Change in rheotactic behavior patterns of dinoflagellates in response to different microfluidic environments.

Plankton live in dynamic fluid environments. Their ability to change in response to different hydrodynamic cues is critical to their energy allocation and resource uptake. This study used a microfluidic device to evaluate the rheotactic behaviors of a model dinoflagellate species, Karlodinium veneficum, in different flow conditions. Although dinoflagellates experienced forced alignment in strong shear (i.e. "trapping"), fluid straining did not play a decisive role in their rheotactic movements. Moderate hydrodynamic magnitude (20 < |uf| < 40 µm s-1) was found to induce an orientation heading towards an oncoming current (positive rheotaxis), as dinoflagellates switched to cross-flow swimming when flow speed exceeded 50 µm s-1. Near the sidewalls of the main channel, the steric mechanism enabled dinoflagellates to adapt upstream orientation through vertical migration. Under oscillatory flow, however, positive rheotaxis dominated with occasional diversion. The varying flow facilitated upstream exploration with directional controlling, through which dinoflagellates exhibited avoidance of both large-amplitude perturbance and very stagnant zones. In the mixed layer where water is not steady, these rheotactic responses could lead to spatial heterogeneity of dinoflagellates. The outcome of this study helps clarify the interaction between swimming behaviors of dinoflagellates and the hydrodynamic environment they reside in.

Iron Availability Modulates the Response of Endosymbiotic Dinoflagellates to Heat Stress.

Warming and nutrient limitation are stressors known to weaken the health of microalgae. In situations of stress, access to energy reserves can minimize physiological damage. Because of its widespread requirements in biochemical processes, iron is an important trace metal, especially for photosynthetic organisms. Lowered iron availability in oceans experiencing rising temperatures may contribute to the thermal sensitivity of reef-building corals, which rely on mutualisms with dinoflagellates to survive. To test the influence of iron concentration on thermal sensitivity, the physiological responses of cultured symbiotic dinoflagellates (genus Breviolum; family Symbiodiniaceae) were evaluated when exposed to increasing temperatures (26 to 30°C) and iron concentrations ranging from replete (500 pM Fe') to limiting (50 pM Fe') under a diurnal light cycle with saturating radiance. Declines in photosynthetic efficiency at elevated temperatures indicated sensitivity to heat stress. Furthermore, five times the amount of iron was needed to reach exponential growth during heat stress (50 pM Fe' at 26-28°C vs. 250 pM Fe' at 30°C). In treatments where exponential growth was reached, Breviolum psygmophilum grew faster than B.minutum, possibly due to greater cellular contents of iron and other trace metals. The metal composition of B.psygmophilum shifted only at the highest temperature (30°C), whereas changes in B.minutum were observed at lower temperatures (28°C). The influence of iron availability in modulating each alga's response to thermal stress suggests the importance of trace metals to the health of coral-algal mutualisms. Ultimately, a greater ability to acquire scarce metals may improve the tolerance of corals to physiological stressors and contribute to the differences in performance associated with hosting one symbiont species over another.

RNA-seq profiling of Fugacium kawagutii reveals strong responses in metabolic processes and symbiosis potential to deficiencies of iron and other trace metals.

A healthy symbiotic relationship between corals and Symbiodiniaceae relies on suitable temperature and adequate nutrients including trace metals. Besides global warming, trace metal deficiency has been shown to cause coral bleaching, a phenomenon responsible for extensive coral reef degradation around the world. How trace metal deficiency impacts Symbiodiniaceae and coral symbiosis is poorly understood, however. In this study, we applied RNA-seq to investigate how Fugacium kawagutii responds to the deficiency of five trace metals (Fe2+, Zn2+, Cu2+, Mn2+, Ni2+). We identified 685 to 2805 differentially expressed genes (DEGs) from these trace metal deficiency conditions, among which 372 were commonly regulated by all the five trace metals and were significantly enriched in energy metabolism (e.g. fatty acid synthesis). Furthermore, genes associated with extracellular matrix (ECM), cell surface structure and cell adhesion were impacted, suggesting that the ability of recognition and adhesion of F. kawagutii may be altered by trace metal deficiencies. In addition, among the five metals, Fe2+ deficiency exhibited the strongest influence, with Fe-rich redox elements and many antioxidant synthesis genes being markedly down-regulated, indicative of adaptive reduction of Fe demand but a compromised ability to combat oxidative stress. Overall, deficiency of trace metals (especially Fe) seems to repress growth and ability of ROS scavenging, elevate energy metabolism and innate immunity, and alter cell adhesion capability, with implications in symbiosis disruption and coral bleaching.

Metatranscriptomics of the bacterial community in response to atmospheric deposition in the Western North Pacific Ocean.

Atmospheric deposition represents a major vector of both macro- and micro-nutrients to the oligotrophic open oceans, potentially imposing a profound impact on the functioning of the microbial community. Whereas bacterial responses to atmospheric deposition are being studied at the community level, corresponding functional changes are essentially unknown. Here we conducted a microcosm experiment coupled with metatranscriptomic analyses to elucidate taxonomic and functional profiles of the bacterial community in response to East Asian aerosols in the Western North Pacific Ocean (WNPO). While the abundance of heterotrophic bacteria showed a minor change, cyanobacterial cell count number decreased dramatically, with Prochlorococcus and Synechococcus counts reduced by 83.2% and 21.5% in the aerosol treatment in relation to the control. Expression of transcripts related with Prochlorococcus, Synechococcus, Trichodesmium and Crocosphaera both were lower in the treatment (5.7%, 2.3%, 0.5% and 0.02%, respectively) than in the control (18.6%, 2.7%, 9.8% and 0.14%, respectively). Aerosol addition led to an increase in transcripts involved in iron metabolism (tonB, feoB, irr, exbB), indicating Fe limitation. Heavy metal toxicity was evidenced by an elevated expression of resistance genes, such as czcC, czcB, czcA and a probable Co/Zn/Cd efflux protein, and a range of genes functioning against oxidative stress. Our findings provide insights into an inhibitory effect of high-flux East Asian aerosols on cyanobacteria in the WNPO likely due to Fe scavenging and heavy metal toxicity.

Trace Metal Requirements and Interactions in Symbiodinium kawagutii.

Photosynthetic organisms need trace metals for various biological processes and different groups of microalgae have distinctive obligate necessities due to their respective biochemical requirements and ecological niches. We have previously shown that the dinoflagellate Symbiodinium kawagutii requires high concentrations of bioavailable Fe to achieve optimum growth. Here, we further explored the trace metal requirements of S. kawagutii with intensive focus on the effect of individual metal and its interaction with other divalent metals. We found that low Zn availability significantly decreases growth rates and results in elevated intracellular Mn, Co, Ni, and Fe quotas in the dinoflagellate. The results highlight the complex interaction among trace metals in S. kawagutii and suggest either metal replacement strategy to counter low Zn availability or enhanced uptake of other metals by non-specific divalent metal transporters. In this work, we also examined the Fe requirement of S. kawagutii using continuous cultures. We validated that 500 pM of Fe' was sufficient to support maximum cell density during steady state growth period either at 26 or 28°C. This study shows that growth of S. kawagutii was limited by metal availability in the following order, Fe > Zn > Mn > Cu > Ni > Co. The fundamental information obtained for the free-living Symbiodinium shall provide insights into how trace metal availability, either from ambient seawater or hosts, affects growth and proliferation of symbiotic dinoflagellates and the interaction between symbiont and their hosts.

Interactive effects of spectral quality and trace metal availability on the growth of Trichodesmium and Symbiodinium.

Light and trace metals are critical growth factors for algae but how the interdependence of light quality and metal availability affects algal growth remains largely unknown. Our previous studies have demonstrated the importance of Ni and Fe on the growth of Trichodesmium and Symbiodinium, respectively, two important marine primary producers inhabiting environments with high light intensities. Here, we investigated the effects of light quality and intensity with availability of either Ni or Fe on their growth. For Trichodesmium, we found that specific growth rates for high Ni treatments were all significantly higher than in corresponding low Ni treatments with varying light quality and intensity. The inhibitory effect of low intensity red light was also countered by sufficient Ni supply. For Symbiodinium, we found that growth rates and biomass were reduced by 75% under low intensity red light and the stress can only be partially relieved by sufficient Fe supply. The results show that trace metal availability plays an important role in relieving the stress induced by low red light condition for both Trichodesmium and Symbiodinium although the cyanobacterium performs better in this growth condition. The difference may be attributed to the presence of phycocyanin, a unique pigment attuned to absorption of red light, in Trichodesmium. Our study shows that the concerted effects of light intensity and quality compounded with trace metal availability may influence the growth of photosynthetic organisms in the ocean.

Effects of Trace Metal Concentrations on the Growth of the Coral Endosymbiont Symbiodinium kawagutii.

Symbiodinium is an indispensable endosymbiont in corals and the most important primary producer in coral reef ecosystems. During the past decades, coral bleaching attributed to the disruption of the symbiosis has frequently occurred resulting in reduction of coral reef coverage globally. Growth and proliferation of corals require some specific trace metals that are essential components of pertinent biochemical processes, such as in photosynthetic systems and electron transport chains. In addition, trace metals are vital in the survival of corals against oxidative stress because these metals serve as enzymatic cofactors in antioxidative defense mechanisms. The basic knowledge about trace metal requirements of Symbiodinium is lacking. Here we show that the requirement of Symbiodinium kawagutii for antioxidant-associated trace metals exhibits the following order: Fe >> Cu/Zn/Mn >> Ni. In growth media with Cu, Zn, Mn, and varying Fe concentrations, we observed that Cu, Zn, and Mn cellular quotas were inversely related to Fe concentrations. In the absence of Cu, Zn, and Mn, growth rates increased with increasing inorganic Fe concentrations up to 1250 pM, indicating the relatively high Fe requirement for Symbiodinium growth and potential functional complementarity of these metals. These results demonstrate the relative importance of trace metals to sustain Symbiodinium growth and a potential metal inter replacement strategy in Symbiodinium to ensure survival of coral reefs in an oligotrophic and stressful environment.

Influence of Co and B 12 on the growth and nitrogen fixation of Trichodesmium.

We investigated the influence of varying cobalt (Co) and B12 concentrations to growth and nitrogen fixation of Trichodesmium, a major diazotroph in the tropical and subtropical oligotrophic ocean. Here we show that sufficient inorganic Co, 20 pmol L(-1), sustains the growth of Trichodesmium either with or without an additional B12 supply. We also found that in these culture conditions, nitrogen levels fixed by Trichodesmium were higher in treatments with insufficient B12 than in treatments with higher B12 availability. Under limited inorganic Co availability, ranging from 0.2 to 2 pmol L(-1), Trichodesmium growth was significantly compromised in cultures without B12. In these low Co concentrations, addition of 400 pmol L(-1) of B12 supported phytoplankton growth indicating that B12 supply augmented for the low Co concentrations. Our study demonstrates that Trichodesmium has an absolute Co requirement, which is not replaceable with Zn, and that B12 supply alleviates stress in cases where Co is limiting. These results show that the interlocking availabilities of Co and B12 may influence the growth and nitrogen fixation of Trichodesmium in the ocean.

Trace metal determination in natural waters by automated solid phase extraction system and ICP-MS: the influence of low level Mg and Ca.

A fully automated high pressure pretreatment system with Nobias Chelate-PA1 resin (PA1) was developed for trace metal determination by ICP-MS in natural waters. By varying the concentrations of Mg and Ca to mimic the concentrations in the eluate obtained by PA1 or iminodiacetate type resins, the overall analytical performance of the system was assessed for the determination of Al, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, Y, Cd, Ag, Pb and REE. Comparing with the low mM level Mg and Ca (both ranging from 1 to 4mM) eluted by iminodiacetate type resins, the eluate obtained by PA1 contains sub-µM level Mg and Ca, which remarkably decrease matrix effect in ICP-MS analysis and significantly improve the analytical performance. With recovery better than 90% for most the trace metals examined, the accuracy was further verified through the analysis of five natural water reference materials with salinity spanning from 0 to 35‰. We have successfully applied the pretreatment system to determine trace metals in the seawater samples collected in the Western Philippine Sea through Taiwan GEOTRACES cruise.

Diel nitrogen fixation pattern of Trichodesmium: the interactive control of light and Ni.

Trichodesmium, a nonheterocystous cyanobacterium widely abundant in the surface water of the tropical and subtropical ocean, fixes dinitrogen under high light conditions while concurrently undergoing photosynthesis. The new production considerably influences the cycling of nitrogen and carbon in the ocean. Here, we investigated how light intensity and nickel (Ni) availability interplay to control daily rates and diel patterns of N2 fixation in Trichodesmium. We found that increasing Ni concentration increased N2 fixation rates by up to 30-fold in the high light treatment. Cultures subjected to high Ni and light levels fixed nitrogen throughout most of the 24 H light:dark regime with the highest rate coinciding with the end of the 12 H light period. Our study demonstrates the importance of Ni on nitrogen fixation rates for Trichodesmium under high light conditions.

Comparison of metal accumulation in the azooxanthellate scleractinian coral (Tubastraea coccinea) from different polluted environments.

The response of metal accumulation in coral Tubastraea coccinea to various degrees of metal enrichment was investigated from the Yin-Yang Sea (YYS) receiving abandoned mining effluents, the Kueishan Islet (KI) hydrothermal vent field, and the nearshore area of remoted Green Island (GI). The concentrations of most dissolved metals were highest in seawater at YYS, followed by KI, and then GI, showing the effects of anthropogenic and venting inputs on metal levels. Five metals (Co, Fe, Mn, Ni, and Zn) yielded significant differences (p<0.05) among the skeleton samples. We identified similar patterns in the metal-Ca ratios, indicating that the elevated metals in skeletons was a consequence of external inputs. The coral tissues were relatively sensitive in monitoring metal accumulation, showing significant differences among three locations for Cd, Co, Cu, Fe, Pb, Ni, and Zn. Specific bioconcentration factors provided strong support for the differential metal accumulation in skeletons and tissues.

Interrelated influence of light and Ni on Trichodesmium growth.

Our previous laboratory study revealed that insufficient Ni supply can limit nitrogen fixation in Trichodesmium, a primary diazotrophic phytoplankton in the tropical and subtropical oceans. Here we show that light intensity and Ni availability interrelate to influence Trichodesmium growth. Trichodesmium growth is severely inhibited under high light (670 µE m(-2) s(-1)) and insufficient Ni condition. On the contrary, the sufficient supply of Ni in seawater can sustain the growth of Trichodesmium under either high or low light conditions. We also observed elevated intracellular Ni uptake in Trichodesmium grown under relatively high light condition, supporting that the Ni requirement is used for removing superoxide generated through photosynthetic electron transport. This study shows that light saturation condition for Trichodesmium growth is Ni concentration dependent. This finding may exhibit implications for interpreting temporal and spatial distributions and activities of Trichodesmium in both modern and ancient oceans when light intensity and Ni concentrations have significantly varied.

Determination of trace metals in seawater by an automated flow injection ion chromatograph pretreatment system with ICPMS.

A novel flow injection ion chromatograph (FI-IC) system has been developed to fully automate pretreatment procedures for multi-elemental analysis of trace metals in seawater by inductively coupled plasma mass spectrometer (ICPMS). By combining 10-port, 2 position and 3-way valves in the FI-IC manifold, the system effectively increase sample throughput by simultaneously processing three seawater samples online for: sample loading, injection, buffering, preconcentration, matrix removal, metal elution, and sample collection. Forty-two seawater samples can be continuously processed without any manual handing. Each sample pretreatment takes about 10 min by consuming 25 mL of seawater and producing 5 mL of processed concentrated samples for multi-elemental offline analysis by ICPMS. The offline analysis improve analytical precision and significantly increase total numbers of isotopes determined by ICPMS, which include the metals Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Ti, V, and Zn. The blank value and detection limits of trace metals using the system with ICPMS analysis all range from 0.1 to 10 parts per trillion (ppt), except Al, Fe, and Zn. The accuracy of the pretreatment system was validated by measuring open-ocean and coastal reference seawater, NASS-5 and CASS-4. Using the system with ICPMS analysis, we have obtained reliable trace metal concentrations in the water columns of the South China Sea. Possessing the features of full automation, high throughput, low blank, and low reagent volume used, the system automates and simplifies rigorous and complicated pretreatment procedures for multi-elemental analysis of trace metals in seawater and effectively enhances analytical capacity for trace metal analysis in environmental and seawater samples.

The evolutionary inheritance of elemental stoichiometry in marine phytoplankton.

Phytoplankton is a nineteenth century ecological construct for a biologically diverse group of pelagic photoautotrophs that share common metabolic functions but not evolutionary histories. In contrast to terrestrial plants, a major schism occurred in the evolution of the eukaryotic phytoplankton that gave rise to two major plastid superfamilies. The green superfamily appropriated chlorophyll b, whereas the red superfamily uses chlorophyll c as an accessory photosynthetic pigment. Fossil evidence suggests that the green superfamily dominated Palaeozoic oceans. However, after the end-Permian extinction, members of the red superfamily rose to ecological prominence. The processes responsible for this shift are obscure. Here we present an analysis of major nutrients and trace elements in 15 species of marine phytoplankton from the two superfamilies. Our results indicate that there are systematic phylogenetic differences in the two plastid types where macronutrient (carbon:nitrogen:phosphorus) stoichiometries primarily reflect ancestral pre-symbiotic host cell phenotypes, but trace element composition reflects differences in the acquired plastids. The compositional differences between the two plastid superfamilies suggest that changes in ocean redox state strongly influenced the evolution and selection of eukaryotic phytoplankton since the Proterozoic era.