Detection of Mixed Populations of<i>Clostridium difficile</i>from Symptomatic Patients Using Capillary-Based Polymerase Chain Reaction Ribotyping

Behroozian, Adam; Chludzinski, Jeffrey; Lo, Edward C M; Ewing, Sarah; Waslawski, Sheila; Newton, Duane W.; Young, Vincent B.; Aronoff, David M.; Walk, Seth T.

doi:10.1086/671728

Cited by 31 publications

(36 citation statements)

References 19 publications

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“…However, those patients were also exposed to antimicrobials and chemotherapeutics, making it unclear whether NTCD was the true cause of the diarrhea. In addition, our group and others have cultured NTCD isolates from toxin positive patients [18, 34]. This observation suggests that NTCD isolates may be involved in mixed (toxigenic and non-toxigenic) infections and such cases could be misidentified as solely associated with the non-toxigenic strain.…”

Section: Can Non-toxigenic C Difficile Cause Disease?mentioning

confidence: 77%

A clinical and epidemiological review of non-toxigenic Clostridium difficile

et al. 2013

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Clostridium difficile is a significant nosocomial threat to human health and is the most commonly identified cause of antibiotic associated diarrhea. The development of C. difficile colitis requires production of toxins A and/or B, but some strains do not express these proteins. These non-toxigenic C. difficile (NTCD) have garnered attention for their capacity to colonize humans and potentially reduce the risk for symptomatic colitis caused by toxigenic strains. Isolates of NTCD have been obtained from the environment as well as from animal and human sources. Studies in a hamster CDI model have demonstrated a protective effect of NTCD against toxigenic infection. The extent to which this protective effect of NTCD occurs in humans remains to be defined. Evidence for a therapeutic or preventive role for NTCD is limited but clinical prophylaxis studies are ongoing. NTCD potentially represents an exciting new tool in preventing CDI and its recurrences.

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Section: Can Non-toxigenic C Difficile Cause Disease?mentioning

confidence: 77%

A clinical and epidemiological review of non-toxigenic Clostridium difficile

et al. 2013

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“…Mixed-species C. difficile infections have been found to occur in 7 to 13% of patients infected with disease (references 55 and 56 and references therein). However, it has been difficult to explore the impact of mixed infection on disease progression due to limitations in the ability to accurately quantify the rate at which mixed infections occur or the dynamics of the mixed strains over time within individual patients (55,56). Therefore, it is unclear whether direct competition between strains may be occurring during coinfection in patients.…”

Section: Discussionmentioning

confidence: 99%

Epidemic Clostridium difficile Strains Demonstrate Increased Competitive Fitness Compared to Nonepidemic Isolates

et al. 2014

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Clostridium difficile infection is the most common cause of severe cases of antibiotic-associated diarrhea (AAD) and is a significant health burden. Recent increases in the rate of C. difficile infection have paralleled the emergence of a specific phylogenetic clade of C. difficile strains (ribotype 027; North American pulsed-field electrophoresis 1 [NAP1]; restriction endonuclease analysis [REA] group BI). Initial reports indicated that ribotype 027 strains were associated with increased morbidity and mortality and might be hypervirulent. Although subsequent work has raised some doubt as to whether ribotype 027 strains are hypervirulent, the strains are considered epidemic isolates that have caused severe outbreaks across the globe. We hypothesized that one factor that could lead to the increased prevalence of ribotype 027 strains would be if these strains had increased competitive fitness compared to strains of other ribotypes. We developed a moderate-throughput in vitro model of C. difficile infection and used it to test competition between four ribotype 027 clinical isolates and clinical isolates of four other ribotypes (001, 002, 014, and 053). We found that ribotype 027 strains outcompeted the strains of other ribotypes. A similar competitive advantage was observed when two ribotype pairs were competed in a mouse model of C. difficile infection. Based upon these results, we conclude that one possible mechanism through which ribotype 027 strains have caused outbreaks worldwide is their increased ability to compete in the presence of a complex microbiota.

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“…It is also possible that more than one toxigenic C. difficile ribotype was present in the same sample. Such cases are not necessarily rare and may comprise up to 13% of samples in some situations (15). Future studies that incorporate deeper sampling are needed to understand the rate of mixed CDI among health care institutions.…”

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Clostridium difficile Ribotype Diversity at Six Health Care Institutions in the United States

Waslawski

Ewing

et al. 2013

J Clin Microbiol

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j Capillary-based PCR ribotyping was used to quantify the presence/absence and relative abundance of 98 Clostridium difficile ribotypes from clinical cases of disease at health care institutions in six states of the United States. Regionally important ribotypes were identified, and institutions in close proximity did not necessarily share more ribotype diversity than institutions that were farther apart. National and international studies strongly suggest that Clostridium difficile populations are geographically distinct (1-5), meaning the composition of C. difficile genotypes varies from region to region. In the United States, there are currently few data concerning the composition, or structure, of the C. difficile population and whether health care institutions that are in close proximity are more similar with respect to ribotype diversity than those that are farther apart. The goals of this study were to quantify the abundance and diversity of C. difficile ribotypes from cases of C. difficile infection (CDI) at health care institutions in six states across the United States and to determine whether ribotype diversity was structured geographically.Stool samples from CDI-positive patients were collected from diagnostic laboratories at six health care institutions (West Roxbury, MA; York, PA; Ann Arbor, MI; St. Louis, MO; Portland, OR; and North Hollywood, CA) between 15 February 2011 and 9 September 2011 and shipped in small batches (typically 5 to 10 samples) overnight on ice to the University of Michigan for processing (Ann Arbor, MI, samples were obtained directly from the Clinical Microbiology Laboratory at the University of Michigan Health System and were not shipped). A variety of CDI diagnostic methods were represented across the institutions (Table 1), and CDI cases were defined based on center-specific diagnostic results. Institutional review board (IRB) approval was obtained as appropriate by the participating institutions. Culture was attempted for all samples submitted by each center. On three occasions, samples from three centers were unable to be processed (i.e., plated) within 24 h of receiving due to the lack of available study personnel. These samples were not included in the study. An aliquot from each sample was cultured under anaerobic conditions on prereduced taurocholate cefoxitin cycloserine fructose agar (TCCFA) plates to select for individual C. difficile colonies (6). All TCCFA plates were prepared at the University of Michigan. All samples were frozen at Ϫ80°C and recultured a second time if the first attempt at culture was unsuccessful. A single colony was analyzed from each stool sample, and the toxigenicity of isolates was inferred based on results of PCR for a C. difficile-specific region of the 16S rRNA-encoding gene (7) and tcdA (toxin A) or tcdB (toxin B) (8).The absence of tcdA or tcdB was not confirmed for all isolates (i.e., tcdA can be variably present/absent and primers may or may not amplify all tcdA/tcdB alleles), but all toxigenic isolates were positive for at least on...

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Detection of Mixed Populations ofClostridium difficilefrom Symptomatic Patients Using Capillary-Based Polymerase Chain Reaction Ribotyping

Cited by 31 publications

References 19 publications

A clinical and epidemiological review of non-toxigenic Clostridium difficile

A clinical and epidemiological review of non-toxigenic Clostridium difficile

Epidemic Clostridium difficile Strains Demonstrate Increased Competitive Fitness Compared to Nonepidemic Isolates

Clostridium difficile Ribotype Diversity at Six Health Care Institutions in the United States

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