Discrimination between several diterpenoid compounds in feeding by Gambusia affinis.

The alcyonacean soft coral Sinularia flexibilis Quoy and Gaimard produces a number of bioactive complementary (secondary) metabolites. The ability of three of these diterpenes - flexibilide, sinulariolide and dihydroflexibilide - to elicit differential discriminatory feeding behavior in Gambusia affinis was assessed in a feeding deterrence study. Terpene-impregnated fish flakes were offered to fish trained to feed on such food, and their responses (acceptance, rejection, avoidance, or no response) were assessed as indicative of feeding deterrence. Food treated with sinulariolide (a compound previously determined to be non-bioactive) was generally accepted at a 1% concentration. It was avoided and rejected, however, at a concentration of 10%, a concentration level generally restricted to polyp-rich branchlets. This indicated negative olfactory and palatability cues, respectively. Flexibilide-treated flakes were accepted to some extent by the fish at the 1% concentration, but strongly avoided at the 10% concentration, indicating effective feeding deterrence via olfaction. Dihydroflexibilide-impregnated flakes were strongly rejected even at low concentrations (1%) after tasting, indicating a negative palatibility cue. They were strongly avoided or rejected at higher concentrations (10%), indicating a negative olfactory cue as well. This response at higher concentrations indicates that sinulariolide and flexibilide become effective at concentrations between 1% and 10%. Such concentrations may be found in the polypary (polyp-bearing portion) of the soft coral colonies. Dihydroflexibilide elicited the strongest negative palatability response from these test fish. The feeding deterrence characteristics of the compounds determined here represent the potentials of individual compounds to elicit differential feeding responses in organisms like Gambusia which are capable of discriminating between different but closely related complementary (secondary) metabolites.

Antimicrobial activity of the diterpenes flexibilide and sinulariolide derived from Sinularia flexibilis Quoy and Gaimard 1833 (Coelenterata: Alcyonacea, Octocorallia).

The soft coral Sinularia flexibilis is rarely overgrown by bacteria and algae. Various studies have shown that it contains diterpenes that protect it from competitors and predators. However, of the many diterpenoids isolated from S. flexibilis, only sinulariolide has been studied for antibiotic properties. Samples of soft corals were collected from Orpheus Island and freeze-dried for chemical extraction and isolation of pure diterpenes. Antimicrobial activity of the diterpenes was determined using the disc assay method with antibiotics as controls and the minimum inhibitory concentrations of the diterpenes were determined using the Tube Dilution Technique. Two out of the five diterpenes tested (sinulariolide and flexibilide), showed marked antimicrobial activity and inhibited growth of Gram-positive bacteria. Flexibilide was effective even at concentrations as low as 5 ppm, whereas sinulariolide was effective at concentrations of 10 ppm. These compounds show potential as antibiotics.

Cardiac and vascular responses of isolated rat tissues treated with diterpenes from Sinularia flexibilis (coelenterata: octocorallia).

The marine environment is a rich source of compounds with cardiovascular activity. This study characterizes the cardiac and vascular responses in isolated rat tissues of flexibilide, dihydroflexibilide and sinulariolide, three diterpenes isolated from the soft coral Sinularia flexibilis. On rat left ventricular papillary muscles, dihydroflexibilide and flexibilide showed similar potencies (-log EC50 = 4.69 +/- 0.05 and 4.66 +/- 0.06, respectively); the maximal response to dihydroflexibilide of 1.4 +/- 0.2 mN was 35 +/- 7% that of calcium chloride in the same muscles. All diterpenes relaxed rat thoracic aortic rings precontracted with KC1 (100 mM) with similar potencies (flexibilide, -log EC50 = 4.17 +/- 0.06). Flexibilide was further characterized and shown to increase force in isolated rat left atria by 0.8 +/- 0.5 mN at 1 x 10(-4) M, to increase rate of contraction in isolated rat right atria by 18 +/- 5 beta/min at 3 x 10(-5) M and to completely relax endothelium-denuded rat thoracic aortic rings (-log EC50 = 4.14 +/- 0.05). Toxicity as indicated by the occurrence of ectopic beats was not observed with the diterpenes at concentrations which produced complete relaxation of blood vessels, maximal positive inotropic activity and minor positive chronotropic responses. Thus, these compounds may be useful lead compounds in the search for improved treatment of cardiovascular disease, especially heart failure.

Lack of predictability in terpenoid function Multiple roles and integration with related adaptations in soft corals.

Terpenoids are commonly found in alcyonacean soft corals of the Great Barrier Reef (Coelenterata, Octocorallia), but they are highly variable in their distribution within this order. Such secondary metabolites in these organisms vary widely in type, concentration, and function. Thus far, they have been found to play roles in predator defense, competitor defense, anti-fouling, and reproduction. The effectiveness of compounds derived from individual species also varies widely with respect to interactions with other members of the community. These chemical characters function in concert with other morphological, physiological, behavioral, or developmental adaptations that have similar functions. Integration of these adaptations is highly species-specific. The multiple functions of terpenoid compounds and the high degree of species specificity in their ecological roles greatly reduces the ability to predict either the presence or absence of a terpenoid from other related characters. In addition, simple indicators of terpenoid function appear not to exist. If such indicators do exist, they are more likely to be found at higher levels of taxonomic resolution (within families or genera).

Localized dispersal and recruitment in great barrier reef corals: the helix experiment.

To examine the problem of how far coral larvae disperse from their natal reef, coral recruitment densities were experimentally determined at distances up to 5 kilometers from a small, relatively isolated platform reef, Helix Reef, on the central Great Barrier Reef for 7 months. High concentrations of recruits, accounting for up to 40 percent of all recruitment, were found downstream of the reef in areas of high water residence time, suggesting that near-field(proximal) circulation has a profound influence on dispersal and recruitment of coral larvae. Coral recruitment declined logarithmically with distance from the reef, decreasing by an order of magnitude at radial distances of only 600 to 1200 meters. On an ecological time scale, advective dispersal of semipassive marine larvae with relatively short planktonic lives(minimally days) may be extensive, but success of recruitment is highly limited. Through evolutionary time, sufficient dispersal occurs to ensure gene flow to reef tracts hundreds or possibly thousands of kilometers apart. In the short term, however, coral reefs appear to be primarily self-seeded with respect to coral larvae.

Defensive strategies of soft corals (Coelenterata: Octocorallia) of the Great Barrier Reef : III. The relationship between ichthyotoxicity and morphology.

The relationship between ichthyotoxicity and predation-related defensive functional morphology was examined in alcyonacean soft corals of the central and northern regions of the Great Barrier Reef (GBR), Australia. Approximately 170 specimens were assessed encompassing a number of genera within three families: 1) the Alcyoniidae (Lobophytum, Sarcophytum, Sinularia, Cladiella, Parerythropodium, and Alcyonium); 2) Neptheidae (Lemnalia, Paralemnalia, Capnella, Lithophyton, Nephthea, Dendronephthya, Scleronephthya, and Stereonephthya), and 3) Xeniidae (Anthelia, Efflatounaria, Cespitularia, Heteroxenia, and Xenia). Ichthyotoxicity data were derived from earlier studies which used Gambusia affinis Baird and Girard (Vertebrata, Pisces) as a test organism. These data were compared to morphological data collected from specimens in the field and laboratory. Three sets of statistical analyses were performed, each considering a progressively narrower group of taxa. The first included 68 specimens and considered 16 morphological characters in each, falling into the general categories of gross colony form, colony texture, presence of mucus, colony color, polyp retractility, and sclerite morphology and distribution. These were tested for independence against ichthyotoxicity data. The second set of analyses involved a more restricted morphological data set derived from 28 species of Sinularia in combination with 28 species within the Nephtheidae, comparing them to their respective toxicity ranks. The third analysis considered the previous two taxonomic groups separately in relation to their toxicity levels.The attempt to consider many morphological characters in a taxonomically diverse collection did not reveal any general association in the Alcyonacea between defensive morphology and toxicity, and those associations which did emerge were clearly erroneous. The second analysis, considering only Sinularia spp. and nephtheids, demonstrated a negative association between ichthyotoxicity and the morphological characters of a) polypary armament, b) microarmament of the individual polyp, and c) strong mineralization of the coenenchyme. The third analysis revealed that the negative association found between toxicity and the first two characters was derived entirely from the nephtheids while the association detected between toxicity and the third character was restricted to Sinularia. It is concluded that a relationship between toxicity and morphology can be demonstrated, but it is heavily dependent upon which specific morphological characters are being considered and at what level of taxonomic resolution the analysis is being performed. An approach utilizing many characters over many taxa is unlikely to yield significant, reliable, or meaningful results.