Snail 3G: genomics, genetics, and gene-editing of Biomphalaria glabrata

This review highlights recent advances and ongoing challenges in the genomics, genetics, and gene-editing (3G) of the freshwater snail <i>Biomphalaria glabrata</i>, based on insights gained from a novel model system we initiated two decades ago. <i>B. glabrata</i> is an intermediate host of the human blood fluke <i>Schistosoma mansoni</i> and serves as the principal model organism for schistosomiasis research. We developed two homozygous lines of <i>B. glabrata</i>, the iM line and iBS90, through 81 and 41 generations of selfing the commonly used M line and BS90, respectively. These lines display contrasting infection phenotypes: susceptibility or resistance to <i>S. mansoni</i>. High-quality scaffold-based genome assemblies were generated for both lines, followed by a chromosome-level assembly of the iM line genome. An F₂ segregating population derived from these lines enabled the identification of three loci, two linked to resistance or susceptibility and one associated with pigmentation, using a double digest restriction-site associated DNA sequencing (ddRADseq) approach. Recombinant inbred lines (RILs) were developed through two crosses and ten generations of selfing. Genetic mapping with RILs refined the resistance locus on chromosome 5 from 8 to 3 Mb through individual-based whole-genome sequencing. Ongoing work includes comparative transcriptome analyses of the two homozygous lines and RILs in response to schistosome infection, along with fine-scale mapping of advanced intercross lines to elucidate the molecular basis of the snail's anti-schistosome defenses. Over the past 10 years, we have made extensive efforts to achieve germline delivery and generate genetically modified snails. Although pantropic lentiviral and yolk protein-mediated germline delivery methods were unsuccessful, these pioneering experiments provide valuable insights for future research. Finally, we successfully generated germline-edited <i>B. glabrata</i>, the first genetically modified schistosomiasis vector snail, by microinjecting CRISPR/Cas9 and guide RNA (gRNA) targeting the <i>fibrinogen-related protein 3.1</i> (<i>FREP3.1</i>) gene into decapsulated embryos, followed by ex ovo culture. This breakthrough establishes a foundation for innovative genetic strategies to control this neglected tropical disease.