Molecular Detection of Linezolid Resistance in Enterococcus faecium and Enterococcus faecalis by Use of 5' Nuclease Real-Time PCR Compared to a Modified Classical Approach

The new BD Max GC real-time PCR assay showed high clinical and analytical sensitivity and specificity. It can be an effective and accurate supplementary test for the BD ProbeTec GC Qx amplified DNA assay, which had suboptimal specificity, and might also be used for initial detection ofNeisseria gonorrhoeae.

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The effect of sublethal photodynamic therapy on the expression of Enterococcal surface protein (esp) encoding gene in Enterococcus faecalis: Quantitative real-time PCR assessment

Photodiagnosis and Photodynamic Therapy ◽

10.1016/j.pdpdt.2018.10.008 ◽

2018 ◽

Vol 24 ◽

pp. 311-317 ◽

Cited By ~ 5

Author(s):

Nasim Chiniforush ◽

Maryam Pourhajibagher ◽

Steven Parker ◽

Stefano Benedicenti ◽

Sima Shahabi ◽

...

Keyword(s):

Photodynamic Therapy ◽

Real Time ◽

Enterococcus Faecalis ◽

Real Time Pcr ◽

Surface Protein ◽

Quantitative Real Time Pcr ◽

Encoding Gene

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Duplex Real-Time PCR Assay for Rapid Detection of Ampicillin-Resistant Enterococcus faecium

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.2.556-560.2004 ◽

2004 ◽

Vol 48 (2) ◽

pp. 556-560 ◽

Cited By ~ 14

Author(s):

Stein Christian Mohn ◽

Arve Ulvik ◽

Roland Jureen ◽

Rob J. L. Willems ◽

Janetta Top ◽

...

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Enterococcus Faecium ◽

Rapid Detection ◽

Resistant Bacteria ◽

Pcr Assay ◽

Culture Methods ◽

Accurate Identification ◽

Antibiotic Resistant ◽

Highly Correlated

ABSTRACT Rapid and accurate identification of carriers of resistant microorganisms is an important aspect of efficient infection control in hospitals. Traditional identification methods of antibiotic-resistant bacteria usually take at least 3 to 4 days after sampling. A duplex real-time PCR assay was developed for rapid detection of ampicillin-resistant Enterococcus faecium (ARE). Primers and probes that are used in this assay specifically detected the d-Ala-d-Ala ligase gene of E. faecium and the modified penicillin-binding protein 5 gene (pbp5) carrying the Glu-to-Val substitution at position 629 (Val-629) in a set of 129 tested E. faecium strains with known pbp5 sequence. Presence of the Val-629 in the strain set from 11 different countries was highly correlated with ampicillin resistance. In a screening of hospitalized patients, the real-time PCR assay yielded a sensitivity and a specificity for the detection of ARE colonization of 95% and 100%, respectively. The results were obtained 4 h after samples were harvested from overnight broth of rectal swab samples, identifying both species and the resistance marker mutation in pbp5. This novel assay reliably identifies ARE 2 to 3 days more quickly than traditional culture methods, thereby increasing laboratory throughput, making it useful for rectal screening of ARE. The assay demonstrates the advantages of real-time PCR for detection of nosocomial pathogens.

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Molecular detection of Pneumocystis jirovecii by real-time PCR: Sensitivity depends on the target selection

International Journal of Infectious Diseases ◽

10.1016/j.ijid.2014.03.1188 ◽

2014 ◽

Vol 21 ◽

pp. 372 ◽

Cited By ~ 1

Author(s):

P. Jayaratne ◽

L. Dalle Vedove ◽

D. Yamamura

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Molecular Detection ◽

Target Selection ◽

Pneumocystis Jirovecii

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Serologic and Molecular Detection of Mycoplasma gallisepticum in Layer Flocks in South Marmara Region of Turkey

Journal of Advances in Microbiology ◽

10.9734/jamb/2021/v21i1030390 ◽

2021 ◽

pp. 29-37

Author(s):

Elçin Günaydın ◽

Özlem Kardoğan ◽

Gülşen Goncagül ◽

Yavuz Çokal

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Molecular Detection ◽

Preventive Measures ◽

Mycoplasma Gallisepticum ◽

Marmara Region ◽

Time Loss ◽

Serological Tests ◽

Primary Screening ◽

Pooled Samples

Background: Due to the economic impacts of Mycoplasma gallisepticum (MG) infection in poultry, it is essential to have a fast, reliable and accurate diagnostic test to diagnose the infection. Aims: It was aimed to examine the presence of MG in the South Marmara Region of Turkey where extensive commercial layer flocks exist by RPA, ELISA and real-time PCR. Materials and Methods: In the study, 981 sera and 160 tracheal swab samples (20 swabs per each flock) obtained from eight layer flocks were examined for the presence of MG-antibody by RPA, ELISA, and the presence of MG by real-time PCR, respectively. Results: MG-seropositive flock rate was determined to be 100% by RPA. Twenty-three of the RPA positive sera in each flock LA, LB, LC, LD, LF, LG, and 17 RPA positive sera in flock LE (due to 17 positive RPA sera obtained) were examined for the presence of MG antibody by ELISA, and MG-seropositive flock rate was determined to be 87.5%. As a result of the examination of a total of 32 tracheal swab samples (20 swabs perflock/5 swabs=4 pooled samples, 8 flocksX4 pooled samples= 32 samples) for the presence of MG, real-time PCR positive flock rate was found to be 75%. Conclusion: To decide the flock whether it is infected or not and the initiate effective preventive measures against MG infection as soon as possible; serology should be applied simultaneously with bacteriology and/or PCR to prevent time loss due to shortcomings of serological tests used as primary screening test such as cross reactions, sensitivity and specificity problems.

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

Microorganisms ◽

10.3390/microorganisms8122021 ◽

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.

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

Microorganisms ◽

10.3390/microorganisms8111839 ◽

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.

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