Evaluation of fluorescence-based viability stains in cells dissociated from scleractinian coral Pocillopora damicornis
Liza M. RogerYaa Adarkwa DarkoTytus BernaśFrances J. WhiteMonsurat OlaosebikanLenore CowenJudith Klein‐SeetharamanNastassja A. Lewinski
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Abstract The application of established cell viability assays such as the commonly used trypan blue staining method to coral cells is not straightforward due to different culture parameters and different cellular features specific to mammalian cells compared to marine invertebrates. Using Pocillopora damicornis as a model, we characterized the autofluorescence and tested different fluorescent dye pair combinations to identify alternative viability indicators. The cytotoxicity of different representative molecules, namely small organic molecules, proteins and nanoparticles (NP), was measured after 24 h of exposure using the fluorescent dye pair Hoechst 33342 and SYTOX orange. Our results show that this dye pair can be distinctly measured in the presence of fluorescent proteins plus chlorophyll. P. damicornis cells exposed for 24 h to Triton-X100, insulin or titanium dioxide (TiO 2 ) NPs, respectively, at concentrations ranging from 0.5 to 100 µg/mL, revealed a LC50 of 0.46 µg/mL for Triton-X100, 6.21 µg/mL for TiO 2 NPs and 33.9 µg/mL for insulin. This work presents the approach used to customize dye pairs for membrane integrity-based cell viability assays considering the species- and genotype-specific autofluorescence of scleractinian corals, namely: endogenous fluorescence characterization followed by the selection of dyes that do not overlap with endogenous signals.Keywords:
Pocillopora damicornis
Summary A few studies have holistically examined successive changes in coral holobionts in response to increased temperatures. Here, responses of the coral host Pocillopora damicornis , its Symbiodiniaceae symbionts, and associated bacteria to increased water temperatures were investigated. High temperatures induced bleaching, but no coral mortality was observed. Transcriptome analyses showed that P . damicornis responded more quickly to elevated temperatures than its algal symbionts. Numerous genes putatively associated with apoptosis, exocytosis, and autophagy were upregulated in P . damicornis , suggesting that Symbiodiniaceae can be eliminated or expelled through these mechanisms when P . damicornis experiences heat stress. Furthermore, apoptosis in P . damicornis is presumably induced through tumour necrosis factor and p53 signalling and caspase pathways. The relative abundances of several coral disease‐associated bacteria increased at 32°C, which may affect immune responses in heat‐stressed corals and potentially accelerates the loss of algal symbionts. Additionally, consistency of Symbiodiniaceae community structures under heat stress suggests non‐selective loss of Symbiodiniaceae. We propose that heat stress elicits interrelated response mechanisms in all parts of the coral holobiont.
Pocillopora damicornis
Holobiont
Coral bleaching
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Temporal patterns in the planulation of the scleractinian coral, Pocillopora damicornis (Linnaeus 1816), common on Singapore reefs were observed from August to November 1990.Widely distributed throughout the Indo-Pacific, the species is known to release planula larvae after internal fertilisation.At any one time, six colonies collected from the field were maintained in clear, sheltered outdoor aquaria and daily planula counts observed.Planulation was observed to be directly influenced by the lunar cycle, commencing 2 to 3 days before the new moon, peaking at around the new moon and ending close to the first quarter.This periodicity is similar to the planulation patterns observed in P. damicornis populations of the Central Pacific, particularly those of Palau, as well as the populations in the Great Barrier Reef during the Austral summer.
Pocillopora damicornis
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Pocillopora damicornis
Hermatypic coral
Zooxanthellae
Porites
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Roseobacter
Alphaproteobacteria
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This study investigates the effect of hyposalinity on coral bleaching in a species of hard coral (Pocillopora damicornis) and a soft coral (Sarcophyton sp.). Corals were exposed to ambient seawater (32 psu) and reduced-salinity seawater (15 psu and 25 psu) for 32 days. Both the hard coral and soft coral were highly affected by the salinity level of 15 psu. Fragments of Pocillopora damicornis were 90% bleached within four days and died after five days of exposure to salinity levels of 15 psu. Similarly, Sarcophyton sp. was 50%-95% bleached within four days and died after seven days of exposure. Both corals were less impacted by exposure to salinity levels of 25 psu and 32 psu; P. damicornis and Sarcophyton sp. were partially bleached but survived until the end of the experiment. We report that P. damicornis is more susceptible to hyposalinity than Sarcophyton sp. The results of this study can serve as basic information for managers regarding the release of freshwater into areas of the ocean with coral reefs. The study also suggests that salinity monitoring during rainy seasons or flooding may allow prediction of future coral bleaching caused by these low-salinity events.
Keywords: rainfall, bleaching, scleractinian, soft corals, hyposalinity
Pocillopora damicornis
Coral bleaching
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Ultraviolet radiation (UVR, 280-400 nm) is one of the potential factors involved in the induction of coral bleaching, loss of the endosymbiotic dinoflagellate Symbiodinium or their photosynthetic pigments. However, little has been documented on its effects on the behavior and recruitment of coral larvae, which sustains coral reef ecosystems. Here, we analyzed physiological changes in larvae of the scleractinian coral Pocillopora damicornis and examined the photophysiological performance of the symbiont algae, following exposure to incident levels of UVR and subsequently observed the development of coral larvae. The endosymbiotic algae exhibited a high sensitivity to UV-B (295-320 nm) during a 6 h exposure, showing lowered photosynthetic performance per larva and per algal cell, whereas the presence of UV-A (320-395 nm) significantly stimulated photosynthesis. UVR decreased chlorophyll a concentration only at higher surface temperature or at the higher doses or intensities of UVR. Correlations between UV-absorbing compound (UVAC) contents or UVR sensitivity and temperature were identified, implying that UVACs might act as a screen or antioxidants in Pocillopora damicornis larvae. Larvae reared under UVR exposures showed lower levels of survivorship, metamorphosis and settlement, with inhibition by UV-A being much greater than that caused by UV-B.
Pocillopora damicornis
Symbiodinium
Zooxanthellae
Dinoflagellate
Coral bleaching
Mariculture
Photoprotection
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AbstractHeterotrophy is essential for meeting the metabolic requirements of scleractinian corals but the feeding mechanisms of coral juveniles are poorly understood. In this study, we observed that two-day-old Pocillopora damicornis recruits were able to immobilize and capture Artemia salina nauplii that come into direct contact with their tentacles. The recruits were able to consume up to six nauplii h−1 and the shortest time required to fully ingest one nauplius was 1 min 25 s. This is the first study to describe the feeding mechanisms employed by coral recruits, and provided both observational and quantitative evidence to show that coral juveniles are able to commence heterotrophy at an early stage.Keywords: coralscleractinianjuvenilefeeding strategieszooplanktonArtemia salinavideo sequences
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