Toward transgenic multicellular parasites: CRISPR-Cas12a germline editing of Schistosoma mansoni eggs as key to success

Information from the abstract

Lack of transgenesis methods impedes functional studies in platyhelminths. We present a substantially improved ribonucleic protein complex (RNP)–based genome editing approach for Schistosoma mansoni comparing Cas9 and Cas12a, the latter attained more efficient homology-directed insertion of a 5′-C6-PEG10–modified double-stranded donor template bearing 50-nucleotide microhomology arms into a genome safe-harbor site (GSH1). Electroporation of eggs recovered from host livers resulted in enhanced green fluorescent protein (EGFP) reporter activity, driven by an S. mansoni ubiquitin promoter and terminator. Hamster infection with transgenic cercariae, the infectious stage of this parasitic platyhelminth, recovered from miracidial infection of snails, or from sporocysts generated in vitro from transgenic miracidia and implantation in snails, produced EGFP-positive adult worms. Sequencing of the edited GSH1 and analyses of filial generations demonstrated chromosomal integration and the stable transformation of S. mansoni . Together, this approach advances schistosome transgenesis and may be pioneering for other platyhelminths, which lack functional genomics and transformation methods.

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Related topics: Parasites and Host Interactions · Genetics, Aging, and Longevity in Model Organisms · CRISPR and Genetic Engineering