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Use of single molecule sequencing for comparative genomics of an environmental and a clinical isolate of Clostridium difficile ribotype 078.

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posted on 2017-01-17, 12:59 authored by Katherine R. Hargreaves, Anisha M. Thanki, Bethany R. Jose, Marco R. Oggioni, Martha R. J. Clokie
BACKGROUND: How the pathogen Clostridium difficile might survive, evolve and be transferred between reservoirs within the natural environment is poorly understood. Some ribotypes are found both in clinical and environmental settings. Whether these strains are distinct from each another and evolve in the specific environments is not established. The possession of a highly mobile genome has contributed to the genetic diversity and ongoing evolution of C. difficile. Interpretations of genetic diversity have been limited by fragmented assemblies resulting from short-read length sequencing approaches and by a limited understanding of epigenetic regulation of diversity. To address this, single molecule real time (SMRT) sequencing was used in this study as it produces high quality genome sequences, with resolution of repeat regions (including those found in mobile elements) and can generate data to determine methylation modifications across the sequence (the methylome). RESULTS: Chromosomal rearrangements and ribosomal operon duplications were observed in both genomes. The rearrangements occurred at insertion sites within two mobile genetic elements (MGEs), Tn6164 and Tn6293, present only in the M120 and CD105HS27 genomes, respectively. The gene content of these two transposons differ considerably which could impact upon horizontal gene transfer; differences include CDSs encoding methylases and a conjugative prophage only in Tn6164. To investigate mechanisms which could affect MGE transfer, the methylome, restriction modification (RM)  and the CRISPR/Cas systems were characterised for each strain. Notably, the environmental isolate, CD105HS27, does not share a consensus motif for (m4)C methylation, but has one additional spacer  when compared to the clinical isolate M120. CONCLUSIONS: These findings show key differences between the two strains in terms of their genetic capacity for MGE transfer. The carriage of horizontally transferred genes appear to have genome wide effects based on two different methylation patterns. The CRISPR/Cas system appears active although perhaps slow to evolve. Data suggests that both mechanisms are functional and impact upon horizontal gene transfer and genome evolution within C. difficile.

Funding

This work has been funded by a NERC grant NBAF896.This research used the ALICE High Performance Computing Facility at the University of Leicester.

History

Citation

BMC Genomics, 2016 17:1020

Author affiliation

/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/School of Medicine/Department of Infection, Immunity and Inflammation

Version

  • VoR (Version of Record)

Published in

BMC Genomics

Publisher

BioMed Central

eissn

1471-2164

Acceptance date

2016-11-25

Available date

2017-01-17

Publisher version

http://bmcgenomics.biomedcentral.com/articles/10.1186/s12864-016-3346-2

Notes

The two SMRT generated genomes of M120 and CD105HS27 have been deposited in the ENA in study PRJEB13565 and have accession numbers ERS1242840 and ERS1242839, respectively. The methylation data has been deposited for each isolate in REBASE as organisms #19242 and #19243, respectively.

Language

en

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