High natural gene expression variation in the reef-building coral Acropora millepora: potential for acclimative and adaptive plasticity

Background: Ecosystems worldwide are suffering the consequences of anthropogenic impact. The diverse ecosystem of coral reefs, for example, are globally threatened by increases in sea surface temperatures due to global warming. Studies to date have focused on determining genetic diversity, the sequence variability of genes in a species, as a proxy to estimate and predict the potential adaptive response of coral populations to environmental changes linked to climate changes. However, the examination of natural gene expression variation has received less attention. This variation has been implicated as an important factor in evolutionary processes, upon which natural selection can act. Results: We acclimatized coral nubbins from six colonies of the reef-building coral Acropora millepora to a common garden in Heron Island (Great Barrier Reef, GBR) for a period of four weeks to remove any site-specific environmental effects on the physiology of the coral nubbins. By using a cDNA microarray platform, we detected a high level of gene expression variation, with 17% (488) of the unigenes differentially expressed across coral nubbins of the six colonies (jsFDR-corrected, p < 0.01). Among the main categories of biological processes found differentially expressed were transport, translation, response to stimulus, oxidation-reduction processes, and apoptosis. We found that the transcriptional profiles did not correspond to the genotype of the colony characterized using either an intron of the carbonic anhydrase gene or microsatellite loci markers. Conclusion: Our results provide evidence of the high inter-colony variation in A. millepora at the transcriptomic level grown under a common garden and without a correspondence with genotypic identity. This finding brings to our attention the importance of taking into account natural variation between reef corals when assessing experimental gene expression differences. The high transcriptional variation detected in this study is interpreted and discussed within the context of adaptive potential and phenotypic plasticity of reef corals. Whether this variation will allow coral reefs to survive to current challenges remains unknown.

Coral Gables Virtual History

Florida International University has partnered with the City of Coral Gables to create a geospatially-enabled Virtual Historic City. With this tool, users may navigate to any point in the city at a selected time period (e.g. the Biltmore Hotel from the 1920s to 1940s) and experience the city as it was, through a wide variety of cultural artifacts and textual materials. A virtual walking tour of Coral Gables was also created, which includes an audio narration and 3D simulations providing the façades of historic buildings and landscape. Students, educators, historians, public policy administrators, architects, sociologists, environmental analysts, urban planners, and the general public can explore over 8,000 historical documents, photographs, oral histories, and maps that have been spatially registered to their relevant locations and time.

Disturbance Driven Colony Fragmentation as a Driver of a Coral Disease Outbreak

In September of 2010, Brewer's Bay reef, located in St. Thomas (U.S. Virgin Islands), was simultaneously affected by abnormally high temperatures and the passage of a hurricane that resulted in the mass bleaching and fragmentation of its coral community. An outbreak of a rapid tissue loss disease among coral colonies was associated with these two disturbances. Gross lesion signs and lesion progression rates indicated that the disease was most similar to the Caribbean coral disease white plague type 1. Experiments indicated that the disease was transmissible through direct contact between colonies, and five-meter radial transects showed a clustered spatial distribution of disease, with diseased colonies being concentrated within the first meter of other diseased colonies. Disease prevalence and the extent to which colonies were bleached were both significantly higher on unattached colony fragments than on attached colonies, and disease occurred primarily on fragments found in direct contact with sediment. In contrast to other recent studies, disease presence was not related to the extent of bleaching on colonies. The results of this study suggest that colony fragmentation and contact with sediment played primary roles in the initial appearance of disease, but that the disease was capable of spreading among colonies, which suggests secondary transmission is possible through some other, unidentified mechanism.

Coral Thermal Tolerance: Tuning Gene Expression to Resist Thermal Stress

The acclimatization capacity of corals is a critical consideration in the persistence of coral reefs under stresses imposed by global climate change. The stress history of corals plays a role in subsequent response to heat stress, but the transcriptomic changes associated with these plastic changes have not been previously explored. In order to identify host transcriptomic changes associated with acquired thermal tolerance in the scleractinian coralAcropora millepora, corals preconditioned to a sub-lethal temperature of 3°C below bleaching threshold temperature were compared to both non-preconditioned corals and untreated controls using a cDNA microarray platform. After eight days of hyperthermal challenge, conditions under which non-preconditioned corals bleached and preconditioned corals (thermal-tolerant) maintained Symbiodinium density, a clear differentiation in the transcriptional profiles was revealed among the condition examined. Among these changes, nine differentially expressed genes separated preconditioned corals from non-preconditioned corals, with 42 genes differentially expressed between control and preconditioned treatments, and 70 genes between non-preconditioned corals and controls. Differentially expressed genes included components of an apoptotic signaling cascade, which suggest the inhibition of apoptosis in preconditioned corals. Additionally, lectins and genes involved in response to oxidative stress were also detected. One dominant pattern was the apparent tuning of gene expression observed between preconditioned and non-preconditioned treatments; that is, differences in expression magnitude were more apparent than differences in the identity of genes differentially expressed. Our work revealed a transcriptomic signature underlying the tolerance associated with coral thermal history, and suggests that understanding the molecular mechanisms behind physiological acclimatization would be critical for the modeling of reefs in impending climate change scenarios.

Aspects Allogenic Memory in the Gorgonian Coral Swiftia Exserta

Accelerated graft rejection can be used to determine immune memory in the gorgonian coral swiftia exserta. The extent ofthe persistence of immune memory will be determined in this experiment using replicate sets that are time elapsed from 1, 3, and 6 month. Although corals lack circulatory systems which can be a component of adaptive systemic immunity, this study will attempt to determine whether this gorgonian coral is capable of transmitting immune information throughout its colonial body. Results showed that at each of the time points; one, three, and six months the secondary response group and the primary response group were significantly different (at p=0.001) therefore, demonstrating long term immune memory. While the primary response group and the 3rd party specificity response group were similar, both were significantly different (at p=O. 001) from the secondary response group which shows the response to be specific, with memory applicable to the original antigen. Systemic immunity was not determined to be present for 15 em and one week after initial sensitization.

Density-Dependent Effects of Coral Reef Restoration on Herbivory and Community Interactions

Coral reefs are among the most productive ecosystems in the world. Yet, with their recent declines due to disease, climate change, and overfishing, restoration of these habitats is one of the main concerns for ecologists, resource managers, and government organizations. Coral reef restoration aims to promote key ecosystem processes to shift these habitats to their historical state of high coral cover, but few studies have focused on effective ways to promote resilience. In addition, little is known about the impact of restoration on the fish communities. The aim of this study is to understand how the community of herbivorous fishes is affected by the density of coral outplants inside a special protection area located in the Florida Keys National Marine Sanctuary. Grazing rates, number of visits and time spent foraging were compared using video footage of sites previously devoid of corals, and six months after coral restorations had occurred. Coral transplantations did not appear to attract herbivores nor increase grazing rates of fishes. Instead Sparisoma and Acanthurus fishes appear to respond to changes in the environment by modifying their grazing behavior. However, there was an observed increase in visits by Acanthurus species after transplantation for all the sites sampled within the reef. These fishes seemed to prefer low coral cover sites for grazing. This study highlights the importance of examining coral restorations impacts at the community level. Understanding how restoration influences herbivores and other guilds of reef fishes will allow individuals to not only determine if these habitats are returning to their “original” state, but provide more information on the ways these systems cope with changes in the environment.

Contribution of lipophilic secondary metabolites to the toxicity of strains of freshwater cyanobacterial harmful algal blooms, identified using the zebrafish (Danio rerio) embryo as a model for vertebrate development

Cyanobacteria ("blue-green algae") are known to produce a diverse repertoire of biologically active secondary metabolites. When associated with so-called "harmful algal blooms", particularly in freshwater systems, a number of these metabolites have been associated—as "toxins", or commonly "cyanotoxins"—with human and animal health concerns. In addition to the known water-soluble toxins from these genera (i.e. microcystins, cylindrospermopsin, and saxitoxins), our studies have shown that there are metabolites within the lipophilic extracts of these strains that inhibit vertebrate development in zebrafish embryos. Following these studies, the zebrafish embryo model was implemented in the bioassay-guided purification of four isolates of cyanobacterial harmful algal blooms, namely Aphanizomenon, two isolates of Cylindrospermopsis, and Microcystis, in order to identify and chemically characterize the bioactive lipophilic metabolites in these isolates. ^ We have recently isolated a group of polymethoxy-1-alkenes (PMAs), as potential toxins, based on the bioactivity observed in the zebrafish embryos. Although PMAs have been previously isolated from diverse cyanobacteria, they have not previously been associated with relevant toxicity. These compounds seem to be widespread across the different genera of cyanobacteria, and, according to our studies, suggested to be derived from the polyketide biosynthetic pathway which is a common synthetic route for cyanobacterial and other algal toxins. Thus, it can be argued that these metabolites are perhaps important contributors to the toxicity of cyanobacterial blooms. In addition to the PMAs, a set of bioactive glycosidic carotenoids were also isolated because of their inhibition of zebrafish embryonic development. These pigmented organic molecules are found in many photosynthetic organisms, including cyanobacteria, and they have been largely associated with the prevention of photooxidative damage. This is the first indication of these compounds as toxic metabolites and the hypothesized mode of action is via their biotransformation to retinoids, some of which are known to be teratogenic. Additional fractions within all four isolates have been shown to contain other uncharacterized lipophilic toxic metabolites. This apparent repertoire of lipophilic compounds may contribute to the toxicity of these cyanobacterial harmful algal blooms, which were previously attributed primarily to the presence of the known water-soluble toxins.^

PROTEOMIC ANALYSIS OF WOLBACHIA SYMBIOSIS WITHIN THE DROSOPHILA OVARY

Wolbachia pipientis are bacterial endosymbionts carried by millions of invertebrate species, including ~40% of insect species and some filarial nematodes. In insects, basic Wolbachia research has potential applications in controlling vector borne disease. Conversely, Wolbachia of filarial nematodes are causative agents of neglected tropical diseases such as lymphatic filariasis and African river blindness. However, remarkably little is known about how Wolbachia interact with their hosts at the molecular level. Understanding this is important to inform the basis for symbiosis and help prevent human disease. I used a high-throughput proteomics approach to study how Drosophila host cells are modified by Wolbachia infection. This analysis identified 23 Drosophila proteins that significantly changed in amount as a result of Wolbachia infection. A subset of differentially abundant host proteins were consistent with Wolbachia-associated phenotypes reported previously. This study also provides the first ever discovery-based evidence for a Wolbachia-associated change in maternal germline histone loads, which has possible implications in Rescue of a common Wolbachia-induced reproductive manipulation known as Cytoplasmic Incompatibility.

A Comparative Study of Concurrent Acoustic and Diver Survey Data, and Fish Community Descriptions of a High Latitude Coral Reef, Florida, USA

Fisheries independent data on relatively unstudied nekton communities were used to explore the efficacy of new tools to be applied in the investigation of shallow coastal coral reef habitats. These data obtained through concurrent diver visual and acoustic surveys provided descriptions of spatial community distribution patterns across seasonal temporal scales in a previously undocumented region. Fish density estimates by both diver and acoustic methodologies showed a general agreement in ability to detect distributional patterns across reef tracts, though magnitude of density estimates were different. Fish communities in southeastern Florida showed significant trends in spatial distribution and seasonal abundance, with higher estimates of biomass obtained in the dry season. Further, community composition shifted across reef tracts and seasons as a function of the movements of several key reef species.

Non-consumptive effects of predators in coral reef communities and the indirect consequences of marine protected areas

Predators exert strong direct and indirect effects on ecological communities by intimidating their prey. Non-consumptive effects (NCEs) of predators are important features of many ecosystems and have changed the way we understand predator-prey interactions, but are not well understood in some systems. For my dissertation research I combined a variety of approaches to examine the effect of predation risk on herbivore foraging and reproductive behaviors in a coral reef ecosystem. In the first part of my dissertation, I investigated how diet and territoriality of herbivorous fish varied across multiple reefs with different levels of predator biomass in the Florida Keys National Marine Sanctuary. I show that both predator and damselfish abundance impacted diet diversity within populations for two herbivores in different ways. Additionally, reef protection and the associated recovery of large predators appeared to shape the trade-off reef herbivores made between territory size and quality. In the second part of my dissertation, I investigated context-dependent causal linkages between predation risk, herbivore foraging behavior and resource consumption in multiple field experiments. I found that reef complexity, predator hunting mode, light availability and prey hunger influenced prey perception of threat and their willingness to feed. This research argues for more emphasis on the role of predation risk in affecting individual herbivore foraging behavior in order to understand the implications of human-mediated predator removal and recovery in coral reef ecosystems.^

Characterizations of the Major Coral Diseases of the Philippines: Ulcerative White Spot Disease and Novel Growth Anomalies of Porites

Coral reefs are in decline worldwide and coral disease is a significant contributing factor. However, etiologies of coral diseases are still not well understood. In contrast with the Caribbean, extremely little is known about coral diseases in the Philippines. In 2005, off Southeast Negros Island, Philippines, I investigated relationships between environmental parameters and prevalence of the two most common coral diseases, ulcerative white spot (UWS) and massive Porites growth anomalies (MPGAs). Samples were collected along a disease prevalence gradient 40.5 km long. Principal component analyses showed prevalence of MPGAs was positively correlated with water column nitrogen, organic carbon of surface sediments, and colony density. UWS was positively correlated with water column phosphorus. This is the first quantitative evidence linking anthropogenically-impacted water and sediment to a higher prevalence of these diseases. Histological and cytological alterations were investigated by comparing tissues from two distinct types of MPGA lesions (types 1 and 2) and healthy coral using light and electron microscopy. Skeletal abnormalities and sloughing, swelling, thinning, and loss of tissues in MPGAs resembled tissues exposed to bacterial or fungal toxins. Both lesion types had decreases in symbiotic zooxanthellae, which supply nutrients to corals. Notable alterations included migrations of chromophore cells (amoebocytes) (1) nocturnally to outer epithelia to perform wound-healing, including plugging gaps and secreting melanin in degraded tissues, and (2) diurnally to the interior of the tissue possibly to prevent shading zooxanthellae in order to maximize photosynthate production. Depletion of melanin (active in wound healing) in type 2 lesions suggested type 2 tissues were overtaxed and less stable. MPGAs contained an abundance of endolithic fungi and virus-like particles, which may result from higher nutrient levels and play roles in disease development. Swollen cells and mucus frequently blocked gastrovascular canals (GVCs) in MPGAs. Type 1 lesions appeared to compensate for impeded flow of wastes and nutrients through these canals with proliferation of new GVCs, which were responsible for the observed thickened tissues. In contrast, type 2 tissues were thin and more degraded. Dysplasia and putative neoplasia were also observed in MPGAs which may result from the tissue regeneration capacity being overwhelmed.

Proteomic Analysis of Wolbachia Symbiosis within the Drosophila

Wolbachia pipientis are bacterial endosymbionts of arthropods and in some filarial nematodes. Wolbachia are of particular interest because nematodeWolbachia have been shown to cause the diseases African river blindness and Lymphatic Filariasis. Doxycycline can be used to eliminate nematode Wolbachia, however, more efficient treatments are needed. Ideally, we would like to repurpose another FDA approved drug that helps to shorten treatment duration. Vitamins are one of the best classes of FDA approved compounds, generally recognized as safe. Interestingly, prior work by Serbus and colleagues found that dietary yeast, which is highly enriched in vitamins, dramatically reducesWolbachia titer in Drosophila melanogaster ovarian tissue. Imaging data indicated that the Wolbachia nucleoids were disrupted in response to yeast. This raised the possibility that yeast cells contain a bio-reactive, anti-Wolbachiacompound. Our close examination of yeast nutritional information identified which vitamins are most highly enriched in yeast. We then administered several of these to D. melanogaster, and saw that two of these led to reduced ovarianWolbachia titers, analogous to yeast-fed flies. This was especially interesting, as both vitamins are critical for functioning of the same biochemical pathway. We used retested effect of one of these vitamins in oogenesis by performing a dilution series, and achieved positive correlation from this dilution series. This opens up the avenue for clarifying the mechanism of how vitamins suppressWolbachia titer, and for testing enhancement of Doxycycline, to hopefully provide faster, more affordable treatment for millions of patients.