Microbial Community Succession Along a Chronosequence in Constructed Salt Marsh Soils.

In this study, we examined the succession of soil microbial communities across a chronosequence of newly constructed salt marshes constructed primarily of fine-grained dredge material, using 16S rRNA amplicon sequences. Alpha diversity in the subsurface horizons was initially low and increased to reference levels within 3 years of marsh construction, while alpha diversity in the newly accumulating organic matter-rich surface soils was initially high and remained unchanged. Microbial community succession was fastest in the surface horizon (~ 24 years to reference equivalency) and became progressively slower with depth in the subsurface horizons (~ 30-67 years). Random forest linear regression analysis was used to identify important taxa driving the trajectories toward reference conditions. In the parent material, putative sulfate-reducers (Desulfobacterota), methanogens (Crenarchaeota, especially Methanosaeta), and fermenters (Chloroflexi and Clostridia) increased over time, suggesting an enrichment of these metabolisms over time, similar to natural marshes. Concurrently in the surface soils, the relative abundances of putative methane-, methyl-, and sulfide oxidizers, especially among Gammaproteobacteria, increased over time, suggesting the co-development of sulfide and methane removal metabolisms in marsh soils. Finally, we observed that the surface soil communities at one of the marshes did not follow the trajectory of the others, exhibiting a greater relative abundance of anaerobic taxa. Uniquely in this dataset, this marsh was developing signs of excessive inundation stress in terms of vegetation coverage and soil geochemistry. Therefore, we suggest that soil microbial community structure may be effective bioindicators of salt marsh inundation and are worthy of further targeted investigation.

Key respiratory genes elucidate bacterial community respiration in a seasonally anoxic estuary.

Intense annual spring phytoplankton blooms and thermohaline stratification lead to anoxia in Chesapeake Bay bottom waters. Once oxygen becomes depleted in the system, microbial communities use energetically favourable alternative electron acceptors for respiration. The extent to which changes in respiration are reflected in community gene expression have only recently been investigated. Metatranscriptomes prepared from near-bottom water plankton over a 4-month time series in central Chesapeake Bay demonstrated changes consistent with terminal electron acceptor availability. The frequency of respiration-related genes in metatranscriptomes was examined by BLASTx against curated databases of genes intimately and exclusively involved in specific electron acceptor utilization pathways. The relative expression of genes involved in denitrification and dissimilatory nitrate reduction to ammonium were coincident with changes in nitrate, nitrite and ammonium concentrations. Dissimilatory iron and manganese reduction transcript ratios increase during anoxic conditions and corresponded with the highest soluble reactive phosphate and manganese concentrations. The sulfide concentration peaked in late July and early August and also matched dissimilatory sulfate reduction transcript ratios. We show that rather than abrupt transitions between terminal electron acceptors, there is substantial overlap in time and space of these various anaerobic respiratory processes in Chesapeake Bay.

Metatranscriptomic analyses of plankton communities inhabiting surface and subpycnocline waters of the Chesapeake Bay during oxic-anoxic-oxic transitions.

We used metatranscriptomics to study the gene transcription patterns of microbial plankton (0.2 to 64 µm) at a mesohaline station in the Chesapeake Bay under transitions from oxic to anoxic waters in spring and from anoxic to oxic waters in autumn. Samples were collected from surface (i.e., above pycnocline) waters (3 m) and from waters beneath the pycnocline (16 to 22 m) in both 2010 and 2011. Metatranscriptome profiles based on function and potential phylogeny were different between 2010 and 2011 and strongly variable in 2011. This difference in variability corresponded with a highly variable ratio of eukaryotic to bacterial sequences (0.3 to 5.5), reflecting transient algal blooms in 2011 that were absent in 2010. The similarity between metatranscriptomes changed at a lower rate during the transition from oxic to anoxic waters than after the return to oxic conditions. Transcripts related to photosynthesis and low-affinity cytochrome oxidases were significantly higher in shallow than in deep waters, while in deep water genes involved in anaerobic metabolism, particularly sulfate reduction, succinyl coenzyme A (succinyl-CoA)-to-propionyl-CoA conversion, and menaquinone synthesis, were enriched relative to in shallow waters. Expected transitions in metabolism between oxic and anoxic deep waters were reflected in elevated levels of anaerobic respiratory reductases and utilization of propenediol and acetoin. The percentage of archaeal transcripts increased in both years in late summer (from 0.1 to 4.4% of all transcripts in 2010 and from 0.1 to 6.2% in 2011). Denitrification-related genes were expressed in a predicted pattern during the oxic-anoxic transition. Overall, our data suggest that Chesapeake Bay microbial assemblages express gene suites differently in shallow and deep waters and that differences in deep waters reflect variable redox states.

Historical contamination of the Anacostia River, Washington, D.C.

The tidal Anacostia River in Washington DC has long been impacted by various sources of chemical pollution over the past 200 years. To explore more recent inputs of various chemicals, six sediment cores were collected for dating and chemical analysis in the downstream section of the tidal Anacostia River. Profiles of contaminants in sediment cores can be useful in determining management direction and effectiveness of pollution controls over time. There were two main objectives for this investigation: (1) determine current sediment contaminant levels; (2) determine a historical perspective of the sediment changes in contamination using (137)Cs and (210)Pb dating. The determination of an age-depth relationship using (210)Pb and (137)Cs dating gave somewhat different results, suggesting that the assumptions of (210)Pb dating were not met. Using the (137)Cs horizon allowed an assignment of approximate sediment accumulation rates and hence an age-depth relationship to contaminant events in the upper portions of the cores. Total PAHs showed higher concentrations at depth and lower surface concentrations. In the upper sections, PAHs were a mixture of combustion and petrogenic sources, while at depth the signature appeared to be of natural origins. Total PCBs, DDTs and chlordane concentrations showed a maximum in recent sediments, decreasing towards the surface. PCBs had lower molecular weight congeners near the surface and higher molecular weights at depth. A phthalate ester, DEHP, appeared in the mid 1940-1950s, and decreased towards the surface. Trace elements fell roughly into three groups. Fe, Mn, and As were in approximately constant proportion to Al, except in some deeper, sandy sediments, where they showed enrichments linked to redox conditions. Ag, Cd, Cu, Hg, Pb, and Zn had low concentrations in the deepest sediments, high concentrations at mid-depths, and declines to intermediate levels at the surface. Ni and Cr followed neither of these patterns closely. We observed that many contaminants appeared in the Anacostia sediments at various times, and reached relatively high concentrations in the past, but are now showing declines in loadings. In some cases, such as PCBs, DDT, chlordane, and Pb from leaded gasoline, these declines can be clearly linked to the discontinuation of their use for environmental reasons. For other contaminants (e.g., PAHs, DEHP, selected metals) these declines are more likely the result of changes in production, usage and waste control.

Mitigation of CyanoHABs Using Phoslock® to Reduce Water Column Phosphorus and Nutrient Release from Sediment.

Cyanobacterial blooms can be stimulated by excessive phosphorus (P) input, especially when diazotrophs are the dominant species. A series of mesocosm experiments were conducted in a lake dominated by a cyanobacteria bloom to study the effects of Phoslock®, a phosphorus adsorbent. The results showed that the addition of Phoslock® lowered the soluble reactive phosphate (SRP) concentrations in water due to efficient adsorption and mitigated the blooms. Once settled on the sediments, Phoslock® serves as a barrier to reduce P diffusion from sediments into the overlying waters. In short-term (1 day) incubation experiments, Phoslock® diminished or reversed SRP effluxes from bottom sediments. At the same time, the upward movement of the oxic-anoxic interface through the sediment column slightly enhanced NH4+ release and depressed N2 release, suggesting the inhibition of nitrification and denitrification. In a long-term (28 days) experiment, Phoslock® hindered the P release, reduced the cyanobacterial abundance, and alleviated the bloom-driven enhancements in the pH and oxygen. These results suggest that, through suppression of internal nutrient effluxes, Phoslock® can be used as an effective control technology to reduce cyanobacteria blooms common to many freshwater systems.

Influence of plant communities on denitrification in a tidal freshwater marsh of the Potomac River, United States.

We investigated whether marsh surface elevation, plant community composition (annuals vs. perennials), and organic matter quantity/quality were associated with differences in denitrification rates in an urban tidal freshwater marsh of the Potomac River, United States. We measured denitrification rates using both denitrification enzyme activity (DEA) with acetylene inhibition (June: n = 38, 3234 +/- 303; October: n = 38, 1557 +/- 368 ng N g dry soil(-1) h(-1)) and direct N(2) flux measurements with membrane inlet mass spectrometry (MIMS) (November: n = 6, 147 +/- 24 mumol m(-2) h(-1)). Organic carbon content and nitrate concentrations in soil, and plant community composition were correlated with elevation, but DEA rates did not differ across marsh surface elevation. Soil organic carbon was highest in plots dominated by perennial graminoids, but DEA rates did not differ across plant community types. The DEA rates increased with increasing soil ammonium concentrations and total N content, and DEA rates differed between summer and fall sampling. The MIMS rates did not differ across plant community types, but were correlated with soil organic N content. Denitrification rates suggest that potential N removal at the site could be substantial. In addition, denitrification rates measured in Dyke Marsh were higher than rates for sediments measured in the adjacent Potomac River. Tidal freshwater marshes can represent an important site for denitrification, and factors fostering denitrification should be considered when restoring urban tidal freshwater wetlands as they are faced with pressures from increasing land use change and sea level rise.

Comparison of methods for determining biogeochemical fluxes from a restored oyster reef.

Oyster reef restoration can significantly increase benthic denitrification rates. Methods applied to measure nutrient fluxes and denitrification from oyster reefs in previous studies include incubations of sediment cores collected adjacent to oyster clumps, benthic chambers filled with intact reef segments that have undergone in situ equilibration and ex situ incubation, and cores with single oysters. However, fluxes of nutrients vary by orders of magnitude among oyster reefs and methods. This study compares two methods of measuring nutrient and metabolic fluxes on restored oyster reefs: incubations including intact segments of oyster reef and incubations containing oyster clumps without underlying sediments. Fluxes of oxygen (O2), dissolved inorganic carbon (DIC), ammonium (NH4+), combined nitrate and nitrite (NO2/3-), di-nitrogen (N2), and soluble reactive phosphorus (SRP) were determined in June and August in Harris Creek, a tributary of the Chesapeake Bay, Maryland, USA. Regression of fluxes measured from clumps alone against those measured from intact reef segments showed significant positive relationships for O2, DIC, NH4+, and SRP (R2 = 0.920, 0.61, 0.26, and 0.52, respectively). Regression of clump fluxes against the oyster tissue biomass indicates significant positive relationships for O2 and NH4+, marginally significant and positive relationships for DIC and N2, and no significant relationship for NO2/3- or SRP. Although these results demonstrate that the incubation of oyster clumps without underlying sediments does not accurately represent biogeochemical fluxes measured from the whole oyster and sediment community, this work supports the need to understand the balance between the metabolism of oysters and local sediments to correctly estimate biogeochemical rates.

Redox reactions and weak buffering capacity lead to acidification in the Chesapeake Bay.

The combined effects of anthropogenic and biological CO2 inputs may lead to more rapid acidification in coastal waters compared to the open ocean. It is less clear, however, how redox reactions would contribute to acidification. Here we report estuarine acidification dynamics based on oxygen, hydrogen sulfide (H2S), pH, dissolved inorganic carbon and total alkalinity data from the Chesapeake Bay, where anthropogenic nutrient inputs have led to eutrophication, hypoxia and anoxia, and low pH. We show that a pH minimum occurs in mid-depths where acids are generated as a result of H2S oxidation in waters mixed upward from the anoxic depths. Our analyses also suggest a large synergistic effect from river-ocean mixing, global and local atmospheric CO2 uptake, and CO2 and acid production from respiration and other redox reactions. Together they lead to a poor acid buffering capacity, severe acidification and increased carbonate mineral dissolution in the USA's largest estuary.The potential contribution of redox reactions to acidification in coastal waters is unclear. Here, using measurements from the Chesapeake Bay, the authors show that pH minimum occurs at mid-depths where acids are produced via hydrogen sulfide oxidation in waters mixed upward from anoxic depths.

The Benthic Exchange of O2, N2 and Dissolved Nutrients Using Small Core Incubations.

The measurement of sediment-water exchange of gases and solutes in aquatic sediments provides data valuable for understanding the role of sediments in nutrient and gas cycles. After cores with intact sediment-water interfaces are collected, they are submerged in incubation tanks and kept under aerobic conditions at in situ temperatures. To initiate a time course of overlying water chemistry, cores are sealed without bubbles using a top cap with a suspended stirrer. Time courses of 4-7 sample points are used to determine the rate of sediment water exchange. Artificial illumination simulates day-time conditions for shallow photosynthetic sediments, and in conjunction with dark incubations can provide net exchanges on a daily basis. The net measurement of N2 is made possible by sampling a time course of dissolved gas concentrations, with high precision mass spectrometric analysis of N2:Ar ratios providing a means to measure N2 concentrations. We have successfully applied this approach to lakes, reservoirs, estuaries, wetlands and storm water ponds, and with care, this approach provides valuable information on biogeochemical balances in aquatic ecosystems.

Transitions in nirS-type denitrifier diversity, community composition, and biogeochemical activity along the Chesapeake Bay estuary.

Chesapeake Bay, the largest estuary in North America, can be characterized as having steep and opposing gradients in salinity and dissolved inorganic nitrogen along the main axis of the Bay. In this study, the diversity of nirS gene fragments (encoding cytochrome cd 1-type nitrite reductase), physical/chemical parameters, and benthic N2-fluxes were analyzed in order to determine how denitrifier communities and biogeochemical activity vary along the estuary salinity gradient. The nirS gene fragments were PCR-amplified, cloned, and sequenced from sediment cores collected at five stations. Sequence analysis of 96-123 nirS clones from each station revealed extensive overall diversity in this estuary, as well as distinct spatial structure in the nirS sequence distributions. Both nirS-based richness and community composition varied among stations, with the most dramatic shifts occurring between low-salinity (oligohaline) and moderate-salinity (mesohaline) sites. For four samples collected in April, the nirS-based richness, nitrate concentrations, and N2-fluxes all decreased in parallel along the salinity gradient from the oligohaline northernmost station to the highest salinity (polyhaline) station near the mouth of the Bay. The vast majority of the 550 nirS sequences were distinct from cultivated denitrifiers, although many were closely related to environmental clones from other coastal and estuarine systems. Interestingly, 8 of the 172 OTUs identified accounted for 42% of the total nirS clones, implying the presence of a few dominant and many rare genotypes, which were distributed in a non-random manner along the salinity gradient of Chesapeake Bay. These data, comprising the largest dataset to investigate nirS clone sequence diversity from an estuarine environment, also provided information that was required for the development of nirS microarrays to investigate the interaction of microbial diversity, environmental gradients, and biogeochemical activity.

Respiratory succession and community succession of bacterioplankton in seasonally anoxic estuarine waters.

Anoxia occurs in bottom waters of stratified estuaries when respiratory consumption of oxygen, primarily by bacteria, outpaces atmospheric and photosynthetic reoxygenation. Once water becomes anoxic, bacterioplankton must change their metabolism to some form of anaerobic respiration. Analysis of redox chemistry in water samples spanning the oxycline of Chesapeake Bay during the summer of 2004 suggested that there was a succession of respiratory metabolism following the loss of oxygen. Bacterial community doubling time, calculated from bacterial abundance (direct counts) and production (anaerobic leucine incorporation), ranged from 0.36 to 0.75 day and was always much shorter than estimates of the time that the bottom water was anoxic (18 to 44 days), indicating that there was adequate time for bacterial community composition to shift in response to changing redox conditions. However, community composition (as determined by PCR-denaturing gradient gel electrophoresis analysis of 16S rRNA genes) in anoxic waters was very similar to that in surface waters in June when nitrate respiration was apparent in the water column and only partially shifted away from the composition of the surface community after nitrate was depleted. Anoxic water communities did not change dramatically until August, when sulfate respiration appeared to dominate. Surface water populations that remained dominant in anoxic waters were Synechococcus sp., Gammaproteobacteria in the SAR86 clade, and Alphaproteobacteria relatives of Pelagibacter ubique, including a putative estuarine-specific Pelagibacter cluster. Populations that developed in anoxic water were most similar (<92% similarity) to uncultivated Firmicutes, uncultivated Bacteroidetes, Gammaproteobacteria in the genus Thioalcalovibrio, and the uncultivated SAR406 cluster. These results indicate that typical estuarine bacterioplankton switch to anaerobic metabolism under anoxic conditions but are ultimately replaced by different organisms under sulfidic conditions.