June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
Comparative Genome Sequencing to Identify Staphylococcus aureus Virulence and Antibiotic Resistance Genes causing Keratitis
Author Affiliations & Notes
  • Irmgard Behlau
    Molecular Biology & Microbiology, Sackler School of Biomedical Sciences - Tufts University School of Medicine, Boston, MA
    Ophthalmology, Massachusetts Eye and Ear Infirmary/Schepens Eye Research Institute - Harvard Medical School, Boston, MA
  • David Lazinski
    Molecular Biology & Microbiology, Sackler School of Biomedical Sciences - Tufts University School of Medicine, Boston, MA
  • Jacqueline Martin
    Molecular Biology & Microbiology, Sackler School of Biomedical Sciences - Tufts University School of Medicine, Boston, MA
    Ophthalmology, Massachusetts Eye and Ear Infirmary/Schepens Eye Research Institute - Harvard Medical School, Boston, MA
  • Susan Heimer
    Ophthalmology, Massachusetts Eye and Ear Infirmary/Schepens Eye Research Institute - Harvard Medical School, Boston, MA
  • Elizabeth Leonard
    Ophthalmology, Massachusetts Eye and Ear Infirmary/Schepens Eye Research Institute - Harvard Medical School, Boston, MA
  • Andrew Wright
    Molecular Biology & Microbiology, Sackler School of Biomedical Sciences - Tufts University School of Medicine, Boston, MA
  • Michael Gilmore
    Ophthalmology, Massachusetts Eye and Ear Infirmary/Schepens Eye Research Institute - Harvard Medical School, Boston, MA
  • Claes Dohlman
    Ophthalmology, Massachusetts Eye and Ear Infirmary/Schepens Eye Research Institute - Harvard Medical School, Boston, MA
  • Andrew Camilli
    Molecular Biology & Microbiology, Sackler School of Biomedical Sciences - Tufts University School of Medicine, Boston, MA
  • Footnotes
    Commercial Relationships Irmgard Behlau, None; David Lazinski, None; Jacqueline Martin, None; Susan Heimer, None; Elizabeth Leonard, None; Andrew Wright, None; Michael Gilmore, Bausch & Lomb (F); Claes Dohlman, None; Andrew Camilli, Tufts University (P)
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 1726. doi:
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      Irmgard Behlau, David Lazinski, Jacqueline Martin, Susan Heimer, Elizabeth Leonard, Andrew Wright, Michael Gilmore, Claes Dohlman, Andrew Camilli; Comparative Genome Sequencing to Identify Staphylococcus aureus Virulence and Antibiotic Resistance Genes causing Keratitis. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1726.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: S. aureus is a normal commensal of the human skin and nasopharynx, yet S. aureus infection appears to be predominantly caused by only a subset of the strains. Currently available sequence typing methods are only able to identify predefined lineages. They do not allow identification of new virulence or antibiotic resistance genes. Whole genome sequencing can identify such genes, and therefore may give us insight into novel virulence traits associated with keratitis.

Methods: S. aureus clinical isolates were prospectively collected from the Boston area. The diagnosis of clinical keratitis and associated risk factors was by medical record review. Keratitis-associated S. aureus strains were assessed for: 1) antibiotic susceptibility by CSLI standards, 2) biofilm robustness by gentian violet staining, 3) genetic lineage by multi-locus sequence typing (MLST), and 4) whole genome sequencing was performed using Illumina sequencing technology.

Results: 32 keratitis isolates were identified. Risk factors included trauma, prior surgery, soft contact lens wear, and the presence of a surgical implant or environmental foreign body; 25% had no identifiable risk factor. All isolates were tetracycline- and trimethoprim-sulfamethoxazole-sensitive. Prior antibiotic usage did correlate strongly with methicillin-resistance; all MRSA strains were found to be ciprofloxacin-resistant. More than one-third of all keratitis-associated isolated were in the same lineage, ST-5, with both methicillin-sensitive and -resistant S. aureus strains represented. Using a novel linker and paired-end library construction protocol, whole genome sequencing was performed at a cost of $100 USD per bacterial genome; analysis is currently underway.

Conclusions: These results suggest that there may be specific S.aureus lineages that possess genotypic characteristics that enable S. aureus to more effectively cause sight-threatening keratitis and other ocular infections. Additionally, we believe this new genome sequencing technology will give us insight into novel virulence traits that may be uniquely associated with different ocular infections. Further work will explore the feasibility of cost-effective, real-time genomic sequencing technology for clinical application in hospital microbiology laboratories.

Keywords: 720 Staphylococcus • 573 keratitis • 593 microbial pathogenesis: clinical studies  
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