Ex situ culture of coral species Porites lobata (Scleractinia: Poritidae) and Pocillopora damicornis (Scleractinia: Pocilloporidae), Costa Rica: first assessment and implications

Introduction: Coral reefs worldwide decline has prompted coral restoration as a viable strategy to rewild vulnerable, foundational coral species. Stony corals are now propagated by thousands in both in-water and ex situ (land-based) coral nurseries, the latter being unexplored in Costa Rica, despite their potential benefits as a reef management tool. Objective: To analyze the viability of ex situ culturing of the Pacific reef-building corals Porites lobata and Pocillopora damicornis at Parque Marino del Pacífico (PMP), Puntarenas, Costa Rica, aquaculture facilities. Methods: From May to October 2018 a total of 180 coral fragments were kept in an aquaculture recirculated system. Survival, growth, and fragment yield in relation to culture medium (physicochemical parameters) were recorded. Results: Survival and growth rate varied between species and culture tanks. On average, surviving P. lobata fragments (68.89 %) placed in Tank 1 (T1) grew 216 %, while fragments placed in Tank 2 (T2) had a survival rate of 71.11 % and an increase of 277 % in live tissue area. P. damicornis fragments survival, basal and crown area percentage increase were: 71.11 %, 980 % and 366 % in T1, and 100 %, 976 % and 287 % in T2. Although fragments survival and growth were net positive, the yield in terms of culture was low, due to culture conditions in the tanks not meeting coral culture optimal requirements. Conclusions: Survival and growth of both species varied depending on the tank in which they were placed. Survival was similar to that found in other ex situ studies and growth was similar to those reported in the wild, however culture performance in terms of yield was low. Aquaculture systems at PMP constitute a good base for the cultivation of corals, however for the culture effort to achieve maximum yield, current systems must be optimized according to the requirements of the target coral species.

Introducción: El declive mundial de los arrecifes de coral, ha impulsado a la restauración coralina como una estrategia viable para recuperar especies de coral fundacionales, en estado vulnerable. Los corales pétreos se propagan por miles, tanto en viveros subacuáticos como ex situ (en tierra). Siendo el segundo método poco explorado en Costa Rica, a pesar de sus potenciales beneficios como medida como herramienta de manejo arrecifal. Objetivo: Analizar la viabilidad del cultivo ex situ de las especies de coral constructoras de arrecifes Porites lobata y Pocillopora damicornis en el módulo de acuicultura del Parque Marino del Pacífico (PMP), Puntarenas, Costa Rica. Métodos: Desde el 17 de mayo hasta el 17 de octubre de 2018, se mantuvieron un total de 180 fragmentos de coral en un sistema de recirculación de acuicultura. Se registraron la supervivencia, el crecimiento y el rendimiento de los fragmentos en relación con el medio de cultivo (parámetros fisicoquímicos). Resultados: La tasa de supervivencia y crecimiento varió entre especies y tanques de cultivo. En promedio, los fragmentos de P. lobata supervivientes (68.89 %) colocados en el tanque 1 (T1) crecieron un 216 %. En contraste con los fragmentos colocados en el tanque 2 (T2) que mostraron una tasa de supervivencia del 71.11 % y un aumento del 277 % en el área de tejido vivo. En el caso de P. damicornis, los porcentajes de supervivencia, de aumento del área basal y del área de la corona fueron: 71.11 %, 980 %, y 366 %, y 100 %, 976 %, y 287 % para los fragmentos colocados en T1 y T2, respectivamente. Aunque la supervivencia y el crecimiento de los fragmentos fueron positivos, el rendimiento en términos de cultivo fue bajo, debido a que las condiciones en los tanques no cumplían con las condiciones ideales para el cultivo de corales. Conclusiones: La supervivencia y el crecimiento de ambas especies variaron en función del tanque en el que se colocaron. La supervivencia fue similar a la observada en otros estudios ex situ y el crecimiento fue similar al reportado en la naturaleza, pero el rendimiento del cultivo fue bajo. Los sistemas de acuicultura del PMP constituyen una buena base para el cultivo de corales, sin embargo, para que el esfuerzo de cultivo alcance un máximo de rendimiento, los sistemas actuales deben optimizarse en función de los requisitos de las especies de coral objetivo.

Coral micro-fragmentation assays for optimizing active reef restoration efforts.

The global decline of coral reefs has driven considerable interest in active coral restoration. Despite their importance and dominance on mature reefs, relatively few coral restoration projects use slower growth forms like massive and encrusting coral species. Micro-fragmentation can increase coral cover by orders of magnitude faster than natural growth, which now allows cultivation of slow growing massive forms and shows promise and flexibility for active reef restoration. However, the major causes of variation in growth and survival of outplanted colonies remain poorly understood. Here, we report simple outplanting assays to aid in active reef restoration of slower growing species and increase the likelihood of restoration success. We used two different micro-fragmentation assays. Pyramid assays were used to examine variation associated with fragment size (ranging from ≈1-9 cm2), nursery residence time (for both in-situ and ex-situ nurseries), and 2D vs. 3D measurements of growth. Block assays were used to examine spatial variation among individual performance at outplanting sites in the field. We found 2D and 3D measurements correlated well, so measured survivorship and growth using top-down planar images for two of the main Hawaiian reef building corals, the plating Montipora capitata and the massive Porites compressa. Pyramid assays housed and outplanted from the in-situ nursery showed no effect of residence time or size on overall survivorship or growth for either species. Results from the ex-situ nursery, however, varied by species, with P. compressa again showing no effect of nursery residence time or size on survivorship or growth. In contrast, nursery culture resulted in improved survivorship of small M. capitata fragments, but net growth showed a weak positive effect of nursery time for medium fragments. Small fragments still suffered higher mortality than either medium or large fragments. Due to their lower mortality, medium fragments (≈3 cm2) appear to be the best compromise between growth and survivorship for outplanting. Likewise, given weak positive gains relative to the investment, our results suggest that it could be more cost-effective to simply outplant medium fragments as soon as possible, without intermediate culture in a nursery. Furthermore, the block assay revealed significant differences in survivorship and growth among sites for individuals of both species, emphasizing the importance of considering spatial variation in coral survival and growth following outplanting. These results highlight the value of using short-term micro-fragmentation assays prior to outplanting to assess size, and location specific performance, optimizing the efficiency of active reef restoration activities and maximizing the probability of success for active coral restoration projects.

Reduced thermal tolerance of massive coral species in a highly variable environment.

Coral bleaching events are increasing in frequency and severity, resulting in widespread losses in coral cover. However, branching corals native to highly variable (HV) thermal environments can have higher bleaching resistance than corals from more moderate habitats. Here, we investigated the response of two massive corals, Porites lobata and Goniastrea retiformis, from a moderately variable (MV) and a low variability (LV) pool transplanted into a HV pool on Ofu Island in American Samoa. Paired transplant and native ramets were exposed to an acute thermal stress after 6 and 12 months of exposure to the HV pool to evaluate changes in thermal tolerance limits. For both species, photosynthetic efficiency and chlorophyll loss following acute heat stress did not differ between ramets transplanted into the HV pool and respective native pool. Moreover, HV native P. lobata exhibited the greatest bleaching susceptibility compared to MV and LV natives and there was no effect of acute heat stress on MV P. lobata. There was also a thermal anomaly during the study, where Ofu's backreef thermal regime surpassed historical records-2015 had 8 degree heating weeks (DHW) and 2016 had up to 5 DHW (in comparison to less than or equal to 3 over the last 10 years)-which may have exceeded the upper thermal limits of HV native P. lobata. These results strongly contrast with other research on coral tolerance in variable environments, potentially underscoring species-specific mechanisms and regional thermal anomalies that may be equally important in shaping coral responses to extreme temperatures.

Global gene expression patterns in Porites white patch syndrome: Disentangling symbiont loss from the thermal stress response in reef-building coral.

The mechanisms resulting in the breakdown of the coral symbiosis once the process of bleaching has been initiated remain unclear. Distinguishing the process of symbiont loss from the thermal stress response may shed light on the cellular and molecular pathways involved in each process. This study examined physiological changes and global gene expression patterns associated with white patch syndrome (WPS) in Porites lobata, which manifests in localized bleaching independent of thermal stress. In addition, a meta-analysis of global gene expression studies in other corals and anemones was used to contrast differential regulation as a result of disease and thermal stress from patterns correlated with symbiotic state. Symbiont density, chlorophyll a content, holobiont productivity, instant calcification rate, and total host protein content were uniformly reduced in WPS relative to healthy tissue. While expression patterns associated with WPS were secondary to fixed effects of source colony, specific functional enrichments combined with a lack of immune regulation suggest that the viral infection putatively giving rise to this condition affects symbiont rather than host cells. Expression in response to WPS also clustered independently of patterns in white syndrome impacted A. hyacinthus, further supporting a distinct aetiology of this syndrome. Expression patterns in WPS-affected tissues were significantly correlated with prior studies that examined short-term thermal stress responses independent of symbiotic state, suggesting that the majority of expression changes reflect a nonspecific stress response. Across studies, the magnitude and direction of expression change among particular functional enrichments suggests unique responses to stressor duration and highlights distinct responses to bleaching in an anemone model.

Expression and Characterization of a Bright Far-red Fluorescent Protein from the Pink-Pigmented Tissues of Porites lobata.

Members of the anthozoan green fluorescent protein (GFP) family display a diversity of photo-physical properties that can be associated with normal and damaged coral tissues. Poritid coral species often exhibit localized pink pigmentation in diseased or damaged tissues. Our spectral and histological analyses of pink-pigmented Porites lobata lesions show co-localization of bright red fluorescence with putative amoebocytes concentrating in the epidermis, suggesting an activated innate immune response. Here we report the cloning, expression, and characterization of a novel red fluorescent protein (plobRFP) from the pink-pigmented tissues associated with lesions on Porites lobata. In vitro, the recombinant plobRFP exhibits a distinct red emission signal of 614 nm (excitation maximum: 578 nm), making plobRFP the furthest red-shifted natural fluorescent protein isolated from a scleractinian coral. The recombinant protein has a high molar extinction coefficient (84,000 M-1 cm-1) and quantum yield (0.74), conferring a notable brightness to plobRFP. Sequence analysis suggests the distinct brightness and marked red shift may be inherent features of plobRFP's chromophore conformation. While plobRFP displays a tendency to aggregate, its high pH stability, photostability, and spectral properties make it a candidate for cell imaging applications and a potential template for engineering optimized RFPs. The association of plobRFP with a possible immune response furthers its potential use as a visual diagnostic and molecular biomarker for monitoring coral health.

The complete mitochondrial genome of the lobe coral Porites lobata (Anthozoa: Scleractinia) sequenced using ezRAD.

The mitochondrial genome of the coral Porites lobata was sequenced using ezRAD. The assembled genome consists of 18,647 bp, including 13 protein-coding genes, two ribosomal RNA genes and two transfer RNA genes. The gene arrangement was consistent with other scleractinian coral mitochondrial genomes. The sequence was strikingly similar to Porites okinawensis, indicating the necessity for further systematic work to resolve phylogenetic relationships in the genus Porites.

Role of bacterial communities in coral’s defence against a causative agent of coral bleaching: Vibrio coralliilyticus

Aims: Different studies have shown that members of the Vibrio such as Vibrio coralliilyticus and Vibrio shiloi are opportunistic pathogens which can cause coral lysis. The aims of this study were to assess whether this results of the virulence of V. coralliilyticus are transmittable to Acropora hyacinthus and Porites lobata, and what role the microbiome of the corals plays during exposure to V. coralliilyticus. Methodology and results: In laboratory-based experiments, we examined the impact of V. coralliilyticus (ATCC BAA- 450) to the microbiome of Acropora hyacinthus and Porites lobata. A. hyacinthus and P. lobata were exposed to ampicillin, V. coralliilyticus, and a combination of both. Results indicate a resistance of A. hyacinthus to V. coralliilyticus through the microbiome and underpin the importance of the microbiome for the coral’s health. Conclusion, significance and impact study: Further studies are needed to identify the bacteria responsible for the coral resistance and could in future lead to the development of a probiotic treatment or prevention of bleaching for sensitive corals.

Growing coral larger and faster: micro-colony-fusion as a strategy for accelerating coral cover.

Fusion is an important life history strategy for clonal organisms to increase access to shared resources, to compete for space, and to recover from disturbance. For reef building corals, fragmentation and colony fusion are key components of resilience to disturbance. Observations of small fragments spreading tissue and fusing over artificial substrates prompted experiments aimed at further characterizing Atlantic and Pacific corals under various conditions. Small (∼1-3 cm(2)) fragments from the same colony spaced regularly over ceramic tiles resulted in spreading at rapid rates (e.g., tens of square centimeters per month) followed by isogenic fusion. Using this strategy, we demonstrate growth, in terms of area encrusted and covered by living tissue, of Orbicella faveolata, Pseudodiploria clivosa, and Porites lobata as high as 63, 48, and 23 cm(2) per month respectively. We found a relationship between starting and ending size of fragments, with larger fragments growing at a faster rate. Porites lobata showed significant tank effects on rates of tissue spreading indicating sensitivity to biotic and abiotic factors. The tendency of small coral fragments to encrust and fuse over a variety of surfaces can be exploited for a variety of applications such as coral cultivation, assays for coral growth, and reef restoration.

High genotypic diversity of the reef-building coral Porites lobata (Scleractinia: Poritidae) in Isla del Coco National Park, Costa Rica / Alta diversidad genotípica del coral constructor de arrecifes Porites lobata (Scleractinia: Poritidae) en el Parque Nacional Isla del Coco, Costa Rica

The isolated Isla del Coco experiences periodic, extreme disturbances which devastate coral reefs surrounding the island. Scleractinian corals build the physical structure of the reef therefore ecosystem recovery relies on coral species recovery. Coral recruits can be of sexual or asexual origin, and the relative success of the two recruit types influences the speed and spread of recovery processes. Here we focus on the massive coral, Porites lobata, because it is the main reef-builder around Isla del Coco to describe the relative contribution of asexual and sexual recruits to population maintenance. P. lobata samples were collected using a spatially explicit random sampling design in three plots at Isla del Coco: Punta Ulloa (n=17), Bahía Weston (n=20) and Punta María (n=20) and samples were genotyped with 11 microsatellite markers. Additional sampling was conducted at three “coastal” sites near the Costa Rican mainland (Isla del Caño Biological Reserve): Caño1 (n=8), Caño2 (n=10), Caño5 (n=11) to compare the contributions of asexual and sexual recruits at Isla del Coco sites to coastal sites. Isla del Coco sites were characterized by small colony size (>60% of colonies <0.5m2) and high sexual reproduction. Sites were either mostly or entirely sexual,consisting of only unique genotypes (N G/N= 0.90-1.00; G O/G E=0.83-1.00; D=0.99-1.00). Although there were no significant differences in genetic diversity (number of alleles per locus, number of private alleles) or colony size between Isla del Coco and the coastal sites, the coastal sites exhibited a greater range of genotypic diversity from moderately asexual (N G/N=0.5; G O/G E=0.36; D=0.8) to purely sexual (N G/N=1.0; G O/G E=1.0; D=1.0). The mode of asexual reproduction in P. lobata is likely fragmentation of adult colonies rather than asexual larval production because ramets of P. lobata occurred close together and asexually produced larvae have not been reported in gonochoric broadcast spawners like P. lobata. Frequent sexual reproduction at Isla del Coco National Park might represent a resource for rapid recovery following extreme El Niño-Southern Oscillation (ENSO) disturbance events. In contrast, larger, asexually-produced fragments rather than smaller, sexually-produced larvae appear to have the advantage at some coastal sites. The high frequency of sexual reproduction at Isla del Coco indicates that not only are sexual partners available but also current conditions are favorable for the delivery of larvae and the rate of predation on small larval recruits must be moderate.

Los ambientes marinos del Parque Nacional Isla del Coco experimentan perturbaciones extremas periódicamente como por ejemplo El Niño-Oscilación del Sur (ENOS) que han devastado las comunidades coralinas. La cobertura coralino se redujo drásticamente durante El Niño de 1982-83. Los corales escleractinios construyen la estructura física de los arrecifes así que la recuperación de estos ecosistemas depende de que los corales se recuperen. Los corales pueden reproducirse sexual y asexualmente, y el éxito relativo de cada forma de reproducción va a guiar el proceso de recuperación con implicaciones potenciales a la diversidad de las comunidades asociadas. En la mayoría de los arrecifes alrededor de la Isla del Coco, el coral masivo, Porites lobata, es la especie constructora predominante. Si la diversidad genotípica (clonal) de esta especie es alta, la diversidad de individuos va a ser más alta resultando en mayor resilencia frente a condiciones ambientales extremas. Alternativamente, una diversidad genotípica baja es indicativo de una estrategia reproductiva asexual posiblemente resultando en el mantenimiento de genotipos bien adaptados aunque la población decline. Aquí, usamos 11 marcadores microsatélite para investigar la contribución relativa de la reproducción sexual o asexual en la recuperación de los arrecifes en el Parque Nacional Isla del Coco. En la Isla del Coco se recolectaron muestras en: Punta Ulloa (n=17), Bahía Weston (n=20) y Punta María (n=20), y para comparar, se recolectaron muestras en localidades cerca de o en la costa continental de Costa Rica; Reserva Biológica Isla del Caño: Caño1 (n=8), Caño2 (n=10) y Caño5 (n=11), y Tres Hermanas, Parque Nacional Marino Ballena (n=4), utilizando un diseño de muestreo espacialmente explícito. Las colonias de la Isla del Coco son generalmente pequeñas (>60% de las colonias <0.5m2) y se observó poca reproducción asexual. En la mayoría de los sitios la reproducción era mayormente sexual (NG/N= 0.90-0.94; GO/GE =0.63-0.74; D=0.99) o totalmente sexual (N G/N= 1.0; G O/G E =1.0; D=1.0), por lo que consiste decolonias con genotipos únicos. En contraste, los sitios costeros tenían un ámbito de predominantemente asexual y genotípicamente pobre (N G/N =0.5; G O/G E =0.11; D=0.5) a totalmente sexual (N G/N= 1.0; G O/G E =1.0; D=1.0). No hubo diferencia en diversidad genética (número de alelos por locus, número de alelos privados) o distribución de tamaño por región. La alta diversidad genotípica del Parque Nacional Isla del Coco indica reproducción sexual frecuente y el potencial para la recuperación rápida de los arrecifes después de perturbaciones severas. En los sitios costeros, la fragmentación asexual de colonias grandes (>2.5m2) y por lo tanto, potencialmente colonias bien adaptadas, puede aumentar las resistencia y resilencia de los sistemas.