Pulsed-Field Gel Electrophoresis of Degradation-Sensitive DNAs from <b> <i>Clostridium difficile</i> PCR Ribotype 1 Strains </b>

Abstract

Clostridium difficile is a causative agent of nosocomially acquired antibiotic-associated diarrhea and pseudomembranous colitis (3). Typing schemes include serogrouping, pyrolysis mass spectroscopy, restriction endonuclease analysis, whole-cell protein profiling, arbitrarily primed PCR, pulsed-field gel electrophoresis (PFGE), and PCR ribotyping (2). PFGE is a highly discriminatory method, but a number of strains of C. difficile are reported to be untypeable due to degradation of DNA during the procedure (4, 5). Such strains can be typed by other methods and have been assigned predominantly to PCR ribotype 1 and serogroup G (4, 5, 7). In the United Kingdom, PCR ribotype 1 accounts for 57% of isolates from hospitalized patients and is endemic in 33 of 58 hospitals surveyed (2). To date all PCR ribotype 1 isolates tested by the Anaerobe Reference Laboratory (Cardiff, United Kingdom) have been untypeable by PFGE; therefore, a method capable of enhanced discrimination of this prevalent ribotype would improve the epidemiology of C. difficile. The aim of this study was to develop a refined PFGE method capable of analyzing previously untypeable strains.