Dondersia festiva Hubrecht, 1888, by monotypy. Type locality. Mediterranean Sea (northern Gulf of Naples); 60 m.

Holotype : SH20364 (USNM 1718003). Gulf of Mexico. 28°5'42.18"N, 92°0'20.38"W. 82 m depth. Serial sections (23 slides), light microscopy preparations of the sclerites (two slides, sclerite from mid-body); SEM stub with sclerites; COI, 16S, and CytB sequences (NCBI PQ246886, PQ249005, and PQ241521, respectively).

Named after Dr. Sarah Tweedt who provided us with the material and for her outstanding work studying invertebrate biodiversity using ARMS; tweedtae is feminine in the genitive.

Elongate animal (~ 14 mm), bright pink with a yellow dorsal keel bearing 17 distinct lobes. Smooth, scaled appearance with three distinct scale-like sclerite types. Large anterior pedal glands. Atrium with about 14 trilobed papillae. Mouth separated from the atrium. Ventrolateral foregut glands of type A. Monoserial radula with two denticles joined at their apex. Midgut with a short dorsal caecum, without lateral constrictions. With five dorsoterminal sensory organs. Without accessory copulatory structures.

Description based on the holotype. Reconstruction of the internal anatomy based on manual reconstruction of the histological sections (Fig. 8A, A’).

Habitus. Long animal (14 mm, 0.55 mm wide in the midbody), pink color aside from the bright yellow, continuous dorsal keel composed of 17 serially arranged lobes (Fig. 2A). Body with shiny and slightly scaly appearance Coloration fades to off-white in 95% ethanol (Fig. 2B). Animal extends and contracts the body significantly; it varies in length, ranging from ~ 6–14 mm, and its width spans from 0.8–2 mm. (Fig. 2A). Tapered anterior. Posterior with a slight finger-like projection. Pedal groove, mantle cavity and mouth apertures visible externally (Fig. 2B).

Mantle. Thin epidermis (17.54–36.57 μm thick, thickness decreases to ~ 10 μm in areas of the posterior end of the body) without epidermal papillae. Three types of sclerites as scales inserted in one layer (Fig. 3): 1) Oval-shaped scales, relatively small (14–17.61 μm long, 7.69–9.79 μm wide) with a proximal rim and an elongated distal end (Fig. 3B, D), most common type, which forms a base layer across the entire body; 2) Lanceolate scales, long and narrow (38.57–39.75 μm long, 5.28–6.86 μm wide) with an acute distal end (Fig. 3B, C, F), distributed intermittently among the oval-shaped sclerites and are less abundant and shorter on the lobes of the dorsal keel; and 3) Pedunculated paddle-like (i.e., oar-shaped scales (Fig. 3B, E; 38.57–39.75 μm long, 5.28–6.86 μm wide), ‘paddle’ portion with a proximal rim, distal edge finely serrated ending in an acuminate point. Paddle-like sclerites inserted in the cuticle amongst the oval-shaped scales, found in the dorsal keel. Scales of the pedal groove not observed.

Pedal groove and mantle cavity. Pedal pit (100 μm long, 165 μm wide, 100–140 μm high) located posteriorly to the mouth. Pedal glands very large, reaching the dorsal part of the body, surrounding the foregut (Fig. 4B–E). Well-defined pedal groove with a single triangular fold (30–60 μm wide, 40–65 μm tall). Mantle cavity (170 μm long, 320 μm high in the middle region) opens ventrally, with posterior pouch (Fig. 4O). Without respiratory folds, walls of the mantle cavity appear slightly folded and ciliated (Fig. 4N).

Nervous system and sensory organs. Cerebral ganglion circular to oval shape in cross section (85 μm long, 50–180 μm wide, 57–110 μm high; Fig. 4C, D). Atrium (182 μm long, 120–200 μm wide 100 μm–260 μm high) opens ventrally with about 24 atrial papillae distally trilobed (27.5–52.5 μm long, 2.5–7.5 μm wide). Five dorsoterminal sensory organs observed both externally and in the serial sections (Fig. 4O).

Digestive system. Mouth opens ventrally, separated from the atrium (Figs 4B, C, 8A). Foregut rounded and narrow (50–70 μm diameter), surrounded by a glandular epithelium and a thin muscular layer. Monoserial radula composed of a broad, non-serrated base (~ 20–25 μm wide, 5–10 μm high) and two long and narrow denticles that join at their apex (20–25 μm high, 2.5–5 μm wide; Fig. 4E, E’). Fragments of what seems to be two small lateral teeth observed in the edges of the base (Fig. 4E’). Radular sac extends posteriorly (Fig. 4F; 35 μm long, and up to 45 μm wide, 75 μm high). Ventrolateral foregut glands of type A (García-Álvarez and Salvini-Plawen 2007) join the foregut via a common opening (Fig. 4E). Esophagus (95 μm long, 35–40 μm in diameter) forms a sphincter as it joins the midgut centrally (Fig. 8A). Midgut with a single dorso-anterior caecum (Fig. 8A) that contains cnidocytes (Fig. 4F, F’), also found in the midgut. Rectum (80–150 μm in diameter) discharges dorsally into the mantle cavity.

Gonopericardial system. Mature animal. Large pericardium (640 μm long, 100 to 530 μm diameter; significantly narrow in its posterior region: 70 μm diameter) (Fig. 8A’), closely associated with gonads, separated only by a thin tissue layer without defined gonoducts (Fig. 4G, H). Heart attached to the dorsal wall of the pericardium (Fig. 4H). Short pericardioducts (60 μm long, 10–20 μm diameter) that connect to the very posterior end of the pericardium and with the spawning ducts in their origin (Fig. 4I). One seminal vesicle attached to each pericardioduct (Fig. 4I). Fused region of the spawning ducts (400 μm long, up to 320 μm in diameter) about double the length of the paired region (Fig. 4J). Spawning ducts terminate into the antero-dorsal region of the mantle cavity (Fig. 8A’) as a single duct (Fig. 4M), with two glandular lateral pouches in its posterior region (Figs 4K, L, 8A’).

Anatomy of the dorsal keel. Continuous cuticular dorsal keel made up of 17 lobes covered by cuticle and sclerites. The serially arranged lobes are connected as can be seen externally through the yellow coloration in the living specimen (Fig. 2A). Lateral view of the animal shows how the region between lobes is a bit elevated and thus constitutes a continuous keel. Serial sections show stained dark brown cells (with Mallory’s Trichrome) contained in the lobes. This stained content is concentrated in the cavity of the lobules, but also continues into the cuticle. The fact that this can be seen in all the series of sections is an additional proof to the morphological continuity of the keel (Fig. 4).

Considering the traditional classification of Solenogastres (sensu Salvini-Plawen 1978), the order Pholidoskepia is characterized by a thin cuticle and sclerites as scales. Some authors have identified issues within this order calling for a taxonomic revision (Scheltema 1999; Scheltema and Schander 2000; Scheltema et al. 2012; Bergmeier et al. 2016, 2019; Yap-Chiongco et al. 2024). Nevertheless, the grouping of Pholidoskepia sensu stricto (Yap-Chiongco et al. 2024) is well-supported by the mentioned mantle characteristics and molecular data. Thus, we follow the traditional classification here. Within Pholidoskepia, the mantle sclerites, radula, and type of ventrolateral foregut glands, as well as some posterior organs, are important characters used to classify specimens into a family (García-Álvarez and Salvini-Plawen 2007). Particularly important for the identification of Dondersiidae species is the types of sclerites (Scheltema et al. 2012; Cobo and Kocot 2021). The sclerites of the specimen studied here can be compared to those described previously for species of Dondersia, especially with those of the type species: Dondersia festiva (Hubrecht 1888: fig. 13-2a; Scheltema et al. 2012: figs 1–3). This, with the structure of the radula, justify the classification of the new species within this genus. Moreover, our phylogenetic analysis also supports this classification (see below). The coloration of living specimens is unknown for most solenogasters as most of the species have been described based on preserved material. Within Dondersia, two described species are known to have bright colorations: D. festiva (bright purple) and D. annulata Nierstrasz, 1902 (hot pink with white stripes). Despite similarities, there are clear differences between Dondersia tweedtae sp. nov. and these two species and the remaining species of the genus (reviewed in Cobo and Kocot 2021). Particularly, the combination of pink and yellow coloration, along with the dorsal cuticular lobes, is exclusive to D. tweedtae sp. nov. Moreover, this constitutes the first Dondersiidae from the Gulf of Mexico (Table 3).

Since this new species bears a cuticular keel, the diagnosis of the family, which states the absence of dorsal keel (Scheltema et al. 2012; Cobo and Kocot 2021), needs to be amended to: “elongate body with or without keel. Anterior end tapered, posterior end with a finger-like projection. Leaf-shaped scales as most abundant type of sclerite, with oar-shaped (= pallet-shaped) or laminar scales scattered between them. With or without common atrio-buccal cavity. Monoserial radula; teeth with four denticles; two central denticles fused and curved distally; two lateral, curved denticles arising from a rounded base. With or without dorsoterminal sensory organs. With or without copulatory stylets. Without respiratory folds. With seminal vesicles and with or without seminal receptacles.”

Paramenia sierra Pruvot, 1890, by monotypy. Type locality. Costa Brava (Mediterranean Sea); 80 m. Type material missing (García-Álvarez and Salvini-Plawen 2007).

Holotype : SH20192-A (USNM 1718004) Gulf of Mexico. CYCLE 2021 event ID CYCLE_2021_ARMS_01_DIAback: 28.088295, -90.701405. 82 m depth. Serial sections (16 slides 5 µm), light microscopy preparation of the sclerites (1 slide); SEM stub with sclerites; COI, 16S, and CytB Sequences (NCBI PQ246885, PQ249006, and PQ241520, respectively). Paratype SH20192-B (USNM 1718005) Gulf of Mexico. 28.088295, -90.701405. 82 m depth. Animal preserved in 95% ethanol.

From Latin bullesco, bullescis, bullescere; meaning “to bubble” or “to form bubbles” due to the aspect given by the protrusion of the dorsal keel.

Elongate animal (~ 12 mm), light orange with a discontinuous dorsal keel with protrusions as lobes (number variable, protrusions simple or trilobed). Sclerites as hollow acicular spines, with hook-shaped and harpoon-shaped sclerites. Without epidermal papillae. Mouth and atrium partially separated. Atrium with numerous (≤20) single and branched papillae. Distichous radula. Ventrolateral foregut gland of type A / Pararrhopalia type. Foregut with a dorso-pharyngeal papilla gland. With 12 respiratory folds. With abdominal spicules. With one dorsoterminal sensory organ.

Description based on the holotype, external aspect of the paratype also considered. Reconstruction of the internal anatomy (Fig. 8B, B’) built from the manual reconstruction based on serial sections of the holotype.

Habitus. Elongate animal (10–12 × 0.5–1 mm), light orange in life (Fig. 5A, B), but white after preservation in ethanol (Fig. 5C). With a dorsal, discontinuous keel formed by nearly spherical lobes of different sizes. Lobes without organized arrangement, which varies between the holotype and paratype and depending on the degree of extension of the body. Lobes single or grouped as pairs or groups of three.

Mantle. Thin cuticle (18.31–27.6 μm) without distinct papillae and with five main types of hollow acicular sclerites protruding from it (Fig. 6): 1) Hook-shaped sclerites (Fig. 6A–C; 80–90 × 9–6 µm; the inner part of the hook is 30 µm long) with a small distal protrusion and a short internal region of the hook are particularly abundant in the dorsal region of the body and on the dorsal lobes; 2) Harpoon-shaped sclerites (Fig. 6A, D; 200–210 × 8 µm), present all over the body and are the dominant sclerite type in the mid-ventral region, also larger than elsewhere on the body (Fig. 6E; 200–300 × 8–10 µm); 3) Very thin and long acicular sclerites (Fig. 6D, E; 80–150 × 2 µm), distributed all over the surface of the body, but are less abundant in the dorsal lobes; 4) Acicular sclerites that look almost flat, but are hollow and elliptical in cross-section (Fig. 6A, D–F; 100–160 × 10 µm), present all over the body; 5) Slightly curved acicular sclerites of varying length (Fig. 6A, F; 80–160 × 6–7 µm), present all over the body. With knife-shaped scales characteristic of the pedal groove.

Pedal groove and mantle cavity. Small pedal pit (90 µm long, 16 µm wide, 4–6 µm high). Pedal grove well marked, extending along the entire length of the body, with a single wide triangular pedal fold (Fig. 7H–L; 5–10 µm wide in the middle region of the fold × 5–12 µm high). Mantle cavity with 12 unbranched respiratory folds (Fig. 7M, N).

Nervous system and sensory organs . Cerebral ganglion of circular shape in cross section (Fig. 7B–D; 35 μm long, 20 to 22 μm wide, 10 to 14 μm high). Atrium (160 μm long, 17 to 34 μm wide, 8 to 10 μm high) with numerous (≤20) single and branched papillae (Fig. 7A, B). Without dorsoterminal sensory organ.

Digestive system. Mouth and atrium partially separated (mouth separated from the atrium by a ridge with musculature but without cuticle; Fig. 7C). Mouth (Fig. 7D) leads to a rounded foregut that enlarges dorsally, where it forms a connection with a dorso-pharyngeal papilla gland (Fig. 7D–F). Short radular sac (Figs 7G, 8B). Ventrolateral foregut glands of type-A (García-Álvarez and Salvini-Plawen 2007) / Pararrhopalia-type (Handl and Todt 2005 that are very glandular posteriorly (Fig. 7H). Radula distichous, formed by hook-shaped teeth (radula broken in the sections so the number of middle denticles, if present, cannot be estimated). Midgut with a dorsal caecum that projects anteriorly above the foregut and dorsal pharyngeal gland (Figs 7G, 8B) and marked lateral constrictions (Fig. 7I). Rectum ends dorsally in the mantle cavity (Fig. 7L).

Gonopericardial system. Mature animal. Gonoducts connect with a large pericardium (540 µm long, 40 to 200 µm high). Heart not evident in most of the serial sections. Pericardioducts (340 μm long, 10–20 μm diameter) connect to the posterior end of the pericardium and the mid-posterior spawning duct (Figs 7J, 8B’). Spawning duct paired in most of its longitude (400 μm), ending as a single tube (160 μm long) in the middle of the mantle cavity (Fig. 7K, L). Seminal receptacles attached dorsally to each of the spawning ducts (Fig. 7J), posterior to the fusion of the pericardioducts with the spawning ducts (Fig. 8B’). Without seminal vesicles. Without copulatory stylets. With abdominal spicules.

Anatomy of the dorsal keel. Dorsal keel consists of a discontinuous series of cuticular lobes. Number of lobes variable among individuals (~ 30 in the holotype and 24 in paratype 1; Fig. 5A, B). In living specimens, lobes protrude less from the cuticle when the animal expands the body (Fig. 5B’). This and the orientation of the animal makes it difficult to get an exact number of lobes. Concentration of lobes along the body seems uniform in preserved specimens (Fig. 5C), but in living specimens the density of lobes is higher in the posterior region of the body (Fig. 5A, B). This variation of the dorsal keel in the living specimens and after fixation was already described for Eleutheromenia sierra Pruvot, 1890 (Cobo et al. 2024). The lobes are mostly single in the mid body, but they can occur as pairs or groups of three, especially toward the anterior and posterior ends. In the decalcified animal it was evident that the lobes have a proximal peduncle in connection with the body (Fig. 5D). Both in the living and fixed specimens, and after decalcification it was observed that the lobes are easily detached from the body, breaking at the peduncular area (Fig. 5D). Histology of the dorsal keel is reminiscent of what has been described for the keel of E. sierra (Pruvot 1890; Salvini-Plawen 2003b: fig. 11): connection between lobes not evident externally nor in the histological series. study of the lobes under the microscope after decalcification (Fig. 5D) revealed that they contain an unidentified material that is in some way connected with the internal organs. Content not easily characterized, although diverse types of cells could be observed (Fig. 7P), including one that was tentatively identified as a cnidocyte (Fig. 7O).

The presence of hollow sclerites with a hook-shaped distal end is characteristic of two subfamilies within the family Pruvotinidae: Pruvotininae Heath, 1911 and Eleutheromeniinae Salvini-Plawen, 1978 (García-Álvarez and Salvini-Plawen 2007). The main distinguishing feature between these subfamilies is the presence of a dorso-pharyngeal papilla gland in Pruvotininae (García-Álvarez and Salvini-Plawen 2007; Pedrouzo et al. 2022). Although several works have shown that some of the diagnostic characters of the family are somehow ambiguous and the group is in need of systematic revision (García-Álvarez and Salvini-Plawen 2007; Zamarro et al. 2013; Pedrouzo et al. 2022; Martinez-Sanjuán 2024), the presence/absence of the dorso-pharyngeal papilla gland has been considered as a good diagnostic character to distinguish these subfamilies (Cobo et al. 2024). The new species described here has a dorso-pharyngeal papilla gland, which would place it within Pruvotininae.

The subfamily Pruvotininae includes three genera: Pruvotina Cockerell, 1903; Pararrhopalia Simroth, 1893 and Labidoherpia Salvini-Plawen, 1978. Traditionally, these three genera are distinguished by a combination of internal morphological characters including the presence/absence of atrio-buccal cavity (García-Álvarez and Salvini-Plawen 2007; Zamarro et al. 2013; Pedrouzo et al. 2022). Nevertheless, it was recently concluded (Cobo et al. 2024) that this is not a valid character to differentiate between genera in the subfamily Pruvotininae, and that the only apparently reliable defining morphological characteristics among these genera are the respiratory folds and copulatory stylets: respiratory folds are present both Pruvotina and Labidoherpia, but absent in Pararrhopalia (García-Álvarez and Salvini-Plawen 2007; Pedrouzo et al. 2022) and Pruvotina is the only genus in the subfamily that lacks copulatory stylets. The species described here has respiratory folds and lacks copulatory stylets and thus it would be classified as Pruvotina. However, the results of our phylogenetic analysis (see below and Fig. 9) place the new species as the sister taxon to a species of Eleutheromenia Salvini-Plawen, 1967 (Eleutheromeniinae) with maximal support and thus a classification of the new species based just on internal morphological characters is called into question.

Externally, the new species resembles Eleutheromenia sierra (Pruvot, 1890) due to the lobular dorsal keel (Pruvot 1890; Salvini-Plawen and Ozturk 2006; Zamarro et al. 2013; Cobo et al. 2024). Prior to the discovery of the species described here, E. sierra was the only species within Pruvotinidae with a distinct dorsal keel. A subtle dorsal keel has been described for Pruvotina peniculata Salvini-Plawen, 1978 (Pedrouzo et al. 2022), but while a slight keel can be seen in the drawings included in the original description of the species (Salvini-Plawen 1978: figs 129, 130), this character was not apparent in our study of the sections deposited at the Smithsonian National Museum of Natural History (USNM 1604160, 1604162, 1604163, and 1604174). Further, the study of the syntypes of P. peniculata preserved in ethanol revealed a spiny habitus without a dorsal keel (USNM 749729). Considering the sclerites, the new species has harpoon-shaped hollow acicular sclerites, which is also a characteristic of E. sierra (Pruvot, 1890). This sclerite type has been reported just for one of the 16 species of Pruvotina, P. harpagone Pedrouzo, Garcia-Alvarez & Urgorri, 2022 (Pedrouzo et al. 2022). Therefore, considering the external aspect and sclerites along with the results of our molecular phylogenetic analysis, the new species is classified in the subfamily Eleutheromeniinae, despite the presence of a dorso-pharyngeal papilla gland.

Eleutheromeniinae includes two genera. No radula is present in the monospecific Luitfriedia García-Álvarez & Urgorri, 2001 while a distichous radula was described for the two accepted species of Eleutheromenia, supporting placement of the new species in this genus. Eleutheromenia bullescens sp. nov. can be clearly differentiated from the two known species of the genus. The dorsal keel distinguishes it clearly from E. antarctica (Salvini-Plawen, 1978), which lacks a keel. Despite the similarities in their external aspect, E. bullescens sp. nov. can be easily distinguished from E. sierra. The new species is orange while E. sierra is white to cream when alive (Pruvot 1890; Cobo et al. 2024). Moreover, the arrangement of the ≤ 30 lobes in the new species differs from what was described for E. sierra, which has ~ 17 single lobes, none of which are grouped, and they are spaced more regularly along the body (Cobo et al. 2024). Internally, E. bullescens sp. nov. has a dorso-pharyngeal papilla gland and lacks a glandular esophagus, in contrast to E. sierra (Salvini-Plawen 1978; Pruvot 1890; Pedrouzo et al. 2022). With the new species included here the distribution of Pruvotinidae is extended to the Gulf of Mexico (Table 3).

Considering all the above, a thorough re-evaluation of the systematics of Pruvotinidae is required. In particular, the generation and analysis of molecular data from already described species seem essential, along with a better characterization of the habitus, sclerites, radula, and digestive glands. The currently accepted classification of the family and diagnoses of the subfamilies and genera, if they prove to represent monophyletic groups, need to be amended, but more research is needed to do this adequately. Given the need for a systematic revision of Pruvotinidae, we refrain from formally amending the diagnosis of Eleutheromeniinae but note that the presence of a dorso-pharyngeal papilla gland in the new species is contrary to the current diagnosis of the group.