Novel insights into the rhizosphere and seawater microbiome of Zostera marina in diverse mariculture zones.

BACKGROUND: Seagrasses offer various ecosystem services and possess high levels of primary productivity. However, the development of mariculture has affected the homeostasis of seagrass meadow ecosystems. Plant-microbiome associations are essential for seagrasses health, but little is known about the role of environmental microbiomes and how they affect seagrass in a mariculture environment. In this study, we investigated the influence of mariculture on the rhizosphere and seawater microbiome surrounding Zostera marina and focused on the bacterial, eukaryotic, and fungal components in the composition, diversity, metabolism, and responses to mariculture-related environmental factors. RESULTS: Significant differences in the composition, richness, diversity, and internal relations of the bacterial community between the seawater and rhizosphere sediment surrounding Z. marina were observed, while differences in the eukaryotic and fungal communities were less significant. More complex bacterial and fungal co-occurrence networks were found in the seawater and rhizosphere sediment of the Saccharina japonica (SJ) and sea cucumber (SC) culture zones. The seawater in the SJ zone had higher levels of dissimilatory and assimilatory nitrate reduction, denitrification, and nitrogen fixation processes than the other three zones. The assimilatory sulfate reduction enzymes were higher in the rhizosphere sediments of the SJ zone than in the other three zones. Tetracycline, sulfonamide, and diaminopyrimidine resistance genes were enriched in the mariculture SJ and SC zones. CONCLUSIONS: Our findings might contribute to a better understanding of the effects of mariculture on the seagrass and the meadow ecosystems and thus revealing their potential operating mechanisms. These insights may serve to raise awareness of the effects of human activities on natural ecosystems, regulation of antibiotic usage, and environmental restoration. Video Abstract.

Adaptive responses of eelgrass (Zostera marina L.) to ocean warming and acidification.

Ocean warming (OW) and ocean acidification (OA), driven by rapid global warming accelerating at unprecedented rates, are profoundly impacting the stability of seagrass ecosystems. Yet, our current understanding of the effects of OW and OA on seagrass remains constrained. Herein, we investigated the response of eelgrass (Zostera marina L.), a representative seagrass species, to OW and OA through comprehensive transcriptomic and metabolomic analyses. The results showed notable variations in plant performance under varying conditions: OW, OA, and OWA (a combination of both conditions). Specifically, under average oceanic temperature conditions for eelgrass growth over the past 20 years -from May to November-OA promoted the production of differentially expressed genes and metabolites associated with alanine, aspartate, and glutamate metabolism, as well as starch and sucrose metabolism. Under warming condition, eelgrass was resistant to OA by accelerating galactose metabolism, along with glycine, serine, and threonine metabolism, as well as the tricarboxylic acid (TCA) cycle. Under the combined OW and OA condition, eelgrass stimulated fructose and mannose metabolism, glycolysis, and carbon fixation, in addition to galactose metabolism and the TCA cycle to face the interplay. Our findings suggest that eelgrass exhibits adaptive capacity by inducing different metabolites and associated genes, primarily connected with carbon and nitrogen metabolism, in response to varying degrees of OW and OA. The data generated here support the exploration of mechanisms underlying seagrass responses to environmental fluctuations, which hold critical significance for the future conservation and management of these ecosystems.

Invasive macroalgae in native seagrass beds: vectors of spread and impacts.

BACKGROUND AND AIMS: Worldwide, invasive species are spreading through marine systems at an unprecedented rate with both positive and negative consequences for ecosystems and the biological functioning of organisms. Human activities from shipping to habitat damage and modification are known vectors of spread, although biological interactions including epibiosis are increasingly recognized as potentially important to introduction into susceptible habitats. METHODS: We assessed a novel mechanism of spread - limpets as transporters of an invasive alga, Sargassum muticum, into beds of the seagrass Zostera marina - and the physiological impact of its invasion. The association of S. muticum with three limpet species and other habitats was assessed using intertidal surveys on rocky shores and snorkelling at two seagrass sites in the UK. A 4-year field study tested the effect of S. muticum on Z. marina shoot density, dry weight and phenolic compounds (caffeic and tannic acid) content, and a laboratory experiment tested the impact of S. muticum on nutrient partitioning (C/H/N/P/Si), photosynthetic efficiency (Fv/Fm) and growth of Z. marina. RESULTS: On rocky shores 15 % of S. muticum occurrences were attached to the shells of live limpets. In seagrass beds 5 % of S. muticum occurrences were attached to the shells of dead limpets. The remainder were attached to rock, to cobblestones, to the seagrass matrix or embedded within the sand. Z. marina density and phenolics content was lower when S. muticum co-occurred with it. Over 3 years, photosynthetic responses of Z. marina to S. muticum were idiosyncratic, and S. muticum had no effect on nutrient partitioning in Z. marina. CONCLUSIONS: Our results show limpets support S. muticum as an epibiont and may act as a previously unreported transport mechanism introducing invaders into sensitive habitats. S. muticum reduced production of phenolics in Z. marina, which may weaken its defensive capabilities and facilitate proliferation of S. muticum. The effect of S. muticum on Z. marina photosynthesis requires further work but having no effect on the capacity of Z. marina to sequester nutrients suggests a degree of resilience to this invader.

Heterozostera nigricaulis from the south-East Pacific coast of Chile: First insights into its physiology and growth.

A short stretch (27°S and 30°S) along the coast of Chile is habitat for the seagrass Heterozostera nigricaulis. The seagrass is classified as endangered and grows only clonally, but there are no data on its physiology and growth. However, this information is important to gain insights into its acclimation potential and how disturbances may affect them. We therefore studied H. nigricaulis at 27° and 30°S, and determined their growth and physiology among seasons and depths over one year. Biomass was higher at 27° than at 30°S, and was always higher in summer than in autumn and winter. Increased photosynthesis supported growth in summer, and in winter carbonic anhydrase activity was in place to maintain these evergreen meadows. Our results suggest that these seagrass meadows are adapted to local conditions, which, together with their asexual reproduction, could make them more vulnerable to disturbance. Therefore, our results serve as a basis for future studies on seagrass growth dynamics, and are important for protection and management plans.

Organic matter composition regulates residual potential of organic carbon of the seagrass Zostera marina L. during its decomposition process in seawater.

Although the seagrass Zostera marina L. (Z. marina) is expected to significantly contribute to environmental carbon dynamics, the residual potential of organic carbon (Corg) in this plant during its decomposition process in seawater remains insufficiently understood. In this study, the factors regulating this potential were examined by evaluating Z. marina decomposition in seawater and conducting a multiple regression analysis on data from the experiment as well as the original plant organic matter composition. The residual ratio during the decomposition experiment (RDEC) of carbon in Z. marina for 111 days ranged from 0.30 to 0.84 (n = 16), and the variation resulted from differences in the sampling season of this plant. Regression analysis was conducted to explain the RDEC of carbon using the RDEC of nitrogen, concentrations of carbon, nitrogen, HCl-extractable Corg and nitrogen, contributions of carbohydrate (CCAR) of total and non-structural fractions and lipid (CLIP) to carbon concentration. The RDEC of carbon was most adequately fitted using a multiple regression including three parameters, carbon concentration, CLIP, and CCAR of total carbohydrate with a significant determination coefficient. The former two parameters (carbon concentration and CLIP) have negative coefficients, indicating that these parameters correspond to bacterially available Corg in Z. marina in the regression analysis. The latter parameter (CCAR of total carbohydrate) shows a positive coefficient that indicates recalcitrant Corg in the regression. Parameters related to nitrogen were not included in the regression formula, although this element is reportedly an important parameter regulating the environmental decomposition rate of Corg. Finally, we suggested that total carbon, carbohydrate, and lipid are especially important factors to regulate the environmental residual potential of Z. marina Corg among organic components of this plant.

The combined effect of planting density and sediment fertilization on survival, growth and physiology of eelgrass Zostera marina.

We subjected shoots of eelgrass Zostera marina to different combinations of planting density [300 (control), 450, 600, 750, 900 shoots m-2] and sediment fertilization [0 (control), 35, 70, 105, 140 g m-2] for 6 weeks under controlled conditions. We measured eelgrass responses in terms of survivorship, growth, productivity, leaf pigmentation and carbohydrate concentrations. The ramet frequency of Z. marina reached 47.8 % when exposed to the combination of 600 shoots m-2 and 70 g m-2, which was 3.2 times higher than that of shoots under the control. Survival analysis combined with growth assessment suggested that the optimal ranges of planting density and sediment fertilization for the propagation of Z. marina shoots were 501 to 530 shoots m-2 and 51 to 60 g m-2, respectively. The promotion of survival and propagation of Z. marina that stemmed from planting density and sediment fertilization mainly depended on the increase of chlorophyll content and accumulation of non-structural carbohydrate. The total chlorophyll content of Z. marina leaves exposed to the combination of 600 shoots m-2 and 70 g m-2 was 2.1 times higher than that of shoots under the control. The results will provide data that could prove helpful in the development of efficient artificial propagation technology for Z. marina shoots.

Effects of depth and overgrowth of ephemeral macroalgae on a remote subtidal NE Atlantic eelgrass (Zostera marina) community.

We conducted a short-term field sampling complemented with time integrating stable isotope analysis to holistically investigate status and ecological interactions in a remote NE Atlantic Zostera marina meadow. We found high nutrient water concentrations, large biomass of fast-growing, ephemeral macroalgae, low abundance, and biodiversity of epifauna and a food web with thornback ray (Raja clavata) as intermediate and cod (Gadus morhua) as top predator. We observed no variation with increasing depth (3.5-11 m) except for decreasing shoot density and biomass of Zostera and macroalgae. Our results indicate that the Finnøya Zostera ecosystem is eutrophicated. During the past three to four decades, nutrients from aquaculture have steadily increased to reach 75% of anthropogenic input while the coastal top predator cod has decreased by 50%. We conclude that bottom-up regulation is a predominant driver of change since top-down regulation is generally weak in low density and exposed Zostera ecosystems such as Finnøya.

Mimicry of emergent traits amplifies coastal restoration success.

Restoration is becoming a vital tool to counteract coastal ecosystem degradation. Modifying transplant designs of habitat-forming organisms from dispersed to clumped can amplify coastal restoration yields as it generates self-facilitation from emergent traits, i.e. traits not expressed by individuals or small clones, but that emerge in clumped individuals or large clones. Here, we advance restoration science by mimicking key emergent traits that locally suppress physical stress using biodegradable establishment structures. Experiments across (sub)tropical and temperate seagrass and salt marsh systems demonstrate greatly enhanced yields when individuals are transplanted within structures mimicking emergent traits that suppress waves or sediment mobility. Specifically, belowground mimics of dense root mats most facilitate seagrasses via sediment stabilization, while mimics of aboveground plant structures most facilitate marsh grasses by reducing stem movement. Mimicking key emergent traits may allow upscaling of restoration in many ecosystems that depend on self-facilitation for persistence, by constraining biological material requirements and implementation costs.

Arabinogalactan-proteins of Zostera marina L. contain unique glycan structures and provide insight into adaption processes to saline environments.

Seagrasses evolved from monocotyledonous land plants that returned to the marine habitat. This transition was accomplished by substantial changes in cell wall composition, revealing habitat-driven adaption to the new environment. Whether arabinogalactan-proteins (AGPs), important signalling molecules of land plants, are present in seagrass cell walls is of evolutionary and plant development interest. AGPs of Zostera marina L. were isolated and structurally characterised by analytical and bioinformatics methods as well as by ELISA with different anti-AGP antibodies. Calcium-binding capacity of AGPs was studied by isothermal titration calorimetry (ITC) and microscopy. Bioinformatic searches of the Z. marina proteome identified 9 classical AGPs and a large number of chimeric AGPs. The glycan structures exhibit unique features, including a high degree of branching and an unusually high content of terminating 4-O-methyl-glucuronic acid (4-OMe GlcA) residues. Although the common backbone structure of land plant AGPs is conserved in Z. marina, the terminating residues are distinct with high amounts of uronic acids. These differences likely result from the glycan-active enzymes (glycosyltransferases and methyltransferases) and are essential for calcium-binding properties. The role of this polyanionic surface is discussed with regard to adaption to the marine environment.

Heat-stress induced expression of stress-inducible nucleotide exchange factor Fes1 in seagrass Zostera japonica.

Seagrass meadows are among the four most productive marine natural ecosystems in the world. Zostera japonica (Z. japonica) is the most widely distributed species of seagrass in China. Nucleotide exchange factors (NEFs) promote the release of ADP during heat stress, accelerating the rate-limiting step of Heat shock protein 70 (Hsp70). Although NEFs play an important role in abiotic stress tolerance of plants, NEFs in seagrass have not been studied. In this study, we cloned Fes1 from Z. japonica (ZjFes1) by rapid amplification of the cDNA ends using RACE, and full length ZjFes1 was 1171 bp. It contained an 81 bp 5'-terminal untranslated region (UTR), 109 bp 3'-UTR and 981 bp open reading frame (ORF). The ORF (ZjFes1) was predicted to encode a polypeptide of 326 amino acids with theoretical molecular weight (MW) of 36.10 kDa and pI of 5.22. ZjFes1 shared 89% amino acid identity with Fes1 from Zostera marina (Z. marina). The transcriptional levels of ZjFes1 increased significantly 1 h after heat treatment. ZjFes1 was localized to the cytoplasm. Taken together, we found that ZjFes1 was a stress-inducible gene that may be involved in heat stress response. This study lays the foundation for further studies on the role of ZjFes1 in heat resistance.

Metabolic Profiling Reveals Biochemical Pathways Responsible for Eelgrass Response to Elevated CO2 and Temperature.

As CO2 levels in Earth's atmosphere and oceans steadily rise, varying organismal responses may produce ecological losers and winners. Increased ocean CO2 can enhance seagrass productivity and thermal tolerance, providing some compensation for climate warming. However, the metabolic shifts driving the positive response to elevated CO2 by these important ecosystem engineers remain unknown. We analyzed whole-plant performance and metabolic profiles of two geographically distinct eelgrass (Zostera marina L.) populations in response to CO2 enrichment. In addition to enhancing overall plant size, growth and survival, CO2 enrichment increased the abundance of Calvin Cycle and nitrogen assimilation metabolites while suppressing the abundance of stress-related metabolites. Overall metabolome differences between populations suggest that some eelgrass phenotypes may be better suited than others to cope with an increasingly hot and sour sea. Our results suggest that seagrass populations will respond variably, but overall positively, to increasing CO2 concentrations, generating negative feedbacks to climate change.

Strong leaf surface basification and CO2 limitation of seagrass induced by epiphytic biofilm microenvironments.

Coastal eutrophication is a growing problem worldwide, leading to increased epiphyte overgrowth of seagrass leaves. Yet little is known about how epiphytes affect key biogeochemical conditions and processes in the seagrass phyllosphere. We used electrochemical microsensors to measure microgradients of O2 , pH, and CO2 at the bare and epiphyte-covered leaf surface of seagrass (Zostera marina L.) to determine effects of epiphytes on the leaf chemical microenvironment. Epiphytes result in extreme daily fluctuations in pH, O2 , and inorganic carbon concentrations at the seagrass leaf surface severely hampering the plant's performance. In light, leaf epiphyte biofilms and their diffusive boundary layer lead to strong basification, markedly reducing the CO2 and HCO3- availability at the leaf surface, leading to reduced photosynthetic efficiency as a result of carbon limitation and enhanced photorespiration. With epiphytes, leaf surface pH increased to >10, thereby exceeding final pH levels (~9.62) and CO2 compensation points for active photosynthesis. In darkness, epiphyte biofilms resulted in increased CO2 and hypoxia at the leaf surface. Epiphytes can lead to severe carbon limitation in seagrasses owing to strong phyllosphere basification leading to CO2 depletion and costly, yet limiting, HCO3- utilization, increasing the risk of plant starvation.

The influence of hydrodynamics and ecosystem engineers on eelgrass seed trapping.

Propagule dispersal is an integral part of the life cycle of seagrasses; important for colonising unvegetated areas and increasing their spatial distribution. However, to understand recruitment success, seed dispersal and survival in habitats of different complexity remains to be quantified. We tested the single and synergistic effects of three commonly distributed ecosystem engineers-eelgrass (Zostera marina), oysters (Magellana gigas) and blue mussels (Mytilus edulis)-on trapping of Z. marina seeds in a hydraulic flume under currents. Our results suggest that seed retention increases with habitat complexity and further reveal insights into the underlying mechanisms. In eelgrass canopy, trapping occurred mostly through direct blocking of a seed's pathway, while trapping in bivalve patches was mainly related to altered hydrodynamics in the lee side, i.e. behind each specimen. With increasing flow velocity (24-30 cm s-1 in eelgrass canopy, 18-30 cm s-1 in bivalve patches), modifications of the sediment surface through increased turbulence and erosive processes became more important and resulted in high seed trapping rates. Furthermore, we show that while monospecific patches of seagrass and bivalves had different trapping optima depending on flow velocities, intermixing resulted in consistently high trapping rates throughout the investigated hydrodynamic gradient. Our results highlight the importance of positive interactions among ecosystem engineers for seed retention and patch emergence in eelgrass.

Interaction of short-term copper pollution and ocean acidification in seagrass ecosystems: Toxicity, bioconcentration and dietary transfer.

We aimed to show how the predicted pH decrease in the ocean would alter the toxicity, bioconcentration and dietary transfer of trace metal copper on seagrass ecosystems, on a short-term basis. Seagrass Zostera noltei was exposed to two pH levels (8.36 and 8.03) and three copper levels (nominal concentrations, <3, 30 and 300 µg Cu L-1) in a factorial design during 21 days, while Gammarus locusta amphipods were continuously fed with the treated seagrass leaves. We found that the toxicity and bioconcentration of copper in seagrasses were not affected by pH, yet complex copper-pH interactions were observed in the seagrass photosynthesis. We demostrated that seagrasses can act as a copper source in the food web via direct consumption by herbivores. Future research need to investigate the interactive effects on a long-term basis, and to include biochemical and molecular endpoints to provide additional insights to the complex phisiological interactions observed.

Secondary dispersal of seagrass seeds in complex microtopographies.

Motivated by observational and experimental evidence, a theoretical model is proposed to relate the secondary dispersal of seagrass seeds with the complexity of microtopography in natural environments. Complexity is encoded in terms of the Hurst exponent of a fractal description of the microtopographical geometry. The percentage of a seafloor transect where secondary dispersal of seagrass seeds occurs, is quantified in terms of the mainstream velocity, bottom complexity and properties of the seeds. Theoretical expressions are validated considering the cases of Zostera marina and Posidonia oceanica seeds and using computational fluid dynamics (CFD). A total of 200 CFD simulations with different bottom complexities and flow conditions, were done for each seagrass genus to validate the theoretical model. Numerical results agree with theoretical predictions. This finding provides a management tool to assess the degree of seed retention in seed-based restoration areas.

Environmental heterogeneity mediates scale-dependent declines in kelp diversity on intertidal rocky shores.

Biodiversity loss is driven by interacting factors operating at different spatial scales. Yet, there remains uncertainty as to how fine-scale environmental conditions mediate biological responses to broad-scale stressors. We surveyed intertidal rocky shore kelp beds situated across a local gradient of wave action and evaluated changes in kelp diversity and abundance after more than two decades of broad scale stressors, most notably the 2013-2016 heat wave. Across all sites, species were less abundant on average in 2017 and 2018 than during 1993-1995 but changes in kelp diversity were dependent on wave exposure, with wave exposed habitats remaining stable and wave sheltered habitats experiencing near complete losses of kelp diversity. In this way, wave exposed sites have acted as refugia, maintaining regional kelp diversity despite widespread local declines. Fucoids, seagrasses and two stress-tolerant kelp species (Saccharina sessilis, Egregia menziesii) did not decline as observed in other kelps, and the invasive species Sargassum muticum increased significantly at wave sheltered sites. Long-term monitoring data from a centrally-located moderate site suggest that kelp communities were negatively impacted by the recent heatwave which may have driven observed losses throughout the region. Wave-sheltered shores, which saw the largest declines, are a very common habitat type in the Northeast Pacific and may be especially sensitive to losses in kelp diversity and abundance, with potential consequences for coastal productivity. Our findings highlight the importance of fine-scale environmental heterogeneity in mediating biological responses and demonstrate how incorporating differences between habitat patches can be essential to capturing scale-dependent biodiversity loss across the landscape.

Towards a sampling design for characterizing habitat-specific benthic biodiversity related to oxygen flux dynamics using Aquatic Eddy Covariance.

The Aquatic Eddy Covariance (AEC) technique has emerged as an important method to quantify in situ seafloor metabolism over large areas of heterogeneous benthic communities, enabling cross-habitat comparisons of seafloor productivity. However, the lack of a corresponding sampling protocol to perform biodiversity comparisons across habitats is impeding a full assessment of marine ecosystem metabolism. Here, we study a range of coastal benthic habitats, from rocky-bed communities defined by either perennial macroalgae or blue mussel beds to soft-sediment communities comprised of either seagrass, patches of different macrophyte species or bare sand. We estimated that the maximum contribution to the AEC metabolic flux can be found for a seafloor area of approximately 80 m2 with a 5 meter upstream distance of the instrument across all the habitats. We conducted a sampling approach to characterize and quantify the dominant features of biodiversity (i.e., community biomass) within the main seafloor area of maximum metabolic contribution (i.e., gross primary production and community respiration) measured by the AEC. We documented a high biomass contribution of the macroalgal Fucus vesiculosus, the seagrass Zostera marina and the macroinvertebrate Mytilus edulis to the net ecosystem metabolism of the habitats. We also documented a significant role of the bare sediments for primary productivity compared to vegetated canopies of the soft sediments. The AEC also provided insight into dynamic short-term drivers of productivity such as PAR availability and water flow velocity for the productivity estimate. We regard this study as an important step forward, setting a framework for upcoming research focusing on linking biodiversity metrics and AEC flux measurements across habitats.

Surface chemical defence of the eelgrass Zostera marina against microbial foulers.

Plants rely on both mechanical and chemical defence mechanisms to protect their surfaces against microorganisms. The recently completed genome of the eelgrass Zostera marina, a marine angiosperm with fundamental importance for coastal ecosystems, showed that its re-adaptation from land to the sea has led to the loss of essential genes (for chemical communication and defence) and structural features (stomata and thick cuticle) that are typical of terrestrial plants. This study was designed to understand the molecular nature of surface protection and fouling-control strategy of eelgrass against marine epiphytic yeasts. Different surface extraction methods and comparative metabolomics by tandem mass spectrometry (LC-MS/MS) were used for targeted and untargeted identification of the metabolite profiles of the leaf surface and the whole tissue extracts. Desorption electrospray ionization-imaging mass spectrometry (DESI-IMS) coupled with traditional bioassays revealed, for the first time, the unique spatial distribution of the eelgrass surface-associated phenolics and fatty acids, as well as their differential bioactivity against the growth and settlement of epiphytic yeasts. This study provides insights into the complex chemical defence system of the eelgrass leaf surface. It suggests that surface-associated metabolites modulate biotic interactions and provide chemical defence and structural protection to eelgrass in its marine environment.

Physiological responses of eelgrass (Zostera marina) to ambient stresses such as herbicide, insufficient light, and high water temperature.

This study aimed to elucidate the biological responses of eelgrass (Zostera marina) to artificially induced stresses such as herbicide (Irgarol 1051, Irg) exposure, insufficient light, and high water temperature (27 ± 1.0 °C) by evaluating growth inhibition, photosynthetic activity, and metabolomic profiles. After 14 days, all treatments inhibited growth, but photosynthetic activity was only reduced in the Irg-exposed group. In the Irg-exposed and insufficient light groups, the metabolomic profiles were characterized by decreased levels of sugar (sucrose) and increased levels of amino acids such as glutamine, glycine, and leucine. Biochemical and ultrastructural analyses revealed that the loss of sugar-derived metabolic energy was compensated for by energy generated during autophagic protein degradation. Furthermore, the level of myo-inositol, which has various biological roles and participates in several cellular processes such as cell wall synthesis, stress response, and mineral nutrient storage, was markedly increased in the Irg-exposed and insufficient light groups. A combination of metabolomic analysis with other analyses such as measurement of photosynthetic activity might further elucidate the response of eelgrass to ambient stresses in the natural environment.

Oxygen loss from seagrass roots coincides with colonisation of sulphide-oxidising cable bacteria and reduces sulphide stress.

Seagrasses thrive in anoxic sediments where sulphide can accumulate to phytotoxic levels. So how do seagrasses persist in this environment? Here, we propose that radial oxygen loss (ROL) from actively growing root tips protects seagrasses from sulphide intrusion not only by abiotically oxidising sulphides in the rhizosphere of young roots, but also by influencing the abundance and spatial distribution of sulphate-reducing and sulphide-oxidising bacteria. We used a novel multifaceted approach combining imaging techniques (confocal fluorescence in situ hybridisation, oxygen planar optodes, and sulphide diffusive gradients in thin films) with microbial community profiling to build a complete picture of the microenvironment of growing roots of the seagrasses Halophila ovalis and Zostera muelleri. ROL was restricted to young root tips, indicating that seagrasses will have limited ability to influence sulphide oxidation in bulk sediments. On the microscale, however, ROL corresponded with decreased abundance of potential sulphate-reducing bacteria and decreased sulphide concentrations in the rhizosphere surrounding young roots. Furthermore, roots leaking oxygen had a higher abundance of sulphide-oxidising cable bacteria; which is the first direct observation of these bacteria on seagrass roots. Thus, ROL may enhance both abiotic and bacterial sulphide oxidation and restrict bacterial sulphide production around vulnerable roots, thereby helping seagrasses to colonise sulphide-rich anoxic sediments.