Mechanism of barotaxis in marine zooplankton.

Hydrostatic pressure is a dominant environmental cue for vertically migrating marine organisms but the physiological mechanisms of responding to pressure changes remain unclear. Here, we uncovered the cellular and circuit bases of a barokinetic response in the planktonic larva of the marine annelid Platynereis dumerilii. Increased pressure induced a rapid, graded, and adapting upward swimming response due to the faster beating of cilia in the head multiciliary band. By calcium imaging, we found that brain ciliary photoreceptors showed a graded response to pressure changes. The photoreceptors in animals mutant for ciliary opsin-1 had a smaller sensory compartment and mutant larvae showed diminished pressure responses. The ciliary photoreceptors synaptically connect to the head multiciliary band via serotonergic motoneurons. Genetic inhibition of the serotonergic cells blocked pressure-dependent increases in ciliary beating. We conclude that ciliary photoreceptors function as pressure sensors and activate ciliary beating through serotonergic signalling during barokinesis.

Exploring the Antimicrobial Potential of Hallachrome, a Defensive Anthraquinone from the Marine Worm Halla parthenopeia (Polychaeta).

Antimicrobial resistance is a critical global health issue, with rising resistance among bacteria and fungi. Marine organisms have emerged as promising, but underexplored, sources of new antimicrobial agents. Among them, marine polychaetes, such as Halla parthenopeia, which possess chemical defenses, could attract significant research interest. This study explores the antimicrobial properties of hallachrome, a unique anthraquinone found in the purple mucus of H. parthenopeia, against Gram-negative bacteria (Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 9027), Gram-positive bacteria (Enterococcus faecalis ATCC 29212, Staphylococcus aureus ATCC 6538, Staphylococcus epidermidis ATCC 12228), and the most common human fungal pathogen Candida albicans ATCC 10231. Antibacterial susceptibility testing revealed that Gram-negative bacteria were not inhibited by hallachrome at concentrations ≤2 mM. However, Gram-positive bacteria showed significant growth inhibition at 0.12-0.25 mM, while C. albicans was inhibited at 0.06 mM. Time-kill studies demonstrated dose-dependent growth inhibition of susceptible strains by hallachrome, which exerted its effect by altering the membrane permeability of C. albicans, E. faecalis, and S. epidermidis after 6 h and S. aureus after 24 h. Additionally, hallachrome significantly reduced biofilm formation and mature biofilm in S. aureus, E. faecalis, and C. albicans. Additionally, it inhibited hyphal growth in C. albicans. These findings highlight hallachrome's potential as a novel antimicrobial agent, deserving further exploration for clinical experimentation.

Nanoplastics enhanced the developmental toxicity of aromatic disinfection byproducts to a marine polychaete at non-feeding early life stage.

Micro/nanoplastics can act as vectors for organic pollutants and enhance their toxicity, which has been attributed to the ingestion by organisms and the "Trojan horse effect". In this study, we disclosed a non-ingestion pathway for the toxicity enhancement effect of nanoplastics. Initially, the combined toxicity of polystyrene microplastics (40 µm) or nanoplastics (50 nm) with three disinfection byproducts (DBPs) to a marine polychaete, Platynereis dumerilii, was investigated. No toxic effect was observed for the micro/nanoplastics alone. The microplastics showed no effect on the toxicity of the three DBPs, whereas the nanoplastics significantly enhanced the toxicity of two aromatic DBPs when the polychaete was in its non-feeding early life stage throughout the exposure period. The microplastics showed no interaction with the P. dumerilii embryos, whereas the nanoplastics agglomerated strongly on the embryonic chorion and fully encapsulated the embryos. This could contribute to higher actual exposure concentrations in the microenvironment around the embryos, as the concentrations of the two aromatic DBPs on the nanoplastics were 1200 and 120 times higher than those in bulk solution. Our findings highlight an important and previously overlooked mechanism by which nanoplastics and organic pollutants, such as DBPs, pose a higher risk to marine species at their vulnerable early life stages. This study may contribute to a broader understanding of the environmental impacts of plastic pollution and underscore the necessity to mitigate their risks associated with DBPs.

A new genus of Herpyllobiidae (Copepoda: Cyclopoida) from a deep-living annelid (Polychaeta: Sigalionidae).

A taxonomic study of deep-sea polychaetes collected at a depth of 2,805 m off the northern coast of California revealed a scaleworm of the family Sigalionidae with an attached parasitic copepod. The copepod represents an undescribed genus of the family Herpyllobiidae, comprising mesoparasitic copepods chiefly recorded from polychaetes of the family Polynoidae. Blakerius gen. nov. diverges from the other herpyllobiid genera by its possession of 1) a chalice-shaped ectosoma with several protuberances along the posterior margin and a long cylindrical shaft with a hyaline coating and integumental sculpturing, a short stalk with a small, anteriorly placed sclerotized ring, 2) a relatively large, discoid-shaped endosoma with digitiform process, and 3) attached male copepodids with 3-segmented antennules, containing limbless sac-like males. The new genus is compared with other herpyllobiids. This discovery increases the number of known herpyllobiid genera to six and is the first record of a herpyllobiid parasitizing a sigalionid polychaete.urn: lsid: zoobank.org:pub:5E31FEED-D3EB-460E-AEA4-02A9D3A778D6.

Transfer and bioaccumulation of chemical and biological contaminants in the marine polychaete Hediste diversicolor (OF müller 1776) when reared on salmon aquaculture sludge.

Side streams from aquaculture production such as fish sludge poses ample opportunities for biological upcycling, as the sludge contains high amounts of nutrients, energy and valuable biochemicals, making it an ideal food for extractive species. Sludge has been proposed as a feed stock for polychaete production, which in turn can be utilized live in shrimp aquaculture or as an aquafeed ingredient. However, the biosafety of such value chains has not yet been addressed. We conducted an experiment exposing the polychaete Hediste diversicolor to aquaculture sludge spiked with four different fish pathogens (Mycobacterium salmoniphilum, Yersinia ruckeri, Infectious Pancreatic Necrosis (IPN) and Infectious Salmon Anaemia (ISA)) known to cause diseases in Atlantic salmon (Salmo salar L.). Moreover, we assessed whether heavy metals and other potentially hazardous elements present in fish sludge bioaccumulates in the polychaetes. Neither of the bacteria nor viruses could be detected in the polychaetes after 14 days of continuous exposure. Seven of the 15 elements we analysed showed bioaccumulation factors significantly below one, meaning biodilution, while the other eight did not differ from one, meaning no bioaccumulation. None of the elements showed a significant bioaccumulation. Further on, none of the heavy metals found in the polychaetes at the end of our experiment exceeded the EU regulatory maximum levels for fish feed ingredients. The current results suggest that a H. diversicolor can reared on aquaculture sludge, and aquaculture sludge may serve as feed stock for polychaete production without the product exceeding EU regulations for contaminants in animal feed.

Biodegradation of four polyolefin plastics in superworms (Larvae of Zophobas atratus) and effects on the gut microbiome.

Recent studies have demonstrated superworms (larvae of Zophobas atratus) ability to degrade polyethylene (PE), polystyrene (PS), polyvinyl chloride (PVC), and polypropylene (PP) within their digestive system. This study aimed to compare the ability of superworms to degrade the above four polyolefin plastics over a duration of 30 days. In this study, the degradation rate of PE was the highest, and the final average weight of superworms, as well as the final plastic mass loss consumed by them, significantly increased (73.38 % and 52.33 %, respectively) when PE was fed with wheat bran (1:1 [w/w]). FTIR and TGA indicated the occurrence of oxidation and biodegradation processes in the four polyolefin plastics when exposed to superworms. In addition, the molecular weights (Mw and Mn) of excreted polymer residues decreased by 3.1 % and 2.87 % in PE-fed superworms, suggesting that the depolymerization of PE was not entirely dependent on the gut microbial community. The analysis of the gut microbial communities revealed that the dominant microbial community were different for each type of plastic. The results indicate that the gut microbiome of superworms exhibited remarkable adaptability in degrading various types of plastics, and the intake preferences and efficiency of different plastics are associated with different dominant microbial community species.

Fauna - Microplastics interactions: Empirical insights from benthos community exposure to marine plastic waste.

Microplastic deposition in soft marine sediments raises concerns on their role in sediment habitats and unknown effects on resident macrobenthic communities. To assess the reciprocal influence that MPs and macrobenthos might have on each other, we performed a mesocosm experiment with ambient concentrations of environmental Polyethylene (PE) and a non-manipulated, natural macrobenthic community from the Belgian part of the North Sea (BPNS). Our results show that PE fragments increase mortality of abundant bivalves (specifically Abra alba) after 30 days of exposure but not for the most abundant polychaete Owenia fusiformis, possibly due to its predominant suspension feeding behavior. Fast burial of surface MPs exposes deep-dwelling burrowers to the pollutant, however reducing the amount of MPs interacting with (sub) surface living fauna. We conclude that macrobenthos promotes the sequestration of deposited MPs, counteracting resuspension, and can have cascading effects on biodiversity due to their effect on abundant and functionally important species.

Aggregative versus solitary settlement in Spirobranchus cariniferus gray, 1843 (serpulinae). What is the trade-off?

Sessile marine invertebrates usually follow a distinct pattern of living in dense aggregations or as solitary individuals. However, at least some serpulins, including Spirobranchus cariniferus, seem to be able to settle aggregative or solitary. To understand how living solitary or in aggregation is beneficial, it is essential to understand the advantages and disadvantages of both settlement strategies for sessile invertebrates. Benefits of living in aggregations include securing suitable habitat, improving the probability of survival by mitigating physical stress and increasing reproductive success. However, living in patches also comes with some disadvantages for the individual, such as higher intra- and interspecific competition for food, space and oxygen. Increased physiological stress can lead to increased mortality and decreased reproductive success, whereas solitary individuals could produce more gametes because of a lack of competition for food and space. On the other hand, predators would have easier access to them, and the possibility of fertilisation success may be lower because of a lack of synchronisation and a greater distance between individuals of different sexes. These issues have not been sufficiently addressed, particularly for serpulids. Individuals of the New Zealand endemic polychaete Spirobranchus cariniferus can be found solitary and aggregative in the same habitat. Therefore, this study is the first on serpulids comparing the growth and mortality of individuals living alone or in aggregations. Hence, bi-monthly observation of mortality and growth measurements were conducted on tagged individuals in the field, and weekly observations were conducted in a laboratory-based study on individuals of both settlement configurations. A final comparison of body metrics to tube dimensions was made by removing an individual from their tube. My findings revealed that while solitary and aggregative individuals elongate their tubes at a similar rate, further correlations of the body to tube sizes lead to the conclusion that solitary worms focus more of their energy on tube growth rather than body size increment than aggregative conspecifics. Mortality is highly variable and seems not to differ between both configurations. However, individuals living in a patch can better recover from damage to their tubes. Here presented observations hopefully initiated further studies into the effects of aggregation size and density on individual growth. Results of this and subsequent studies can inform the management efforts for reefs of serpulins, bivalves and other invertebrates.

Metabolomic and biochemical disorders reveal the toxicity of environmental microplastics and benzo[a]pyrene in the marine polychaete Hediste diversicolor.

Recently, the abundance of environmental microplastics (MPs) has become a global paramount concern. Besides the danger of MPs for biota due to their tiny size, these minute particles may act as vectors of other pollutants. This study focused on evaluating the toxicity of environmentally relevant concentrations of MPs (10 and 50 mg/kg sediment) and benzo[a]pyrene (B[a]P, 1 µg/kg sediment), alone and in mixture, for 3 and 7 days in marine polychaete Hediste diversicolor, selected as a benthic bioindicator model. The exposure period was sufficient to confirm the bioaccumulation of both contaminants in seaworms, as well as the potential capacity of plastic particles to adsorb and vehiculate the B[a]P. Interestingly, increase of acidic mucus production was observed in seaworm tissues, indicative of a defense response. The activation of oxidative system pathways was demonstrated as a strategy to prevent lipid peroxidation. Furthermore, the comprehensive Nuclear Magnetic Resonance (NMR)-based metabolomics revealed significant disorders in amino acids metabolism, osmoregulatory process, energetic components, and oxidative stress related elements. Overall, these findings proved the possible synergic harmful effect of MPs and B[a]P even in small concentrations, which increases the concern about their long-term presence in marine ecosystems, and consequently their transfer and repercussions on marine fauna.

Hexavalent chromium damages intestinal cells and coelomocytes and impairs immune function in the echiuran worm Urechis unicinctus by causing oxidative stress and apoptosis.

Hexavalent chromium (Cr(VI)) is a common pollutant in the marine environment, which impairs immunity and causes reproductive and heredity disorders in organisms. To clarify the immunotoxic effects of Cr (VI) on the marine worm Urechis unicinctus, we analyzed tissue damage and immune dysfunction caused by Cr (VI) in this organism at histopathologic, zymologic, apoptotic and molecular levels. The results indicated that the bioaccumulation of Cr (VI) bioaccumulation levels in coelomocytes was significantly higher than in the intestines and muscles. Pathological observation showed that Cr (VI) caused damage to the respiratory intestine, stomach and midgut. Cr (VI) also increased the replication of goblet cells and a reduction in the replication of epithelial cells. Meanwhile, Cr (VI) induced apoptosis of intestinal cells and coelomocytes, accompanied by an increase in the expression of Caspase-3, COX-2, and MyD88 in the intestine and coelomocytes. At the same time, Cr (VI) significantly affected the activities of antioxidant enzymes such as SOD, ACP, CAT, CAT, and GST, and increased H2O2 and MDA contents in U. unicinctus. Moreover, Cr (VI) exposure also up-regulated the transcription of hsc70, mt and jnk genes but decreased that of sod in the intestines. In contrast, Cr (VI) down-regulated the expression of sod, hsc70, mt, and jnk genes in coelomocytes. Collectively, Cr (VI) bioaccumulated in U. unicinctus cells and tissues, causing several histopathological changes, oxidative stress, and apoptosis of several cells in the organism, resulting in intestinal and coelomocyte damage and immune dysfunctioning.

A new genus and species of nudibranch-mimicking Syllidae (Annelida, Polychaeta).

Nudibranch mollusks, which are well-known for their vivid warning coloration and effective defenses, are mimicked by diverse invertebrates to deter predation through both Müllerian and Batesian strategies. Despite extensive documentation across different taxa, mimickers have not been detected among annelids, including polychaetes, until now. This study described a new genus and species of polychaete living on Dendronephthya octocorals in Vietnam and Japan. Belonging to Syllidae, it exhibits unique morphological adaptations such as a low number of body segments, simple chaetae concealed within the parapodia and large and fusiform antennae and cirri. Moreover, these appendages are vividly colored, featuring an internal dark red area with numerous terminal white spots and bright yellow tips, effectively contributing to mimicking the appearance of a nudibranch. This discovery not only documents the first known instance of such mimicry among annelids, but also expands our understanding of evolutionary adaptation and ecological strategies in marine invertebrates.

Variations in cell plasticity and proliferation underlie distinct modes of regeneration along the antero-posterior axis in the annelid Platynereis.

The capacity to regenerate lost tissues varies significantly among animals. Some phyla, such as the annelids, display substantial regenerating abilities, although little is known about the cellular mechanisms underlying the process. To precisely determine the origin, plasticity and fate of the cells participating in blastema formation and posterior end regeneration after amputation in the annelid Platynereis dumerilii, we developed specific tools to track different cell populations. Using these tools, we find that regeneration is partly promoted by a population of proliferative gut cells whose regenerative potential varies as a function of their position along the antero-posterior axis of the worm. Gut progenitors from anterior differentiated tissues are lineage restricted, whereas gut progenitors from the less differentiated and more proliferative posterior tissues are much more plastic. However, they are unable to regenerate the stem cells responsible for the growth of the worms. Those stem cells are of local origin, deriving from the cells present in the segment abutting the amputation plane, as are most of the blastema cells. Our results favour a hybrid and flexible cellular model for posterior regeneration in Platynereis relying on different degrees of cell plasticity.

Toxic effect of fluorene on Perinereis aibuhitensis body wall and its corresponding defense mechanisms: A metabolomics perspective.

Fluorene is a coastal sediment pollutant with high ecological risk. Perinereis aibuhitensis is an ecotoxicological model used for polycyclic aromatic hydrocarbon bioremediation; however, the effects of fluorene on the physiological metabolism of P. aibuhitensis and its corresponding responses remain unclear. This study explored the tolerance and defense responses of P. aibuhitensis in sediments with different fluorene concentrations using histology, ecological biomarkers, and metabolic responses. Metabolomics analyses revealed that P. aibuhitensis has high tolerance to fluorene in sediments. Fluorene stress disrupted the normal metabolism of the P. aibuhitensis body wall, resulting in excessive glycosphospholipid and stearamide accumulation and elevated oxygen consumption rates. To mitigate this, P. aibuhitensis has adopted tail cutting, yellowing, and modulation of metabolite contents in the body wall. This study provides novel insights into the potential ecological risk of fluorene pollution in marine sediments and proposes the use of P. aibuhitensis in the bioremediation of fluorene-contaminated sediments.

Bacterial envelope polysaccharide cues settlement and metamorphosis in the biofouling tubeworm Hydroides elegans.

Metamorphosis for many marine invertebrates is triggered by external cues, commonly produced by bacteria. For larvae of Hydroides elegans, lipopolysaccharide (LPS) from the biofilm-dwelling bacterium Cellulophaga lytica induces metamorphosis. To determine whether bacterial LPS is a common metamorphosis-inducing factor for this species, we compare larval responses to LPS from 3 additional inductive Gram-negative marine biofilm bacteria with commercially available LPS from 3 bacteria not known to induce metamorphosis. LPS from all the inductive bacteria trigger metamorphosis, while LPS from non-inductive isolated marine bacteria do not. We then ask, which part of the LPS is the inductive element, the lipid (Lipid-A) or the polysaccharide (O-antigen), and find it is the latter for all four inductive bacteria. Finally, we examine the LPS subunits from two strains of the same bacterial species, one inductive and the other not, and find the LPS and O-antigen to be inductive from only the inductive bacterial strain.

Single-cell RNA-seq reveals distinct metabolic "microniches" and close host-symbiont interactions in deep-sea chemosynthetic tubeworm.

Vestimentiferan tubeworms that thrive in deep-sea chemosynthetic ecosystems rely on a single species of sulfide-oxidizing gammaproteobacterial endosymbionts housed in a specialized symbiotic organ called trophosome as their primary carbon source. While this simple symbiosis is remarkably productive, the host-symbiont molecular interactions remain unelucidated. Here, we applied an approach for deep-sea in situ single-cell fixation in a cold-seep tubeworm, Paraescarpia echinospica. Single-cell RNA sequencing analysis and further molecular characterizations of both the trophosome and endosymbiont indicate that the tubeworm maintains two distinct metabolic "microniches" in the trophosome by controlling the availability of chemosynthetic gases and metabolites, resulting in oxygenated and hypoxic conditions. The endosymbionts in the oxygenated niche actively conduct autotrophic carbon fixation and are digested for nutrients, while those in the hypoxic niche conduct anaerobic denitrification, which helps the host remove ammonia waste. Our study provides insights into the molecular interactions between animals and their symbiotic microbes.

Critical role of benthic fauna in enhancing nanoplastics removal in constructed wetland: Performance, fate and mechanism.

The presence of nanoplastics (NPs) in wastewater poses a considerable risk to ecosystems. Although constructed wetlands (CWs) have the potential to removal NPs, their efficiency is limited by insufficient consideration of ecosystem integrity. Herein, three typical benthic fauna (Corbicula fluminea, Chironomus riparius and Tubifex tubifex) were added to CWs to improve the ecological integrity of CWs, and further enhance the ecological benefits. Results indicated that the addition of C. fluminea, C. riparius and T. tubifex increased NPs removal by 19.14 %, 17.02 %, and 15.76 % than that without benthic faunas, respectively. Based on fluorescence signal analysis, the presence of benthic fauna could intake NPs, and enhanced the adsorption of NPs by plants. The addition of C. fluminea significantly increased catalase (1541.82 ± 41.35 U/g), glutathione S-transferase (0.34 ± 0.02 U/g), and superoxide dismutase (116.33 ± 6.91 U/g) activities (p < 0.05) as a defense mechanism against NPs-induced oxidative stress. Metagenomic analysis revealed that the abundances of key enzymes involved in glycolysis, the tricarboxylic acid cycle, and polystyrene metabolism pathways were increased when C. fluminea was added, corresponding to the microbial degradation of NPs. Overall, the results of this study implied that the benthic fauna can efficiently remove NPs from wastewater in CWs.

Discovery and characterization of a transient chaetal gland during the development of Capitella teleta (Sedentaria: Annelida).

Chaetae are among the most extensively studied structures in polychaetes, serving as a defining morphological trait for annelids. Capitella teleta stands out as one of the few established annelid models for developmental and morphological studies, thus receiving significant scholarly attention. In this study, we unveil a previously unnoticed glandular structure associated with chaetae within the larvae of C. teleta. Our investigations demonstrate the absence of comparable structures in the chaetal follicles of adults and juveniles (older than 1 week), as well as during active chaetogenesis, underscoring the transient nature of these glands. This indicates that larval chaetal follicles transform into a gland that later disappears. Utilizing histology and transmission electron microscopy, we characterized these glands. Our findings underscore the diversity of chaetal ultrastructure in annelids and show that, even in well-studied species, novel morphological details can be found. We emphasize the importance of examining various life-history stages to capture such transient morphological features. This work lays a crucial morphological foundation and deepens our understanding of chaetae and chaetogenesis in C. teleta, paving the way for more accurate interpretations of future experimental studies on chaetogenesis in this species.

Co-expression analysis reveals distinct alliances around two carbon fixation pathways in hydrothermal vent symbionts.

Most autotrophic organisms possess a single carbon fixation pathway. The chemoautotrophic symbionts of the hydrothermal vent tubeworm Riftia pachyptila, however, possess two functional pathways: the Calvin-Benson-Bassham (CBB) and the reductive tricarboxylic acid (rTCA) cycles. How these two pathways are coordinated is unknown. Here we measured net carbon fixation rates, transcriptional/metabolic responses and transcriptional co-expression patterns of Riftia pachyptila endosymbionts by incubating tubeworms collected from the East Pacific Rise at environmental pressures, temperature and geochemistry. Results showed that rTCA and CBB transcriptional patterns varied in response to different geochemical regimes and that each pathway is allied to specific metabolic processes; the rTCA is allied to hydrogenases and dissimilatory nitrate reduction, whereas the CBB is allied to sulfide oxidation and assimilatory nitrate reduction, suggesting distinctive yet complementary roles in metabolic function. Furthermore, our network analysis implicates the rTCA and a group 1e hydrogenase as key players in the physiological response to limitation of sulfide and oxygen. Net carbon fixation rates were also exemplary, and accordingly, we propose that co-activity of CBB and rTCA may be an adaptation for maintaining high carbon fixation rates, conferring a fitness advantage in dynamic vent environments.

An effective and rapidly degradable disinfectant from disinfection byproducts.

Chloroxylenol is a worldwide commonly used disinfectant. The massive consumption and relatively high chemical stability of chloroxylenol have caused eco-toxicological threats in receiving waters. We noticed that chloroxylenol has a chemical structure similar to numerous halo-phenolic disinfection byproducts. Solar detoxification of some halo-phenolic disinfection byproducts intrigued us to select a rapidly degradable chloroxylenol alternative from them. In investigating antimicrobial activities of disinfection byproducts, we found that 2,6-dichlorobenzoquinone was 9.0-22 times more efficient than chloroxylenol in inactivating the tested bacteria, fungi and viruses. Also, the developmental toxicity of 2,6-dichlorobenzoquinone to marine polychaete embryos decreased rapidly due to its rapid degradation via hydrolysis in receiving seawater, even without sunlight. Our work shows that 2,6-dichlorobenzoquinone is a promising disinfectant that well addresses human biosecurity and environmental sustainability. More importantly, our work may enlighten scientists to exploit the slightly alkaline nature of seawater and develop other industrial products that can degrade rapidly via hydrolysis in seawater.

Integrative taxonomy in Syllis prolifera (Annelida, Syllidae): from a unique cosmopolitan species to a complex of pseudocryptic species.

Syllis prolifera (Syllidae, Syllinae) is an abundant species of marine annelids commonly found in warm to temperate waters worldwide. Although morphological variability occurs among populations, S. prolifera has long been considered a cosmopolitan species, widely distributed in coastal environments, including acidified and polluted areas. However, the increasing number of cases of cryptic and pseudocryptic speciation in several polychaete families in recent years has led us to question whether S. prolifera represents a single globally distributed taxon or is a species complex. To address this question, we conducted an integrative study, combining morphological, ecological and molecular data of 52 S. prolifera specimens collected in different localities across the western Mediterranean Sea and the Gulf of Cadiz. Our phylogenetic and species delimitation analyses that included two mitochondrial DNA markers (COI and 16S rRNA ) were congruent in not considering S. prolifera a unique entity. Five distinct lineages that can also be recognised by certain morphological and ecological traits were identified from these analyses instead. Overall, our study does not support the homogeneity of S. prolifera across the Mediterranean Sea, providing a new example of pseudocrypticism in marine invertebrates.