1. Microbiology and Infectious Disease
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Programmed genome editing of the omega-1 ribonuclease of the blood fluke, Schistosoma mansoni

  1. Wannaporn Ittiprasert
  2. Victoria H Mann
  3. Shannon E Karinshak
  4. Avril Coghlan
  5. Gabriel Rinaldi
  6. Geetha Sankaranarayanan
  7. Apisit Chaidee
  8. Toshihiko Tanno
  9. Chutima KumKhaek
  10. Pannathee Prangtaworn
  11. Margaret M Mentink-Kane
  12. Christina J Cochran
  13. Patrick Driguez
  14. Nancy Holroyd
  15. Alan Tracey
  16. Rutchannee Rodpai
  17. Bart Everts
  18. Cornelis H Hokke
  19. Karl F Hoffmann
  20. Matthew Berriman
  21. Paul J Brindley  Is a corresponding author
  1. George Washington University, United States
  2. Wellcome Trust Sanger Institute, United Kingdom
  3. University of Maryland, United States
  4. National Heart, Lungs and Blood Institute, National Institutes of Health, United States
  5. Biomedical Research Institute, United States
  6. Leiden University Medical Center, Netherlands
  7. Aberystwyth University, United Kingdom
Research Article
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Cite this article as: eLife 2019;8:e41337 doi: 10.7554/eLife.41337

Abstract

CRISPR/Cas9-based genome editing has yet to be reported in species of the Platyhelminthes. We tested this approach by targeting omega-1 (ω1) of Schistosoma mansoni as a proof of principle. This secreted ribonuclease is crucial for Th2 polarization and granuloma formation. Schistosome eggs were exposed to Cas9 complexed with guide RNA complementary to ω1 by electroporation or by transduction with lentiviral particles. Some eggs were also transfected with a single stranded donor template. Sequences of amplicons from gene-edited parasites exhibited Cas9-catalyzed mutations including homology directed repaired alleles, and other analyses revealed depletion of ω1 transcripts and the ribonuclease. Gene-edited eggs failed to polarize Th2 cytokine responses in macrophage/T-cell co-cultures, while the volume of pulmonary granulomas surrounding ω1-mutated eggs following tail-vein injection into mice was vastly reduced. Knock-out of ω1 and the diminished levels of these cytokines following exposure showcase the novel application of programmed gene editing for functional genomics in schistosomes.

Article and author information

Author details

  1. Wannaporn Ittiprasert

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Victoria H Mann

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Shannon E Karinshak

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2079-0973
  4. Avril Coghlan

    Parasite Genomics, Wellcome Trust Sanger Institute, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Gabriel Rinaldi

    Parasite Genomics, Wellcome Trust Sanger Institute, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  6. Geetha Sankaranarayanan

    Parasite Genomics, Wellcome Trust Sanger Institute, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  7. Apisit Chaidee

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Toshihiko Tanno

    Institute of Human Virology, School of Medicine, Department of Surgery, University of Maryland, Baltimore, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Chutima KumKhaek

    Cellular and Molecular Therapies, National Heart, Lungs and Blood Institute, National Institutes of Health, Bethesda, United States
    Competing interests
    The authors declare that no competing interests exist.
  10. Pannathee Prangtaworn

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    Competing interests
    The authors declare that no competing interests exist.
  11. Margaret M Mentink-Kane

    Schistosomiasis Resource Center, Biomedical Research Institute, Rockville, United States
    Competing interests
    The authors declare that no competing interests exist.
  12. Christina J Cochran

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    Competing interests
    The authors declare that no competing interests exist.
  13. Patrick Driguez

    Parasite Genomics, Wellcome Trust Sanger Institute, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  14. Nancy Holroyd

    Parasite Genomics, Wellcome Trust Sanger Institute, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  15. Alan Tracey

    Parasite Genomics, Wellcome Trust Sanger Institute, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  16. Rutchannee Rodpai

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    Competing interests
    The authors declare that no competing interests exist.
  17. Bart Everts

    Department of Parasitology, Leiden University Medical Center, Leiden, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
  18. Cornelis H Hokke

    Department of Parasitology, Leiden University Medical Center, Leiden, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
  19. Karl F Hoffmann

    Institute of Biological, Environmental and Rural Sciences, Aberystwyth University, Aberystwyth, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  20. Matthew Berriman

    Parasite Genomics, Wellcome Trust Sanger Institute, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9581-0377
  21. Paul J Brindley

    Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington DC, United States
    For correspondence
    pbrindley@gwu.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1765-0002

Funding

Wellcome (107475/Z/15/Z)

  • Karl F Hoffmann

National Institute of Allergy and Infectious Diseases (R21AI109532)

  • Gabriel Rinaldi

Thailand Research Fund (PHD/0011/2555)

  • Apisit Chaidee

Royal Golden Jubilee Ph.D Program, Thailand (PHD/0047/2556)

  • Pannathee Prangtaworn

Wellcome (WT 098051)

  • Matthew Berriman

MaxMind Inc

  • Paul J Brindley

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: Mice experimentally infected with S. mansoni, obtained from the Biomedical Research Institute (BRI), Rockville, MD were housed at the Animal Research Facility of the George Washington University Medical School, which is accredited by the American Association for Accreditation of Laboratory Animal Care (AAALAC no. 000347) and has an Animal Welfare Assurance on file with the National Institutes of Health, Office of Laboratory Animal Welfare, OLAW assurance number A3205-01. All procedures employed were consistent with the Guide for the Care and Use of Laboratory Animals. The Institutional Animal Care and Use Committee (IACUC) of the George Washington University approved the protocol used for maintenance of mice and recovery of schistosomes. Studies with BALB/c mice involving tail vein injection of schistosome eggs and subsequent euthanasia using overdose of sodium pentobarbital was approved by the IACUC of BRI, protocol 18-04, AAALAC no. 000779 and OLAW no. A3080-01.

Reviewing Editor

  1. James B Lok, University of Pennsylvania, United States

Publication history

  1. Received: August 24, 2018
  2. Accepted: December 12, 2018
  3. Accepted Manuscript published: January 15, 2019 (version 1)
  4. Version of Record published: January 31, 2019 (version 2)

Copyright

This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.

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