Comparison of an Automated Repetitive-Sequence-Based PCR Microbial Typing System with Pulsed-Field Gel Electrophoresis for Molecular Typing of Vancomycin-Resistant
            <i>Enterococcus faecium</i>

Chuang, Yu‐Chung; Wang, Jann-Tay; Chen, Mei‐Ling; Chen, Yee‐Chun

doi:10.1128/jcm.00136-10

Cited by 33 publications

(25 citation statements)

References 25 publications

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“…The DL system is a rapid and semiautomated rep-PCR commercial system that uses standard PCR kits and high-resolution amplicon detection by microfluidic capillary electrophoresis. Its discriminatory power and concordance with other typing methods have been previously evaluated for a number of bacterial pathogens (5,7,11,13,16,19,21,23,24). A recent study analyzed its usefulness in identifying hospital outbreaks caused by different bacterial pathogens (12).…”

Section: Discussionmentioning

confidence: 99%

Controlled Performance Evaluation of the DiversiLab Repetitive-Sequence-Based Genotyping System for Typing Multidrug-Resistant Health Care-Associated Bacterial Pathogens

Deplano¹,

Denis²,

Rodríguez-Villalobos³

et al. 2011

J Clin Microbiol

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Fast, reliable, and versatile typing tools are essential to differentiate among related bacterial strains for epidemiological investigation and surveillance of health care-associated infection with multidrug-resistant (MDR) pathogens. The DiversiLab (DL) system is a semiautomated repetitive-sequence-based PCR system designed for rapid genotyping. The DL system performance was assessed by comparing its reproducibility, typeability, discriminatory power, and concordance with those of pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) and by assessing its epidemiological concordance on well-characterized MDR bacterial strains (n ‫؍‬ 165). These included vanA Enterococcus faecium, extended-spectrum ␤-lactamase (ESBL)-producing strains of Klebsiella pneumoniae, Escherichia coli, and Acinetobacter baumannii, and ESBL-or metallo-␤-lactamase (MBL)-producing Pseudomonas aeruginosa strains. The DL system showed very good performance for E. faecium and K. pneumoniae and good performance for other species, except for a discrimination index of <95% for A. baumannii and E. coli (93.9% and 93.5%, respectively) and incomplete concordance with MLST for P. aeruginosa (78.6%) and E. coli (97.0%). Occasional violations of MLST assignment by DL types were noted for E. coli. Complete epidemiological concordance was observed for all pathogens, as all outbreak-associated strains clustered in identical DL types that were distinct from those of unrelated strains. In conclusion, the DL system showed good to excellent performance, making it a reliable typing tool for investigation of outbreaks caused by study pathogens, even though it was generally less discriminating than PFGE analysis. For E. coli and P. aeruginosa, MLST cannot be reliably inferred from DL type due to phylogenetic group violation or discordance.Rapid assessment of microbial clonal relationships enables the tracking of the spread of multidrug-resistant (MDR) pathogens and guides transmission control measures in the hospital setting. The ideal typing method should be rapid, easy to perform, high throughput, and applicable to a wide range of microorganisms and should meet performance criteria, such as full typeability, reproducibility, high discriminatory power, and concordance with validated typing methods as well as consistency with underlying subspecies genetic population structures (29, 31). In the hospital setting, identification of epidemic strains is usually based on fingerprinting methods, such as pulsed-field gel electrophoresis (PFGE), a labor-intensive method that requires 3 to 4 days to produce results. The DiversiLab (DL) system (bioMérieux, Marcy l'Etoile, France) is a repetitive-sequence-based PCR (rep-PCR) technology which offers semiautomated, easy-to-use, high-throughput, and rapid bacterial strain typing. It could be a suitable alternative to PFGE analysis for outbreak investigation of health care-associated pathogens. This method was recently validated for typing of Acinetobacter spp., Escherichia coli, and Klebsiella spp., but...

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Section: Discussionmentioning

confidence: 99%

Controlled Performance Evaluation of the DiversiLab Repetitive-Sequence-Based Genotyping System for Typing Multidrug-Resistant Health Care-Associated Bacterial Pathogens

Deplano¹,

Denis²,

Rodríguez-Villalobos³

et al. 2011

J Clin Microbiol

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“…Recently, a commercial system for fingerprinting strains by the rep-PCR method (DiversiLab, Bacterial Barcodes, Athens, GA) has become available, offering both standardization and automated data analysis. This approach has been successfully used with several bacterial genera/species, such as Salmonella, Serratia, Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, Enterococcus, and Clostridium difficile (Ross et al, 2005;Doléans-Jordheim et al, 2009;Ben-Darif et al, 2010;Chuang et al, 2010;Fluit et al, 2010;Grisold et al, 2010;Ligozzi et al, 2010;Pasanen et al, 2011). In addition, several E. coli strains such as uropathogenic E. coli, extended-spectrum b-lactamase, and AmpC-producing E. coli and strains isolated from neonatal meningitis cases have been analyzed with the DiversiLab method (Bonacorsi et al, 2009;Pitout et al, 2009;Bogaerts et al, 2010;Brolund et al, 2010;Lau et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

Applicability of DiversiLab Repetitive Sequence-Based PCR Method in Epidemiological Typing of EnterohemorrhagicEscherichia coli(EHEC)

Lukinmaa-Åberg

Horsma

Pasanen

et al. 2013

Foodborne Pathogens and Disease

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Enterohemorrhagic Escherichia coli (EHEC) causes diarrhea, often with severe complications. Rapid and discriminatory typing of EHEC using advanced molecular methods is needed for determination of the genetic relatedness of clones responsible for foodborne outbreaks and for finding out the transmission sources of the outbreaks. This study evaluated the potential of DiversiLab, a semiautomated repetitive sequence-based polymerase chain reaction method for the genotyping of EHEC strains. A set of 52 EHEC strains belonging to 15 O:H serotypes was clustered into 10 DiversiLab groups. All of the O157 strains and one O55 strain were classified into the same group based on a 90% similarity threshold. The other serotypes were classified to their own DiversiLab group, with the exception of one R:H(-) strain that was grouped with O5:H(-) strains. In addition, O26 and O111 strains were grouped together but ultimately subdivided according to their O-serotypes based on a 95% similarity threshold. The O104 strain, which was associated with a major outbreak of hemolytic uremic syndrome in Germany in May 2011, was also classified independently. The DiversiLab performed well in identifying isolates, but the discriminatory power of the repetitive sequence-based polymerase chain reaction method was lower than that of pulsed-field gel electrophoresis. Analysis of 15 enteropathogenic E. coli (EPEC) strains revealed that some EPEC strains clustered together with EHEC strains. Therefore, the DiversiLab system cannot be used to discriminate between these pathogroups. In conclusion, DiversiLab is a rapid and easy system for the primary exclusion of unrelated EHEC strains based on their serotypes, but more discriminatory methods, such as pulsed-field gel electrophoresis, are needed for accurate typing of the EHEC strains.

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“…Therefore, infection monitoring is the most important strategy for preventing colonization and limiting hospital spread of VRE (Orsi & Ciorba, 2013;De Angelis et al, 2014). Molecular typing methods based on analysis of chromosomal DNA provide very useful information about the epidemiology of VRE infections and also for the development of effective strategies to limit the spread of VRE (Chuang et al, 2010). Typing methods such as random-amplified PCR, DNA restriction fragment analysis, ribotyping, PFGE and DNA sequencing that vary in their reproducibility and discriminatory abilities are frequently used.…”

Section: Introductionmentioning

confidence: 99%

Molecular characterization of vancomycin-resistant Enterococcus faecium strains isolated from carriage and clinical samples in a tertiary hospital, Turkey

Gözalan¹,

Coşkun-Ari²,

Özdem³

et al. 2015

Journal of Medical Microbiology

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This study aimed to determine the presence of vancomycin resistance (vanA and vanB) and virulence genes (esp, asa1, gelE, ace, hyl, cylA, cpd and ebpA) in vancomycin-resistant Enterococcus faecium (VREfm) strains and to analyse the clonal relationships among the strains. E. faecium strains were identified from rectal and clinical specimens by biochemical tests and the API-20 Strep kit. Susceptibility testing was performed using disc-diffusion and broth-dilution methods. PFGE was used for molecular typing of the VREfm strains. The vancomycin resistance and virulence genes were amplified by two-step multiplex PCR. All 55 VREfm isolates were resistant to penicillin G, ampicillin and high-level gentamicin but were susceptible to quinupristin/dalfopristin and linezolid. Multiplex PCR analysis indicated that all isolates harboured vanA and that 41 (75 %) were positive for virulence genes. The esp gene was the most common virulence factor and was detected in nine (41 %) invasive and 32 (96.7 %) non-invasive isolates. Multiple virulence genes were observed only in two non-invasive isolates; one harboured esp and ebpA and the other harboured esp, ebpA, asa1, gelE and cpd. PFGE typing yielded 16 different types, seven of which were clusters with two to 14 strains each. The clustering rates of the rectal swab, blood and urine isolates were 72.7 %, 61.5 % and 87.5 %, respectively. The genetic similarity observed among the VREfm isolates indicated crosstransmission in the hospital. Further studies on the virulence factors present in the strains might provide insight into the acquisition of these traits and their contribution to increased prevalence of VREfm.

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Comparison of an Automated Repetitive-Sequence-Based PCR Microbial Typing System with Pulsed-Field Gel Electrophoresis for Molecular Typing of Vancomycin-Resistant Enterococcus faecium

Cited by 33 publications

References 25 publications

Controlled Performance Evaluation of the DiversiLab Repetitive-Sequence-Based Genotyping System for Typing Multidrug-Resistant Health Care-Associated Bacterial Pathogens

Controlled Performance Evaluation of the DiversiLab Repetitive-Sequence-Based Genotyping System for Typing Multidrug-Resistant Health Care-Associated Bacterial Pathogens

Applicability of DiversiLab Repetitive Sequence-Based PCR Method in Epidemiological Typing of EnterohemorrhagicEscherichia coli(EHEC)

Molecular characterization of vancomycin-resistant Enterococcus faecium strains isolated from carriage and clinical samples in a tertiary hospital, Turkey

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