Molecular typing of Clostridioides difficile from frozen stool samples to investigate cross-transmissions: A proof of concept

Dominique S Blanc; Fabrice Poncet; Bruno Grandbastien; Guy Prod'hom; Gilbert Greub; Laurence Senn

doi:10.1016/j.ijmmb.2022.07.018

Molecular typing of Clostridioides difficile from frozen stool samples to investigate cross-transmissions: A proof of concept

Indian J Med Microbiol. 2022 Oct-Dec;40(4):531-535. doi: 10.1016/j.ijmmb.2022.07.018. Epub 2022 Aug 23.

Authors

Dominique S Blanc¹, Fabrice Poncet², Bruno Grandbastien³, Guy Prod'hom⁴, Gilbert Greub⁴, Laurence Senn³

Affiliations

¹ Infection Prevention and Control Unit, Infectious Diseases Service, Lausanne University Hospital and University of Lausanne, 1011 Lausanne, Switzerland; Swiss National Reference Center for Emerging Antibiotic Resistance (NARA), University of Fribourg, Fribourg, Switzerland. Electronic address: Dominique.Blanc@chuv.ch.
² Infection Prevention and Control Unit, Infectious Diseases Service, Lausanne University Hospital and University of Lausanne, 1011 Lausanne, Switzerland; Swiss National Reference Center for Emerging Antibiotic Resistance (NARA), University of Fribourg, Fribourg, Switzerland.
³ Infection Prevention and Control Unit, Infectious Diseases Service, Lausanne University Hospital and University of Lausanne, 1011 Lausanne, Switzerland.
⁴ Institute of Microbiology, Lausanne University Hospital and University of Lausanne, 1011 Lausanne, Switzerland.

PMID: 36008194
DOI: 10.1016/j.ijmmb.2022.07.018

Abstract

Purpose: Toxigenic Clostridioides difficile is responsible for up to one third of post antibiotic diarrhea and for more than 95% of pseudomembranous colitis. Nowadays, diagnosis relies on the documentation of the presence of the toxin in stools by specific antigenic or PCR tests. Stool cultures have been mostly abandoned, leading to the absence of isolates for further epidemiological analyses.

Methods: Aliquots of stool samples, frozen for up to two years, were thawed and inoculated onto commercial C. difficile media. Eighteen stools were recovered from patients hospitalized in the pediatric ward where at that time a chain of transmission was suspected. Eleven stools were recovered from patients hospitalized in a medical ward over a three months period with no suspected transmission event. Up to 16 characteristic colonies were isolates per culture. PCR of toxins genes and molecular typing by Double Locus Sequence Typing (DLST) were performed on these colonies. Whole genome multi locus sequence typing (wgMLST) was performed on selected isolates.

Results: Among the 29 stool specimens, no growth was observed for four stools and only one colony grew for one stool. Except the latter, all 16 colonies of the 24 stools showed identical toxin genes profiles than the original stool. However, variant DLST genotypes was observed within 20% of investigated stools. The majority of variants were single locus variant due to an IN/DEL of the repeat in one of the two DLST locus. Despite this variation, results of molecular typing overrule the putative transmission chain in the pediatric ward and revealed undetected chains of transmission in the medical ward. These results were confirmed with wgMLST.

Conclusions: The developed protocol allows prospective and retrospective molecular and genomic epidemiological investigation of C. difficile infections for infection control purpose.

Keywords: Anaerobic culture; Clostridioides difficile; Genotyping; Molecular typing; Outbreak investigation.

MeSH terms

Anti-Bacterial Agents
Child
Clostridioides
Clostridioides difficile* / genetics
Clostridium Infections* / diagnosis
Feces
Humans
Multilocus Sequence Typing / methods
Prospective Studies
Retrospective Studies

Substances

Anti-Bacterial Agents