Noncovalent bonds are key mechanisms for the cohesion of barnacle (Balanus crenatus) adhesive proteins

Cement of the barnacle species, Balanus crenatus, was isolated and separated by SDS-PAGE. Reductive and nonreductive conditions yielded the same result of six major proteins (in the range up to 100 kDa) occurring as two triplets. A third triplet of lower molecular mass was faintly visible. The presence of a reductive agent had only little impact on the solubility of B. crenatus cement. The strong effect of the denaturing conditions (the presence of SDS and the incubation at elevated temperature) on the solubility of the barnacle cement and the absence of disulfide signals in Raman spectra suggest a complex structure of the barnacle cement noncovalently bound. It can be speculated that this protein complex in its quaternary structure is formed by small subunits and is equivalent to the globular cement structures previously detected.     

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.     

Influence of activity and salinity on the weight-dependent oxygen consumption of the rainbow trout Salmo gairdneri

Two types of secretory cell have been found in the cement glands of the cypris larva of Balanus balanoides (L.). Histochemical tests show the presence of proteins, phenols and the enzyme polyphenol oxidase in both the glands and secreted cement. It is proposed that cyprid cement is composed of tanned protein, all the components of which are derived from the cement glands.     

Ecophysiology of Limonium axillare and Avicennia marina from the coastline of Arabian Gulf-Qatar

Attention is being focused on the coastline from Doha to Ras Laffan in Qatar since higher activities in the development of land and establishment of roads, highways and new buildings and houses is not coupled by serious studies on habitat destruction, fragmentation or disturbances. Ecophysiological study was carried out to investigate the adaptation of two halophytes (Limonium axillare and Avicennia marina) in this area, with special emphasis on the ultrastructure of salt glands found in the leaves. Soils in these locations accumulated much Na⁺ and Cl^? as compared to other cations like K⁺, Ca²⁺ and Mg²⁺. Both plants accumulated higher concentrations of Na⁺ , Cl^?, and Ca²⁺ and lower concentrations of K⁺ and Mg²⁺. Organic compounds found in leaves of these plants under their natural habitats including proline, soluble sugars and nitrogen, and photosynthetic pigments were determined. Scanning electron micrographs of the surface of leaves showed that salt glands of these plants are well developed. It is urgently required that exact vegetation maps, and monitoring exercises will be conducted, in order to document exactly the state of the vegetation in Qatar. Only this will allow the environment authorities to bring forward suggestions for vegetation and ecosystem management to the decision makers.     

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.     

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.     

Attachment strength and colonization patterns of two macrofouling species on substrata with different surface tension (in situ studies)

Attachment strength and colonization patterns of one barnacle (Balanus c.f. variegatus) and one polychaete species (Pomatoleios kraussii) on seven artificial substrata with surface tensions between 19 and 64.5 mNm^-1 were studied between June 1991 and January 1992 at Laem Than (Chonburi province, Gulf of Thailand). The purpose of the present study was to demonstrate the role that surface tension plays under natural conditions in colonization success by these species. Does stronger adhesion to one substratum result in higher densities of specimens on that particular surface? Although both species adhere much better on substrata with higher surface tension than on surfaces with low surface tension (B. c.f. variegatus: between 0.04×10⁵ and 16.35×10⁵ Nm^-2 on surfaces of 22 and 33.5 mNm^-1, respectively; P. kraussii: between 0.22×10⁵ and 1.73×10⁵ Nm^-2 on 19 and 33.5 mNm^-1, respectively), colonization pattern is not influenced by surface tension. The major factors influencing settlement patterns seem to be space competition for B. c.f. variegatus and attraction to already settled adults by P. kraussii. The results show that the importance of surface tension in marine fouling control is limited because organisms colonize substrata successfully despite lower adhesion. Attachment strength is not a limiting factor under natural conditions.     

Some observations of factors affecting embryonic and larval growth of Pollicipes polymerus (Cirripedia: Lepadomorpha) in vitro

Observations on the effects of several environmental conditions in embryonic and larval cultures are reported for the gooseneck barnacle Pollicipes polymerus (Sowerby, 1833). When the growth rates of control embryos kept in finger bowls at ambient-temperature sea water (ca. 12°–15°C) were compared to those of embryos grown under variously modified circumstances, it was found that growth rates were faster under conditions closely simulating the adult barnacle mantle-cavity (e.g. darkness and aeration). Addition of antimicrobial drugs and reduction of egg-mass size also promoted fast growth and development. Nauplii were fed several species of algae; only three species promoted growth to Stage V or older. Naupliar growth was fastest in larvae fed the combination of Prorocentrum micans/Platymonas sp. Larvae fed Prococentrum micans/pennate diatom grew slowest, and most larvae died at Naupliar Stage III. Interactions between algal species may have affected their nutritional value for Pollicipes polymerus nauplii. Less larval activity, slower growth rates, and higher mortality rates were observed in individuals given small amounts of food. The development of embryos and larvae in culture depends greatly on the culture conditions. These conditions should be described if comparison of timetables and envents are to be made between studies.     

Effect of the insect growth regulator diflubenzuron (Dimilin®) on the uptake of glucose and N-acetylglucosamine into the cuticle of crab larvae

Incorporation of ³H-labelled glucose and ³H-labelled N-acetylglucosamine (NAGA) — both precursors to chitin —into the cuticle of Rhithropanopeus harrisii (Gould) larvae (Crustacea: Brachyura) has been examined at different stages of the moult cycle in control larvae as well as in larvae treated with the insect growth regulator diflubenzuron (Dimilin^®). As far as the control larvae were concerned, the incorporation of both precursors was high at the postmoult stage when endocuticle was secreted. NAGA appeared to be a more specific precursor of cuticular material than glucose during the premoult stage when exocuticle was produced. Incorporation of both precursors was low immediately before ecdysis and during the intermoult stage when secretion of the cuticle is complete. The results show that incorporation of glucose into chitin was greatly inhibited by the pollutant during the postmoult stage when endocuticle is produced, while incorporation of NAGA was reduced to a lesser extent at this stage. Diflubenzuron treatment markedly affected the incorporation of both NAGA and glucose in the premoult stage during secretion of exocuticle.     

Behavioural and electrophysiological responses of the olive fruit fly, Bactrocera oleae (Rossi) (Diptera: Tephritidae), to male- and female-borne sex attractants

In the olive fruit fly, Bactrocera oleae, females attract males by producing 1,7-dioxaspiro[5.5]undecane (olean), the main component of the sex pheromone secreted by rectal glands. It has been recently demonstrated that males are able to produce (Z)-9-tricosene (muscalure) in rectal glands, a compound that selectively attracts females. In this study, a male grooming reaction that may transfer the male-borne compounds from rectal to urotergal glands was observed, suggesting that urotergal glands could be involved in B. oleae sexual communication. GC/MS, EAG, GC/EAD analyses and behavioural assays were carried out to compare the role of male rectal and urotergal glands during courtship. In both male glands, olean and muscalure amounts were age dependent. Extracts of rectal glands contained higher amounts of olean and/or muscalure than urotergal ones. Extracts of rectal and urotergal glands of males and females elicited EAG responses in both sexes. GC/EAD showed that female EAG response to male rectal extracts was mainly due to olean and muscalure. Synthetic compounds evoked EAG dose-dependent responses in both sexes, and the EAG response to muscalure was higher as compared to olean. Rectal and urotergal glands from old males were able to attract females, while urotergal glands from young males attracted only males. Overall, our results add knowledge to the mating system of B. oleae, giving first evidences on the electrophysiological activity of muscalure towards both sexes, as well as on the involvement of male urotergal glands in the chemical sexual communication of this pest.     

Activity patterns and predatory behavior of an intertidal nemertean from rocky shores: Prosorhochmus nelsoni (Hoplonemertea) from the Southeast Pacific

Understanding the impact of environmental stressors on predator activity is a prerequisite to understanding the underlying mechanisms shaping community structure. The nemertean Prosorhochmus nelsoni is a common predator in the mid-intertidal zone on rocky shores along the Chilean coast, where it can reach very high abundances (up to 260 ind m⁻²) in algal turfs, algal crusts, barnacle crusts, and mixed substrata. Tidal and diurnal scans revealed that the activity of P. nelsoni is primarily restricted to night and early-morning low tides and is relatively low when air temperatures are high. On average, larger worms crawled faster than smaller worms, with their maximum velocity being influenced by substratum type. Their estimated rate of predation is 0.092 prey items nemertean⁻¹ day⁻¹, just below the laboratory rate of ~0.2 amphipods nemertean⁻¹ day⁻¹ previously estimated for this species. P. nelsoni consumes a diverse spectrum of prey items (i.e., amphipods, isopods, decapods, barnacles, and dipterans) and is possibly exerting a significant influence on its prey populations. We suggest that the opportunistic predatory behavior of this intertidal predator is caused by the trade-off between immediate persistence (e.g., avoidance of desiccation) and long-term survival through successful foraging.     

Initial contact,exploration and attachment of barnacle (Balanus amphitrite) cyprids settling in flow

Settlement responses of barnacle (Balanus amphitrite) cyprids to boundary-layer flows were examined in laboratory flume-experiments. The leading-edge configuration of flat plates was altered in order to manipulate flows without changing surface topography or freestream velocity. Settlement along the plates correlated strongly with downstream gradients in shear stress. Analyses of video images taken during the experiments indicate that cyprids first contact plates in regions where plate-ward advection is high, and subsequent exploratory movement along the plate is oriented with flow direction at the plate surface. After exploration, cyprids reject a surface more frequently in a fast flow (10 cm s^–1 freestream velocity) than in a slow flow (5 cm s^–1), but rejection occurs in shear stresses well below the threshold that would prevent attachment and exploration. A higher rejection rate does not result in lower settlement, however, since contact rate is higher in fast than slow flows. The movement of cyprids in flow thus appears to be a passive transport process during the initial contact stage of settlement, but an active behavioral response to flow direction and shear stress during later stages of exploration and attachment.     

Phenotypic variation in shell form in the intertidal acorn barnacle Chthamalus montagui: distribution,response to predators and life history trade-offs

The acorn barnacle Chthamalus montagui can present strong variation in shell morphology, ranging from flat conic to a highly bent form, caused by a substantial overgrowth of the rostrum plate. Shell shape distribution was investigated between January and May 2004 from geographical to microhabitat spatial scales along the western coast of Britain. Populations studied in the north (Scotland and Isle of Man) showed a higher degree of shell variation compared to those in the south (Wales and south-west England). In the north, C. montagui living at lower tidal levels and in proximity to the predatory dogwhelk, Nucella lapillus, were more bent in profile. Laboratory experiments were conducted to examine behavioural responses, and vulnerability of bent and conic barnacles to predation by N. lapillus. Dogwhelks did not attack one morphotype more than the other, but only 15 % of attacks on bent forms were successful compared to 75 % in conic forms. Dogwhelk effluent reduced the time spent feeding by C. montagui (11 %), but there was no significant difference between conic and bent forms. Examination of barnacle morphology indicated a trade-off in investment in shell structure and feeding appendages associated with being bent, but none with egg or somatic tissue mass. These results are consistent with C. montagui showing an induced defence comparable to that found in its congeners Chthamalus anisopoma and Chthamalus fissus on the Pacific coast of North America, but further work to demonstrate inducibility is required.     

Development of the gooseneck barnacle Pollicipes polymerus (Cirripedia: Lepadomorpha): Fertilization through settlement

Pollicipes polymerus (Sowerby, 1833) is a common barnacle in rocky intertidal zones of the West Coast of North America whose development has been previously undescribed in detail. In the present study, certain aspects of the development of P. polymerus are recorded from fertilization through settlement. In vitro fertilization has been accomplished using already-implanted sperm plus ovulating eggs and oviducal gland fluid. The eggs are small (about 100 μ diameter at fertilization; 90 x 140 μ at first cleavage) and somewhat yolky. Embryos obtained from brooding adults and from in vitro fertilization were reared in glass dishes until hatching. Cleavage was total, asynchronous and unequal. Division of the D macromere lagged behind division of the other blastomeres, and gastrulation occurred by epiboly so that blastoderm surrounded the macromeres. The nauplii hatched an average of 25.4 days (range was 20.3 to 30.1 days) after fertilization, and molted within 1 day to Stage II nauplii. From Stages II through VI the nauplii required food. As they grew from stage to stage, the number of setae per appendage increased. Nauplii have feathery and hispid-type setae, previously described only in Chthamalus aestuarii. In contrast to other pedunculate larvae which have been described, P. polymerus nauplii are small and have specialized feathery setae. Cypris-larva settlement was stimulated only when healthy adult peduncles were available, thus showing an affinity for its own species.     

Description of a Tetraclita stalactifera panamensis community on a rocky intertidal Pacific shore of Panama

The midlittoral zone at Paitilla Beach, Panama, Is occupied by a group of organisms closely associated with Tetraclita stalactifera panamensis. Upon death, this cirripede provides housing for a number of species bound by common substrata and interacting with each other due to their close physical proximity; the faunistic group associated with each barnacle qualifies then, as a small, well-defined community which represents a subsample of the more comprehensive midlittoral community dominated by T. stalactifera panamensis. In patches where it is abundant, T. stalactifera panamensis occupies 28% of the surface area available. It has a relative abundance of only 1% of the specimens found in the zone, but all the other 95 species (with an average of 26, 810 individuals/m²) identified from the area are associated either with the parietal canals, or the external or internal surfaces of T. stalactifera panamensis tests. Each one of these structures represents a habitat which provides surface for attachment, shelter, or both, to 37 species of polychaetes, 32 of mollusks, 20 of crustaceans, 3 of actiniarians, 2 of pycogonids, 2 of sipunculans and an unknown number of tanaid, nemertean and turbellarian species. The single most diverse space-niche is represented by the parietal canals (H=0.783), occupied mostly by polychaetes. dead barnacles harbor an average of 131 individuals per test, which compose a fauna of higher diversity (H=0.678) than that associated with live barnacles (H=0.163). The latter contain 53 specimens per test on the average. Although there is considerable overlapping of species associated with the different T. stalactifera panamensis space-niches, mollusks are clearly more abundant on internal surfaces, polychaetes in parietal, canals; Balanus spp. on external surfaces, and anemones at the base of the parapet on the external surfaces of the tests. Over 220 species of macro-invertebrates were identified from the rocky intertidal at Paitilla. Most of the species are concentrated in the lower (157 species, 1824 specimens/m²) and midlittoral (96 species, 26, 810 specimens/m²) intertidal zones. Splash zone is depauperate (6 species, 1578 specimens/m²). Of the entire fauna identified from the intertidal, 73 species (33%) occur in more than one zone and 147 (67%) are restricted to a particular belt. This last category of zone-restricted species breaks down into 42% for the lower intertidal, 17% for the midlittoral, 6% for the intermediate zone, and 2% for the splash zone.     

Toxic effects of enrofloxacin on Scenedesmus obliquus

In this article, the toxic effects of Enrofloxacin (ENFX) on Scenedesmus obliquus were studied, through investigating the growth, photosynthetic pigments, and protein contents. The possible toxic mechanisms of ENFX were analyzed by determining the superoxide dismutase (SOD) activity, malondialdehyde (MDA) content, proline content, and superoxide anion (O ₂ ^? ) generation rate. Results showed that the growth of algae was inhibited by ENFX and the 50% effective concentration (EC₅₀) values for 24, 48, 72, and 96 h of ENFX were 88.39, 63.86, 45.10, and 59.16 mg·L^?1, respectively. After treated with ENFX for 96 h, the contents of photosynthetic pigments decreased with the increase of ENFX concentration, the content of soluble protein and the activity of SOD increased and then decreased, and the generation rate of superoxide anion (O ₂ ^? ) increased continually. The contents of MDA and proline changed little in lower ENFX concentration groups, but increased rapidly when treated with higher concentration groups. These results suggested that ENFX affected the growth of S. obliquus, and the main toxicity mechanism was that algal cells generated the reactive oxygen species under ENFX stress, and then the reactive oxygen species (ROS) induced the oxidation damages of biologic macromolecules and changed the biomembrane permeability further.     

Balanus amphitrite (Cirripedia: Thoracica) — A potential indicator of fluoride,copper, lead,chromium and mercury in North Adriatic lagoons

The capacity of the barnacle Balanus amphitrite Darwin to accumulate pollutants above ambient levels was examined in two North Adriatic lagoons with respect to fluoride, copper, lead, chromium and mercury. Levels in soft tissues ranged from 138 to 312 ppm (dry weight) for fluoride, 41 to 109 ppm for copper, 7.1 to 11.7 ppm for lead, 2.10 to 3.89 ppm for chromium, and from 0.96 to 1.35 ppm for mercury. The concentration factor was of the order of 10² for fluoride, 10³ for copper, and possibly higher than 10³ for lead, chromium and mercury. The differences of about 2:1 in the fluoride and copper concentrations in the waters of the two lagoons were clearly reflected by the levels in the barnacles. B. amphitrite that had set on experimental panels had levels similar to those found in specimens collected from long-term natural populations as early as 42 days after immersion of the panels. Compared with literature data, the accumulation levels found in B. amphitrite for fluoride were among the highest known for invertebrates; the values for copper and lead were considerable, but exceeded by others published for B. balanoides; the results for chromium and mercury were one order of magnitude lower than values reported for other suspension-feeders or indicator organisms. B. amphitrite appears to possess most of the properties considered essential for a biological indicator. Eventual determination of the response time of the barnacle to changes in environmental level could profitably be carried out utilizing experimental panels.     

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.     

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.