Single-cell atavism reveals an ancient mechanism of cell type diversification in a sea anemone.

Cnidocytes are the explosive stinging cells unique to cnidarians (corals, jellyfish, etc). Specialized for prey capture and defense, cnidocytes comprise a group of over 30 morphologically and functionally distinct cell types. These unusual cells are iconic examples of biological novelty but the developmental mechanisms driving diversity of the stinging apparatus are poorly characterized, making it challenging to understand the evolutionary history of stinging cells. Using CRISPR/Cas9-mediated genome editing in the sea anemone Nematostella vectensis, we show that a single transcription factor (NvSox2) acts as a binary switch between two alternative stinging cell fates. Knockout of NvSox2 causes a transformation of piercing cells into ensnaring cells, which are common in other species of sea anemone but appear to have been silenced in N. vectensis. These results reveal an unusual case of single-cell atavism and expand our understanding of the diversification of cell type identity.

Skimming for barcodes: rapid production of mitochondrial genome and nuclear ribosomal repeat reference markers through shallow shotgun sequencing.

DNA barcoding is critical to conservation and biodiversity research, yet public reference databases are incomplete. Existing barcode databases are biased toward cytochrome oxidase subunit I (COI) and frequently lack associated voucher specimens or geospatial metadata, which can hinder reliable species assignments. The emergence of metabarcoding approaches such as environmental DNA (eDNA) has necessitated multiple marker techniques combined with barcode reference databases backed by voucher specimens. Reference barcodes have traditionally been generated by Sanger sequencing, however sequencing multiple markers is costly for large numbers of specimens, requires multiple separate PCR reactions, and limits resulting sequences to targeted regions. High-throughput sequencing techniques such as genome skimming enable assembly of complete mitogenomes, which contain the most commonly used barcoding loci (e.g., COI, 12S, 16S), as well as nuclear ribosomal repeat regions (e.g., ITS1&2, 18S). We evaluated the feasibility of genome skimming to generate barcode references databases for marine fishes by assembling complete mitogenomes and nuclear ribosomal repeats. We tested genome skimming across a taxonomically diverse selection of 12 marine fish species from the collections of the National Museum of Natural History, Smithsonian Institution. We generated two sequencing libraries per species to test the impact of shearing method (enzymatic or mechanical), extraction method (kit-based or automated), and input DNA concentration. We produced complete mitogenomes for all non-chondrichthyans (11/12 species) and assembled nuclear ribosomal repeats (18S-ITS1-5.8S-ITS2-28S) for all taxa. The quality and completeness of mitogenome assemblies was not impacted by shearing method, extraction method or input DNA concentration. Our results reaffirm that genome skimming is an efficient and (at scale) cost-effective method to generate all mitochondrial and common nuclear DNA barcoding loci for multiple species simultaneously, which has great potential to scale for future projects and facilitate completing barcode reference databases for marine fishes.

Cassiosomes are stinging-cell structures in the mucus of the upside-down jellyfish Cassiopea xamachana.

Snorkelers in mangrove forest waters inhabited by the upside-down jellyfish Cassiopea xamachana report discomfort due to a sensation known as stinging water, the cause of which is unknown. Using a combination of histology, microscopy, microfluidics, videography, molecular biology, and mass spectrometry-based proteomics, we describe C. xamachana stinging-cell structures that we term cassiosomes. These structures are released within C. xamachana mucus and are capable of killing prey. Cassiosomes consist of an outer epithelial layer mainly composed of nematocytes surrounding a core filled by endosymbiotic dinoflagellates hosted within amoebocytes and presumptive mesoglea. Furthermore, we report cassiosome structures in four additional jellyfish species in the same taxonomic group as C. xamachana (Class Scyphozoa; Order Rhizostomeae), categorized as either motile (ciliated) or nonmotile types. This inaugural study provides a qualitative assessment of the stinging contents of C. xamachana mucus and implicates mucus containing cassiosomes and free intact nematocytes as the cause of stinging water.

Scientific gear as a vector for non-native species at deep-sea hydrothermal vents.

The fauna of deep-sea hydrothermal vents are among the most isolated and inaccessible biological communities on Earth. Most vent sites can only be visited by subsea vehicles, which can and do move freely among these communities. Researchers assume individuals of the regionally homogeneous vent fauna are killed by the change in hydrostatic pressure the animals experience when the subsea vehicles, which collected them, rise to the surface. After an Alvin dive, we found 38 apparently healthy individuals of a vent limpet in a sample from a hydrothermally inactive area. Prompted by our identification of these specimens as Lepetodrilus gordensis, a species restricted to vents 635 km to the south of our dive site, we tested whether they were from a novel population or were contaminants from the dive made 36 h earlier. The 16S gene sequences, morphology, sex ratio, bacterial colonies, and stable isotopes uniformly indicated the specimens came from the previous dive. We cleaned the sampler, but assumed pressure changes would kill any organisms we did not remove and that the faunas of the 2 areas were nearly identical and disease-free. Our failure to completely clean the gear on the subsea vehicle meant we could have introduced the species and any diseases it carried to a novel location. Our findings suggest that the nearly inaccessible biological communities at deep-sea vents may be vulnerable to anthropogenic alteration, despite their extreme physical conditions.

Morphology, distribution, and evolution of apical structure of nematocysts in hexacorallia.

Cnidae are complex intracellular capsules made by all cnidarians. The most diverse of these capsules are nematocysts, which are made by all members of the phylum; spirocysts and ptychocysts are made only by members of some lineages, and they show less functional and structural diversity. In nematocysts, the apex has been shown to be either a hinged cap (operculum) or three flaps that flex outward during discharge. The operculum is known only from medusozoan nematocysts; flaps are known only from nematocysts of members of the hexacorallian order Actiniaria, although they have been inferred to be characteristic of Anthozoa, the group to which Actiniaria belongs. Using scanning and transmission electron microscopy, we discover a third apical morphology in nematocysts, an apical cap, which we find in all nonactiniarian anthozoans examined. This apical cap is identical structurally to the apical cap of spirocysts, and it resembles the apical structure of ptychocysts, whose apex is documented here for the first time. Additionally, a full survey of nematocysts from all body structures of two actiniarians demonstrates that a particular type of nematocyst, the microbasic p-mastigophore of the mesenterial filaments, does not have apical flaps. The observed variation does not correspond to conventional categorization of capsule morphology and raises questions about the function and structure of capsules across Cnidaria. Despite some ambiguity in optimization of ancestral states across cnidae, we determine that the apical cap is the plesiomorphic structure for anthozoan cnidae and that apical flaps are a synapomorphy of Actiniaria. At present, the operculum is interpreted as a synapomorphy for Medusozoa, but either it or an apical cap is the ancestral state for nematocysts.

Phylogenetic signal in mitochondrial and nuclear markers in sea anemones (Cnidaria, Actiniaria).

The mitochondrial genome of basal animals is generally more slowly evolving than that of bilaterians. This difference in rate complicates the study of relationships among members of these lineages and the discovery of cryptic species or the testing of morphological species concepts within them. We explore the properties of mitochondrial and nuclear ribosomal genes in the cnidarian order Actiniaria, using both an ordinal- and familial-scale sample of taxa. Although the markers do not show significant incongruence, they differ in their phylogenetic informativeness and the kinds of relationships they resolve. Among the markers studied here, the fragments of 12S rDNA and 18S rDNA most effectively recover well-supported nodes; those of 16S rDNA and 28S rDNA are less effective. The general patterns we observed are similar to those in other hexacorallians, although Actiniaria alone show saturation of transitions for ordinal-scale analyses.

Formation of the apical flaps in nematocysts of sea anemones (cnidaria: actiniaria).

Using scanning and transmission electron microscopy, we studied formation of the structure at the apical end of sea anemone nematocysts through which the tubule everts at discharge. In anemones of the genus Metridium, we found that each of the three solid triangular apical flaps comprises two layers that are continuous with those of the capsule wall: the electron-lucent inner layer is bound to the electron-dense outer layer. The two-layer structure is obvious in some discharged capsules in which, perhaps due to fixation, the layers part at the flap's periphery. Before the nematocyst discharges, a channel leads from a pore at the tip of the joined flaps into the lumen of the inverted tubule. The thin laminate layer that coats each flap lines the channel. The base of the nematocyst tubule adheres to the capsule wall near the capsule's apical end, and a branch of the tubule underlies part of the laminate layer that coats the flaps. Thus the tubule is not continuous with the capsule wall but structurally separate from it. This helps reconcile differences in understanding of the number of layers constituting the capsule wall, and makes clear that the tubule should be considered part of the capsule contents.