Grazing effects on nitrogen fixation in coral reef algal turfs

This study addressed whether grazing by the sea urchin Diadema antillarum influenced rates of nitrogen fixation by algal turf communities on Caribbean coral reefs. Because the turfs were nitrogen-limited, we also assessed whether newly-fixed nitrogen was important for supporting net primary productivity by the turfs. We measured acetylene reduction in turfs grown in treatments excluding or including D. antillarum in the presence of other herbivores at 3 m water depth on Tague Bay forereef, St. Croix, U.S. Virgin Islands. These were the first measurements of acetylene reduction on coral reefs under quasi-natural conditions of high water-flow and photosynthetic oxygen generation. Rates of acetylene reduction under these conditions were as high as any measured previously in coral reef communities (mean 7.6 nmol C₂H₄ cm⁻² h⁻¹). Algal turfs grazed by D. antillarum and other herbivores had chlorophyll-specific acetylene reduction rates up to three times higher than when D. antillarum was excluded. High rates of nitrogen fixation by the turfs were sufficient to meet <2% of the nitrogen required to support net chlorophyll-specific primary productivity over 24 h. Grazer-mediated increases in nitrogen fixation do not appear responsible for a parallel enhancement of net primary productivity. Algal turfs at this site must be dependent primarily on external sources of nitrogen. Received: 1 July 1997 / Accepted: 5 September 1997     

Nitrogen fixation (acetylene reduction) associated with macroalgae in a coral-reef community in the Bahamas

N₂ fixation (C₂H₂ reduction) was associated with several species of macroalgae on a coral reef near Grand Bahama Island. The highest rates were associated with Microdictyon sp. (Chlorophyceae) and Dictyota sp. (Phaeophyceae). Extensive mats of filamentous blue-green algae, not heterotrophic bacteria, were the N₂ fixing agents: in experiments with samples of Microdictyon sp., the activity was lightdependent and not stimulated by organic compounds under either aerobic or anaerobic conditions. Assays in situ, at 20 m depth, and on shipboard, gave similar rates of N₂ fixation; the cyanophytes presumably have pigment adaptations to function in blue light. The maximum rate of N₂ fixation, associated with Microdictyon sp., was 3.8 g N fixed g dry weight^-1 h^-1. Coral-reef communities flourish in nutrientimpoverished waters, and therefore any input of nitrogen is probably important in stabilizing such ecosystems.     

Nitrogen fixation in the rhizosphere of marine angiosperms

High rates of acetylene reduction were observed in systems containing excised rhizomes of the Caribbean marine angiosperms Thalassia testudinum, Syringodium filiforme and Diplanthera wrightii, and the temperate marine angiosperm Zostera marina. For 4 plant and plant-sediment systems the ratio of acetylene reduced/N₂ fixed varied from 2.6 to 4.6. For T. testudinum the estimated rates of nitrogen fixation are in agreement with estimated requirements of the plant for nitrogen. For a typical T. testudinum stand, N₂ fixation is estimated to be 100 to 500 kg N/hectare per year. Numbers of N₂-fixing bacteria in the rhizosphere sediments were roughly 50 to 300 times more abundant than those in the nonrhizosphere sediments, and in both types of sediments were of the same orders as the estimated numbers of heterotrophic aerobes.Canadian IBP Contribution No. 137.     

Nitrogen fixation on a coral reef

Acetylene reduction was used to assess nitrogen fixation on all major substrates at all major areas over a period of 1 to 6 yr (1980–1986) at One Tree Reef (southern Great Barrier Reef). Experiments using ¹⁵N₂ gave a ratio of 3.45:1.0 for C₂H₂ reduced:N₂ fixed. Acetylene reduction was largely light-dependent, saturated at 0.15 ml C₂H₂ per ml seawater, and linear over 6 h. High fixation was associated with two emergent cyanophyte associations, Calothrix crustacea and Scytonema hofmannii, of limited distribution. Subtidally, the major contribution to nitrogen fixation came from well-grazed limestone substrates with an epilithic algal community in the reef flat and patch reefs (3 to 15 nmol C₂H₄ cm^-2 h^-1). Similar substrates from the outer reef slope showed lower rates. Nitrogen fixation on beach rock, intertidal coral rubble, reef crest and lagoon sand was relatively small (0.3 to 1.0 nmol C₂H₄ cm^-2 h^-1). Seasonal changes in light-saturated rates were small, with slight reduction only in winter. Rates are also reported for experimental coral blocks (13 to 39 nmol cm^-2 h^-1) and for branching coral inside and outside territories of gardening damselfish (3 to 28 nmol cm^-2 h^-1). This work supports the hypothesis that the high nitrogen fixation on the reef flat and patch reefs of the lagoon (34 to 68 kg N ha^-1 yr^-1) is because these subtidal areas support highly disturbed communities with the greatest abundance of nitrogen-fixing cyanophyte algae. It is calculated from a budget of all areas that One Tree Reef has an annual nitrogen fixation rate of 8 to 16 kg N ha^-1 yr^-1.     

Aerobic nitrogenase activity measured as acetylene reduction in the marine non-heterocystous cyanobacterium Trichodesmium spp. grown under artificial conditions

Aerobic nitrogenase activity in the marine non-heterocystous cyanobacterium Trichodesmium spp. NIBB 1067, isolated off the Izu Peninsula, Japan in 1983 and grown under artificial conditions, was assayed by the acetylene reduction method. This strain exhibited acetylene reduction activity under aerobic conditions when cells had been grown in the medium free of combined nitrogen. Activity was markedly enhanced by light, and dependent on the growth phase being higher during the exponential growth phase and lower during the late linear and stationary growth phases. Since typical colony formation occurred during the last growth phase, the present results contradict the idea that N₂-fixation depends on colony formation. The photosynthesis inhibitor DCMU at 10^-6 M inhibited light-dependent acetylene reduction completely. Acetylene reduction by Trichodesmium spp. was tolerant of O₂ as strongly as that in the heterocystous cyanobacteria. Even at a partial pressure of oxygen (p_{O 2}) of 3 atm, the activity still remained as high as half of the maximum. It was almost under anaerobic conditions. Maximum activity was obtained at pO₂ of ca. 0.1 atm.     

High rates of nitrogen fixation on coral skeletons after predation by the crown of thorns starfish Acanthaster planci

At One Tree Reef, Great Barrier Reef, Australia, between 1983 and 1985, corals killed by the crown of thorns seastar Acanthaster planci L. gave rise to skeletons which were colonised rapidly by blue-green and other algae. For the next 3 to 9 mo these coral skeletons showed over three times more nitrogen fixation (acetylene reduction) than control substratum rates (9 to 32 nmol vs 3 to 10 nmol C₂H₂ cm^-2 h^-1, over all seasons). These values convert to relatively high annual fixation rates of 37 to 127 kg N ha^-1 yr^-1 but, at the low densities of A. planci on One Tree Reef (ca. 0.65 ha^-1), this has little impact on the total nitrogen fixation rate and, as a result, on the level of organic nitrogen in the system. However, it is suggested that on reefs subjected to high aggregations of a. planci such an effect would enhance the level of organic nitrogen and lead to greater primary and secondary production throughout the reef system.     

Nitrogen fixation (acetylene reduction) in the phyllosphere of Thalassia testudinum

N₂ fixation (C₂H₂ reduction) associated with the leaves of the sea grass Thalassia testudinum was investigated at 5 sites in South Florida (Biscayne Bay) and one site in the Bahamas (Bimini Harbor). Significant activities were correlated with the occurrence of a heterocystous blue-green alga (Calothrix sp.) on the leaves. C₂H₂ reduction was not stimulated by organic compounds, either aerobically or anaerobically in the light or dark. Therefore, other physiological types of microbes were not important in N₂ fixation. Diurnal and seasonal variations in N₂ fixation occurred, with maximal rates during the daytime and in the late spring and early summer. N₂ fixation was negligible at four stations in Biscayne Bay. At the fifth station, near Fowey Rock, about 5 kg N ha^-1 year^-1 was fixed. In the summer, the N₂ fixed per day (4–5 mg N m^-2) could provide 4 to 23% of the foliar productivity demands of T. testudinum at this site and the station in Bimini Harbor. N₂ fixation at the periphery of a sea-grass patch, near Fowey Rock, could provide 8 to 38% of the daily nitrogen requirement for leaf production, and thereby might compensate for a less effective trapping and recycling of nitrogen from dead leaves in such regions.     

Nitrogen fixation in the North Pacific Ocean

Nitrogen fixation in the euphotic zone of the ocean was measured by C₂H₂ reduction and ¹⁵N₂ incorporation associated with Trichodesmium sp. and also with Richelia intracellularis occurring within the cells of Rhizosolenia styliformis var. longispina, and R. cylindrus. The vertical distribution of N₂ fixation activity, N₂-fixing species, particulate matter and dissolved nutrients was measured. The effects of light intensity, sample concentration, length of incubation, and nutrient enrichment on the rates of C₂H₂ reduction were determined. Estimates of the importance of N₂ fixation in adding previously uncycled nitrogen to the euphotic zone are given.     

Use of the “Acetylene blockage” technique for assaying denitrification in a salt marsh

The acetylene blockage technique was evaluated for measurement of denitrification in salt-marsh sediments (near Halifax, Nova Scotia, Canada). N₂O in the gas phase of closed Spartina alterniflora marsh-sediment systems was analyzed with use of a thermal conductivity gas chromatograph sensitive to approximately 0.1 nmoles ml^-1 gas. No N₂O was detected for unfertilized sediment samples taken through the growing season and incubated in sealed buckets with 10% C₂H₂. For sediment samples amended with nitrate and for enrichments, initial rates of N₂O evolution were higher in the presence of 10% C₂H₂ than in the absence of C₂H₂, but after longterm incubation N₂O was consumed in some samples containing C₂H₂ as well as in samples without C₂H₂. In addition, total gaseous nitrogen (N₂ and N₂O) production in the absence of C₂H₂ was higher than in the presence of C₂H₂. Acetylene appears to be an inconsistent inhibitor of N₂O reduction in salt-marsh sediments. The usefulness of the acetylene-denitrification technique in this habitat is, therefore, questionable.     

Spatial variations in the N2O emissions and denitrification potential of riparian buffer strips in a contaminated urban river

Spatial variations in the N₂O emissions and denitrification potential of riparian buffer strips (RBS) in a polluted river were examined. The river received large pollutant inputs from urban runoff and wastewater discharge, resulting in impaired water quality in the river and downstream reservoir. The potential for nitrogen removal by RBS was evaluated by measuring in situ N₂O emission fluxes in static closed chambers and sediment denitrification potentials with acetylene inhibition techniques. The results showed that N₂O emission fluxes decreased from the upstream (16.39 μg/(m²·h)) to downstream (0.30 μg/(m²·h)) sites and from the water body to upland sites. The trend in decreasing N₂O emission fluxes in the downstream direction was mainly associated with sediment/soil textures (clay loam→sandy soil) and sediment/soil water contents and was also related to the vegetation along the RBS and nutrients in the sediments/soils. The correlation coefficient was highest (r=0.769) between the N₂O emission flux and sediment/soil water content. Sediment/soil denitrification potentials under N-amended and ambient conditions were higher (highest 32.86 mg/(kg·h)) for the upstream sites, which were consistent with in situ N₂O flux rates.     

Mineralization of particulate organic matter derived from coral-reef organisms in reef sediments of the Gulf of Aqaba

In situ and laboratory incubation experiments in a fringing reef in the Gulf of Aqaba were performed to study degradation rates of particulate organic matter in reef sediments. Coral mucus, clam eggs, and zooxanthellae were used as model particulate organic compounds for these experiments. Aerobic and anaerobic mineralization rates were calculated by dissolved inorganic carbon (DIC) and O₂ fluxes from the sediments under different particulate organic matter additions. Fast enhancement (approximately twofold) of O₂ and DIC fluxes were found with the addition of coral mucus and clam eggs compared with control incubations without addition. Most of the degradation is believed to have occurred anaerobically rather than aerobically (DIC:O₂ ratios were 4.3–28.1). Higher degradation rates of coral mucus and clam eggs were estimated in carbonate sediment than in silicate sediment (1.2–1.6-fold), which was attributed to the different physical and chemical properties of both sediments. Our study shows the significance of the reef sediment as a suitable site for microbial degradation of particulate organic material excreted from different reef community organisms. This may increase the regeneration of nutrients in the reef environment necessary to sustain high biological productivity.     

Seasonal variations in nitrogen-fixation (acetylene reduction) and sulphate-reduction rates in the rhizosphere of Zostera noltii: nitrogen fixation by sulphate-reducing bacteria

Nitrogen-fixation (acetylene reduction) rates were measured over an annual cycle in meadows of the seagrass Zostera noltii Hornem in the Bassin d'Arcachon, south-west France, between March 1994 and February 1995, using both slurry and whole-core techniques. Measured rates using the slurry technique consistently overestimated those determined on whole cores, probably due to the release of labile organic carbon sources as a result of root damage during preparation of the slurries. Thus, the whole-core technique may provide a more accurate estimate of in situ activity, since disturbance of physicochemical gradients of oxygen, sulphide, nutrients and the relationship between the plant roots and the rhizosphere microflora is minimised. Rates measured by the whole-core method were 1.8- to 4-fold greater (dependent upon season) in the light than those measured during dark incubations, indicating that organic carbon diffusing from the plant roots during photosynthesis was an important factor in regulating nitrogen fixation in the rhizosphere. Additions of sodium molybdate, a specific inhibitor of sulphate-reducing bacteria (SRB) inhibited acetylene-reduction activity by >80% as measured by both the slurry and whole-core techniques throughout the year, inferring that SRB were the dominant component of the nitrogen-fixing microflora. A mutualistic relationship between Z. noltii and nitrogen-fixing SRB in the rhizosphere, based on the exchange of organic carbon and fixed nitrogen is proposed. Acetylene- and sulphate-reduction rates showed distinct summer peaks which correlated with a reduced availability of ammonium in the sediment and the annual growth cycle of Z. noltii in the basin. Overall, these data indicate that acetylene reduction (nitrogen fixation) activity in the rhizosphere of Z. noltii was regulated both by the availability of organic carbon from the plant roots and maintenance of a low NH ₄ ⁺ concentration in the vicinity of the plant roots due to efficient assimilation of NH ₄ ⁺ by Z. noltii during the growth season. Nitrogenfixation rates determined from acetylene-reduction rates measured using the whole-core technique ranged from 0.1 to 7.3 mg N m^-2d^-1, depending on season, and were calculated to contribute between 0.4 and 1.1 g N m^-2yr^-1, or 6.3 to 12% of the annual fixed nitrogen requirement of Z. noltii.     

Effects of dissolved ammonium addition and host feeding with Artemia salina on photoacclimation of the hermatypic coral Stylophora pistillata

 Effects of nutrient treatments on photoacclimation of the hermatypic coral Stylophora pistillata (Esper) were studied. Studies on photoacclimation of colonies from different light regimes in the field were evaluated and used to design laboratory experiments. Coral colonies were collected in the Gulf of Eilat (Israel) from January to March 1993. Exterior branches of colonies from different depths (1 to 40 m) displayed different trends in production characteristics at reduced and very low levels of illumination. From 24 ± 3% to 12 ± 2% of incident surface photosynthetic active radiation (PAR_o), zooxanthella population density and chlorophyll a+c per 10⁶ zooxanthellae increased, a trend seen in the range of light levels optimal for coral growth (90 to 30% PAR_o). The P _max of CO₂ per 10⁶ zooxanthellae decreased, while P _max of CO₂ per 10³ polyps increased, indicating an increase in zooxanthella population density at low light levels. Proliferous zooxanthella frequency (PZF, a measure of zooxanthella division) declined significantly at light levels <18 ± 3% PAR_o. At the lowest levels of illumination (<5% PAR_o), zooxanthella population density decreased, as did the PZF; chl a+c per 10⁶ zooxanthellae was unchanged. In 28-d experiments, exterior coral branches from the upper surfaces of colonies from 3 m depth (65 ± 4% PAR_o) were incubated in aquaria under bright (80 to 90% PAR_o), reduced (20 to 30% PAR_o), and extremely low (2 to 4% PAR_o) light intensities. At each light intensity, the corals were maintained in three feeding treatments: sea water (SW); ammonium enriched SW (SW + N); SW with Artemia salina nauplii (SW + A). An increase in P _max of CO₂ per 10³ polyps was found in corals acclimated to reduced light (20 to 30% PAR_o) in nutrient-enriched SW, while in SW, where the increase in zooxanthella population density was smaller, it did not occur. Nutrient enrichments (SW + N at 2 to 4% PAR_o and SW + A at 20 to 30% PAR_o) increased zooxanthella population density, but had no effect on chl a+c per 10⁶ zooxanthellae. Acclimation for 14 d to reduced (10 to 20% PAR_o) and extremely low (1 to 3% PAR_o) light intensities shifted ¹⁴C photoassimilation into glycerol and other compounds (probably glycerides), rather than sugars. Both ammonium addition and feeding with Artemia salina nauplii resulted in an increase in photosynthetic assimilation of ¹⁴C into amino acids. We conclude that acclimation to reduced light consists of two processes: an increase in photosynthetic pigments and in zooxanthella population density. Both processes require nitrogen, the increase in zooxanthella population density needing more; this adaptation is therefore limited in nitrogen-poor sea water. Received: 19 June 1998 / Accepted: 13 June 2000     

Aerobic nitrogen fixation by the marine non-heterocystous cyanobacterium Trichodesmium (Oscillatoria) spp.: Its protective mechanism against oxygen

Nitrogen fixation (acetylene reduction) by the marine non-heterocystous cyanobacteria, Trichodesmium thiebautii and T. erythraeum, is sensitive to oxygen. Its sensitivity to oxygen was intensified when the colonies of T. thiebautii were disintegrated, but the separate trichomes yielded still retained the capacity for light dependent acetylene reduction. Trichodesmium colonies evolved hydrogen under argon in the light. The addition of carbon monoxide with DCMU [3-(3,4-dichlorophenyl)-1,1-dimethylurea] enhanced the rate of hydrogen evolution to approximately the same level as that of the maximum acetylene reduction on an electron basis. This probably results from the inhibition of the uptake hydrogenase. We propose that the uptake hydrogenase functions to protect nitrogenase from damage by oxygen.     

Atmospheric dinitrogen fixation by benthic communities of Tikehau Lagoon (Tuamotu Archipelago, French Polynesia) and its contribution to benthic primary production

Acetylene reduction rates were measured in lagoonal sediments, cyanobacterial mats and limestone surfaces between 1991 and 1995 at many sites, depths and seasons; all the studied substrata contained cyanobacteria. The acetylene reduction/¹⁵N₂ fixation ratio was measured for the different communities and varied between 1.8 and 4.8, depending on substratum. Fixation rates were 1.7 to 7 times higher during daylight compared to night-time rates. N₂ fixation rates ranged from 0.4 to 3.9 mg N m^-2 day^-1 for the lagoonal sediment/mat communities, and the rate was about 2 mg N m^-2 day^-1 for the lagoonal limestone substrata. Total lagoonal benthic N₂ fixation contributed 24.4% of the total nitrogen requirement for the benthic primary production of benthic communities of the lagoon. The input of N₂ fixation by the microbial planktonic communities (including cyanobacteria) of the lagoon, which are highly productive, is unquantified but is likely to be large.     

Effects of oxygen,mannitol and ammonium concentrations on nitrogenase [C2H2] activity in a marine skeletal carbonate sand

Following a lag of 3 to 18 h, acetylene reduction in mannitol-amended sand systems proceeded at approximately constant and high rates for periods up to 4 days. Carbon dioxide production and O₂ consumption were low in these systems in comparison to similar systems additionally amended with ammonium, indicating N-limitation of growth in the former. Thus, long-term acetylene assays of mannitol-amended sand and suspensions from the sand incubated at various partial pressures of oxygen could be used to characterize the O₂-sensitivity of the N₂-fixing bacterial population as a whole, in batch-type systems with a minimal degree of enrichment or change in pO₂ during the course of the assays. Results of various studies suggested that aerobic or microaerophilic N₂-fixing bacteria were absent or scarce in the sand, and that nitrogenase activity occurring in aerobically incubated systems occurred in anaerobic microenvironments. Hydrogen stimulated acetylene-reducing activity, but the time course differed from that of mannitol-supported activity, and proceeded with shorter lags in systems incubated at 0.2 and 0.05 atm O₂ than in systems incubated anaerobically. Efficiency of N₂ fixation [C₂H₂] increased with decreasing initial mannitol concentration. For sand washed with seawater to remove native combined inorganic nitrogen, and amended with 0.015% mannitol, 374 μmoles added NH₄-N/kg wet sand caused almost complete repression of nitrogenase activity, while concentrations as low as 12 μmoles added NH₄-N/kg wet sand appeared to cause at least partial repression of nitrogenase activity. Some implications of these results for the existence of anaerobic microenvironments in the cavities of skeletal carbonates, and for N₂-fixation in the seagrass rhizosphere are discussed.     

Different niches of the Halophila stipulacea seagrass bed harbor distinct populations of nitrogen fixing bacteria

Halophila stipulacea beds in the northern Gulf of Elat (Red Sea) may be subdivided into three different sub-habitats, each harboring a distinct population of diazotrophs. Nitrogen (N₂) fixation in the phyllosphere and in the rhizomes/upper-sediment niche was light dependent, suggesting its dependence on photosynthesis. N₂ fixation in the phyllosphere was not affected by the addition of either glucose or 3-3,4-dichloro-phenyl-1,1-dimethyl-urea (DCMU), indicating that the diazotrophs involved carried out non-oxygenic photosynthesis. They may, thus, have been photosynthetic bacteria. N₂ fixation in the rhizomes/upper-sediment niche, however, was greatly enhanced by the addition of glucose, but was suppressed in the presence of DCMU. This indicates that the diazotrophs involved here probably possess two photosystems (I and II) and may be Cyanobacteria. The anaerobic rhizosphere, in which fixation rates in light were very slow but were greatly enhanced by the addition of glucose, is probably populated by heterotrophic diazotrophs. Plant and sediment samples used in the present study were collected from the Gulf of Elat between 1990 and 1992.     

Acetylene reduction (nitrogen fixation) in a Delaware,USA salt marsh

Acetylene reduction (nitrogen fixation) was measured in several vegetational areas in a Delaware, USA salt marsh. Samples were collected for 1 yr and the results showed a seasonally variable pattern of acetylene reduction at all stations. Peak rates were generally recorded during the later summer and early fall (September–October). The seasonality was influenced mainly, although not exclusively, by the soil temperature. In addition, samples collected in short Spartina alterniflora stands exhibited rates which were up to 20-fold higher than those found in samples from tall S. alterniflora stands. Over 50% of the total yearly ethylene production occurred from mid-August until the beginning of December at the tall and short S. alterniflora stations. Maximum activity occurred at 5 cm depth for all stations. Surface activity accounted for only 3–4% of the total measured in the top 20 cm. Addition of glucose or mannitol resulted in considerable increases in activity, thus suggesting that heterotrophic acetylene reduction is carbon and/or energy limited. The results obtained in this study indicate that the measured rates are only potential rates and that considerable caution must be used in extrapolating from acetylene reduction rates to nitrogen fixation rates in situ.     

Gas-exchange studies with the staghorn coral Acropora acuminata and its zooxanthellae

Compensation point and light-saturation values were determined from oxygen-exchange experiments with branches and isolated zooxanthellae from the staghorn coral Acropora acuminata. Branches and dense suspensions of zooxanthellae showed similar lightresponse curves for oxygen exchange, with light saturation at about 23 Klux (300 W. m^-2) and compensation point occurring between 4 and 6 Klux (60–80 W. m^-2). Zooxanthellae appear to be mutually shaded in dense suspensions and coral tissues. The effects of metabolic inhibitors, including photosynthetic and respiratory inhibitiors, on oxygen exchange in coral branches and isolated zooxanthellae are presented. Bubbles formed on coral tissues and on several macroalgae under conditions of high illumination contained large amounts of oxygen, suggesting that a high oxygen tension may occur in coral tissues during the day. Photorespiration and dissolved organic carbon production by suspensions of zooxanthellae are discussed in relation to a high oxygen tension which probably occurs in coral tissues during daylight.     

Defensive mechanisms and ecology of some tropical holothurians

The ecology, defensive behavior and toxicity of three species of reef flat holothurians (Actinopyga mauritiana, Holothuria atra and Holothuria difficilis) were studied at Eniwetok Atoll, Marshall Islands. The average diurnal population density of H. difficilis ranged from 1.4 to 32 holothurians/900 cm²; resting respiratory rates (0.05 ml O₂/g wet wt/h) were comparable during day and night; nourishment in H. difficilis may be primarily from bacteria and foraminifera in which about 2% of the dry weight of sediment consumed is utilized, and the species probably passes at least 3 g dry wt of sediment/m²/day (>1 kg/m²/year). A fundamental difference in energy flow is suggested: considerably more energy is passed from benthic algae to grazing and browsing fishes to predatory fishes on coral reefs whereas, in extra-tropical latitudes, more energy is shunted from benthic algae to invertebrates to predators. The effects of holothurin leading to death in fishes are irreversible. Holothuria difficilis is best protected from predation. Its body wall is toxic and it can accurately eject Cuvierian tubules, which are also toxic. The discharge of tubules was regulated by a circadian rhythm in May. Studies on holothurians and sponges suggest that many exposed coral reef invertebrates have evolved effective defensive mechanisms in association with high intensity predation.Supported by A.E.C. Contract AT (29-2)-226 with the University of Hawaii.