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Three quantitative energy flow models have been constructed for the Sylt-Romo Bight representing the stocks and flows in the ecosystem during 1995, 2007 and 2010. The 1995 model consists of 59 compartments, while the 2007 and 2010 models have 63 compartments each. The 4 additional compartments in the 2007 and 2010 models are 2 native barnacle species Semi- balanus balanoides and Balanus crenatus and 2 invasive species namely the Pacific oyster Crasso strea gigas and the Australasian barnacle Austrominius modestus. Oyster and A. modestus biomass increased from virtually zero in 1995 to 15 and 1.3 gC m �2 , respectively, in 2007, and then subsequently decreased to 0.6 and 0.12 gC m �2 , respectively, in 2010. Reasons for the increases in oysters and A. modestus are related to climate change which favored strong recruitment of the invasive species during unseasonably warm summers, and the subsequent decline to weak recruitment of the invasive species and an unseasonably cold 2009/2010 winter. We used eco - logical network analysis (ENA) to quantify ecosystem structure and function through analyses of energy (or carbon) transfer between living and non-living compartments in each of the 3 network models. Results from ENA show an increase in most of the system attributes, but a decline in number of cycles, the trophic efficiency, the ascendency ratios, and relative redundancy from 1995 to 2007; these are ascribed to the impact of the invasive species on system organization and function. Following the dramatic decline of the invasive species from 2007 to 2010 most of the system attributes decreased. The variance in the biomass and associated flows of other compart- ments in addition to that of the invasive species has also played a role in the changes in system attributes and function.

The barnacle Balanus amphitrite Darwin is a dominant intertidal and major fouling organism. As its life cycle includes both sessile and pelagic phases, adaptation is a necessity to toler- ate harsh conditions that are faced in the intertidal and pelagic environment. A downward trend in dissolved oxygen levels in coastal areas has been recently observed, which would have a more prominent effect on sessile organisms inhabiting the intertidal habitats. The effects of reduced dis- solved oxygen levels (hypoxia) and no oxygen (anoxia) on B. amphitrite adults and nauplii was assessed. Their response was evaluated by estimating the levels of the antioxidant enzymes catalase and superoxide dismutase. The impact of desiccation on antioxidant enzymes in adult B. amphitrite collected from different tidal heights was also evaluated. When nauplii were starved, 100% mortal- ity was observed after 64 h under hypoxia, and between 24 and 32 h under anoxia. However, when nauplii were exposed to hypoxia and anoxia along with food, their tolerance level increased. A decrease in naupliar feeding rate and oxygen consumption was observed when they were exposed to hypoxia and anoxia. A significant difference in the levels of antioxidant enzymes in adult B. amphitrite collected at different tidal levels was observed. Enzyme activity increased with increase in the duration of tidal exposure, suggesting a capability to adapt to stress in different niches. Nauplii showed antioxidant defense (increased enzyme activity) under hypoxia and anoxia. The magnitude of change in enzyme levels suggests that these enzymes could be potential biomarkers of stress.

Multiple invasions by ecologically similar species can be viewed as ‘natural’ addition experiments in which access to key resources might be reduced. Possible outcomes might include: (1) the extirpation of a species already present in the ecosystem, (2) the exclusion of a new invader, or (3) niche compression, with each species using less of the shared resource. Chthamalus proteus, a barnacle that arrived in the Hawaiian Islands ~30 yr ago, is now the most abundant and widespread non-native barnacle in the intertidal zone on the island of Oahu. In a series of field experiments, I demonstrate that the abundance of an earlier invader — the larger, faster growing barnacle Balanus reticulatus — is reduced via substrate pre-emption in the zone of overlap between the 2 barnacle species. A third barnacle, Balanus amphitrite, which invaded Hawaii earlier than the other two, is now virtually absent from locations where it was once abundant and where C. proteus is now the numerical dominant. B. amphitrite did not settle on plates from which C. proteus was removed, suggesting that the presence of C. proteus is not the proximal cause of its decline. B. amphitrite is still present on Oahu, particularly in lower salinity settings. While successively invading barnacles have reportedly replaced each other in other locations around the world, it appears that invasive barnacles on Oahu are undergoing niche compression rather than complete replacement.

This study investigated the impacts of nutrient enrichment on intertidal biofilm micro- bial communities and on larval metamorphosis of invertebrates. We modulated the nutrient levels in the water column during biofilm development and examined the resulting biofilms for bacterial and diatom community composition using 2 DNA fingerprinting techniques and microscopic examina- tion, respectively. Results of both terminal-restriction fragment length polymorphism (T-RFLP) and denaturing gradient gel electrophoresis (DGGE) analyses indicated that the nutrient enrichment treatment altered the biofilm bacterial communities relative to the control. Biofilms that had different microbial compositions were tested for their effects on larval metamorphosis of the polychaete Hydroides elegans, the barnacle Balanus amphitrite, and the slipper limpet Crepidula onyx. We showed that nutrient enrichment in the water column effectively changed the biofilm composition in ways that increased its inductiveness for larval metamorphosis of C. onyx and B. amphitrite, but not of H. elegans. This study highlights the apparent importance of nutrient enrichment in influencing the intertidal biofilm composition directly and the invertebrates indirectly.

Sessile invertebrates and algae are ubiquitous as hosts for a range of fouling organisms in marine benthic habitats. Fouling can sometimes impose negative fitness effects, and hosts may decrease the amount of fouling through production of chemical defenses. Waterborne chemical defenses may rapidly become diluted and should be released continuously in order to inhibit fouling; this process may be costly for the host. Therefore, it has been hypothesized that natural selection should favor the evolution of defense metabolites that stay on the host surface in aquatic environ- ments. We investigated chemical deterrence of fouling in the marine sessile invertebrate Halichon- dria panicea (breadcrumb sponge), through both field observations and a series of laboratory bio- assays. Natural populations of H. panicea had very few macrofouling organisms. Cyprid larvae of the barnacle Balanus improvisus were deterred from attaching to the substratum in the presence of water-soluble compounds that were rapidly exuded from the sponges both under laboratory condi- tions and in the field. Artificial substrata placed near H. panicea in the field received less fouling by barnacles than control substrata placed farther away from sponges. These results clearly show that H. panicea releases water-soluble compounds that deter attachment of B. improvisus. However, these compounds may not have evolved as a chemical defense against fouling in the sponges. H. panicea can release epidermal cell layers, and therefore the barnacles may have evolved an ability to recog- nize water-soluble compounds from the sponges in order to avoid attaching to unsuitable substrata.

Delayed metamorphosis can affect post-metamorphic performance in a variety of marine invertebrate species, possibly through effects on larval energy reserves. Nevertheless, the consequences of the 'latent effects' of natural environmental conditions and the proximate causes of the effects have rarely been examined. We documented the combined effect of delayed metamorpho- sis and sub-lethal exposure of larvae to low-salinity stress on depletion of energy reserves, meta- morphic success and juvenile growth rates for the estuarine barnacle Balanus amphitrite, both in the laboratory and in field outplant experiments. Acute (24 h) exposure of 0 d old cyprids to low salinity (10 psu) dramatically reduced juvenile growth rates (measured as basal diameter, dry weight and ash-free dry weight) for the first 5 d after metamorphosis, in both laboratory and field conditions, and the effects were similar to those resulting from delayed metamorphosis. There was an interaction between delayed metamorphosis and salinity effects on juvenile tissue growth rates. To determine how the negative effects of delayed metamorphosis observed in the laboratory might be modulated in the field under different nutrient conditions, we also outplanted juveniles from cyprids that had delayed metamorphosis for either 0 d (control) or 4 d at 2 intertidal sites that experienced different nutrient conditions. Surprisingly, surplus food availability in the juvenile habitat could not entirely compensate for the negative effects imposed by delayed metamorphosis on juvenile growth rates. Additionally, we examined whether or not the detrimental effects of delayed metamorphosis on growth were mediated through effects on juvenile feeding rates and feeding appendage (cirri) mor- phology. Interestingly, although 2 out of the 3 juvenile feeding cirri lengths were shorter than expected in delayed individuals, mean juvenile filtration rate was significantly higher in delayed individuals than in control individuals. Overall, our results emphasise that acute exposure of compe- tent larvae to low-salinity stress negatively affects juvenile growth rates as severely as delayed meta- morphosis, which may ultimately influence juvenile and adult population dynamics in the field. The cause of reduced post-metamorphic performance in young barnacles remains unclear, but does not seem to be caused by reduced capacity for feeding.

The possible importance of larval settlement inhibition by Fucus vesiculosus in explain- ing the low abundance of the barnacle Balanus improvisus on algal fronds compared to adjacent rock was investigated by a combination of field and laboratory studies. A field survey showed that the abundance of adult barnacles was significantly lower on F. vesiculosus than on rocks and settlement preference experiments strongly suggested that settlement avoidance for algal fronds during the natural settlement season was chemically mediated. By using a multi-step approach, whereby both exuded and surface associated metabolites were tested on larval settlement, we further investigated the possible existence of chemical antifouling processes in F. vesiculosus. For exuded metabolites, seawater was conditioned for 1 and 3 h with both submerged and desiccated algae. Surface extracts at natural concentrations and seawater conditioned with submerged algae had no significant effect on larval settlement, although there was a tendency for reduced settlement in the 3 h samples. Sea- water conditioned with desiccated algae consistently inhibited settlement and chemical analyses showed that the mean phlorotannin level in the water was 31.5 µg ml -1 , which is ca. 30× higher than concentrations previously shown to inhibit barnacle settlement. Seawater conditioned with sub- merged algae had a mean phlorotannin content of 0.85 µg ml -1 and in samples conditioned for 3 h there was a strong and significant negative relationship between phlorotannin concentration and larval settlement, suggesting that the phlorotannin concentration varied around a level inhibitory to barnacle settlement. Altogether, the results from the laboratory experiments strongly suggested that the low larval preference for algae was caused by exudation of waterborne metabolites, primarily phlorotannins. Finally, field measurements showed that phlorotannins exuded from F. vesiculosus can under natural conditions reach concentrations inhibitory to settlement of barnacle larvae.

This study examined the attachment response of Balanus amphitrite larvae to bacteria- dominated biofilms originating from 4 sites of varying environmental conditions in the intertidal region of subtropical Hong Kong waters and during 2 seasons (winter and summer), under both laboratory and field conditions. Using multiple fingerprinting techniques (terminal restriction frag- ment length polymorphism, denaturing gradient gel electrophoresis and fluorescence in situ hybri- dization), we observed differences in the bacterial community composition of biofilms originating from the 4 sites. These biofilm samples were used to study the linkage between spatial changes in bacterial communities of biofilms and larval choice at the time of attachment. It was hypothesized that cyprids can distinguish biofilms originating from habitats that support higher recruitment. Both laboratory and field multiple-choice bioassays demonstrated that cyprids preferred to attach on biofilms originating from habitats where recruitment, juvenile growth and survival were the highest, thereby accepting the hypothesis proposed. This study did not identify particular bacterial species or groups in biofilms that attract or inhibit larval attachment, but we could correlate site-specific varia- tions in bacterial community composition with larval choice, whereas bacterial abundance in biofilms was less important in this regard. Overall, this study highlights the significance of site-specific variation in biofilms on larval recruitment and demonstrated the discriminative behavior of barnacle larvae to biofilms originating from contrasting environments in the intertidal region. Thus, attach- ment cues from biofilms may also play a significant role in generating spatial variation in larval recruitment.

A field experiment was used to assess the relationship between the physiological condition of barnacle larvae and habitat selectivity at settlement. Three experiments were carried out on methacrylate (Plexiglas) disks precolonized with biofilms of different ages (0, 7, 14 and 21 d) that were arranged within 3 blocks and placed at 3 intertidal positions correspond- ing to the upper limit, mid zone and lower limit of the adult Balanus amphitrite (Darwin) habitat. To limit the effect of post-settlement mortality, each experiment was conducted over 2 full tidal cycles only. During these periods, larvae in the water column were sampled and abundance estimated at each high tide at the depths corresponding to the 3 intertidal positions. Lar- val physiological condition was assessed using lipid classes analyzed by liquid chromatography. Ratios of total triacylglycerols to sterol content (TAG/ST) were used to quantify the energy status of larvae. Our results indicate that the cyprids' TAG levels reflect the level of settlement success independent of cyprid abundance in the water column. Intertidal position and biofilm char- acteristics were significant factors determining the settlement success with respect to cyprid energy con- tent. The number and the energy content of settling cyprids were maximal on clean surfaces and decreased gradually with the degree of biofilm precolonization. Our study shows (1) the importance of lipid levels, particularly TAG, in habitat discrimination at small temporal scales in B. amphitrite cyprids and (2) that larval energy content is a critical variable for under- standing benthic-pelagic coupling.

Microbial communities of the sponges Callyspongia sp. from Hong Kong and Cally- spongia plicifera (Porifera: Demospongia) from the Bahamas were compared with each other and with those from reference substrata using a terminal restriction fragment length polymorphism (T-RFLP) analysis. The least number of bacterial ribotypes and bacterial isolates were retrieved from Bahamas reference and sponge surfaces, while the bacterial communities from Hong Kong Callyspongia sp. and reference surfaces were more diverse. Microbial communities from the 2 sponges were different from each other and from reference substrata. Gas chromatographic-mass spectrometric (GC-MS) analysis of dichloromethane extracts revealed that more than 60% of the compounds were similar in the 2 species Callyspongia sp. and C. plicifera, compared to the com- pounds of Halichondria spp. At tissue level (TL) concentrations, both sponge extracts predominantly inhibited the growth of bacteria from reference substrata. Multifactor ANOVA revealed that the source of bacteria (sponge surface, interior, or reference substrata), the geographic location of iso- lates (Hong Kong or the Bahamas), the sponge extract (from Callyspongia sp. or from C. plicifera), and combinations of these factors contributed significant effects in disc diffusion assay experiments. Sponge extracts at both TL concentrations and 10× dilutions were toxic to larvae of the polychaete Hydroides elegans and the barnacle Balanus amphitrite. Our results suggest that the 2 congeneric sponges Callyspongia spp. from different biogeographic regions have different bacterial associates, while producing relatively similar secondary metabolites. It remains to be explored whether differ- ences in sponge-associated bacterial communities will also hold for other congeneric sponge species from different regions.

Using factorial laboratory and field-transplant experiments, we have examined the effects of cyprid energy reserve and habitat on the performance of juvenile Balanus amphitrite. The cyprid energy reserve (quantified as total lipid content) increased linearly with increasing food (the diatom Chaetoceros gracilis) concentrations, i.e. cyprids from nauplii fed the highest (1 × 10 6 cells ml -1 ), high (5 × 10 5 cells ml -1 ), low (1 × 10 5 cells ml -1 ) and the lowest (5 × 10 4 cells ml -1 ) food concen- trations had the highest (0.81 µg cyprid -1 ), high (0.65 µg cyprid -1 ), low (0.45 µg cyprid -1 ) and the lowest (0.18 µg cyprid -1 ) energy reserves, respectively. These cyprids were induced to metamorphose in the laboratory and then transplanted to 4 field sites differing in food availability. Their perfor- mance (measured as growth and survival) was assessed on Day 6. Regardless of the site, both survival and growth were lower for juveniles obtained from cyprids with the lowest energy reserve compared with juveniles obtained from cyprids with low, high and the highest energy reserves. However, the magnitude of the effect of cyprid energy reserve on growth varied among sites. For example, juve- niles obtained from cyprids with the lowest energy reserve grew larger (~2- to 3-fold) at sites with a high chlorophyll a and low C/N ratio (indicators of surplus and high-quality food) than their siblings at other sites. This was interpreted as the result of the interaction between the negative effects of the lowest energy reserve in cyprids and the positive effect of surplus food. Adverse effects of the lowest energy reserve in cyprids on juvenile growth, however, could not be completely compensated by sur- plus food. Regardless of the cyprid energy reserve, juvenile growth (not survival) increased as chloro- phyll a increased. This study suggests that the in situ growth of juvenile barnacles may depend on both independent and interactive effects of cyprid energy reserve (larval nutritional condition) and habitat.

This study investigated (1) the effects of salinity and temperature on the bacterial community composition of developing biofilms, and (2) the responses of marine invertebrate larvae (the polychaete Hydroides elegans and the barnacles Balanus amphitrite and B. trigonus) to these biofilms during settlement (i.e. attachment to a surface and metamorphosis into juveniles). Biofilms developed in a 3 × 3 array of salinity and temperature treatments resulted in different bacterial community compositions (revealed by DGGE and T-RFLP), bacterial densities and total biomasses. Larval settlement of B. amphitrite and B. trigonus was induced by biofilms developed at high temperatures (23 and 30°C), but was unaffected (B. amphitrite) or inhibited (B. trigonus) by those developed at a low temperature (16°C). The settlement response of these barnacles did not correlate with the biomass or the bacterial density of the biofilms, but did coincide with the marked differences in bacterial community composition between the biofilms developed at different temperatures. In contrast, larval settlement of H. elegans differed slightly among biofilms developed in different salinities, but not among those developed at different temperatures. This settlement response was moderately correlated with bacterial density but had no apparent relationship with bacterial community composition of the biofilms. Our results implied that the community composition and cell density of bacteria in biofilms, which can vary with local environmental conditions, may allow larvae of the 2 barnacles and H. elegans, respectively, to distinguish between habitats with different environmental conditions.

Natural biofilms constitute a complex network of microorganisms (bacteria, diatoms, protozoa, fungi) and their extracellular polymeric substances (EPS), which influence settlement in benthic invertebrates. We investigated the influence of diatom (Bacillariophyceae) films and EPS (>1000 molecular weight) on metamorphosis in the acorn barnacle Balanus amphitrite Darwin, a dominant fouling organism, using axenic and non-axenic films, and free and biofilm EPS, of 5 species of pennate diatoms: Amphora coffeaeformis and A. rostrata, and Navicula transitans var. derasa f. delicatula, N. crucicula and N. subinflata. Fourier-transformed infrared spectroscopy (FTIR) spectra revealed that the EPS produced by diatoms are of similar nature; however, the proportion of mono- saccharides varies with species, indicating a potential role for influencing larval metamorphosis. Free EPS of axenic diatoms had no effect, whereas biofilm EPS induced larvae to metamorphose. Amphora spp. produced more biofilm EPS, but had relatively weaker effects than Navicula spp. Axenic diatom films also facilitated larval metamorphosis (depending on species and cell density) in some cases, suggesting that the cues provided by the diatoms themselves can also mediate invertebrate larval metamorphosis in the absence of microbial films. Non-axenic diatom films (which had higher cell densities) and biofilm EPS promoted metamorphosis to a greater degree than those of axenic diatoms. Enhancement of metamorphosis depended on diatom species and on their density in the films, as well as on the composition of their EPS. Differential responses of barnacle larvae to different diatom species and their EPS indicate that each diatom species provides a different set of physico-chemical signals to settling larvae.

This study investigated whether the nature of the binding of the trace metals cadmium, silver and zinc accumulated by phytoplankton can affect their subsequent assimilation efficiencies (AE) in 3 filter-feeding benthic invertebrates, the green mussel Perna viridis, the clam Ruditapes philippinarum and the barnacle Balanus amphitrite. Seven phytoplankton species were chosen from a wide systematic range to ensure large differences in the partitioning of their accumulated trace metals into 3 fractions: (1) exchangeable metal adsorbed on the outside of the cells, as defined by extraction with the chelating agent 8-hydroxyquinoline-5-sulphonate; (2) incorporated metal that is in a soluble form; and (3) insoluble incorporated metal. There were few significant correlations between AE and the percentage of phytoplankton metal incorporated into any 1 fraction or combination of fractions. There is no support, therefore, for a generalised conclusion that any of the 3 fractions isolated represents the sole form of phytoplankton metal that is bioavailable for trophic transfer to a herbivore. Even trace metals bound to the insoluble fraction in phytoplankton may be bioavailable to herbivores. Furthermore, there were no consistent effects of phytoplankton concentration on metal AEs in 1 of the herbivores-P. viridis. There was no evidence that the AE of any of the 3 trace metals was changed when the herbivores were feeding on the phytoplankton species (Thalassiosira weissflogii) on which they were fed during acclimation.

Many marine invertebrate larvae have the capacity to reject or accept settlement sites based on a broad range of cues. Species-specific settlement responses to different cues are often inferred from final settlement choice in the field. Little is known about species-specific larval behav- iour in response to different cues and, in particular, how the behaviour is linked to final settlement. Rejection of unsuitable substrata may be an important driving force that leads to aggregated settle- ment patterns. This study examines rejection responses in relation to surface attractiveness for set- tlement under field and laboratory conditions in the barnacle Balanus improvisus. The attractiveness for settlement was manipulated by varying surface texture in combination with crude extract from conspecific adults. Active rejection behaviour was examined as a function of surface texture and con- specific pheromones in the field and then related to behavioural responses under static and flowing conditions in the laboratory. Recruitment was heavily reduced on substrata with ribbed microtexture compared to smooth substrata and unaffected by crude extract from conspecific adults. On average, 28% of the cyprids that encountered smooth settlement panels recruited. The proportion of cyprids recruiting on 2 microtextured substrata after encounter was 5 and 1% respectively. In behavioural experiments cyprids showed higher motion speed and dispersal rate on textured substrata, which indicated less exploratory behaviour than on smooth substrata, while an addition of conspecific extract increased intensities of surface exploration on all types of substrata. Flume experiments fur- ther demonstrated that cyprids are more prone to leave textured substrata and that the rejection rate was independent of conspecific extract. This work emphasises the role of larval behaviour as a poten- tially powerful mechanism determining final recruitment pattern. It is concluded that the choice of settlement site is an important factor in the settlement process of B. improvisus, and the results sug- gest that surface topography may be a stronger cue for settlement than chemical attraction by con- specific adults in this species. This study presents an example whereby rejection of unsuitable sub- strata leads to an increased larval pool on adjacent substrata that are suitable for settlement, and indicates that this process may drive aggregated settlement in the barnacle B. improvisus.

The relationship between bacterial community profile in biofilm and attachment of the acorn barnacle Balanus amphitrite was investigated using a double-dish choice larval attachment bioassay and the DNA fingerprinting technique T-RFLP (terminal-restriction fragment length poly- morphism). Biofilms for bioassays were either developed at 3 intertidal heights (i.e. high, mid and low) for 6 d or at the mid-intertidal height for 3 to 12 d. A clear distinction among biofilm communi- ties at the 3 intertidal heights was revealed in the bacterial community profiles (determined by T- RFLP), biomass (determined by total organic carbon analysis), and abundance of bacteria and diatoms. Overall, cyprids of B. amphitrite preferred intertidal biofilms (i.e. 6 d old) over unfilmed sur- faces for attachment. Moreover, cyprids also preferred to attach on biofilms of mid-intertidal height over high-intertidal or subtidal heights. There was no correlation between cypris attachment and any of the 3 biofilm attributes (i.e. biomass, abundance of bacteria and diatoms). Therefore, it was con- cluded that changes in the bacterial community profile in the biofilm affect the attractiveness of the biofilm to barnacle larvae and, thus, may determine the behavior (accepting or rejecting a surface) of settling larvae. We hypothesize that the temporal and spatial changes in the microbial community profile lead to the temporal and spatial recruitment pattern of marine invertebrates at a microscopic scale.

A common littoral barnacle, Balanus glandula Darwin, 1854 (Crustacea, Cirripedia) from the west coast of North America has become established in the mid-littoral zone of harbors on the Pacific side of northern Honshu, Japan. The new immigrant occupies space in the lower half of the Chthamalus spp. and Balanus albicostatus zones, and in the upper fringe of the Semibalanus cariosus zone. However, the species has not been detected in harbors along the Sea of Japan, south of the Ozika Peninsula, northern Honshu, and to the north and east of Kushiro, Hokkaido. Considering its present distribution, shell sizes, and abundance, it is most likely that it was introduced into 3 separate port areas in NE Honshu, Japan, during the last 20 to 40 yr, probably by anthropogenic activities. It was apparently also introduced to the La Plata estuary, Argentina, during the same period. B. glandula is becoming a dominant littoral zone species, especially in embayments, largely at the expense of the tropical endemic B. albicostatus in the upper littoral fringe. Taking into account the biogeographical distribution of common Japanese barnacles in relation to water temperature, the future distribution of the newly introduced species is considered.

Gregarious settlement in barnacles has been related to the settlement-inducing compounds from adult conspecifics, bacteria in the biofilms, and their interaction. Elucidation of larval settlement cues from these sources is limited. The effectiveness of larval settlement cues under different environmental conditions (salinity, temperature) needs evaluation. Pseudomonas aeruginosa, a bacterium isolated from the shell surface of Balanus amphitrite Darwin, was used as a candidate. The influence of bacterial film, culture supernatant and its molecular-weight fractions, and bacterial extract was investigated along with the conspecific adult extract (AE). The influence of culture supernatants and exopolysaccharides obtained from the bacterium cultivated in different nutrient media, effectiveness of leachants and adsorbed (surface-bound) compounds on the metamorphosis of cyprids of B. amphitrite was also assessed. The influence of P. aeruginosa on cyprid metamorphosis varied with salinity and temperature. The differences were not significant as the film and the cyprids aged. When the bacterial film was examined in the presence of an active substance (agonist) such as AE, metamorphosis was facilitated, suggesting the role of competitive antagonism in cue perception. The higher molecular-weight fraction of the bacterial-culture supernatant was inductive at higher salinity. Conversely, the lower molecular-weight fraction of the culture supernatant showed maximum inhibition when the adsorbed (surface-bound) compounds were assessed along with the leachants. Bacterial extract showed the presence of ketonic compounds, and its influence differed with salinity. The inhibitory effect of the extract was nullified in the presence of AE. When the extract was examined in the presence of leachants, a 2-fold increase in the metamorphosis rates was evident where only surface-bound components were inhibitory. Fourier transformed infrared spectroscopy analysis revealed that bacteria grown in different nutrient media yielded culture supernatants with different chemical composition, thus altering their ability to induce metamorphosis of cyprids. Maximum inducement was provoked by the culture supernatant obtained from semi-solid culture, and this positive effect was protein concentration dependent. The exopolysaccharides obtained from bacteria grown in basal salt solution facilitated metamorphosis similar to that of the bacterial film and AE. The response of the cyprids to bacteria and its products seems to be regulated by both contact chemoreception and olfaction, depending on the properties of the settlement-inducing compounds. The need to characterize and distinguish the receptors, which act via different signaling systems on a particular settlement cue, may be a step ahead to resolve the complexities of invertebrate larval recruitment.

Both surface texture and littorinid grazing are known to influence the establishment of many shallow-water benthic hard-bottom communities. However, the effects of these factors and their interactions have not yet been investigated in a quantifiable manner. This investigation aims to assess the interactive effect of both factors in a strictly standardized manner. Natural recruitment by diatoms, the barnacle Balanus improvisus and the tube-building polychaete Polydora sp. was monitored under a 2-factorial treatment: grazing by the periwinkle Littorina littorea (Factor 1; 2 levels: I or no snails per plate) on artificial recruitment plates of different initial surface rugosities (Factor 2; 5 levels: smooth, 0.1, 0.5, 1 and 5 mm rugosity elements), In the absence of grazers, barnacle recruitment decreased with increasing initial rugosity, polychaete recruitment peaked at intermediate rugosities, and diatoms exhibited contrasting recruitment patterns in an in vitro and an in situ experiment. When preferred recruitment sites coincided for Polydora and B. improvisus, a competition for space could be inferred from a negative correlation between the 2 species. However, when the overlap of requirements weakened on the 5 rugosities, the relationship was positive, but was not statistically significant. Grazing efficiency by L. littorea depended on initial rugosity, generally showing minimum values on intermediate rugosities which is attributable to a mismatch between radula dimensions and surface structures in these rugosity classes. Additionally, grazing effects tended to increase with higher prey densities. As all factors-initial rugosity, grazing, colonizer species-interact with each other, the outcome of recruitment under combined factors is difficult to predict from single factor effects.

Assimilation from the diet is established as a key factor in the accumulation of very high trace metal concentrations by the barnacle Elminius modestus. Assimilation efficiencies (AE) of Cd, Cr, Se and Zn were measured from a diet of different phytoplankton (2 diatoms, a dinoflagellate, a prasinophyte and a chlorophyte) and of zooplankton (the copepod Acartia spinicauda with metal accumulated from solution or diet). AEs of Cd, Se and Zn (but not Cr) varied greatly with phyto- plankton type, and for Cd and Se the AE was correlated with the percentage of metal burden held in the cytoplasm of the phytoplankton diet. AE was generally higher from the zooplankton diet than from a phytoplankton diet. Variation in AEs of Cd, Se and Zn was not explained by any correlation with the percentage of copepod metal burden held in the soft tissues, nor was there variation for either Cd or Zn according to whether the copepod prey had accumulated metal from dissolved or food sources. Comparisons of the assimilation and efflux of accumulated metals by the archaeo- balanid barnacle E. modestus and literature data for (phylogenetically younger) balanid species of the genus Balanus indicate some differences in digestive physiology of barnacles from the 2 families, tending towards higher AEs in the balanids. Modeling of the accumulation of Cd and Zn by E. mod- estus predicts that for each metal >97% of accumulated metal has been derived from dietary inges- tion. The dominance of dietary ingestion in trace metal accumulation is a function of the conspicu- ously high assimilation efficiencies and high ingestion activity of barnacles. This study adds to the small but growing list of examples highlighting the significance of trophic transfer in metal accumu- lation by aquatic invertebrates.

Epibiotic assemblages have been shown previously to differ between pontoons and rocky reefs. This may occur for a variety of reasons, one of which is that pontoons move, whereas reefs do not. Effects of movement of the substratum are particularly pertinent to studies involving settlement panels because these experimental units are often suspended from pontoons such that they can move up and down, or attached to ropes such that they can rotate. The development of epibiotic assemblages on panels that were fixed, moved up and down, or rotated, was studied to test hypotheses about the effects of movement on sessile organisms. The covers of barnacles, sponges and ascidians increased greatly with increasing movement and/or rotation of the substratum, whereas species of red and brown foliose algae and tubiculous polychaetes generally decreased with increasing movement and/or rotation of the substratum. In general, assemblages on panels that moved up and down were most similar to those on rotating panels. One of the most obvious differences was the 2 to 3-fold greater biomass on rotating panels compared to fixed panels, which was attributed to the greater abundance of barnacles. It was concluded that, although movement of a surface can influence the composition of assemblages, this factor on its own may explain only some of the differences reported between pontoons and reefs. Furthermore, the method of deployment of settlement panels was shown to have great effects on the types of assemblages that develop. Differences in water flow were proposed to explain some of the observed patterns, but this idea needs to be investigated further.

Behavior of laboratory-reared larvae of the bar- nacle Balanus amphitrite was examined in Beaufort. North Carolina (USA), relation to their physiological condition. Cyprid substratum exploration was monitored by means of video endoscopy using various experimental surface types (clean, biofilm, 1 and 2 wk fouled) and 2 water flow regirnes (still water and ca 5 cm s-') at room temperature (21°C). We used the triacylglyceroYDNA (TAG/DNA) ratio of small batches of larvae as a measure of physiologicai condition in 0 to 12 d old cyprids. The physiologicai condition of cyprids decreased significantly with age (p < 0.001), ratios severely dropping between 5 and 8 d. Although exploration behavior did not show much variation with age, the overail number of active exploring cyprids appeared to be age dependent. Additionally, the relationship between surface exploration behavior and age also appeared to vary with substratum type as weil as flow rate. For example, fewer young cyprids (0 to 5 d old) explored unfavorable substrata (clean and biofilm treatments) than older ones in still water. Exploration re- sponses, however, appeared to differ in relation to flow regirnes (still vs moving water trials). Time spent by cyprids on surfaces before returning to the water column (non- exploratory behavior) appeared to vary in relation to age and substratum type in stiu water trials. Duration of exploration and distance explored by cyprids in flow generally peaked with cyprids from the 3 d cohort. Overall, our results showed that habitat selection in barnacle cyprids results from a com- plex relationship involving substratum type, hydrodynamics and larval age (i.e. physiological condition and competency).

Settlement, attachment, and metamorphosis of larvae of marine invertebrates is extremely variable in the laboratory and in nature. The relative contributions of genetic, ontogenetic and environmental effects to this variation are not well understood. We tested for genetic, maternal and ontogenetic effects on permanent attachment of cyprids of the barnacle Balanus amphitrite (Darwin). Larvae were assayed for attachment in response to a control treatment and 3 artificial inducers: 20-hydroxyecdysone (20-HE), 3-isobutyl-1-methylxanthine (IBMX), and phorbol 12,13- dibutyrate (PDB). We observed strong family effects on attachment in the control, 20-HE, and IBMX treatments, suggesting that attachment in response to these treatments is influenced by genetic or maternal factors. In addition, attachment responses were correlated, indicating that the same genetic or maternal factors may affect attachment in each treatment. Family effects on attachment in the control treatments decreased with cypris age, suggesting the loss of genetically- or maternally- influenced discriminatory behavior in older larvae. The mechanisms underlying genetic or maternal effects on attachment may act via the external receptors and signal-transduction pathways regulat- ing larval responses. Alternatively, there may be variation in larval 'vigor' or the rate at which cyprids attain metamorphic competence.

Tubes constructed of different materials were used to examine the role of hydrodynam- ics and surface characteristics, as measured by contact angles, on larval attachment of marine poly- chaetes, bryozoans and barnacles. Tubes (10 mm inner diameter) of 7 different materials, ranging in contact angle from 25° to 113°, were used. Laboratory and field experiments were conducted using a range of flow rates from 1.7 to 84.9 cm s -1 . For all 3 taxa of organisms, the results of field experiments were similar to those of the laboratory experiments. Larval settlement of the polychaete Hydroides elegans and the bryozoan Bugula neritina was the highest at a flow rate of  2.1 cm s -1 , and decreased when the flow rate was increased. H. elegans larvae settled within the narrowest range of flow rates (1.7 to 9.1 cm s -1 ), B. neritina larvae settled over the widest range of flow rates (1.7 to 53.0 cm s -1 ), and barnacle larvae (Balanus spp.) settled in an intermediate range of flow rates (4.2 to 21.2 cm s -1 ). Bar- nacle larvae did not settle when the flow rate was 21.2 cm s -1 , and the highest settlement was recorded at the flow rate of 10.6 cm s -1 . Barnacle larvae showed the highest settlement in glass tubes (smallest contact angle) and the lowest in Teflon tubes (largest contact angle), while B. neritina showed the highest larval settlement in Teflon tubes and the lowest settlement in glass tubes. H. ele- gans also showed the highest larval settlement in Teflon tubes, but the lowest settlement in polyurethane and polyethylene tubes. Biofilm formation changed the contact angles of all the tubes, the degree of change varied among the 7 types of tubes. Bacterial densities of biofilm on the 7 types of tubes were significantly different and were not correlated with the initial contact angles of tubes. The bacterial densities on the Teflon tubes were the same as on glass tubes. Our results suggest a very complex interaction among substratum characteristics, flow rates, and larval settlement behav- ior and indicate the necessity for considering the interaction when one interprets laboratory bioassay data on larval settlement of marine invertebrates.

A set of 3 field experiments lasting 24 h was conducted during April 1998 at the Duke University Marine Laboratory (Beaufort, North Carolina, USA) to: (1) assess the influence of larval supply, intertidal height, quantity and quality of biofilm and age of the larvae on the settlement of Balanus amphitrite Darwin and (2) examine the correspondence between small-scale planktonic distribution of larvae, the initial spatial pattern of newly settled larvae and the vertical distribution of adult barnacles. Precolonized methacrylate (Plexiglas) disks, arranged within 3 blocks and established so as to eliminate edge effects within 3 large experimental panels, were placed at 3 predetermined tidal heights (High, Medium, Low) corresponding to the upper limit, modal zone and the lower limit of adults of B. amphitrite. Split-split-plot ANOVAs were performed on densities of newly attached larvae (metamorphosis not completed) to test their habitat selection behavior to surfaces which had been precolonized by microbiota (bacteria and diatoms) at 3 heights (origin factor) for 0, 7, 14 or 21 d (age factor). The physical environment (salinity, temperature, current flow) was stable and comparable during the 3 experiments. B. amphitrite cyprids were uniformly distributed in the water column. Larval supply was poorly correlated with the intensity of settlement over the 1 wk experimental period. In fact, the same larval supply could induce either high (4×) or low (1×) settlement after 2 tidal cycles, and, inversely, similar settlement intensities were associated with planktonic larval abundance varying significantly at 2 d intervals (109 to 171 cyprids 923 1 -1 ). Settlement was homogeneous on each experimental unit (no significant block effect). Tidal height, however, was a significant factor in determining the vertical patterns of newly settled larvae during the first experiment where larvae were abundant but not during subsequent experiments for which fewer larvae were collected. The degree of microbial precolonization was the main parameter affecting the settlement of B. amphitrite. For the first 2 experiments, 'weighed cyprid settlement' significantly decreased as the age of the biofilm increased, revealing a strong preference of settlers for clean surfaces and avoidance of biofouled surfaces of all intertidal origins. Further analysis of biofilm samples showed that free-space availability in the microbial film and bacterial densities were significantly inversely correlated to settlement intensity. Moreover, settlement to 'favorable' substrata decreased by nearly 1/2 during our experimental period, suggesting changes in the selectivity of settling larvae. Our experiments confirm the role of larval supply in determining the vertical intertidal distribution of adults of B. amphitrite, but the short-term variability in the larval supply/settlement coupling observed over a 1 wk period must be integrated in models of recruitment dynamics of barnacles.

We determined the assimilation efficiencies (AEs) and efflux rate constants of Cd, Cr, and Se in barnacles Balanus amphitrite feeding on diverse planktonic prey. The measured AEs of Cd, Cr, and Se for zooplankton prey (brine shrimp Artemia salina larvae and copepods Canthocalanus pauper and Temora turbinata) were 53 to 88, 32 to 59, and 63 to 76%, respectively, and for diatom diets were 35 to 86, 22 to 26, and 79%, respectively. Distribution of metals in the soft tissues of zooplankton could not account for the variability of AE for each metal, but did explain the variability of AE among different metals. Metal distribution in the cytoplasm of diatoms determined the variability of AEs among metals. There was a significant correlation between Cd and Zn in AE and efflux rate constant. No relationship in AE or efflux rate was found for the other metals. The efflux rate constants in B. amphitrite were 0.007, 0.020, and 0.014 d -1 for Cd, Cr, and Se, respectively. Cd concentrations in barnacles, predicted by a simple bioenergetic-based kinetic model, were comparable to the actual concentrations measured in the field when phytoplankton was considered as the primary Cd source. Cd concentrations predicted by the model were much higher than the field measurements when copepods were assumed as the sole food source for barnacles. Trophic transfer appeared to be responsible for Cd accumulation in barnacles. Many biological and geochemical processes can affect metal accumulation in barnacles. Our study demonstrated that biological processes must be considered to interpret metal concentrations in barnacles when the barnacles are used to monitor coastal contamination.

We conducted 4 experiments to study the effects of salinity and temperature on the barnacle Balanus amphitrite amphitrite Darwin, with particular focus on the effects of stress experienced in one life-stage on the performance of the next life-stage. At 15°C, typical winter water temperature in Hong Kong, larvae exhibited low survivorship, adults molted infrequently, and only a low percentage of individuals had developing ovaries and embryos. However, at 30°C, typical summertime temperature in Hong Kong, larvae developed rapidly, survivorship was high, adults molted frequently, and a high percentage of individuals had developing ovaries and embryos. These results suggest that low winter temperature may be a limiting factor responsible for cessation of recruitment, whereas high summer temperature is unlikely to be the cause for the decline in recruitment. Salinity produced significant detrimental effects on both survival and development at ≤10‰. In the 15 to 35‰ S range, however, none of the stages tested exhibited signs of stress. Salinity is a limiting factor for the survival and development of B. a. amphitrite in Hong Kong only during mid-summer when salinity in the surface water can drop to below 10‰. Exposing embryos to different salinities produced differential effects on larvae. For larvae cultured at 10‰ S, both survivorship and time of development were independent of the salinity that the embryos had experienced; for larvae cultured at 15 and 35‰ S, exposing embryos to 10‰ S led to lower larval survivorship and longer larval development times. Exposing cypris larvae to 10‰ S did not alter juvenile growth but did result in lower survivorship. Osmotic stress experienced in one life-stage can be passed over to the next life-stage. In bioassays involving the use of B. a. amphitrite, results of the tested life-stage may be affected by stress experienced in a previous life-stage.

In the marine environment, competent larvae of sessile invertebrates are influenced by water flow and a variety of biological, chemical and physical cues. Most research has focused on how these biotic and abiotic factors influence where individual larvae ultimately settle. Much less is known about post-contact exploration prior to metamorphosis. This is, in part, due to limitations associated with directly observing small larvae (100 to 500 μm) in flowing seawater. A study was conducted in Beaufort, North Carolina, USA to understand how larvae of the barnacle Balanus amphitrite and the bryozoan Bugula neritina respond to a variety of flow rates (0, 1.3, 6.1 and 8.3 cm s -1 ) and surface types (clean, biofilmed, 1 and 2 wk fouled). Larval behavior was studied by means of endoscopy in a running-seawater chamber. Larval movements were observed at 30 frames s -1 for individuals that remained in contact with surfaces from

Intake pipes are unique habitats that provide an experimental environment for studying the role of hydrodynamics and larval settlement in community development. In this study, we used 5 and 10 mm (inner diameter) tubes as experimental models to mimic intake pipe environments to study the settlement patterns at different flow rates of the bryozoan Bugula neritina, the polychaete Hydroides elegans, and barnacles of the genus Balanus. Clean, unfilmed PVC tubes were used to examine settlement of B. neritina. PVC tubes, on which biofilm had been allowed to develop for 48 h, were used for studying attachment of H. elegans, while clean tubes were used for investigating settlement of Balanus spp. In all but very low flows, the flow velocity and Reynolds number were poorly correlated with patterns of larval settlement. A hydrodynamic measure, which is theoretically independent of the size of the tube, the so called 'velocity gradient', was well correlated with the highest settlement for all 3 species. Comparisons of results from field and laboratory experiments reveal slight differences. Settlement of the small elongate larvae of H. elegans was offset to higher shear values in the tubes of smaller diameters. Flow velocities for the highest settlement were from 1 to 3 cm s-1 for B. neritina and H. elegans, and from 3 to 15 cm s -1 for barnacles. Velocity gradients for the highest settlement of tubeworms and bryozoans ranged from 8 to 25 s -1 , while those for barnacles ranged from 50 to 120 s -1 . Barnacles, as reported previously by other authors, did not settle in high numbers when velocity gradients were too low or too high. Barnacles did not settle at 400 s-1. H. elegans had the narrowest range of settlement in relation to flow as settlement of this species was the highest from 8 to 20 s -1 and hardly occurred above 200 s -1 . In general, larval settlement in response to flow is species specific. We suggest that this species specificity is related to larval morphology, swimming ability, and behavior.

Recruitment of the barnacle Balanus amphitrite into a macrofouling community was examined in a semi-enclosed coastal environment (Hamana Bay, Japan), where water exchange with the adjacent open sea is very limited. The recruitment period of these barnacles was shorter than the period in which this species can breed and in which the cirripede larvae are present in the environment. Rearing of B. amphitrite larvae at different temperatures (15 to 30*C) and salinities (10, 20 and 300/00) revealed that its development is euryhaline. The influence of temperature was found to be greatest on the second instar. Mortality rates at 15*C temperature ranged from 43% (300/00 salinity) to 99% (100/00 salinity). Rearing experiments indicated that larvae released to the environment during autumn and winter failed to establish themselves in the macrofouling community because of adverse environmental conditions. These results also show that loss of planktonic larvae through starvation and misrouting may well be the main cause of reproductive loss.

Boards of CCA-treated wood, control (untreated) wood, and recycled plastic 'lumber' were placed into an estuary (Fort Pond, Southampton, NY, USA) in early May and examined monthly for abundance of species. One set of boards was left in the water continuously for 5 mo. Another set was renewed each month; after counting, organisms were removed and analyzed for dry weight and metal concentrations. A further set, placed in Accabonac Harbor, East Hampton, NY, in early July, compared CCA-treated wood that had not been impregnated well (green did not extend to inner portions), wood that had been well impregnated, and control wood. These boards were left in the water for 2 mo, and examined every 2 wk. Treated wood had a reduced number of species, lower diversity index, fewer barnacles (Balanus eburneus) and reduced growth of the barnacles that settled, fewer Membranipora and reduced growth of Membranipora colonies, fewer macroalgae, and fewer amphipods, compared to control wood and plastic The bryozoan Bugula turnta, however, was found at great density on the treated wood Biomass was lower on treated wood compared to control wood and plastic, and accumulations of Cu, Cr, and As were found Barnacles in~tially avoided settling on plastic, but after 1 mo exceeded those on control wood The communities in the 2 estuaries differed: e.g. at Accabonac species number was lower, there was less algae, bryozoans were absent, and barnacles were more dominant.