Dead molluscan shells from multiple trophic groups as archives of nitrogen isotopic evidence of wastewater gradients in estuaries.

Increased coastal urbanization worldwide has resulted in increased nitrogen inputs to ecosystems, leading to eutrophication and other negative effects. We assessed δ15N in the dead-collected shells of three molluscan species in two estuaries in order to evaluate their ability to identify known gradients in wastewater nitrogen input, namely from private septic systems feeding directly into Waquoit Bay and from a groundwater plume reflecting wastewater injection at a municipal treatment plant in West Falmouth Harbor, Massachusetts, USA. Shells of a suspension-feeder (Geukensia demissa), a micro-algal grazer (Littorina littorea), and an omnivore (Nassarius obsoletus) were collected from lower intertidal sediments near the taxon's life habitat. We found that δ15N in dead-collected shells decreased significantly along the gradients of wastewater pollution in both estuaries, with similar results across all three trophic groups. These positive results demonstrate the potential of dead-shell assemblages for detecting spatial gradients of wastewater pollution.

A partner-switching system controls activation of mixed-linkage ß-glucan synthesis by c-di-GMP in Sinorhizobium meliloti.

Sinorhizobium meliloti synthesizes a linear mixed-linkage (1 → 3)(1 → 4)-ß-d-glucan (ML ß-glucan, MLG) in response to high levels of cyclic diguanylate (c-di-GMP). Two proteins BgsA and BgsB are required for MLG synthesis, BgsA being the glucan synthase which is activated upon c-di-GMP binding to its C-terminal domain. Here we report that the product of bgrR (SMb20447) is a diguanylate cyclase (DGC) that provides c-di-GMP for the synthesis of MLG by BgsA. bgrR is the first gene of a hexacistronic bgrRSTUWV operon, likely encoding a partner-switching regulatory network where BgrR is the final target. Using different approaches, we have determined that the products of genes bgrU (containing a putative PP2C serine phosphatase domain) and bgrW (with predicted kinase effector domain), modulate the phosphorylation status and the activity of the STAS domain protein BgrV. We propose that unphosphorylated BgrV inhibits BgrR DGC activity, perhaps through direct protein-protein interactions as established for other partner switchers. A bgrRSTUWV operon coexists with MLG structural bgsBA genes in many rhizobial genomes but is also present in some MLG non-producers, suggesting a role of this partner-switching system in other processes besides MLG biosynthesis.

Stable isotopic evidence of nitrogen sources and C4 metabolism driving the world's largest macroalgal green tides in the Yellow Sea.

During recent years, rapid seasonal growth of macroalgae covered extensive areas within the Yellow Sea, developing the world's most spatially extensive "green tide". The remarkably fast accumulation of macroalgal biomass is the joint result of high nitrogen supplies in Yellow Sea waters, plus ability of the macroalgae to optionally use C4 photosynthetic pathways that facilitate rapid growth. Stable isotopic evidence shows that the high nitrogen supply is derived from anthropogenic sources, conveyed from watersheds via river discharges, and by direct atmospheric deposition. Wastewater and manures supply about half the nitrogen used by the macroalgae, fertiliser and atmospheric deposition each furnish about a quarter of the nitrogen in macroalgae. The massive green tides affecting the Yellow Sea are likely to increase, with significant current and future environmental and human consequences. Addressing these changing trajectories will demand concerted investment in new basic and applied research as the basis for developing management policies.

Transient coastal landscapes: Rising sea level threatens salt marshes.

Salt marshes are important coastal environments that provide key ecological services. As sea level rise has accelerated globally, concerns about the ability of salt marshes to survive submergence are increasing. Previous estimates of likely survival of salt marshes were based on ratios of sea level rise to marsh platform accretion. Here we took advantage of an unusual, long-term (1979-2015), spatially detailed comparison of changes in a representative New England salt marsh to provide an empirical estimate of habitat losses based on actual measurements. We show prominent changes in habitat mosaic within the marsh, consistent and coincident with increased submergence and coastal erosion. Model results suggest that at current rates of sea level rise, marsh platform accretion, habitat loss, and with the limitation of the widespread "coastal squeeze", the entire ecosystem might disappear by the beginning of the next century, a fate that might be likely for many salt marshes elsewhere. Meta-analysis of available data suggests that 40 to 95% of the world's salt marshes will be submerged, depending on whether sea level rise remains at current or reaches anticipated rates for the end of this century.

Long-term nutrient addition increases respiration and nitrous oxide emissions in a New England salt marsh.

Salt marshes may act either as greenhouse gas (GHG) sources or sinks depending on hydrological conditions, vegetation communities, and nutrient availability. In recent decades, eutrophication has emerged as a major driver of change in salt marsh ecosystems. An ongoing fertilization experiment at the Great Sippewissett Marsh (Cape Cod, USA) allows for observation of the results of over four decades of nutrient addition. Here, nutrient enrichment stimulated changes to vegetation communities that, over time, have resulted in increased elevation of the marsh platform. In this study, we measured fluxes of carbon dioxide (CO 2), methane (CH 4) and nitrous oxide (N2O) in dominant vegetation zones along elevation gradients of chronically fertilized (1,572 kg N ha-1 year-1) and unfertilized (12 kg N ha-1 year-1) experimental plots at Great Sippewissett Marsh. Flux measurements were performed using darkened chambers to focus on community respiration and excluded photosynthetic CO 2 uptake. We hypothesized that N-replete conditions in fertilized plots would result in larger N2O emissions relative to control plots and that higher elevations caused by nutrient enrichment would support increased CO 2 and N2O and decreased CH 4 emissions due to the potential for more oxygen diffusion into sediment. Patterns of GHG emission supported our hypotheses. Fertilized plots were substantially larger sources of N2O and had higher community respiration rates relative to control plots, due to large emissions of these GHGs at higher elevations. While CH 4 emissions displayed a negative relationship with elevation, they were generally small across elevation gradients and nutrient enrichment treatments. Our results demonstrate that at decadal scales, vegetation community shifts and associated elevation changes driven by chronic eutrophication affect GHG emission from salt marshes. Results demonstrate the necessity of long-term fertilization experiments to understand impacts of eutrophication on ecosystem function and have implications for how chronic eutrophication may impact the role that salt marshes play in sequestering C and N.

Tropical land-sea couplings: Role of watershed deforestation, mangrove estuary processing, and marine inputs on N fluxes in coastal Pacific Panama.

We review data from coastal Pacific Panama and other tropical coasts with two aims. First, we defined inputs and losses of nitrogen (N) mediating connectivity of watersheds, mangrove estuaries, and coastal sea. N entering watersheds-mainly via N fixation (79-86%)-was largely intercepted; N discharges to mangrove estuaries (3-6%), small compared to N inputs to watersheds, nonetheless significantly supplied N to mangrove estuaries. Inputs to mangrove estuaries (including watershed discharges, and marine inputs during flood tides) were matched by losses (mainly denitrification and export during ebb tides). Mangrove estuary subsidies of coastal marine food webs take place by export of forms of N [DON (62.5%), PN (9.1%), and litter N (12.9%)] that provide dissimilative and assimilative subsidies. N fixation, denitrification, and tidal exchanges were major processes, and DON was major form of N involved in connecting fluxes in and out of mangrove estuaries. Second, we assessed effects of watershed forest cover on connectivity. Decreased watershed forest cover lowered N inputs, interception, and discharge into receiving mangrove estuaries. These imprints of forest cover were erased during transit of N through estuaries, owing to internal N cycle transformations, and differences in relative area of watersheds and estuaries. Largest losses of N consisted of water transport of energy-rich compounds, particularly DON. N losses were similar in magnitude to N inputs from sea, calculated without considering contribution by intermittent coastal upwelling, and hence likely under-estimated. Pacific Panama mangrove estuaries are exposed to major inputs of N from land and sea, which emphasizes the high degree of bi-directional connectivity in these coupled ecosystems. Pacific Panama is still lightly affected by human or global changes. Increased deforestation can be expected, as well as changes in ENSO, which will surely raise watershed-derived loads of N, as well as significantly change marine N inputs affecting coastal coupled ecosystems.

Eutrophication of Cape Cod estuaries: Effect of decadal changes in global-driven atmospheric and local-scale wastewater nutrient loads.

Nitrogen (N) supply by atmospheric deposition, wastewater, and fertilizers controls estuarine eutrophication. In New England, atmospheric N loads recently decreased by 50% and land-derived contributions rose about 80%, owing to national-scale emission controls and local urban development. The decrease in atmospheric deposition was large enough to balance increases in land-derived N loads, so total N loads to Waquoit Bay estuaries in Cape Cod did not change significantly between 1990 and 2014. Unchanged N regimes were corroborated by finding no differences in estuarine nutrient concentrations and macrophyte biomass between pre-2005 and in 2015. Coastal zones, subject to reasonably rapid changes in global and local driver variables, will require that assessment and management of eutrophication include adaptive strategies that capture effects of changing baselines. Management initiatives will be constrained by spatial scale of driver variables: local efforts may address wastewater and fertilizer N sources, but atmospheric sources require national or international attention.

Assessment of marine ecosystem services indicators: Experiences and lessons learned from 14 European case studies.

This article shares the experiences, observations, and discussions that occurred during the completing of an ecosystem services (ES) indicator framework to be used at European Union (EU) and Member States' level. The experience base was drawn from 3 European research projects and 14 associated case study sites that include 13 transitional-water bodies (specifically 8 coastal lagoons, 4 riverine estuaries, and 1 fjord) and 1 coastal-water ecosystem. The ES pertinent to each case study site were identified along with indicators of these ES and data sources that could be used for mapping. During the process, several questions and uncertainties arose, followed by discussion, leading to these main lessons learned: 1) ES identification: Some ES that do not seem important at the European scale emerge as relevant at regional or local scales; 2) ES indicators: When direct indicators are not available, proxies for indicators (indirect indicators) might be used, including combined data on monitoring requirements imposed by EU legislation and international agreements; 3) ES mapping: Boundaries and appropriate data spatial resolution must be established because ES can be mapped at different temporal and spatial scales. We also acknowledge that mapping and assessment of ES supports the dialogue between human well-being and ecological status. From an evidence-based marine planning-process point of view, mapping and assessment of marine ES are of paramount importance to sustainable use of marine natural capital and to halt the loss of marine biodiversity. Integr Environ Assess Manag 2016;12:726-734. © 2016 SETAC.

An integrated Pan-European perspective on coastal Lagoons management through a mosaic-DPSIR approach.

A decision support framework for the management of lagoon ecosystems was tested using four European Lagoons: Ria de Aveiro (Portugal), Mar Menor (Spain), Tyligulskyi Liman (Ukraine) and Vistula Lagoon (Poland/Russia). Our aim was to formulate integrated management recommendations for European lagoons. To achieve this we followed a DPSIR (Drivers-Pressures-State Change-Impacts-Responses) approach, with focus on integrating aspects of human wellbeing, welfare and ecosystem sustainability. The most important drivers in each lagoon were identified, based on information gathered from the lagoons' stakeholders, complemented by scientific knowledge on each lagoon as seen from a land-sea perspective. The DPSIR cycles for each driver were combined into a mosaic-DPSIR conceptual model to examine the interdependency between the multiple and interacting uses of the lagoon. This framework emphasizes the common links, but also the specificities of responses to drivers and the ecosystem services provided. The information collected was used to formulate recommendations for the sustainable management of lagoons within a Pan-European context. Several common management recommendations were proposed, but specificities were also identified. The study synthesizes the present conditions for the management of lagoons, thus analysing and examining the activities that might be developed in different scenarios, scenarios which facilitate ecosystem protection without compromising future generations.

Linking Seed Photosynthesis and Evolution of the Australian and Mediterranean Seagrass Genus Posidonia.

Recent findings have shown that photosynthesis in the skin of the seed of Posidonia oceanica enhances seedling growth. The seagrass genus Posidonia is found only in two distant parts of the world, the Mediterranean Sea and southern Australia. This fact led us to question whether the acquisition of this novel mechanism in the evolution of this seagrass was a pre-adaptation prior to geological isolation of the Mediterranean from Tethys Sea in the Eocene. Photosynthetic activity in seeds of Australian species of Posidonia is still unknown. This study shows oxygen production and respiration rates, and maximum PSII photochemical efficiency (Fv : Fm) in seeds of two Australian Posidonia species (P. australis and P. sinuosa), and compares these with previous results for P. oceanica. Results showed relatively high oxygen production and respiratory rates in all three species but with significant differences among them, suggesting the existence of an adaptive mechanism to compensate for the relatively high oxygen demands of the seeds. In all cases maximal photochemical efficiency of photosystem II rates reached similar values. The existence of photosynthetic activity in the seeds of all three species implicates that it was an ability probably acquired from a common ancestor during the Late Eocene, when this adaptive strategy could have helped Posidonia species to survive in nutrient-poor temperate seas. This study sheds new light on some aspects of the evolution of marine plants and represents an important contribution to global knowledge of the paleogeographic patterns of seagrass distribution.

Novel mixed-linkage ß-glucan activated by c-di-GMP in Sinorhizobium meliloti.

An artificial increase of cyclic diguanylate (c-di-GMP) levels in Sinorhizobium meliloti 8530, a bacterium that does not carry known cellulose synthesis genes, leads to overproduction of a substance that binds the dyes Congo red and calcofluor. Sugar composition and methylation analyses and NMR studies identified this compound as a linear mixed-linkage (1 → 3)(1 → 4)-ß-D-glucan (ML ß-glucan), not previously described in bacteria but resembling ML ß-glucans found in plants and lichens. This unique polymer is hydrolyzed by the specific endoglucanase lichenase, but, unlike lichenan and barley glucan, it generates a disaccharidic → 4)-ß-D-Glcp-(1 → 3)-ß-D-Glcp-(1 → repeating unit. A two-gene operon bgsBA required for production of this ML ß-glucan is conserved among several genera within the order Rhizobiales, where bgsA encodes a glycosyl transferase with domain resemblance and phylogenetic relationship to curdlan synthases and to bacterial cellulose synthases. ML ß-glucan synthesis is subjected to both transcriptional and posttranslational regulation. bgsBA transcription is dependent on the exopolysaccharide/quorum sensing ExpR/SinI regulatory system, and posttranslational regulation seems to involve allosteric activation of the ML ß-glucan synthase BgsA by c-di-GMP binding to its C-terminal domain. To our knowledge, this is the first report on a linear mixed-linkage (1 → 3)(1 → 4)-ß-glucan produced by a bacterium. The S. meliloti ML ß-glucan participates in bacterial aggregation and biofilm formation and is required for efficient attachment to the roots of a host plant, resembling the biological role of cellulose in other bacteria.

The Sinorhizobium fredii HH103 Genome: A Comparative Analysis With S. fredii Strains Differing in Their Symbiotic Behavior With Soybean.

Sinorhizobium fredii HH103 is a fast-growing rhizobial strain infecting a broad range of legumes including both American and Asiatic soybeans. In this work, we present the sequencing and annotation of the HH103 genome (7.25 Mb), consisting of one chromosome and six plasmids and representing the structurally most complex sinorhizobial genome sequenced so far. Comparative genomic analyses of S. fredii HH103 with strains USDA257 and NGR234 showed that the core genome of these three strains contains 4,212 genes (61.7% of the HH103 genes). Synteny plot analysis revealed that the much larger chromosome of USDA257 (6.48 Mb) is colinear to the HH103 (4.3 Mb) and NGR324 chromosomes (3.9 Mb). An additional region of the USDA257 chromosome of about 2 Mb displays similarity to plasmid pSfHH103e. Remarkable differences exist between HH103 and NGR234 concerning nod genes, flavonoid effect on surface polysaccharide production, and quorum-sensing systems. Furthermore a number of protein secretion systems have been found. Two genes coding for putative type III-secreted effectors not previously described in S. fredii, nopI and gunA, have been located on the HH103 genome. These differences could be important to understand the different symbiotic behavior of S. fredii strains HH103, USDA257, and NGR234 with soybean.

The symbiotic biofilm of Sinorhizobium fredii SMH12, necessary for successful colonization and symbiosis of Glycine max cv Osumi, is regulated by Quorum Sensing systems and inducing flavonoids via NodD1.

Bacterial surface components, especially exopolysaccharides, in combination with bacterial Quorum Sensing signals are crucial for the formation of biofilms in most species studied so far. Biofilm formation allows soil bacteria to colonize their surrounding habitat and survive common environmental stresses such as desiccation and nutrient limitation. This mode of life is often essential for survival in bacteria of the genera Mesorhizobium, Sinorhizobium, Bradyrhizobium, and Rhizobium. The role of biofilm formation in symbiosis has been investigated in detail for Sinorhizobium meliloti and Bradyrhizobium japonicum. However, for S. fredii this process has not been studied. In this work we have demonstrated that biofilm formation is crucial for an optimal root colonization and symbiosis between S. fredii SMH12 and Glycine max cv Osumi. In this bacterium, nod-gene inducing flavonoids and the NodD1 protein are required for the transition of the biofilm structure from monolayer to microcolony. Quorum Sensing systems are also required for the full development of both types of biofilms. In fact, both the nodD1 mutant and the lactonase strain (the lactonase enzyme prevents AHL accumulation) are defective in soybean root colonization. The impairment of the lactonase strain in its colonization ability leads to a decrease in the symbiotic parameters. Interestingly, NodD1 together with flavonoids activates certain quorum sensing systems implicit in the development of the symbiotic biofilm. Thus, S. fredii SMH12 by means of a unique key molecule, the flavonoid, efficiently forms biofilm, colonizes the legume roots and activates the synthesis of Nod factors, required for successfully symbiosis.

Genome sequence of the soybean symbiont Sinorhizobium fredii HH103.

Sinorhizobium fredii HH103 is a fast-growing rhizobial strain that is able to nodulate legumes that develop determinate nodules, e.g., soybean, and legumes that form nodules of the indeterminate type. Here we present the genome of HH103, which consists of one chromosome and five plasmids with a total size of 7.22 Mb.

The contribution of benthic macrofauna to the nutrient filter in coastal lagoons.

Human activities in coastal areas have increased the occurrence of eutrophication events, especially in vulnerable ecosystems such as coastal lagoons. Although we have a general knowledge of the consequences of eutrophication in these ecosystems, some efforts need to be made to understand biotic feedbacks that could modify the response of the environment to nutrient enrichment. The plant-mediated 'coastal filter' is one of the main factors that determine lagoonal efficiency in processing excess nutrients. In this context, the present paper examined the relative contribution of benthic macrofauna to the 'coastal filter' of a Mediterranean lagoon. The analysis of macrofaunal assemblages in the Mar Menor lagoon led to a clear differentiation between shallow areas of net nutrient recycling and exportation and deeper areas of net retention. These differences enhance nutrient removal from the water column, thus increasing the ecosystem's resistance to eutrophication.

Symbiotic properties and first analyses of the genomic sequence of the fast growing model strain Sinorhizobium fredii HH103 nodulating soybean.

Glycine max (soybean) plants can be nodulated by fast-growing rhizobial strains of the genus Sinorhizobium as well as by slow-growing strains clustered in the genus Bradyrhizobium. Fast-growing rhizobia strains with different soybean cultivar specificities have been isolated from Chinese soils and from other geographical regions. Most of these strains have been clustered into the species Sinorhizobium fredii. The S. fredii strain HH103 was isolated from soils of Hubei province, Central China and was first described in 1985. This strain is capable to nodulate American and Asiatic soybean cultivars and many other different legumes and is so far the best studied fast-growing soybean-nodulating strain. Additionally to the chromosome S. fredii HH103 carries five indigenous plasmids. The largest plasmid (pSfrHH103e) harbours genes for the production of diverse surface polysaccharides, such as exopolysaccharides (EPS), lipopolysaccharides (LPS), and capsular polysaccharides (KPS). The second largest plasmid (pSfrHH103d) is a typical symbiotic plasmid (pSym), carrying nodulation and nitrogen fixation genes. The present mini review focuses on symbiotic properties of S. fredii HH103, in particular on nodulation and surface polysaccharides aspects. The model strain S. fredii HH103 was chosen for genomic sequencing, which is currently in progress. First analyses of the draft genome sequence revealed an extensive synteny between the chromosomes of S. fredii HH103 and Rhizobium sp. NGR234.

The Sinorhizobium meliloti RNA chaperone Hfq influences central carbon metabolism and the symbiotic interaction with alfalfa.

BACKGROUND: The bacterial Hfq protein is able to interact with diverse RNA molecules, including regulatory small non-coding RNAs (sRNAs), and thus it is recognized as a global post-transcriptional regulator of gene expression. Loss of Hfq has an extensive impact in bacterial physiology which in several animal pathogens influences virulence. Sinorhizobium meliloti is a model soil bacterium known for its ability to establish a beneficial nitrogen-fixing intracellular symbiosis with alfalfa. Despite the predicted general involvement of Hfq in the establishment of successful bacteria-eukaryote interactions, its function in S. meliloti has remained unexplored. RESULTS: Two independent S. meliloti mutants, 2011-3.4 and 1021Deltahfq, were obtained by disruption and deletion of the hfq gene in the wild-type strains 2011 and 1021, respectively, both exhibiting similar growth defects as free-living bacteria. Transcriptomic profiling of 1021Deltahfq revealed a general down-regulation of genes of sugar transporters and some enzymes of the central carbon metabolism, whereas transcripts specifying the uptake and metabolism of nitrogen sources (mainly amino acids) were more abundant than in the wild-type strain. Proteomic analysis of the 2011-3.4 mutant independently confirmed these observations. Symbiotic tests showed that lack of Hfq led to a delayed nodulation, severely compromised bacterial competitiveness on alfalfa roots and impaired normal plant growth. Furthermore, a large proportion of nodules (55%-64%) elicited by the 1021Deltahfq mutant were non-fixing, with scarce content in bacteroids and signs of premature senescence of endosymbiotic bacteria. RT-PCR experiments on RNA from bacteria grown under aerobic and microoxic conditions revealed that Hfq contributes to regulation of nifA and fixK1/K2, the genes controlling nitrogen fixation, although the Hfq-mediated regulation of fixK is only aerobiosis dependent. Finally, we found that some of the recently identified S. meliloti sRNAs co-inmunoprecipitate with a FLAG-epitope tagged Hfq protein. CONCLUSIONS: Our results support that the S. meliloti RNA chaperone Hfq contributes to the control of central metabolic pathways in free-living bacteria and influences rhizospheric competence, survival of the microsymbiont within the nodule cells and nitrogen fixation during the symbiotic interaction with its legume host alfalfa. The identified S. meliloti Hfq-binding sRNAs are predicted to participate in the Hfq regulatory network.

Gene SMb21071 of plasmid pSymB is required for osmoadaptation of Sinorhizobium meliloti 1021 and is implicated in modifications of cell surface polysaccharides structure in response to hyperosmotic stress.

Megaplasmid pSymB of the nitrogen-fixing symbiont Sinorhizobium meliloti, implicated in adaptation to hyperosmotic stress, contains 11 gene clusters that apparently encode surface polysaccharides. However, only 2 of these clusters, containing the exo and exp genes, have been associated with the synthesis of the acidic exopolysaccharides succinoglycan and galactoglucan, respectively. The functions of the other 9 clusters remain unsolved. The involvement of one of those regions, pSymB cluster 3, on surface polysaccharide synthesis and its possible implication in osmoadaptation were investigated. In silico analysis of cluster 3 showed that it putatively encodes for the synthesis and transport of a methylated surface polysaccharide. Mutants affected in this cluster were symbiotically effective but showed defects in growth under saline and nonsaline osmotic stress. The gene SMb21071, encoding a putative initiating glycosyltransferase, is transcriptionally induced under hyperosmotic conditions. Sodium dodecyl sulfate - polyacrylamide gel electrophoresis and silver staining showed that osmotic stresses changed the profiles of surface polysaccharides of wild-type and mutants strains in different ways. The overall results suggest that cluster 3 is important for growth under saline stress and essential for growth under nonsaline hyperosmotic stress, and it appears to be implicated in maintaining and (or) modifying surface polysaccharides in response to osmotic stress.

The role of benthic macrophytes and their associated macroinvertebrate community in coastal lagoon resistance to eutrophication.

Eutrophication is widely recognised as one of the major menaces to coastal environments, particularly enclosed bays and lagoons. Although there is a general understanding of the consequences of eutrophication in these systems, there is a lack of sufficient knowledge concerning biotic feedbacks that influence eutrophication patterns and the resistance capacity of coastal environments. In this paper, the isotope ratios of main producers and consumers of a Mediterranean lagoon were examined in order to elucidate the fate of anthropogenic inputs from the main watercourse flowing into the lagoon. The results of the study of stable isotope data in the Mar Menor lagoon reflected that the whole benthic community plays an important role as a natural 'filter' that removes excess nutrients from the water column and stores them in the sediments, thereby enhancing lagoon resistance to eutrophication.