Characterization of barnacle (Balanus eburneus and B. cenatus ) adhesive proteins

Cements from two species of barnacles, Balanuseburneus Gould and Balanus crenatus Bruguiére, were analyzed to identify the number and nature of proteins present. B. crenatus cement was composed mainly of a small peptide cross-linked into aggregates ranging in size from ˜ 3 kD to 40–50 kD. These aggregates could be reduced with 2-mercaptoethanol (2-ME) in the presence of sodium dodecyl sulfate (SDS) but only after incubation at 40 °C for 12 to 24 h and only when the cement was recently formed. Reductive alkylation of cysteine residues with 4-vinylpyridine produced a water-soluble peptide of less than 5 kD. By comparison, the cement of B. eburneus could be dissolved only partially in SDS and 2-ME when heated at 100 °C for 10 min. Five major proteins were identified by SDS-PAGE: 7, 22, 36 and 58 kD bands for which N-terminal sequence and amino acid compositions are presented; and a 52 kD band for which sequence data are given. A minor protein band of ˜ 80 kD has the same N-terminus as the 36 kD band. CNBr digests of individual proteins produced peptides for which sequence and composition data are also presented. The study was conducted during 1993 to 1995. In general, the proteins identified from B. crenatus cement are similar to those characterized from B. eburneus, and they are different in composition and sequence than those previously reported from Mytilus edulis. Received: 7 August 1996 / Accepted: 8 October 1996     

Desiccation as a factor in the intertidal zonation of barnacles

Four species of balanomorph barnacles, Balanus crenatus Brugière, B. balanoides (L.), Elminius modestus Darwin and Chthamalus stellatus (Poli), were studied to assess the susceptibility of intertidal barnacle species to desiccation. Known sized samples of barnacles were exposed to controlled desiccating conditions and subsequent survival and water loss were determined. It is clear that the ability to live high on the shore is dependent on a reduction of the overall permeability to water loss. Because of greater surface area to volume ratios, small stages are particularly prone to desiccation. In normal intertidal emersion periods, small stages of B. crenatus particularly, and also of B. balanoides and E. modestus which are similar in their desiccation resistance, would be susceptible to desiccation at normal temperatures and low humidities. Large barnacles would be more prone to death from high temperatures when the tide is out. The spat of C. stellatus, although surviving much longer than spat of larger dimensions of the other species, must also be prone to prolonged emersion conditions at high shore levels.     

The effect of heavy metals on photosynthesis and loss of cell potassium in two species of marine algae,Dunaliella tertiolecta and Phaeodactylum tricornutum

Observations on sixth-stage nauplius (pre-cyprid) of the barnacle Balanus eberneus Gould treated with L-proline-3,4-H³ suggest that the cement gland in this stage is a functional organ with considerable biosynthetic activity; the activity of the cement glands in the cyprid similarly treated is relatively low, ceasing gradually during, and completely after, metamorphosis. In the early phases of this stage, the glands become highly vacuolated (degenerate), and disappear completely in 3 days. The origin of the cement apparatus in the newly metamorphosed barnacle is discussed.     

A new method for obtaining barnacle cement in the liquid state for polymerization studies

A new method for collecting barnacle cement in the liquid state is described. This permits studies on the transformation of cement from liquid to solid states not heretofore possible. Preliminary analyses showed that polymerization, i.e., the in vitro change from a clear liquid to an opaque solid mass is not affected by temperatures ranging from-10° to 45°C, by diluting with distilled water up to two-fold, or by treating with selective enzymes; some chemical inhibitors induce the formation of a white precipitate, possibly indicating denaturation of the protein. It is concluded that the liquid cement contains all the necessary components for self-assembly, progressively changing from a clear liquid to an opaque rubbery insoluble mass. The polymerization process is time-dependent and requires no exogenous catalyst. A mechanism that may be involved in the adhesion process of the barnacle is suggested.     

Environmental control of the breeding of three boreo-arctic cirripedes

Three species of barnacles with boreo-aretic distribution were shown to require maintenance for several weeks below a critical temperature before the breeding condition could be attained. The temperatures critical for Balanus balanoides (L.) Balanus balanus (L.) and Balanus crenatus (Bruguière) were found to be between 10° and 12°C, 10° and 14°C, and at about 17°C, respectively. Although the strong influence of continuous light and the weak influence of continued feeding in delaying the onset of breeding in B. balanoides were confirmed, there remained some outstanding anomalies between the breeding behaviour of this species under laboratory conditions and between the tide marks. It was found impossible to initiate breeding by the application of conditioning procedures significantly in advance of the time of the normal Autumn breeding season in B. balanoides. Breeding appears to be inhibited, independently of external conditions, for a set period after the preceding brood cycle. Evidence points to a similar, largely endogenous, control of breeding in B. balanus which also breeds once annually, but not in B. crenatus which breeds continuously so long as food and temperature levels permit.     

Influence of substratum heterogeneity and settled barnacle density on the settlement of cypris larvae

On the Atlantic coast of Canada, Semibalanus balanoides (L.) is widely distributed in the mid-intertidal zone, whereas in the Gulf of St. Lawrence, this species is mostly limited to crevices. We tested the hypothesis of regional differences in microhabitat selection by barnacle larvae at settlement in 1984 and 1985 at St. Andrews, New Brunswick, Canada. Since larvae settle in microhabitats already colonized by adults, the relative influence of settled barnacle density and of different scales of substratum heterogeneity on settlement were evaluated experimentally at Capucins, Québec, (Gulf of St. Lawrence) and at St. Andrews, New Brunswick (Atlantic coast). On a large scale (>10 cm deep crevices) of heterogeneity, results show that, in the Gulf, cypris larvae settled nearly exclusively (93%) in natural crevices rather than on adjacent horizontal surfaces. On the Atlantic coast, settlement was more important outside than inside of crevices, when the substrata were either natural or artificial. This result is unique and contrasts sharply with all known reports on barnacle settlement in relation to surface contour. The influence of barnacle density on settlement was greater than that of large scale heterogeneity. On a small scale (<1.5 cm deep cracks), the presence of conspecifics had a stronger effect on settlement than heterogeneity in both regions. Field observations showed a relationship between larval settlement density and percentage of adult cover. Settlement increased up to 22 or 30% (Gulf and Atlantic coast) of adult cover and decreased afterwards. The results confirm the hypothesis of larval selection for cryptic habitats in the Gulf and the opposite behaviour (preferences for horizontal surfaces) on the Atlantic coast. This microhabitat selection is apparent at large scales of heterogeneity, whereas at small scales, the presence of conspecifics is the predominant factor.Contribution to the programme of GIROQ (Groupe interuniversitaire de recherches océanographiques du Québec)     

Tolerance of high temperatures by some intertidal barnacles

The median lethal times of survival of the barnacles Elminius modestus, Balanus crenatus and B. balanoides, when continuously submerged at high temperatures, were determined for individuals collected in the summer and winter. In E. modestus and B. crenatus there was no seasonal change in high temperature tolerance. In B. balanoides, however, the adults were more susceptible in the winter than in the summer to temperatures a little below the upper lethal temperature. Using less comprehensive data for other British species of barnacles, it is concluded that, in general, the order of tolerance to high temperatures corresponds to the order of temperatures within the geographical and the intertidal distributions of the species. From the time-temperature-survival curves, intertidal barnacles are living closer to environmental temperatures than would be supposed on the basis of the measurement of the upper lethal temperature, which has been commonly measured for many species, but is of less ecological significance.     

Scanning electron microscopy of metamorphosis in four species of barnacles (Cirripedia Thoracica Balanomorpha)

Metamorphosis and early juvenile development was followed in the laboratory in Balanus amphitrite and B. improvisus (family Balanidae) and in Semibalanus balanoides and Elminius modestus (family Archaeobalanidae) from cyprid settlement until 24 h after ecdysis. Stages of development were studied in vivo and with scanning electron microscopy. Events in metamorphosis and early juvenile development are very similar in all four species and can be interpreted in terms of a highly modified moult. Notably, there was no indication whatsoever of the amorphous decorticated settler previously described from metamorphosis of B. amphitrite. The shape of a juvenile barnacle with cirri and incipient shell plates is apparent immediately after the shedding of the cypris carapace, and rudimentary peduncle can be distinguished below the developing wall plates. A basal row of cuticular hairs encircles the peduncle in all species except S. balanoides. These hairs seem to serve a restraining function during early development, when the juvenile barnacle is only attached by the initially secreted cyprid cement. Similarly situated hairs are also present in very young juveniles of the lepadomorph Scalpellum scalpellum. In Semibalanus balanoides nothing indicates that the rostrum originates from the fusion of two latera as previously claimed, since this plate is single as soon as it can be distinguished.     

Hypoxic life of intertidal acorn barnacles

Oxygen levels were monitored within the mantle cavities of three barnacle species ( Chthamalus stellatus, Semibalanus balanoides, Elminius modestus), using optode microsensors. Conditions were always hypoxic, even when barnacles were actively using the prosoma and cirri to pump aerated seawater into the mantle cavity. Mantle fluid oxygen concentrations were extremely variable; behaviour and oxygen concentrations were not closely coupled. Ventilation of the mantle cavity depended partially on external water flow, with higher and more stable mantle fluid oxygen concentrations being sustained when the water around barnacles was agitated. During emersion, barnacles initially pumped seawater between the mantle cavity and the cone above the opercular plates to achieve ventilation. As water was lost it was replaced by air bubbles, eventually resulting in an air-filled mantle cavity. In S. balanoides and E. modestus, once the mantle cavity was filled with air, the barnacle usually used up the oxygen within the bubble within 2–3 h and did not regain oxic conditions until the barnacle was reimmersed in seawater. In C. stellatus, the air bubble was repeatedly refreshed for many hours by pneumostome formation. In response to low environmental salinity, all three species closed the opercular plates firmly and rapidly used up oxygen within the mantle fluid.     

Sugar compound products of the periphytic diatom <Emphasis Type="Italic">Navicula ramosissima</Emphasis> induce larval settlement in the barnacle, <Emphasis Type="Italic">Amphibalanus amphitrite</Emphasis>

This study investigated the response of cyprids of the barnacle Amphibalanus amphitrite to 23 strains of laboratory cultured periphytic diatoms isolated from microbial biofilms that formed on glass slides immersed in Tachibana bay, Nagasaki, and those from mass-production tanks in the Fisheries Center of Nagasaki City, Japan. In addition, periphytic diatoms were subjected to various treatments, in order to investigate the nature of the chemical cue in periphytic diatoms. Cyprids of A. amphitrite responded differently to the 23 different periphytic diatom strains and settled in high percentages on Cocconeis sp. and Navicula ramosissima strain A. On the other hand, nine strains of diatoms significantly inhibited settlement. The settlement inducing activity of N. ramosissima strain A increased linearly with diatom density, and its activity was enhanced by culturing the diatom under a bacteria-free condition, suggesting that specific diatom species, i.e., N. ramosissima strain A, may play an important role on larval settlement of the barnacle. Subjecting N. ramosissima strain A biofilm to hydrochloric acid (HCl) and ethanol (EtOH) treatments or heating it at 100°C did not inactivate the film, indicating that the settlement cue was a stable surface bound compound that did not decompose from the above treatments. Moreover, of the various lectins, enzymes, and drugs [H₅IO₆ and sodium dodecyl sulfate or (SDS)] used, only Lentil Agglutinin (LCA) treatment of N. ramosissima strain A biofilm resulted in the reduction its settlement inducing activity. A positive correlation was observed between the settlement inducing activity and the amount of LCA conjugated fluorescein isothiocyanate (FITC-LCA) of N. ramosissima strain A. On the other hand, subjecting biofilms of N. ramosissima strain B, an inactive strain, to various types of treatments resulted in the induction of A. amphitrite larval settlement but LCA treatment also reduced the activity of these treated N. ramosissima strain B biofilms. These findings suggest that a cue containing an LCA-binding sugar chain is present in both A and B strains of N. ramosissima but the large amount of mucous substance covering N. ramosissima strain B biofilm probably makes the sugar chain containing active subunit in strain B unavailable to A. amphitrite cyprids. In conclusion, periphytic diatoms such as N. ramosissima play an important role in larval settlement of the barnacle A. amphitrite. The cue in the diatom was an LCA-binding sugar chain(s) compound that may have similarities to the settlement inducing protein complex (SIPC) from adult shell of the barnacle.     

Thraustochytrid protists as a component of marine microbial films

Although ubiquitous in the marine environment, the presence and importance of thraustochytrid fungoid protists in primary film formation of freshly immersed surfaces in the sea have not been investigated. We isolated thraustochytrids from surfaces of glass, aluminium, mild steel and fibre glass panels immersed in sea water for 1 to 4 d, but not from those of copper and cupro-nickel. Microscopic examination of the primary film on glass surfaces revealed the presence of thraustochytrids. Thraustochytrids grew to varying population densities on surfaces of glass, aluminium and fibre glass in the laboratory. Scanning electron microscopy of the growth of Ulkenia profunda Gaertner, isolated from the primary film, on surfaces of glass and aluminium suggested that cells of the thraustochytrid attached directly to the surfaces, without producing copious extracellular polysaccharides. The presence of ectoplasmic net elements was not a prerequisite for attachment of cells to surfaces. Cell surface hydrophobicity of the thraustochytrid, as estimated by the MATH assay, might play an important role in adhesion. The presence of thraustochytrid cells on a polystyrene surface markedly induced settlement of barnacle larvae (Balanus amphitrite), as compared to barnacle extract and a control. Received: 26 July 1999 / Accepted: 21 January 2000     

Community-level and species-specific responses of coastal phytoplankton to the presence of two mesozooplanktonic grazers

The response of phytoplankton to the presence of two mesozooplanktonic grazers was studied in a 44-h laboratory experiment from the phytoplankton perspective. The <45-μm-filtered plankton community, as well as females of the copepod Eurytemora affinis and nauplii of the barnacle Balanus improvisus originated from the northern Baltic Sea. The phytoplankton community was dominated by the dinoflagellate Heterocapsa triquetra, which was preferred as food by the mesozooplanktonic grazers, especially E. affinis. Cryptophytes were eaten by the B. improvisus nauplii, while heterotrophic nanoflagellates increased in the presence of B. improvisus nauplii, and colonial cyanobacteria increased in the presence of E. affinis. Because of the strong selective feeding on the dominant, large-sized phytoplankton species, the negative effect of the mesozooplanktonic grazers on total chlorophyll a was stronger than any cascade effect releasing phytoplankton from protozoan grazing.     

High intertidal community organization on a rocky headland in Maine,USA

A mosaic patchwork of the barnacle Balanus balanoides L., the mussel Mytilus edulis L., and the alga Fucus vesiculosus L. was found in the transitional region between the mid and high intertidal zones on a rocky headland on Mount Desert Island, Maine, USA. The development of the mosaic was observed by following recruitment and survival of B. balanoides in denuded patches located at the same tidal level along a 60 m stretch of shore. Barnacle recruitment was least under canopies of F. vesiculosus and greatest in open areas kept moist at low tide by surf. Barnacle survival after settlement was least under the F. vesiculosus canopy due to the whiplash effect of the algal fronds in the surf and greatest in open areas free from competition from mussels. In open areas, early mortality was correlated with settlement density. In areas of dense settlement (60 spat cm^-2) up to 90% mortality resulted within 5 months from crowding associated with growth. In older individuals crowding produced hummocks of elongated, weakly attached barnacles which were more prone to removal by surf than uncrowded barnacles. Mussels exerted competitive dominance over barnacles for space and the presence of mussel beds prevented further barnacle recruitment. Mussels suffered extensive mortality during winter storms when surf removed dense mats of weakly attached mussels. The patchy distribution of mussels and barnacles results from irregular rock substrata producing numerous environmental patches with respect to wave exposure and drainage at low tide, and from densitydependent mortality of both mussels and barnacles which creates patches of new colonizable space within each environmental patch.     

Inhibition of attachment of larval barnacles,Balanus amphitrite,by bacterial surface films

Films of bacteria on solid substrata can positively or negatively influence the attachment of marine invertebrate larvae. Effects of marine bacteria on the attachment of cypris larvae of the barnacle Balanus amphitrite Darwin were examined in the laboratory. Bacteria, grown to mid-exponential phase and allowed to adsorb irreversibly to polystyrene petri dishes, attached in densities of 10⁷ cells cm^-2. Assays (22h) were used to compare the effects of adsorbed cells of 18 different bacterial species on larval barnacle attachment. Most of the adsorbed bacteria either inhibited or had no effect on larval attachment compared to clean surfaces. Experiments testing the effect of larval age on barnacle attachment were conducted with six species of bacteria and showed that older larvae attached in higher percentages to clean surfaces and that bacterial films generally inhibited larval attaschment. Both the species of bacteria and the in situ age of the adsorbed bacteria affected barnacle attachment response: older films of Deleya (Pseudomonas) marina were more inhibitory. Bacterial extracellular materials may be involved in the inhibitory process.     

Are barnacle larvae able to escape from the threat of UV?

Earlier laboratory experiments suggested that environmental levels of UV-B radiation can damage the eyes of barnacle naupliar larvae and impair their phototactic behaviors. However, since barnacle larvae may avoid UV by migrating to deeper waters, it is not known whether such impairment would actually occur under field conditions. For the first time, this study provides both field and laboratory evidences to show that prevailing UV-B in the natural habitat of barnacle larvae could be an important environmental factor affecting natural barnacle populations. We here showed that although barnacle nauplii may avoid UV-B irradiation by downward migration, the amount of UV energy (9.8 × 10⁻⁶ J) received by a naupliar eye during downward migration in the natural water column is within the same order of magnitude as the total energy (7.5 × 10⁻⁶ J) sufficient to cause damages to naupliar eye and impair their phototactic responses. It is possible that solar UV-B prevailing at shallow waters would pose a similar threat to other zooplankton species over large geographic scale.     

Physiological tolerance predicts species composition at different scales in a barnacle guild

This study examined how the species composition of an intertidal barnacle guild varied according to physical gradients in the environment at small scales governed by microclimates, medium scales of wave exposure and large scales of latitude. Barnacle distributions at small and medium scales were sampled in Ireland between 51°29′ and 52°44′N and 6°50′ and 10°08′W. Sampling on European shores spanned ~18° latitude from 37°05′ to 55°16′N. Barnacle surveys mainly took place in 2003–2004. An index of wave fetch was calculated along the wave exposure gradient using a digital coastline-based model that was supported by a biological exposure scale. A ‘dryness’ index was defined according to mean monthly wind speed, fetch along the average wind direction and mean monthly air or sea surface temperatures for 2 years (January 2001–December 2002) which is the period when the most recent adults in the barnacle community would have settled and grown to adulthood. The proportion of the dry-loving barnacle Chthamalus montagui Southward increased within the barnacle guild at all scales as the habitat became warmer and drier. Barnacle densities were high in all habitats, mean densities ranged from a minimum of 4.16 cm⁻² on moderately exposed shores to a maximum of 6.27 cm⁻² in sunlit or south-facing microclimates. Percentage cover of barnacles across the gradient of latitudes was usually >70%. The results suggest that the distribution and abundance of interacting barnacle species on European coasts is strongly controlled by abiotic factors, most likely temperature and desiccation.     

Zoogeography of fish parasites of the pearlside (Maurolicus muelleri), with genetic evidence of Anisakis simplex (s.s.) from the Mid-Atlantic Ridge

A total of 200 Maurolicus muelleri from the Mid-Atlantic Ridge (MAR) and the Norwegian Deep (ND) were studied for parasitic helminths. Two different metazoans were isolated from the MAR and five species from the ND. The predominant parasite species in both areas were tetraphyllidean cestode larvae (Scolex pleuronectis) and the anisakid nematode Anisakis simplex sensu stricto (s.s.), the latter being identified using genetic analysis of the internal transcribed spacer (ITS-1, ITS-2) and 5.8S regions of the rDNA. The parasite fauna of M. muelleri from the MAR was less species rich in comparison to ND, due to the deep-sea and oceanic environment. The digeneans Brachyphallus crenatus and Lecithaster confusus as well as the raphidascarid fish nematode Hysterothylacium aduncum were only collected from the ND. This can be explained either by the deep origin of the sampled fish specimens or the lack of suitable intermediate or final hosts in the region. Based on the frequent occurrence of A. simplex (s.s.) around the MAR and the ND, a pelagic life cycle is suggested at both localities, involving baleen and toothed whales as final and pelagic and mesopelagic fish and invertebrates as intermediate or paratenic hosts.     

Studies of marine fouling and boring off Kodiak Island,Alaska

Marine fouling and boring off Kodiak Island, Alaska, was studied at depths of 5, 15, and 30 m by exposing wood/asbestos test panels and retrieving them at monthly and cumulatively longer intervals. Fouling was moderate between June and October and negligible between November and May. The fouling communities at 15 and 30 m were quite dissimilar, probably due to a large surface-to-bottom salinity gradient. The community at the 30 m level was dominated by Balanus crenatus and Pseudochitinopoma occidentalis, that at 15 m by Alcyonidium polyoum and Obelia borealis. At the 5 m deep, pierside site, the most common organisms were B. crenatus, O. borealis and Desmacystis sandalia. Borer attack was a trace at 5 and 15 m and slightly more severe at 30 m.     

Responses of cirripede larvae to light. I. Experiments with white light

The phototactic response of the nauplius larva of Balanus balanoides, B. crenatus and Elminius modestus shows darkadaptation; the response of the cyprid of B. balanoides shows both phototaxis and low photokinesis. The phototactic responses and the orientation of the cyprid to white light at settlement require an intensity of illumination slightly above 10^-5 lux. The ability to select a shaded position by cyprids of R. balanoides requires a higher intensity of 10^-2 to 10^-4 lux; hence a different mechanism may be involved. Barnacle larvae are sufficiently sensitive to be able to respond to light beneath the sea surface, even on cloudy, moonless nights.     

Sponge-inhabiting barnacles on Red Sea coral reefs

In this study eight different species of barnacles were found within nine species of sponges from the Red Sea. This brings to 11 the number of sponge-symbiotic barnacles reported from the Red Sea, two of these are new Acasta species (not described herein) and one (A. tzetlini Kolbasov) is a new record for this sea. This number is much higher than that of symbiotic barnacles found within sponges from either the N. Atlantic (2) or the Mediterranean (4). Two possible explanations for this are the presence of numerous predators in coral reefs and scarcity of available substrate for settlement. These factors can lead to high incidence of symbiotic relationships. Of the nine sponge species, only one (Suberites cf. clavatus) had previously been known to contain barnacles. Even at the family level, this is the first record of symbiotic barnacles in two out of the seven sponge families (Latrunculiidae, Theonellidae). Our present findings strengthen the apparent rule that the wider the openings in a barnacle shell, the fewer the host taxa with which it will associate, usually from one or two closely related families, and the more frequent it will associate with elastic sponges. Most Neoacasta laevigata found on Carteriospongia foliascens were located on the same side as the sponge's ostia, i.e. facing the incoming water. This adaptation allows the barnacles to catch more suspended particles from the water, provides them with more oxygen and prevents their exposure to discharged sponge waste. The highest density of barnacles observed on one face of a “leaf ” (with ostia) was 0.389 barnacles cm⁻² (one barnacle per 2.57 cm²) and on average 0.181 ± 0.68, while the average on the other side was only 0.068 ± 0.52 barnacles cm⁻². As indicated by the Morisita index, these barnacles most frequently (58%, n = 12) had a clumped spatial distribution (while the rest were randomly distributed), as is to be expected from such sessile organisms with internal fertilization via copulation. The presence of N. laevigata induced the growth of secondary perpendicular projections of its host C. foliascens. Of the N. laevigata examined, 17% brooded 324 ± 41 embryos each, of 286 ± 17 μm total length; only 5.7% (n = 123) were found to be dead. Size distribution analysis of skeletal elements from dead barnacles showed them to be significantly larger than the skeletal elements of the population of live barnacles ( p < 0.05). Received: 26 June 1998 / Accepted: 1 December 1998