Cory J. Krediet

The tropical sea anemone Exaiptasia diaphana (commonly referred to as 'Aiptasia'), serves as a model system for the coral holobiont. Previous research has shown that exposure to elevated salinity conveyed increased thermotolerance in Aiptasia. Based on this observation, we aimed to determine if acclimation to elevated salinity could lead to improved pathogen resilience. Aiptasia were acclimated to seawater at either 35 ppt or 42 ppt over a 'short-term' period (14-16 days) or 'long-term' period (90 + days). After the acclimation period, Aiptasia were exposed to Serratia marcescens PDL100 (a known coral pathogen) at 108 CFU/mL, 107 CFU/mL, or remained un-exposed. The exposed anemones were monitored for 10 days, and their mortality was scored daily. Aiptasia acclimated to 42 ppt, over both short- and long-term periods, exhibited a higher survival than that of control anemones. We then tested whether pre-exposure to sub-lethal concentra-tions of S. marcescens could lead to increased survival rates in comparison to naive anemones at both salinities. This resulted in 100% survival of the anemones exposed to the pathogen, suggesting priming of the Aiptasia immune system in response to exposure to non-lethal concentrations of the pathogen. Evidence of immune priming was confirmed by dif-ferential gene expression of known stress-response genes by qPCR. Our results show, for the first time, increased pathogen resilience due to exposure to elevated salinity in Aiptasia. Since our results are based on a salinity effect in symbiotic sea anemones, it remains unclear whether this salinity effect might also occur in stony corals.

Corals and dinoflagellate algae form a unique mutualistic symbiosis that provides the energetic and structural foundation for shallow coral reef ecosystems. Despite the long success of this partnership in oligotrophic seas, coral reefs are in decline due to increasing threats from rising seawater temperatures and disease, both of which can lead to bleaching and mortality. In order to better understand the mechanisms that underpin this mutualism, it may be necessary to dismantle the coral-algal symbiosis. Previous studies have experimentally bleached corals using thermal stress, photosynthetic inhibitors (DCMU), and menthol. We compared lab-induced bleaching of staghorn coral Acropora cervicornis by menthol treatment to traditional thermal stress. The larger aim was to adapt existing bleaching protocols for this important coral species, providing a guide for future studies. Bleaching in corals treated with menthol or exposed to elevated temperature stress (31°C) was monitored by measuring photosynthetic activity determined by Fv/Fm using pulse-amplitude modulated (PAM) fluorescence and compared to untreated conspecifics. Corals were also monitored for symbiont density and overall health using the CoralWatch Coral Health Chart card throughout the experiment. We found that A. cervicornis bleached in response to both menthol treatment and thermal stress, but menthol treatment was more effective at reducing algal symbiont photosynthetic capacity (Fv/Fm) without negatively affecting the health of the coral. Our results indicate that menthol treatment at 0.38 mM rendered staghorn coral aposymbiotic within fourteen days without any visual or physiological damage to the coral. This study provides a simple and effective menthol-bleaching treatment protocol for future studies on staghorn coral.

Symbiotic cnidarians such as corals and anemones form highly productive and biodiverse coral reef ecosystems in nutrient-poor ocean environments, a phenomenon known as Darwin’s paradox. Resolving this paradox requires elucidating the molecular bases of efficient nutrient distribution and recycling in the cnidarian-dinoflagellate symbiosis. Using the sea anemone Aiptasia, we show that during symbiosis, the increased availability of glucose and the presence of the algae jointly induce the coordinated up-regulation and relocalization of glucose and ammonium transporters. These molecular responses are critical to support symbiont functioning and organism-wide nitrogen assimilation through glutamine synthetase/glutamate synthase–mediated amino acid biosynthesis. Our results reveal crucial aspects of the molecular mechanisms underlying nitrogen conservation and recycling in these organisms that allow them to thrive in the nitrogen-poor ocean environments. Whole-organism nitrogen assimilation fueled by glucose from symbiotic algae enables corals to flourish in oligotrophic waters.

The sensitivity of reef-building coral to elevated temperature is a function of their symbiosis with dinoflagellate algae in the family Symbiodiniaceae. Changes in the composition of the endosymbiont community in response to thermal stress can increase coral thermal tolerance. Consequently, this mechanism is being investigated as a human-assisted intervention for rapid acclimation of coral in the face of climate change. Successful establishment of novel symbioses that increase coral thermal tolerance have been demonstrated in laboratory conditions; however, it is unclear how long these heterologous relationships persist in nature. Here, we test the persistence of a novel symbiosis between Acropora palmata and Durusdinium spp. from Mote Marine Laboratory's ex situ nursery by outplanting clonal replicates (ramets) of five A. palmata host genotypes to natural reefs in the lower Florida Keys. Amplicon sequencing analysis of ITS2-type profiles revealed that the majority of surviving ramets remained dominated by Durusdinium spp. two years after transplantation. However, 15% of ramets, including representatives of all genotypes, exhibited some degree of symbiont shuffling or switching at six of eight sites, including complete takeover by site-specific strains of the native symbiont, Symbiodinium fitti. The predominant long-term stability of the novel symbiosis supports the potential effectiveness of symbiont modification as a management tool. Although, the finding that 6-7 year-old coral can alter symbiont community composition in the absence of bleaching indicates that Symbiodiniaceae communities are indeed capable of great flexibility under ambient conditions.

Genotype-by-environment interactions (GxE) indicate that variation in organismal traits cannot be explained by fixed effects of genetics or site-specific plastic responses alone. For tropical coral reefs experiencing dramatic environmental change, identifying the contributions of genotype, environment, and GxE on coral performance will be vital for both predicting persistence and developing restoration strategies. We quantified the impacts of G, E, and GxE on the morphology and survival of the endangered coral, , through an in situ transplant experiment exposing common garden (nursery)-raised clones of ten genotypes to nine reef sites in the Florida Keys. By fate-tracking outplants over one year with colony-level 3D photogrammetry, we uncovered significant GxE on coral size, shape, and survivorship, indicating that no universal winner exists in terms of colony performance. Rather than differences in mean trait values, we found that individual-level morphological plasticity is adaptive in that the most plastic individuals also exhibited the fastest growth and highest survival. This indicates that adaptive morphological plasticity may continue to evolve, influencing the success of and resulting reef communities in a changing climate. As focal reefs are active restoration sites, the knowledge that variation in phenotype is an important predictor of performance can be directly applied to restoration planning. Taken together, these results establish as a system for studying the ecoevolutionary dynamics of phenotypic plasticity that also can inform genetic- and environment-based strategies for coral restoration.

Microbiome studies across taxa have established the influence of host genotype on microbial recruitment and maintenance. However, research exploring host-specific epibionts in scleractinian corals is scant, and the influence of intraspecific differences across environments remains unclear. Here, we studied ten Acropora cervicornis genotypes to investigate the relative roles of host genotype and environment in structuring the epibiome. Coral mucus was sampled in a common garden nursery from replicate ramets of distinct genotypes (T-0). Coral fragment replicates (n = 3) of each genotype were then transplanted to nine different field sites in the Lower Florida Keys, and mucus was again sampled one year later from surviving ramets (T-12). 16S rRNA amplicon sequencing was used to assess microbial composition, richness, and beta-diversity. The most abundant and consistent amplicon sequencing variants (ASVs) in all samples belonged to Midichloriaceae (MD3-55 genus) and Cyanobacteria (Synechococccus). The relative abundances of these bacterial taxa varied consistently between genotypes, whereas neither the composition nor taxonomic relative abundance were significantly different among field sites. Interestingly, several high MD3-55 hosting genotypes showed rapid diversification and an increase in MD3-55 following transplantation. Overall, our results indicate healthy A. cervicornis genotypes retain distinct epibiome signatures through time, suggesting a strong host component. Lastly, our results show that differences in MD3-55 abundances can be consistently detected in the epibiome of distinct host genotypes of A. cervicornis. As this organism (sensu Aquarickettsia rohweri) has been implicated as a marker of disease resistance, this finding reinforces the potential use of microbial indicators in reef restoration efforts via non-invasive surface/mucus sampling.

Loss of endosymbiotic algae ("bleaching") under heat stress has become a major problem for reef-building corals worldwide. To identify genes that might be involved in triggering or executing bleaching, or in protecting corals from it, we used RNAseq to analyze gene-expression changes during heat stress in a coral relative, the sea anemone Aiptasia. We identified >500 genes that showed rapid and extensive up-regulation upon temperature increase. These genes fell into two clusters. In both clusters, most genes showed similar expression patterns in symbiotic and aposymbiotic anemones, suggesting that this early stress response is largely independent of the symbiosis. Cluster I was highly enriched for genes involved in innate immunity and apoptosis, and most transcript levels returned to baseline many hours before bleaching was first detected, raising doubts about their possible roles in this process. Cluster II was highly enriched for genes involved in protein folding, and most transcript levels returned more slowly to baseline, so that roles in either promoting or preventing bleaching seem plausible. Many of the genes in clusters I and II appear to be targets of the transcription factors NFκB and HSF1, respectively. We also examined the behavior of 337 genes whose much higher levels of expression in symbiotic than aposymbiotic anemones in the absence of stress suggest that they are important for the symbiosis. Unexpectedly, in many cases, these expression levels declined precipitously long before bleaching itself was evident, suggesting that loss of expression of symbiosis-supporting genes may be involved in triggering bleaching.

The bacterium Vibrio coralliilyticus has been implicated as the causative agent of coral tissue loss diseases (collectively known as white syndromes) at sites across the Indo-Pacific and represents an emerging model pathogen for understanding the mechanisms linking bacterial infection and coral disease. In this study, we used a mini-Tn7 transposon delivery system to chromosomally label a strain of V. coralliilyticus isolated from a white syndrome disease lesion with a green fluorescent protein gene (GFP). We then tested the utility of this modified strain as a research tool for studies of coral host–pathogen interactions. A suite of biochemical assays and experimental infection trials in a range of model organisms confirmed that insertion of the GFP gene did not interfere with the labeled strain’s virulence. Using epifluorescence video microscopy, the GFP-labeled strain could be reliably distinguished from non-labeled bacteria present in the coral holobiont, and the pathogen’s interactions with the coral host could be visualized in real time. This study demonstrates that chromosomal GFP labeling is a useful technique for visualization and tracking of coral pathogens and provides a novel tool to investigate the role of V. coralliilyticus in coral disease pathogenesis.