Programmed knockout mutation of liver fluke granulin attenuates virulence of infection-induced hepatobiliary morbidity
- Khon Kaen University, Thailand
- George Washington University, United States
- James Cook University, Australia
- The University of Melbourne, Australia
Abstract
Infection with the food-borne liver fluke Opisthorchis viverrini is the principal risk factor for cholangiocarcinoma (CCA) in the Lower Mekong River Basin countries including Thailand, Lao PDR, Vietnam and Cambodia. We exploited this link to explore the role of the secreted growth factor termed liver fluke granulin (Ov-grn-1) in pre-malignant lesions by undertaking programmed knockout of the Ov-grn-1 gene from the liver fluke genome. Deep sequencing of amplicon libraries from genomic DNA of gene-edited parasites revealed Cas9-catalyzed mutations within Ov-grn-1. Gene editing resulted in rapid depletion of Ov-grn-1 transcripts and the encoded Ov-grn-1 protein. Gene-edited parasites colonized the biliary tract of hamsters and developed into adult flukes, but the infection resulted in reduced pathology as evidenced by attenuated biliary hyperplasia and fibrosis. Not only does this report pioneer programmed gene-editing in parasitic flatworms, but the striking, clinically-relevant pathophysiological phenotype confirms the role for Ov-grn-1 in virulence morbidity during opisthorchiasis.
Data availability
The Illumina sequencing reads and related project details are available at GenBank: Bioproject PRJNA385864, Biosample SAMN07287348, SRA study SRP110673, accessions SRR5764463-5764618 and SRR8187484-SRR8187487, at https://www.ncbi.nlm.nih.gov/Traces/study/?acc=SRP110673, Bioproject, www.ncbi.nlm.nih.gov/bioproject/PRJNA385864.
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Opisthorchis viverrini Raw sequence readsGenBank, PRJNA385864.
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Sequence: FJ436341.1European Nucleotide Archive, FJ436341.
Article and author information
Author details
Shannon E Karinshak
Funding
National Cancer Institute (R01CA164719)
- Paul J Brindley
Thailand Research Fund (PHD/0111/2557)
- Thewarach Laha
Wellcome (107475/Z/15/Z)
- Paul J Brindley
National Health and Medical Research Council (APP1085309)
- Alex Loukas
Royal Golden Jubilee PhD Program, Thailand (PHD/0111/2557)
- Patpicha Arunsan
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- James B Lok, University of Pennsylvania, United States
Ethics
Animal experimentation: The Animal Ethics Committee of Khon Kaen University approved the study, approval number ACUC-KKU-61/60, which adhered to standard guidelines of the National Research Council of Thailand for the Ethics of Animal Experimentation.
Version history
- Received: August 27, 2018
- Accepted: December 12, 2018
- Accepted Manuscript published: January 15, 2019 (version 1)
- Version of Record published: January 31, 2019 (version 2)
Copyright
© 2019, Arunsan et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Programmed knockout mutation of liver fluke granulin attenuates virulence of infection-induced hepatobiliary morbidity
eLife 8:e41463.
https://doi.org/10.7554/eLife.41463
Further reading
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- Microbiology and Infectious Disease
The first reports of CRISPR/Cas9 genome editing in flatworms could usher in a new era of research on these dangerous human parasites.
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