Marine Colloids Boost Nitrogen Fixation in Trichodesmium erythraeum by Photoelectrophy.

Nitrogen fixation by the diazotrophic cyanobacterium Trichodesmium contributes up to 50% of the bioavailable nitrogen in the ocean. N2 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., Fv/Fm and carboxysomes). As a result, MC increase the N2 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.

Spatial and Temporal Resolution of Cyanobacterial Bloom Chemistry Reveals an Open-Ocean Trichodesmium thiebautii as a Talented Producer of Specialized Metabolites.

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.

Discovery of nondiazotrophic Trichodesmium species abundant and widespread in the open ocean.

Filamentous and colony-forming cells within the cyanobacterial genus Trichodesmium might account for nearly half of nitrogen fixation in the sunlit ocean, a critical mechanism that sustains plankton's primary productivity. Trichodesmium has long been portrayed as a diazotrophic genus. By means of genome-resolved metagenomics, here we reveal that nondiazotrophic Trichodesmium species not only exist but also are abundant and widespread in the open ocean, benefiting from a previously overlooked functional lifestyle to expand the biogeography of this prominent marine genus. Near-complete environmental genomes for those closely related candidate species reproducibly shared functional features including a lack of genes related to nitrogen fixation, hydrogen recycling, and hopanoid lipid production concomitant with the enrichment of nitrogen assimilation genes. Our results elucidate fieldwork observations of Trichodesmium cells fixing carbon but not nitrogen. The Black Queen hypothesis and burden of low-oxygen concentration requirements provide a rationale to explain gene loss linked to nitrogen fixation among Trichodesmium species. Disconnecting taxonomic signal for this genus from a microbial community's ability to fix nitrogen will help refine our understanding of the marine nitrogen balance. Finally, we are reminded that established links between taxonomic lineages and functional traits do not always hold true.

Why Environmental Biomarkers Work: Transcriptome-Proteome Correlations and Modeling of Multistressor Experiments in the Marine Bacterium Trichodesmium.

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 CO2, 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 CO2. 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.

Long-Term m5C Methylome Dynamics Parallel Phenotypic Adaptation in the Cyanobacterium Trichodesmium.

A major challenge in modern biology is understanding how the effects of short-term biological responses influence long-term evolutionary adaptation, defined as a genetically determined increase in fitness to novel environments. This is particularly important in globally important microbes experiencing rapid global change, due to their influence on food webs, biogeochemical cycles, and climate. Epigenetic modifications like methylation have been demonstrated to influence short-term plastic responses, which ultimately impact long-term adaptive responses to environmental change. However, there remains a paucity of empirical research examining long-term methylation dynamics during environmental adaptation in nonmodel, ecologically important microbes. Here, we show the first empirical evidence in a marine prokaryote for long-term m5C methylome modifications correlated with phenotypic adaptation to CO2, using a 7-year evolution experiment (1,000+ generations) with the biogeochemically important marine cyanobacterium Trichodesmium. We identify m5C methylated sites that rapidly changed in response to high (750 µatm) CO2 exposure and were maintained for at least 4.5 years of CO2 selection. After 7 years of CO2 selection, however, m5C methylation levels that initially responded to high-CO2 returned to ancestral, ambient CO2 levels. Concurrently, high-CO2 adapted growth and N2 fixation rates remained significantly higher than those of ambient CO2 adapted cell lines irrespective of CO2 concentration, a trend consistent with genetic assimilation theory. These data demonstrate the maintenance of CO2-responsive m5C methylation for 4.5 years alongside phenotypic adaptation before returning to ancestral methylation levels. These observations in a globally distributed marine prokaryote provide critical evolutionary insights into biogeochemically important traits under global change.

Alphaproteobacteria facilitate Trichodesmium community trimethylamine utilization.

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 N2 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 (NH4 + ), which was subsequently used by Trichodesmium and the rest of the community. The results implicate microbiome-mediated carbon (C) and N transformations in modulating N2 fixation and thus highlight the involvement of host-associated heterotrophs in global biogeochemical cycling.

A Rise in ROS and EPS Production: New Insights into the Trichodesmium erythraeum Response to Ocean Acidification.

The diazotrophic cyanobacterium Trichodesmium is thought to be a major contributor to the new N in parts of the oligotrophic, subtropical, and tropical oceans. In this study, physiological and biochemical methods and transcriptome sequencing were used to investigate the influences of ocean acidification (OA) on Trichodesmium erythraeum (T. erythraeum). We presented evidence that OA caused by CO2 slowed the growth rate and physiological activity of T. erythraeum. OA led to reduced development of proportion of the vegetative cells into diazocytes which included up-regulated genes of nitrogen fixation. Reactive oxygen species (ROS) accumulation was increased due to the disruption of photosynthetic electron transport and decrease in antioxidant enzyme activities under acidified conditions. This study showed that OA increased the amounts of (exopolysaccharides) EPS in T. erythraeum, and the key genes of ribose-5-phosphate (R5P) and glycosyltransferases (Tery_3818) were up-regulated. These results provide new insight into how ROS and EPS of T. erythraeum increase in an acidified future ocean to cope with OA-imposed stress.

Fluorescent proteins for in vivo imaging, where's the biliverdin?

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.

Projected expansion of Trichodesmium's geographical distribution and increase in growth potential in response to climate change.

Estimates of marine N2 fixation range from 52 to 73 Tg N/year, of which we calculate up to 84% is from Trichodesmium based on previous measurements of nifH gene abundance and our new model of Trichodesmium growth. Here, we assess the likely effects of four major climate change-related abiotic factors on the spatiotemporal distribution and growth potential of Trichodesmium for the last glacial maximum (LGM), the present (2006-2015) and the end of this century (2100) by mapping our model of Trichodesmium growth onto inferred global surface ocean fields of pCO2 , temperature, light and Fe. We conclude that growth rate was severely limited by low pCO2 at the LGM, that current pCO2 levels do not significantly limit Trichodesmium growth and thus, the potential for enhanced growth from future increases in CO2 is small. We also found that the area of the ocean where sea surface temperatures (SST) are within Trichodesmium's thermal niche increased by 32% from the LGM to present, but further increases in SST due to continued global warming will reduce this area by 9%. However, the range reduction at the equator is likely to be offset by enhanced growth associated with expansion of regions with optimal or near optimal Fe and light availability. Between now and 2100, the ocean area of optimal SST and irradiance is projected to increase by 7%, and the ocean area of optimal SST, irradiance and iron is projected to increase by 173%. Given the major contribution of this keystone species to annual N2 fixation and thus pelagic ecology, biogeochemistry and CO2 sequestration, the projected increase in the geographical range for optimal growth could provide a negative feedback to increasing atmospheric CO2 concentrations.

Isolation of Isotrichophycin C and Trichophycins G-I from a Collection of Trichodesmium thiebautii.

The trichophycin family of compounds are chlorinated polyketides first discovered from environmental collections of a bloom-forming Trichodesmium sp. cyanobacterium. In an effort to fully capture the chemical space of this group of metabolites, the utilization of MS/MS-based molecular networking of a Trichodesmium thiebautii extract revealed a metabolome replete with halogenated compounds. Subsequent MS-guided isolation resulted in the characterization of isotrichophycin C and trichophycins G-I (1-4). These new metabolites had intriguing structural variations from those trichophycins previously characterized, which allowed for a comparative study to examine structural features that are associated with toxicity to murine neuroblastoma cells. Additionally, we propose the absolute configuration of the previously characterized trichophycin A (5). Overall, the metabolome of the Trichodesmium bloom is hallmarked by an unprecedented amount of chlorinated molecules, many of which remain to be structurally characterized.