Seagrass-mediated rhizosphere redox gradients are linked with ammonium accumulation driven by diazotrophs.

Seagrasses can enhance nutrient mobilization in their rhizosphere via complex interactions with sediment redox conditions and microbial populations. Yet, limited knowledge exists on how seagrass-derived rhizosphere dynamics affect nitrogen cycling. Using optode and gel-sampler-based chemical imaging, we show that radial O2 loss (ROL) from rhizomes and roots leads to the formation of redox gradients around below-ground tissues of seagrass (Zostera marina), which are co-localized with regions of high ammonium concentrations in the rhizosphere. Combining such chemical imaging with fine-scale sampling for microbial community and gene expression analyses indicated that multiple biogeochemical pathways and microbial players can lead to high ammonium concentration within the oxidized regions of the seagrass rhizosphere. Symbiotic N2-fixing bacteria (Bradyrhizobium) were particularly abundant and expressed the diazotroph functional marker gene nifH in Z. marina rhizosphere areas with high ammonium concentrations. Such an association between Z. marina and Bradyrhizobium can facilitate ammonium mobilization, the preferred nitrogen source for seagrasses, enhancing seagrass productivity within nitrogen-limited environments. ROL also caused strong gradients of sulfide at anoxic/oxic interfaces in rhizosphere areas, where we found enhanced nifH transcription by sulfate-reducing bacteria. Furthermore, we found a high abundance of methylotrophic and sulfide-oxidizing bacteria in rhizosphere areas, where O2 was released from seagrass rhizomes and roots. These bacteria could play a beneficial role for the plants in terms of their methane and sulfide oxidation, as well as their formation of growth factors and phytohormones. ROL from below-ground tissues of seagrass, thus, seems crucial for ammonium production in the rhizosphere via stimulation of multiple diazotrophic associations. IMPORTANCE: Seagrasses are important marine habitats providing several ecosystem services in coastal waters worldwide, such as enhancing marine biodiversity and mitigating climate change through efficient carbon sequestration. Notably, the fitness of seagrasses is affected by plant-microbe interactions. However, these microscale interactions are challenging to study and large knowledge gaps prevail. Our study shows that redox microgradients in the rhizosphere of seagrass select for a unique microbial community that can enhance the ammonium availability for seagrass. We provide first experimental evidence that Rhizobia, including the symbiotic N2-fixing bacteria Bradyrhizobium, can contribute to the bacterial ammonium production in the seagrass rhizosphere. The release of O2 from rhizomes and roots also caused gradients of sulfide in rhizosphere areas with enhanced nifH transcription by sulfate-reducing bacteria. O2 release from seagrass root systems thus seems crucial for ammonium production in the rhizosphere via stimulation of multiple diazotrophic associations.

New chemical and microbial perspectives on vitamin B1 and vitamer dynamics of a coastal system.

Vitamin B1 (thiamin, B1) is an essential micronutrient for cells, yet intriguingly in aquatic systems most bacterioplankton are unable to synthesize it de novo (auxotrophy), requiring an exogenous source. Cycling of this valuable metabolite in aquatic systems has not been fully investigated and vitamers (B1-related compounds) have only begun to be measured and incorporated into the B1 cycle. Here, we identify potential key producers and consumers of B1 and gain new insights into the dynamics of B1 cycling through measurements of B1 and vitamers (HMP: 4-amino-5-hydroxymethyl-2-methylpyrimidine, HET: 4-methyl-5-thiazoleethanol, FAMP: N-formyl-4-amino-5-aminomethyl-2-methylpyrimidine) in the particulate and dissolved pool in a temperate coastal system. Dissolved B1 was not the primary limiting nutrient for bacterial production and was relatively stable across seasons with concentrations ranging from 74-117 pM, indicating a balance of supply and demand. However, vitamer concentration changed markedly with season as did transcripts related to vitamer salvage and transport suggesting use of vitamers by certain bacterioplankton, e.g. Pelagibacterales. Genomic and transcriptomic analyses showed that up to 78% of the bacterioplankton taxa were B1 auxotrophs. Notably, de novo B1 production was restricted to a few abundant bacterioplankton (e.g. Vulcanococcus, BACL14 (Burkholderiales), Verrucomicrobiales) across seasons. In summer, abundant picocyanobacteria were important putative B1 sources, based on transcriptional activity, leading to an increase in the B1 pool. Our results provide a new dynamic view of the players and processes involved in B1 cycling over time in coastal waters, and identify specific priority populations and processes for future study.

Use and detection of a vitamin B1 degradation product yields new views of the marine B1 cycle and plankton metabolite exchange.

Vitamin B1 (thiamin) is a vital nutrient for most cells in nature, including marine plankton. Early and recent experiments show that B1 degradation products instead of B1 can support the growth of marine bacterioplankton and phytoplankton. However, the use and occurrence of some degradation products remains uninvestigated, namely N-formyl-4-amino-5-aminomethyl-2-methylpyrimidine (FAMP), which has been a focus of plant oxidative stress research. We investigated the relevance of FAMP in the ocean. Experiments and global ocean meta-omic data indicate that eukaryotic phytoplankton, including picoeukaryotes and harmful algal bloom species, use FAMP while bacterioplankton appear more likely to use deformylated FAMP, 4-amino-5-aminomethyl-2-methylpyrimidine. Measurements of FAMP in seawater and biomass revealed that it occurs at picomolar concentrations in the surface ocean, heterotrophic bacterial cultures produce FAMP in the dark-indicating non-photodegradation of B1 by cells, and B1-requiring (auxotrophic) picoeukaryotic phytoplankton produce intracellular FAMP. Our results require an expansion of thinking about vitamin degradation in the sea, but also the marine B1 cycle where it is now crucial to consider a new B1-related compound pool (FAMP), as well as generation (dark degradation-likely via oxidation), turnover (plankton uptake), and exchange of the compound within the networks of plankton. IMPORTANCE Results of this collaborative study newly show that a vitamin B1 degradation product, N-formyl-4-amino-5-aminomethyl-2-methylpyrimidine (FAMP), can be used by diverse marine microbes (bacteria and phytoplankton) to meet their vitamin B1 demands instead of B1 and that FAMP occurs in the surface ocean. FAMP has not yet been accounted for in the ocean and its use likely enables cells to avoid B1 growth deficiency. Additionally, we show FAMP is formed in and out of cells without solar irradiance-a commonly considered route of vitamin degradation in the sea and nature. Altogether, the results expand thinking about oceanic vitamin degradation, but also the marine B1 cycle where it is now crucial to consider a new B1-related compound pool (FAMP), as well as its generation (dark degradation-likely via oxidation), turnover (plankton uptake), and exchange within networks of plankton.

Ammonia-oxidizing archaea release a suite of organic compounds potentially fueling prokaryotic heterotrophy in the ocean.

Ammonia-oxidizing archaea (AOA) constitute a considerable fraction of microbial biomass in the global ocean, comprising 20%-40% of the ocean's prokaryotic plankton. However, it remains enigmatic to what extent these chemolithoautotrophic archaea release dissolved organic carbon (DOC). A combination of targeted and untargeted metabolomics was used to characterize the exometabolomes of three model AOA strains of the Nitrosopumilus genus. Our results indicate that marine AOA exude a suite of organic compounds with potentially varying reactivities, dominated by nitrogen-containing compounds. A significant fraction of the released dissolved organic matter (DOM) consists of labile compounds, which typically limit prokaryotic heterotrophic activity in open ocean waters, including amino acids, thymidine and B vitamins. Amino acid release rates corresponded with ammonia oxidation activity and the three Nitrosopumilus strains predominantly released hydrophobic amino acids, potentially as a result of passive diffusion. Despite the low contribution of DOC released by AOA (~0.08%-1.05%) to the heterotrophic prokaryotic carbon demand, the release of physiologically relevant metabolites could be crucial for microbes that are auxotrophic for some of these compounds, including members of the globally abundant and ubiquitous SAR11 clade.

Proteomic Response of Three Marine Ammonia-Oxidizing Archaea to Hydrogen Peroxide and Their Metabolic Interactions with a Heterotrophic Alphaproteobacterium.

Ammonia-oxidizing archaea (AOA) play an important role in the nitrogen cycle and account for a considerable fraction of the prokaryotic plankton in the ocean. Most AOA lack the hydrogen peroxide (H2O2)-detoxifying enzyme catalase, and some AOA have been shown to grow poorly under conditions of exposure to H2O2 However, differences in the degrees of H2O2 sensitivity of different AOA strains, the physiological status of AOA cells exposed to H2O2, and their molecular response to H2O2 remain poorly characterized. Further, AOA might rely on heterotrophic bacteria to detoxify H2O2, and yet the extent and variety of costs and benefits involved in these interactions remain unclear. Here, we used a proteomics approach to compare the protein profiles of three Nitrosopumilus strains grown in the presence and absence of catalase and in coculture with the heterotrophic alphaproteobacterium Oceanicaulis alexandrii We observed that most proteins detected at a higher relative abundance in H2O2-exposed Nitrosopumilus cells had no known function in oxidative stress defense. Instead, these proteins were putatively involved in the remodeling of the extracellular matrix, which we hypothesize to be a strategy limiting the influx of H2O2 into the cells. Using RNA-stable isotope probing, we confirmed that O. alexandrii cells growing in coculture with the Nitrosopumilus strains assimilated Nitrosopumilus-derived organic carbon, suggesting that AOA could recruit H2O2-detoxifying bacteria through the release of labile organic matter. Our results contribute new insights into the response of AOA to H2O2 and highlight the potential ecological importance of their interactions with heterotrophic free-living bacteria in marine environments.IMPORTANCE Ammonia-oxidizing archaea (AOA) are the most abundant chemolithoautotrophic microorganisms in the oxygenated water column of the global ocean. Although H2O2 appears to be a universal by-product of aerobic metabolism, genes encoding the hydrogen peroxide (H2O2)-detoxifying enzyme catalase are largely absent in genomes of marine AOA. Here, we provide evidence that closely related marine AOA have different degrees of sensitivity to H2O2, which may contribute to niche differentiation between these organisms. Furthermore, our results suggest that marine AOA rely on H2O2 detoxification during periods of high metabolic activity and release organic compounds, thereby potentially attracting heterotrophic prokaryotes that provide this missing function. In summary, this report provides insights into the metabolic interactions between AOA and heterotrophic bacteria in marine environments and suggests that AOA play an important role in the biogeochemical carbon cycle by making organic carbon available for heterotrophic microorganisms.