Photosensitive behaviors and circadian rhythms are well documented in reef-building corals and their larvae, yet photoreceptive structures and opsins have not been described in these organisms. Here I provide evidence for red sensitivity in several species of coral larvae. Behavioral experiments with two Caribbean corals, Porites astreoides and Acropora palmata demonstrated that larvae settle and metamorphose at a greater frequency on red substrata than on similar substrata of other colors. Attachment to red substrata was not observed when larvae were maintained in the dark, suggesting that red sensitivity was responsible for the observed behavior. Extracellular recordings confirm photosensitivity and indicate that the peak sensitivity of coral photoreceptors are shifted towards the orange-red region of the visible light spectrum, similar to the spectra (fluorescence and reflectance) of preferred artificial (plastic) and natural (crustose coralline algae) settlement substrata. Using Blast analyses and a PCRbased approach, I have identified, sequenced and cloned two full-length opsin cDNAs from A. palmata larvae. One cDNA (Acropsin 1) encodes an opsin protein that is similar to a vertebrate melanopsin; the second (Acropsin 2) encodes a protein that is most similar to cephalopod rod opsin. I have successfully developed synthetic peptide antibodies against each Acropsin 1 and Acropsin 2. Western blots of adult A. palmata and A. cervicornis protein detect a 37kDa and 40kDa band, corresponding to the predicted molecular weights of Acropsins 1 and 2, respectively. Immunohistochemistry confirms expression of both opsins in A. palmata larvae. Staining of sectioned larvae demonstrates that Acropsin 1 is localized in the larval gastroderm while Acropsin 2 is localized in solitary epithelial cells, scattered throughout the larval ectoderm but with a polarized distribution and higher concentration in the aboral epidermis. This research provides several lines of evidence to support the existence, and demonstrate one potential ecological function, of opsin-based photosensitivity in corals.

Made available in DSpace on 2015-03-26T13:47:08Z (GMT). No. of bitstreams: 1 texto completo.pdf: 407930 bytes, checksum: 2e2e384d18c87e7272273f070c122bed (MD5) Previous issue date: 2006-09-29 / The necessity to treat different etiologies, magnitudes and localization of bone defects has stimulated the search and development of materials capable to substitute the bone. Autogenous bone graft, although considered ideal because his behavior, in the majority of the situations, presents intrinsic morbidity of attainment, and limitation as for the amount and morphology. The bone s bank allows to provide great part of the current necessities, but it possess related problems with cost, infrastructure necessity and, in human medicine, legal issues. The ideal bone substitute must be osteogenic, biocompatible, capable to provide scaffolds, to propagate other substances, to be easily usable in the clinic and have adjusted ratio cost-benefit. On the other side, in determined applications, it is desirable that some of these characteristics predominate on others, depending on the necessity of the case to treat. This paper has for objective to evaluate the Porites astreoides coral as alternative of bone substitute in the fulfillment of small defects, associate or not to the autogenous bone marrow, and to compare its morphology and composition with normal bone tissue. With this intention there was accomplish a comparative study of the structural and morphologic properties of Porites astreoides coral samples, synthetic hydroxyapatite and bone of young and adult dogs, by x-rays diffraction; and an experimental study, comparing the samples previously, in the repair bone defects in dogs tibia , by radiographic and histology evaluations. / A necessidade de tratar defeitos ósseos de diferentes etiologias, magnitudes e localizações têm estimulado a busca e desenvolvimento de materiais capazes de substituir o osso. O autoenxerto, mesmo sendo considerado o enxerto ideal por seu comportamento, na maioria das situações, apresenta morbidade intrínseca a sua obtenção, e limitação quanto à quantidade e morfologia. Os bancos de ossos permitem resolver grande parte das necessidades atuais, mas possui problemas relacionados a custo, necessidade de infra-estrutura e, em medicina humana, de ordem médico-legal. O substituto ósseo ideal deve ser osteogênico, biocompatível, capaz de proporcionar suporte estrutural e de veicular outras substâncias, facilmente utilizável na clínica e com adequada proporção custo-benefício. Por outro lado, em determinadas aplicações, é desejável que algumas dessas características predominem sobre outras, dependendo da necessidade do caso a tratar. Esta dissertação tem por objetivo avaliar o coral Porites astreoides como alternativa de substituto ósseo no preenchimento de pequenos defeitos, associado ou não à medula óssea autógena, e comparar a sua morfologia e composição com a do tecido ósseo normal. Para tal propósito foi realizado um estudo comparativo das propriedades estruturais e morfológicas de amostras do coral Porites astreoides, hidroxiapatita sintética e de ossos de cães jovens e adultos, por meio da difração de raios-x; e um estudo experimental, comparando as amostras citadas anteriormente, na reparação de falhas ósseas realizadas na tíbia de cães, por meio de avaliações radiográficas e histológicas.

Coral reef biodiversity is threatened by rapidly changing anthropogenic activities and natural perturbations, leading to massive ecological and economic consequences ranging from the loss of fisheries to coastal erosion. It is necessary to understand corals responses to environmental changes in order to determine management programs on appropriate spatial and temporal scales to address these issues. Coral larvae are the product of sexual reproduction, have the potential to recruit to new areas, and are fundamental in maintaining genetic diversity. These larvae are subjected to variations in local environmental conditions until they settle, inducing specific larval molecular response patterns. One factor that influences coral health is salinity. Low salinities can alter cell homeostasis creating stress in cells. In the natural environment larvae may be exposed to low salinities due to heavy rainfall or run-off. This study investigated larvae responses to low salinity and characterized gene expression in the reef-building coral Porites astreoides using a coral stress-focused microarray. Nine batches of 250+ larvae from three different colonies were collected and immediately exposed in an acute hyposalinity experiment. Samples from two treatments of 25 and 30 ppt, and a control at 35 ppt were used in this study. After experimental exposure these samples were stored in RNAlater® and molecular analysis was performed. The RNA from the samples was extracted, purified and hybridized to a coral stress-focused microarray. Statistical analysis indicates 72 genes were differentially expressed across treatments (p<0.003, analysis of variance). The hierarchical cluster analysis groups together the larvae exposed to salinities of 30 and 35 ppt indicating both treatments induced similar patterns of gene expression. Larvae responses to 30 ppt are minimal, suggesting larvae can tolerate acute exposures to 30 ppt salinity levels. In contrast, the lower salinity (25 ppt) induced a strong response in both the coral and zooxanthellae. The coral larvae up-regulated stress response genes and down-regulated genes associated with normal cell functioning. Additionally, the zooxanthellae down-regulated genes associated with photosynthesis. These results suggest larvae may be vulnerable to bleaching, which may affect the ability of larvae to successfully undergo metamorphosis and survive at low salinities. However, this has yet to be confirmed with complementary techniques. Long-term studies are recommended to examine the effects of hyposalinity on larvae at different time scales and life history stages.

Multiple stressors have caused a decline in coral populations. Broadcast spawning corals once dominated the Florida Reef Tract (FRT), but since their decline, smaller brooding corals, soft corals, and macroalgae are replacing them. Brooding corals are more resilient to current threats in part because they are reproductive throughout much of the year and their larvae are competent to settle after release. Despite the ubiquity of brooders on Florida reefs, much of their reproductive strategy remains unknown. This study aimed to examine paternity as a function of colony size and density in Porites astreoides, a common brooding coral in the FRT. Porites astreoides colonies were configured in arrays at three densities that were replicated three times. A focal colony was surrounded by six other colonies, separated from the focal colony at different distances (1m, 7m, and 15m) representing high, moderate, and low population densities, respectively. All arrays were placed in the field but were separated from the reef and naturally occurring P. astreoides colonies by at least 50 m. Four days before the new moon, colonies were transported to the laboratory for larval collection. Over a four day period, a total of 3,184 larvae were collected from 24 colonies, 13 of which released larvae over consecutive days. The resulting larvae were genotyped using seven microsatellite markers. All larvae had the exact genotypes of the colony from which the larvae were collected, i.e. maternal- egg donor. This suggested the larvae were parthenogenically produced and no sperm was used to fertilize the eggs. This is the first study to suggest that parthenogenesis is occurring in P. astreoides. In today's oceans that have been depleted of corals, parthenogenesis may be an advantageous reproductive strategy used to boost populations. However, parthenogenesis reduces the genetic diversity which could hinder successful sexual reproduction in the future causing fragmented populations.

Restoration efforts are being implemented in many of the world’s coral reefs due to damages from anthropogenic sources such as ship groundings and anchor damage. One restoration technique involves attempts to save dislodged and fragmented coral colonies by transplanting them back to damage sites. Research has shown that survivorship and growth of transplanted colonies is comparable to that of natural, control colonies. What remains unknown is to what extent transplantation affects the ecological success and reproduction of dislodged and fragmented coral colonies. The purpose of this study was twofold. Reproduction and spawning information is sparse for S. intersepta and Solenastrea bournoni, so the first purpose was to describe gamete development of these two species and assess correlations between environmental dynamics and spawning of each species. Tissue samples were collected throughout Broward County, Florida and processed for histological examination. Gametes were counted, and development was assessed. For S. intersepta and S. bournoni, late stage oocyte abundance was compared with environmental factors of mean daily water temperatures at depth, lunar phase, semidiurnal tides and solar insolation for correlative evidence to predict future spawning events. Findings indicated that both S. bournoni and S. intersepta are gonochoric broadcast spawners. Solenastrea bournoni spawns annually after the full moon in September when sea temperatures are at a maximum. Stephanocoenia intersepta spawns annually after the full moon of August or September, depending on the timing of the full moon. The second purpose was to determine if previously transplanted Porites astreoides, Montastraea cavernosa, Siderastrea siderea and Stephanocoenia intersepta corals produce gametes and spawn similarly to naturally occurring colonies and to address the issue of transplantation as a suitable resource management tool to aid in reef recovery for future coral generations. Results indicated no significant difference in fecundity between transplants and controls of M. cavernosa, S. siderea or S. intersepta. A significant difference was found in fecundity between P. astreoides transplants and controls, but it is thought that it is due to a difference in depth of collected samples. Overall, this study shows that transplantation of coral colonies after damage and fragmentation events does not have adverse effects on the long-term fecundity of coral colonies.

Corals combine photosynthesis and calcification in an intricate and delicately balanced relationship to form large biomineralized structures that are dominant features of tropical coastlines worldwide. Coral reefs have great scientific and economic importance but have recently experienced widespread decline attributed to increasing anthropogenic pressure on reef systems. Physical damage events, such as ship groundings, when coupled with existing nutrient stress and changing global climate present a poor outlook for successful natural recovery of reef communities. The main goal of the proposed research is to better understand how environmental factors, both local and global, affect the coral holobiont and influence overall coral fitness. The target species of this research, Montastraea cavernosa and Porites astreoides, are important and widespread Caribbean reef-builders. While it has been shown that nutrient and pCO2 stress affect coral growth and calcification, study of specific effects on coral tissue and reproductive success has not received significant attention in the literature despite considerable current interest. This study addresses this data gap in quantitatively examining the effect of elevated nutrients and pCO2 on 1) P. astreoides recruit survivorship, development, early calcification, and symbiotic zooxanthellae morphology; 2) M. cavernosa and P. astreoides wound regeneration, tissue characteristics over time at the histological and ultrastructural level, and trends in symbiotic zooxanthellae morphology; and 3) survival, growth and histological/ultrastructural characteristics of M. cavernosa and P. astreoides fragments transplanted to the field and in the laboratory. Histological and ultrastructural observations from corals transplanted to the field are then compared to ex-situ laboratory experimental corals. In the fleshy and large-polyped faviid M. cavernosa, healing of a linear wound was characterized by granulation of new tissue across the wound site, facilitated by coalescent granular amoebocytes. The wound healing strategy of this species appears to progress with wound closure and re-epithelialization before calcification resumes, as actively calcifying calicodermis was generally not observed at the healing front. Tissue regeneration in the small-polyped P. astreoides was characterized by formation of multiple islands of eosinophilic healing fronts along the depth of the wound track, and an accumulation of granular amoebocyte cells in regenerating tissue. The wound healing strategy of this species appeared to result in re-epithelialization of exposed body wall without necessarily closing the wound. Elevated pCO2 significantly reduced survivorship in P. astreoides recruits, and both nutrient enrichment and elevated pCO2 significantly reduced wound regeneration rate in M. cavernosa and P. astreoides. In both species, phosphate enrichment had the greatest deleterious effect on wound repair. A significant application of this study is the identification of possible zooxanthellar morphological indices of elevated nutrients and ocean acidification. The similarity in starch, lipid and uric acid accumulation patterns in Symbiodinium sp. from P. astreoides recruits and coral fragments of both species indicate a correlation between these anthropogenic stressors and the intracellular accumulation of excess carbon and nitrogen by the symbiont. Zooxanthellar carbon accumulation, in the form of starch and/or lipid, was the greatest under elevated nitrate. Zooxanthellar nitrogen accumulation, in the form of uric acid, was the greatest under elevated CO2. Comparison of zooxanthellar metrics between the field corals (P. astreoides, and M. cavernosa) and ex-situ corals and recruits indicated that carbon accumulation in Symbiodinium from field corals was consistently significantly less than in the ex-situ experimental P. astreoides recruits and M. cavernosa fragments exposed to elevated nitrate. This indicates that the field corals were likely not exposed to elevated nitrate at the time of collection. Both M. cavernosa and P. astreoides adults in the field accumulated significantly less uric acid than their counterparts in the tissue repair experiment, indicating that the field corals were exposed to higher pH and lower CO2 than the ex-situ corals. These results suggest that the field corals were not exposed to nutrient concentration profiles similar to the experimental treatments, particularly elevated nitrate. However, histological metrics indicated that the transplanted corals were subjected to increasing sedimentation stress over time. Overall, nitrate was found to affect recruits and adults on a similar scale, while phosphate and pCO2 affected carbon and nitrogen storage more in recruits compared to adults. While nutrients and pCO2 had no mechanistic effect on regeneration at histological level, ultrastructural metrics indicate an impact on the mutualistic energy exchange between the symbiotic partners, partially decoupling symbiosis. Effects were generally found to be greater in P. astreoides compared to M. cavernosa, and the unique life history strategy of the subject species and differences in their endosymbiont physiology reveal distinct responses to elevated nutrients and pCO2. Although the laboratory findings were not necessarily applicable to field observations, they provide insight into factors that may influence fragment success in the field. Quantitative assessment of the effect of elevated nutrients and pCO2 is thus useful in management decisions involving water quality standards, and is essential in the prediction of future coral condition and resilience.

Reef-building coral populations throughout the world are being threatened by numerous stressors and continue to decline. As potent endocrine-disrupting compounds, exogenous sex steroid contamination has been a largely overlooked stressor to corals. Previous research indicates these compounds are prevalent in marine environments, fluctuate annually along with reproductive cycles, can bioaccumulate, and have had variable effects on growth and reproduction in several cnidarian species. This project had three primary objectives: (1) establish environmental estradiol and progesterone concentrations in Broward County and lower Florida Keys reef environments, (2) conduct 17 β-estradiol and progesterone larval assays on P. astreoides larvae to determine the effects of these compounds on settlement and viability, and (3) conduct 17 β-estradiol and progesterone dosing experiments on adult Acropora cervicornis and Porites astreoides fragments to determine the effects on growth, zooxanthellae, reproduction, and overall tissue health. Estradiol was detected in surface and at-depth water samples from Broward County and lower Keys reef sites at effect level concentrations for marine organisms. Broward County larvae treated with low progesterone (5 ng/L) had decreased survival, while lower Keys larvae in low estradiol treatments (1 ng/L) had increased on-disc settlement. No other treatment effects were observed, however, lower Keys larvae had greater overall survival in comparison to Broward County larvae. There were no significant differences between estradiol and progesterone treatments in the adult-dosing experiment for growth, zooxanthellae density, reproduction, and overall tissue health. This is the first study to detect estradiol at Broward County reefs sites and our results, while inconclusive, indicate these compounds may have the potential to affect coral reef ecosystems.