177 The Presence of Clostridioides difficile Strains in the Gut by Next Generation Shotgun Sequencing: Innocent Bystander or Villain? : Official journal of the American College of Gastroenterology | ACG

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177 The Presence of Clostridioides difficile Strains in the Gut by Next Generation Shotgun Sequencing: Innocent Bystander or Villain?

Papoutsis, Andreas PhD1; Campbell, Jordana BSc2; Hazan-Steinberg, Sabine MD3

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The American Journal of Gastroenterology 114():p S109, October 2019. | DOI: 10.14309/01.ajg.0000590240.42359.f8
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Pathogenic Clostridioides difficile (C. difficile) is the most common cause of nosocomial infection in the United States. However, the prevalence of C. difficile colonization in the general population is poorly understood. We sought to determine the presence and nature of various C. difficile strains colonizing a representative sample of 17 asymptomatic adult volunteers, consisting of 10 healthy and 7 stable Crohn's patients.


Next-generation (NGS) shotgun sequencing was performed on fecal samples. Following fecal collection, DNA was extracted from stool samples, before being quantitated and normalized for downstream library fabrication utilizing shotgun methodology. Prepared and indexed libraries were subsequently pooled and sequenced on the Illumina NextSeq 550 System. Sample FASTQ files were analyzed with a computational tool (Kraken) profiling the microbial communities from metagenomic sequencing data with species level resolution. Individual microbiome profiles were then analyzed for the presence of C. difficile and specific categorical strains (Figure 1).


All 17/17 subjects (100%) were found to possess ≥1 strain of C. difficile (Figure 1). The ATCC 43255 strain was the most common strain (94.1% 16/17), followed by QCD-63q42 (88.2% 15/17), and then M120 and QCD-37x79 strains (both identified in 10/17 [58.8%] individuals).


In our representative sample of 17 adult volunteers, all (100%) were found to possess at least one C. difficile strain in their gut, with many colonized by several strains. While it is recognized that some C. difficile strains are pathogenic, our findings indicate that various non-pathogenic C. difficile species make up an important component of the commensal gut microbiome, and may perhaps play a protective role. Although symptomatic toxigenic CDI is a clear indication for therapy, C. difficile colonization with non-toxigenic strains is not believed to be a direct precursor for CDI. These findings demonstrate our need for awareness that numerous strains of C. difficile exist and the relevance of sequencing (a) prior to hospitalization or antibiotic treatment to help predict those at risk of CDI, and (b) after treatment to identify any loss of potentially protective components of our microbiome.

© 2019 by The American College of Gastroenterology