Coelomocytes are the cellular workhorses of the echinoderm immune system. They circulate in the coelomic fluids that fill the body cavity, performing functions that in vertebrates are distributed among blood cells and immune cells of many kinds. For decades, biologists have classified these cells by looking at their shapes under the microscope, and sea cucumbers, or holothuroids, stand out among echinoderms for displaying the greatest variety of coelomocyte morphotypes of any class in the phylum. Yet morphology alone has proved a poor guide to function. While the overall immune role of these cells is broadly accepted, the specific jobs performed by each cell type have remained murky, and molecular data specific to the different morphotypes have been scarce in the literature.

Single-cell RNA sequencing, or scRNA-seq, has transformed how biologists untangle cellular heterogeneity, allowing researchers to profile the gene expression of thousands of individual cells and group them into transcriptionally distinct populations. The technique, however, has been largely underutilised in studies of non-model organisms, particularly among invertebrates. The new study set out to close that gap for sea cucumbers by applying the method to coelomocytes harvested from the perivisceral fluid of Holothuria forskali, a species collected with the help of the collection service of the Roscoff Biological Station in France. Sequencing was carried out at the Genomic Platform of the GIGA Institute in Liège.

The analysis identified ten distinct clusters of cells, each assumed to correspond to a separate transcriptional coelomocyte population. The clustering revealed a striking architecture: one cluster, designated cluster 0, occupied a central position relative to all the others on the two-dimensional map of gene expression, suggesting that it may represent undifferentiated cells, a reservoir of progenitors from which more specialised populations arise. At the opposite extreme, cluster 6 sat markedly apart from every other group, hinting at a fundamentally different biological identity and function.

To work out what each cluster might actually do, the team performed functional enrichment analyses and hunted for immune marker genes among the transcripts enriched in each population. The results showed that several clusters carry out key immune functions, including the recognition of pathogens, phagocytosis, the engulfing and destruction of foreign particles, the activation of complement-like pathways, and the regulation of redox balance, the delicate chemistry of oxidation and reduction that cells must manage to survive an immune response. These findings provide the first tentative clues about which transcriptional populations correspond to which defensive duties, offering a molecular handle on a system that has until now been described mostly by cell shape.

One of the most intriguing results concerns a recently discovered cell type called the carotenocyte. These cells, rich in carotenoids, the pigments responsible for many of the reds, oranges and yellows in nature, were only recently described in sea cucumbers, and their presence in the perivisceral fluid was confirmed when the researchers examined their processed samples under the microscope. By drawing on transcriptomic data previously generated for this cell type using bulk RNA sequencing, the team was able to confidently assign cluster 6, the most divergent group on the map, to the carotenocyte lineage, and to deepen the picture of what these pigment-laden cells express.

The convergence of two independent datasets proved decisive. Marker genes supported by both the bulk RNA sequencing of carotenocyte-enriched samples and the new single-cell data gave the researchers confidence in the identity of cluster 6, an approach that illustrates how single-cell and bulk methods can be combined to anchor cell identities in non-model organisms where no reference atlas exists. The study’s supplementary tables catalogue the marker genes for each cluster, their functional annotations against multiple databases, and the results of enrichment analyses against KEGG pathways and gene ontology categories, providing a resource for other researchers working on echinoderm immunity.

Rigorous quality control underpinned the analysis. The authors filtered cells on the basis of the number of unique molecular identifiers and detected genes per cell, examined the expression of mitochondrial genes to flag stressed or damaged cells, and used the DoubletFinder tool to detect and remove doublets, artefacts in which two cells are captured and sequenced as one. Sensitivity analyses testing different filtering parameters showed that while individual clusters could merge or split depending on the settings, the overall configuration of the map remained stable, with cluster 0 staying central and cluster 6 remaining the most divergent population, a reassuring sign that the biological signal is robust.

The implications reach well beyond sea cucumbers. Echinoderms are deuterostomes, the same major lineage as chordates, which makes their immune cells valuable comparators for understanding how immune cell lineages evolved on the branch of the tree that eventually produced our own adaptive immune system. A better grasp of holothuroid coelomocyte diversity could also help interpret stress responses in these animals, which are ecologically important grazers and the target of a rapidly expanding aquaculture industry in Asia. Knowing which cells respond to pathogens or environmental stressors, and through which genes, could inform disease management in farmed populations and shed light on how wild sea cucumbers cope with a changing ocean.

The authors are careful to frame the study as a pioneering first step. Linking the remaining transcriptional clusters to the morphotypes described in the historical literature will require further analyses, and the functional assignments drawn from enrichment analyses remain hypotheses to be tested experimentally. Still, the study demonstrates that single-cell transcriptomics can be successfully deployed on a non-model invertebrate with no reference-grade tools, and it delivers a preliminary but detailed map of the functional diversity of holothuroid coelomocytes. For a phylum whose immune system has been studied mostly through a microscope lens, the jump to single-cell genomics marks the beginning of a new era, one in which the hidden division of labour among sea cucumber immune cells can finally be read directly from their genes.

Subject of Research: Single-cell transcriptomic profiling of coelomocyte immune cell diversity in the sea cucumber Holothuria forskali

Article Title: Single-cell transcriptomics reveals transcriptional diversity of sea cucumber perivisceral fluid coelomocytes

Article References: Wambreuse, N., Lavergne, A., Fievez, L., Bureau, F., Zhang, L., Deng, B., Caulier, G., Eeckhaut, I., & Delroisse, J. (2026). Single-cell transcriptomics reveals transcriptional diversity of sea cucumber perivisceral fluid coelomocytes. BMC Genomics. https://doi.org/10.1186/s12864-026-13356-9

Image Credits: AI Generated

DOI: 10.1186/s12864-026-13356-9

Keywords: sea cucumber, coelomocytes, single-cell RNA sequencing, echinoderm immunity, carotenocytes, Holothuria forskali, transcriptomics, immune cells, deuterostomes, phagocytosis, BMC Genomics, non-model organisms