Fast and pervasive transcriptomic resilience and acclimation of extremely heat-tolerant coral holobionts from the northern Red Sea.

Corals from the northern Red Sea and Gulf of Aqaba exhibit extreme thermal tolerance. To examine the underlying gene expression dynamics, we exposed Stylophora pistillata from the Gulf of Aqaba to short-term (hours) and long-term (weeks) heat stress with peak seawater temperatures ranging from their maximum monthly mean of 27 °C (baseline) to 29.5 °C, 32 °C, and 34.5 °C. Corals were sampled at the end of the heat stress as well as after a recovery period at baseline temperature. Changes in coral host and symbiotic algal gene expression were determined via RNA-sequencing (RNA-Seq). Shifts in coral microbiome composition were detected by complementary DNA (cDNA)-based 16S ribosomal RNA (rRNA) gene sequencing. In all experiments up to 32 °C, RNA-Seq revealed fast and pervasive changes in gene expression, primarily in the coral host, followed by a return to baseline gene expression for the majority of coral (>94%) and algal (>71%) genes during recovery. At 34.5 °C, large differences in gene expression were observed with minimal recovery, high coral mortality, and a microbiome dominated by opportunistic bacteria (including Vibrio species), indicating that a lethal temperature threshold had been crossed. Our results show that the S. pistillata holobiont can mount a rapid and pervasive gene expression response contingent on the amplitude and duration of the thermal stress. We propose that the transcriptomic resilience and transcriptomic acclimation observed are key to the extraordinary thermal tolerance of this holobiont and, by inference, of other northern Red Sea coral holobionts, up to seawater temperatures of at least 32 °C, that is, 5 °C above their current maximum monthly mean.

Heat stress destabilizes symbiotic nutrient cycling in corals.

Recurrent mass bleaching events are pushing coral reefs worldwide to the brink of ecological collapse. While the symptoms and consequences of this breakdown of the coral-algal symbiosis have been extensively characterized, our understanding of the underlying causes remains incomplete. Here, we investigated the nutrient fluxes and the physiological as well as molecular responses of the widespread coral Stylophora pistillata to heat stress prior to the onset of bleaching to identify processes involved in the breakdown of the coral-algal symbiosis. We show that altered nutrient cycling during heat stress is a primary driver of the functional breakdown of the symbiosis. Heat stress increased the metabolic energy demand of the coral host, which was compensated by the catabolic degradation of amino acids. The resulting shift from net uptake to release of ammonium by the coral holobiont subsequently promoted the growth of algal symbionts and retention of photosynthates. Together, these processes form a feedback loop that will gradually lead to the decoupling of carbon translocation from the symbiont to the host. Energy limitation and altered symbiotic nutrient cycling are thus key factors in the early heat stress response, directly contributing to the breakdown of the coral-algal symbiosis. Interpreting the stability of the coral holobiont in light of its metabolic interactions provides a missing link in our understanding of the environmental drivers of bleaching and may ultimately help uncover fundamental processes underpinning the functioning of endosymbioses in general.

A modern scleractinian coral with a two-component calcite-aragonite skeleton.

One of the most conserved traits in the evolution of biomineralizing organisms is the taxon-specific selection of skeletal minerals. All modern scleractinian corals are thought to produce skeletons exclusively of the calcium-carbonate polymorph aragonite. Despite strong fluctuations in ocean chemistry (notably the Mg/Ca ratio), this feature is believed to be conserved throughout the coral fossil record, spanning more than 240 million years. Only one example, the Cretaceous scleractinian coral Coelosmilia (ca. 70 to 65 Ma), is thought to have produced a calcitic skeleton. Here, we report that the modern asymbiotic scleractinian coral Paraconotrochus antarcticus living in the Southern Ocean forms a two-component carbonate skeleton, with an inner structure made of high-Mg calcite and an outer structure composed of aragonite. P. antarcticus and Cretaceous Coelosmilia skeletons share a unique microstructure indicating a close phylogenetic relationship, consistent with the early divergence of P. antarcticus within the Vacatina (i.e., Robusta) clade, estimated to have occurred in the Mesozoic (ca. 116 Mya). Scleractinian corals thus join the group of marine organisms capable of forming bimineralic structures, which requires a highly controlled biomineralization mechanism; this capability dates back at least 100 My. Due to its relatively prolonged isolation, the Southern Ocean stands out as a repository for extant marine organisms with ancient traits.

Correlated cryo-SEM and CryoNanoSIMS imaging of biological tissue.

BACKGROUND: The development of nanoscale secondary ion mass spectrometry (NanoSIMS) has revolutionized the study of biological tissues by enabling, e.g., the visualization and quantification of metabolic processes at subcellular length scales. However, the associated sample preparation methods all result in some degree of tissue morphology distortion and loss of soluble compounds. To overcome these limitations an entirely cryogenic sample preparation and imaging workflow is required. RESULTS: Here, we report the development of a CryoNanoSIMS instrument that can perform isotope imaging of both positive and negative secondary ions from flat block-face surfaces of vitrified biological tissues with a mass- and image resolution comparable to that of a conventional NanoSIMS. This capability is illustrated with nitrogen isotope as well as trace element mapping of freshwater hydrozoan Green Hydra tissue following uptake of 15N-enriched ammonium. CONCLUSION: With a cryo-workflow that includes vitrification by high pressure freezing, cryo-planing of the sample surface, and cryo-SEM imaging, the CryoNanoSIMS enables correlative ultrastructure and isotopic or elemental imaging of biological tissues in their most pristine post-mortem state. This opens new horizons in the study of fundamental processes at the tissue- and (sub)cellular level. TEASER: CryoNanoSIMS: subcellular mapping of chemical and isotopic compositions of biological tissues in their most pristine post-mortem state.

Non-invasive investigation of the morphology and optical properties of the upside-down jellyfish Cassiopea with optical coherence tomography.

The jellyfish Cassiopea largely cover their carbon demand via photosynthates produced by microalgal endosymbionts, but how holobiont morphology and tissue optical properties affect the light microclimate and symbiont photosynthesis in Cassiopea remain unexplored. Here, we use optical coherence tomography (OCT) to study the morphology of Cassiopea medusae at high spatial resolution. We include detailed 3D reconstructions of external micromorphology, and show the spatial distribution of endosymbionts and white granules in the bell tissue. Furthermore, we use OCT data to extract inherent optical properties from light-scattering white granules in Cassiopea, and show that granules enhance local light-availability for symbionts in close proximity. Individual granules had a scattering coefficient of µs = 200-300 cm-1, and scattering anisotropy factor of g = 0.7, while large tissue-regions filled with white granules had a lower µs = 40-100 cm-1, and g = 0.8-0.9. We combined OCT information with isotopic labelling experiments to investigate the effect of enhanced light-availability in whitish tissue regions. Endosymbionts located in whitish tissue exhibited significantly higher carbon fixation compared to symbionts in anastomosing tissue (i.e. tissue without light-scattering white granules). Our findings support previous suggestions that white granules in Cassiopea play an important role in the host modulation of the light-microenvironment.

Generalized size scaling of metabolic rates based on single-cell measurements with freshwater phytoplankton.

Kleiber's law describes the scaling of metabolic rate with body size across several orders of magnitude in size and across taxa and is widely regarded as a fundamental law in biology. The physiological origins of Kleiber's law are still debated and generalizations of the law accounting for deviations from the scaling behavior have been proposed. Most theoretical and experimental studies of Kleiber's law, however, have focused on the relationship between the average body size of a species and its mean metabolic rate, neglecting intraspecific variation of these 2 traits. Here, we propose a theoretical characterization of such variation and report on proof-of-concept experiments with freshwater phytoplankton supporting such framework. We performed joint measurements at the single-cell level of cell volume and nitrogen/carbon uptake rates, as proxies of metabolic rates, of 3 phytoplankton species using nanoscale secondary ion mass spectrometry (NanoSIMS) and stable isotope labeling. Common scaling features of the distribution of nutrient uptake rates and cell volume are found to hold across 3 orders of magnitude in cell size. Once individual measurements of cell volume and nutrient uptake rate within a species are appropriately rescaled by a function of the average cell volume within each species, we find that intraspecific distributions of cell volume and metabolic rates collapse onto a universal curve. Based on the experimental results, this work provides the building blocks for a generalized form of Kleiber's law incorporating intraspecific, correlated variations of nutrient-uptake rates and body sizes.

Surface Analysis by Secondary Ion Mass Spectrometry (SIMS): Principles and Applications from Swiss laboratories.

Secondary Ion Mass Spectrometry (SIMS) extracts chemical, elemental, or isotopic information about a localized area of a solid target by performing mass spectrometry on secondary ions sputtered from its surface by the impact of a beam of charged particles. This primary beam sputters ionized atoms and small molecules (as well as many neutral particles) from the upper few nanometers of the sample surface. The physical basis of SIMS has been applied to a large range of applications utilizing instruments optimized with different types of mass analyzer, either dynamic SIMS with a double focusing mass spectrometer or static SIMS with a Time of Flight (TOF) analyzer. Here, we present a short review of the principles and major applications of three different SIMS instruments located in Switzerland.

Photosynthesis from stolen chloroplasts can support sea slug reproductive fitness.

Some sea slugs are able to steal functional chloroplasts (kleptoplasts) from their algal food sources, but the role and relevance of photosynthesis to the animal host remain controversial. While some researchers claim that kleptoplasts are slowly digestible 'snacks', others advocate that they enhance the overall fitness of sea slugs much more profoundly. Our analysis shows light-dependent incorporation of 13C and 15N in the albumen gland and gonadal follicles of the sea slug Elysia timida, representing translocation of photosynthates to kleptoplast-free reproductive organs. Long-chain polyunsaturated fatty acids with reported roles in reproduction were produced in the sea slug cells using labelled precursors translocated from the kleptoplasts. Finally, we report reduced fecundity of E. timida by limiting kleptoplast photosynthesis. The present study indicates that photosynthesis enhances the reproductive fitness of kleptoplast-bearing sea slugs, confirming the biological relevance of this remarkable association between a metazoan and an algal-derived organelle.

Tracking the cargo of extracellular symbionts into host tissues with correlated electron microscopy and nanoscale secondary ion mass spectrometry imaging.

Extracellular bacterial symbionts communicate biochemically with their hosts to establish niches that foster the partnership. Using quantitative ion microprobe isotopic imaging (nanoscale secondary ion mass spectrometry [NanoSIMS]), we surveyed localization of 15 N-labelled molecules produced by the bacterium Vibrio fischeri within the cells of the symbiotic organ of its host, the Hawaiian bobtail squid, and compared that with either labelled non-specific species or amino acids. In all cases, two areas of the organ's epithelia were significantly more 15 N enriched: (a) surface ciliated cells, where environmental symbionts are recruited, and (b) the organ's crypts, where the symbiont population resides in the host. Label enrichment in all cases was strongest inside host cell nuclei, preferentially in the euchromatin regions and the nucleoli. This permissiveness demonstrated that uptake of biomolecules is a general mechanism of the epithelia, but the specific responses to V. fischeri cells recruited to the organ's surface are due to some property exclusive to this species. Similarly, in the organ's deeper crypts, the host responds to common bacterial products that only the specific symbiont can present in that location. The application of NanoSIMS allows the discovery of such distinct modes of downstream signalling dependent on location within the host and provides a unique opportunity to study the microbiogeographical patterns of symbiotic dialogue.

Sub-cellular imaging shows reduced photosynthetic carbon and increased nitrogen assimilation by the non-native endosymbiont Durusdinium trenchii in the model cnidarian Aiptasia.

Hosting different symbiont species can affect inter-partner nutritional fluxes within the cnidarian-dinoflagellate symbiosis. Using nanoscale secondary ion mass spectrometry (NanoSIMS), we measured the spatial incorporation of photosynthetically fixed 13 C and heterotrophically derived 15 N into host and symbiont cells of the model symbiotic cnidarian Aiptasia (Exaiptasia pallida) when colonized with its native symbiont Breviolum minutum or the non-native Durusdinium trenchii. Breviolum minutum exhibited high photosynthetic carbon assimilation per cell and translocation to host tissue throughout symbiosis establishment, whereas D. trenchii assimilated significantly less carbon, but obtained more host nitrogen. These findings suggest that D. trenchii has less potential to provide photosynthetically fixed carbon to the host despite obtaining considerable amounts of heterotrophically derived nitrogen. These sub-cellular events help explain previous observations that demonstrate differential effects of D. trenchii compared to B. minutum on the host transcriptome, proteome, metabolome and host growth and asexual reproduction. Together, these differential effects suggest that the non-native host-symbiont pairing is sub-optimal with respect to the host's nutritional benefits under normal environmental conditions. This contributes to our understanding of the ways in which metabolic integration impacts the benefits of a symbiotic association, and the potential evolution of novel host-symbiont pairings.

Intracellular competition for nitrogen controls dinoflagellate population density in corals.

The density of dinoflagellate microalgae in the tissue of symbiotic corals is an important determinant for health and productivity of the coral animal. Yet, the specific mechanism for their regulation and the consequence for coral nutrition are insufficiently understood due to past methodological limitations to resolve the fine-scale metabolic consequences of fluctuating densities. Here, we characterized the physiological and nutritional consequences of symbiont density variations on the colony and tissue level in Stylophora pistillata from the Red Sea. Alterations in symbiont photophysiology maintained coral productivity and host nutrition across a broad range of symbiont densities. However, we demonstrate that density-dependent nutrient competition between individual symbiont cells, manifested as reduced nitrogen assimilation and cell biomass, probably creates the negative feedback mechanism for symbiont population growth that ultimately defines the steady-state density. Despite fundamental changes in symbiont nitrogen assimilation, we found no density-related metabolic optimum beyond which host nutrient assimilation or tissue biomass declined, indicating that host nutrient demand is sufficiently met across the typically observed range of symbiont densities under ambient conditions.

Amoebocytes facilitate efficient carbon and nitrogen assimilation in the Cassiopea-Symbiodiniaceae symbiosis.

The upside-down jellyfish Cassiopea engages in symbiosis with photosynthetic microalgae that facilitate uptake and recycling of inorganic nutrients. By contrast to most other symbiotic cnidarians, algal endosymbionts in Cassiopea are not restricted to the gastroderm but are found in amoebocyte cells within the mesoglea. While symbiont-bearing amoebocytes are highly abundant, their role in nutrient uptake and cycling in Cassiopea remains unknown. By combining isotopic labelling experiments with correlated scanning electron microscopy, and Nano-scale secondary ion mass spectrometry (NanoSIMS) imaging, we quantified the anabolic assimilation of inorganic carbon and nitrogen at the subcellular level in juvenile Cassiopea medusae bell tissue. Amoebocytes were clustered near the sub-umbrella epidermis and facilitated efficient assimilation of inorganic nutrients. Photosynthetically fixed carbon was efficiently translocated between endosymbionts, amoebocytes and host epidermis at rates similar to or exceeding those observed in corals. The Cassiopea holobionts efficiently assimilated ammonium, while no nitrate assimilation was detected, possibly reflecting adaptation to highly dynamic environmental conditions of their natural habitat. The motile amoebocytes allow Cassiopea medusae to distribute their endosymbiont population to optimize access to light and nutrients, and transport nutrition between tissue areas. Amoebocytes thus play a vital role for the assimilation and translocation of nutrients in Cassiopea, providing an interesting new model for studies of metabolic interactions in photosymbiotic marine organisms.

Ammonium is the preferred source of nitrogen for planktonic foraminifer and their dinoflagellate symbionts.

The symbiotic planktonic foraminifera Orbulina universa inhabits open ocean oligotrophic ecosystems where dissolved nutrients are scarce and often limit biological productivity. It has previously been proposed that O. universa meets its nitrogen (N) requirements by preying on zooplankton, and that its symbiotic dinoflagellates recycle metabolic 'waste ammonium' for their N pool. However, these conclusions were derived from bulk 15N-enrichment experiments and model calculations, and our understanding of N assimilation and exchange between the foraminifer host cell and its symbiotic dinoflagellates remains poorly constrained. Here, we present data from pulse-chase experiments with 13C-enriched inorganic carbon, 15N-nitrate, and 15N-ammonium, as well as a 13C- and 15N- enriched heterotrophic food source, followed by TEM (transmission electron microscopy) coupled to NanoSIMS (nanoscale secondary ion mass spectrometry) isotopic imaging to visualize and quantify C and N assimilation and translocation in the symbiotic system. High levels of 15N-labelling were observed in the dinoflagellates and in foraminiferal organelles and cytoplasm after incubation with 15N-ammonium, indicating efficient ammonium assimilation. Only weak 15N-assimilation was observed after incubation with 15N-nitrate. Feeding foraminifers with 13C- and 15N-labelled food resulted in dinoflagellates that were labelled with 15N, thereby confirming the transfer of 15N-compounds from the digestive vacuoles of the foraminifer to the symbiotic dinoflagellates, likely through recycling of ammonium. These observations are important for N isotope-based palaeoceanographic reconstructions, as they show that δ15N values recorded in the organic matrix in symbiotic species likely reflect ammonium recycling rather than alternative N sources, such as nitrates.

Kleptoplastidic benthic foraminifera from aphotic habitats: insights into assimilation of inorganic C, N and S studied with sub-cellular resolution.

The assimilation of inorganic compounds in foraminiferal metabolism compared to predation or organic matter assimilation is unknown. Here, we investigate possible inorganic-compound assimilation in Nonionellina labradorica, a common kleptoplastidic benthic foraminifer from Arctic and North Atlantic sublittoral regions. The objectives were to identify the source of the foraminiferal kleptoplasts, assess their photosynthetic functionality in light and darkness and investigate inorganic nitrogen and sulfate assimilation. We used DNA barcoding of a ~ 830 bp fragment from the SSU rDNA to identify the kleptoplasts and correlated transmission electron microscopy and nanometre-scale secondary ion mass spectrometry (TEM-NanoSIMS) isotopic imaging to study 13 C-bicarbonate, 15 N-ammonium and 34 S-sulfate uptake. In addition, respiration rate measurements were determined to assess the response of N. labradorica to light. The DNA sequences established that over 80% of the kleptoplasts belonged to Thalassiosira (with 96%-99% identity), a cosmopolitan planktonic diatom. TEM-NanoSIMS imaging revealed degraded cytoplasm and an absence of 13 C assimilation in foraminifera exposed to light. Oxygen measurements showed higher respiration rates under light than dark conditions, and no O2 production was detected. These results indicate that the photosynthetic pathways in N. labradorica are not functional. Furthermore, N. labradorica assimilated both 15 N-ammonium and 34 S-sulfate into its cytoplasm, which suggests that foraminifera might have several ammonium or sulfate assimilation pathways, involving either the kleptoplasts or bona fide foraminiferal pathway(s) not yet identified.

Developmental carryover effects of ocean warming and acidification in corals from a potential climate refugium, the Gulf of Aqaba.

Coral reefs are degrading from the effects of anthropogenic activities, including climate change. Under these stressors, their ability to survive depends upon existing phenotypic plasticity, but also transgenerational adaptation. Parental effects are ubiquitous in nature, yet empirical studies of these effects in corals are scarce, particularly in the context of climate change. This study exposed mature colonies of the common reef-building coral Stylophora pistillata from the Gulf of Aqaba to seawater conditions likely to occur just beyond the end of this century during the peak planulae brooding season (Representative Concentration Pathway 8.5: pH -0.4 and +5°C beyond present day). Parent and planulae physiology were assessed at multiple time points during the experimental incubation. After 5 weeks of incubation, the physiology of the parent colonies exhibited limited treatment-induced changes. All significant time-dependent changes in physiology occurred in both ambient and treatment conditions. Planulae were also resistant to future ocean conditions, with protein content, symbiont density, photochemistry, survival and settlement success not significantly different compared with under ambient conditions. High variability in offspring physiology was independent of parental or offspring treatments and indicate the use of a bet-hedging strategy in this population. This study thus demonstrates weak climate-change-associated carryover effects. Furthermore, planulae display temperature and pH resistance similar to those of adult colonies and therefore do not represent a larger future population size bottleneck. The findings add support to the emerging hypothesis that the Gulf of Aqaba may serve as a coral climate change refugium aided by these corals' inherent broad physiological resistance.

Cellular Uptake and Intracellular Trafficking of Poly( N-(2-Hydroxypropyl) Methacrylamide).

Cellular uptake and intracellular trafficking of polymer conjugates or polymer nanoparticles is typically monitored using fluorescence-based techniques such as confocal microscopy. While these methods have provided a wealth of insight into the internalization and trafficking of polymers and polymer nanoparticles, they require fluorescent labeling of the polymer or polymer nanoparticle. Because in biological media fluorescent dyes may degrade, be cleaved from the polymer or particle, or even change uptake and trafficking pathways, there is an interest in fluorescent label-free methods to study the interactions between cells and polymer nanomedicines. This article presents a first proof-of-concept that demonstrates the feasibility of NanoSIMS to monitor the intracellular localization of polymer conjugates. For the experiments reported here, poly( N-(2-hydroxypropyl) methacrylamide)) (PHPMA) was selected as a prototypical polymer-drug conjugate. This PHPMA polymer contained a 19F-label at the α-terminus, which was introduced in order to allow NanoSIMS analysis. Prior to the NanoSIMS experiments, the uptake and intracellular trafficking of the polymer was established using confocal microscopy and flow cytometry. These experiments not only provided detailed insight into the kinetics of these processes but were also important to select time points for the NanoSIMS analysis. For the NanoSIMS experiments, HeLa cells were investigated that had been exposed to the PHPMA polymer for a period of 4 or 15 h, which was known to lead to predominant lysosomal accumulation of the polymer. NanoSIMS analysis of resin-embedded and microtomed samples of the cells revealed a punctuated fluorine signal, which was found to colocalize with the sulfur signal that was attributed to the lysosomal compartments. The localization of the polymer in the endolysosomal compartments was confirmed by TEM analysis on the same cell samples. The results of this study illustrate the potential of NanoSIMS to study the uptake and intracellular trafficking of polymer nanomedicines.

The Differential Distribution of RAPTA-T in Non-Invasive and Invasive Breast Cancer Cells Correlates with Its Anti-Invasive and Anti-Metastatic Effects.

Nanoscale secondary ion mass spectrometry (NanoSIMS) combined with transmission electron microscopy (TEM) can be a powerful approach to visualize the exact distribution of drugs at the sub-cellular level. In this work, we exploit this approach to identify the distribution and localisation of the organometallic ruthenium(II)-arene drug Ru(η6-C6H5Me)(pta)Cl2, termed RAPTA-T, in MDA-MB-231 and MCF-7 human breast cancer cells. These cell lines have been chosen because the former cell lines are highly invasive and resistant to most chemotherapeutic agents and the latter ones are very sensitive to hormonal-based therapies. In the MDA-MB-231 cells, RAPTA-T was found to predominantly localise on the cell membrane and to a lesser extent in the nucleolus. These findings are consistent with the previously reported anti-metastatic properties of RAPTA-T and the observation that once internalized RAPTA-T is associated with chromatin. RAPTA-T shows a lack of membrane accumulation on the non-invasive MCF-7 cells, which correlates well with its selective anti-metastatic properties on invasive cell lines.

Cell proliferation and migration during early development of a symbiotic scleractinian coral.

In scleractinian reef-building corals, patterns of cell self-renewal, migration and death remain virtually unknown, limiting our understanding of cellular mechanisms underlying initiation of calcification, and ontogenesis of the endosymbiotic dinoflagellate relationship. In this study, we pulse-labelled the coral Stylophora pistillata for 24 h with BrdU at four life stages (planula, early metamorphosis, primary polyp and adult colony) to investigate coral and endosymbiont cell proliferation during development, while simultaneously recording TUNEL-positive (i.e. apoptotic) nuclei. In the primary polyp, the fate of BrdU-labelled cells was tracked during a 3-day chase. The pharynx and gastrodermis were identified as the most proliferative tissues in the developing polyp, and BrdU-labelled cells accumulated in the surface pseudostratified epithelium and the skeletogenic calicodermis during the chase, revealing cell migration to these epithelia. Surprisingly, the lowest cell turnover was recorded in the calicodermis at all stages, despite active, ongoing skeletal deposition. In dinoflagellate symbionts, DNA synthesis was systematically higher than coral host gastrodermis, especially in planula and early metamorphosis. The symbiont to host cell ratio remained constant, however, indicating successive post-mitotic control mechanisms by the host of its dinoflagellate density in early life stages, increasingly shifting to apoptosis in the growing primary polyp.

A nanoscale secondary ion mass spectrometry study of dinoflagellate functional diversity in reef-building corals.

Nutritional interactions between corals and symbiotic dinoflagellate algae lie at the heart of the structural foundation of coral reefs. Whilst the genetic diversity of Symbiodinium has attracted particular interest because of its contribution to the sensitivity of corals to environmental changes and bleaching (i.e. disruption of coral-dinoflagellate symbiosis), very little is known about the in hospite metabolic capabilities of different Symbiodinium types. Using a combination of stable isotopic labelling and nanoscale secondary ion mass spectrometry (NanoSIMS), we investigated the ability of the intact symbiosis between the reef-building coral Isopora palifera, and Symbiodinium C or D types, to assimilate dissolved inorganic carbon (via photosynthesis) and nitrogen (as ammonium). Our results indicate that Symbiodinium types from two clades naturally associated with I. palifera possess different metabolic capabilities. The Symbiodinium C type fixed and passed significantly more carbon and nitrogen to its coral host than the D type. This study provides further insights into the metabolic plasticity among different Symbiodinium types in hospite and strengthens the evidence that the more temperature-tolerant Symbiodinium D type may be less metabolically beneficial for its coral host under non-stressful conditions.

Imaging liver and brain glycogen metabolism at the nanometer scale.

In mammals, glycogen synthesis and degradation are dynamic processes regulating blood and cerebral glucose-levels within a well-defined physiological range. Despite the essential role of glycogen in hepatic and cerebral metabolism, its spatiotemporal distribution at the molecular and cellular level is unclear. By correlating electron microscopy and ultra-high resolution ion microprobe (NanoSIMS) imaging of tissue from fasted mice injected with (13)C-labeled glucose, we demonstrate that liver glycogenesis initiates in the hepatocyte perinuclear region before spreading toward the cell membrane. In the mouse brain, we observe that (13)C is inhomogeneously incorporated into astrocytic glycogen at a rate ~25 times slower than in the liver, in agreement with prior bulk studies. This experiment, using temporally resolved, nanometer-scale imaging of glycogen synthesis and degradation, provides greater insight into glucose metabolism in mammalian organs and shows how this technique can be used to explore biochemical pathways in healthy and diseased states.