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PCR fragment length polymorphism analysis of vancomycin-resistant Enterococcus faecium
S Donabedian1, E Hershberger, L A Thal
1Departments of Medicine, Wayne State University, Detroit, Michigan, USA.
Abstract:
In this study, the glycopeptide resistance element, Tn1546, in 124 VanA Enterococcus faecium clinical isolates from 13 Michigan hospitals was evaluated using PCR fragment length polymorphism. There were 26 pulsed-field gel electrophoresis (PFGE) types, which consisted of epidemiologically related and unrelated isolates from separate patients (1992 to 1996). Previously published oligonucleotides specific for regions in the vanA gene cluster of Tn1546 were used to amplify vanRS, vanSH, vanHAX, vanXY, and vanYZ. The glycopeptide resistance element, Tn1546, of E. faecium 228 was used as the basis of comparison for all the isolates in this study. Five PCR fragment length patterns were found, as follows. (i) PCR amplicons were the same size as those of EF228 for all genes in the vanA cluster in 19.4% of isolates. (ii) The PCR amplicon for vanSH was larger than that of EF228 (3.7 versus 2.3 kb) due to an insertion between the vanS and vanH genes (79.2% of isolates). (iii) One isolate in a unique PFGE group had a vanSH amplicon larger than that of EF228 (5.7 versus 2.3 kb) due to an insertion in the vanS gene and an insertion between the vanS and vanH genes. (iv) One isolate did not produce a vanSH amplicon, but when vanS and vanH were amplified separately, both amplicons were the same size as those as EF228. (v) One isolate had a vanYZ PCR product larger than that of EF228 (2.8 versus 1.6 kb). This study shows that in a majority of the VanA E. faecium isolates, Tn1546 is altered compared to that of EF228. A total of 79.2% of the study isolates had the same-size insertion between the vanS and vanH genes. The results of this study show dissemination of an altered Tn1546 in heterologous VanA E. faecium in Michigan hospitals.
Insights
This study found that the Tn1546 element, responsible for glycopeptide resistance in VanA Enterococcus faecium, is frequently altered in clinical isolates. Most isolates showed changes in the vanSH region, indicating widespread dissemination of modified resistance genes.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- VanA Enterococcus faecium is a significant cause of hospital-acquired infections.
- Glycopeptide resistance in Enterococcus faecium is often mediated by the Tn1546 element.
- Understanding variations in Tn1546 is crucial for infection control.
Purpose of the Study:
- To investigate the genetic variations of the Tn1546 glycopeptide resistance element in clinical isolates of VanA Enterococcus faecium.
- To determine the prevalence of different Tn1546 variants among isolates from Michigan hospitals.
Main Methods:
- Utilized PCR fragment length polymorphism to analyze the vanA gene cluster within the Tn1546 element.
- Employed pulsed-field gel electrophoresis (PFGE) to assess the genetic relatedness of isolates.
- Compared PCR amplicons to a reference Tn1546 element from E. faecium 228.
Main Results:
- Five distinct PCR fragment length patterns were identified, indicating alterations in Tn1546.
- A significant majority (79.2%) of isolates exhibited an insertion between the vanS and vanH genes, resulting in a larger vanSH amplicon.
- Other variations included insertions within the vanS gene and the vanYZ region.
Conclusions:
- The Tn1546 element is frequently altered in VanA Enterococcus faecium clinical isolates.
- The prevalence of an insertion between vanS and vanH suggests the dissemination of a common, modified Tn1546 element.
- These findings highlight the dynamic nature of antimicrobial resistance genes in hospital settings.