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Programmed genome editing of the omega-1 ribonuclease of the blood fluke, Schistosoma mansoni

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      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 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.

      eLife digest

      Schistosomiasis is a tropical disease that can cause serious health problems, including damage to the liver and kidneys, infertility and bladder cancer. Nearly a quarter billion people are currently infected, mostly in poor regions of sub-Saharan Africa, the Philippines and Brazil.

      A freshwater worm known as Schistosoma mansoni causes the disease. These parasites enter the human body by burrowing into the skin; once in the bloodstream, they move to various organs where they rapidly start to reproduce. Their eggs release several molecules, including a protein known as omega-1 ribonuclease, which can damage the surrounding tissues.

      A gene editing technique called CRISPR/Cas9 allows scientists to precisely target and then deactivate the genetic information a cell needs to produce a given protein. While the tool has been used in other species before, it was unknown if it could be applied to S. mansoni. Here, Ittiprasert et al. harnessed CRISPR/Cas9 to deactivate the gene that codes for omega-1 ribonuclease and create parasites that do not produce the protein, or only very little of it. The experiments showed that mice infected with the gene-edited worm eggs displayed far fewer symptoms of schistosomiasis compared to those that carry the non-edited parasites.

      Alongside this work, Arunsan et al. used CRISPR/Cas9 to inactivate a gene in another species of worm that can cause liver cancer in humans. Together, these findings demonstrate for the first time that the gene editing method can be adapted for use in parasitic flatworms, which are a major public health problem in tropical climates. This tool should help scientists understand how the parasites invade and damage our bodies, and provide new ideas for treatment and disease control.

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            Author and article information

            Affiliations
            [1 ]deptDepartment of Microbiology, Immunology and Tropical Medicine, School of Medicine and Health Sciences George Washington University Washington, DCUnited States
            [2 ]deptResearch Center for Neglected Diseases of Poverty, School of Medicine and Health Sciences George Washington University Washington, DCUnited States
            [3 ]Wellcome Sanger Institute, Wellcome Genome Campus HinxtonUnited Kingdom
            [4 ]deptDepartment of Parasitology, Faculty of Medicine Khon Kaen University Khon KaenThailand
            [5 ]deptDepartment of Surgery University of Maryland BaltimoreUnited States
            [6 ]deptInstitute of Human Virology University of Maryland BaltimoreUnited States
            [7 ]deptCellular and Molecular Therapeutics Laboratory National Heart, Lungs and Blood Institute, National Institutes of Health BethesdaUnited States
            [8 ]deptDepartment of Parasitology, Faculty of Medicine Siriraj Hospital Mahidol University BangkokThailand
            [9 ]deptSchistosomiasis Resource Center Biomedical Research Institute RockvilleUnited States
            [10 ]deptDepartment of Parasitology Leiden University Medical Center LeidenNetherlands
            [11 ]deptInstitute of Biological, Environmental and Rural Sciences Aberystwyth University AberystwythUnited Kingdom
            University of Pennsylvania United States
            Harvard T.H. Chan School of Public Health United States
            University of Pennsylvania United States
            Contributors
            ORCID: http://orcid.org/0000-0002-9581-0377
            ORCID: https://orcid.org/0000-0003-1765-0002
            Role: Reviewing Editor,
            University of Pennsylvania United States
            Role: Senior Editor,
            Harvard T.H. Chan School of Public Health United States
            Journal
            eLife
            Elife
            eLife
            eLife
            eLife Sciences Publications, Ltd
            2050-084X
            15 January 2019
            2019
            : 8
            30644357
            6355194
            41337
            10.7554/eLife.41337
            (Reviewing Editor), (Senior Editor)

            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.

            Product
            Funding
            Funded by: FundRef http://dx.doi.org/10.13039/100000060, National Institute of Allergy and Infectious Diseases;
            Award ID: R21AI109532
            Award Recipient :
            Funded by: FundRef http://dx.doi.org/10.13039/501100004396, Thailand Research Fund;
            Award ID: PHD/0011/2555
            Award Recipient :
            Funded by: FundRef http://dx.doi.org/10.13039/501100004396, Thailand Research Fund;
            Award ID: PHD/0047/2556
            Award Recipient :
            Funded by: FundRef http://dx.doi.org/10.13039/501100004396, Thailand Research Fund;
            Award ID: PHD/00531/2556
            Award Recipient :
            Funded by: FundRef http://dx.doi.org/10.13039/100004440, Wellcome;
            Award ID: 107475/Z/15/Z
            Award Recipient :
            Funded by: FundRef http://dx.doi.org/10.13039/100004440, Wellcome;
            Award ID: WT 098051
            Award Recipient :
            Funded by: MaxMind Inc;
            Award Recipient :
            The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
            Categories
            Research Article
            Microbiology and Infectious Disease
            Custom metadata
            Gene knock-out of the omega-1 ribonuclease of Schistosoma mansoni eggs resulted in immunologically impaired phenotype, showcasing the novel application of CRISPR/Cas9 genome editing and utility for functional genomics in schistosomes.

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