Targeted insertion and reporter transgene activity at a gene safe harbor of the human blood fluke, Schistosoma mansoni

Wannaporn Ittiprasert; Max F Moescheid; Cristian Chaparro; Victoria H Mann; Thomas Quack; Rutchanee Rodpai; André Miller; Prapakorn Wisitpongpun; Watunyoo Buakaew; Margaret Mentink-Kane; Sarah Schmid; Anastas Popratiloff; Christoph G Grevelding; Christoph Grunau; Paul J Brindley

doi:10.1016/j.crmeth.2023.100535

Targeted insertion and reporter transgene activity at a gene safe harbor of the human blood fluke, Schistosoma mansoni

Cell Rep Methods. 2023 Jul 24;3(7):100535. doi: 10.1016/j.crmeth.2023.100535.

Authors

Wannaporn Ittiprasert¹, Max F Moescheid^{1

2}, Cristian Chaparro³, Victoria H Mann¹, Thomas Quack², Rutchanee Rodpai^{1

4}, André Miller⁵, Prapakorn Wisitpongpun^{1

6}, Watunyoo Buakaew^{1

7}, Margaret Mentink-Kane⁵, Sarah Schmid⁵, Anastas Popratiloff⁸, Christoph G Grevelding², Christoph Grunau³, Paul J Brindley¹

Affiliations

¹ Department of Microbiology, Immunology & Tropical Medicine, School of Medicine & Health Sciences, George Washington University, Washington, DC 20037, USA.
² Institute of Parasitology, Biomedical Research Center Seltersberg, Justus Liebig University Giessen, Giessen, Germany.
³ IHPE, University of Perpignan Via Domitia, CNRS, IFREMER, University Montpellier, Perpignan, France.
⁴ Department of Parasitology and Excellence in Medical Innovation, and Technology Research Group, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
⁵ Schistosomiasis Resource Center, Biomedical Research Institute, Rockville, MD 20850, USA.
⁶ Faculty of Medical Technology, Rangsit University, Pathum Thani 12000, Thailand.
⁷ Department of Microbiology, Faculty of Medicine, Srinakharinwirot University, Bangkok 10110, Thailand.
⁸ Nanofabrication and Imaging Center, Science & Engineering Hall, George Washington University, Washington, DC 20052, USA.

Abstract

The identification and characterization of genomic safe harbor sites (GSHs) can facilitate consistent transgene activity with minimal disruption to the host cell genome. We combined computational genome annotation and chromatin structure analysis to predict the location of four GSHs in the human blood fluke, Schistosoma mansoni, a major infectious pathogen of the tropics. A transgene was introduced via CRISPR-Cas-assisted homology-directed repair into one of the GSHs in the egg of the parasite. Gene editing efficiencies of 24% and transgene-encoded fluorescence of 75% of gene-edited schistosome eggs were observed. The approach advances functional genomics for schistosomes by providing a tractable path for generating transgenics using homology-directed, repair-catalyzed transgene insertion. We also suggest that this work will serve as a roadmap for the development of similar approaches in helminths more broadly.

Keywords: Schistosoma mansoni; gene safe harbor; human blood fluke; overlapping CRISPR target; reporter transgene.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Animals, Genetically Modified / genetics
Gene Editing*
Humans
Schistosoma mansoni* / genetics
Transgenes / genetics

Grants and funding

WT_/Wellcome Trust/United Kingdom