Determining indicator taxa across spatial and seasonal gradients in the Columbia River coastal margin

Bacterioplankton communities are deeply diverse and highly variable across space and time, but several recent studies demonstrate repeatable and predictable patterns in this diversity. We expanded on previous studies by determining patterns of variability in both individual taxa and bacterial communities across coastal environmental gradients. We surveyed bacterioplankton diversity across the Columbia River coastal margin, USA, using amplicon pyrosequencing of 16S rRNA genes from 596 water samples collected from 2007 to 2010. Our results showed seasonal shifts and annual reassembly of bacterioplankton communities in the freshwater-influenced Columbia River, estuary, and plume, and identified indicator taxa, including species from freshwater SAR11, Oceanospirillales, and Flavobacteria groups, that characterize the changing seasonal conditions in these environments. In the river and estuary, Actinobacteria and Betaproteobacteria indicator taxa correlated strongly with seasonal fluctuations in particulate organic carbon (ρ=−0.664) and residence time (ρ=0.512), respectively. In contrast, seasonal change in communities was not detected in the coastal ocean and varied more with the spatial variability of environmental factors including temperature and dissolved oxygen. Indicator taxa of coastal ocean environments included SAR406 and SUP05 taxa from the deep ocean, and Prochlorococcus and SAR11 taxa from the upper water column. We found that in the Columbia River coastal margin, freshwater-influenced environments were consistent and predictable, whereas coastal ocean community variability was difficult to interpret due to complex physical conditions. This study moves beyond beta-diversity patterns to focus on the occurrence of specific taxa and lends insight into the potential ecological roles these taxa have in coastal ocean environments.     

Tracking differential incorporation of dissolved organic carbon types among diverse lineages of Sargasso Sea bacterioplankton

Bacterioplankton are the primary trophic conduit for dissolved organic carbon (DOC) and linking community structure with DOC utilization is central to understanding global carbon cycling. We coupled stable isotope probing (SIP) with 16S rRNA pyrosequencing in dark seawater culture experiments on euphotic and mesopelagic communities from the Sargasso Sea. Parallel cultures were amended with equimolar quantities of four DO¹³C substrates to simultaneously evaluate community utilization and population‐specific incorporation. Of the substrates tested – two cyanobacterial products (exudates or lysates from a culture of Synechococcus) and two defined monosaccharides (glucose or gluconic acid) – the cyanobacterial exudates were incorporated by the greatest diversity of oligotrophic bacterioplankton populations in surface waters, including taxa from > 10 major subclades within the Flavobacteria, Actinobacteria, Verrucomicrobia and Proteobacteria (including SAR11). In contrast, the monosaccharide glucose was not incorporated by any taxa belonging to extant oligotrophic oceanic clades. Conversely, proteobacterial copiotrophs, which were rare in the ambient water (< 0.1% of sequences), grew rapidly on all DOC amendments at both depths, but with different substrate preferences among lineages. We present a new analytical framework for using SIP to detect DOC incorporation across diverse oligotrophic bacterioplankton and discuss implications for the ecology of bacterial–DOC interactions among populations of diverging trophic strategies.     

Bacterioplankton production in freshwater Antarctic lakes

1. Bacterioplankton production was measured in the water columns of two ultra‐oligotrophic, freshwater Antarctic lakes (Crooked Lake and Lake Druzhby) during an annual cycle. In both lakes bacterial production, measured by the incorporation of [³H] thymidine, continued in winter and showed a cycle over the year. The range of production was between 0 and 479 ng C L^?1 h^?1 in Crooked Lake and 0–354 ng L^?1 h^?1 in Lake Druzhby. 2. Abundance and mean cell volume both varied, producing marked changes in biomass during the year, with highest biomass occurring in the winter and early spring. Biomass showed similar seasonal trends in both lakes. 3. For most of the year inorganic forms of nitrogen and phosphorus were detectable in the water columns of the lakes and were unlikely to have limited bacterial production. Dissolved organic carbon (DOC) was below 3000 μg L^?1. Dissolved amino acids and carbohydrates contributed 5–25% of the DOC pool in Crooked Lake and 5–64% in Lake Druzhby. Dissolved carbohydrates were consistently low, suggesting that this may have been the preferred carbon substrate for bacterioplankton. 4. Aggregate associated bacteria had higher mean cell volume, abundances and production than freely suspended bacteria in Lake Druzhby, while in Crooked Lake aggregate associated bacteria consistently had higher mean cell volumes than free bacteria, but abundance and production were on occasion higher in free bacteria compared with aggregate associated communities. 5. The data indicated that production is limited by continuous low temperatures and the limited availability of suitable DOC substrate. However, the bacterioplankton functions year round, responding to factors other than temperature.     

The microbial carbon pump: from genes to ecosystems

The majority of marine dissolved organic carbon (DOC) is resistant to biological degradation and thus can remain in the water column for thousands of years, constituting carbon sequestration in the ocean. To date the origin of such recalcitrant DOC (RDOC) is unclear. A recently proposed conceptual framework, the microbial carbon pump (MCP), emphasizes the microbial transformation of organic carbon from labile to recalcitrant states. The MCP is concerned with both microbial uptakes and outputs of DOC compounds, covering a wide range from gene to ecosystem levels. In this minireview, the ATP binding cassette (ABC) transporter is used as an example for the microbial processing of DOC at the genetic level. The compositions of the ABC transporter genes of the two major marine bacterial clades Roseobacter and SAR11 demonstrate that they have distinct patterns in DOC utilization: Roseobacter strains have the advantage of taking up carbohydrate DOC, while SAR11 bacteria prefer nitrogen-containing DOC. At the ecosystem level, bacterially derived RDOC based on d-amino acid biomarkers is reported to be responsible for about a quarter of the total marine RDOC pool. Under future global warming scenarios, partitioning of primary production into DOC could be enhanced, and thus the MCP could play an even more important role in carbon sequestration by the ocean. Joint efforts to study the MCP from multiple disciplines are required to obtain a better understanding of ocean carbon cycle and its coupling with global change.     

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Dissolved organic carbon (DOC) and nitrogen (DON) derived from aquatic and terrestrial vascular plants provide a major energy and nutrient source for freshwater and coastal marine biota. The bioavailability of this material may to a large extent depend on plant species. In this study, we have compared the bioavailability of DOC and DON sampled in two distinct stands of Typha domingensisand Eleocharis mutatain a coastal tropical lake and in the adjacent ocean in the state of Rio de Janeiro, Brazil. Bioavailability of organic matter was assessed by regrowth bioassays using natural bacterial inocula. Nutrients were added to achieve carbon or nitrogen limitation. At all sampling sites, DON comprised over 95% of the total bioavailable nitrogen, suggesting its dominant role as a nitrogen source. The bioavailability of lacustrine DON (22% in the Typhastand and 34% in the Eleocharisstand) exceeded the bioavailability of DOC (8 and 10%, respectively) and exhibited a larger difference between the stands. ³H-leucine incorporation studies showed that lake bacterioplankton had a well balanced supply of C, N and P. Therefore, an accumulation of labile DON due to an excess nitrogen supply is not probable. We propose that a substantial part of the lake DON was newly formed within the macrophyte stands, while DOC was predominantly of terrestrial origin and more diagenetically changed.     

Composition and variations in the occurrence of dissolved free simple organic compounds of an unproductive lake ecosystem in northern Sweden

Low molecular weight organic carbon compounds are potentially important carbon and energy substrates to heterotrophic production in the aquatic environment. We studied the occurrence of dissolved free amino acids (AA), monosaccharides (CHO), and carboxylic acids (CA) in the subarctic Lake Diktar-Erik. The lake is unproductive with slightly humic water, and receives water via one major inlet stream draining a birch forest environment. The concentration of dissolved organic carbon (DOC) in the inlet stream was strongly correlated with the discharge. This relationship changed from season to season, indicating changes in the sources of the DOC entering the stream. AA and CHO each accounted for an average of less than 0.5% of the DOC. After high discharge events during the ice-free period, AA and CHO occurred in especially high concentrations. CA occurred in higher concentrations during the ice-free period, when it generally accounted for 20–30% of the DOC pool. The CA content relative to the total DOC pool was strongly inversely correlated with overall DOC concentration, and at low DOC levels the relative content of CA was high and vice versa. This followed a seasonal trend, with CA accounting for a smaller proportion of the DOC in winter and a larger part in spring/early summer. A conservative estimate suggested that the studied simple organic carbon compounds potentially could cover 30% of the bacterial gross production in the lake and therefore potentially also was an important source of CO₂ that occur in supersaturated concentrations in the lake.     

Bacterial Contribution to Dissolved Organic Matter in Eutrophic Lake Kasumigaura,Japan

Incubation experiments using filtered waters from Lake Kasumigaura were conducted to examine bacterial contribution to a dissolved organic carbon (DOC) pool. Bacterial abundance, bacterial production, concentrations of DOC, total dissolved amino acids (TDAA), and total dissolved neutral sugars (TDNS) were monitored during the experiments. Bacterial production during the first few days was very high (20 to 35 μg C liter⁻¹ day⁻¹), accounting for 40 to 70% of primary production. The total bacterial production accounted for 34 to 55% of the DOC loss during the experiment, indicating high bacterial activities in Lake Kasumigaura. The DOC degradation was only 12 to 15%, whereas the degradation of TDAA and TDNS ranged from 30 to 50%, suggesting the preferential usage of TDAA and TDNS. The contribution of bacterially derived carbon to a DOC pool in Lake Kasumigaura was estimated using d-amino acids as bacterial biomarkers and accounted for 30 to 50% of the lake DOC. These values were much higher than those estimated for the open ocean (20 to 30%). The ratio of bacterially derived carbon to bulk carbon increased slightly with time, suggesting that the bacterially derived carbon is more resistant to microbial degradation than bulk carbon. This is the first study to estimate the bacterial contribution to a DOC pool in freshwater environments. These results indicate that bacteria play even more important roles in carbon cycles in freshwater environments than in open oceans and also suggests that recent increases in recalcitrant DOC in various lakes could be attributed to bacterially derived carbon. The potential differences in bacterial contributions to dissolved organic matter (DOM) between freshwater and marine environments are discussed.     

Latitudinal gradients in degradation of marine dissolved organic carbon

Heterotrophic microbial communities cycle nearly half of net primary productivity in the ocean, and play a particularly important role in transformations of dissolved organic carbon (DOC). The specific means by which these communities mediate the transformations of organic carbon are largely unknown, since the vast majority of marine bacteria have not been isolated in culture, and most measurements of DOC degradation rates have focused on uptake and metabolism of either bulk DOC or of simple model compounds (e.g. specific amino acids or sugars). Genomic investigations provide information about the potential capabilities of organisms and communities but not the extent to which such potential is expressed. We tested directly the capabilities of heterotrophic microbial communities in surface ocean waters at 32 stations spanning latitudes from 76°S to 79°N to hydrolyze a range of high molecular weight organic substrates and thereby initiate organic matter degradation. These data demonstrate the existence of a latitudinal gradient in the range of complex substrates available to heterotrophic microbial communities, paralleling the global gradient in bacterial species richness. As changing climate increasingly affects the marine environment, changes in the spectrum of substrates accessible by microbial communities may lead to shifts in the location and rate at which marine DOC is respired. Since the inventory of DOC in the ocean is comparable in magnitude to the atmospheric CO(2) reservoir, such a change could profoundly affect the global carbon cycle.     

Bromodeoxyuridine labelling and fluorescence‐activated cell sorting of polyamine‐transforming bacterioplankton in coastal seawater

Polyamines (PAs) are a group of nitrogen‐rich dissolved organic nitrogen (DON) compounds that are ubiquitously distributed in marine environments. To identify bacteria that are involved in PA transformations, coastal bacterioplankton microcosms were amended with a single PA model compound, i.e. putrescine (PUT) or spermidine (SPD), or with no addition as controls (CTRs). Bromodeoxyuridine (BrdU) was added to all the microcosms to label newly synthesized DNAs. Fluorescence‐activated cell sorting (FACS) analysis indicated significant increases in numbers of total cells and cells with both high and low levels of BrdU incorporation in the PUT and SPD microcosms, but not in the CTRs. 16S rDNA pyrotag sequencing of FACS‐sorted cells indicated that PUT‐ and SPD‐transforming bacteria were composed similarly of a diverse group of taxa affiliated with Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria (especially Roseobacter of its alpha lineage). Broad taxonomic distribution of PA‐transforming bacteria was also indicated by the abundance and distribution of PA transporter gene homologues in a survey of sequenced marine bacterial genomes. Our results suggest that PAs may be common DON substrates for marine bacterioplankton, in line with the hypothesis that bacterially mediated PA transformation accounts for an important proportion of marine DON flux.     

Testing the metabolic theory of ecology with marine bacteria: different temperature sensitivity of major phylogenetic groups during the spring phytoplankton bloom

Although temperature is a key driver of bacterioplankton metabolism, the effect of ocean warming on different bacterial phylogenetic groups remains unclear. Here, we conducted monthly short‐term incubations with natural coastal bacterial communities over an annual cycle to test the effect of experimental temperature on the growth rates and carrying capacities of four phylogenetic groups: SAR11, Rhodobacteraceae, Gammaproteobacteria and Bacteroidetes. SAR11 was the most abundant group year‐round as analysed by CARD‐FISH, with maximum abundances in summer, while the other taxa peaked in spring. All groups, including SAR11, showed high temperature‐sensitivity of growth rates and/or carrying capacities in spring, under phytoplankton bloom or post‐bloom conditions. In that season, Rhodobacteraceae showed the strongest temperature response in growth rates, estimated here as activation energy (E, 1.43 eV), suggesting an advantage to outcompete other groups under warmer conditions. In summer E values were in general lower than 0.65 eV, the value predicted by the Metabolic Theory of Ecology (MTE). Contrary to MTE predictions, carrying capacity tended to increase with warming for all bacterial groups. Our analysis confirms that resource availability is key when addressing the temperature response of heterotrophic bacterioplankton. We further show that even under nutrient‐sufficient conditions, warming differentially affected distinct bacterioplankton taxa.     

Direct effects of organic pollutants on the growth and gene expression of the Baltic Sea model bacterium Rheinheimera sp. BAL341

Organic pollutants (OPs) are critically toxic, bioaccumulative and globally widespread. Moreover, several OPs negatively influence aquatic wildlife. Although bacteria are major drivers of the ocean carbon cycle and the turnover of vital elements, there is limited knowledge of OP effects on heterotrophic bacterioplankton. We therefore investigated growth and gene expression responses of the Baltic Sea model bacterium Rheinheimera sp. BAL341 to environmentally relevant concentrations of distinct classes of OPs in 2-h incubation experiments. During exponential growth, exposure to a mix of polycyclic aromatic hydrocarbons, alkanes and organophosphate esters (denoted MIX) resulted in a significant decrease (between 9% and 18%) in bacterial abundance and production compared with controls. In contrast, combined exposure to perfluorooctanesulfonic acids and perfluorooctanoic acids (denoted PFAS) had no significant effect on growth. Nevertheless, MIX and PFAS exposures both induced significant shifts in gene expression profiles compared with controls in exponential growth. This involved several functional metabolism categories (e.g. stress response and fatty acids metabolism), some of which were pollutant-specific (e.g. phosphate acquisition and alkane-1 monooxygenase genes). In stationary phase, only two genes in the MIX treatment were significantly differentially expressed. The substantial direct influence of OPs on metabolism during bacterial growth suggests that widespread OPs could severely alter biogeochemical processes governed by bacterioplankton.     

Bacterioplankton assemblages transforming dissolved organic compounds in coastal seawater

To characterize bacterioplankton functional assemblages that transform specific components of the coastal seawater dissolved organic carbon (DOC) pool, bromodeoxyuridine (BrdU) was used to label the bacterioplankton cells that were active following addition of single-DOC model compounds: two organic osmolytes [dimethylsulfoniopropionate (DMSP) and glycine betaine (GlyB)] and two aromatic monomers [para-hydroxybenzoic acid (pHBA) and vanillic acid (VanA)]. Bacterial populations were analysed based on in situ fluorescent immunodetection of BrdU incorporation followed by fluorescence-activated cell sorting (FACS). Sorted cells were then characterized by 16S rDNA-based analysis. Populations with high BrdU incorporation level (HI) developed within 8 h of introduction of 100 nM model compound. Terminal restriction fragment length polymorphisms (T-RFLP) analysis indicated that the HI populations in all four amendments were composed of bacteria from the same major taxa (phylum and subphylum levels), but the relative abundance of each differed. High-resolution clone libraries (each containing approximately 200 clones) showed that the HI populations in the GlyB and VanA amendments consisted of both metabolic generalists and specialists within the alpha-Proteobacteria (mainly members of the Roseobacter clade), beta-Proteobacteria and gamma-Proteobacteria (mainly members of Altermonadaceae, Chromatiaceae, Oceanospirillaceae and Pseudomonadaceae). The presence of members of OM60/241, OM185, SAR11, SAR86 and SAR116 in the HI populations indicated that members of these groups can assimilate the model DOC compounds, providing some of the first glimpses into heterotrophy by members of these poorly understood environmental clusters.     

Depleted dissolved organic carbon and distinct bacterial communities in the water column of a rapid-flushing coral reef ecosystem

Coral reefs are highly productive ecosystems bathed in unproductive, low-nutrient oceanic waters, where microbially dominated food webs are supported largely by bacterioplankton recycling of dissolved compounds. Despite evidence that benthic reef organisms efficiently scavenge particulate organic matter and inorganic nutrients from advected oceanic waters, our understanding of the role of bacterioplankton and dissolved organic matter (DOM) in the interaction between reefs and the surrounding ocean remains limited. In this study, we present the results of a 4-year study conducted in a well-characterized coral reef ecosystem (Paopao Bay, Moorea, French Polynesia) where changes in bacterioplankton abundance and dissolved organic carbon (DOC) concentrations were quantified and bacterial community structure variation was examined along spatial gradients of the reef:ocean interface. Our results illustrate that the reef is consistently depleted in concentrations of both DOC and bacterioplankton relative to offshore waters (averaging 79 μmol l⁻¹ DOC and 5.5 × 10⁸ cells l⁻¹ offshore and 68 μmol l⁻¹ DOC and 3.1 × 10⁸ cells l⁻¹ over the reef, respectively) across a 4-year time period. In addition, using a suite of culture-independent measures of bacterial community structure, we found consistent differentiation of reef bacterioplankton communities from those offshore or in a nearby embayment across all taxonomic levels. Reef habitats were enriched in Gamma-, Delta-, and Betaproteobacteria, Bacteriodetes, Actinobacteria and Firmicutes. Specific bacterial phylotypes, including members of the SAR11, SAR116, Flavobacteria, and Synechococcus clades, exhibited clear gradients in relative abundance among nearshore habitats. Our observations indicate that this reef system removes oceanic DOC and exerts selective pressures on bacterioplankton community structure on timescales approximating reef water residence times, observations which are notable both because fringing reefs do not exhibit long residence times (unlike those characteristic of atoll lagoons) and because oceanic DOC is generally recalcitrant to degradation by ambient microbial assemblages. Our findings thus have interesting implications for the role of oceanic DOM and bacterioplankton in the ecology and metabolism of reef ecosystems.     

Primary production and phytoplankton composition in relation to DOC input and bacterioplankton production in humic Lake Örträsket

1. The biomass and production of picophytoplankton, large phytoplankton and heterotrophic bacterioplankton were measured in humic Lake Örträsket, northern Sweden during four consecutive summers.
2. High flow episodes, carrying fresh dissolved organic carbon (DOC) into the lake, always stimulated heterotrophic bacterial production at the expense of primary production. Primary production never exceeded bacterial production for approximately 20 days after such an episode had replenished epilimnial DOC. We suggest that allochthonous DOC is an energy source that stimulates bacterioplankton that, because of their efficient uptake of inorganic nutrients, are then able to outcompete phytoplankton. After the exhaustion of readily available DOC, phytoplankton were able to dominate epilimnion production in Lake Örträsket.
3. Biomass production was higher when dominated by phytoplankton than by bacterioplankton, despite a similar utilization of nutrients in the epilimnion throughout the summer. We propose that different C : N : P ratios of bacterioplankton and phytoplankton permit the latter to produce more carbon (C) biomass per unit of available inorganic nutrients than bacterioplankton.     

Particulate matter composition and bacterial distribution in Terra Nova Bay (Antarctica) during summer 1989–1990

Spatial distributions of particulate organic matter (POM) and microbes were investigated during the summer of 1989–1990 in the coastal waters of Terra Nova Bay (Antarctica). The elemental (organic carbon and nitrogen) and biochemical (lipids, proteins, carbohydrates, DNA and RNA) composition of organic matter was related to bacterioplankton abundance, and pico-phytoplankton density. The ATP concentrations were also measured to gather information about the relationships between particulate matter composition and microbial distribution in Antarctic waters. Total seston was characterized by little spatial variation and was unrelated to the distance from the coast. Suspended particulate matter included some terrestrial components but was mostly composed of autochthonous material. POM was characterized by a uniform distribution and homogeneous composition (mostly of phytoplanktonic origin), and was associated with a relatively scarce microbial community characterized at the surface by high picophytoplankton density. The increase with depth of the living carbon fraction suggested an increase in the microheterotrophic community in the deeper water layers. A significant positive relationship between total bacterioplankton density, and carbohydrate and RNA concentrations was found. Similar significant relationships between pico-phytoplankton abundance and lipids, proteins, carbohydrates and nucleic acids were observed. On the basis of the close coupling found between microbiological and chemical compartments, it seems that, in Terra Nova Bay, bacterial distribution depends on suspended matter and in particular to the labile fraction of the organic detritus.     

Limitation of lowland riverine bacterioplankton by dissolved organic carbon and inorganic nutrients

Flow regulation in lowland rivers has reduced the amount of allochthonous dissolved organic carbon (DOC) entering main channels through less frequent wetting of benches, flood runners and floodplains. The hypothesis tested was that lowland riverine bacterioplankton are DOC limited when flow events are absent and simulating an increase in assimilable DOC similar to that expected during an environmental flow will lead to heterotrophic dominance. Experiments took place in the Namoi River, a highly regulated lowland river in Australia. Specifically, in situ microcosms were used to examine the responses of bacterioplankton and phytoplankton to various additions of DOC as glucose or leaf leachate, with and without additions of inorganic nutrients. The results indicated that ambient DOC availability limited the bacterioplankton for the three seasons over which we conducted the experiments. When DOC was added alone, dissolved oxygen concentrations decreased primarily because of increased bacterial respiration and bacterioplankton growth generally increased relative to controls. Additions of DOC alone led to a pattern of decreased chlorophyll a concentration relative to controls, except for willow leachate. Additions of inorganic nutrients alone increased chlorophyll a concentrations above controls, indicating limitation of phytoplankton. These findings support our hypothesis. Based on the present results, environmental flows should increase the duration of allochthonously driven heterotrophic dominance, thus shifting regulated lowland rivers to more natural (pre-regulation) conditions for greater periods.     

Temporal patterns of microbial community structure in the Mid-Atlantic Bight

Although open ocean time-series sites have been areas of microbial research for years, relatively little is known about the population dynamics of bacterioplankton communities in the coastal ocean on kilometer spatial and seasonal temporal scales. To gain a better understanding of microbial community variability, monthly samples of bacterial biomass were collected in 1995-1996 along a 34-km transect near the Long-Term Ecosystem Observatory (LEO-15) off the New Jersey coast. Surface and bottom sampling was performed at seven stations along a transect line with depths ranging from 1 to 35 m (n=178). Microbial populations were fingerprinted using ribosomal 16S rRNA genes and terminal restriction fragment length polymorphism analysis. Results from cluster analysis revealed distinct temporal patterns among the bacterioplankton communities in the Mid-Atlantic Bight rather than grouping by sample location or depth. Principal components analysis models supported the temporal patterns. In addition, partial least squares regression modeling could not discern a significant correlation from traditional oceanographic physical and phytoplankton nutrient parameters on overall bacterial community variability patterns at LEO-15. These results suggest factors not traditionally measured during oceanographic studies are structuring coastal microbial communities.