Epidemiological Fingerprinting of Klebsiella pneumoniae by Small-Fragment-Restriction-Endonuclease-Analysis (SF-REA)

Abstract

Epidemiological fingerprinting of Klebsiella pneumoniae was performed by restriction endonuclease analysis (REA) of whole cell DNA. 11 isolates from 4 patients in an intensive care unit and 80 unrelated strains were examined in this study. DNA was cleaved with restriction endonuclease EcoR I, electrophoresed on 10% polyacrylamide gels, and restriction fragment patterns were visualized by silver staining. The analysis of small fragments within the cleavage patterns (SF-REA) yielded sufficient information for reliable strain identification. The gel patterns of unrelated strains exhibited marked differences by direct visual comparison. In contrast, the isolates from the ICU could only be subdivided into 2 types, supporting our suspicion of nosocomial infections in some of these patients. SF-REA was evaluated with regard to interstrain discriminatory ability, reproducibility, and practicability. Our results indicate that SF-REA may be used as a rapid, precise and reliable technique in typing K. pneumoniae strains.

Epidemiological fingerprinting of Enterobacter cloacae by small-fragment restriction endonuclease analysis and pulsed-field gel electrophoresis of genomic restriction fragments

“…However, the limited diversity of patterns in this method may be restrictive in large epidemiological studies, thus making further analysis by restriction endonuclease digestion necessary. Instability of profiles due to acquisition, loss, or transfer of plasmids may be another handicap of this technique, and identical plasmid profiles may not reflect the overall genetic relatedness of isolates (7,19,26).…”

“…Chromosomal analysis by SF-REA. Whole-cell DNA for SF-REA was prepared by a modified guanidine thiocyanate- Triton X-100 lysis procedure (4) followed by phenol-chloroform extraction and isopropanol precipitation as previously described (19). Restriction endonucleases were used in the appropriate buffers under the conditions recommended by the manufacturer (New England Biolabs).…”

Genetic variability and molecular typing of Shigella sonnei strains isolated in Canada

“…The results revealed a high degree of RFLP pattern heterogeneity among epidemiologically independent S. sonnei strains. Among 20 strains associated with sporadic cases of shigellosis in various regions of Canada, 15 distinct patterns 1 2 3 4 5 6 7 8 9 1011121314FIG. 2.…”

“…Recently, Haertl and colleagues (3,4) have described an improved restriction endonuclease analysis (REA) technique for the detection of relatively small genomic DNA restriction fragments from Klebsiella pneumoniae and Enterobacter cloacae. In the present study, we used a modified version of this procedure to investigate the genetic variability of epidemiologically related and unrelated S. sonnei strains isolated in Canada, and we evaluated this technique as a potential molecular typing tool for this species.…”

Randomly amplified polymorphic DNA typing: a useful tool for rapid epidemiological typing ofKlebsiella pneumoniae

“…Whereas capsular serotyping, phage typing and bacteriocin typing have more discriminatory power, all three are time consuming and expensive methods and not all strains of K. pneumoniae are successfully typed by these methods [2,[4][5][6]. At present the most discriminatory method of typing K. pneumoniae involves the use of restriction fragment length polymorphism (RFLP) techniques [6][7][8]. However, these techniques are costly, labour-intensive and have lengthy processing times [9][10][11].…”