scholarly journals Enzymatic and Structural Characterization of Alanine Racemase from Enterococcus faecium by Kinetic and Computational Studies

2021 ◽  
Vol 18 (1) ◽  
pp. 3-12
Author(s):  
Arie Van Wieren ◽  
Emma Bouchard ◽  
Sudipta Majumdar
Keyword(s):  
Steady State ◽  
Homology Modeling ◽  
Enterococcus Faecium ◽  
Catalytic Efficiency ◽  
Homology Model ◽  
Alanine Racemase ◽  
Active Site Residues ◽  
Ph Optimum ◽  
Vancomycin Resistant Enterococci ◽  
Vancomycin Resistant

The surge in vancomycin-resistant enterococci (VRE) strains poses a serious threat to public and clinical health. VRE strains are the leading cause of multi-drug resistant enterococcal infections and are commonly acquired from medical devices. Therefore, it is essential to discover new antibacterial targets and drugs for this pathogen. Alanine racemase could be a valuable drug target due to its crucial role in E. faecium survival. Alr from E. faecium (EF_Alr) was heterologously produced and purified from E. coli., and the steady-state kinetic constants were determined at different pH values. Using a coupled reaction with L-alanine dehydrogenase, rate of production of NADH was measured at 340 nm to observe EF_Alr activity in the D- to L-alanine direction. The highest catalytic efficiency, 8.61 ± 0.5 s-1 mM-1, was found at pH 9. Additionally, the tentative active site residues, Lys40 and Tyr268, for the alanine racemization reaction were assigned by homology modeling and sequence comparison studies. Using UCSF Chimera, the structure of the EF_Alr homology model was superimposed and compared to the crystal structure of Alr from E. faecalis. KEYWORDS: Alanine Racemase; Enterococcus faecium; Vancomycin-Resistant Enterococci; Homology Modeling; pH Optimum; Kinetic Characterization; SWISS-MODEL Server; Steady-State Kinetics

scholarly journals Evaluation of a vanA-Specific PCR Assay for Detection of Vancomycin-Resistant Enterococcus faecium during a Hospital Outbreak

1999 ◽  
Vol 37 (10) ◽  
pp. 3348-3349 ◽  
Author(s):  
Michel Roger ◽  
Marie-Claude Faucher ◽  
Pierre Forest ◽  
Pierre St-Antoine ◽  
François Coutlée
Keyword(s):  
Enterococcus Faecium ◽  
Pcr Assay ◽  
Culture Methods ◽  
Agar Culture ◽  
Enrichment Broth ◽  
Specific Pcr ◽  
Hospital Outbreak ◽  
Vancomycin Resistant Enterococci ◽  
Vancomycin Resistant ◽  
Specific Pcr Assay

We investigated the use of PCR as an alternative to culture of fecal samples for detection of vanA-containingEnterococcus faecium during a recent hospital outbreak. Rectal swabs collected consecutively from 223 patients were analyzed by culture with and without enrichment broth and byvanA-specific PCR of enrichment broth samples. Fifty-five specimens were positive for vanA-containing E. faecium by at least one method. The sensitivities of thevanA-specific PCR assay and agar culture with and without enrichment broth were 94.5, 98, and 89%, respectively. All three methods were 100% specific. Final results were obtained much more rapidly by PCR (within 24 to 30 h of specimen submission) than by the culture methods (4 to 5 days). Thus, PCR is an accurate and rapid alternative to culture for detection of vancomycin-resistant enterococci during hospital outbreaks.

scholarly journals Draft Genome Sequence of a Vancomycin-Resistant and Vancomycin-Dependent Enterococcus faecium Isolate

2016 ◽  
Vol 4 (2) ◽  
Author(s):  
Marion Blaschitz ◽  
Sarah Lepuschitz ◽  
Laura Wagner ◽  
Franz Allerberger ◽  
Alexander Indra ◽  
...  
Keyword(s):  
Genome Sequence ◽  
Enterococcus Faecium ◽  
Prolonged Exposure ◽  
Draft Genome ◽  
Vancomycin Resistance ◽  
Draft Genome Sequence ◽  
Nosocomial Pathogens ◽  
Vancomycin Resistant Enterococci ◽  
Vancomycin Resistant ◽  
Faecium Isolate

Vancomycin-resistant enterococci have emerged as major nosocomial pathogens worldwide. While antimicrobial pressure promotes nosocomial colonization with these enterococci, prolonged exposure to vancomycin may foster the transition from vancomycin resistance to vancomycin dependence. Here, we report the draft genome sequence of a vancomycin-dependent Enterococcus faecium isolate showing partial teicoplanin dependence.

scholarly journals High Prevalence of VanA-Type Vancomycin-Resistant Enterococci in Austrian Poultry

2005 ◽  
Vol 71 (10) ◽  
pp. 6407-6409 ◽  
Author(s):  
Alexandra Eisner ◽  
Gebhard Feierl ◽  
Gregor Gorkiewicz ◽  
Franz Dieber ◽  
Harald H. Kessler ◽  
...  
Keyword(s):  
Enterococcus Faecium ◽  
Broiler Chickens ◽  
Carriage Rate ◽  
Fecal Samples ◽  
Vancomycin Resistant Enterococci ◽  
High Prevalence ◽  
Vancomycin Resistant

ABSTRACT Fecal samples from humans and food-producing animals were analyzed for the presence of vancomycin-resistant enterococci (VRE). The VRE carriage rate in humans was 6%, and there was a predominance of VanC-type resistance. Enterococcus faecium with vanA-mediated resistance was frequent in broiler chickens (42%) but rare in cattle and pig samples.

scholarly journals Molecular epidemiology of vancomycin-resistant Enterococcus faecium strains isolated from haematological malignancy patients in a research hospital in Turkey

2010 ◽  
Vol 59 (6) ◽  
pp. 660-664 ◽  
Author(s):  
S. Kirdar ◽  
A. G. Sener ◽  
U. Arslan ◽  
S. G. Yurtsever
Keyword(s):  
Enterococcus Faecium ◽  
Haematological Malignancy ◽  
Research Hospital ◽  
Vancomycin Resistant Enterococci ◽  
Time Period ◽  
Genes Encoding ◽  
Resistance Patterns ◽  
Training And Research ◽  
Vancomycin Resistant ◽  
Short Time

Infections and outbreaks of vancomycin-resistant enterococci (VRE) still appear to be rare in Turkey. In the present study, VRE strains isolated during an outbreak in a haematology unit of a training and research hospital in Turkey were typed and their antimicrobial-resistance patterns were characterized by molecular methods. Twelve vancomycin-resistant Enterococcus faecium strains isolated from patients with haematological malignancies were investigated by PCR for the presence of genes encoding resistance to vancomycin, tetracycline, chloramphenicol, gentamicin and erythromycin. Their clonal relationship was evaluated by PFGE and multilocus sequence typing. All strains were resistant to vancomycin and erythromycin, and had the vanA and ermB genes, respectively. PFGE was used to determine the presence of two pulsotypes and determine their subtypes. Pulsotype A belonged to sequence type (ST) 17 and pulsotype B belonged to ST 78. All strains with the vanA gene were not the same clone, indicating multiple acquisitions of resistant isolates, even over such a short time period.

scholarly journals ddcP, pstB, and excess D-lactate impact synergism between vancomycin and chlorhexidine against Enterococcus faecium 1,231,410

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0249631
Author(s):  
Pooja Bhardwaj ◽  
Moutusee Z. Islam ◽  
Christi Kim ◽  
Uyen Thy Nguyen ◽  
Kelli L. Palmer
Keyword(s):  
Enterococcus Faecium ◽  
Whole Genome ◽  
Vancomycin Resistant Enterococci ◽  
Control Hospital ◽  
Skin Antisepsis ◽  
Life Threatening ◽  
Membrane Bound ◽  
Vancomycin Resistant ◽  
Hospital Associated Infections

Vancomycin-resistant enterococci (VRE) are important nosocomial pathogens that cause life-threatening infections. To control hospital-associated infections, skin antisepsis and bathing utilizing chlorhexidine is recommended for VRE patients in acute care hospitals. Previously, we reported that exposure to inhibitory chlorhexidine levels induced the expression of vancomycin resistance genes in VanA-type Enterococcus faecium. However, vancomycin susceptibility actually increased for VanA-type E. faecium in the presence of chlorhexidine. Hence, a synergistic effect of the two antimicrobials was observed. In this study, we used multiple approaches to investigate the mechanism of synergism between chlorhexidine and vancomycin in the VanA-type VRE strain E. faecium 1,231,410. We generated clean deletions of 7 of 11 pbp, transpeptidase, and carboxypeptidase genes in this strain (ponA, pbpF, pbpZ, pbpA, ddcP, ldtfm, and vanY). Deletion of ddcP, encoding a membrane-bound carboxypeptidase, altered the synergism phenotype. Furthermore, using in vitro evolution, we isolated a spontaneous synergy escaper mutant and utilized whole genome sequencing to determine that a mutation in pstB, encoding an ATPase of phosphate-specific transporters, also altered synergism. Finally, addition of excess D-lactate, but not D-alanine, enhanced synergism to reduce vancomycin MIC levels. Overall, our work identified factors that alter chlorhexidine and vancomycin synergism in a model VanA-type VRE strain.

scholarly journals Enterococcus faecium of thevanAGenotype in Rural Drinking Water, Effluent, and the Aqueous Environment

2011 ◽  
Vol 78 (2) ◽  
pp. 596-598 ◽  
Author(s):  
Dearbháile Morris ◽  
Sandra Galvin ◽  
Fiona Boyle ◽  
Paul Hickey ◽  
Martina Mulligan ◽  
...  
Keyword(s):  
Drinking Water ◽  
Wastewater Treatment ◽  
Environmental Contamination ◽  
Wastewater Treatment Plant ◽  
Enterococcus Faecium ◽  
Treatment Plant ◽  
Aqueous Environment ◽  
Vancomycin Resistant Enterococci ◽  
Vancomycin Resistant ◽  
Rural Drinking Water Supply

ABSTRACTTotal enterococci and vancomycin-resistant enterococci (VRE) were enumerated in samples of effluent (n= 50) and water (n= 167) from a number of sources. VRE were detected in the outflow of a wastewater treatment plant and in a single rural drinking water supply, suggesting potential for transmission to humans through environmental contamination.

scholarly journals First Detection of Vancomycin Resistant Enterococcus faecium in Latvia

2009 ◽  
Vol 63 (4-5) ◽  
pp. 234-236
Author(s):  
Dace Rudzīte ◽  
Arta Balode ◽  
Uga Dumpis ◽  
Edvīns Miklaševičs
Keyword(s):  
High Risk ◽  
Antimicrobial Resistance ◽  
Enterococcus Faecium ◽  
Pathogenic Bacteria ◽  
Advanced Treatment ◽  
Vancomycin Resistant Enterococcus ◽  
Vancomycin Resistant Enterococci ◽  
Treatment Facilities ◽  
The Subject ◽  
Vancomycin Resistant

First Detection of Vancomycin Resistant Enterococcus faecium in Latvia Enterococci have become one of the most important nosocomial pathogens in advanced treatment facilities. Though they are not considered as very pathogenic bacteria, their high levels of antimicrobial resistance are the subject of major concern. Particularly epidemiologically important are vancomycin-resistant enterococci (VRE) due to high risk of transmission of vancomycin resistance genes to staphylococci. We describe the first outbreak of VRE in a Latvian multidisciplinary hospital.

scholarly journals Impact of Daptomycin Dose Exposure Alone or in Combination with β-Lactams or Rifampin against Vancomycin-Resistant Enterococci in an In Vitro Biofilm Model

2020 ◽  
Vol 64 (5) ◽  
Author(s):  
Seyedehameneh Jahanbakhsh ◽  
Nivedita B. Singh ◽  
Juwon Yim ◽  
Razieh Kebriaei ◽  
Jordan R. Smith ◽  
...  
Keyword(s):  
Bactericidal Activity ◽  
Enterococcus Faecium ◽  
Biofilm Reactor ◽  
Content Type ◽  
Biofilm Model ◽  
Vancomycin Resistant Enterococci ◽  
Dosing Regimens ◽  
Vancomycin Resistant ◽  
Reactor Models

ABSTRACT Enterococcus faecium strains are commonly resistant to vancomycin and β-lactams. In addition, E. faecium often causes biofilm-associated infections and these infections are difficult to treat. In this context, we investigated the activity of dosing regimens using daptomycin (DAP) (8, 10, 12, and 14 mg/kg of body weight/day) alone and in combination with ceftaroline (CPT), ampicillin (AMP), ertapenem (ERT), and rifampin (RIF) against 2 clinical strains of biofilm-producing vancomycin-resistant Enterococcus faecium (VREfm), namely, strains S447 and HOU503, in an in vitro biofilm model. HOU503 harbors common LiaS and LiaR substitutions, whereas S447 lacks mutations associated with the LiaFSR pathway. MIC results demonstrated that both strains were susceptible to DAP and resistant to CPT, AMP, ERT, and RIF. The 168-h pharmacokinetic/pharmacodynamic (PK/PD) CDC biofilm reactor models (simulating human antibiotic exposures) were used with titanium and polyurethane coupons to evaluate the efficacy of antibiotic combinations. DAP 12 and 14 achieved bactericidal activity against S447 but lacked such effect against HOU503. Addition of ERT and RIF enhanced DAP activity, allowing DAP 8 and 10 plus ERT or RIF to produce bactericidal activity against both strains at 168 h. While DAP 8 and 10 plus CPT improved killing, they did not reach bactericidal reduction against S447. Combination of AMP, CPT, ERT, or RIF resulted in enhanced and bactericidal activity for DAP against HOU503 at 168 h. Our data provide further support for the use of combinations of DAP with AMP, ERT, CPT, and RIF in infections caused by biofilm producing VREfm. Further research involving DAP combinations against biofilm-producing enterococci is warranted.

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