Performance of Core Genome Multilocus Sequence Typing Compared to Capillary-Electrophoresis PCR Ribotyping and SNP Analysis of Clostridioides difficile

bioRxiv (Cold Spring Harbor Laboratory)

Abstract

ABSTRACT Clostridioides difficile is the most common cause of antibiotic-associated gastrointestinal infections. Capillary-electrophoresis (CE)-PCR ribotyping is currently the gold standard for C. difficile typing but lacks discriminatory power to study transmission and outbreaks in detail. New molecular methods have the capacity to differentiate better, but backward compatibility with CE-PCR ribotyping must be assessed. Using a well-characterized collection of diverse strains (N=630; 100 unique ribotypes [RTs]), we aimed to investigate PCR ribotyping prediction from core genome multilocus sequence typing (cgMLST). Additionally, we compared the discriminatory power of cgMLST (SeqSphere & EnteroBase) and whole genome MLST (wgMLST) (EnteroBase) with single nucleotide polymorphism (SNP) analysis). A unique cgMLST profile (>6 allele differences) was observed in 82/100 ribotypes, indicating sufficient backward compatibility. Intra-RT allele difference varied per ribotype and MLST clade. Application of cg/wgMLST and SNP analysis in two outbreak settings with ribotypes RT078 and RT181 (known with a low intra-ribotype allele difference) showed no distinction between outbreak- and non-outbreak strains, in contrast to wgMLST and SNP analysis. We conclude that cgMLST has the potential to be an alternative to CE-PCR ribotyping. The method is reproducible, easy to standardize and offers higher discrimination. However, in some ribotype complexes adjusted cut-off thresholds and epidemiological data are necessary to recognize outbreaks. We propose to decrease the current threshold of 6 to 3 alleles to better identify outbreaks.

Authors 11

1. Amoe Baktash Aachen Department of Medical Microbiology Section Experimental Bacteriology

   Leiden University Medical Center

   Affiliation as printed

   Department of Medical Microbiology, Section Experimental Bacteriology, Leiden University Medical Center, Leiden, the Netherlands

2. Jeroen Corver Aachen Department of Medical Microbiology Section Experimental Bacteriology

   Leiden University Medical Center

   Affiliation as printed

   Department of Medical Microbiology, Section Experimental Bacteriology, Leiden University Medical Center, Leiden, the Netherlands

3. Céline Harmanus Aachen Department of Medical Microbiology Section Experimental Bacteriology National Reference Laboratory Clostridium difficile National Institute of Public Health and the Environment

   Leiden University Medical Center

   Affiliation as printed

   Department of Medical Microbiology, Section Experimental Bacteriology, Leiden University Medical Center, Leiden, the Netherlands

   National Reference Laboratory Clostridium difficile, National Institute of Public Health and the Environment, Leiden University Medical Center, Leiden, the Netherlands

4. Wiep Klaas Smits Aachen Department of Medical Microbiology Section Experimental Bacteriology

   Leiden University Medical Center

   Affiliation as printed

   Department of Medical Microbiology, Section Experimental Bacteriology, Leiden University Medical Center, Leiden, the Netherlands

5. Warren N. Fawley

   University of Leeds · Public Health England

   Affiliation as printed

   National Infection Service, Public Health England, and University of Leeds, Leeds, United Kingdom

6. MH Wilcox

   University of Leeds · Leeds Teaching Hospitals NHS Trust

   Affiliation as printed

   Department of Microbiology, Leeds Teaching Hospitals & University of Leeds, Leeds, United Kingdom

7. Nitin Kumar

   Wellcome Sanger Institute

   Affiliation as printed

   Microbiota Interactions Laboratory, Wellcome Sanger Institute, Hinxton, UK

8. David W. Eyre

   University of Oxford

   Affiliation as printed

   Big Data Institute, Nuffield Department of Population Health, University of Oxford, Oxford, United Kingdom

9. A. Indra

   Paracelsus Medical University

   Affiliation as printed

   Paracelsus Medical University of Salzburg, Salzburg, Austria

10. Alexander Mellmann

    University of Münster · University Hospital Münster

    Affiliation as printed

    Institute of Hygiene, University Hospital Münster, and National Reference Center for C. difficile, Münster Branch, Münster, Germany

11. EJ Kuijper corresponding Aachen Department of Medical Microbiology Section Experimental Bacteriology National Reference Laboratory Clostridium difficile National Institute of Public Health and the Environment

    Leiden University Medical Center

    Affiliation as printed

    Department of Medical Microbiology, Section Experimental Bacteriology, Leiden University Medical Center, Leiden, the Netherlands

    National Reference Laboratory Clostridium difficile, National Institute of Public Health and the Environment, Leiden University Medical Center, Leiden, the Netherlands