Prediction of antimicrobial resistance in clinical Enterococcus faecium isolates using a rules-based analysis of whole genome sequences

2021 ◽  
Author(s):  
Melis N. Anahtar ◽  
Juliet T. Bramante ◽  
Jiawu Xu ◽  
Lisa A. Desrosiers ◽  
Jeffrey M. Paer ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Predictive Value ◽  
Enterococcus Faecium ◽  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Clinical Isolates ◽  
Automated System ◽  
Whole Genome ◽  
Prediction Rules ◽  
Phenotypic Susceptibility

Background: Enterococcus faecium is a major cause of clinical infections, often due to multidrug-resistant (MDR) strains. Whole genome sequencing (WGS) is a powerful tool to study MDR bacteria and their antimicrobial resistance (AMR) mechanisms. Here we use WGS to characterize E. faecium clinical isolates and test the feasibility of rules-based genotypic prediction of AMR. Methods: Clinical isolates were divided into derivation and validation sets. Phenotypic susceptibility testing for ampicillin, vancomycin, high-level gentamicin, ciprofloxacin, levofloxacin, doxycycline, tetracycline, and linezolid was performed using the VITEK 2 automated system, with confirmation and discrepancy resolution by broth microdilution, disk diffusion, or gradient diffusion when needed. WGS was performed to identify isolate lineage and AMR genotype. AMR prediction rules were derived by analyzing the genotypic-phenotypic relationship in the derivation set. Results: Phylogenetic analysis demonstrated that 88% of isolates in the collection belonged to hospital-associated clonal complex 17. Additionally, 12% of isolates had novel sequence types. When applied to the validation set, the derived prediction rules demonstrated an overall positive predictive value of 98% and negative predictive value of 99% compared to standard phenotypic methods. Most errors were falsely resistant predictions for tetracycline and doxycycline. Further analysis of genotypic-phenotypic discrepancies revealed potentially novel pbp5 and tet (M) alleles that provide insight into ampicillin and tetracycline class resistance mechanisms. The prediction rules demonstrated generalizability when tested on an external dataset. Conclusions: Known AMR genes and mutations can predict E. faecium phenotypic susceptibility with high accuracy for most routinely tested antibiotics, providing opportunities for advancing molecular diagnostics.

scholarly journals Antimicrobial Susceptibility and Clonality of Vaginally Derived Multidrug-Resistant Mobiluncus Isolates in China

2020 ◽  
Vol 64 (8) ◽  
Author(s):  
Xueying Zhang ◽  
Yongying Bai ◽  
Long Zhang ◽  
Mohamed S. Draz ◽  
Zhi Ruan ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Antimicrobial Resistance ◽  
Transposable Element ◽  
Genome Sequencing ◽  
Antimicrobial Susceptibility ◽  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Whole Genome ◽  
Content Type ◽  
High Diversity

ABSTRACT Here, the antimicrobial susceptibility, resistance mechanisms, and clonality of Mobiluncus sp. isolates recovered from gynecological outpatients in China were investigated. Compared to M. mulieris, M. curtisii exhibited higher antimicrobial resistance to metronidazole, clindamycin, and tetracycline. Whole-genome sequencing indicated that the clindamycin resistance gene erm(X) was located on a transposable element, Tn5432, which was composed of two IS1249 sequences. Phylogenetic analysis indicated that Mobiluncus spp. had high diversity, with isolates being grouped into several sporadic clades.

scholarly journals 1459. Whole Genome Sequencing Analysis of Enterococcus faecium Clinical Isolates Reveals High Strain Diversity and High Accuracy Prediction of Antimicrobial Resistance

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S731-S732
Author(s):  
Melis N Anahtar ◽  
Juliet Bramante ◽  
Jiawu Xu ◽  
Lisa Desrosiers ◽  
Eric Rosenberg ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Antimicrobial Susceptibility ◽  
Predictive Value ◽  
Enterococcus Faecium ◽  
High Accuracy ◽  
Whole Genome ◽  
Strain Diversity ◽  
Validation Set

Abstract Background Whole genome sequencing (WGS) is a powerful tool to uncover transmission patterns and antimicrobial resistance (AMR) mechanisms of Enterococcus faecium, a major cause of hospital-acquired infections. Most E. faecium genomic studies include isolates from outbreak investigations rather than routine sampling. Additionally, the use of WGS to predict E. faecium AMR has not been tested systematically. Here we use WGS to characterize over 400 E. faecium clinical isolates to assess their strain diversity and AMR mechanisms. Methods Clinical E. faecium isolates from the MGH Microbiology Laboratory were collected at random from 1/2016-12/2017 (derivation set; 193 isolates) and with enrichment for more resistant isolates from 1/2018-9/2019 (validation set; 226 isolates). Species identification was performed using the bioMérieux VITEK MS instrument. Susceptibility testing was performed using the AST-GP75 card (bioMérieux VITEK 2), with confirmation by disk diffusion or ETEST when needed. Bacterial DNA from isolates was extracted, purified, sequenced (Illumina NextSeq), and quality filtered. Samples with >20x genome coverage were analyzed with SRST2 and AliView. Results MLST analysis of the derivation set demonstrated strikingly high diversity compared to previously published studies, with the three most frequent types (ST412, ST18, ST736) comprising fewer than half of samples. We identified and confirmed four novel MLST types comprising 12% of samples. We next analyzed the derivation isolate set to determine which genes and SNPs, if applicable, predicted resistance to seven antibiotics routinely tested at our institution: ampicillin, ciprofloxacin, doxycycline, high-level gentamicin, levofloxacin, tetracycline, and vancomycin. These rules were uniformly applied to the validation isolate set and demonstrated that genotypic AMR prediction has an overall positive predictive value of 97.0% and negative predictive value of 97.1% compared to standard susceptibility methods. Table 1. Summary of validation set predictions of antimicrobial susceptibility based on defined genotypic features. * The intermediate category is considered with the susceptible category. Conclusion In a diverse and challenging set of clinical E. faecium isolates, known AMR genes and SNPs can be simply applied to predict phenotypic susceptibility with high accuracy for seven routinely tested antibiotics. Further testing will be performed to resolve phenotype-genotype discrepancies. Summary of validation set predictions of antimicrobial susceptibility based on defined genotypic features. * The intermediate category is considered with the susceptible category. Disclosures Melis N. Anahtar, MD, PhD, Day Zero Diagnostics (Other Financial or Material Support, Co-founder, consultant, equity holder) Virginia M. Pierce, MD, Selux Diagnostics, Inc. (Grant/Research Support) Douglas S. Kwon, MD, PhD, Day Zero Diagnostics (Consultant, Shareholder, Other Financial or Material Support, co-founder)

scholarly journals 1280. In-Vitro Activity of Cefiderocol, Imipenem/Relebactam, & Ceftazidime/Avibactam in Ceftolozane/Tazobactam Resistant Strains of Multidrug Resistant Pseudomonas aeruginosa

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S655-S655
Author(s):  
Daniel Navas ◽  
Angela Charles ◽  
Amy Carr ◽  
Jose Alexander
Keyword(s):  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Vitro Activity ◽  
Clinical Isolates ◽  
Resistance Testing ◽  
Clinical Samples ◽  
In Vitro Activity ◽  
Carbapenemase Gene ◽  
Resistant Strains

Abstract Background The activity of imipenem/relebactam (I/R), ceftazidime/avibactam (CZA) and cefiderocol (FDC) were evaluated against clinical isolates of multidrug resistant (MDR) strains of P. aeruginosa which was resistant to ceftolozane/tazobactam (C/T). The recent increase of MDR P. aeruginosa strains isolated from clinical samples has prompted research and development of new antimicrobials that can withstand its multiple resistance mechanisms. C/T is an effective option for treatment of MDR P. aeruginosa in our facility with only 10% of resistance in MDR strains, but the emergence of resistance may occur due to the presence of a carbapenemase gene or an ampC mutation. Methods Antimicrobial susceptibility testing for C/T Etest® (bioMérieux, Inc.) were performed on all MDR strains initially screened by the VITEK2® (bioMérieux, Inc.). 10% (n=20) of all MDR isolates were resistant to C/T by the CLSI 2019 breakpoints. These resistant isolates were tested for presence of a carbapenemase gene using the GeneXpert CARBA-R (Cepheid®) PCR and against CZA Etest® (bioMérieux, Inc.) I/R gradient strips (Liofilchem®) and FDC broth microdilution (Thermo Scientific™ Sensititre™). Results A total of 20 clinical isolates of MDR P. aeruginosa resistant to C/T were tested following standardized CLSI protocols and techniques. All 20 isolates were screened for the presence of a carbapenemase gene (blaVIM, blaNDM, blaKPC, blaOXA-48, blaIMP). A blaVIM gene was detected in 6 (30%) out of 20 isolates. FDC demonstrated the greatest activity with 85% (n=17) of susceptible isolates (CLSI MIC <4µg/dL). CZA (CLSI MIC <8µg/dL) and I/R (FDA MIC <2µg/dL) showed 15% (n=3) and 10% (n=2) of susceptible isolates respectively. FDC was active against all 6 blaVIM isolates, where all 6 strains were resistant to CZA and I/R as expected. 3 isolates tested non-susceptible against FDC; additional characterization was not performed at this time. Conclusion Based on these results, FDC demonstrated the greatest in-vitro activity against C/T resistant strains of MDR P. aeruginosa. FDC also demonstrated activity against all 6 MDR P. aeruginosa carrying blaVIM gene. FDC is a strong option to consider on MDR P. aeruginosa strains based on a resistance testing algorithm and a cost/effective protocol. Disclosures All Authors: No reported disclosures

scholarly journals Staphylococcus aureus Infections in Malaysia: A Review of Antimicrobial Resistance and Characteristics of the Clinical Isolates, 1990–2017

Antibiotics ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 128 ◽  
Author(s):  
Ainal Mardziah Che Hamzah ◽  
Chew Chieng Yeo ◽  
Suat Moi Puah ◽  
Kek Heng Chua ◽  
Ching Hoong Chew
Keyword(s):  
Staphylococcus Aureus ◽  
Antimicrobial Resistance ◽  
Sccmec Type ◽  
Epidemiological Data ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Type Iv ◽  
Resistant Strains ◽  
Vancomycin Resistant ◽  
Comprehensive Surveillance

Staphylococcus aureus is an important nosocomial pathogen and its multidrug resistant strains, particularly methicillin-resistant S. aureus (MRSA), poses a serious threat to public health due to its limited therapeutic options. The increasing MRSA resistance towards vancomycin, which is the current drug of last resort, gives a great challenge to the treatment and management of MRSA infections. While vancomycin resistance among Malaysian MRSA isolates has yet to be documented, a case of vancomycin resistant S. aureus has been reported in our neighboring country, Indonesia. In this review, we present the antimicrobial resistance profiles of S. aureus clinical isolates in Malaysia with data obtained from the Malaysian National Surveillance on Antimicrobial Resistance (NSAR) reports as well as various peer-reviewed published records spanning a period of nearly three decades (1990–2017). We also review the clonal types and characteristics of Malaysian S. aureus isolates, where hospital-associated (HA) MRSA isolates tend to carry staphylococcal cassette chromosome mec (SCCmec) type III and were of sequence type (ST)239, whereas community-associated (CA) isolates are mostly SCCmec type IV/V and ST30. More comprehensive surveillance data that include molecular epidemiological data would enable further in-depth understanding of Malaysian S. aureus isolates.

scholarly journals Mechanisms of Antimicrobial Resistance in ESKAPE Pathogens

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Sirijan Santajit ◽  
Nitaya Indrawattana
Keyword(s):  
Public Health ◽  
Antimicrobial Resistance ◽  
Antimicrobial Agents ◽  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Global Public Health ◽  
Drug Usage ◽  
Inappropriate Use ◽  
Public Health Challenges ◽  
Eskape Pathogens

The ESKAPE pathogens (Enterococcus faecium,Staphylococcus aureus,Klebsiella pneumoniae,Acinetobacter baumannii,Pseudomonas aeruginosa, andEnterobacterspecies) are the leading cause of nosocomial infections throughout the world. Most of them are multidrug resistant isolates, which is one of the greatest challenges in clinical practice. Multidrug resistance is amongst the top three threats to global public health and is usually caused by excessive drug usage or prescription, inappropriate use of antimicrobials, and substandard pharmaceuticals. Understanding the resistance mechanisms of these bacteria is crucial for the development of novel antimicrobial agents or other alternative tools to combat these public health challenges. Greater mechanistic understanding would also aid in the prediction of underlying or even unknown mechanisms of resistance, which could be applied to other emerging multidrug resistant pathogens. In this review, we summarize the known antimicrobial resistance mechanisms of ESKAPE pathogens.

scholarly journals Rapid in vivo development of resistance to daptomycin in vancomycin-resistant Enterococcus faecium due to genomic rearrangements

2021 ◽  
Author(s):  
Sarah Mollerup ◽  
Christine Elmeskov ◽  
Heidi Gumpert ◽  
Mette Pinholt ◽  
Tobias Steen Sejersen ◽  
...  
Keyword(s):  
Cell Wall ◽  
Enterococcus Faecium ◽  
Chromosomal Rearrangements ◽  
Resistance Mechanisms ◽  
Resistant Isolate ◽  
Wall Structure ◽  
Whole Genome ◽  
Cyclic Lipopeptide ◽  
Vancomycin Resistant

AbstractBackgroundDaptomycin is a cyclic lipopeptide used in the treatment of vancomycin-resistant Enterococcus faecium (VREfm). However, the development of daptomycin-resistant VREfm challenges the treatment of nosocomial VREfm infections. Resistance mechanisms of daptomycin are not fully understood. Here we analysed the genomic changes leading to a daptomycin-susceptible VREfm isolate becoming resistant after 40 days of daptomycin and linezolid combination therapy.MethodsThe two isogenic VREfm isolates (daptomycin-susceptible and daptomycin-resistant) were analysed using whole genome sequencing with Illumina and Nanopore.ResultsWhole genome comparative analysis identified the loss of a 46.5 kb fragment and duplication of a 29.7 kb fragment in the daptomycin-resistant isolate, with many implicated genes involved in cell wall synthesis. Two plasmids of the daptomycin-susceptible isolate were also found integrated in the chromosome of the resistant isolate. One nonsynonymous SNP in the rpoC gene was identified in the daptomycin-resistant isolate.ConclusionsDaptomycin resistance developed through chromosomal rearrangements leading to altered cell wall structure. Such novel types of resistance mechanisms can only be identified by comparing closed genomes of isogenic isolates.

scholarly journals Retrospective Analysis on Antimicrobial Resistance Trends and Prevalence of β-lactamases in Escherichia coli and ESKAPE Pathogens Isolated from Arabian Patients during 2000–2020

2020 ◽  
Vol 8 (10) ◽  
pp. 1626
Author(s):  
Mahfouz Nasser ◽  
Snehal Palwe ◽  
Ram Naresh Bhargava ◽  
Marc G. J. Feuilloley ◽  
Arun S. Kharat
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Antimicrobial Resistance ◽  
Enterococcus Faecium ◽  
Arab Countries ◽  
Clinical Isolates ◽  
Arab Region ◽  
E Coli ◽  
Extended Spectrum ◽  
Eskape Pathogens

The production of diverse and extended spectrum β-lactamases among Escherichia coli and ESKAPE pathogens is a growing threat to clinicians and public health. We aim to provide a comprehensive analysis of evolving trends of antimicrobial resistance and β-lactamases among E. coli and ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acine to bacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) in the Arabian region. A systematic review was conducted in Medline PubMed on papers published between January 2000 and February 2020 on countries in the Arab region showing different antibiotic resistance among E. coli and ESKAPE pathogens. A total of n = 119,144 clinical isolates were evaluated for antimicrobial resistance in 19 Arab countries. Among these clinical isolates, 74,039 belonged to E. coli and ESKAPE pathogen. Distribution of antibiotic resistance among E. coli and ESKAPE pathogens indicated that E. coli (n = 32,038) was the predominant pathogen followed by K. pneumoniae (n = 17,128), P. aeruginosa (n = 11,074), methicillin-resistant S. aureus (MRSA, n = 4370), A. baumannii (n = 3485) and Enterobacter spp. (n = 1574). There were no reports demonstrating Enterococcus faecium producing β-lactamase. Analyses revealed 19 out of 22 countries reported occurrence of ESBL (Extended-Spectrum β-Lactamase) producing E. coli and ESKAPE pathogens. The present study showed significantly increased resistance rates to various antimicrobial agents over the last 20 years; for instance, cephalosporin resistance increased from 37 to 89.5%, fluoroquinolones from 46.8 to 70.3%, aminoglycosides from 40.2 to 64.4%, mono-bactams from 30.6 to 73.6% and carbapenems from 30.5 to 64.4%. An average of 36.9% of the total isolates were reported to have ESBL phenotype during 2000 to 2020. Molecular analyses showed that among ESBLs and Class A and Class D β-lactamases, blaCTX-M and blaOXA have higher prevalence rates of 57% and 52.7%, respectively. Among Class B β-lactamases, few incidences of blaVIM 27.7% and blaNDM 26.3% were encountered in the Arab region. Conclusion: This review highlights a significant increase in resistance to various classes of antibiotics, including cephalosporins, β-lactam and β-lactamase inhibitor combinations, carbapenems, aminoglycosides and quinolones among E. coli and ESKAPE pathogens in the Arab region.

scholarly journals Association of Alterations in ParC and GyrA Proteins with Resistance of Clinical Isolates of Enterococcus faecium to Nine Different Fluoroquinolones

1999 ◽  
Vol 43 (10) ◽  
pp. 2513-2516 ◽  
Author(s):  
Sylvain Brisse ◽  
Ad C. Fluit ◽  
Ulrich Wagner ◽  
Peter Heisig ◽  
Dana Milatovic ◽  
...  
Keyword(s):  
Enterococcus Faecium ◽  
Resistance Mechanisms ◽  
Clinical Isolates

ABSTRACT The parC and gyrA genes of 73 ciprofloxacin-resistant and 6 ciprofloxacin-susceptibleEnterococcus faecium clinical isolates were partly sequenced. Alterations in ParC and GyrA, possibly in combination with other resistance mechanisms, severely restricted the in vitro activities of the nine quinolones tested. For all isolates, clinafloxacin and sitafloxacin showed the best activities.

scholarly journals Integrated Genome-Wide Analysis of an Isogenic Pair of Pseudomonas aeruginosa Clinical Isolates with Differential Antimicrobial Resistance to Ceftolozane/Tazobactam, Ceftazidime/Avibactam, and Piperacillin/Tazobactam

2020 ◽  
Vol 21 (3) ◽  
pp. 1026 ◽  
Author(s):  
Weihua Huang ◽  
Joelle El Hamouche ◽  
Guiqing Wang ◽  
Melissa Smith ◽  
Changhong Yin ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Pseudomonas Aeruginosa ◽  
Antimicrobial Resistance ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Epigenetic Mechanism ◽  
Phenotypic Change ◽  
Genome Wide ◽  
Family Protein ◽  
Significant Difference

Multidrug-resistant (MDR) Pseudomonas aeruginosa is one of the main causes of morbidity and mortality in hospitalized patients and the leading cause of nosocomial infections. We investigated, here, two MDR P. aeruginosa clinical isolates from a hospitalized patient with differential antimicrobial resistance to ceftazidime/avibactam (CZA), ceftolozane/tazobactam (C/T), and piperacillin/tazobactam (P/T). Their assembled complete genomes revealed they belonged to ST235, a widespread MDR clone; and were isogenic with only a single nucleotide variant, causing G183D mutation in AmpC β-lactamase, responsible for a phenotypic change from susceptible to resistant to CZA and C/T. Further epigenomic profiling uncovered two conserved DNA methylation motifs targeted by two distinct putative methyltransferase-containing restriction-modification systems, respectively; more intriguingly, there was a significant difference between the paired isolates in the pattern of genomic DNA methylation and modifications. Moreover, genome-wide gene expression profiling demonstrated the inheritable genomic methylation and modification induced 14 genes being differentially regulated, of which only toxR (downregulated), a regulatory transcription factor, had its promoter region differentially methylate and modified. Since highly expressed opdQ encodes an OprD porin family protein, therefore, we proposed an epigenetic regulation of opdQ expression pertinent to the phenotypic change of P. aeruginosa from resistant to susceptible to P/T. The disclosed epigenetic mechanism controlling phenotypic antimicrobial resistance deserves further experimental investigation.

scholarly journals The Resistome of Pseudomonas aeruginosa in Relationship to Phenotypic Susceptibility

2014 ◽  
Vol 59 (1) ◽  
pp. 427-436 ◽  
Author(s):  
Veronica N. Kos ◽  
Maxime Déraspe ◽  
Robert E. McLaughlin ◽  
James D. Whiteaker ◽  
Paul H. Roy ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Genetic Determinants ◽  
Content Type ◽  
Susceptibility Data ◽  
Next Generation Sequencing Ngs ◽  
Sequence Types ◽  
Generation Sequencing ◽  
Phenotypic Susceptibility

ABSTRACTMany clinical isolates ofPseudomonas aeruginosacause infections that are difficult to eradicate due to their resistance to a wide variety of antibiotics. Key genetic determinants of resistance were identified through genome sequences of 390 clinical isolates ofP. aeruginosa, obtained from diverse geographic locations collected between 2003 and 2012 and were related to microbiological susceptibility data for meropenem, levofloxacin, and amikacin. β-Lactamases and integron cassette arrangements were enriched in the established multidrug-resistant lineages of sequence types ST111 (predominantly O12) and ST235 (O11). This study demonstrates the utility of next-generation sequencing (NGS) in defining relevant resistance elements and highlights the diversity of resistance determinants withinP. aeruginosa. This information is valuable in furthering the design of diagnostics and therapeutics for the treatment ofP. aeruginosainfections.

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