Towards a cell atlas for the liver fluke Fasciola hepatica

Abstract text

The worldwide prevalent liver fluke Fasciola hepatica causes fascioliasis, a neglected tropical disease and zoonosis. Comprehensive knowledge on the parasite"s cell types and cell-specific gene expression repertoire is missing to date. This kind of insight would take research on drug target genes, developmentally, and metabolically important genes to a next level. Recent technological advances, like the introduction of microfluidics in single-cell RNA sequencing (scRNA-seq) have greatly enhanced and may be transferred to multicellular parasites. Here, we provide the first whole-organism cell atlas of F. hepatica as a basis for expression analysis of genes on a single-cell level.

We established a digestion protocol to obtain high-quality single-cell preparations from adult F. hepatica worms. Cells were then enriched by flow cytometry and finally 19.000 cells processed by 10X Genomics scRNA-seq technology. Cell clusters and marker genes were identified by using the software package Seurat in R. In-situ hybridization (ISH) was used to confirm the sites of gene expression. Gene ontology (GO) enrichment analysis revealed specific biological processes, molecular functions, or cellular components associated with the identified cell clusters.

We successfully identified more than 17 cell clusters that represent distinct cell types, including gastrodermal cells expressing cathepsins, somatic stem cells (neoblasts) expressing nanos2, and neuronal cells. Marker genes for each cluster were identified and their expression confirmed by ISH. Some clusters were enriched for genes of important biological processes, like signal transduction in neuronal cells and proliferation in the neoblast cluster. The resolution of this dataset allowed uncovering potential lineages spanning from the central neoblast cluster into other clusters like the vitellarium, spermatogonia and the female germline. Additionally, we were also able to identify a tissue-specific expression of several protein kinase genes in the tegument as well as reproductive organs, a gene family that is well studied in other helminths with respect to druggable targets.

We present the first transcriptome for the liver fluke F. hepatica on a single-cell resolution. This dataset can serve as treasure trove for the discovery and study of tissue-type and cell-type specific genes in this parasite.