Marine particle microbiomes during a spring diatom bloom contain active sulfate-reducing bacteria.

Phytoplankton blooms fuel marine food webs with labile dissolved carbon and also lead to the formation of particulate organic matter composed of living and dead algal cells. These particles contribute to carbon sequestration and are sites of intense algal-bacterial interactions, providing diverse niches for microbes to thrive. We analyzed 16S and 18S ribosomal RNA gene amplicon sequences obtained from 51 time points and metaproteomes from 3 time points during a spring phytoplankton bloom in a shallow location (6-10 m depth) in the North Sea. Particulate fractions larger than 10 µm diameter were collected at near daily intervals between early March and late May in 2018. Network analysis identified two major modules representing bacteria co-occurring with diatoms and with dinoflagellates, respectively. The diatom network module included known sulfate-reducing Desulfobacterota as well as potentially sulfur-oxidizing Ectothiorhodospiraceae. Metaproteome analyses confirmed presence of key enzymes involved in dissimilatory sulfate reduction, a process known to occur in sinking particles at greater depths and in sediments. Our results indicate the presence of sufficiently anoxic niches in the particle fraction of an active phytoplankton bloom to sustain sulfate reduction, and an important role of benthic-pelagic coupling for microbiomes in shallow environments. Our findings may have implications for the understanding of algal-bacterial interactions and carbon export during blooms in shallow-water coastal areas.

Particle-attached bacteria act as gatekeepers in the decomposition of complex phytoplankton polysaccharides.

BACKGROUND: Marine microalgae (phytoplankton) mediate almost half of the worldwide photosynthetic carbon dioxide fixation and therefore play a pivotal role in global carbon cycling, most prominently during massive phytoplankton blooms. Phytoplankton biomass consists of considerable proportions of polysaccharides, substantial parts of which are rapidly remineralized by heterotrophic bacteria. We analyzed the diversity, activity, and functional potential of such polysaccharide-degrading bacteria in different size fractions during a diverse spring phytoplankton bloom at Helgoland Roads (southern North Sea) at high temporal resolution using microscopic, physicochemical, biodiversity, metagenome, and metaproteome analyses. RESULTS: Prominent active 0.2-3 µm free-living clades comprised Aurantivirga, "Formosa", Cd. Prosiliicoccus, NS4, NS5, Amylibacter, Planktomarina, SAR11 Ia, SAR92, and SAR86, whereas BD1-7, Stappiaceae, Nitrincolaceae, Methylophagaceae, Sulfitobacter, NS9, Polaribacter, Lentimonas, CL500-3, Algibacter, and Glaciecola dominated 3-10 µm and > 10 µm particles. Particle-attached bacteria were more diverse and exhibited more dynamic adaptive shifts over time in terms of taxonomic composition and repertoires of encoded polysaccharide-targeting enzymes. In total, 305 species-level metagenome-assembled genomes were obtained, including 152 particle-attached bacteria, 100 of which were novel for the sampling site with 76 representing new species. Compared to free-living bacteria, they featured on average larger metagenome-assembled genomes with higher proportions of polysaccharide utilization loci. The latter were predicted to target a broader spectrum of polysaccharide substrates, ranging from readily soluble, simple structured storage polysaccharides (e.g., laminarin, α-glucans) to less soluble, complex structural, or secreted polysaccharides (e.g., xylans, cellulose, pectins). In particular, the potential to target poorly soluble or complex polysaccharides was more widespread among abundant and active particle-attached bacteria. CONCLUSIONS: Particle-attached bacteria represented only 1% of all bloom-associated bacteria, yet our data suggest that many abundant active clades played a pivotal gatekeeping role in the solubilization and subsequent degradation of numerous important classes of algal glycans. The high diversity of polysaccharide niches among the most active particle-attached clades therefore is a determining factor for the proportion of algal polysaccharides that can be rapidly remineralized during generally short-lived phytoplankton bloom events. Video Abstract.

Spatial insurance against a heatwave differs between trophic levels in experimental aquatic communities.

Climate change-related heatwaves are major threats to biodiversity and ecosystem functioning. However, our current understanding of the mechanisms governing community resistance to and recovery from extreme temperature events is still rudimentary. The spatial insurance hypothesis postulates that diverse regional species pools can buffer ecosystem functioning against local disturbances through the immigration of better-adapted taxa. Yet, experimental evidence for such predictions from multi-trophic communities and pulse-type disturbances, like heatwaves, is largely missing. We performed an experimental mesocosm study to test whether species dispersal from natural lakes prior to a simulated heatwave could increase the resistance and recovery of plankton communities. As the buffering effect of dispersal may differ among trophic groups, we independently manipulated the dispersal of organisms from lower (phytoplankton) and higher (zooplankton) trophic levels. The experimental heatwave suppressed total community biomass by having a strong negative effect on zooplankton biomass, probably due to a heat-induced increase in metabolic costs, resulting in weaker top-down control on phytoplankton. While zooplankton dispersal did not alleviate the negative heatwave effects on zooplankton biomass, phytoplankton dispersal enhanced biomass recovery at the level of primary producers, providing partial evidence for spatial insurance. The differential responses to dispersal may be linked to the much larger regional species pool of phytoplankton than of zooplankton. Our results suggest high recovery capacity of community biomass independent of dispersal. However, community composition and trophic structure remained altered due to the heatwave, implying longer-lasting changes in ecosystem functioning.

DNA extraction bias is more pronounced for microbial eukaryotes than for prokaryotes.

DNA extraction and preservation bias is a recurring topic in DNA sequencing-based microbial ecology. The different methodologies can lead to distinct outcomes, which has been demonstrated especially in studies investigating prokaryotic community composition. Eukaryotic microbes are ubiquitous, diverse, and increasingly a subject of investigation in addition to bacteria and archaea. However, little is known about how the choice of DNA preservation and extraction methodology impacts perceived eukaryotic community composition. In this study, we compared the effect of two DNA preservation methods and six DNA extraction methods on the community profiles of both eukaryotes and prokaryotes in phototrophic biofilms on seagrass (Zostera marina) leaves from the Baltic Sea. We found that, whereas both DNA preservation and extraction method caused significant bias in perceived community composition for both eukaryotes and prokaryotes, extraction bias was more pronounced for eukaryotes than for prokaryotes. In particular, soft-bodied and hard-shelled eukaryotes like nematodes and diatoms, respectively, were differentially abundant depending on the extraction method. We conclude that careful consideration of DNA preservation and extraction methodology is crucial to achieving representative community profiles of eukaryotes in marine biofilms and likely all other habitats containing diverse eukaryotic microbial communities.

Tissue- and Population-Level Microbiome Analysis of the Wasp Spider Argiope bruennichi Identified a Novel Dominant Bacterial Symbiont.

Many ecological and evolutionary processes in animals depend upon microbial symbioses. In spiders, the role of the microbiome in these processes remains mostly unknown. We compared the microbiome between populations, individuals, and tissue types of a range-expanding spider, using 16S rRNA gene sequencing. Our study is one of the first to go beyond targeting known endosymbionts in spiders and characterizes the total microbiome across different body compartments (leg, prosoma, hemolymph, book lungs, ovaries, silk glands, midgut, and fecal pellets). Overall, the microbiome differed significantly between populations and individuals, but not between tissue types. The microbiome of the wasp spider Argiope bruennichi features a novel dominant bacterial symbiont, which is abundant in every tissue type in spiders from geographically distinct populations and that is also present in offspring. The novel symbiont is affiliated with the Tenericutes, but has low sequence identity (<85%) to all previously named taxa, suggesting that the novel symbiont represents a new bacterial clade. Its presence in offspring implies that it is vertically transmitted. Our results shed light on the processes that shape microbiome differentiation in this species and raise several questions about the implications of the novel dominant bacterial symbiont on the biology and evolution of its host.

Drying and Rainfall Shape the Structure and Functioning of Nitrifying Microbial Communities in Riverbed Sediments.

Non-flow periods in fluvial ecosystems are a global phenomenon. Streambed drying and rewetting by sporadic rainfalls could drive considerable changes in the microbial communities that govern stream nitrogen (N) availability at different temporal and spatial scales. We performed a microcosm-based experiment to investigate how dry period duration (DPD) (0, 3, 6, and 9 weeks) and magnitude of sporadic rewetting by rainfall (0, 4, and 21 mm applied at end of dry period) affected stocks of N in riverbed sediments, ammonia-oxidizing bacteria (AOB) and archaea (AOA) and rates of ammonia oxidation (AO), and emissions of nitrous oxide (N2O) to the atmosphere. While ammonium (NH4 +) pool size decreased, nitrate (NO3 -) pool size increased in sediments with progressive drying. Concomitantly, the relative and absolute abundance of AOB and, especially, AOA (assessed by 16S rRNA gene sequencing and quantitative PCR of ammonia monooxygenase genes) increased, despite an apparent decrease of AO rates with drying. An increase of N2O emissions occurred at early drying before substantially dropping until the end of the experiment. Strong rainfall of 21 mm increased AO rates and NH4 + in sediments, whereas modest rainfall of 4 mm triggered a notable increase of N2O fluxes. Interestingly, such responses were detected only after 6 and 9 weeks of drying. Our results demonstrate that progressive drying drives considerable changes in in-stream N cycling and the associated nitrifying microbial communities, and that sporadic rainfall can modulate these effects. Our findings are particularly relevant for N processing and transport in rivers with alternating dry and wet phases - a hydrological scenario expected to become more important in the future.

Light availability impacts structure and function of phototrophic stream biofilms across domains and trophic levels.

Phototrophic biofilms are ubiquitous in freshwater and marine environments where they are critical for biogeochemical cycling, food webs and in industrial applications. In streams, phototrophic biofilms dominate benthic microbial life and harbour an immense prokaryotic and eukaryotic microbial biodiversity with biotic interactions across domains and trophic levels. Here, we examine how community structure and function of these biofilms respond to varying light availability, as the crucial energy source for phototrophic biofilms. Using metatranscriptomics, we found that under light limitation-dominant phototrophs, including diatoms and cyanobacteria, displayed a remarkable plasticity in their photosynthetic machinery manifested as higher abundance of messenger RNAs (mRNAs) involved in photosynthesis and chloroplast ribosomal RNA. Under higher light availability, bacterial mRNAs involved in phosphorus metabolism, mainly from Betaproteobacteria and Cyanobacteria, increased, likely compensating for nutrient depletion in thick biofilms with high biomass. Consumers, including diverse ciliates, displayed community shifts indicating preferential grazing on algae instead of bacteria under higher light. For the first time, we show that the functional integrity of stream biofilms under variable light availability is maintained by structure-function adaptations on several trophic levels. Our findings shed new light on complex biofilms, or "microbial jungles", where in analogy to forests, diverse and multitrophic level communities lend stability to ecosystem functioning. This multitrophic level perspective, coupling metatranscriptomics to process measurements, could advance understanding of microbial-driven ecosystems beyond biofilms, including planktonic and soil environments.

Eelgrass Leaf Surface Microbiomes Are Locally Variable and Highly Correlated with Epibiotic Eukaryotes.

Eelgrass (Zostera marina) is a marine foundation species essential for coastal ecosystem services around the northern hemisphere. Like all macroscopic organisms, it possesses a microbiome (here defined as an associated prokaryotic community) which may play critical roles in modulating the interaction of eelgrass with its environment. For example, its leaf surface microbiome could inhibit or attract eukaryotic epibionts which may overgrow the eelgrass leading to reduced primary productivity and subsequent eelgrass meadow decline. We used amplicon sequencing of the 16S and 18S rRNA genes of prokaryotes and eukaryotes to assess the leaf surface microbiome (prokaryotes) as well as eukaryotic epibionts in- and outside lagoons on the German Baltic Sea coast. Prokaryote microbiomes varied substantially both between sites inside lagoons and between open coastal and lagoon sites. Water depth, leaf area and biofilm chlorophyll a concentration explained a large amount of variation in both prokaryotic and eukaryotic community composition. The prokaryotic microbiome and eukaryotic epibiont communities were highly correlated, and network analysis revealed disproportionate co-occurrence between a limited number of eukaryotic taxa and several bacterial taxa. This suggests that eelgrass leaf surfaces are home to a mosaic of microbiomes of several epibiotic eukaryotes, in addition to the microbiome of the eelgrass itself. Our findings thereby underline that eukaryotic diversity should be taken into account in order to explain prokaryotic microbiome assembly and dynamics in aquatic environments.

The ecology and biogeochemistry of stream biofilms.

Streams and rivers form dense networks, shape the Earth's surface and, in their sediments, provide an immensely large surface area for microbial growth. Biofilms dominate microbial life in streams and rivers, drive crucial ecosystem processes and contribute substantially to global biogeochemical fluxes. In turn, water flow and related deliveries of nutrients and organic matter to biofilms constitute major constraints on microbial life. In this Review, we describe the ecology and biogeochemistry of stream biofilms and highlight the influence of physical and ecological processes on their structure and function. Recent advances in the study of biofilm ecology may pave the way towards a mechanistic understanding of the effects of climate and environmental change on stream biofilms and the biogeochemistry of stream ecosystems.

Light availability affects stream biofilm bacterial community composition and function, but not diversity.

Changes in riparian vegetation or water turbidity and browning in streams alter the local light regime with potential implications for stream biofilms and ecosystem functioning. We experimented with biofilms in microcosms grown under a gradient of light intensities (range: 5-152 µmole photons s(-1) m(-2) ) and combined 454-pyrosequencing and enzymatic activity assays to evaluate the effects of light on biofilm structure and function. We observed a shift in bacterial community composition along the light gradient, whereas there was no apparent change in alpha diversity. Multifunctionality, based on extracellular enzymes, was highest under high light conditions and decoupled from bacterial diversity. Phenol oxidase activity, involved in the degradation of polyphenolic compounds, was twice as high on average under the lowest compared with the highest light condition. This suggests a shift in reliance of microbial heterotrophs on biofilm phototroph-derived organic matter under high light availability to more complex organic matter under low light. Furthermore, extracellular enzyme activities correlated with nutrient cycling and community respiration, supporting the link between biofilm structure-function and biogeochemical fluxes in streams. Our findings demonstrate that changes in light availability are likely to have significant impacts on biofilm structure and function, potentially affecting stream ecosystem processes.

Functional and structural responses of hyporheic biofilms to varying sources of dissolved organic matter.

Headwater streams are tightly connected with the terrestrial milieu from which they receive deliveries of organic matter, often through the hyporheic zone, the transition between groundwater and streamwater. Dissolved organic matter (DOM) from terrestrial sources (that is, allochthonous) enters the hyporheic zone, where it may mix with DOM from in situ production (that is, autochthonous) and where most of the microbial activity takes place. Allochthonous DOM is typically considered resistant to microbial metabolism compared to autochthonous DOM. The composition and functioning of microbial biofilm communities in the hyporheic zone may therefore be controlled by the relative availability of allochthonous and autochthonous DOM, which can have implications for organic matter processing in stream ecosystems. Experimenting with hyporheic biofilms exposed to model allochthonous and autochthonous DOM and using 454 pyrosequencing of the 16S rRNA (targeting the "active" community composition) and of the 16S rRNA gene (targeting the "bulk" community composition), we found that allochthonous DOM may drive shifts in community composition whereas autochthonous DOM seems to affect community composition only transiently. Our results suggest that priority effects based on resource-driven stochasticity shape the community composition in the hyporheic zone. Furthermore, measurements of extracellular enzymatic activities suggest that the additions of allochthonous and autochthonous DOM had no clear effect on the function of the hyporheic biofilms, indicative of functional redundancy. Our findings unravel possible microbial mechanisms that underlie the buffering capacity of the hyporheic zone and that may confer stability to stream ecosystems.

No evidence of aquatic priming effects in hyporheic zone microcosms.

The priming effect refers to quantitative changes in microbial decomposition of recalcitrant organic matter upon addition of labile organic matter and is a phenomenon that mainly has been reported and debated in soil science. Recently, priming effects have been indicated in aquatic ecosystems and have received attention due to the potential significance for ecosystem carbon budgets. Headwater stream biofilms, which are important degraders of both allochthonous, presumably recalcitrant, organic matter and labile autochthonous organic matter, may be sites where priming effects are important in aquatic environments. We have experimentally tested for priming effects in stream biofilms within microcosms mimicking the stream hyporheic zone. A (13)C labeled model allochthonous carbon source was used in combination with different carbon sources simulating autochthonous inputs. We did not detect changes in respiration, removal or incorporation of allochthonous organic matter in response to autochthonous treatments, thus not supporting the occurrence of priming effects under the experimental conditions. This study is the first to address priming effects in the hyporheic zone, and one of very few studies quantitatively assessing aquatic priming effects. The results contrast with existing studies, which highlights the need for quantitative approaches to determine the importance of priming effects in aquatic environments.

Bacterial diversity in relation to secondary production and succession on surfaces of the kelp Laminaria hyperborea.

Kelp forests worldwide are known as hotspots for macroscopic biodiversity and primary production, yet very little is known about the biodiversity and roles of microorganisms in these ecosystems. Secondary production by heterotrophic bacteria associated to kelp is important in the food web as a link between kelp primary production and kelp forest consumers. The aim of this study was to investigate the relationship between bacterial diversity and two important processes in this ecosystem; bacterial secondary production and primary succession on kelp surfaces. To address this, kelp, Laminaria hyperborea, from southwestern Norway was sampled at different geographical locations and during an annual cycle. Pyrosequencing (454-sequencing) of amplicons of the 16S rRNA gene of bacteria was used to study bacterial diversity. Incorporation of tritiated thymidine was used as a measure of bacterial production. Our data show that bacterial diversity (richness and evenness) increases with the age of the kelp surface, which corresponds to the primary succession of its bacterial communities. Higher evenness of bacterial operational taxonomical units (OTUs) is linked to higher bacterial production. Owing to the dominance of a few abundant OTUs, kelp surface biofilm communities may be characterized as low-diversity habitats. This is the first detailed study of kelp-associated bacterial communities using high-throughput sequencing and it extends current knowledge on microbial community assembly and dynamics on living surfaces.

Exploring the composition and diversity of microbial communities at the Jan Mayen hydrothermal vent field using RNA and DNA.

DNA sequencing technology has proven very valuable for analysing the microbiota of poorly accessible ecosystems such as hydrothermal vents. Using a combination of amplicon and shotgun sequencing of small-subunit rRNA and its gene, we examined the composition and diversity of microbial communities from the recently discovered Jan Mayen vent field, located on Mohn's Ridge in the Norwegian-Greenland Sea. The communities were dominated by the epsilonproteobacterial genera Sulfurimonas and Sulfurovum. These are mesophiles involved in sulphur metabolism and typically found in vent fluid mixing zones. Composition and diversity predictions differed systematically between extracted DNA and RNA samples as well as between amplicon and shotgun sequencing. These differences were more substantial than those between two biological replicates. Amplicon vs. shotgun sequencing differences could be explained to a large extent by bias introduced during PCR, caused by preferential primer-template annealing, while DNA vs. RNA differences were thought to be caused by differences between the activity levels of taxa. Further, predicted diversity from RNA samples was consistently lower than that from DNA. In summary, this study illustrates how different methods can provide complementary ecological insights.

Planctomycetes dominate biofilms on surfaces of the kelp Laminaria hyperborea.

BACKGROUND: Bacteria belonging to Planctomycetes display several unique morphological and genetic features and are found in a wide variety of habitats on earth. Their ecological roles in these habitats are still poorly understood. Planctomycetes have previously been detected throughout the year on surfaces of the kelp Laminaria hyperborea from southwestern Norway. We aimed to make a detailed investigation of the abundance and phylogenetic diversity of planctomycetes inhabiting these kelp surfaces. RESULTS: Planctomycetes accounted for 51-53% of the bacterial biofilm cells in July and September and 24% in February according to fluorescence in situ hybridization (FISH) results. Several separate planctomycetes lineages within Pirellulae, Planctomyces and OM190 were represented in 16S rRNA gene clone libraries and the most abundant clones belonged to yet uncultured lineages. In contrast to the abundance, the diversity of the planctomycete populations increased from July to February and was probably influenced by the aging of the kelp tissue. One planctomycete strain that was closely related to Rhodopirellula baltica was isolated using selective cultivation techniques. CONCLUSIONS: Biofilms on surfaces of L. hyperborea display an even higher proportion of planctomycetes compared to other investigated planctomycete-rich habitats such as open water, sandy sediments and peat bogs. The findings agree well with the hypothesis of the role of planctomycetes as degraders of sulfated polymeric carbon in the marine environment as kelps produce such substances. In addition, the abundant planctomycete populations on kelp surfaces and in association with other eukaryotes suggest that coexistence with eukaryotes may be a key feature of many planctomycete lifestyles.