and cfr linezolid resistance genes in linezolid-resistant enterococci from Irish hospitals and to characterize associated plasmids. Methods One hundred and fifty-four linezolid-resistant isolates recovered in 14 hospitals between June 2016 and August 2019 were screened for resistance genes by PCR. All isolates harbouring resistance genes, and 20 without, underwent Illumina MiSeq WGS. Isolate relatedness was assessed using enterococcal whole-genome MLST. MinION sequencing (Oxford Nanopore) and hybrid assembly were used to resolve genetic environments/plasmids surrounding resistance genes. Results optrA and/or poxtA were identified in 35/154 (22.7%) isolates, the highest prevalence reported to date. Fifteen isolates with diverse STs harboured optrA only; one Enterococcus faecium isolate harboured optrA (chromosome) and poxtA (plasmid). Seven Enterococcus faecalis and one E. faecium harboured optrA on a 36 331 bp plasmid with 100% identity to the previously described optrA-encoding conjugative plasmid pE349. Variations around optrA were also observed, with optrA located on plasmids in five isolates and within the chromosome in three isolates. Nine E. faecium and 10 E. faecalis harboured poxtA, flanked by IS1216E, within an identical 4001 bp region on plasmids exhibiting 72.9%–100% sequence coverage to a 21 849 bp conjugative plasmid. E. faecalis isolates belonged to ST480, whereas E. faecium isolates belonged to diverse STs. Of the remaining 119 linezolid-resistant isolates without linezolid resistance genes, 20 investigated representatives all harboured the G2576T 23S RNA gene mutation associated with linezolid resistance. Conclusions This high prevalence of optrA and poxtA in diverse enterococcal lineages in Irish hospitals indicates significant selective pressure(s) for maintenance.

scholarly journals Detection of Oxazolidinone Resistance Genes and Characterization of Genetic Environments in Enterococci of Swine Origin, Italy

2020 ◽  
Vol 8 (12) ◽  
pp. 2021
Author(s):  
Simona Fioriti ◽  
Gianluca Morroni ◽  
Sonia Nina Coccitto ◽  
Andrea Brenciani ◽  
Alberto Antonelli ◽  
...  
Keyword(s):  
Resistance Gene ◽  
Enterococcus Faecalis ◽  
Resistance Genes ◽  
Enterococcus Faecium ◽  
Central Italy ◽  
Great Variability ◽  
Linezolid Resistance ◽  
Pig Farms ◽  
Clonality Analysis

One hundred forty-five florfenicol-resistant enterococci, isolated from swine fecal samples collected from 76 pig farms, were investigated for the presence of optrA, cfr, and poxtA genes by PCR. Thirty florfenicol-resistant Enterococcus isolates had at least one linezolid resistance gene. optrA was found to be the most widespread linezolid resistance gene (23/30), while cfr and poxtA were detected in 6/30 and 7/30 enterococcal isolates, respectively. WGS analysis also showed the presence of the cfr(D) gene in Enterococcus faecalis (n = 2 isolates) and in Enterococcus avium (n = 1 isolate). The linezolid resistance genes hybridized both on chromosome and plasmids ranging from ~25 to ~240 kb. Twelve isolates were able to transfer linezolid resistance genes to enterococci recipient. WGS analysis displayed a great variability of optrA genetic contexts identical or related to transposons (Tn6628 and Tn6674), plasmids (pE035 and pWo27-9), and chromosomal regions. cfr environments showed identities with Tn6644-like transposon and a region from p12-2300 plasmid; cfr(D) genetic contexts were related to the corresponding region of the plasmid 4 of Enterococcus faecium E8014; poxtA was always found on Tn6657. Circular forms were obtained only for optrA- and poxtA-carrying genetic contexts. Clonality analysis revealed the presence of E. faecalis (ST16, ST27, ST476, and ST585) and E. faecium (ST21) clones previously isolated from humans. These results demonstrate a dissemination of linezolid resistance genes in enterococci of swine origin in Central Italy and confirm the spread of linezolid resistance in animal settings.

Multiple-Antibiotic Resistance of Enterococcus faecalis and Enterococcus faecium from Cecal Contents in Broiler Chicken and Turkey Flocks Slaughtered in Canada and Plasmid Colocalization of tetO and ermB Genes

2011 ◽  
Vol 74 (10) ◽  
pp. 1639-1648 ◽  
Author(s):  
CINDY-LOVE TREMBLAY ◽  
ANN LETELLIER ◽  
SYLVAIN QUESSY ◽  
MARTINE BOULIANNE ◽  
DANIELLE DAIGNAULT ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Enterococcus Faecalis ◽  
Resistance Genes ◽  
Enterococcus Faecium ◽  
Broiler Chicken ◽  
Multidrug Resistant ◽  
Poultry Meat ◽  
Antimicrobial Resistance Genes ◽  
High Level ◽  
Level Resistance

This study was conducted to characterize the antimicrobial resistance determinants and investigate plasmid colocalization of tetracycline and macrolide genes in Enterococcus faecalis and Enterococcus faecium from broiler chicken and turkey flocks in Canada. A total of 387 E. faecalis and E. faecium isolates were recovered from poultry cecal contents from five processing plants. The percentages of resistant E. faecalis and E. faecium isolates, respectively, were 88.1 and 94% to bacitracin, 0 and 0.9% to chloramphenicol, 0.7 and 14.5% to ciprofloxacin, 72.6 and 80.3% to erythromycin, 3.7 and 41% to flavomycin, 9.6 and 4.3% (high-level resistance) to gentamicin, 25.2 and 17.1% (high-level resistance) to kanamycin, 100 and 94% to lincomycin, 0 and 0% to linezolid, 2.6 and 20.5% to nitrofurantoin, 3 and 27.4% to penicillin, 98.5 and 89.7% to quinupristin-dalfopristin, 7 and 12.8% to salinomycin, 46.7 and 38.5% (high-level resistance) to streptomycin, 95.6 and 89.7% to tetracycline, 73 and 75.2% to tylosin, and 0 and 0% to vancomycin. One predominant multidrug-resistant phenotypic pattern was identified in both E. faecalis and E. faecium (bacitracin, erythromycin, lincomycin, quinupristin-dalfopristin, tetracycline, and tylosin). These isolates were further examined by PCR and sequencing for the genes encoding their antimicrobial resistance. Various combinations of vatD, vatE, bcrR, bcrA, bcrB, bcrD, ermB, msrC, linB, tetM, and tetO genes were detected, and ermB, tetM, and bcrB were the most common antimicrobial resistance genes identified. For the first time, plasmid extraction and hybridization revealed colocalization of tetO and ermB genes on a ca. 11-kb plasmid in E. faecalis isolates, and filter mating experiments demonstrated its transferability. Results indicate that the intestinal enterococci of healthy poultry, which can contaminate poultry meat at slaughter, could be a reservoir for quinupristin-dalfopristin, bacitracin, tetracycline, and macrolide resistance genes.

scholarly journals Conjugative delivery of CRISPR-Cas9 for the selective depletion of antibiotic-resistant enterococci

2019 ◽  
Author(s):  
Marinelle Rodrigues ◽  
Sara W. McBride ◽  
Karthik Hullahalli ◽  
Kelli L. Palmer ◽  
Breck A. Duerkop
Keyword(s):  
Antibiotic Resistance ◽  
Enterococcus Faecalis ◽  
Resistance Genes ◽  
Bacterial Infections ◽  
Antibiotic Resistance Genes ◽  
Multidrug Resistant ◽  
Conjugative Plasmid ◽  
Antibiotic Resistant ◽  
Resistance Determinants

AbstractThe innovation of new therapies to combat multidrug-resistant (MDR) bacteria is being outpaced by the continued rise of MDR bacterial infections. Of particular concern are hospital-acquired infections (HAIs) recalcitrant to antibiotic therapies. The Gram-positive intestinal pathobiontEnterococcus faecalisis associated with HAIs and some strains are MDR. Therefore, novel strategies to controlE. faecalispopulations are needed. We previously characterized anE. faecalisType II CRISPR-Cas system and demonstrated its utility in the sequence-specific removal of antibiotic resistance determinants. Here we present work describing the adaption of this CRISPR-Cas system into a constitutively expressed module encoded on a pheromone-responsive conjugative plasmid that efficiently transfers toE. faecalisfor the selective removal of antibiotic resistance genes. Usingin vitrocompetition assays, we show that these CRISPR-Cas-encoding delivery plasmids, or CRISPR-Cas antimicrobials, can reduce the occurrence of antibiotic resistance in enterococcal populations in a sequence-specific manner. Furthermore, we demonstrate that deployment of CRISPR-Cas antimicrobials in the murine intestine reduces the occurrence of antibiotic-resistantE. faecalisby several orders of magnitude. Finally, we show thatE. faecalisdonor strains harboring CRISPR-Cas antimicrobials are immune to uptake of antibiotic resistance determinantsin vivo. Our results demonstrate that conjugative delivery of CRISPR-Cas antimicrobials may be adaptable for future deployment from probiotic bacteria for exact targeting of defined MDR bacteria or for precision engineering of polymicrobial communities in the mammalian intestine.ImportanceCRISPR-Cas nucleic acid targeting systems hold promise for the amelioration of multidrug-resistant enterococci, yet the utility of such tools in the context of the intestinal environment where enterococci reside is understudied. We describe the development of a CRISPR-Cas antimicrobial, deployed on a conjugative plasmid, for the targeted removal of antibiotic resistance genes from intestinalEnterococcus faecalis. We demonstrate that CRISPR-Cas targeting reduces antibiotic resistance ofE. faecalisby several orders of magnitude in the intestine. Although barriers exist that influence the penetrance of the conjugative CRISPR-Cas antimicrobial among target recipientE. faecaliscells, the removal of antibiotic resistance genes inE. faecalisupon uptake of the CRISPR-Cas antimicrobial is absolute. In addition, cells that obtain the CRISPR-Cas antimicrobial are immunized against the acquisition of new antibiotic resistance genes. This study suggests a potential path toward plasmid based CRISPR-Cas therapies in the intestine.

scholarly journals Detection of the Phenicol–Oxazolidinone Resistance Gene poxtA in Enterococcus faecium and Enterococcus faecalis from Food-Producing Animals during 2008–2018 in Korea

2020 ◽  
Vol 8 (11) ◽  
pp. 1839
Author(s):  
Seok-Hyeon Na ◽  
Dong-Chan Moon ◽  
Mi-Hyun Kim ◽  
Hee-Young Kang ◽  
Su-Jeong Kim ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Ribosomal Protein ◽  
Resistance Gene ◽  
Enterococcus Faecalis ◽  
Enterococcus Faecium ◽  
Gel Electrophoresis ◽  
Pulsed Field Gel Electrophoresis ◽  
Molecular Characteristics ◽  
Pfge Analysis ◽  
Linezolid Resistance

We aimed to investigate the presence of the phenicol–oxazolidinone resistance gene poxtA in linezolid-resistant enterococci from food-producing animals and analyze its molecular characteristics. We collected 3941 Enterococcus faecium and 5088 E. faecalis isolates from all provinces of South Korea from 2008 to 2018. We found linezolid resistance in 0.79% (94/3941) of E. faecium and 1.22% (62/5088) of E. faecalis isolates. Overall, 23.1% (36/156) of the linezolid-resistant isolates had the poxtA gene, including 31 E. faecium and five E. faecalis isolates. The poxtA-positive enterococci were mainly isolated from chicken (86.1%; 26/36). Fifteen poxtA-harboring isolates co-carried another linezolid-resistance gene, optrA. Eight E. faecium isolates had an N130K mutation in the ribosomal protein L4, while no mutations were observed in E. faecalis isolates. The poxtA gene was transferred into 10 enterococci by conjugation. Multi-locus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) analysis indicated that poxtA-carrying isolates were heterogeneous. Three E. faecium isolates belonged to CC17 (ST32, ST121, and ST491). To our knowledge, this is the first report on the poxtA gene in Korea. Prudent use of antimicrobials and active surveillance on antimicrobial resistance are urgently needed to reduce the risk of dissemination of the linezolid-resistant isolates in humans and animals.

scholarly journals Gene Dosage and Linezolid Resistance in Enterococcus faecium and Enterococcus faecalis

2002 ◽  
Vol 46 (10) ◽  
pp. 3334-3336 ◽  
Author(s):  
S. H. Marshall ◽  
C. J. Donskey ◽  
R. Hutton-Thomas ◽  
R. A. Salata ◽  
L. B. Rice
Keyword(s):  
Enterococcus Faecalis ◽  
Enterococcus Faecium ◽  
Gene Dosage ◽  
Clinical Isolates ◽  
Rrna Genes ◽  
23S Rrna ◽  
Linezolid Resistance ◽  
Clear Association ◽  
23S Rrna Genes ◽  
Domain V

ABSTRACT Resistance to linezolid has been associated with a G2576U mutation in domain V of the 23S rRNA. We analyzed nine clinical isolates of linezolid-resistant enterococci and showed a clear association between the number of 23S rRNA genes containing this mutation and the level of linezolid resistance expressed.

scholarly journals Linezolid Resistance in Vancomycin-Resistant Enterococcus faecalis and Enterococcus faecium Isolates in a Brazilian Hospital

2014 ◽  
Vol 58 (5) ◽  
pp. 2993-2994 ◽  
Author(s):  
L. M. de Almeida ◽  
M. R. E. de Araujo ◽  
M. F. Iwasaki ◽  
A. G. Sacramento ◽  
D. Rocha ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Enterococcus Faecium ◽  
Vancomycin Resistant Enterococcus ◽  
Linezolid Resistance ◽  
Vancomycin Resistant

scholarly journals Phenotypic and genotypic characterization of linezolid-resistant Enterococcus faecium from the USA and Pakistan

2019 ◽  
Vol 74 (12) ◽  
pp. 3445-3452 ◽  
Author(s):  
Kate E Wardenburg ◽  
Robert F Potter ◽  
Alaric W D’Souza ◽  
Tahir Hussain ◽  
Meghan A Wallace ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Enterococcus Faecium ◽  
Antibiotic Resistance Genes ◽  
23S Rrna ◽  
Linezolid Resistance ◽  
Geographical Regions ◽  
The Us ◽  
The Usa ◽  
Acquired Antibiotic Resistance

Abstract Objectives Linezolid is an important therapeutic option for the treatment of infections caused by VRE. Linezolid is a synthetic antimicrobial and resistance to this antimicrobial agent remains relatively rare. As a result, data on the comparative genomics of linezolid resistance determinants in Enterococcus faecium are relatively sparse. Methods To address this knowledge gap in E. faecium, we deployed phenotypic antibiotic susceptibility testing and Illumina WGS on hospital surface (environmental) and clinical isolates from the USA and Pakistan. Results We found complete concordance between isolate source country and mechanism of linezolid resistance, with all the US isolates possessing a 23S rRNA gene mutation and the Pakistan isolates harbouring two to three acquired antibiotic resistance genes. These resistance genes include the recently elucidated efflux-pump genes optrA and poxtA and a novel cfr-like variant. Although there was no difference in the linezolid MIC between the US and Pakistan isolates, there was a significant difference in the geometric mean of the MIC between the Pakistan isolates that had two versus three of the acquired antibiotic resistance genes. In five of the Pakistan E. faecium that possessed all three of the resistance genes, we found no difference in the local genetic context of poxtA and the cfr-like gene, but we identified different genetic contexts surrounding optrA. Conclusions These results demonstrate that E. faecium from different geographical regions employ alternative strategies to counter selective pressure of increasing clinical linezolid use.

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