Effect of 2020-21 and 2021-22 Highly Pathogenic Avian Influenza H5 Epidemics on Wild Birds, the Netherlands.

The number of highly pathogenic avian influenza (HPAI) H5-related infections and deaths of wild birds in Europe was high during October 1, 2020-September 30, 2022. To quantify deaths among wild species groups with known susceptibility for HPAI H5 during those epidemics, we collected and recorded mortality data of wild birds in the Netherlands. HPAI virus infection was reported in 51 bird species. The species with the highest numbers of reported dead and infected birds varied per epidemic year; in 2020-21, they were within the Anatidae family, in particular barnacle geese (Branta leucopsis) and in 2021-22, they were within the sea bird group, particularly Sandwich terns (Thalasseus sandvicensis) and northern gannet (Morus bassanus). Because of the difficulty of anticipating and modeling the future trends of HPAI among wild birds, we recommend monitoring live and dead wild birds as a tool for surveillance of the changing dynamics of HPAI.

Finding food: how generalist predators use contact-chemosensory information to guide prey preferences.

Understanding the processes that guide carnivores in finding and selecting prey is a fundamental, unresolved challenge in sensory biology. To our knowledge, no published work has yet revealed the complete structural identities of compounds that cue preferences by generalist predators for different prey species. With this research imperative in mind, we determined the chemistry driving consumer preferences for live intact prey using two generalist predatory species (sea stars, Pisaster ochraceus; whelks, Acanthinucella spirata), along with two foundation prey species (mussels, Mytilus californianus; barnacles, Balanus glandula), inhabiting rocky, wave-swept shores. Each prey species is known to secrete either a 29.6 kDa (named 'KEYSTONEin') or a 199.6 kDa (named 'MULTIFUNCin') glycoprotein as a contact-chemical cue. Here, experimental manipulations utilized faux prey consisting of cleaned barnacle or mussel shells infused with KEYSTONEin, MULTIFUNCin or seawater (control) gels. Whelks exhibited a strong penchant for MULTIFUNCin over KEYSTONEin, irrespective of shell type. In contrast, sea stars generally preferred KEYSTONEin over MULTIFUNCin, but this preference shifted depending on the experimental context in which they encountered physical (shell) and chemical (glycoprotein) stimuli. This study ultimately demonstrates clear and contrasting chemical preferences between sea stars and whelks. It highlights the importance of experimental setting in determining chemical preferences. Finally, it shows that prey preferences by these predators hinge only on one or two contact-protein cues, without the need for quality coding via fluid-borne compounds, low-molecular-weight substances or mixture blends.

Contrasting effects of shooting disturbance on the movement and behavior of sympatric wildfowl species.

Human-wildlife conflict is a global conservation issue, necessitating effective mitigation strategies. Hunting is a common management approach to reduce conflict, but the indirect consequences are often overlooked. Chronic hunting-related disturbance can reduce fitness and redistribute species. In recent decades, goose-agricultural conflict has intensified due to increasing abundance and shifts towards agricultural foraging. On Islay, Scotland, escalating conflict culminated in shooting Greenland barnacle geese Branta leucopsis to reduce damage to agricultural grassland. In this study, we contrast the impact of shooting disturbance on the movement, behavior, energy expenditure and habitat selection of the target species (Greenland barnacle goose) and a vulnerable nontarget species (Greenland white-fronted goose, Anser albifrons flavirostris) using biologging devices (target species: n = 33; nontarget species: n = 94). Both species were displaced by shooting, and greater distances were subsequently traveled by the target species (1.71 km when directly targeted). When disturbed at any distance, total daily movement increased significantly by 1.18 km for the target species but not for the nontarget species. The target species exhibited no accompanying change in diurnal energy expenditure (measured via accelerometery) but foraged in improved grasslands further from roads after shooting disturbance, where disturbance from all sources was likely lower. The significant increases in movement and changes in foraging site selection of the target species could reduce fitness but given the infrequency of shooting disturbances (0.09 per day) there is likely capacity for compensatory feeding to recoup energetic losses. The nontarget species expectedly showed no significant change in energy expenditure, behavior or habitat selection following shooting disturbance, suggesting mitigation strategies have been effective at minimizing fitness impacts. Refuge areas with a 3.5 km diameter (three times the maximum distance from shooting that displacement was detectable) could provide undisturbed foraging for the target species, minimizing compensatory feeding and further agricultural damage. Wildlife managers should, where possible, consider the fitness implications of shooting disturbance, and whether compensatory feeding and redistribution could hamper conflict mitigation. Management strategies should also include species-specific monitoring and mitigation as we have demonstrated differing responses potentially due to imposed mitigation but also differing species ecology and "landscapes of fear."

The role of indirect interspecific effects in the stochastic dynamics of a simple trophic system.

Indirect interspecific effects (IIEs) occur when one species affects another through a third intermediary species. Understanding the role of IIEs in population dynamics is key for predicting community-level impacts of environmental change. Yet, empirically teasing apart IIEs from other interactions and population drivers has proven challenging and data-demanding, particularly in species-rich communities. We used stochastic population models parameterized with long-term time series of individual data to simulate population trajectories and examine IIEs in a simple high-arctic vertebrate food chain consisting of the wild Svalbard reindeer, its scavenger (the Arctic fox) and the barnacle goose, a migratory prey of the fox. We used the simulated population trajectories to explore co-fluctuations between the species within the food chain. Additionally, we adjusted the model in two ways: first, to isolate the impact of fluctuations in the abundance of a species by keeping its abundance constant; and second, to isolate the impact of a trophic interaction on the dynamics of other species by setting the abundance of the influencing species to zero. We found that fluctuations in reindeer carcasses shaped fox abundance fluctuations, which subsequently affected goose population dynamics. Reindeer and goose population growth rates were nevertheless only weakly correlated, probably in part due to demographic and environmental stochasticity, density dependence and lagged dynamics in the geese. However, removing the fluctuations in reindeer abundance or setting reindeer abundance to zero indeed demonstrated strong underlying IIEs on goose population dynamics and extinction probability. This study thus highlights the importance of species interactions, including IIEs, on species coexistence and communities in the long-term, that is beyond immediate effects and covariation in short-term fluctuations.

Rapid discovery of a new antifoulant: From in silico studies targeting barnacle chitin synthase to efficacy against barnacle settlement.

Due to the adverse environmental impacts of toxic heavy metal-based antifoulants, the screening of environmentally friendly antifoulants has become important for the development of marine antifouling technology. Compared with the traditional lengthy and costly screening method, computer-aided drug design (CADD) offers a promising and efficient solution that can accelerate the screening process of green antifoulants. In this study, we selected barnacle chitin synthase (CHS, an important enzyme for barnacle settlement and development) as the target protein for docking screening. Three CHS genes were identified in the barnacle Amphibalanus amphitrite, and their encoded proteins were found to share a conserved glycosyltransferase domain. Molecular docking of 31,561 marine natural products with AaCHSs revealed that zoanthamine alkaloids had the best binding affinity (-11.8 to -12.6 kcal/mol) to AaCHSs. Considering that the low abundance of zoanthamine alkaloids in marine organisms would limit their application as antifoulants, a marine fungal-derived natural product, mycoepoxydiene (MED), which has a similar chemical structure to zoanthamine alkaloids and the potential for large-scale production by fermentation, was selected and validated for stable binding to AaCHS2L2 using molecular docking and molecular dynamics simulations. Finally, the efficacy of MED in inhibiting cyprid settlement of A. amphitrite was confirmed by a bioassay that demonstrated an EC50 of 1.97 µg/mL, suggesting its potential as an antifoulant candidate. Our research confirmed the reliability of using AaCHSs as antifouling targets and has provided insights for the efficient discovery of green antifoulants by CADD.

More management, less damage? With increasing population size, economic costs of managing geese to minimize yield losses may outweigh benefits.

Conflicts between farmers and geese are intensifying; yet, it remains unclear how interactions between goose population size and management regimes affect yield loss and economic costs. We investigate the cost-effectiveness of accommodation and scaring areas in relation to barnacle goose (Branta leucopsis) population size. We use an existing individual-based model of barnacle geese foraging in nature, accommodation, and scaring areas in Friesland, the Netherlands, to study the most cost-effective management under varying population sizes (i.e., between 20 and 200% of the current size). Our study shows that population size non-linearly affects yield loss costs and total costs per goose. The most cost-effective management scenario for intermediate to large populations is to avoid scaring of geese. For small populations, intensive scaring resulted in minimized yield loss costs and total costs, but also substantially lower goose body mass. Our results strongly suggest that scaring becomes a less effective management measure as goose populations increase.

Gene co-option, duplication and divergence of cement proteins underpin the evolution of bioadhesives across barnacle life histories.

Acquisition of new genes often results in the emergence of novel functions and is a key step in lineage-specific adaptation. As a group of sessile crustaceans, barnacles establish permanent attachment through initial cement secretion at the larval phase followed by continuous cement secretion in juveniles and adults. However, the origins and evolution of barnacle larval and adult cement proteins remain poorly understood. By performing microdissection of larval cement glands, transcriptome and shotgun proteomics and immunohistochemistry validation, we identified 30 larval and 27 adult cement proteins of the epibiotic turtle barnacle Chelonibia testudinaria, of which the majority are stage- and barnacle-specific. While only two proteins, SIPC and CP100K, were expressed in both larvae and adults, detection of protease inhibitors and the cross-linking enzyme lysyl oxidase paralogs in larvae and adult cement. Other barnacle-specific cement proteins such as CP100k and CP52k likely share a common origin dating back at least to the divergence of Rhizocephala and Thoracica. Different CP52k paralogues could be detected in larval and adult cement, suggesting stage-specific cement proteins may arise from duplication followed by changes in expression timing of the duplicates. Interestingly, the biochemical properties of larval- and adult-specific CP52k paralogues exhibited remarkable differences. We conclude that barnacle larval and adult cement systems evolved independently, and both emerged from co-option of existing genes and de novo formation, duplication and functional divergence of lineage-specific cement protein genes. Our findings provide important insights into the evolutionary mechanisms of bioadhesives in sessile marine invertebrates.

Mercury-Modulated Immune Responses in Arctic Barnacle Goslings (Branta leucopsis) upon a Viral-Like Immune Challenge.

Historical mining activities in Svalbard (79°N/12°E) have caused local mercury (Hg) contamination. To address the potential immunomodulatory effects of environmental Hg on Arctic organisms, we collected newborn barnacle goslings (Branta leucopsis) and herded them in either a control or mining site, differing in Hg levels. An additional group at the mining site was exposed to extra inorganic Hg(II) via supplementary feed. Hepatic total Hg concentrations differed significantly between the control (0.011 ± 0.002 mg/kg dw), mine (0.043 ± 0.011 mg/kg dw), and supplementary feed (0.713 ± 0.137 mg/kg dw) gosling groups (average ± standard deviation). Upon immune challenge with double-stranded RNA (dsRNA) injection, endpoints for immune responses and oxidative stress were measured after 24 h. Our results indicated that Hg exposure modulated the immune responses in Arctic barnacle goslings upon a viral-like immune challenge. Increased exposure to both environmental as well as supplemental Hg reduced the level of natural antibodies, suggesting impaired humoral immunity. Hg exposure upregulated the expression of proinflammatory genes in the spleen, including inducible nitric oxide synthase (iNOS) and interleukin 18 (IL18), suggesting Hg-induced inflammatory effects. Exposure to Hg also oxidized glutathione (GSH) to glutathione disulfide (GSSG); however, goslings were capable of maintaining the redox balance by de novo synthesis of GSH. These adverse effects on the immune responses indicated that even exposure to low, environmentally relevant levels of Hg might affect immune competence at the individual level and might even increase the susceptibility of the population to infections.

Sacculina-Induced Morphological Feminization in the Grapsid Crab Pachygrapsus crassipes.

Rhizocephalan barnacles (Thecostraca: Cirripedia) are parasitic crustaceans that lack appendages, segmentation, and a digestive system in adults, while instead infiltrating their hosts with a nutrient-absorbing system of rootlets. Sacculinids, belonging to the Rhizocephala order, are known for their various parasitization-induced effects on their decapod hosts, such as parasitic castration, reduction in the growth of secondary sexual characteristics, feminization of male crabs, and alteration of host behavior. In this study, we conducted field surveys in Japan at Manazuru Town (Kanagawa) on the Pacific coast, and on Sado Island and Noto Peninsula on the Sea of Japan side, and found that sacculinid-parasite-ratios on the grapsid crab Pachygrapsus crassipes were particularly high on the Sea of Japan coast. Molecular phylogenetic analysis revealed that the Manazuru population forms a single clade with Sacculina yatsui, and both Sado and Noto populations form a single clade with S. confragosa. We further demonstrated that external morphologies of male P. crassipes parasitized by sacculinids were changed to female phenotypes. This host-parasite interaction will be a useful model for understanding molecular mechanisms underlying rhizocephalan-driven morphological and behavioral feminization and castration.

Efficacy of amorphous TiOx-coated surfaces against micro- and macrofouling through laboratory microcosms and field studies.

In this study, Soda Lime Glass (SLG) and Stainless Steel (SS316L) substrata coated with Titanium oxide (TiOx) were tested for their efficacy in the laboratory microcosms and in field against micro- and macrofouling. Laboratory microcosm studies were conducted for five days using natural biofilms, single-species diatom (Navicula sp.), and bacterial biofilms, whereas field observations were conducted for 30 days. The TiOx-coating induced change in the mean contact angle of the substratum and rendered SS316L more hydrophilic and SLG hydrophobic, which influenced the Navicula sp. biofilm, and bacterial community structure of the biofilm. Overall, the TiOx-coated SS316L showed minimal microfouling, whereas non-coated SLG exhibited greater efficacy in deterring/preventing macrofouling organisms. Moreover, the reduction in macrofouling could be attributed to high abundance of Actinobacteria. Unraveling the mechanism of action needs future studies emphasizing biochemical processes and pathways.

Bioactive Peptides from Barnacles and Their Potential for Antifouling Development.

Barnacles, a prevalent fouler organism in intertidal zones, has long been a source of annoyance due to significant economic losses and ecological impacts. Numerous antifouling approaches have been explored, including extensive research on antifouling chemicals. However, the excessive utilization of small-molecule chemicals appears to give rise to novel environmental concerns. Therefore, it is imperative to develop new strategies. Barnacles exhibit appropriate responses to environmental challenges with complex physiological processes and unique sensory systems. Given the assumed crucial role of bioactive peptides, an increasing number of peptides with diverse activities are being discovered in barnacles. Fouling-related processes have been identified as potential targets for antifouling strategies. In this paper, we present a comprehensive review of peptides derived from barnacles, aiming to underscore their significant potential in the quest for innovative solutions in biofouling prevention and drug discovery.

Anti-Barnacle Activities of Isothiocyanates Derived from ß-Citronellol and Their Structure-Activity Relationships.

Antifouling agents with low toxicity are in high demand for sustaining marine industries and the environment. This study aimed to synthesize 15 isothiocyanates derived from ß-citronellol and evaluate their antifouling activities and toxicities against cypris larvae of the barnacle Amphibalanus amphitrite. The synthesized isothiocyanates exhibited effective antifouling activities (EC50 =0.10-3.33 µg mL-1 ) with high therapeutic ratios (LC50 /EC50 >30). Four isothiocyanates with an amide or isocyano group showed great potential as effective antifouling agents (EC50 =0.10-0.32 µg mL-1 , LC50 /EC50 =104-833). The enantiomers of the isothiocyanates only slightly differed in their antifouling activities. These results may serve as a basis for further research and development of ß-citronellol-derived isothiocyanates as effective low-toxic antifouling agents. To the best of our knowledge, this study is the first to report the antifouling activities of isothiocyanates derived from accessible natural products.

Postnatal growth rate varies with latitude in range-expanding geese: The role of plasticity and day length.

The postnatal growth period is a crucial life stage, with potential lifelong effects on an animal's fitness. How fast animals grow depends on their life-history strategy and rearing environment, and interspecific comparisons generally show higher growth rates at higher latitudes. However, to elucidate the mechanisms behind this gradient in growth rate, intraspecific comparisons are needed. Recently, barnacle geese expanded their Arctic breeding range from the Russian Barents Sea coast southwards, and now also breed along the Baltic and North Sea coasts. Baltic breeders shortened their migration, while barnacle geese breeding along the North Sea stopped migrating entirely. We collected cross-sectional data on gosling tarsus length, head length and body mass, and constructed population-specific growth curves to compare growth rates among three populations (Barents Sea, Baltic Sea and North Sea) spanning 17° in latitude. Growth rate was faster at higher latitudes, and the gradient resembled the latitudinal gradient previously observed in an interspecific comparison of precocial species. Differences in day length among the three breeding regions could largely explain the observed differences in growth rate. In the Baltic, and especially in the Arctic population, growth rate was slower later in the season, most likely because of the stronger seasonal decline in food quality. Our results suggest that differences in postnatal growth rate between the Arctic and temperate populations are mainly a plastic response to local environmental conditions. This plasticity can increase the individuals' ability to cope with annual variation in local conditions, but can also increase the potential to re-distribute and adapt to new breeding environments.

Isotopes from fossil coronulid barnacle shells record evidence of migration in multiple Pleistocene whale populations.

Migration is an integral feature of modern mysticete whale ecology, and the demands of migration may have played a key role in shaping mysticete evolutionary history. Constraining when migration became established and assessing how it has changed through time may yield valuable insight into the evolution of mysticete whales and the oceans in which they lived. However, there are currently few data which directly assess prehistoric mysticete migrations. Here we show that calcite δ18O profiles of two species of modern whale barnacles (coronulids) accurately reflect the known migration routes of their host whales. We then analyze well-preserved fossil coronulids from three different locations along the eastern Pacific coast, finding that δ18O profiles from these fossils exhibit trends and ranges similar to modern specimens. Our results demonstrate that migration is an ancient behavior within the humpback and gray whale lineages and that multiple Pleistocene populations were undertaking migrations of an extent similar to those of the present day.

Histology and transcriptomic analyses of barnacles with different base materials and habitats shed lights on the duplication and chemical diversification of barnacle cement proteins.

BACKGROUND: Barnacles are sessile crustaceans that attach to underwater surfaces using barnacle cement proteins. Barnacles have a calcareous or chitinous membranous base, and their substratum varies from biotic (e.g. corals/sponges) to abiotic surfaces. In this study, we tested the hypothesis that the cement protein (CP) composition and chemical properties of different species vary according to the attachment substrate and/or the basal structure. We examined the histological structure of cement glands and explored the variations in cement protein homologs of 12 barnacle species with different attachment habitats and base materials. RESULTS: Cement gland cells in the rocky shore barnacles Tetraclita japonica formosana and Amphibalanus amphitrite are eosinophilic, while others are basophilic. Transcriptome analyses recovered CP homologs from all species except the scleractinian coral barnacle Galkinia sp. A phylogenomic analysis based on sequences of CP homologs did not reflect a clear phylogenetic pattern in attachment substrates. In some species, certain CPs have a remarkable number of paralogous sequences, suggesting that major duplication events occurred in CP genes. The examined CPs across taxa show consistent bias toward particular sets of amino acid. However, the predicted isoelectric point (pI) and hydropathy are highly divergent. In some species, conserved regions are highly repetitive. CONCLUSIONS: Instead of developing specific cement proteins for different attachment substrata, barnacles attached to different substrata rely on a highly duplicated cementation genetic toolkit to generate paralogous CP sequences with diverse chemical and biochemical properties. This general CP cocktail might be the key genetic feature enabling barnacles to adapt to a wide variety of substrata.

Environmental change reduces body condition, but not population growth, in a high-arctic herbivore.

Environmental change influences fitness-related traits and demographic rates, which in herbivores are often linked to resource-driven variation in body condition. Coupled body condition-demographic responses may therefore be important for herbivore population dynamics in fluctuating environments, such as the Arctic. We applied a transient Life-Table Response Experiment ('transient-LTRE') to demographic data from Svalbard barnacle geese (Branta leucopsis), to quantify their population-dynamic responses to changes in body mass. We partitioned contributions from direct and delayed demographic and body condition-mediated processes to variation in population growth. Declines in body condition (1980-2017), which positively affected reproduction and fledgling survival, had negligible consequences for population growth. Instead, population growth rates were largely reproduction-driven, in part through positive responses to rapidly advancing spring phenology. The virtual lack of body condition-mediated effects indicates that herbivore population dynamics may be more resilient to changing body condition than previously expected, with implications for their persistence under environmental change.

On Sea Turtle-associated Craspedostauros (Bacillariophyta), with Description of Three Novel Species.

The current study focuses on four species from the primarily marine diatom genus Craspedostauros that were observed growing attached to numerous sea turtles and sea turtle-associated barnacles from Croatia and South Africa. Three of the examined taxa, C. danayanus sp. nov., C. legouvelloanus sp. nov., and C. macewanii sp. nov., are described based on morphological and, whenever possible, molecular characteristics. The new taxa exhibit characters not previously observed in other members of the genus, such as the presence of more than two rows of cribrate areolae on the girdle bands, shallow perforated septa, and a complete reduction of the stauros. The fourth species, C. alatus, itself recently described from museum sea turtle specimens, is reported for the first time from loggerhead sea turtles rescued in Europe. A 3-gene phylogenetic analysis including DNA sequence data for three sea turtle-associated Craspedostauros species and other marine and epizoic diatom taxa indicated that Craspedostauros is monophyletic and sister to Achnanthes. This study, being based on a large number of samples and animal specimens analyzed and using different preservation and processing methods, provides new insights into the ecology and biogeography of the genus and sheds light on the level of intimacy and permanency in the host-epibiont interaction within the epizoic Craspedostauros species.

Design of RGDS Peptide-Immobilized Self-Assembling ß-Strand Peptide from Barnacle Protein.

We designed three types of RGD-containing barnacle adhesive proteins using self-assembling peptides. In the present study, three types of RGD-containing peptides were synthesized by solid-phase peptide synthesis, and the secondary structures of these peptides were analyzed by CD and FT-IR spectroscopy. The mechanical properties of peptide hydrogels were characterized by a rheometer. We discuss the correlation between the peptide conformation, and cell attachment and cell spreading activity from the viewpoint of developing effective tissue engineering scaffolds. We created a peptide-coated cell culture substrate by coating peptides on a polystyrene plate. They significantly facilitated cell adhesion and spreading compared to a non-coated substrate. When the RGDS sequence was modified at N- or C-terminal of R-Y, it was found that the self-assembling ability was dependent on the strongly affects hydrogel formation and cell adhesion caused by its secondary structure.

Comparative transcriptomic analysis of deep- and shallow-water barnacle species (Cirripedia, Poecilasmatidae) provides insights into deep-sea adaptation of sessile crustaceans.

BACKGROUND: Barnacles are specialized marine organisms that differ from other crustaceans in possession of a calcareous shell, which is attached to submerged surfaces. Barnacles have a wide distribution, mostly in the intertidal zone and shallow waters, but a few species inhabit the deep-sea floor. It is of interest to investigate how such sessile crustaceans became adapted to extreme deep-sea environments. We sequenced the transcriptomes of a deep-sea barnacle, Glyptelasma gigas collected at a depth of 731 m from the northern area of the Zhongjiannan Basin, and a shallow-water coordinal relative, Octolasmis warwicki. The purpose of this study was to provide genetic resources for investigating adaptation mechanisms of deep-sea barnacles. RESULTS: Totals of 62,470 and 51,585 unigenes were assembled for G. gigas and O. warwicki, respectively, and functional annotation of these unigenes was made using public databases. Comparison of the protein-coding genes between the deep- and shallow-water barnacles, and with those of four other shallow-water crustaceans, revealed 26 gene families that had experienced significant expansion in G. gigas. Functional annotation showed that these expanded genes were predominately related to DNA repair, signal transduction and carbohydrate metabolism. Base substitution analysis on the 11,611 single-copy orthologs between G. gigas and O. warwicki indicated that 25 of them were distinctly positive selected in the deep-sea barnacle, including genes related to transcription, DNA repair, ligand binding, ion channels and energy metabolism, potentially indicating their importance for survival of G. gigas in the deep-sea environment. CONCLUSIONS: The barnacle G. gigas has adopted strategies of expansion of specific gene families and of positive selection of key genes to counteract the negative effects of high hydrostatic pressure, hypoxia, low temperature and food limitation on the deep-sea floor. These expanded gene families and genes under positive selection would tend to enhance the capacities of G. gigas for signal transduction, genetic information processing and energy metabolism, and facilitate networks for perceiving and responding physiologically to the environmental conditions in deep-sea habitats. In short, our results provide genomic evidence relating to deep-sea adaptation of G. gigas, which provide a basis for further biological studies of sessile crustaceans in the deep sea.

Sponge symbiosis is facilitated by adaptive evolution of larval sensory and attachment structures in barnacles.

Symbiotic relations and range of host usage are prominent in coral reefs and crucial to the stability of such systems. In order to explain how symbiotic relations are established and evolve, we used sponge-associated barnacles to ask three questions. (1) Does larval settlement on sponge hosts require novel adaptations facilitating symbiosis? (2) How do larvae settle and start life on their hosts? (3) How has this remarkable symbiotic lifestyle involving many barnacle species evolved? We found that the larvae (cyprids) of sponge-associated barnacles show a remarkably high level of interspecific variation compared with other barnacles. We document that variation in larval attachment devices are specifically related to properties of the surface on which they attach and metamorphose. Mapping of the larval and sponge surface features onto a molecular-based phylogeny showed that sponge symbiosis evolved separately at least three times within barnacles, with the same adaptive features being found in all larvae irrespective of phylogenetic relatedness. Furthermore, the metamorphosis of two species proceeded very differently, with one species remaining superficially on the host and developing a set of white calcareous structures, the other embedding itself into the live host tissue almost immediately after settlement. We argue that such a high degree of evolutionary flexibility of barnacle larvae played an important role in the successful evolution of complex symbiotic relationships in both coral reefs and other marine systems.