Molecular epidemiology of endemic Clostridium difficile infection and the significance of subtypes of the United Kingdom epidemic strain (PCR ribotype 1)

Abstract

We previously identified two subtypes of the epidemic strain Clostridium difficile PCR ribotype 1, one clindamycin-sensitive strain (arbitrarily primed PCR [AP-PCR] type Ia) and a closely related clindamycin-resistant strain (AP-PCR type Ib) in our institution. We have now carried out prospective epidemiological surveillance for 4 years, immediately following the relocation of two acute medicine wards for elderly patients (wards A and B), to determine the clinical epidemiology of subtypes of the epidemic C. difficile PCR ribotype 1 group. To maximize the chance of strain discrimination, we used three DNA fingerprinting methods, AP-PCR, ribospacer PCR (RS-PCR), and pulsed-field gel electrophoresis (PFGE), to analyze C. difficile isolates recovered from symptomatic patients and from repeated environmental samplings. On ward B the incidence of C. difficile infection correlated significantly with the prevalence of environmental C. difficile both in ward areas closely associated with patients and health care personnel (r = 0.53; P < 0.05) and in high-reach sites (r = 0.85; P < 0.05). No such relationships were found on ward A. Seventeen distinct C. difficile genotypes were identified, 17 by AP-PCR, 12 by PFGE, and 11 by RS-PCR, but only 4 of 17 genotypes caused patient infection. Isolates recovered from the hospital ward environment were much more diverse (14 genotypes). AP-PCR type Ia represented >90% of the C. difficile isolates. In addition to this genotype, only two others were isolated from both patient feces and environmental surfaces. AP-PCR type Ib (clindamycin-resistant PCR ribotype 1 clone) was not associated with any cases of C. difficile infection and was isolated from the environment on only two occasions, after having been implicated in a cluster of six C. difficile infections 5 months before this study. The disappearance of this strain implies that differences in virulence and/or selective pressures may exist for this strain and the closely related, widespread C. difficile AP-PCR type Ia strain. Our findings emphasize the need to understand the epidemiology and virulence of clinically significant strains to determine successful control measures for C. difficile infections.

FIG. 1.

Distribution of AP-PCR genotype III isolates on ward A. Open circles, AP-PCR genotype IIIa isolates; stippled circles, AP-PCR genotype IIIb isolates; closed circles, AP-PCR genotype IIIc isolates; closed ovals, environmental sites commonly associated with patients and health care workers (sampled monthly), including floors (FL), radiators (R), bed frames (BF), curtain rails (CR), and commodes (C); closed diamonds, high-reach environmental sites (samples every 6 months), including overbed lamps (OL), window frames (WF), curtain rails (CR), bay partitions (BP), door tops (DT), door frames (DF), storage cupboards (SC), fire hoses (FH), and smoke detectors (SD).

FIG. 2.

Frequency of C. difficile culture-positive environmental sites commonly associated with patients and health care workers (A) and high-reach sites (B) on study wards A (grey bars) and B (white bars).

FIG. 3.

Analysis of AP-PCR profiles of C. difficile strains isolated from the stools of patients with C. difficile infection, the hands of health care workers, and the environments of two medicine hospital wards for elderly patients. The dendrogram includes a small representative type of the predominant hospital genotype (type Ia) and all other genotypes found in the study. Strains isolated from the hands of health care workers are marked with an asterisk. DNA profiles were analyzed by using BioNumerics software (Applied Maths, BioSystematica). Dendrograms were constructed by the unweighted pair group method with arithmetic mean clustering by using the Dice correlation coefficient. The percentage level of similarity chosen for type assignments (roman numerals) is indicated by the thick black line and was based on the guidelines recommended by Tenover et al. (29).

FIG. 4.

Analysis of PFGE profiles of C. difficile strains isolated from the stools of patients with C. difficile infection, the hands of health care workers, and the environment of two medicine hospital wards for elderly patients. The dendrogram includes a small representative type of the predominant hospital genotype (subtype 1a) and all other genotypes found in the study. Strains isolated from the hands of health care workers are marked with an asterisk. The DNA profiles were analyzed by using BioNumerics software (Applied Maths, BioSystematica). Dendrograms were constructed by the unweighted pair group method with arithmetic mean clustering by using the Dice correlation coefficient. The percentage level of similarity chosen for group assignments (roman numerals) is indicated by the thick black line and was based on the guidelines recommended by Tenover et al. (29).

FIG. 5.

Analysis of RS-PCR profiles of C. difficile strains isolated from the stools of patients with C. difficile infection, the hands of health care workers, and the environment of two medicine hospital wards for elderly patients. The dendrogram includes a small representative group of the predominant hospital genotype (type I) and all other genotypes found in the study. Strains isolated from the hands of health care workers are marked with an asterisk. The DNA profiles were analyzed by using BioNumerics software (Applied Maths, BioSystematica). Dendrograms were constructed by unweighted pair group method with arithmetic mean clustering by using the Dice correlation coefficient. The percentage level of similarity chosen for group assignments (roman numerals) is indicated by the thick black line and was based on the guidelines recommended by Tenover et al. (29).

FIG. 6.

C. difficile infection and environmental culture positivity for study wards A and B; ▪, patient isolates; •, environmental isolates from sites regularly in contact with patients and ward staff; ⋄, environmental isolates from high-reach sites.