Genotypic characterization of Clostridium difficile strains isolated from patients with C.difficile infection in a tertiary hospital in Spain

Abstract number: P1718

Marin M., Martin A., Alcala L., Insa R., Cercenado E., Sanchez-Somolinos M., Bouza E.

Objectives:Clostridium difficile (CD) is the main aetiological agent of antibiotic-associated diarrhoea and a common nosocomial pathogen. In recent years, epidemic strains belonging to the 027 and 078 ribotypes have emerged (both binary toxin+). Genotypic characterization of isolates is essential for recognition of these strains and complements epidemiological investigations. Data on the genotypic characteristics of CD strains circulating in Spain are scarce. We genotyped CD strains circulating in our hospital and isolated from patients with CD infection (CDI) during the year 2007.

Methods: Only strains from patients with a new episode of CDI at least 60 days after a previous episode were considered for study. Strains were cultured and identified by conventional microbiological methods. DNA was obtained from pure cultures using Chelex resin (Instagene matrix, BioRad). The tcdA gene (toxin A), tcdB gene (toxin B), and binary-toxin genes cdtA and cdtB were detected by multiplex PCR (Persson, 2008). Isolates were characterized by PCR-ribotyping (Stubbs, 1999). Phylogenetic analysis of ribotyping profiles was conducted using Bionumerics software 5.0. tcdC gene PCR and sequencing were performed only for clustered strains as previously described (Rupnik, 1998 and Spigaglia, 2002).

Results: Seven hundred and fifty-six CD strains were isolated during 2007. Non-toxigenic CD isolates (96 strains) and those belonging to the same episode (212 strains) were excluded from the study. Overall, 448 CD strains from 435 patients with CD-associated diarrhoea were studied. Two toxigenic profiles were detected: tox A+B+bin- (386 isolates, 86.2%, 45 ribotypes) and tox A+B+bin+ (62 strains, 13.8%; 7 ribotypes). CD strains were clustered in 22 different ribotypes (418 strains, 93.3%) and 30 ribotypes were non-shared. The most frequent ribotypes were 001 (222 strains, 49.5%) and 014 (75 strains, 16.7%). Ribotype 078 was the third most frequent (48 strains, 10.7%). Only 6 isolates (2 patients) belonged to ribotype 027. Analysis of the tcdC gene revealed deletions of 18 bp to 54 bp in 62 strains. All but 2 strains were tox A+B+bin+.

Conclusions: Most of our toxigenic CD isolates were distributed in two ribotypes. Ribotype 078, which is a frequent cause of diarrhoea in our patients, accounted for 10% of toxigenic CD isolated at our institution during 2007. Epidemic strains of ribotype 027 were detected in only 2 patients. All the bin+ CD isolates had deletions in the tcdC gene.

Session Details

Date: 10/04/2010
Time: 00:00-00:00
Session name: Abstracts 20th European Congress of Clinical Microbiology and Infectious Diseases
Location: Vienna, Austria, 10 - 13 April 2010
Presentation type:
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