Barnacle Larvae Research Articles

Antifouling activities of proteinase K and α-amylase enzymes: Laboratory bioassays and in silico analysis

The application of enzymes as antifoulants is one of the environment-friendly strategies in biofouling management. In this study, antifouling activities of commercially available proteinase K and α-amylase enzymes were evaluated using barnacle larva and biofilm-forming bacteria as test organisms. The enzymes were also tested against barnacle cement protein through in silico analysis. The results showed that both enzymes inhibited the attachment of bacteria and settlement of barnacle larvae on the test surface. The lowest minimum inhibitory concentration of 0.312 mg ml−1 was exhibited by proteinase K against biofilm-forming bacteria. The calculated LC50 values for proteinase K and α-amylase against the barnacle nauplii were 91.8 and 230.96 mg ml−1 respectively. While α-amylase showed higher antibiofilm activity, proteinase K exhibited higher anti-larval settlement activity. Similarly, in silico analysis of the enzymes revealed promising anti-settlement activity, as the enzymes showed good binding scores with barnacle cement protein. Overall, the results suggested that the enzymes proteinase K and α-amylase could be used in antifouling coatings to reduce the settlement of biofouling on artificial materials in the marine environment.

Reducing Gut Dissolution of Zinc Oxide Nanoparticles by Secondary Microplastics with Consequent Impacts on Barnacle Larvae.

The environmental impact of sunscreen is a growing concern, yet the combined effects of its components on marine animals are poorly understood. In this study, we investigated the combined effects of sunscreen-extracted zinc oxide nanoparticles (nZnO) and microplastics (MPs) on the development of barnacle larvae, focusing on the different roles played by primary microplastics (PMPs) and secondary microplastics (SMPs) generated through the phototransformation of PMPs. Our findings revealed that a lower concentration of nZnO (50 μg/L) enhanced molting and eye development in barnacle larvae, while a higher concentration (500 μg/L) inhibited larval growth. Co-exposure to PMPs had no significant effect on larval development, whereas SMPs mitigated the impact of nZnO by restricting the in vivo transformation to ionic Zn. Accumulated SMPs reduced gut dissolution of nZnO by up to 40%, lowering gut acidity by 85% and buffering the in vivo dissolution of nZnO. We further identified a rough-surfaced Si-5 fragment in SMPs that damaged larval guts, resulting in decreased acidity. Another Si-32 resisted phototransformation and had no discernible effects. Our study presented compelling evidence of the impacts of SMPs on the bioeffect of nZnO, highlighting the complex interactions between sunscreen components and their combined effects on marine organisms.

Inhibition of biofilm formation and settlement of barnacle larvae by a biosurfactant produced from a marine biofilm-forming Exiguobacterium sp. R58

Natural product based antifouling strategies are promising alternatives for achieving biofouling control with minimal environmental burden. Although microbial biosurfactants have shown antimicrobial and antibiofilm activities, their efficacy on settlement of barnacles, among the most problematic of biofouling organisms in the marine environment, has not be evaluated. Biofilm-forming bacteria isolated from the seawater cooling system of a power plant were screened for biosurfactant production and one of the promising strains was identified as Exiguobacterium sp. R58. Production, characterization and anti-biofouling activity of the biosurfactant was determined. Emulsifying activity of 68–89% was achieved at 4−100 °C, 2–12% salinity and 2–12 pH. Biofilm formation by Escherichia coli, Staphylococcus aureus, Salinococcus sp., and Marinobacter sp. was strongly inhibited (80–92%) in the presence of 2000 μg/mL biosurfactant. Cypris larval settlement was impaired by the biosurfactant without causing mortality. Biosurfactant coated surfaces prevented biofilm formation and cypris settlement. Adhesion of bacteria on biosurfactant coated surfaces was 75–93% lower as compared to control. Cypris settlement was inhibited completely in Petri dishes coated with 5.6–22.7 mg/cm2 in no-choice assays and 1.3 mg/cm2 in choice assays. Antibiofouling activity of the biosurfactant was related to surface modification, thus promising for developing non-toxic antifouling materials.

The impact of habitat complexity on the structure of marine sessile communities and larvae supply

Coastal infrastructure replaces complex and heterogeneous natural habitats with flat, two-dimensional concrete walls, reducing refuges against predation, which modifies the composition and identity of the dominant species in sessile communities. This modification in the community structure can also change the reproductive propagules available in plankton, affecting the recruitment dynamics in communities from natural habitats nearby. Here, we tested the combined effects of the habitat type (simple vs. complex with holes) and predation on the diversity, larval production, and structure of sessile communities from a recreational marina. Complex substrates showed a larger biomass and a greater abundance of solitary organisms, mainly ascidians and bivalves, that benefited from refuges. Barnacles and calcified encrusting bryozoans dominated simple, flat substrates. The difference in dominance affected the pool of larvae produced by the communities. After eight months, communities growing on flat substrates produced more barnacle larvae than those from complex substrates, where larvae of ascidians were more abundant. However, this difference disappeared after 18 months of community development. The difference in the pool of larvae between simple and complex substrates did not affect the structure of the community on flat substrates nearby, which was determined by the predation regime. In the studied region, communities in artificial environments are under intense predation control, suppressing eventual recruitment differences in communities developing in flat substrates. Large interventions that modify habitat topography, creating refuges in the subtidal zone, can change the dynamic of the sessile communities in artificial habitats and, consequently, the larval supply in the vicinities. However, differences in larval supply will only translate in distinct sessile communities when the scale of intervention encompasses large areas, and other processes do not buffer the differences in recruitment.

Host specificity and adaptive evolution in settlement behaviour of coral-associated barnacle larvae (Cirripedia: Pyrgomatidae)

Coral-associated organisms often exhibit a continuum of host specificities. We do not know whether the variation in host specificity is related to the settlement organs or preferential settlement behaviours of the larvae. We examined the morphology of attachment discs, the settlement and metamorphosis of coral barnacles—Pyrgoma cancellatum (lives in a single coral species), Nobia grandis (two families of corals), and Armatobalanus allium (six families of corals). Our results revealed that the attachment organ of all three species are spear-shaped with sparse villi, indicating that the morphology of the attachment organs does not vary among species with different host specificities. Larvae of P. cancellatum and N. grandis only settle on their specific hosts, suggesting that chemical cues are involved in the settlement. Cyprids of N. grandis display close searching behaviour before settlement. Cyprids of P. cancellatum settle immediately on their specific host corals, without any exploratory behaviour. The host specificity and exploratory behaviours of coral barnacle cyprids are results of adaptive evolution. We argue that there is a trade-off between exploration and energy conservation for metamorphosis processes. Coral barnacle metamorphosis is longer when compared to free-living species, likely because it involves the development of a tube-shaped base on the coral surface.

Diel variability in the vertical distribution of coastal barnacle larvae in the Bay of Cartagena, Chile

BackgroundVertical distribution of zooplankton is an important biological factor that can modulate zooplankton transport, dispersal, and survival in the ocean. Seawater temperature and the associated formation of a thermocline can affect the spatial distribution of organisms in the water column and possibly modulate larval vertical distribution in coastal waters. In this study, we examined larval vertical distribution over small spatial scales where environmental conditions could exhibit strong and fairly predictable variability.MethodsDiel variation in vertical distributions of barnacle larvae across the thermocline was characterized within the Bay of Cartagena of central Chile. Two intensive 21- and 24-h surveys were conducted in the northern (ECIM) and southern (CTGN) extremes of the bay in summer 2016 and spring 2017. In each survey, vertical tows were conducted at both sites every 3 h above and below the thermocline.ResultsDuring summer 2016, larval vertical distribution differed between taxa (balanids and verrucids), barnacle stages (nauplii, cyprids), and sites within the bay. Of all taxa analyzed Balanid nauplii at ECIM were more abundant in the bottom layer during the day and at the surface during the night, suggesting they can control their vertical distribution and follow a diel pattern.Acording to the results all barnacle larvae were found in diferent positions in the water column between day and night during periods of high stratification in this survey.. In spring 2017 all barnacle larvae were confined to the upper layer, despite the strength of stratification, potentially as a result of the intrusion of hypoxic water (< 2 mL L−1) at the bottom, which reached up to around 5 m deep.ConclusionsWe conclude that diel vertical distribution is not uniform across different barnacle larvae taxa and stages within the Bay of Cartagena and is not affected by stratification. Of all the taxa analyzed, only balanid nauplii showed a pattern of diel vertical migration across the thermocline under certain hydrographic conditions.

Front Cover: Anti‐Barnacle Activities of Isothiocyanates Derived from β‐Citronellol and Their Structure–Activity Relationships (Chem. Biodiversity 2/2023)

Front Cover. Antifouling agents with low toxicity are in high demand for sustaining marine industries and the environment. Fifteen isothiocyanates derived from β-citronellol were synthesized, and their antifouling activities and toxicities were evaluated against the cypris larvae of barnacles. The isothiocyanates with an amide or isocyano group exhibited great potential as effective antifouling agents. They are therefore expected to be useful for further research on the development of environmentally friendly antifouling agents, as reported by Kitano et al. in their research article at 10.1002/cbdv.202200953.

PDMS networks meet barnacles: a complex and often toxic relationship

The biological impact of chemical formulations used in various coating applications is essential in guiding the development of new materials that directly contact living organisms. To illustrate this point, an investigation addressing the impact of chemical compositions of polydimethylsiloxane networks on a common platform for foul-release biofouling management coatings was conducted. The acute toxicity of network components to barnacle larvae, the impacts of aqueous extracts of crosslinker, silicones and organometallic catalyst on trypsin enzymatic activity, and the impact of assembled networks on barnacle adhesion was evaluated. The outcomes of the study indicate that all components used in the formulation of the silicone network alter trypsin enzymatic activity and have a range of acute toxicity to barnacle larvae. Also, the adhesion strength of barnacles attached to PDMS networks correlates to the network formulation protocol. This information can be used to assess action mechanisms and risk–benefit analysis of PDMS networks.

Understanding the biological impact of organic pollutants absorbed by nanoplastics

Many organisms are consuming food contaminated with micro- and nanoparticles of plastics, some of which absorb persistent organic pollutants (POPs) from the environment and acting as carrier vectors for increasing the bioavailability in living organisms. We recently reported that polymethylmethacrylate (PMMA) nanoparticles at low concentrations are not toxic to animal models tested. In this study, the toxicity of diphenylamine (DPA) incorporated PMMA nanoparticles are assessed using barnacle larvae as a model organism. The absorption capacity of DPA from water for commercially available virgin PMMA microparticles is relatively low (0.14 wt%) during a 48 h period, which did not induce exposure toxicity to barnacle nauplii. Thus, PMMA nanoparticles encapsulated with high concentrations of DPA (DPA-enc-PMMA) were prepared through a reported precipitation method to achieve 40% loading of DPA inside the particles. Toxicity of DPA-enc-PMMA nanoparticles were tested using freshly spawned acorn barnacle nauplii. The observed mortality of nauplii from DPA-enc-PMMA exposure was compared to the values obtained from pure DPA exposure in water. The mortality among the exposed acorn barnacle nauplii did not exceed 50% even at a high concentration of DPA inside the PMMA nanoparticles. The results suggest that the slow release of pollutants from polymer nanoparticles may not induce significant toxicity to the organism living in a dynamic environment. The impact of long-term exposure of DPA absorbed plastic nanoparticles need to be investigated in the future.

Experiment on Phototaxis Behavior of Barnacle Larvae

Experiment on Phototaxis Behavior of Barnacle Larvae

Faint chemical traces of conspecifics delay settlement of barnacle larvae

Larval settlement drives population and community dynamics and evolution of marine invertebrates. While it is well known that larvae sense a variety of biological and environmental factors, there is limited information on how they react to specific factors so as to locate settlement sites. Proteinaceous Waterborne Settlement Pheromone (WSP) was purified from adult barnacles, Amphibalanus amphitrite, in a previous study, but it is unclear how WSP guides barnacle cyprids to the source. Here, a series of settlement assays were conducted to investigate the concentration dependence of recombinant WSP. We report that low concentrations of barnacle WSP decrease the probability of settlement of conspecific cyprids, whereas high concentrations have the opposite effect. We suggest that this is because weak WSP informs cyprids that suitable settlement sites with adult barnacles are distant, inducing them to extend the larval phase. The present study clearly shows that larvae use WSP concentration, rather than presence/absence information. In barnacles, several other conspecific chemical/visual cues also attract larvae. This study opens the door to future research to determine how these cues influence larval behavior in nature.

Synthesis and Properties of Self-Polishing Antifouling Coatings Based on BIT-Acrylate Resins

Painting antifouling coatings is one of the most important methods to prevent marine biofouling. Acrylic resin is widely used in marine antifouling because of its excellent stickiness, water resistance, and film-forming capabilities. At present, the widely used acrylate antifouling coatings require a high concentration of cuprous oxide as antifoulant. The release and accumulation of copper ions are the main factors affecting the marine environment. In this study, BIT–allyl methacrylate (BM) and zinc acrylate (ZM) were selected as functional monomers copolymerized with methyl methacrylate (MMA) and butyl acrylate (BA) to prepare a series of BIT acrylate antifouling resins. The inhibitory effects of all resins against marine bacteria (S. aureus, V. coralliilyticus, and V. parahaemolyticus), marine algae (Chlorella, I. galbana, and C. curvisetus), and barnacle larvae were studied. Moreover, marine field tests on the BIT modified resin in coastal waters were conducted. The results demonstrate that the grafted BIT–zinc acrylate resin not only exhibits excellent antifouling properties but also a significant self-polishing performance, providing a novel strategy to design a long-term antifouling resin with stable antifoulant release.

Degradable Vinyl Polymers for Combating Marine Biofouling.

Marine organisms such as barnacle larvae and spores of algae adhere to underwater surfaces leading to marine biofouling. This phenomenon has numerous adverse impacts on marine industries and maritime activities. Due to the diversity of fouling organisms and the complexity of the marine environment, it is a huge challenge to combat marine biofouling, which limits the development and utilization of marine resources. Since the International Marine Organization banned the use of tributyltin self-polishing copolymer (SPC) coatings in 2008, the development of an environmentally friendly and efficient anti-biofouling polymer has been the most important task in this field. Tin-free SPC is a well-established and widely used polymer binder for anti-biofouling coating today. Being a nondegradable vinyl polymer, SPC exhibits poor anti-biofouling performance in static conditions. Even more, such nondegradable polymers were considered to be a source of microplastics by the International Union for the Conservation of Nature in 2019. Recently, numerous degradable polymers, which can form dynamic surface through main chain scission, have been developed for preventing marine biofouling in static conditions. Nevertheless, the regulation of their degradation and mechanical properties is limited, and they are also difficult to functionalize. A new polymer combining the advantages of vinyl polymers and degradable polymers is needed. However, such a combination is a challenge since the former are synthesized via free radical polymerization whereas the latter are synthesized via ring-opening polymerization.In this Account, we review our recent progress toward degradable vinyl polymers for marine anti-biofouling in terms of polymerization methods and structures and properties of polymers. First, we introduce the strategies for preparing degradable vinyl polymers with an emphasis on hybrid copolymerization. Then, we present the synthesis and performance of degradable and hydrolyzable polyacrylates, degradable polyurethanes with hydrolyzable side groups, and surface-fragmenting hyperbranched polymers. Polymers with degradable main chains and hydrolyzable side groups combine the advantages of SPC and degradable polymers, so they are degradable and functional. They are becoming new-generation polymers with great potential for preparing high-efficiency, long-lasting, environmentally friendly and broad-spectrum coatings to inhibit marine biofouling. They can also find applications in wastewater treatment, biomedical materials, and other fields.

The anti-settlement activity of extracts of marine bacteria associated with soft corals against barnacle larvae

The anti-settlement activity of extracts of marine bacteria associated with soft corals against barnacle larvae

Antifouling coating based on biopolymers (PCL/ PLA) and bioactive extract from the sea cucumber Stichopus herrmanni

An important challenge to decrease the toxic effects of the common biocides in marine environments and to achieve suitable ecofriendly natural antifouling coatings is to find effective natural antifoulants and efficient biodegradable coatings. In this study, antifouling activities of nine bioactive extracts (non-polar to polar) from different organs of the sea cucumber Stichopus herrmanni were tested against five bacterial strains, barnacle and brine shrimp larvae. The ethyl acetate extract of the body wall showed the highest in-vitro antifouling activity including high antibacterial and anti-barnacle activities and low toxicity against the brine shrimp as non-target organism. Based on these results, 10 phr of the ethyl acetate extract from S.herrmanni was added to different coatings consisting of polycaprolactone (PCL)/polylactic acid (PLA) blends containing various compositions of PLA (0, 10, and 20 wt.%). Polyvinyl chloride panels were coated with the prepared antifouling coatings and immersed in seawater for three months. Panel coated with PCL 80% /PLA 20% containing 10 phr of the antifoulant (panel-5), showed the highest resistance against fouling settlement with fouling coverage of 41.66% (P < 0.05). In addition, the lowest fouling weight was measured in panel-5 as well (81.00 ± 9.85 g) (P < 0.05). These findings indicate the antibacterial and antifouling potential of semi-polar bioactive extracts from the S. herrmanni body wall as natural antifoulants, as well as the enhanced antifouling performance of PCL/the natural antifoulant coatings by adding PLA.Graphical

Quantification of Cypris Larvae of Barnacles in Sailing, Anchoring, and Berthing Areas of Western Japan as well as in Tokyo Port in Summer

Quantification of Cypris Larvae of Barnacles in Sailing, Anchoring, and Berthing Areas of Western Japan as well as in Tokyo Port in Summer

Independent and adaptive evolution of phenotypic novelties driven by coral symbiosis in barnacle larvae.

The invasion of novel habitats is recognized as a major promotor of adaptive trait evolution in animals. We tested whether similar ecological niches entail independent and adaptive evolution of key phenotypic structures related to larval host invasion in distantly related taxa. We use disparately related clades of coral barnacles as our model system (Acrothoracica: Berndtia and Thoracicalcarea: Pyrgomatidae). We analyze the larval antennular phenotypes and functional morphologies facilitating host invasion. Extensive video recordings show that coral host invasion is carried out exclusively by cypris larvae with spear-shaped antennules. These first exercise a series of complex probing behaviors followed by repeated antennular penetration of the soft host tissues, which subsequently facilitates permanent invasion. Phylogenetic mapping of larval form and function related to niche invasion in 99 species of barnacles (Thecostraca) compellingly shows that the spear phenotype is uniquely associated with corals and penetrative behaviors. These features evolved independently in the two coral barnacle clades and from ancestors with fundamentally different antennular phenotypes. The larval host invasion system in coral barnacles likely evolved adaptively across millions of years for overcoming challenges associated with invading and entering demanding coral hosts.

High Larval Concentrations and Onshore Transport of Barnacle Cyprids Associated With Thermal Stratification

To better understand the hydrodynamic and hydrographic conditions experienced by larvae in the nearshore (within 1 km of shore), and the role that larval behavior plays in mediating shoreward transport to adult benthic habitats, we examined the vertical distribution and concentration of barnacle cyprids in a shallow, nearshore region in southern California, United States. We collected high-resolution physical measurements of currents and temperature at 3 stations (8, 5, and 4 m depths), and high-frequency measurements of barnacle larvae at a 4 m deep station ∼300 m from shore. Larvae were sampled from distinct 1 m depth intervals between the surface and the bottom (0–1 m, 1–2 m, 2–3 m, 3 m-bottom), each hour for overnight periods that ranged between 13 to 24 h in five cruises during the summers of 2017 and 2018. Barnacle cyprids of Chthamalus fissus predominated in all samples. Thermal stratification decreased closer to shore, but when the nearshore-most station remained stratified (Δ°C m–1 ≥ 0.1), C. fissus cyprid concentrations were high to extremely abundant (exceeding 200 and 4,000 individuals m–3, respectively). There were significant positive correlations between thermal stratification and the log-transformed C. fissus concentration at cruise-to-cruise scales, and between stratification and vertical variability in the high-frequency cross-shore currents at 2-day scales. Additionally, estimated larval transport was relatively high and shoreward when nearshore thermal stratification was greatest. Significant, albeit small, diel differences in cyprid distributions were also observed, with the proportion of cyprids increasing near the surface at night, and concentrations greater during the day than at night. Collectively, these results suggest that thermal stratification increases larval supply to the nearshore, and may enhance onshore larval transport to augment chances of successful settlement and recruitment to the intertidal adult habitat.

In-situ repair of marine coatings by a Fe3O4 nanoparticle-modified epoxy resin under seawater

The in-situ repair of organic anticorrosive coatings under seawater is an ideal method to extend their service life once the coatings suffer from unexpected damage. However, the attachment and solidification of the repairing organic binder on the defects under seawater are two big challenging issues, which may lead to serious delamination and bad impermeability in the repaired coatings. Inspired by the attachment of barnacle larvae, lipophilic magnetic Fe3O4 nanoparticles were prepared via a co-precipitation method and then modified with oleic acid in this study. They were mixed with epoxy binder to squeeze away the seawater between the epoxy binder and substrate metals with the aid of magnetic force. The experimental results proved that the Fe3O4 nanoparticles had target-driving and competitive-wetting functions, which could effectively prevent the formation of penetrating micro-channels and enhance the adhesion strength of the organic binder to substrate metals. The new method may provide a repeatable and straightforward route for the in-situ repair of organic anti-corrosion coating under seawater.

Nonconsumptive Predator Effects on Prey Demography: Recent Advances Using Intertidal Invertebrates

Predators influence prey demography through consumption, but the mere presence of predators may trigger behavioural changes in prey that, if persistent or intense, may also influence prey demography. A tractable system to study such nonconsumptive effects (NCEs) of predators involves intertidal invertebrates. This mini review summarises recent research using barnacles and mussels as prey and dogwhelks as predators. The field manipulation of dogwhelk density revealed that pelagic barnacle larvae avoid benthic settlement near dogwhelks, which limits barnacle recruitment, a relevant outcome because recruitment is the only source of population replenishment for barnacles, as they are sessile. This avoidance behaviour is likely triggered by waterborne dogwhelk cues and may have evolved to limit future predation risk. Increasing densities of barnacle recruits and adults can prevent such NCEs from occurring, seemingly because benthic barnacles attract conspecific larvae through chemical cues. Barnacle recruit density increased with the abundance of coastal phytoplankton (food for barnacle larvae and recruits), so barnacle food supply seems to indirectly limit dogwhelk NCEs. By inhibiting barnacle feeding, dogwhelk cues also limited barnacle growth and reproductive output. Wave action weakens dogwhelk NCEs likely through hydrodynamic influences. Dogwhelk cues also limit mussel recruitment, as mussel larvae also exhibit predator avoidance behaviour. The NCEs on recruitment are weaker for mussels than for barnacles, possibly because mussel larvae can detach themselves after initial settlement, an ability that barnacle larvae lack. Overall, these field experiments provide evidence of predator NCEs on prey demography for coastal marine systems.