Ocean acidification stunts molluscan growth at CO2 seeps.

Ocean acidification can severely affect bivalve molluscs, especially their shell calcification. Assessing the fate of this vulnerable group in a rapidly acidifying ocean is therefore a pressing challenge. Volcanic CO2 seeps are natural analogues of future ocean conditions that offer unique insights into the scope of marine bivalves to cope with acidification. Here, we used a 2-month reciprocal transplantation of the coastal mussel Septifer bilocularis collected from reference and elevated pCO2 habitats to explore how they calcify and grow at CO2 seeps on the Pacific coast of Japan. We found significant decreases in condition index (an indication of tissue energy reserves) and shell growth of mussels living under elevated pCO2 conditions. These negative responses in their physiological performance under acidified conditions were closely associated with changes in their food sources (shown by changes to the soft tissue δ13C and δ15N ratios) and changes in their calcifying fluid carbonate chemistry (based on shell carbonate isotopic and elemental signatures). The reduced shell growth rate during the transplantation experiment was further supported by shell δ13C records along their incremental growth layers, as well as their smaller shell size despite being of comparable ontogenetic ages (5-7 years old, based on shell δ18O records). Taken together, these findings demonstrate how ocean acidification at CO2 seeps affects mussel growth and reveal that lowered shell growth helps them survive stressful conditions.

Contrasting life-history responses to climate variability in eastern and western North Pacific sardine populations.

Massive populations of sardines inhabit both the western and eastern boundaries of the world's subtropical ocean basins, supporting both commercial fisheries and populations of marine predators. Sardine populations in western and eastern boundary current systems have responded oppositely to decadal scale anomalies in ocean temperature, but the mechanism for differing variability has remained unclear. Here, based on otolith microstructure and high-resolution stable isotope analyses, we show that habitat temperature, early life growth rates, energy expenditure, metabolically optimal temperature, and, most importantly, the relationship between growth rate and temperature are remarkably different between the two subpopulations in the western and eastern North Pacific. Varying metabolic responses to environmental changes partly explain the contrasting growth responses. Consistent differences in the life-history traits are observed between subpopulations in the western and eastern boundary current systems around South Africa. These growth and survival characteristics can facilitate the contrasting responses of sardine populations to climate change.

Metatranscriptomic Analysis of Corals Inoculated With Tolerant and Non-Tolerant Symbiont Exposed to High Temperature and Light Stress.

Algal symbionts of corals can influence host stress resistance; for example, in the Pacific Ocean, whereas Cladocopium (C-type) is generally dominant in corals, Durusdinium (D-type) is found in more heat-resistant corals. Thus, the presence of D-type symbiont likely increases coral heat tolerance, and this symbiotic relationship potentially provides a hint to increase the stress tolerance of coral-algal symbioses. In this study, transcriptome profiles of Cladocopium- and Durusdinium-harboring Acropora solitaryensis (C-coral and D-coral, respectively) and algal photosystem functioning (F v /F m ) under bleaching conditions (high temperature and light stress) were compared. Stress treatment caused algal photoinhibition that the F v /F m value of Symbiodiniaceae was immediately reduced. The transcriptome analysis of corals revealed that genes involved in the following processes were detected: endoplasmic reticulum (ER) stress, mitophagy, apoptosis, endocytosis, metabolic processes (acetyl-CoA, chitin metabolic processes, etc.), and the PI3K-AKT pathway were upregulated, while DNA replication and the calcium signaling pathway were downregulated in both C- and D-corals. These results suggest that unrepaired DNA and protein damages were accumulated in corals under high temperature and light stress. Additionally, some differentially expressed genes (DEGs) were specific to C- or D-corals, which includes genes involved in transient receptor potential (TRP) channels and vitamin B metabolic processes. Algal transcriptome analysis showed the increased expression of gene encoding photosystem and molecular chaperone especially in D-type symbiont. The transcriptome data imply a possible difference in the stress reactions on C-type and D-type symbionts. The results reveal the basic process of coral heat/light stress response and symbiont-type-specific coral transcriptional responses, which provides a perspective on the mechanisms that cause differences in coral stress tolerance.

Sulfur assimilation in corals with aposymbiotic and symbiotic zooxanthellae.

Although sulfate ions are the main form of sulfur in the ocean, there is limited knowledge on their use by living organisms. Stable isotope labelling and NanoSIMS analysis were used in this study to clarify how sulfate, in seawater, is assimilated by corals and zooxanthellae at the cellular level. Aposymbiotic and symbiotic coral juveniles from the genus Acropora were incubated for 2 days in filtered seawater with 34 S-labelled sulfate. Further, the labelled corals were incubated for additional 2 days in natural seawater. Mapping of sulfur isotopes (34 S/32 S) showed that the 'hotspots' were enriched in 34 S on a sub-micro level and were heterogeneously distributed in the coral soft tissues. Specifically, 34 S hotspots were found in both the symbiotic zooxanthellae and coral host tissues. In aposymbiotic corals, 34 S was detected in the tissues, indicating that the host corals directly assimilated the sulfate ions without any aid from the zooxanthellae. Even after 2 days in normal seawater, the 34 S label was clearly seen in both symbiotic and aposymbiotic corals, indicating that the assimilated sulfur was retained for at least 2 days.

Identification of timing of scallop morphological deformity and mortality from shell oxygen isotope records.

The Yesso scallop, Patinopecten yessoensis (Jay), is one of the most important bivalve species in the Japanese and Chinese mariculture industry. In recent years, however, high incidences of scallop shell deformity and mortality have occurred with increasing frequency, but timing of onset and underlying causes are often unclear. Here, we proposed a promising δ18Oshell-based method for constraining the onset of shell deformity and mortality of P. yessoensis. Following six months of intermediate suspension culture in Funka Bay, Northern Japan, shells from healthy, deformed and dead scallops were randomly sampled. High-resolution seawater temperature time-series computed from healthy scallop shell δ18O profiles were precisely and temporally aligned to the instrumental temperature curve, thus allowing δ18Oshell-derived temperature time-series from deformed and dead scallops to be contextualized and allowing timing of scallop deformity and death to be retrieved. Irrespective of scallop shell length, onsets of deformity were anchored in February, and since then deformed scallops grew slowly in comparison to healthy individuals. Without exception, however, dead scallops had already ceased their shell building and died before February, indicating different underlying causes of scallop deformity and mortality. Perhaps most promisingly, considering that shells do not have any isotopic turn-over and once formed, temperature information is locked in. Thus, this approach holds great promise for identifying time anchor points (onsets of deformity and death) in archived scallops collected over different time scales, especially during massive mortality events.

Differential gene expression in skeletal organic matrix proteins of scleractinian corals associated with mixed aragonite/calcite skeletons under low mMg/Ca conditions.

Although coral skeletons generally comprise aragonite crystals, changes in the molar Mg/Ca ratio (mMg/Ca) in seawater result in the incorporation of calcite crystals. The formation mechanism of aragonite and calcite crystals in the scleractinian coral Acropora tenuis was therefore investigated by RNA-seq analysis, using early growth stage calcite (mMg/Ca = 0.5) and aragonite (mMg/Ca = 5.2)-based corals. As a result, 1,287 genes were up-regulated and 748 down-regulated in calcite-based corals. In particular, sixty-eight skeletogenesis-related genes, such as ectin, galaxin, and skeletal aspartic acid-rich protein, were detected as up-regulated, and six genes, such as uncharacterized skeletal organic matrix protein 5, down-regulated, in low-Mg/Ca conditions. Since the number of down-regulated genes associated with the skeletal organic matrix of aragonite skeletons was much lower than that of up-regulated genes, it is thought that corals actively initiate construction of an aragonite skeleton by the skeletal organic matrix in low-Mg/Ca conditions. In addition, different types of skeletal organic matrix proteins, extracellular matrix proteins and calcium ion binding proteins appeared to change their expression in both calcite-formed and normal corals, suggesting that the composition of these proteins could be a key factor in the selective formation of aragonite or calcite CaCO3.

Retrospective monitoring of salinity in coastal waters with mussel shells.

Sea surface salinity (SSS) is a key parameter to understand and predict many physical, chemical and biological processes in dynamic coastal environments. Yet, in many regions, instrumental measurements are spatially sparse and insufficiently long, hindering our ability to document changes, causes, and consequences of SSS across different time scales. Therefore, there is an need to develop a robust proxy to extend SSS records back in time. Here, we test whether SSS can be reconstructed reliably and quantitatively from shell oxygen isotopic ratios (δ18Oshell) of the mussel Mytilus galloprovincialis (Lamarck, 1819) in Otsuchi Bay, Northern Japan. δ18Oshell ratios vary spatially and temporally and exhibit strong linear correlations with both sea surface temperature (SST) and SSS measurements, indicating that the composite signal recorded by δ18Oshell measurably responds to variations in both parameters. By combining contemporaneous variations of SST and δ18Oshell, SSS records encoded into mussel shells are deconvolved that significantly correlate with in situ SSS values. To further validate the robustness of δ18Oshell as a quantitative SSS proxy, high-resolution and temporally aligned time-series of δ18Oshell-derived SSS are reconstructed that are highly synchronous with the instrumental records. In particular, two lowered SSS scenarios occur concomitantly with periods of the summer monsoon and typhoon events. δ18Oshell-derived SSS time-series are also comparable to those from numerical modeling. In conclusion, our findings demonstrate that mussel δ18Oshell signatures can be used as a useful tool to construct high-resolution records of SSS in the coastal regions.

Sulfur utilization of corals is enhanced by endosymbiotic algae.

Sulfur-containing compounds are important components of all organisms, but few studies have explored sulfate utilization in corals. Our previous study found that the expression of a sulfur transporter (SLC26A11) was upregulated in the presence of Symbiodinium cells in juveniles of the reef-building coral Acropora tenuis In this study, we performed autoradiography using (35)S-labeled sulfate ions ((35)SO4  (2-)) to examine the localization and amount of incorporated radioactive sulfate in the coral tissues and symbiotic algae. Incorporated (35)SO4  (2-) was detected in symbiotic algal cells, nematocysts, ectodermal cells and calicoblast cells. The combined results of (35)S autoradiography and Alcian Blue staining showed that incorporated (35)S accumulated as sulfated glycosaminoglycans (GAGs) in the ectodermal cell layer. We also compared the relative incorporation of (35)SO4  (2-) into coral tissues and endosymbiotic algae, and their chemical fractions in dark versus light (photosynthetic) conditions. The amount of sulfur compounds, such as GAGs and lipids, generated from (35)SO4  (2-) was higher under photosynthetic conditions. Together with the upregulation of sulfate transporters by symbiosis, our results suggest that photosynthesis of algal endosymbionts contributes to the synthesis and utilization of sulfur compounds in corals.

Different Stress Tolerances of Juveniles of the Coral Acropora tenuis Associated with Clades C1 and D Symbiodinium.

Ikuko Yuyama, Takashi Nakamura, Tomihiko Higuchi, and Michio Hidaka (2016) Reef-building corals are often associated with multiple clades of symbiotic dinoflagellate Symbiodinium spp., where the relative composition of Symbiodinium can alter the phylogenetic properties (e.g., stress responsiveness, growth rate) of the host coral. The genus Symbiodinium contains nine clades, some of which behave differently in response to strong light and/or temperature stresses, for example, clade D Symbiodinium are thermally tolerant. However, previous studies are based on corals present in the field, and it is possible that the corals used in previous experiments did not contain single Symbiodinium clades. For an accurate assessment of the effects of each Symbiodinium clade on host thermal stress resistance, monoclonal cultures of clades C1 and D were inoculated into aposymbiotic juvenile polyps. Photosynthetic efficiency (maximum quantum yield: F v /F m) showed a decline at 30°C than at 25°C in both clades. Symbiodinium clade C1 showed a consistently higher rETRmax with larger fluctuations than clade D, with a lower survival rate of juveniles during thermal stress treatment. Under strong light exposure, corals containing clade C1 showed a greater decline in F v /F m (-74%), compared to decline in corals associated with clade D (-50%) after 3 hours. This is the first study to assess stress tolerances of juvenile corals in association with the monoclonal Symbiodinium clades C and D, and our results indicated greater tolerance of corals associated with clade D to strong light (500 µmol m-2 s-1). However, it is difficult to determine the impact of high-temperature stress on coral-algae symbiosis from photosynthetic activity. At high temperatures, clade C1 Symbiodinium exhibited high photosynthetic activity, but host survival rates were higher in corals associated with clade D Symbiodinium. Since clade C1 has a relatively high photosynthetic activity under high temperatures, clade C1 symbiosis at high temperatures might have a negative impact on corals compared with clade D.

The northern limit of corals of the genus Acropora in temperate zones is determined by their resilience to cold bleaching.

The distribution of corals in Japan covers a wide range of latitudes, encompassing tropical to temperate zones. However, coral communities in temperate zones contain only a small subset of species. Among the parameters that determine the distribution of corals, temperature plays an important role. We tested the resilience to cold stress of three coral species belonging to the genus Acropora in incubation experiments. Acropora pruinosa, which is the northernmost of the three species, bleached at 13 °C, but recovered once temperatures were increased. The two other species, A. hyacinthus and A. solitaryensis, which has a more southerly range than A. pruinosa, died rapidly after bleaching at 13 °C. The physiological effects of cold bleaching on the corals included decreased rates of photosynthesis, respiration, and calcification, similar to the physiological effects observed with bleaching due to high temperature stress. Contrasting hot bleaching, no increases in antioxidant enzyme activities were observed, suggesting that reactive oxygen species play a less important role in bleaching under cold stress. These results confirmed the importance of resilience to cold stress in determining the distribution and northern limits of coral species, as cold events causing coral bleaching and high mortality occur regularly in temperate zones.

Comparing the effects of symbiotic algae (Symbiodinium) clades C1 and D on early growth stages of Acropora tenuis.

Reef-building corals switch endosymbiotic algae of the genus Symbiodinium during their early growth stages and during bleaching events. Clade C Symbiodinium algae are dominant in corals, although other clades - including A and D - have also been commonly detected in juvenile Acroporid corals. Previous studies have been reported that only molecular data of Symbiodinium clade were identified within field corals. In this study, we inoculated aposymbiotic juvenile polyps with cultures of clades C1 and D Symbiodinium algae, and investigated the different effect of these two clades of Symbiodinium on juvenile polyps. Our results showed that clade C1 algae did not grow, while clade D algae grew rapidly during the first 2 months after inoculation. Polyps associated with clade C1 algae exhibited bright green fluorescence across the body and tentacles after inoculation. The growth rate of polyp skeletons was lower in polyps associated with clade C1 algae than those associated with clade D algae. On the other hand, antioxidant activity (catalase) of corals was not significantly different between corals with clade C1 and clade D algae. Our results suggested that clade D Symbiodinium algae easily form symbiotic relationships with corals and that these algae could contribute to coral growth in early symbiosis stages.

Biotic control of skeletal growth by scleractinian corals in aragonite-calcite seas.

Modern scleractinian coral skeletons are commonly composed of aragonite, the orthorhombic form of CaCO3. Under certain conditions, modern corals produce calcite as a secondary precipitate to fill pore space. However, coral construction of primary skeletons from calcite has yet to be demonstrated. We report a calcitic primary skeleton produced by the modern scleractinian coral Acropora tenuis. When uncalcified juveniles were incubated from the larval stage in seawater with low mMg/Ca levels, the juveniles constructed calcitic crystals in parts of the primary skeleton such as the septa; the deposits were observable under Raman microscopy. Using scanning electron microscopy, we observed different crystal morphologies of aragonite and calcite in a single juvenile skeleton. Quantitative analysis using X-ray diffraction showed that the majority of the skeleton was composed of aragonite even though we had exposed the juveniles to manipulated seawater before their initial crystal nucleation and growth processes. Our results indicate that the modern scleractinian coral Acropora mainly produces aragonite skeletons in both aragonite and calcite seas, but also has the ability to use calcite for part of its skeletal growth when incubated in calcite seas.

The effects of thermal and high-CO2 stresses on the metabolism and surrounding microenvironment of the coral Galaxea fascicularis.

The effects of elevated temperature and high pCO2 on the metabolism of Galaxea fascicularis were studied with oxygen and pH microsensors. Photosynthesis and respiration rates were evaluated from the oxygen fluxes from and to the coral polyps. High-temperature alone lowered both photosynthetic and respiration rates. High pCO2 alone did not significantly affect either photosynthesis or respiration rates. Under a combination of high-temperature and high-CO2, the photosynthetic rate increased to values close to those of the controls. The same pH in the diffusion boundary layer was observed under light in both (400 and 750 ppm) CO2 treatments, but decreased significantly in the dark as a result of increased CO2. The ATP contents decreased with increasing temperature. The effects of temperature on the metabolism of corals were stronger than the effects of increased CO2. The effects of acidification were minimal without combined temperature stress. However, acidification combined with higher temperature may affect coral metabolism due to the amplification of diel variations in the microenvironment surrounding the coral and the decrease in ATP contents.

Montiporic acid D, a new polyacetylene carboxylic acid from scleractinian coral Montipora digitata.

A new polyacetylene carboxylic acid named montiporic acid D (1) was isolated along with a known polyacetylene alcohol, (Z)-13,15-hexadecadien-2,4-diyn-1-ol (2) from scleractinian coral Montipora digitata. The structures of compounds were determined by analyses of NMR and MS spectra.

Photochemical formation of hydroxyl radicals in tissue extracts of the coral Galaxea fascicularis.

Various stresses induce the formation of reactive oxygen species (ROS) in biological cells. In addition to stress-induced ROS, we studied the photochemical formation of hydroxyl radicals (˙OH), the most potent ROS, in coral tissues using phosphate buffer-extracted solutions and a simulated sunlight irradiation system. ˙OH formation was seen in extracts of both coral host and endosymbiont zooxanthellae. This study is the first to report quantitative measurements of ˙OH photoformation in coral tissue extracts. Our results indicated that whether or not coral bleaching occurred, coral tissues and symbiotic zooxanthellae have the potential to photochemically produce ˙OH under sunlight. However, no significant difference was found in the protein content-normalized formation rates of ˙OH between corals incubated under different temperatures and irradiance conditions. ˙OH formation rates were reduced by 40% by reducing the UV radiation in the illumination. It was indicated that UV radiation strongly affected ˙OH formation in coral tissue and zooxanthellae, in addition to its formation through photoinhibition processes.

Chromone and chromanone glucosides from Hypericum sikokumontanum and their anti-Helicobacter pylori activities.

Chromone glucosides, takanechromones A-C (1, 2 and 5) and chromanone glucosides, named takanechromanones A and B (3 and 4), were isolated from the methanolic extracts of Hypericumsikokumontanum together with 27 known compounds. Their structures were established based on spectroscopic evidence. The isolated compounds and some chromone derivatives were assayed for antimicrobial activity against Helicobacter pylori and cytotoxicity against human cancer cell lines.

Continuous-flow complete-mixing system for assessing the effects of environmental factors on colony-level coral metabolism.

A small-scale chamber experimental system was designed to study the effects of temperature on colony-level coral metabolism. The system continuously supplies fresh seawater to the chamber, where it is mixed immediately and completely with the seawater already present. This continuous-flow complete-mixing system (CFCM system), in conjunction with theoretical equations, allows quantitative determination of chemical uptake and release rates by coral under controlled environmental conditions. We used the massive hermatypic coral Goniastrea aspera to examine variations in pH, total alkalinity, and total inorganic carbon for 16 days at 27 degrees C under controlled light intensities (300 and 0 micromol m(-2) s(-1)). We confirmed the stability of the CFCM system with respect to coral photosynthetic and calcification fluxes. In addition, we obtained daily photosynthetic and calcification rates at different temperatures (27 degrees C, 29 degrees C, 31 degrees C, and 33 degrees C). When seawater temperature was raised from 31 degrees C to 33 degrees C, the gross primary production rate (Pgross) decreased 29.5%, and the calcification rate (G) decreased 85.7% within 2 days. The CFCM system allows quantitative evaluation of coral colony chemical release and uptake rates, and metabolism.

Novel abietane diterpenoids and aromatic compounds from Cladonia rangiferina and their antimicrobial activity against antibiotics resistant bacteria.

From Cladonia rangiferina were isolated two novel abietane diterpenoids, hanagokenols A (1) and B (2). Also in this investigation, four known abitetane diterpenoids (3-6), four known labdane diterpenoids (7-10), one known isopimarane diterpenoid (11), and six known aromatic compounds were isolated. These structures were elucidated primarily through extensive NMR experiments. Hanagokenol A (1) was a unique abietane diterpene having an ether linkage between C-6 and C-18 of sugiol. Hanagokenol B (2) is also a unique secoabietane diterpene, having gamma-lactone which occurred by cleavage and subsequently oxidation between C-6/C-7 of 12-hydroxydehydroabietinol. Furthermore, all the isolated compounds (1-17) were tested for the antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococci (VRE).