In situ ATR–FTIR characterization of primary cement interfaces of the barnacle Balanus amphitrite

A method is presented for characterizing primary cement interfaces of barnacles using in situ attenuated total reflection–Fourier transform infrared spectroscopy. Primary cement of the barnacle, Balanus amphitrite (Amphibalanus amphitrite), was characterized without any disruption to the original cement interface, after settling and growing barnacles directly on double sided polished germanium wafers. High-quality IR spectra were acquired of live barnacle cement interfaces, providing a spectroscopic fingerprint of cured primary cement in vivo with the barnacle adhered to the substratum. Additional spectra were also acquired of intact cement interfaces for which the upper portion of the barnacle had been removed leaving only the base plate and cement layer attached to the substratum. This allowed further characterization of primary cement interfaces that were dried or placed in D₂O. The resulting spectra were consistent with the cement being proteinaceous, and allowed analysis of the protein secondary structure and water content in the cement layer. The estimated secondary structure composition was primarily β-sheet, with additional α-helix, turn and unordered components. The cement of live barnacles, freshly removed from seawater, was estimated to have a water content of 20–50% by weight. These results provide new insights into the chemical properties of the undisturbed barnacle adhesive interface.     

Direct and indirect effects of Littorina littorea (L.) on barnacles growing on mussel beds in the Wadden Sea

On the extensive sedimentary tidal flats of the Wadden Sea, beds of the blue mussel Mytilus edulis represent the only major hard substratum and attachment surface for sessile organisms. On this substratum, the barnacle Semibalanus balanoides is the most frequent epibiont. In summer 1998, it occurred on over 90% of the large mussels (>45 mm shell length) and the dry weight of barnacles reached 65% of mussel dry weight. However, the extent of barnacle overgrowth is not constant and differs widely between years. Periwinkles (Littorina littorea) may reach densities >2000 m^–2 on intertidal mussel beds. Field experiments were conducted to test the effect of periwinkle grazing on barnacle densities. An experimental reduction of grazing and bulldozing pressure by periwinkles resulted in increased recruitment of barnacles, while barnacle numbers decreased with increasing snail density. The highest numbers of barnacles survived in the absence of L. littorea. However, a lack of periwinkle grazing activity also facilitated settlement of ephemeral algae which settled later in the year. Field experiments showed that the growth rate of barnacles decreased in the presence of these ephemeral algae. Thus, L. littorea may reduce initial barnacle settlement, but later may indirectly increase barnacle growth rate by reducing ephemeral algae. It is suggested that periwinkle density may be a key factor in the population dynamics of S. balanoides on intertidal mussel beds in the Wadden Sea.     

Interfacial morphology and nanomechanics of cement of the barnacle,Amphibalanus reticulatus on metallic and non-metallic substrata

The barnacle exhibits a high degree of control over its attachment onto different types of solid surface. The structure and composition of barnacle cement have been reported previously, but mostly for barnacles growing on low surface energy materials. This article focuses on the strategies used by barnacles when they attach to engineering materials such as polymethylmethacrylate (PMMA), titanium (Ti) and stainless steel 316L (SS316L). Adhesion to these substrata is compared in terms of morphological structure, thickness and functional groups of the primary cement, the molting cycle and the nanomechanical properties of the cement. Structural characterization studies using scanning electron microscopy (SEM) and transmission electron microscopy (TEM) in conjunction with nanomechanical characterization and infrared spectroscopy (FTIR) are used to understand the differences in the adhesion of primary barnacle cement to the different substrata. The results provide new insights into understanding the mechanisms at work across the barnacle–substratum interface.     

How whale and dolphin barnacles attach to their hosts and the paradox of remarkably versatile attachment structures in cypris larvae

How larvae of whale and dolphin epibionts settle on their fast-swimming and migrating hosts is a puzzling question in zoology. We successfully reared the larvae of the whale and dolphin barnacle Xenobalanus globicipitis to the cyprid stage. We studied the larval developmental ecology and antennular morphology in an attempt to assess whether an epibiotic lifestyle on this extreme substratum entails any unique larval specializations. Morphological parameters were compared with five other barnacle species that also inhabit extreme substrata. We found no larval specializations to a lifestyle associated with marine mammals. The external morphology of the antennules in Xenobalanus cyprids is morphologically similar to species from strikingly different substrata. We found variation only in the structures that are in physical contact with the substratum, i.e., the third segments carrying the villi-covered attachment disc. The third segments of the Xenobalanus cyprid antennules are not spear-shaped as in the stony coral barnacles, which are here used to penetrate the live tissue of their hosts. The presence of a cyprid cement gland implies that Xenobalanus uses cement protein when attaching to its cetacean host. Naupliar instars developed outside of the mantle cavity, indicating dispersal is planktonic. Our results militate against the idea that the cyprids settle during ocean migrations of their hosts. We suggest cyprids settle during coastal aggregations of the cetacean hosts. We conclude that the ecological success of barnacles has ultimately depended on a larva that with little structural alteration possesses the ability to settle on an amazingly wide array of substrata, including cetaceans.

     

Distribution and abundance of the acmaeid limpet,Patelloida latistrigata,and its interaction with barnacles

Summary At Cape Banks, New South Wales, adults of the small intertidal limpet, Patelloida latistrigata occur exclusively in the barnacle zone, and are primarily associated with the barnacle, Tesseropora rosea. Limpet density increases with barnacle density. Juvenile limpets can be found throughout the barnacle zone, and on patches of bare rock that may be temporarily available at lower levels on the shore. The failure of juveniles to survive and grow in places other than among Tesseropora is due to a combination of factors. These include desication at high levels on the shore, smothering by rapidly growing algae low on the shore, and the grazing activities of the larger limpet, Cellana tramoserica. These latter two factors also reduce the survival of experimentally transplanted adult Patelloida: algae by covering the substratum and smothering the limpets, and Cellana by outcompeting them for food. The density of Cellana is greater on patches of bare rock than among barnacles, and these large limpets may be unable to move and feed effectively over the irregular surface created by Tesseropora. Patelloida, however, is small enough to feed over and among these barnacles, and hence has a refuge from competition with Cellana. Barnacles may also provide shelter from the effects of desiccation and strong wave action, and thus increase the survival of juvenile Patelloida. By being associated with barnacles, however, Patelloida becomes vulnerable to intermittent predation by the whelk, Morula marginalba. This effect may be serious enough to eliminate small local populations of limpets, either by direct predation or by removing the refuge-providing barnacles. The association with barnacles may also limit the maximum size to which Patelloida can grow. Patelloida is not always found with Tesseropora, and adults in different localities can be found in association with other sessile organisms. It may be argued that small species of limpets require a spatial refuge from physical and/or biological pressures. To examine this hypothesis, the relationship between Patelloida and Tesseropora is compared to other published accounts of limpets with specialised modes of life.     

Desiccation,predation, and mussel-barnacle interactions in the northern Gulf of California

Summary Field experiments were conducted in order to determine the potential for desiccation and predation to mediate the effect of mussels (Brachidontes semilaevis) on barnacles (Chthamalus anisopoma) in the highly seasonal northern Gulf of California. We did this by removing both mussels and a common mussel predator (Morula ferruginosa: Gastropoda) and by spraying selected sites with sea water during summertime spring low tides. We also determined the effect of crowding on resistance to desiccation in barnacles, and the effect of barnacles on colonization by mussels. The mussel-barnacle community was not affected by keeping experimental quadrats damp during daytime low tides throughout the summer. Exposure to summertime low tides, however, did affect the survivorship of isolated, but not crowded, barnacles; and barnacle clumps enhanced the recruitment of mussels. Hence crowding in barnacles had a positive effect on both barnacle survivorship and mussel recruitment. Morula had a negative effect on mussel density, and mussels had a negative effect on barnacle density. The effect of Morula on barnacle density was positive, presumably due to its selective removal of mussels. These results suggest an indirect mutualism between barnacles and the gastropod predator, because barnacles attract settlement or enhance the survival of mussels, and the predator reduces the competitive effect of mussels on barnacles.     

Incorporation of silicone oil into elastomers enhances barnacle detachment by active surface strain

Silicone-oil additives are often used in fouling-release silicone coatings to reduce the adhesion strength of barnacles and other biofouling organisms. This study follows on from a recently reported active approach to detach barnacles, which was based on the surface strain of elastomeric materials, by investigating a new, dual-action approach to barnacle detachment using Ecoflex®-based elastomers incorporated with poly(dimethylsiloxane)-based oil additives. The experimental results support the hypothesis that silicone-oil additives reduce the amount of substratum strain required to detach barnacles. The study also de-coupled the two effects of silicone oils (ie surface-activity and alteration of the bulk modulus) and examined their contributions in reducing barnacle adhesion strength. Further, a finite element model based on fracture mechanics was employed to qualitatively understand the effects of surface strain and substratum modulus on barnacle adhesion strength. The study demonstrates that dynamic substratum deformation of elastomers with silicone-oil additives provides a bifunctional approach towards management of biofouling by barnacles.     

An in situ study of the nanomechanical properties of barnacle (Balanus amphitrite) cyprid cement using atomic force microscopy (AFM)

Abstract

Cyprids are the final planktonic stage in the larval dispersal of barnacles and are responsible for surface exploration and attachment to appropriate substrata. The nanomechanical properties of barnacle (Balanus amphitrite) cyprid permanent cement were studied in situ using atomic force microscopy (AFM). Force curves were recorded from the cement disc continually over the course of its curing and these were subsequently analysed using custom software. Results showed a narrowing of the pull-off force distribution with time, as well as a reduction in molecular stretch length over time. In addition, there was a strong correlation between maximum pull-off force and molecular stretch length for the cement, suggesting ‘curing’ of the adhesive; some force curves also contained a ‘fingerprint’ of modular protein unfolding. This study provides the first direct experimental evidence in support of a putative ‘tanning’ mechanism in barnacle cyprid cement.     

The symbiotic relationship between the hydrocoral Millepora dichotoma and the barnacle Savignium milleporum

Translocation of radioactive ¹⁴C and ³²P between the pyrgomatine barnacle Savignium milleporum and the hydrocoral Millepora dichotoma in the Red Sea was investigated in order to discover any mutual nutritional benefits. Translocation of photosynthetic products from endosymbiotic zooxanthellae to the hydrocoral was demonstrated. There was no evidence that carbon was further translocated to the barnacle. However, hydrocorals bearing barnacles accumulated significantly more ¹⁴C and ³²P than those with no barnacles. The possibility that the hydrocorals recycle substances excreted by the barnacles is discussed in the context of the oligotrophic environment of the Red Sea.     

Early life-history processes and their implications for the invasion of the barnacle Balanus glandula

As invasions become increasingly prevalent, it is important to understand how spread may be moderated by environmental conditions. This study considered the effect of location and changing substratum temperature on the early life-history processes of the alien barnacle Balanus glandula and its native comparator, Chthamalus dentatus, along the South African coast. Using settlement plates of different colours, temperature was manipulated to assess settlement, mortality, recruitment and growth of the two species. These variables were tracked over 10 weeks using repeat photography. Unexpectedly, there was no evidence of an effect of temperature on early life-history processes of either species. Settlement by the two barnacles was spatially segregated, with B. glandula occurring only on the West Coast where it is the dominant intertidal barnacle, while C. dentatus settled only on the South Coast, which has only recently been invaded by the alien. Despite this, it was notable that the relative settlement of B. glandula on rock was higher than that of C. dentatus. However, the lack of mortality of the native resulted in comparable levels of recruitment among the species. Nonetheless, the propensity of B. glandula to settle sporadically, coupled with fast growth, suggests that the invader may still possess the ability to become dominant along the newly invaded South Coast. This study highlights that measurements of early life-history parameters may not adequately predict the future range and impacts of alien species unless interpreted within the broader context of the nature of the recipient region and species-specific traits of the invader.     

The Impact of Desiccation on the Adhesion of Barnacles Attached to Non-stick Coatings

Fouling-release coatings prevent fouling of ships' hulls through hydrodynamic forces generated as the ship moves through the water. The effectiveness of such coatings may be evaluated by measuring the adhesion strength of settled organisms, e.g. barnacles. The influence of desiccation of the barnacle adhesive on such measurements was investigated. Shear forces required to remove barnacles of the genus Balanus increased during the course of desiccation up to the point when the barnacles suddenly self-detached. The increase was thought to be due to the rising cohesive strength of the adhesive. Growing tensile forces within the weakly cross-linked adhesive, however, are suggested to have led to self-detachment. The shear forces required to remove barnacles of the genus Elminius were generally low and did not differ significantly during the course of desiccation. The different results may be attributed to specific base morphologies. It was concluded that measuring the adhesion strength of members of the Balanidae on non-stick surfaces in air could produce flawed results due to the influence of desiccation of the barnacle adhesive. The investigations have also provided new insights into the characteristics of barnacle adhesive.     

Prolonged morphometric study of barnacles grown on soft substrata of hydrogels and elastomers

A long-term investigation of the shell shape and the basal morphology of barnacles grown on tough, double-network (DN) hydrogels and polydimethylsiloxane (PDMS) elastomer was conducted in a laboratory environment. The elastic modulus of these soft substrata varied between 0.01 and 0.47 MPa. Polystyrene (PS) (elastic modulus, 3 GPa) was used as a hard substratum control. It was found that the shell shape and the basal plate morphology of barnacles were different on the rigid PS substratum compared to the soft substrata of PDMS and DN hydrogels. Barnacles on the PS substratum had a truncated cone shape with a flat basal plate while on soft PDMS and DN gels, barnacles had a pseudo-cylindrical shape and their basal plates showed curvature. In addition, a large adhesive layer was observed under barnacles on PDMS, but not on DN gels. The effect of substratum stiffness is discussed in terms of barnacle muscle contraction, whereby the relative stiffness of the substratum compared to that of the muscle is considered as the key parameter.     

Differential growth of the barnacle Notobalanus flosculus (Archaeobalanidae) onto artificial and live substrates in the Beagle Channel, Argentina

In the Beagle Channel, southern South America (ca. 55°S 67°W), about 20% of false king crabs (Paralomis granulosa) >80 mm carapace length are fouled with the barnacle Notobalanus flosculus. To evaluate differences in growth rates of barnacles attached to artificial and live substrates, clay tiles were anchored as collectors to the bottom at two different sites in the Beagle Channel in September 1996: in Ushuaia harbour (low currents and high levels of suspended matter) and around the Bridges Islands (strong currents and low level of suspended matter). Another set of collectors was deployed at the same sites in October 1998 to collect barnacles for histological studies. Tiles were removed from each place, approximately, on a monthly basis. Carapaces of P. granulosa with the epizoic N. flosculus were sampled between November 1996 and 1997, and between March 1998 and September 1999, to study sexual maturation of barnacles. Growth of barnacles was compared between the collectors and P. granulosa carapaces following a qualitative approach. A sexual maturity scale was defined, based on the stage of development of the female reproductive apparatus of N. flosculus. Growth rate of barnacles was highest in the harbour, intermediate on P. granulosa, and lowest around the Bridges Islands. Presence of oocytes was registered only in epizoic barnacles, suggesting that at least a proportion of these individuals is able to spawn on the carapaces. The potential advantages of settling on a living substrate, namely increased availability of food particles and decreased predation risks are discussed.     

Proteomic analysis during larval development and metamorphosis of the spionid polychaete Pseudopolydora vexillosa

Background  

While the larval-juvenile transition (metamorphosis) in the spionid polychaete Pseudopolydora vexillosa involves gradual morphological changes and does not require substantial development of juvenile organs, the opposite occurs in the barnacle Balanus amphitrite. We hypothesized that the proteome changes during metamorphosis in the spionids are less drastic than that in the barnacles. To test this, proteomes of pre-competent larvae, competent larvae (ready to metamorphose), and juveniles of P. vexillosa were compared using 2-dimensional gel electrophoresis (2-DE), and they were then compared to those of the barnacle.     

Evaluation of a fully automated method to measure the critical removal stress of adult barnacles

A computer-controlled force gauge designed to measure the adhesive strength of barnacles on test substrata is described. The instrument was evaluated with adult barnacles grown in situ on Silastic T2®-coated microscope slides and epoxy replicas adhered to the same substratum with synthetic adhesive. The force per unit area required to detach the barnacles (critical removal stress) using the new automated system was comparable to that obtained with ASTM D5618 (1994 ASTM D5618. 1994. Standard test method for measurement of barnacle adhesion strength in shear American Standard for Testing and Materials. Paint-Tests for formulated products and applied coatings 06.01 [Google Scholar]) (0.19 and 0.28 MPa compared with 0.18 and 0.27 MPa for two batches of barnacles). The automated method showed a faster rate of force development compared with the manual spring force gauge used for ASTM D5618 (1994 ASTM D5618. 1994. Standard test method for measurement of barnacle adhesion strength in shear American Standard for Testing and Materials. Paint-Tests for formulated products and applied coatings 06.01 [Google Scholar]). The new instrument was as accurate and precise at determining surface area as manual delineation used with ASTM D5618 (1994 ASTM D5618. 1994. Standard test method for measurement of barnacle adhesion strength in shear American Standard for Testing and Materials. Paint-Tests for formulated products and applied coatings 06.01 [Google Scholar]). The method provided significant advantages such as higher throughput speed, the ability to test smaller barnacles (which took less time to grow) and to control the force application angle and speed. The variability in measurements was lower than previously reported, suggesting an improved ability to compare the results obtained by different researchers.     

Effects of tidal height and wave exposure on cirrus and penis morphology of the acorn barnacle Tetraclita stalactifera

Exposure to wave action and other environmental factors can alter the morphology of intertidal barnacles. We tested several hypotheses on the causes of morphological variation in the cirri and penises of the barnacle Tetraclita stalactifera at sites differing in wave exposure, at different heights in the intertidal zone, and at different levels of population density. Unlike many other acorn barnacle species, cirrus and penis characteristics did not correspond to differences in wave exposure or crowding. However, barnacles from higher tidal elevations had thicker cirri and thicker penises than those from lower elevations. Because of reduced time submerged at higher elevations, increased thickness may be a means of compensating for reduced feeding and mating opportunity by allowing for continued feeding and mating attempts during periods of greater wave action. Our observations of differences in cirrus and penis morphology suggest that phenotypic plasticity in penis and cirrus characteristics are adaptations shared by the species T. stalactifera and other acorn barnacles, but that T. stalactifera responds differently to environmental stimuli than do other species.     

Surface elastic modulus of barnacle adhesive and release characteristics from silicone surfaces

The properties of barnacle adhesive on silicone surfaces were studied by AFM indentation, imaging, and other tests and compared to the barnacle shear adhesion strength. A multilayered structure of barnacle adhesive plaque is proposed based on layered modulus regions measured by AFM indentation. The fracture of barnacles from PDMS surfaces was found to include both interfacial and cohesive failure of barnacle adhesive plaque, as determined by protein staining of the substratum after forced barnacle release from the substrate. Data for freshly released barnacles showed that there was a strong correlation between the mean Young's modulus of the outermost (softest) adhesive layer (E<0.3 MPa) and the shear strength of adhesion, but no correlation for other higher modulus regions. Linear, quadratic, and Griffith's failure criterion (based on rough estimate of crack length) regressions were used in the fit, and showed significance.     

Selective settlement of the barnacle Semibalanus balanoides (L.) facilitates its growth and reproduction on mussel beds in the Wadden Sea

On the unstable sedimentary tidal flats of the Wadden Sea, a suitable attachment substrate for sessile organisms is generally lacking. Epibenthic mussel beds (Mytilus edulis L.) provide the only and strongly limited settlement sites available for the barnacle, Semibalanus balanoides (L.). Field investigations showed that barnacles were non-randomly distributed within a mussel bed. They preferentially occurred near the siphonal apertures of living mussels but rarely grew on dead mussels or shell fragments. Field experiments revealed that this was due to selective settlement of barnacle cyprid larvae. Growth of barnacles was significantly higher upon living mussels than on empty mussel shells. Moreover, a higher reproductive output was obtained by individuals on living mussels which produced twice as many nauplii larvae than barnacles attached to empty shells. This study shows that selective settlement of S. balanoides cyprid larvae on living mussels is adaptive with respect to individual fitness. Received in revised form: 15 January 2001 Electronic Publication     

Spatial distribution of turtle barnacles on the green sea turtle, Chelonia mydas

Distribution patterns of epibiotic barnacles on green sea turtles were investigated in waters neighboring Okinawa, Japan. A number of barnacle species were found to coexist on the turtles and were classified into three genera: Chelonibia, Platylepas and Stomatolepas. Attachment sites on the turtles varied among the barnacle species, suggesting that there is niche partitioning with respect to their microhabitat selection. Turtle bodies offer a “patchy” environment for barnacles, so we also analyzed coexistence patterns in the context of an aggregation model. Within each genus, individual barnacles showed a clumped distribution. The different genera do not have mutually exclusive distribution patterns, but instead occur on the same turtle to various degrees. However, when turtles were divided into two size classes, both the level of aggregation and the degree of interspecific overlap among the barnacles was significantly lower on large turtles. We suggest that obtaining basic information on turtle epibionts will shed light on the biology of wild turtles, which is still largely unknown.     

Surface elastic modulus of barnacle adhesive and release characteristics from silicone surfaces

The properties of barnacle adhesive on silicone surfaces were studied by AFM indentation, imaging, and other tests and compared to the barnacle shear adhesion strength. A multilayered structure of barnacle adhesive plaque is proposed based on layered modulus regions measured by AFM indentation. The fracture of barnacles from PDMS surfaces was found to include both interfacial and cohesive failure of barnacle adhesive plaque, as determined by protein staining of the substratum after forced barnacle release from the substrate. Data for freshly released barnacles showed that there was a strong correlation between the mean Young's modulus of the outermost (softest) adhesive layer (E< 0.3 MPa) and the shear strength of adhesion, but no correlation for other higher modulus regions. Linear, quadratic, and Griffith's failure criterion (based on rough estimate of crack length) regressions were used in the fit, and showed significance.