ADSRRS-fingerprinting and PCR MP techniques for studies of intraspecies genetic relatedness in Staphylococcus aureus.

The present study was designed to evaluate the usefulness of two novel molecular typing methods, amplification of DNA fragments surrounding rare restriction sites (ADSRRS-fingerprinting) and the PCR melting profile (PCR MP), for Staphylococcus aureus strain differentiation. Thirty-seven S. aureus strains isolated from patients with a history of furunculosis were studied. The strains were identified by determining several phenotypic properties and were genotyped using three differentiation methods: macrorestriction analysis of the chromosomal DNA by pulsed-field gel electrophoresis (REA-PFGE), ADSRRS-fingerprinting, and PCR MP technique. In some cases the results obtained showed that the S. aureus isolated from the nose was identical to the one from the furuncle of the same patient. The same genotype was also identified for S. aureus strains isolated from two different members of a family with a history of recurrent furunculosis, although the active lesions were present in only one of them when the investigation was done. Results from strain genotyping illustrated that the recently developed ADSRRS-fingerprinting and PCR MP techniques are useful for studies of intraspecies genetic relatedness of S. aureus strains. They are as effective in discriminating closely related strains as the PFGE method, which is currently considered to be "the gold standard" for epidemiological studies.

[PCR MP (PCR melting profile) technique--possibilities of application in epidemiological studies]. / Technika PCR MP (PCR melting profile)--mozliwosci zastosowania w badaniach epidemiologicznych.

The performance and convenience of a PCR melting profile (PCR MP) technique based on using low denaturation temperatures during ligation mediated PCR (LM PCR) of bacterial DNA is shown. We found that PCR MP technique is a rapid method that offers good discriminatory power, excellent reproducibility and may be applied for epidemiological studies. Results from strain genotyping illustrate that PCR MP is useful for the study of intraspecific genetic relatedness of strains and is as effective in discriminating closely related strains as the PFGE method, which is currently considered to be the gold standard for epidemiological studies. The usefulness of the PCR MP for molecular typing was shown for clinical strains of Escherichia coli, Enterococcus faecium VRE and Stapylococcus aureus.

PCR melting profile method for genotyping analysis of vancomycin-resistant Enterococcus faecium isolates from Hematological Unit patients.

A number of Enterococcus strains with high-level inducible resistance to vancomycin have been identified, and the relative incidence of these strains has increased significantly in the last years. The first outbreak caused by vancomycin-resistant enterococci in Poland was reported in 1999. Vancomycin-resistant Enterococcus faecium is known for its propensity to cause infections which are difficult to eradicate. In this study, we determined the genetic similarities between vancomycin-resistant E. faecium isolates consecutively recovered from single patients to assess the duration of infection or colonization. The isolates taken in the study were identified by the conventional methods as E. faecium. PCR melting profile (PCR-MP) and pulsed-field gel electrophoresis (PFGE) typing revealed that the isolates belonged to six distinct genotypes and that two of them were predominant. Consecutive E. faecium isolates with identical genotypes were found in 7 of 12 (58.0%) patients. The delay between the times of recovery of the first and last isolates of identical genotypes from each patient was from 9 days to about 1 year. In six patients, paired blood and non-blood isolates showed identical genotypes. Data presented here demonstrate the complexity of the epidemiological situation concerning vancomycin-resistant enterococci that may occur in a single medical ward. We also show for the first time the evaluation of PCR-MP technique in enterococci strains differentiation and we revealed that there is at least a similar power of discrimination between the present gold-standard REA-PFGE and a PCR-MP method.

Evaluation of a PCR melting profile technique for bacterial strain differentiation.

In search of an effective DNA typing technique for hospital epidemiology use, the performance and convenience of a PCR melting profile (PCR MP) technique based on using low denaturation temperatures during ligation-mediated PCR (LM PCR) of bacterial DNA was tested. A number of Escherichia coli isolates from patients of the Clinical Hospital in Gdansk, Poland, were examined. We found that the PCR MP technique is a rapid method that offers good discriminatory power and excellent reproducibility and may be applied for epidemiological studies. The usefulness of the PCR MP for molecular typing was compared with the pulsed-field gel electrophoresis method, which is currently considered the gold standard for epidemiological studies of isolates recovered from patients and the environment. Clustering of PCR MP fingerprinting data matched pulsed-field gel electrophoresis data. The features of the PCR MP technique are discussed in comparison with conventional methods. Data presented here demonstrate the complexity of the epidemiological situation concerning E. coli that may occur in a hospital.