A single-cell RNAseq atlas of the pathogenic stage ofSchistosoma mansoniidentifies a key regulator of blood feeding

Abstract

Schistosomiasis is an ancient and chronic neglected tropical disease that infects over 240 million people and kills over 200,000 of the world’s poorest people every year1, 2. There are no vaccines and because there is only one drug available, the need for new therapeutics is great. The causative agents of this disease are flatworm parasites that dwell inside the host’s circulation, often for decades, where they feed on blood and lay eggs which are primarily responsible for disease pathology. As metazoans comprised of multiple tissue types, understanding the schistosome’s tissues on a molecular level and their functions during what can be decades of successful parasitism could suggest novel therapeutic strategies. Here, we employ single-cell RNAseq to characterize 43,642 cells from the pathogenic (adult) stage of the schistosome lifecycle. From these data, we characterize 68 molecularly distinct cell populations that comprise nearly all tissues described morphologically, including the nervous and reproductive systems. We further uncover a lineage of somatic stem cells responsible for producing and maintaining the parasite’s gut – the primary tissue responsible for digestion of host blood. Finally, we show that a homologue of hepatocyte nuclear factor 4 (hnf4) is expressed in this gut lineage and required for gut maintenance, blood feeding and inducing egg-associated pathology in vivo. Together, the data highlight the utility of this single-cell RNAseq atlas to understand schistosome biology and identify potential therapeutic interventions.

Large CRISPR-Cas-induced deletions in the oxamniquine resistance locus of the human parasiteSchistosoma mansoni

“…In our study, while amplicon sequencing revealed reads carrying CRISPR-Cas9-induced mutations in SULT-OR, a knock-down of SULT-OR at the mRNA level was not evident, probably because the deletions occurred in only a small fraction of the adult cells that express SULT-OR. Single-cell sequencing data from adults shows SULT-OR is a marker of parenchymal cells [10], while our confocal microscopy data suggest the Cas9-gRNA complex penetrated better into the adult tegument and intestine compared to parenchymal tissue. The same phenomenon was previously described in the liver fluke Fasciola hepatica when delivering fluorescently labelled molecules by electroporation [52,53], suggesting the flatworm intestine as the main point of entry when this delivery approach is employed.…”

“…However, ex vivo SULT-OR RNAi experiments in adult male worms showed no evident phenotypic effects other than becoming resistant to OXA [23]. Single-cell transcriptomic analysis of two-day-old schistosomula [9], adult worms [10], and in vitro-transformed mother sporocysts (unpublished) revealed SULT-OR mRNA is a marker of parenchymal cell clusters ( Supplementary Table S3 and Supplementary Figure S2B), while its top BLASTP hit in the planarian Schmidtea mediterranea, dd_Smed_v6_9472_0, is a marker of intestinal cells [38].…”

“…(B) Uniform Manifold Approximation projection (UMAP) plot of the SULT-OR gene (Smp_089320) expression pattern in adult worms showing an enriched expression in the parenchyma_1 (1), parenchyma_2 (2) and unknown (3) cell clusters. Raw data obtained from [10].…”

“…Expression profile of the SULT-OR gene in single-cell sequencing data. Raw single-cell sequencing data obtained from [9,10] Supplementary Movies. Supplementary Movie S1.…”

“…High-throughput datasets, including high quality reference genomes for the three main species of schistosomes [5][6][7], have been generated. More recently, a thorough transcriptome analysis during the parasite's intra-mammalian development [8], and the identification of different cell types by single-cell RNA sequencing of various life cycle stages [9,10] represent significant steps towards deciphering cell fate and pathways involved in parasite development and host-parasite interactions.…”