Diversity at single nucleotide to pangenome scales among sulfur cycling bacteria in salt marshes.

IMPORTANCE: Salt marshes are known for their significant carbon storage capacity, and sulfur cycling is closely linked with the ecosystem-scale carbon cycling in these ecosystems. Sulfate reducers are key for the decomposition of organic matter, and sulfur oxidizers remove toxic sulfide, supporting the productivity of marsh plants. To date, the complexity of coastal environments, heterogeneity of the rhizosphere, high microbial diversity, and uncultured majority hindered our understanding of the genomic diversity of sulfur-cycling microbes in salt marshes. Here, we use comparative genomics to overcome these challenges and provide an in-depth characterization of sulfur-cycling microbial diversity in salt marshes. We characterize communities across distinct sites and plant species and uncover extensive genomic diversity at the taxon level and specific genomic features present in MAGs affiliated with uncultivated sulfur-cycling lineages. Our work provides insights into the partnerships in salt marshes and a roadmap for multiscale analyses of diversity in complex biological systems.

Parasite manipulation of host phenotypes inferred from transcriptional analyses in a trematode-amphipod system.

Manipulation of host phenotypes by parasites is hypothesized to be an adaptive strategy enhancing parasite transmission across hosts and generations. Characterizing the molecular mechanisms of manipulation is important to advance our understanding of host-parasite coevolution. The trematode (Levinseniella byrdi) is known to alter the colour and behaviour of its amphipod host (Orchestia grillus) presumably increasing predation of amphipods which enhances trematode transmission through its life cycle. We sampled 24 infected and 24 uninfected amphipods from a salt marsh in Massachusetts to perform differential gene expression analysis. In addition, we constructed novel genomic tools for O. grillus including a de novo genome and transcriptome. We discovered that trematode infection results in upregulation of amphipod transcripts associated with pigmentation and detection of external stimuli, and downregulation of multiple amphipod transcripts implicated in invertebrate immune responses, such as vacuolar ATPase genes. We hypothesize that suppression of immune genes and the altered expression of genes associated with coloration and behaviour may allow the trematode to persist in the amphipod and engage in further biochemical manipulation that promotes transmission. The genomic tools and transcriptomic analyses reported provide new opportunities to discover how parasites alter diverse pathways underlying host phenotypic changes in natural populations.

Nitrate is an important nitrogen source for Arctic tundra plants.

Plant nitrogen (N) use is a key component of the N cycle in terrestrial ecosystems. The supply of N to plants affects community species composition and ecosystem processes such as photosynthesis and carbon (C) accumulation. However, the availabilities and relative importance of different N forms to plants are not well understood. While nitrate (NO3-) is a major N form used by plants worldwide, it is discounted as a N source for Arctic tundra plants because of extremely low NO3- concentrations in Arctic tundra soils, undetectable soil nitrification, and plant-tissue NO3- that is typically below detection limits. Here we reexamine NO3- use by tundra plants using a sensitive denitrifier method to analyze plant-tissue NO3- Soil-derived NO3- was detected in tundra plant tissues, and tundra plants took up soil NO3- at comparable rates to plants from relatively NO3--rich ecosystems in other biomes. Nitrate assimilation determined by 15N enrichments of leaf NO3- relative to soil NO3- accounted for 4 to 52% (as estimated by a Bayesian isotope-mixing model) of species-specific total leaf N of Alaskan tundra plants. Our finding that in situ soil NO3- availability for tundra plants is high has important implications for Arctic ecosystems, not only in determining species compositions, but also in determining the loss of N from soils via leaching and denitrification. Plant N uptake and soil N losses can strongly influence C uptake and accumulation in tundra soils. Accordingly, this evidence of NO3- availability in tundra soils is crucial for predicting C storage in tundra.

Not All Nitrogen Is Created Equal: Differential Effects of Nitrate and Ammonium Enrichment in Coastal Wetlands.

Excess reactive nitrogen (N) flows from agricultural, suburban, and urban systems to coasts, where it causes eutrophication. Coastal wetlands take up some of this N, thereby ameliorating the impacts on nearshore waters. Although the consequences of N on coastal wetlands have been extensively studied, the effect of the specific form of N is not often considered. Both oxidized N forms (nitrate, NO3-) and reduced forms (ammonium, NH4+) can relieve nutrient limitation and increase primary production. However, unlike NH4+, NO3- can also be used as an electron acceptor for microbial respiration. We present results demonstrating that, in salt marshes, microbes use NO3- to support organic matter decomposition and primary production is less stimulated than when enriched with reduced N. Understanding how different forms of N mediate the balance between primary production and decomposition is essential for managing coastal wetlands as N enrichment and sea level rise continue to assail our coasts.

Nitrate addition stimulates microbial decomposition of organic matter in salt marsh sediments.

Salt marshes sequester carbon at rates more than an order of magnitude greater than their terrestrial counterparts, helping to mitigate climate change. As nitrogen loading to coastal waters continues, primarily in the form of nitrate, it is unclear what effect it will have on carbon storage capacity of these highly productive systems. This uncertainty is largely driven by the dual role nitrate can play in biological processes, where it can serve as a nutrient-stimulating primary production or a thermodynamically favorable electron acceptor fueling heterotrophic metabolism. Here, we used a controlled flow-through reactor experiment to test the role of nitrate as an electron acceptor, and its effect on organic matter decomposition and the associated microbial community in salt marsh sediments. Organic matter decomposition significantly increased in response to nitrate, even at sediment depths typically considered resistant to decomposition. The use of isotope tracers suggests that this pattern was largely driven by stimulated denitrification. Nitrate addition also significantly altered the microbial community and decreased alpha diversity, selecting for taxa belonging to groups known to reduce nitrate and oxidize more complex forms of organic matter. Fourier Transform-Infrared Spectroscopy further supported these results, suggesting that nitrate facilitated decomposition of complex organic matter compounds into more bioavailable forms. Taken together, these results suggest the existence of organic matter pools that only become accessible with nitrate and would otherwise remain stabilized in the sediment. The existence of such pools could have important implications for carbon storage, since greater decomposition rates as N loading increases may result in less overall burial of organic-rich sediment. Given the extent of nitrogen loading along our coastlines, it is imperative that we better understand the resilience of salt marsh systems to nutrient enrichment, especially if we hope to rely on salt marshes, and other blue carbon systems, for long-term carbon storage.

Similar temperature responses suggest future climate warming will not alter partitioning between denitrification and anammox in temperate marine sediments.

Removal of biologically available nitrogen (N) by the microbially mediated processes denitrification and anaerobic ammonium oxidation (anammox) affects ecosystem N availability. Although few studies have examined temperature responses of denitrification and anammox, previous work suggests that denitrification could become more important than anammox in response to climate warming. To test this hypothesis, we determined whether temperature responses of denitrification and anammox differed in shelf and estuarine sediments from coastal Rhode Island over a seasonal cycle. The influence of temperature and organic C availability was further assessed in a 12-week laboratory microcosm experiment. Temperature responses, as characterized by thermal optima (Topt ) and apparent activation energy (Ea ), were determined by measuring potential rates of denitrification and anammox at 31 discrete temperatures ranging from 3 to 59 °C. With a few exceptions, Topt and Ea of denitrification and anammox did not differ in Rhode Island sediments over the seasonal cycle. In microcosm sediments, Ea  was somewhat lower for anammox compared to denitrification across all treatments. However, Topt  did not differ between processes, and neither Ea  nor Topt  changed with warming or carbon addition. Thus, the two processes behaved similarly in terms of temperature responses, and these responses were not influenced by warming. This led us to reject the hypothesis that anammox is more cold-adapted than denitrification in our study system. Overall, our study suggests that temperature responses of both processes can be accurately modeled for temperate regions in the future using a single set of parameters, which are likely not to change over the next century as a result of predicted climate warming. We further conclude that climate warming will not directly alter the partitioning of N flow through anammox and denitrification.

Functional Responses of Salt Marsh Microbial Communities to Long-Term Nutrient Enrichment.

UNLABELLED: Environmental nutrient enrichment from human agricultural and waste runoff could cause changes to microbial communities that allow them to capitalize on newly available resources. Currently, the response of microbial communities to nutrient enrichment remains poorly understood, and, while some studies have shown no clear changes in community composition in response to heavy nutrient loading, others targeting specific genes have demonstrated clear impacts. In this study, we compared functional metagenomic profiles from sediment samples taken along two salt marsh creeks, one of which was exposed for more than 40 years to treated sewage effluent at its head. We identified strong and consistent increases in the relative abundance of microbial genes related to each of the biochemical steps in the denitrification pathway at enriched sites. Despite fine-scale local increases in the abundance of denitrification-related genes, the overall community structures based on broadly defined functional groups and taxonomic annotations were similar and varied with other environmental factors, such as salinity, which were common to both creeks. Homology-based taxonomic assignments of nitrous oxide reductase sequences in our data show that increases are spread over a broad taxonomic range, thus limiting detection from taxonomic data alone. Together, these results illustrate a functionally targeted yet taxonomically broad response of microbial communities to anthropogenic nutrient loading, indicating some resolution to the apparently conflicting results of existing studies on the impacts of nutrient loading in sediment communities. IMPORTANCE: In this study, we used environmental metagenomics to assess the response of microbial communities in estuarine sediments to long-term, nutrient-rich sewage effluent exposure. Unlike previous studies, which have mainly characterized communities based on taxonomic data or primer-based amplification of specific target genes, our whole-genome metagenomics approach allowed an unbiased assessment of the abundance of denitrification-related genes across the entire community. We identified strong and consistent increases in the relative abundance of gene sequences related to denitrification pathways across a broad phylogenetic range at sites exposed to long-term nutrient addition. While further work is needed to determine the consequences of these community responses in regulating environmental nutrient cycles, the increased abundance of bacteria harboring denitrification genes suggests that such processes may be locally upregulated. In addition, our results illustrate how whole-genome metagenomics combined with targeted hypothesis testing can reveal fine-scale responses of microbial communities to environmental disturbance.

Effects of long-term nutrient additions on Arctic tundra, stream, and lake ecosystems: beyond NPP.

Primary producers form the base of food webs but also affect other ecosystem characteristics, such as habitat structure, light availability, and microclimate. Here, we examine changes caused by 5-30+ years of nutrient addition and resulting increases in net primary productivity (NPP) in tundra, streams, and lakes in northern Alaska. The Arctic provides an important opportunity to examine how ecosystems characterized by low diversity and low productivity respond to release from nutrient limitation. We review how responses of algae and plants affect light availability, perennial biotic structures available for consumers, oxygen levels, and temperature. Sometimes, responses were similar across all three ecosystems; e.g., increased NPP significantly reduced light to the substrate following fertilization. Perennial biotic structures increased in tundra and streams but not in lakes, and provided important new habitat niches for consumers as well as other producers. Oxygen and temperature responses also differed. Life history traits (e.g., longevity) of the primary producers along with the fate of detritus drove the responses and recovery. As global change persists and nutrients become more available in the Arctic and elsewhere, incorporating these factors as response variables will enable better prediction of ecosystem changes and feedbacks in this biome and others.

Marsh sediments chronically exposed to nitrogen enrichment contain degraded organic matter that is less vulnerable to decomposition via nitrate reduction.

Blue carbon habitats, including salt marshes, can sequester carbon at rates that are an order of magnitude greater than terrestrial forests. This ecosystem service may be under threat from nitrate (NO3-) enrichment, which can shift the microbial community and stimulate decomposition of organic matter. Despite efforts to mitigate nitrogen loading, salt marshes continue to experience chronic NO3- enrichment, however, the long-term consequence of this enrichment on carbon storage remains unclear. To investigate the effect of chronic NO3- exposure on salt marsh organic matter decomposition, we collected sediments from three sites across a range of prior NO3- exposure: a relatively pristine marsh, a marsh enriched to ~70 µmol L-1 NO3- in the flooding seawater for 13 years, and a marsh enriched between 100 and 1000 µmol L-1 for 40 years from wastewater treatment effluent. We collected sediments from 20 to 25 cm depth and determined that sediments from the most chronically enriched site had less bioavailable organic matter and a distinct assemblage of active microbial taxa compared to the other two sites. We also performed a controlled anaerobic decomposition experiment to test whether the legacy of NO3- exposure influenced the functional response to additional NO3-. We found significant changes to microbial community composition resulting from experimental NO3- addition. Experimental NO3- addition also increased microbial respiration in sediments collected from all sites. However, sediments from the most chronically enriched site exhibited the smallest increase, the lowest rates of total NO3- reduction by dissimilatory nitrate reduction to ammonium (DNRA), and the highest DNF:DNRA ratios. Our results suggest that chronic exposure to elevated NO3- may lead to residual pools of organic matter that are less biologically available for decomposition. Thus, it is important to consider the legacy of nutrient exposure when examining the carbon cycle of salt marsh sediments.

Nitrogen dynamics in arctic tundra soils of varying age: differential responses to fertilization and warming.

In the foothills of the Brooks Range, Alaska, different glaciation histories have created landscapes with varying soil age. Productivity of most of these landscapes is generally N limited, but varies widely, as do plant species composition and soil properties (e.g., pH). We hypothesized that the projected changes in productivity and vegetation composition under a warmer climate might be mediated through differential changes in N availability across soil age. We compared readily available [water-soluble NH4 (+), NO3 (-), and amino acids (AA)], moderately available (soluble proteins), hydrolyzable, and total N pools across three tussock-tundra landscapes with soil ages ranging from 11.5k to 300k years. The effects of fertilization and warming on these N pools were also compared for the two younger sites. Readily available N was highest at the oldest site, and AA accounted for 80-89 % of this N. At the youngest site, inorganic N constituted the majority (80-97 %) of total readily available N. This variation reflected the large differences in plant functional group composition and soil chemical properties. Long-term (8-16 years) fertilization increased the soluble inorganic N by 20- to 100-fold at the intermediate-age site, but only by twofold to threefold at the youngest site. Warming caused small and inconsistent changes in the soil C:N ratio and AA, but only in soils beneath Eriophorum vaginatum, the dominant tussock-forming sedge. These differential responses suggest that the ecological consequences of warmer climates on these tundra ecosystems are more complex than simply elevated N-mineralization rates, and that the responses of landscapes might be impacted by soil age, or time since deglaciation.

Salt marsh nitrogen cycling: where land meets sea.

Salt marshes sit at the terrestrial-aquatic interface of oceans around the world. Unique features of salt marshes that differentiate them from their upland or offshore counterparts include high rates of primary production from vascular plants and saturated saline soils that lead to sharp redox gradients and a diversity of electron acceptors and donors. Moreover, the dynamic nature of root oxygen loss and tidal forcing leads to unique biogeochemical conditions that promote nitrogen cycling. Here, we highlight recent advances in our understanding of key nitrogen cycling processes in salt marshes and discuss areas where additional research is needed to better predict how salt marsh N cycling will respond to future environmental change.

Dissimilatory nitrate reduction to ammonium (DNRA) is marginal relative to denitrification in emerging-eroding wetlands in a subtropical oligohaline and eutrophic coastal delta.

Nitrate (NO3-) and ammonium (NH4+) are reactive nitrogen (Nr) forms that can exacerbate eutrophication in coastal regions. NO3- can be lost to the atmosphere as N2 gas driven by direct denitrification, coupled nitrification-denitrification and annamox or retained within the ecosystems through conversion of NO3- to NH4+ via dissimilatory nitrate reduction to ammonium (DNRA). Denitrification and DNRA are competitive pathways and hence it is critical to evaluate their functional biogeochemical role. However, there is limited information about the environmental factors driving DNRA in oligohaline habitats, especially within deltaic regions where steep salinity gradients define wetland spatiotemporal distribution. Here we use the Isotope Pairing Technique to evaluate the effect of temperature (10, 20, 30 °C) and in situ soil/sediment organic matter (OM%) on total denitrification (Dtotal = direct + coupled nitrification) and DNRA rates in oligohaline forested/marsh wetlands soils and benthic sediment habitats at two sites representing prograding (Wax Lake Delta, WLD) and eroding (Barataria- Lake Cataouatche, BLC) deltaic stages in the Mississippi River Delta Plain (MRDP). Both sites receive MR water with high NO3- (>40 µM) concentrations during the year via river diversions. Denitrification rates were significantly higher (range: 18.0 ± 0.4-113.0 ± 10.6 µmol m-2 h-1) than DNRA rates (range: 0.7 ± 0.2-9.2 ± 0.3 µmol m-2 h-1). Therefore, DNRA represented on average < 10% of the total NO3- reduction (DNRA + Dtotal). Unlike denitrification, DNRA showed no consistent response to temperature. These results indicate that DNRA in wetland soils and benthic sediment is not a major nitrogen transformation in oligohaline regions across the MRDP regardless of wide range of OM% content in these eroding and prograding delta lobes.

The Diversity and Functional Capacity of Microbes Associated with Coastal Macrophytes.

Coastal marine macrophytes exhibit some of the highest rates of primary productivity in the world. They have been found to host a diverse set of microbes, many of which may impact the biology of their hosts through metabolisms that are unique to microbial taxa. Here, we characterized the metabolic functions of macrophyte-associated microbial communities using metagenomes collected from 2 species of kelp (Laminaria setchellii and Nereocystis luetkeana) and 3 marine angiosperms (Phyllospadix scouleri, P. serrulatus, and Zostera marina), including the rhizomes of two surfgrass species (Phyllospadix spp.), the seagrass Zostera marina, and the sediments surrounding P. scouleri and Z. marina. Using metagenomic sequencing, we describe 63 metagenome-assembled genomes (MAGs) that potentially benefit from being associated with macrophytes and may contribute to macrophyte fitness through their metabolic activity. Host-associated metagenomes contained genes for the use of dissolved organic matter from hosts and vitamin (B1, B2, B7, B12) biosynthesis in addition to a range of nitrogen and sulfur metabolisms that recycle dissolved inorganic nutrients into forms more available to the host. The rhizosphere of surfgrass and seagrass contained genes for anaerobic microbial metabolisms, including nifH genes associated with nitrogen fixation, despite residing in a well-mixed and oxygenated environment. The range of oxygen environments engineered by macrophytes likely explains the diversity of both oxidizing and reducing microbial metabolisms and contributes to the functional capabilities of microbes and their influences on carbon and nitrogen cycling in nearshore ecosystems. IMPORTANCE Kelps, seagrasses, and surfgrasses are ecosystem engineers on rocky shorelines, where they show remarkably high levels of primary production. Through analysis of their associated microbial communities, we found a variety of microbial metabolisms that may benefit the host, including nitrogen metabolisms, sulfur oxidation, and the production of B vitamins. In turn, these microbes have the genetic capabilities to assimilate the dissolved organic compounds released by their macrophyte hosts. We describe a range of oxygen environments associated with surfgrass, including low-oxygen microhabitats in their rhizomes that host genes for nitrogen fixation. The tremendous productivity of coastal seaweeds and seagrasses is likely due in part to the activities of associated microbes, and an increased understanding of these associations is needed.

Abundance of ammonia-oxidizing archaea and bacteria along an estuarine salinity gradient in relation to potential nitrification rates.

Abundance of ammonia-oxidizing Archaea (AOA) was found to be always greater than that of ammonia-oxidizing Bacteria along an estuarine salinity gradient, and AOA abundance was highest at intermediate salinity. However, AOA abundance did not correlate with potential nitrification rates. This lack of correlation may be due to methodological limitations or alternative energy sources.

Wastewater input reductions reverse historic hypereutrophication of Boston Harbor, USA.

This paper documents the changes that followed large nutrient (N and P) and organic matter input reductions to a major metropolitan marine bay, Boston Harbor (USA). Before input reduction, its N and P inputs fell in the upper range of the < 1-> 300 gN m-2 year-1 and < 0.1-> 40 gP m-2 year-1 for coastal systems. Elevated nutrient and organic matter inputs are recognized causes of coastal eutrophication. Treatment upgrades and then diversion of its wastewater discharges offshore, lowered its N, P, and organic C inputs by 80-90%. The input decreases lowered its trophic status from hypereutrophic to eutrophic-mesotrophic. With the reversal of hypereutrophication, pelagic production and phytoplankton biomass decreased, and the nitrogen limitation relative to phosphorus limitation increased. Benthic metabolism and dissolved inorganic N fluxes decreased, and benthic-pelagic coupling was altered. Bottom-water dissolved oxygen, already at healthy levels, increased, and seagrass expanded. Coastal management requires that the changes, following the nutrient and organic matter input reductions implemented to address eutrophication, be understood. Boston Harbor's recovery, because its water column was vertically well mixed and marine, was more pronounced than in many other systems.

Influence of local and regional drivers on spatial and temporal variation of ammonia-oxidizing communities in Gulf of Mexico salt marshes.

We characterized ammonia-oxidizing archaea (AOA) and bacteria (AOB) from salt marsh sediments in the Gulf of Mexico over 5 years to identify environmental drivers of nitrifying community patterns following the Deepwater Horizon oil spill. Samples were collected from oiled and unoiled sites in July of 2012-2016 from 12 marshes spanning three regions on the Louisiana coast. No consistent oil effect was detected for either AOA or AOB abundance or community composition. At the local scale, abundance was correlated with changes in marsh elevation, suggesting that oxygen may be an important driver. Regional differences in abundance were best explained by salinity and soil moisture, while interannual variation may be more linked to changes in climate and Mississippi River discharge. Variation of AOA communities was correlated with organic sediment nutrients, while AOB communities were correlated with soil extractable nutrients. AOA and AOB diversity and AOB abundance decreased in 2014 in all regions, suggesting that broad-scale drivers, such as climate, may explain synchronous shifts throughout the coastal area. Our results provide insights about large-scale disturbances on nitrifying microbes in the Gulf of Mexico, and suggest that nitrogen cycling may be controlled primarily by local factors, but large-scale drivers might override these localized differences at times.

Methods for measuring denitrification: diverse approaches to a difficult problem.

Denitrification, the reduction of the nitrogen (N) oxides, nitrate (NO3-) and nitrite (NO2-), to the gases nitric oxide (NO), nitrous oxide (N2O), and dinitrogen (N2), is important to primary production, water quality, and the chemistry and physics of the atmosphere at ecosystem, landscape, regional, and global scales. Unfortunately, this process is very difficult to measure, and existing methods are problematic for different reasons in different places at different times. In this paper, we review the major approaches that have been taken to measure denitrification in terrestrial and aquatic environments and discuss the strengths, weaknesses, and future prospects for the different methods. Methodological approaches covered include (1) acetylene-based methods, (2) 15N tracers, (3) direct N2 quantification, (4) N2:Ar ratio quantification, (5) mass balance approaches, (6) stoichiometric approaches, (7) methods based on stable isotopes, (8) in situ gradients with atmospheric environmental tracers, and (9) molecular approaches. Our review makes it clear that the prospects for improved quantification of denitrification vary greatly in different environments and at different scales. While current methodology allows for the production of accurate estimates of denitrification at scales relevant to water and air quality and ecosystem fertility questions in some systems (e.g., aquatic sediments, well-defined aquifers), methodology for other systems, especially upland terrestrial areas, still needs development. Comparison of mass balance and stoichiometric approaches that constrain estimates of denitrification at large scales with point measurements (made using multiple methods), in multiple systems, is likely to propel more improvement in denitrification methods over the next few years.

Population Dynamics and Community Composition of Ammonia Oxidizers in Salt Marshes after the Deepwater Horizon Oil Spill.

The recent oil spill in the Gulf of Mexico had significant effects on microbial communities in the Gulf, but impacts on nitrifying communities in adjacent salt marshes have not been investigated. We studied persistent effects of oil on ammonia-oxidizing archaeal (AOA) and bacterial (AOB) communities and their relationship to nitrification rates and soil properties in Louisiana marshes impacted by the Deepwater Horizon oil spill. Soils were collected at oiled and unoiled sites from Louisiana coastal marshes in July 2012, 2 years after the spill, and analyzed for community differences based on ammonia monooxygenase genes (amoA). Terminal Restriction Fragment Polymorphism and DNA sequence analyses revealed significantly different AOA and AOB communities between the three regions, but few differences were found between oiled and unoiled sites. Community composition of nitrifiers was best explained by differences in soil moisture and nitrogen content. Despite the lack of significant oil effects on overall community composition, we identified differences in correlations of individual populations with potential nitrification rates between oiled and unoiled sites that help explain previously published correlation patterns. Our results suggest that exposure to oil, even 2 years post-spill, led to subtle changes in population dynamics. How, or if, these changes may impact ecosystem function in the marshes, however, remains uncertain.

Rhizosphere heterogeneity shapes abundance and activity of sulfur-oxidizing bacteria in vegetated salt marsh sediments.

Salt marshes are highly productive ecosystems hosting an intense sulfur (S) cycle, yet little is known about S-oxidizing microorganisms in these ecosystems. Here, we studied the diversity and transcriptional activity of S-oxidizers in salt marsh sediments colonized by the plant Spartina alterniflora, and assessed variations with sediment depth and small-scale compartments within the rhizosphere. We combined next-generation amplicon sequencing of 16S rDNA and rRNA libraries with phylogenetic analyses of marker genes for two S-oxidation pathways (soxB and rdsrAB). Gene and transcript numbers of soxB and rdsrAB phylotypes were quantified simultaneously, using newly designed (RT)-qPCR assays. We identified a diverse assemblage of S-oxidizers, with Chromatiales and Thiotrichales being dominant. The detection of transcripts from S-oxidizers was mostly confined to the upper 5 cm sediments, following the expected distribution of root biomass. A common pool of species dominated by Gammaproteobacteria transcribed S-oxidation genes across roots, rhizosphere, and surrounding sediment compartments, with rdsrAB transcripts prevailing over soxB. However, the root environment fine-tuned the abundance and transcriptional activity of the S-oxidizing community. In particular, the global transcription of soxB was higher on the roots compared to mix and rhizosphere samples. Furthermore, the contribution of Epsilonproteobacteria-related S-oxidizers tended to increase on Spartina roots compared to surrounding sediments. These data shed light on the under-studied oxidative part of the sulfur cycle in salt marsh sediments and indicate small-scale heterogeneities are important factors shaping abundance and potential activity of S-oxidizers in the rhizosphere.

Functionally distinct communities of ammonia-oxidizing bacteria along an estuarine salinity gradient.

The relationship between ammonia-oxidizing bacteria (AOB) and potential nitrification rates was examined along a salinity gradient in a New England estuary in spring and late summer over 3 years. Ammonia-oxidizing bacteria abundance was estimated by measuring gene copies of the ammonia monooxygenase catalytic subunit (amoA) using real-time polymerase chain reaction. Ammonia-oxidizing bacteria abundance ranged from below detection to 6.0 x 10(7)amoA copies (gdw sediment)(-1). Mean potential nitrification rates ranged from 0.5 to 186.5 nmol N (gdw sediment)(-1) day(-1). Both AOB abundance and potential rates were significantly higher in spring than late summer. Correlations between rates and abundance varied significantly among sites, but showed site-specific ammonia oxidation kinetics related to AOB community structure. The effect of salinity on potential nitrification rates was evaluated by incubating sediment from each site under four salinity conditions (0, 5, 10 and 30 psu). At all sites, rates were generally highest in the intermediate salinity treatments, but rates at the upstream site were inhibited at high salinity, while rates at the two downstream sites were inhibited at the lowest salinity. Although salinity appears to be an important factor in determining AOB distribution, it may not be the primary factor as AOB exhibited a broad range of salinity tolerance in our experiments. Our results indicate that there are significant differences in abundance and community composition of AOB along the salinity gradient, and the differences are reflected in community function.