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Trichodesmium , a globally significant N 2 -fixing marine cyanobacterium, forms extensive surface blooms in nutrient-poor ocean regions. These blooms consist of a dynamic assemblage of Trichodesmium species that form distinct colony morphotypes and are inhabited by diverse microorganisms. Trichodesmium colony morphotypes vary in ecological niche, nutrient uptake, and organic molecule release, differentially impacting ocean carbon and nitrogen biogeochemical cycles. Here, we assessed the poorly studied spatial abundance of metabolites within and between three morphologically distinct Trichodesmium colonies collected from the Red Sea. We also compared these results with two morphotypes of the cultivable Trichodesmium strain IMS101. Using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) coupled with liquid extraction surface analysis (LESA) tandem mass spectrometry (MS 2 ), we identified and localized a wide range of small metabolites associated with single-colony Trichodesmium morphotypes. Our untargeted MALDI-MSI approach revealed 80 unique features (metabolites) shared between Trichodesmium morphotypes. Discrimination analysis showed spatial variations in 57 shared metabolites, accounting for 62% of the observed variation between morphotypes. The greatest variations in metabolite abundance were observed between the cultured morphotypes compared to the natural colony morphotypes, suggesting substantial differences in metabolite production between the cultivable strain IMS101 and the naturally occurring colony morphotypes that the cultivable strain is meant to represent. This study highlights the variations in metabolite abundance between natural and cultured Trichodesmium morphotypes and provides valuable insights into metabolites common to morphologically distinct Trichodesmium colonies, offering a foundation for future targeted metabolomic investigations.IMPORTANCEThis work demonstrates that the application of spatial mass spectrometry imaging at single-colony resolution can successfully resolve metabolite differences between natural and cultured Trichodesmium morphotypes, shedding light on their distinct biochemical profiles. Understanding the morphological differences between Trichodesmium colonies is crucial because they impact nutrient uptake, organic molecule production, and carbon and nitrogen export, and subsequently influence ocean biogeochemical cycles. As such, our study serves as an important initial assessment of metabolite differences between distinct Trichodesmium colony types, identifying features that can serve as ideal candidates for future targeted metabolomic studies., Competing Interests: The authors declare no conflict of interest.

Cyanobacteria of the genus Trichodesmium form extensive blooms that supply new N to nutrient-poor marine ecosystems. Yet little is known about what eats Trichodesmium . In this laboratory study, we show that one of the greatest threats to coral reefs, predatory crown-of-thorns starfish (CoTS), Acanthaster sp., completes their larval phase feeding solely on Trichodesmium . We observed Trichodesmium erythraeum CMP1985 in the stomachs of larvae using florescence microscopy and traced the assimilation of nitrogen from labeled trichomes into larval tissues using stable isotopes. Some larvae fed T. erythraeum were morphologically ready to become benthic juveniles after 19 days. Given that Trichodesmium can be food for CoTS, reported increases in Trichodesmium could be a driving factor in the heightened frequency of CoTS population irruptions that have devastated coral reefs in past decades. Future studies could test this through investigating the diets of wild larvae and incorporating Trichodesmium abundance into models of CoTS population dynamics.

While the ecological role that Trichodesmium sp. play in nitrogen fixation has been widely studied, little information is available on potential specialized metabolites that are associated with blooms and standing stock Trichodesmium colonies. While a collection of biological material from a T. thiebautii bloom event from North Padre Island, Texas, in 2014 indicated that this species was a prolific producer of chlorinated specialized metabolites, additional spatial and temporal resolution was needed. We have completed these metabolite comparison studies, detailed in the current report, utilizing LC-MS/MS-based molecular networking to visualize and annotate the specialized metabolite composition of these Trichodesmium blooms and colonies in the Gulf of Mexico (GoM) and other waters. Our results showed that T. thiebautii blooms and colonies found in the GoM have a remarkably consistent specialized metabolome. Additionally, we isolated and characterized one new macrocyclic compound from T. thiebautii , trichothilone A ( 1 ), which was also detected in three independent cultures of T. erythraeum . Genome mining identified genes predicted to synthesize certain functional groups in the T. thiebautii metabolites. These results provoke intriguing questions of how these specialized metabolites affect Trichodesmium ecophysiology, symbioses with marine invertebrates, and niche development in the global oligotrophic ocean.

Biological nitrogen (N 2 ) fixation is critical in global biogeochemical cycles and in sustaining the productivity of the oceans. There remain many unanswered questions, unresolved hypotheses, and unchallenged paradigms. The fundamental balance of N input and losses has not been fully resolved. One of the major N 2 -fixers, Trichodesmium, remains an enigma with intriguing biological and ecological secrets. Cyanobacterial N 2 fixation, once thought to be primarily due to free-living cyanobacteria, now also appears to be dependent on microbial interactions, from microbiomes to unicellular symbioses, which remain poorly characterized. Nitrogenase genes associated with diverse non-cyanobacterial diazotrophs (NCDs) are prevalent, but their significance remains a huge knowledge gap. Answering questions, new and old, such as those discussed here, is needed to understand the ocean's N and C cycles and their responses to environmental change., Competing Interests: Declaration of interests No interests are declared., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Nitrogen fixation by the diazotrophic cyanobacterium Trichodesmium contributes up to 50% of the bioavailable nitrogen in the ocean. N 2 fixation by Trichodesmium is limited by the availability of nutrients, such as iron (Fe) and phosphorus (P). Although colloids are ubiquitous in the ocean, the effects of Fe limitation on nitrogen fixation by marine colloids (MC) and the related mechanisms are largely unexplored. In this study, we found that MC exhibit photoelectrochemical properties that boost nitrogen fixation by photoelectrophy in Trichodesmium erythraeum . MC efficiently promote photosynthesis in T. erythraeum , thus enhancing its growth. Photoexcited electrons from MC are directly transferred to the photosynthetic electron transport chain and contribute to nitrogen fixation and ammonia assimilation. Transcriptomic analysis revealed that MC significantly upregulates genes related to the electron transport chain, photosystem, and photosynthesis, which is consistent with elevated photosynthetic capacities (e.g., F v / F m and carboxysomes). As a result, MC increase the N 2 fixation rate by 67.5-89.3%. Our findings highlight a proof-of-concept electron transfer pathway by which MC boost nitrogen fixation, broadening our knowledge on the role of ubiquitous colloids in marine nitrogen biogeochemistry.

Diazotrophic cyanobacteria such as Trichodesmium play a crucial role in the nitrogen budget of the oceans due to their capability to bind atmospheric nitrogen. Little is known about their interoceanic transport pathways and their distribution in upwelling regions. Trichodesmium has been detected using a Video Plankton Recorder (VPR) mounted on a remotely operated towed vehicle (TRIAXUS) in the southern and northern Benguela Upwelling System (BUS) in austral autumn, Feb/Mar 2019. The TRIAXUS, equipped with a CTD as well as fluorescence and nitrogen sensors, was towed at a speed of 8 kn on two onshore-offshore transects undulating between 5 and 200 m over distances of 249 km and 372 km, respectively. Trichodesmium was not detected near the coast in areas of freshly upwelled waters but was found in higher abundances offshore on both transects, mainly in subsurface water layers down to 80 m depth with elevated salinities. These salinity lenses can be related to northward moving eddies that most probably have been detached from the warm and salty Agulhas Current. Testing for interaction and species-habitat associations of Trichodesmium colonies with salinity yielded significant results, indicating that Trichodesmium may be transported with Agulhas Rings from the Indian Ocean into the Atlantic Ocean., (© 2024. The Author(s).)

The aim of this study was to investigate how acclimation to medium-level, long-term, non-lethal iron limitation changes the electron flux around the Photosystem II of the oceanic diazotroph Trichodesmium erythraeum IMS101. Fe availability of about 5× and 100× lower than a replete level, i.e. conditions common in the natural environment of this cyanobacterium, were applied in chemostats. The response of the cells was studied not only in terms of growth, but also mechanistically, measuring the chlorophyll fluorescence of dark-adapted filaments via imaging fluorescence kinetic microscopy (FKM) with 0.3 ms time resolution. Combining these measurements with those of metal binding to proteins via online coupling of metal-free HPLC (size exclusion chromatography SEC) to sector-field ICP-MS allowed to track the fate of the photosystems, together with other metalloproteins. General increase of fluorescence has been observed, with the consequent decrease in the quantum yields φ of the PSII, while the efficiency ψ of the electron flux between PSII and the PSI remained surprisingly unchanged. This indicates the ability of Trichodesmium to cope with a situation that makes assembling the many iron clusters in Photosystem I a particular challenge, as shown by decreasing ratios of Fe to Mg in these proteins. The negative effect of Fe limitation on PSII may also be due to its fast turnover. A broader view was obtained from metalloproteomics via HPLC-ICP-MS, revealing a differential protein expression pattern under iron limitation with a drastic down-regulation especially of iron-containing proteins and some increase in low MW metal-binding complexes., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Hendrik Kuepper reports financial support was provided by Czech Academy of Sciences. Hendrik Kuepper reports financial support was provided by Ministry of Education Youth and Sports of the Czech Republic. Hendrik Kuepper reports financial support was provided by COST Association., (Copyright © 2023 Elsevier B.V. All rights reserved.)

There is considerable debate about the benefits and trade-offs for colony formation in a major marine nitrogen fixer, Trichodesmium . To quantitatively analyze the trade-offs, we developed a metabolic model based on carbon fluxes to compare the performance of Trichodesmium colonies and free trichomes under different scenarios. Despite reported reductions in carbon fixation and nitrogen fixation rates for colonies relative to free trichomes, we found that model colonies can outperform individual cells in several cases. The formation of colonies can be advantageous when respiration rates account for a high proportion of the carbon fixation rate. Negative external influence on vital rates, such as mortality due to predation or micronutrient limitations, can also create a net benefit for colony formation relative to individual cells. In contrast, free trichomes also outcompete colonies in many scenarios, such as when respiration rates are equal for both colonies and individual cells or when there is a net positive external influence on rate processes (i.e., optimal environmental conditions regarding light and temperature or high nutrient availability). For both colonies and free trichomes, an increase in carbon fixation relative to nitrogen fixation rates would increase their relative competitiveness. These findings suggest that the formation of colonies in Trichodesmium might be linked to specific environmental and ecological circumstances. Our results provide a road map for empirical studies and models to evaluate the conditions under which colony formation in marine phytoplankton can be sustained in the natural environment. IMPORTANCE Trichodesmium is a marine filamentous cyanobacterium that fixes nitrogen and is an important contributor to the global nitrogen cycle. In the natural environment, Trichodesmium can exist as individual cells (trichomes) or as colonies (puffs and tufts). In this paper, we try to answer a longstanding question in marine microbial ecology: how does colony formation benefit the survival of Trichodesmium ? To answer this question, we developed a carbon flux model that utilizes existing published rates to evaluate whether and when colony formation can be sustained. Enhanced respiration rates, influential external factors such as environmental conditions and ecological interactions, and variable carbon and nitrogen fixation rates can all create scenarios for colony formation to be a viable strategy. Our results show that colony formation is an ecologically beneficial strategy under specific conditions, enabling Trichodesmium to be a globally significant organism.

The photosynthetic cyanobacterium Trichodesmium is widely distributed in the surface low latitude ocean where it contributes significantly to N 2 fixation and primary productivity. Previous studies found nifH genes and intact Trichodesmium colonies in the sunlight-deprived meso- and bathypelagic layers of the ocean (200-4000 m depth). Yet, the ability of Trichodesmium to fix N 2 in the dark ocean has not been explored. We performed 15 N 2 incubations in sediment traps at 170, 270 and 1000 m at two locations in the South Pacific. Sinking Trichodesmium colonies fixed N 2 at similar rates than previously observed in the surface ocean (36-214 fmol N cell -1 d -1 ). This activity accounted for 40 ± 28% of the bulk N 2 fixation rates measured in the traps, indicating that other diazotrophs were also active in the mesopelagic zone. Accordingly, cDNA nifH amplicon sequencing revealed that while Trichodesmium accounted for most of the expressed nifH genes in the traps, other diazotrophs such as Chlorobium and Deltaproteobacteria were also active. Laboratory experiments simulating mesopelagic conditions confirmed that increasing hydrostatic pressure and decreasing temperature reduced but did not completely inhibit N 2 fixation in Trichodesmium. Finally, using a cell metabolism model we predict that Trichodesmium uses photosynthesis-derived stored carbon to sustain N 2 fixation while sinking into the mesopelagic. We conclude that sinking Trichodesmium provides ammonium, dissolved organic matter and biomass to mesopelagic prokaryotes., (© 2022. The Author(s), under exclusive licence to International Society for Microbial Ecology.)

The dominant marine filamentous N 2 fixer, Trichodesmium , conducts photosynthesis and N 2 fixation during the daytime. Because N 2 fixation is sensitive to O 2 , some previous studies suggested that spatial segregation of N 2 fixation and photosynthesis is essential in Trichodesmium . However, this hypothesis conflicts with some observations where all the cells contain both photosystems and the N 2 -fixing enzyme nitrogenase. Here, we construct a systematic model simulating Trichodesmium metabolism, showing that the hypothetical spatial segregation is probably useless in increasing the Trichodesmium growth and N 2 fixation, unless substances can efficiently transfer among cells with low loss to the environment. The model suggests that Trichodesmium accumulates fixed carbon in the morning and uses that in respiratory protection to reduce intracellular O 2 during the mid-daytime, when photosynthesis is downregulated, allowing the occurrence of N 2 fixation. A cell membrane barrier against O 2 and alternative non-O 2 evolving electron transfer also contribute to maintaining low intracellular O 2 . Our study provides a mechanism enabling N 2 fixation despite the presence of photosynthesis across Trichodesmium . IMPORTANCE The filamentous Trichodesmium is a globally prominent marine nitrogen fixer. A long-standing paradox is that the nitrogen-fixing enzyme nitrogenase is sensitive to oxygen, but Trichodesmium conducts both nitrogen fixation and oxygen-evolving photosynthesis during the daytime. Previous studies using immunoassays reported that nitrogenase was limited in some specialized Trichodesmium cells (termed diazocytes), suggesting the necessity of spatial segregation of nitrogen fixation and photosynthesis. However, attempts using other methods failed to find diazocytes in Trichodesmium , causing controversy on the existence of the spatial segregation. Here, our physiological model shows that Trichodesmium can maintain low intracellular O 2 in mid-daytime and achieve feasible nitrogen fixation and growth rates even without the spatial segregation, while the hypothetical spatial segregation might not be useful if substantial loss of substances to the environment occurs when they transfer among the Trichodesmium cells. Our study then suggests a possible mechanism by which Trichodesmium can survive without the spatial segregation.

The filamentous diazotrophic cyanobacterium Trichodesmium is responsible for a significant fraction of marine di-nitrogen (N 2 ) fixation. Growth and distribution of Trichodesmium and other diazotrophs in the vast oligotrophic subtropical gyres is influenced by iron (Fe) and phosphorus (P) availability, while reciprocally influencing the biogeochemistry of these nutrients. Here we use observations across natural inverse gradients in Fe and P in the North Atlantic subtropical gyre (NASG) to demonstrate how Trichodesmium acclimates in situ to resource availability. Transcriptomic analysis identified progressive upregulation of known iron-stress biomarker genes with decreasing Fe availability, and progressive upregulation of genes involved in the acquisition of diverse P sources with decreasing P availability, while genes involved in N 2 fixation were upregulated at the intersection under moderate Fe and P availability. Enhanced N 2 fixation within the Fe and P co-stressed transition region was also associated with a distinct, consistent metabolic profile, including the expression of alternative photosynthetic pathways that potentially facilitate ATP generation required for N 2 fixation with reduced net oxygen production. The observed response of Trichodesmium to availability of both Fe and P supports suggestions that these biogeochemically significant organisms employ unique molecular, and thus physiological responses as adaptations to specifically exploit the Fe and P co-limited niche they construct., (© 2021. The Author(s).)

Ocean microbial communities are important contributors to the global biogeochemical reactions that sustain life on Earth. The factors controlling these communities are being increasingly explored using metatranscriptomic and metaproteomic environmental biomarkers. Using published proteomes and transcriptomes from the abundant colony-forming cyanobacterium Trichodesmium (strain IMS101) grown under varying Fe and/or P limitation in low and high CO 2 , we observed robust correlations of stress-induced proteins and RNAs (i.e., involved in transport and homeostasis) that yield useful information on the nutrient status under low and/or high CO 2 . Conversely, transcriptional and translational correlations of many other central metabolism pathways exhibit broad discordance. A cellular RNA and protein production/degradation model demonstrates how biomolecules with small initial inventories, such as environmentally responsive proteins, achieve large increases in fold-change units as opposed to those with a higher basal expression and inventory such as metabolic systems. Microbial cells, due to their immersion in the environment, tend to show large adaptive responses in both RNA and protein that result in transcript-protein correlations. These observations and model results demonstrate multi-omic coherence for environmental biomarkers and provide the underlying mechanism for those observations, supporting the promise for global application in detecting responses to environmental stimuli in a changing ocean.

In the surface waters of the warm oligotrophic ocean, filaments and aggregated colonies of the nitrogen (N)-fixing cyanobacterium Trichodesmium create microscale nutrient-rich oases. These hotspots fuel primary productivity and harbour a diverse consortium of heterotrophs. Interactions with associated microbiota can affect the physiology of Trichodesmium, often in ways that have been predicted to support its growth. Recently, it was found that trimethylamine (TMA), a globally abundant organic N compound, inhibits N 2 fixation in cultures of Trichodesmium without impairing growth rate, suggesting that Trichodesmium can use TMA as an alternate N source. In this study, 15 N-TMA DNA stable isotope probing (SIP) of a Trichodesmium enrichment was employed to further investigate TMA metabolism and determine whether TMA-N is incorporated directly or secondarily via cross-feeding facilitated by microbial associates. Herein, we identify two members of the marine Roseobacter clade (MRC) of Alphaproteobacteria as the likely metabolizers of TMA and provide genomic evidence that they converted TMA into a more readily available form of N, e.g., ammonium (NH 4 + ), which was subsequently used by Trichodesmium and the rest of the community. The results implicate microbiome-mediated carbon (C) and N transformations in modulating N 2 fixation and thus highlight the involvement of host-associated heterotrophs in global biogeochemical cycling., (© 2021 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Noninvasive fluorescent imaging requires far-red and near-infrared fluorescent proteins for deeper imaging. Near-infrared light penetrates biological tissue with blood vessels due to low absorbance, scattering, and reflection of light and has a greater signal-to-noise due to less autofluorescence. Far-red and near-infrared fluorescent proteins absorb light >600 nm to expand the color palette for imaging multiple biosensors and noninvasive in vivo imaging. The ideal fluorescent proteins are bright, photobleach minimally, express well in the desired cells, do not oligomerize, and generate or incorporate exogenous fluorophores efficiently. Coral-derived red fluorescent proteins require oxygen for fluorophore formation and release two hydrogen peroxide molecules. New fluorescent proteins based on phytochrome and phycobiliproteins use biliverdin IXα as fluorophores, do not require oxygen for maturation to image anaerobic organisms and tumor core, and do not generate hydrogen peroxide. The small Ultra-Red Fluorescent Protein (smURFP) was evolved from a cyanobacterial phycobiliprotein to covalently attach biliverdin as an exogenous fluorophore. The small Ultra-Red Fluorescent Protein is biophysically as bright as the enhanced green fluorescent protein, is exceptionally photostable, used for biosensor development, and visible in living mice. Novel applications of smURFP include in vitro protein diagnostics with attomolar (10-18 M) sensitivity, encapsulation in viral particles, and fluorescent protein nanoparticles. However, the availability of biliverdin limits the fluorescence of biliverdin-attaching fluorescent proteins; hence, extra biliverdin is needed to enhance brightness. New methods for improved biliverdin bioavailability are necessary to develop improved bright far-red and near-infrared fluorescent proteins for noninvasive imaging in vivo., (© 2020 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

During the summer of 2017, recurrent extensive blooms of the diazotrophic cyanobacterium Trichodesmium invaded the beaches and coastal waters of the Canary Islands, causing great social alarm. Some local media and public sectors ascribed, without any strong scientific evidence, the origin and reactivation of these blooms to untreated sewage outfalls distributed along the coasts. In order to test whether sewage outfalls could have any influence on the metabolic activity of Trichodesmium, we performed 13 C and 15 N 2 uptake experiments with colonies experiencing three different bloom development stages, incubated both with clear seawater and sewage water from an outfall south of Gran Canaria island. Our results showed that sewage outfalls did not promote any increase in dinitrogen (N 2 ) fixation in Trichodesmium, supporting the hypothesis that decaying blooms were generated offshore and transported shoreward by local currents and winds, accumulating mostly leeward of the islands. The combination of unusually warm seawater temperatures, enhanced and sustained stratification of the upper water column and recurrent dust deposition events would have favored the development of the Trichodesmium blooms, which lasted for at least four months.

Mixotrophy, the combination of heterotrophic and autotrophic nutrition modes, is emerging as the rule rather than the exception in marine photosynthetic plankton. Trichodesmium, a prominent diazotroph ubiquitous in the (sub)tropical oceans, is generally considered to obtain energy via autotrophy. While the ability of Trichodesmium to use dissolved organic phosphorus when deprived of inorganic phosphorus sources is well known, the extent to which this important cyanobacterium may benefit from other dissolved organic matter (DOM) resources is unknown. Here we provide evidence of carbon-, nitrogen- and phosphorus-rich DOM molecules enhancing N2 fixation rates and nifH gene expression in natural Trichodesmium colonies collected at two stations in the western tropical South Pacific. Sampling at a third station located in the oligotrophic South Pacific Gyre revealed no Trichodesmium but showed presence of UCYN-B, although no nifH expression was detected. Our results suggest that Trichodesmium behaves mixotrophically in response to certain environmental conditions, providing them with metabolic plasticity and adding up to the view that mixotrophy is widespread among marine microbes., (© FEMS 2020.)

Trichodesmium is an important dinitrogen (N 2 )-fixing cyanobacterium in marine ecosystems. Recent nucleic acid analyses indicate that Trichodesmium colonies with their diverse epibionts support various nitrogen (N) transformations beyond N 2 fixation. However, rates of these transformations and concentration gradients of N compounds in Trichodesmium colonies remain largely unresolved. We combined isotope-tracer incubations, micro-profiling and numeric modelling to explore carbon fixation, N cycling processes as well as oxygen, ammonium and nitrate concentration gradients in individual field-sampled Trichodesmium colonies. Colonies were net-autotrophic, with carbon and N 2 fixation occurring mostly during the day. Ten percent of the fixed N was released as ammonium after 12-h incubations. Nitrification was not detectable but nitrate consumption was high when nitrate was added. The consumed nitrate was partly reduced to ammonium, while denitrification was insignificant. Thus, the potential N transformation network was characterised by fixed N gain and recycling processes rather than denitrification. Oxygen concentrations within colonies were ~60-200% air-saturation. Moreover, our modelling predicted steep concentration gradients, with up to 6-fold higher ammonium concentrations, and nitrate depletion in the colony centre compared to the ambient seawater. These gradients created a chemically heterogeneous microenvironment, presumably facilitating diverse microbial metabolisms in millimetre-sized Trichodesmium colonies.

In natural environments, cells live in complex communities and experience a high degree of heterogeneity internally and in the environment. Even in 'ideal' laboratory environments, cells can experience a high degree of heterogeneity in their environments. Unfortunately, most of the metabolic modeling approaches that are currently used assume ideal conditions and that each cell is identical, limiting their application to pure cultures in well-mixed vessels. Here we describe our development of Multiscale Multiobjective Systems Analysis (MiMoSA), a metabolic modeling approach that can track individual cells in both space and time, track the diffusion of nutrients and light and the interaction of cells with each other and the environment. As a proof-of concept study, we used MiMoSA to model the growth of Trichodesmium erythraeum, a filamentous diazotrophic cyanobacterium which has cells with two distinct metabolic modes. The use of MiMoSA significantly improves our ability to predictively model metabolic changes and phenotype in more complex cell cultures.

Growth and dinitrogen (N 2 ) fixation of the globally important diazotrophic cyanobacteria Trichodesmium are often limited by iron (Fe) availability in surface seawaters. To systematically examine the combined effects of Fe limitation and ocean acidification (OA), T. erythraeum strain IMS101 was acclimated to both Fe-replete and Fe-limited concentrations under ambient and acidified conditions. Proteomic analysis showed that OA affected a wider range of proteins under Fe-limited conditions compared to Fe-replete conditions. OA also led to an intensification of Fe deficiency in key cellular processes (e.g., photosystem I and chlorophyll a synthesis) in already Fe-limited T. erythraeum. This is a result of reallocating Fe from these processes to Fe-rich nitrogenase to compensate for the suppressed N 2 fixation. To alleviate the Fe shortage, the diazotroph adopts a series of Fe-based economic strategies (e.g., upregulating Fe acquisition systems for organically complexed Fe and particulate Fe, replacing ferredoxin by flavodoxin, and using alternative electron flow pathways to produce ATP). This was more pronounced under Fe-limited-OA conditions than under Fe limitation only. Consequently, OA resulted in a further decrease of N 2 - and carbon-fixation rates in Fe-limited T. erythraeum. In contrast, Fe-replete T. erythraeum induced photosystem I (PSI) expression to potentially enhance the PSI cyclic flow for ATP production to meet the higher demand for energy to cope with the stress caused by OA. Our study provides mechanistic insight into the holistic response of the globally important N 2 -fixing marine cyanobacteria Trichodesmium to acidified and Fe-limited conditions of future oceans.

Trichodesmium is a globally important marine nitrogen fixing cyanobacteria which forms colonies and utilizes atmospherically derived dust as a source for the limiting micro-nutrient iron. Here we report the identification of metallophores isolated from incubations of natural Trichodesmium colonies collected from the Gulf of Aqaba in the Red Sea. Three of our compounds were identified as the ferrioxamine siderophores B, E, and G. The remaining fifteen metallophores had mass to charge ratios that, to our knowledge, are not common to known siderophores. Putative sum formulas suggest most of these compounds were not structurally related to each other. We also found that the novel metallophores readily formed complexes with aluminium and were less specific for iron than the ferrioxamines. In our incubations of Trichodesmium colonies, the abundance of ten of the novel metallophores positively correlated with Trichodesmium biomass, but not with bacterial biomass, whilst ferrioxamine siderophores were more strongly associated with bacterial biomass. We identified ferrioxamines and our novel metallophores in filtered surface seawater samples from the Gulf of Aqaba. However, our novel metallophores were only observed in the surface seawater sample collected at the time of highest Trichodesmium abundance, while ferrioxamines were observed even when Trichodesmium was not present. We hypothesize that the novel metallophores were specifically associated with Trichodesmium colonies. Together with the bacterially produced ferrioxamines they likely contribute to a distinctive "ligandosphere" surrounding the Trichodesmium colonies, with potential implications for metal homeostasis within the colony environment.

Iron (Fe) bioavailability limits phytoplankton growth in vast ocean regions. Iron-rich dust uplifted from deserts is transported in the atmosphere and deposited on the ocean surface. However, this dust is a poor source of iron for most phytoplankton since dust-bound Fe is poorly soluble in seawater and dust rapidly sinks out of the photic zone. An exception is Trichodesmium , a globally important, N 2 fixing, colony forming, cyanobacterium, which efficiently captures and shuffles dust to its colony core. Trichodesmium and bacteria that reside within its colonies carry out diverse metabolic interactions. Here we show evidence for mutualistic interactions between Trichodesmium and associated bacteria for utilization of iron from dust, where bacteria promote dust dissolution by producing Fe-complexing molecules (siderophores) and Trichodesmium provides dust and optimal physical settings for dissolution and uptake. Our results demonstrate how intricate relationships between producers and consumers can influence productivity in the nutrient starved open ocean., Competing Interests: Competing interestsThe authors declare no competing interests.

The response of the prominent marine dinitrogen (N 2 )-fixing cyanobacteria Trichodesmium to ocean acidification (OA) is critical to understanding future oceanic biogeochemical cycles. Recent studies have reported conflicting findings on the effect of OA on growth and N 2 fixation of Trichodesmium. Here, we quantitatively analyzed experimental data on how Trichodesmium reallocated intracellular iron and energy among key cellular processes in response to OA, and integrated the findings to construct an optimality-based cellular model. The model results indicate that Trichodesmium growth rate decreases under OA primarily due to reduced nitrogenase efficiency. The downregulation of the carbon dioxide (CO 2 )-concentrating mechanism under OA has little impact on Trichodesmium, and the energy demand of anti-stress responses to OA has a moderate negative effect. We predict that if anthropogenic CO 2 emissions continue to rise, OA could reduce global N 2 fixation potential of Trichodesmium by 27% in this century, with the largest decrease in iron-limiting regions.

As atmospheric CO2 concentrations increase, so too does the dissolved CO2 and HCO3- concentrations in the world's oceans. There are still many uncertainties regarding the biological response of key groups of organisms to these changing conditions, which is crucial for predicting future species distributions, primary productivity rates, and biogeochemical cycling. In this study, we established the relationship between gross photosynthetic O2 evolution and light-dependent O2 consumption in Trichodesmium erythraeum IMS101 acclimated to three targeted pCO2 concentrations (180 µmol mol-1=low-CO2, 380 µmol mol-1=mid-CO2, and 720 µmol mol-1=high-CO2). We found that biomass- (carbon) specific, light-saturated maximum net O2 evolution rates (PnC,max) and acclimated growth rates increased from low- to mid-CO2, but did not differ significantly between mid- and high-CO2. Dark respiration rates were five times higher than required to maintain cellular metabolism, suggesting that respiration provides a substantial proportion of the ATP and reductant for N2 fixation. Oxygen uptake increased linearly with gross O2 evolution across light intensities ranging from darkness to 1100 µmol photons m-2 s-1. The slope of this relationship decreased with increasing CO2, which we attribute to the increased energetic cost of operating the carbon-concentrating mechanism at lower CO2 concentrations. Our results indicate that net photosynthesis and growth of T. erythraeum IMS101 would have been severely CO2 limited at the last glacial maximum, but that the direct effect of future increases of CO2 may only cause marginal increases in growth.

Atmospheric nitrogen fixation by photosynthetic cyanobacteria (diazotrophs) strongly influences oceanic primary production and in turn affects global biogeochemical cycles. Species of the genus Trichodesmium are major contributors to marine diazotrophy, accounting for a significant proportion of the fixed nitrogen in tropical and subtropical oceans. However, Trichodesmium spp. are metabolically constrained by the availability of iron, an essential element for both the photosynthetic apparatus and the nitrogenase enzyme. Survival strategies in low-iron environments are typically poorly characterized at the molecular level, because these bacteria are recalcitrant to genetic manipulation. Here, we studied a homolog of the iron deficiency-induced A (IdiA)/ferric uptake transporter A (FutA) protein, Tery_3377, which has been used as an in situ iron-stress biomarker. IdiA/FutA has an ambiguous function in cyanobacteria, with its homologs hypothesized to be involved in distinct processes depending on their cellular localization. Using signal sequence fusions to GFP and heterologous expression in the model cyanobacterium Synechocystis sp. PCC 6803, we show that Tery_3377 is targeted to the periplasm by the twin-arginine translocase and can complement the deletion of the native Synechocystis ferric-iron ABC transporter periplasmic binding protein (FutA2). EPR spectroscopy revealed that purified recombinant Tery_3377 has specificity for iron in the Fe 3+ state, and an X-ray crystallography-determined structure uncovered a functional iron substrate-binding domain, with Fe 3+ pentacoordinated by protein and buffer ligands. Our results support assignment of Tery_3377 as a functional FutA subunit of an Fe 3+ ABC transporter but do not rule out dual IdiA function., (© 2018 Polyviou et al.)

Trichodesmium spp. are diazotrophic cyanobacteria that exist as single filaments (trichomes) and as macroscopic colonies of varying shapes formed by aggregating trichomes. The causes and dynamics of colony formation and disassociation are not yet elucidated. we demonstrate that limited availability of dissolved phosphorus (P) or iron (Fe) stimulated trichome mobility and induced colony formation in Trichodesmium erythraeum IMS101 cultures. The specific nutrient limitation differentially affected the rate of colony formation and morphology of the colonies. Fe starvation promoted rapid colony formation (10-48 h from depletion) while 5-7 days were required for colonies to form in P-depleted cultures. Video analyses confirmed that the probability of trichomes to cluster increased from 12 to 35% when transferred from nutrient replete to Fe-depleted conditions. Moreover, the probability for Fe-depleted aggregates to remain colonial increased to 50% from only 10% in nutrient replete cultures. These colonies were also characterized by stronger attachment forces between the trichomes. Enrichment of nutrient-depleted cultures with the limited nutrient-stimulated colony dissociation into single trichomes. We postulate that limited P and Fe availability enhance colony formation of Trichodesmium and primarily control the abundance and distribution of its different morphologies in the nutrient-limited surface ocean.

The N 2 -fixing cyanobacterium Trichodesmium is intensely studied because of the control this organism exerts over the cycling of carbon and nitrogen in the low nutrient ocean gyres. Although iron (Fe) and phosphorus (P) bioavailability are thought to be major drivers of Trichodesmium distributions and activities, identifying resource controls on Trichodesmium is challenging, as Fe and P are often organically complexed and their bioavailability to a single species in a mixed community is difficult to constrain. Further, Fe and P geochemistries are linked through the activities of metalloenzymes, such as the alkaline phosphatases (APs) PhoX and PhoA, which are used by microbes to access dissolved organic P (DOP). Here we identified significant correlations between Trichodesmium-specific transcriptional patterns in the North Atlantic (NASG) and North Pacific Subtropical Gyres (NPSG) and patterns in Fe and P biogeochemistry, with the relative enrichment of Fe stress markers in the NPSG, and P stress markers in the NASG. We also observed the differential enrichment of Fe-requiring PhoX transcripts in the NASG and Fe-insensitive PhoA transcripts in the NPSG, suggesting that metalloenzyme switching may be used to mitigate Fe limitation of DOP metabolism in Trichodesmium. This trait may underpin Trichodesmium success across disparate ecosystems.

Trichodesmium plays a significant role in the oligotrophic oceans, fixing nitrogen in an area corresponding to half of the Earth's surface, representing up to 50% of new production in some oligotrophic tropical and subtropical oceans. Whilst Trichodesmium blooms at the surface exhibit a strong dependence on diazotrophy, colonies at depth or at the surface after a mixing event could be utilising additional N-sources. We conducted experiments to establish how acclimation to varying N-sources affects the growth, elemental composition, light absorption coefficient, N2 fixation, PSII electron transport rate and the relationship between net and gross photosynthetic O2 exchange in T. erythraeum IMS101. To do this, cultures were acclimated to growth medium containing NH4+ and NO3- (replete concentrations) or N2 only (diazotrophic control). The light dependencies of O2 evolution and O2 uptake were measured using membrane inlet mass spectrometry (MIMS), while PSII electron transport rates were measured from fluorescence light curves (FLCs). We found that at a saturating light intensity, Trichodesmium growth was ~ 10% and 13% lower when grown on N2 than with NH4+ and NO3-, respectively. Oxygen uptake increased linearly with net photosynthesis across all light intensities ranging from darkness to 1100 μmol photons m-2 s-1. The maximum rates and initial slopes of light response curves for C-specific gross and net photosynthesis and the slope of the relationship between gross and net photosynthesis increased significantly under non-diazotrophic conditions. We attribute these observations to a reduced expenditure of reductant and ATP for nitrogenase activity under non-diazotrophic conditions which allows NADPH and ATP to be re-directed to CO2 fixation and/or biosynthesis. The energy and reductant conserved through utilising additional N-sources could enhance Trichodesmium's productivity and growth and have major implications for its role in ocean C and N cycles.

Compared to extensive studies reporting the aluminum (Al) toxicity to terrestrial plants and freshwater organisms, very little is known about how marine phytoplankton responds to Al in the field. Here we report the marine phytoplankton responses to Al enrichment in the South China Sea (SCS) using on-deck bottle incubation experiments during eight cruises from May 2010 to November 2013. Generally, Al addition alone enhanced the growth of diatom and Trichodesmium, and nitrogen fixation, but it inhibited the growth of dinoflagellates and Synechococcus. Nevertheless, Al addition alone did not influence the chlorophyll a concentration of the entire phytoplankton assemblages. By adding nitrate and phosphate simultaneously, Al enrichment led to substantial increases in chlorophyll a concentration (especially that of the picophytoplankton<3μm), and cell abundances of diatom and photosynthetic picoeukaryotes. These results indicate varied responses of phytoplankton in different size fractions and taxonomic groups to Al enrichment. Further, by simultaneously adding different macronutrients and/or sufficient trace metals including iron, we found that the phytoplankton responses to Al enrichment were relevant to nutrients coexisting in the environment. Al enrichment may give some phytoplankton a competitive edge over using nutrients, especially the limited ones. The possible influences of Al on the competitors and grazers (predators) of some phytoplankton might indirectly contribute to the positive responses of the phytoplankton to Al enrichment. Our results indicate that Al may influence marine carbon cycle by impacting phytoplankton growth and structure in natural seawater., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Trichodesmium is a widespread, N 2 fixing marine cyanobacterium that drives inputs of newly fixed nitrogen and carbon into the oligotrophic ecosystems where it occurs. Colonies of Trichodesmium ubiquitously occur with heterotrophic bacteria that make up a diverse microbiome, and interactions within this Trichodesmium holobiont could influence the fate of fixed carbon and nitrogen. Metatranscriptome sequencing was performed on Trichodesmium colonies collected during high-frequency Lagrangian sampling in the North Pacific Subtropical Gyre (NPSG) to identify possible interactions between the Trichodesmium host and microbiome over day-night cycles. Here we show significantly coordinated patterns of gene expression between host and microbiome, many of which had significant day-night periodicity. The functions of the co-expressed genes suggested a suite of interactions within the holobiont linked to key resources including nitrogen, carbon, and iron. Evidence of microbiome reliance on Trichodesmium-derived vitamin B12 was also detected in co-expression patterns, highlighting a dependency that could shape holobiont community structure. Collectively, these patterns of expression suggest that biotic interactions could influence colony cycling of resources like nitrogen and vitamin B12, and decouple activities, like N 2 fixation, from typical abiotic drivers of Trichodesmium physiological ecology.

Distribution of diazotrophs and their nitrogen fixation activity were investigated in the northern South China Sea (nSCS) and the Kuroshio from July 16th to September 1st, 2009. N 2 fixation activities in whole seawater and <10 μm fraction at the surface were measured by acetylene reduction assay. Higher activities were observed at the East China Sea (ECS) Kuroshio and the nSCS shelf. The nSCS basin showed a low N 2 fixation activity. The <10 μm fractions (unicellular diazotrophs) contributed major portion to the whole-water activity in the survey time, indicating that nanoplanktonic cyanobacterias were the major diazotrophs in the survey area. Daily N 2 fixation rates of Trichodesmium ranged from 0.11 to 9.83 pmolNtrichome -1 d -1 with an average of 4.03 pmolNtrichome -1 d -1 . The Luzon Strait and the ECS Kuroshio had higher N 2 fixation rates of Trichodesmium than the nSCS shelf and basin. Calculated activities of Trichodesmium at most stations were moderately low compared with that of the whole-water. The contribution of N 2 fixation by the whole-water to primary production ranged from 1.7% to 18.5%. The estimated amount of new nitrogen introduced by Trichodesmium contributed up to 0.14% of the total primary production and 0.41% of the new production in the Luzon Strait.

Nitrogen-fixing (N 2 ) cyanobacteria provide bioavailable nitrogen to vast ocean regions but are in turn limited by iron (Fe) and/or phosphorus (P), which may force them to employ alternative nitrogen acquisition strategies. The adaptive responses of nitrogen fixers to global-change drivers under nutrient-limited conditions could profoundly alter the current ocean nitrogen and carbon cycles. Here, we show that the globally important N 2 fixer Trichodesmium fundamentally shifts nitrogen metabolism toward organic-nitrogen scavenging following long-term high-CO 2 adaptation under iron and/or phosphorus (co)limitation. Global shifts in transcripts and proteins under high-CO 2 /Fe-limited and/or P-limited conditions include decreases in the N 2 -fixing nitrogenase enzyme, coupled with major increases in enzymes that oxidize trimethylamine (TMA). TMA is an abundant, biogeochemically important organic nitrogen compound that supports rapid Trichodesmium growth while inhibiting N 2 fixation. In a future high-CO 2 ocean, this whole-cell energetic reallocation toward organic nitrogen scavenging and away from N 2 fixation may reduce new-nitrogen inputs by Trichodesmium while simultaneously depleting the scarce fixed-nitrogen supplies of nitrogen-limited open-ocean ecosystems. IMPORTANCE Trichodesmium is among the most biogeochemically significant microorganisms in the ocean, since it supplies up to 50% of the new nitrogen supporting open-ocean food webs. We used Trichodesmium cultures adapted to high-CO 2 conditions for 7 years, followed by additional exposure to iron and/or phosphorus (co)limitation. We show that "future ocean" conditions of high CO 2 and concurrent nutrient limitation(s) fundamentally shift nitrogen metabolism away from nitrogen fixation and instead toward upregulation of organic nitrogen-scavenging pathways. We show that the responses of Trichodesmium to projected future ocean conditions include decreases in the nitrogen-fixing nitrogenase enzymes coupled with major increases in enzymes that oxidize the abundant organic nitrogen source trimethylamine (TMA). Such a shift toward organic nitrogen uptake and away from nitrogen fixation may substantially reduce new-nitrogen inputs by Trichodesmium to the rest of the microbial community in the future high-CO 2 ocean, with potential global implications for ocean carbon and nitrogen cycling., (Copyright © 2018 American Society for Microbiology.)

The diazotrophic cyanobacterium, Trichodesmium, is an integral component of the marine nitrogen cycle and contributes significant amounts of new nitrogen to oligotrophic, tropical/subtropical ocean surface waters. Trichodesmium forms macroscopic, fusiform (tufts), spherical (puffs) and raft-like colonies that provide a pseudobenthic habitat for a host of other organisms including marine invertebrates, microeukaryotes and numerous other microbes. The diversity and activity of denitrifying bacteria found in association with the colonies was interrogated using a series of molecular-based methodologies targeting the gene encoding the terminal step in the denitrification pathway, nitrous oxide reductase (nosZ). Trichodesmium spp. sampled from geographically isolated ocean provinces (the Atlantic Ocean, the Red Sea and the Indian Ocean) were shown to harbor highly similar, taxonomically related communities of denitrifiers whose members are affiliated with the Roseobacter clade within the Rhodobacteraceae (Alphaproteobacteria). These organisms were actively expressing nosZ in samples taken from the mid-Atlantic Ocean and Red Sea implying that Trichodesmium colonies are potential sites of nitrous oxide consumption and perhaps earlier steps in the denitrification pathway also. It is proposed that coupled nitrification of newly fixed N is the most likely source of nitrogen oxides supporting nitrous oxide cycling within Trichodesmium colonies., (© 2017 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

Inorganic phosphate is the major bioavailable form of the essential nutrient phosphorus. However, the concentration of phosphate in most natural habitats is low enough to limit microbial growth. Under phosphate-depleted conditions some bacteria utilise phosphite and hypophosphite as alternative sources of phosphorus, but the molecular basis of reduced phosphorus acquisition from the environment is not fully understood. Here, we present crystal structures and ligand binding affinities of periplasmic binding proteins from bacterial phosphite and hypophosphite ATP-binding cassette transporters. We reveal that phosphite and hypophosphite specificity results from a combination of steric selection and the presence of a P-H…π interaction between the ligand and a conserved aromatic residue in the ligand-binding pocket. The characterisation of high affinity and specific transporters has implications for the marine phosphorus redox cycle, and might aid the use of phosphite as an alternative phosphorus source in biotechnological, industrial and agricultural applications.

Trichodesmium is a genus of marine diazotrophic colonial cyanobacteria that exerts a profound influence on global biogeochemistry, by injecting 'new' nitrogen into the low nutrient systems where it occurs. Colonies of Trichodesmium ubiquitously contain a diverse assemblage of epibiotic microorganisms, constituting a microbiome on the Trichodesmium host. Metagenome sequences from Trichodesmium colonies were analyzed along a resource gradient in the western North Atlantic to examine microbiome community structure, functional diversity and metabolic contributions to the holobiont. Here we demonstrate the presence of a core Trichodesmium microbiome that is modulated to suit different ocean regions, and contributes over 10 times the metabolic potential of Trichodesmium to the holobiont. Given the ubiquitous nature of epibionts on colonies, the substantial functional diversity within the microbiome is likely an integral facet of Trichodesmium physiological ecology across the oligotrophic oceans where this biogeochemically significant diazotroph thrives.

Gradients of oxygen (O 2 ) and pH, as well as small-scale fluxes of carbon (C), nitrogen (N) and O 2 were investigated under different partial pressures of carbon dioxide (pCO 2 ) in field-collected colonies of the marine dinitrogen (N 2 )-fixing cyanobacterium Trichodesmium. Microsensor measurements indicated that cells within colonies experienced large fluctuations in O 2 , pH and CO 2 concentrations over a day-night cycle. O 2 concentrations varied with light intensity and time of day, yet colonies exposed to light were supersaturated with O 2 (up to ~200%) throughout the light period and anoxia was not detected. Alternating between light and dark conditions caused a variation in pH levels by on average 0.5 units (equivalent to 15 nmol l -1 proton concentration). Single-cell analyses of C and N assimilation using secondary ion mass spectrometry (SIMS; large geometry SIMS and nanoscale SIMS) revealed high variability in metabolic activity of single cells and trichomes of Trichodesmium, and indicated transfer of C and N to colony-associated non-photosynthetic bacteria. Neither O 2 fluxes nor C fixation by Trichodesmium were significantly influenced by short-term incubations under different pCO 2 levels, whereas N 2 fixation increased with increasing pCO 2 . The large range of metabolic rates observed at the single-cell level may reflect a response by colony-forming microbial populations to highly variable microenvironments.

Acidification of seawater caused by anthropogenic carbon dioxide (CO 2 ) is anticipated to influence the growth of dinitrogen (N 2 )-fixing phytoplankton, which contribute a large fraction of primary production in the tropical and subtropical ocean. We found that growth and N 2 -fixation of the ubiquitous cyanobacterium Trichodesmium decreased under acidified conditions, notwithstanding a beneficial effect of high CO 2 Acidification resulted in low cytosolic pH and reduced N 2 -fixation rates despite elevated nitrogenase concentrations. Low cytosolic pH required increased proton pumping across the thylakoid membrane and elevated adenosine triphosphate production. These requirements were not satisfied under field or experimental iron-limiting conditions, which greatly amplified the negative effect of acidification., (Copyright © 2017, American Association for the Advancement of Science.)

The globally distributed diazotroph Trichodesmium contributes importantly to nitrogen inputs in the oligotrophic oceans. Sites of dissolved organic matter (DOM) accumulation could promote the mixotrophic nutrition of Trichodesmium when inorganic nutrients are scarce. Nano-scale secondary ion mass spectrometry (nanoSIMS) analyses of individual trichomes sampled in the South Pacific Ocean, showed significant 13 C-enrichments after incubation with either 13 C-labeled carbohydrates or amino acids. These results suggest that DOM could be directly taken up by Trichodesmium or primarily consumed by heterotrophic epibiont bacteria that ultimately transfer reduced DOM compounds to their host trichomes. Although the addition of carbohydrates or amino acids did not significantly affect bulk N 2 fixation rates, N 2 fixation was enhanced by amino acids in individual colonies of Trichodesmium. We discuss the ecological advantages of DOM use by Trichodesmium as an alternative to autotrophic nutrition in oligotrophic open ocean waters.

Background: Computational, genome based predictions of organism phenotypes has enhanced the ability to investigate the biological phenomena that help organisms survive and respond to their environments. In this study, we have created the first genome-scale metabolic network reconstruction of the nitrogen fixing cyanobacterium T. erythraeum and used genome-scale modeling approaches to investigate carbon and nitrogen fluxes as well as growth and equilibrium population composition., Results: We created a genome-scale reconstruction of T. erythraeum with 971 reactions, 986 metabolites, and 647 unique genes. We then used data from previous studies as well as our own laboratory data to establish a biomass equation and two distinct submodels that correspond to the two cell types formed by T. erythraeum. We then use flux balance analysis and flux variability analysis to generate predictions for how metabolism is distributed to account for the unique productivity of T. erythraeum. Finally, we used in situ data to constrain the model, infer time dependent population compositions and metabolite production using dynamic Flux Balance Analysis. We find that our model predicts equilibrium compositions similar to laboratory measurements, approximately 15.5% diazotrophs for our model versus 10-20% diazotrophs reported in literature. We also found that equilibrium was the most efficient mode of growth and that equilibrium was stoichiometrically mediated. Moreover, the model predicts that nitrogen leakage is an essential condition of optimality for T. erythraeum; cells leak approximately 29.4% total fixed nitrogen when growing at the optimal growth rate, which agrees with values observed in situ., Conclusion: The genome-metabolic network reconstruction allows us to use constraints based modeling approaches to predict growth and optimal cellular composition in T. erythraeum colonies. Our predictions match both in situ and laboratory data, indicating that stoichiometry of metabolic reactions plays a large role in the differentiation and composition of different cell types. In order to realize the full potential of the model, advance modeling techniques which account for interactions between colonies, the environment and surrounding species need to be developed.

Nitrogen-fixing cyanobacteria in the genus Trichodesmium play a critical role in the productivity of the tropical and subtropical oligotrophic oceans. The ecological success of these populations is likely associated with the diverse microbial interactions occurring within the Trichodesmium holobiont, especially between Trichodesmium and heterotrophic bacterial epibionts. Yet, the composition of the Trichodesmium holobiont and the processes governing microbial assemblage are not well documented. Here, we used high-resolution 16S rDNA amplicon sequencing to examine the diversity of Trichodesmium and associated epibionts across different ocean regions and colony morphologies (puffs and rafts). Trichodesmium Clade I (i.e., T. thiebautii-like) dominated the colonies in all ocean basins regardless of morphology, although the Trichodesmium community structure significantly varied between morphologies in some regions. On average, Alphaproteobacteria (i.e., Thalassobius), Gammaproteobacteria (i.e., Pseudoalteromonas), Sphingobacteria (i.e., Microscilla and Vibrio) and Flavobacteria dominated the epibiont communities, but community composition and structure significantly differed between regions. Epibionts from the two colony morphologies were taxonomically and functionally distinct in the North Atlantic and North Pacific. These findings suggest that the colony types might define two distinct niches and that epibiont assemblage might be driven in part by selective processes, where epibionts are selected according to their influence on colony metabolism., (© 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.)

The oceanic N 2 -fixing cyanobacterium Trichodesmium spp. form extensive surface blooms and contribute significantly to marine carbon and nitrogen cycles in the oligotrophic subtropical and tropical oceans. Trichodesmium grows in salinities from 27 to 43 parts per thousand (ppt), yet its salt acclimation strategy remains enigmatic because the genome of Trichodesmium erythraeum strain IMS101 lacks all genes for the biosynthesis of any known compatible solute. Using NMR and liquid chromatography coupled to mass spectroscopy, we identified the main compatible solute in T. erythraeum strain IMS101 as the quaternary ammonium compound N,N,N-trimethyl homoserine (or homoserine betaine) and elucidated its biosynthetic pathway. The identification of this compatible solute explains how Trichodesmium spp. can thrive in the marine system at varying salinities and provides further insight into the diversity of microbial salt acclimation., Competing Interests: The authors declare no conflict of interest.

Far-red fluorescent proteins (FPs) are desirable for in vivo imaging because with these molecules less light is scattered, absorbed, or re-emitted by endogenous biomolecules compared with cyan, green, yellow, and orange FPs. We developed a new class of FP from an allophycocyanin α-subunit (APCα). Native APC requires a lyase to incorporate phycocyanobilin. The evolved FP, which we named small ultra-red FP (smURFP), covalently attaches a biliverdin (BV) chromophore without a lyase, and has 642/670-nm excitation-emission peaks, a large extinction coefficient (180,000 M(-1)cm(-1)) and quantum yield (18%), and photostability comparable to that of eGFP. smURFP has significantly greater BV incorporation rate and protein stability than the bacteriophytochrome (BPH) FPs. Moreover, BV supply is limited by membrane permeability, and smURFPs (but not BPH FPs) can incorporate a more membrane-permeant BV analog, making smURFP fluorescence comparable to that of FPs from jellyfish or coral. A far-red and near-infrared fluorescent cell cycle indicator was created with smURFP and a BPH FP.

Australia's tropical waters represent predicted 'hotspots' for nitrogen (N2) fixation based on empirical and modelled data. However, the identity, activity and ecology of diazotrophs within this region are virtually unknown. By coupling DNA and cDNA sequencing of nitrogenase genes (nifH) with size-fractionated N2 fixation rate measurements, we elucidated diazotroph dynamics across the shelf region of the Arafura and Timor Seas (ATS) and oceanic Coral Sea during Austral spring and winter. During spring, Trichodesmium dominated ATS assemblages, comprising 60% of nifH DNA sequences, while Candidatus Atelocyanobacterium thalassa (UCYN-A) comprised 42% in the Coral Sea. In contrast, during winter the relative abundance of heterotrophic unicellular diazotrophs (δ-proteobacteria and γ-24774A11) increased in both regions, concomitant with a marked decline in UCYN-A sequences, whereby this clade effectively disappeared in the Coral Sea. Conservative estimates of N2 fixation rates ranged from <1 to 91 nmol l(-1) day(-1), and size fractionation indicated that unicellular organisms dominated N2 fixation during both spring and winter, but average unicellular rates were up to 10-fold higher in winter than in spring. Relative abundances of UCYN-A1 and γ-24774A11 nifH transcripts negatively correlated to silicate and phosphate, suggesting an affinity for oligotrophy. Our results indicate that Australia's tropical waters are indeed hotspots for N2 fixation and that regional physicochemical characteristics drive differential contributions of cyanobacterial and heterotrophic phylotypes to N2 fixation.

We discuss a mathematical model of growth of two types of phytoplankton, non-nitrogen-fixing and nitrogen-fixing, that both require light in order to grow. We use general functional responses to represent the inhibitory effect their biomass has on the exposure to light. We give conditions for the existence and local stability of all of the possible steady-states (die out, single species survival, and coexistence). We derive conditions for global stability of the die out and single-species steady-states and for persistence of both species when the coexistence steady-state exists. Numerical investigation illustrates the qualitative dynamics demonstrating that even under constant environmental conditions, both stable intrinsic oscillatory behavior and a period doubling route to chaotic dynamics are possible. We also show that competitor-mediated coexistence can occur due to the positive feedback resulting from recycling by the nitrogen-fixing phytoplankton. To show the impact of seasonal change in water depth, we also allow the water depth to vary in an annual cycle and discuss echo blooms in this context.