At hydrothermal vent sites, chimneys consisting of sulfides, sulfates, and oxides are formed upon contact of reduced hydrothermal fluids with oxygenated seawater. The walls and surfaces of these chimneys are an important habitat for vent-associated microorganisms. We used community proteogenomics to investigate and compare the composition, metabolic potential and relative in situ protein abundance of microbial communities colonizing two actively venting hydrothermal chimneys from the Manus Basin back-arc spreading center (Papua New Guinea). We identified overlaps in the in situ functional profiles of both chimneys, despite differences in microbial community composition and venting regime. Carbon fixation on both chimneys seems to have been primarily mediated through the reverse tricarboxylic acid cycle and fueled by sulfur-oxidation, while the abundant metabolic potential for hydrogen oxidation and carbon fixation via the Calvin-Benson-Bassham cycle was hardly utilized. Notably, the highly diverse microbial community colonizing the analyzed black smoker chimney had a highly redundant metabolic potential. In contrast, the considerably less diverse community colonizing the diffusely venting chimney displayed a higher metabolic versatility. An increased diversity on the phylogenetic level is thus not directly linked to an increased metabolic diversity in microbial communities that colonize hydrothermal chimneys.
Characterization of the first “Candidatus Nitrotoga” isolate reveals metabolic versatility and separate evolution of widespread nitrite-oxidizing bacteria
Kitzinger K, Koch H, Lücker S, Sedlacek CJ, Herbold C, Schwarz J, Daebeler A, Mueller AJ, Lukumbuzya M, Romano S, Leisch N, Karst SM, Kirkegaard R, Albertsen M, Nielsen PH, Wagner M, Daims H
2018 - mBio, 9: e01186-18
Nitrification is a key process of the biogeochemical nitrogen cycle and of biological wastewater treatment. The second step, nitrite oxidation to nitrate, is catalyzed by phylogenetically diverse, chemolithoautotrophic nitrite-oxidizing bacteria (NOB). Uncultured NOB from the genus “Candidatus Nitrotoga” are widespread in natural and engineered ecosystems. Knowledge about their biology is sparse, because no genomic information and no pure “Ca. Nitrotoga” culture was available. Here we obtained the first “Ca. Nitrotoga” isolate from activated sludge. This organism, “Candidatus Nitrotoga fabula,” prefers higher temperatures (>20°C; optimum, 24 to 28°C) than previous “Ca. Nitrotoga” enrichments, which were described as cold-adapted NOB. “Ca. Nitrotoga fabula” also showed an unusually high tolerance to nitrite (activity at 30 mM NO2−) and nitrate (up to 25 mM NO3−). Nitrite oxidation followed Michaelis-Menten kinetics, with an apparent Km (Km(app)) of ~89 µM nitrite and a Vmax of ~28 µmol of nitrite per mg of protein per h. Key metabolic pathways of “Ca. Nitrotoga fabula” were reconstructed from the closed genome. “Ca. Nitrotoga fabula” possesses a new type of periplasmic nitrite oxidoreductase belonging to a lineage of mostly uncharacterized proteins. This novel enzyme indicates (i) separate evolution of nitrite oxidation in “Ca. Nitrotoga” and other NOB, (ii) the possible existence of phylogenetically diverse, unrecognized NOB, and (iii) together with new metagenomic data, the potential existence of nitrite-oxidizing archaea. For carbon fixation, “Ca. Nitrotoga fabula” uses the Calvin-Benson-Bassham cycle. It also carries genes encoding complete pathways for hydrogen and sulfite oxidation, suggesting that alternative energy metabolisms enable “Ca. Nitrotoga fabula” to survive nitrite depletion and colonize new niches.
Recognizing Patterns: Spatial Analysis of Observed Microbial Colonization on Root Surfaces
Schmidt H, Nunan N, Höck A, Eickhorst T, Kaiser C, Woebken D, Xaynaud X
2018 - Front Environ Sci, in press
Root surfaces are major sites of interactions between plants and associated microorganisms. Here, plants and microbes communicate via signaling molecules, compete for nutrients, and release substrates that may have beneficial or harmful effects on each other. Whilst the body of knowledge on the abundance and diversity of microbial communities at root-soil interfaces is now substantial, information on their spatial distribution at the microscale is still scarce. In this study, a standardized method for recognizing and analyzing microbial cell distributions on root surfaces is presented. Fluorescence microscopy was combined with automated image analysis and spatial statistics to explore the distribution of bacterial colonization patterns on rhizoplanes of rice roots. To test and evaluate the presented approach, a gnotobiotic experiment was performed using a potential nitrogen-fixing bacterial strain in combination with roots of wetland rice. The automated analysis procedure resulted in reliable spatial data of bacterial cells colonizing the rhizoplane. Among all replicate roots, the analysis revealed an increasing density of bacterial cells from the root tip to the region of root cell maturation. Moreover, bacterial cells showed significant spatial clustering and tended to be located around plant root cell walls. The quantitative data suggest that the structure of the root surface plays a major role in bacterial colonization patterns. Possible adaptations of the presented approach for future studies are discussed along with potential pitfalls such as inaccurate imaging. Our results demonstrate that standardized recognition and statistical evaluation of microbial colonization on root surfaces holds the potential to increase our understanding of microbial associations with roots and of the underlying ecological interactions.
Evaluation of primers targeting the diazotroph functional gene and development of NifMAP – a bioinformatics pipeline for analyzing nifH amplicon data
Angel R, Nepel M, Panhölzl C, Schmidt H, Herbold C, Eichorst SA, Woebken D
2018 - Front Microbiol, 9: 1-15
Diazotrophic microorganisms introduce biologically available nitrogen (N) to the global N cycle through the activity of the nitrogenase enzyme. The genetically conserved dinitrogenase reductase (nifH) gene is phylogenetically distributed across four clusters (I-IV) and is widely used as a marker gene for N2 fixation, permitting investigators to study the genetic diversity of diazotrophs in nature and target potential participants in N2 fixation. To date there have been limited, standardized pipelines for the nifH functional gene, which is in stark contrast to the rRNA gene. Here we present a bioinformatics pipeline for processing nifH amplicon datasets – NifMAP (“NifHMiSeq Illumina amplicon Analysis Pipeline”), which as a novel aspect uses Hidden-Markov models to filter out homologous genes to nifH. By using this pipeline, we evaluated the broadly inclusive primer pairs (Ueda19F-R6, IGK3-DVV, F2-R6) that target the nifH gene. To evaluate any systematic biases, the nifH gene was amplified with the aforementioned primer pairs in a diverse collection of environmental samples (soils, rhizosphere and roots samples, biological soil crusts and estuarine samples), in addition to a nifH mock community consisting of six phylogenetically diverse members. We noted that all primer pairs co-amplified nifH homologs to varying degrees; up to 90% of the amplicons were nifH homologs with IGK3-DVV in some samples (rhizosphere and roots from tall oat-grass). In regards to specificity, we observed some degree of bias across the primer pairs. For example, primer pair F2-R6 discriminated against cyanobacteria (amongst others), yet captured many sequences from subclusters IIIE and IIIL-N. These aforementioned subclusters were largely missing by the primer pair IGK3-DVV, which also tended to discriminate against Alphaproteobacteria, but amplified sequences within clusters IIIC (affiliated with Clostridia) and clusters IVB and IVC. Primer pair Ueda19F-R6 exhibited the least bias and successfully captured diazotrophs in cluster I and subclusters IIIE, IIIL, IIIM and IIIN, but discriminated against Firmicutes and subcluster IIIC. Taken together, our newly established bioinformatics pipeline, NifMAP, along with our systematic evaluations of nifH primer pairs permit more robust, high-throughput investigations of diazotrophs in diverse environments.
Transmission of fungal partners to incipient Cecropia-tree ant colonies
Mayer VE, Nepel M, Blatrix R, Oberhauser FB, Fiedler K, Schönenberger J, Voglmayr H
2018 - PLoS One, 13: e0192207
Ascomycete fungi in the nests of ants inhabiting plants (= myrmecophytes) are very often cultivated by the ants in small patches and used as food source. Where these fungi come from is not known yet. Two scenarios of fungus recruitment are possible: (1) random infection through spores or hyphal fragments from the environment, or (2) transmission from mother to daughter colonies by the foundress queen. It is also not known at which stage of the colony life cycle fungiculture is initiated, and whether the- symbiont fungi serve as food for the ant queen. To clarify these questions, we investigated four Azteca ant species inhabiting three different Cecropia species (C. insignis, C. obtusifolia, and C. peltata). We analysed an rRNA gene fragment from 52 fungal patches produced by founding queens and compared them with those from established Azteca colonies (n = 54). The infrabuccal pockets of winged queens were dissected to investigate whether young queens carry fungi from their mother colony. Additionally, 15N labelling experiments were done to verify whether the queen feeds on the patches until she is nourished by her first worker offspring. We infer from the results that the fungi cultivated in hollow plant structures are transferred from the parental colony of the young queen. First, fungal genotypes/OTU diversity was not significantly different between foundress queen patches and established colonies, and second, hyphal parts were discovered in the infrabuccal pockets of female alates. We could show that fungiculture already starts before queens lay their eggs, and that the queens do not feed on fungal patch material but feed it to the larvae. Our findings suggest that fungiculture may be crucial for successful colony founding of arboreal ants in the tropics.
Ammonia monooxygenase-mediated cometabolic biotransformation and hydroxylamine-mediated abiotic transformation of micropollutants in an AOB/NOB co-culture
Biotransformation of various micropollutants (MPs) has been found to be positively correlated with nitrification in activated sludge communities. To further elucidate the roles played by ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB), we investigated the biotransformation capabilities of an NOB pure culture (Nitrobacter sp.), and an AOB (Nitrosomonas europaea) / NOB (Nitrobacter sp.) co-culture for fifteen MPs, whose biotransformation were reported previously to be associated with nitrification. The NOB pure culture did not biotransform any investigated MP, whereas the AOB/NOB co-culture was capable of biotransforming asulam, and five other MPs. Two transformation products (TPs) of asulam were identified and tentative structures were proposed. Inhibition studies with octyne, an ammonia monooxygenase (AMO) inhibitor, suggested that AMO was the responsible enzyme for asulam transformation that occurred co-metabolically. Interestingly, hydroxylamine, a key intermediate of all aerobic ammonia oxidizers, was found to react with several MPs at concentrations typically occurring in AOB batch cultures. All of these MPs were also biotransformed by the AOB/NOB co-culture. Moreover, the same asulam TPs were detected in both biotransformation and hydroxylamine-treated abiotic transformation experiments. Thus, in addition to cometabolism likely carried out by AMO, an abiotic transformation route indirectly mediated by AMO might also contribute to MP biotransformation by AOB.
Cultivation and genomic analysis of “Candidatus Nitrosocaldus islandicus”, an obligately thermophilic, ammonia-oxidizing thaumarchaeon from a hot spring biofilm in Graendalur valley, Iceland
Ammonia-oxidizing archaea (AOA) within the phylum Thaumarchaeota are the only known aerobic ammonia oxidizers in geothermal environments. Although molecular data indicate the presence of phylogenetically diverse AOA from the Nitrosocaldus clade, group 1.1b and group 1.1a Thaumarchaeota in terrestrial high-temperature habitats, only one enrichment culture of an AOA thriving above 50 °C has been reported and functionally analyzed. In this study, we physiologically and genomically characterized a newly discovered thaumarchaeon from the deep-branching Nitrosocaldaceae family of which we have obtained a high (~85 %) enrichment from biofilm of an Icelandic hot spring (73 °C). This AOA, which we provisionally refer to as “Candidatus Nitrosocaldus islandicus”, is an obligately thermophilic, aerobic chemolithoautotrophic ammonia oxidizer, which stoichiometricall converts ammonia to nitrite at temperatures between 50 °C and 70 °C. “Ca. N. islandicus” encodes the expected repertoire of enzymes proposed to be required for archaeal ammonia oxidation, but unexpectedly lacks a nirK gene and also possesses no identifiable other enzyme for nitric oxide (NO) generation*. Nevertheless, ammonia oxidation by this AOA appears to be NO-dependent as “Ca. N. islandicus” is, like all other tested AOA, inhibited by the addition of an NO scavenger. Furthermore, comparative genomics revealed that “Ca. N. islandicus” has the potential for aromatic amino acid fermentation as its genome encodes an indolepyruvate oxidoreductase (iorAB) as well as a type 3b hydrogenase, which are not present in any other sequenced AOA. A further surprising genomic feature of this thermophilic ammonia oxidizer is the absence of DNA polymerase D genes – one of the predominant replicative DNA polymerases in all other ammonia-oxidizing Thaumarchaeota. Collectively, our findings suggest that metabolic versatility and DNA replication might differ substantially between obligately thermophilic and other AOA.
Genomic insights into the Acidobacteria reveal strategies for their success in terrestrial environments
Eichorst SA, Trojan D, Roux S, Herbold C, Rattei T, Woebken D
2018 - Environ Microbiol, 20: 1041-1063
Members of the phylum Acidobacteria are abundant and ubiquitous across soils. We performed the largest (to date) comparative genome analysis spanning subdivisions 1, 3, 4, 6, 8, and 23 (n=24) with the goal to identify features to help explain their prevalence in soils and understand their ecophysiology. In contrast to earlier studies, our analysis revealed that bacteriophage integration events along with transposable and mobile elements influenced the structure and plasticity of these genomes. Low- and high-affinity respiratory oxygen reductases were detected in multiple genomes, suggesting the capacity for growing across different oxygen gradients. Amongst many genomes, the capacity to use a diverse collection of carbohydrates, as well as inorganic and organic N sources (such as extracellular peptidases), were detected – both advantageous traits in environments with fluctuating nutrient environments. We also identified multiple soil acidobacteria with the potential to scavenge atmospheric concentrations of H2, now encompassing mesophilic soil strains within the subdivision 1 and 3, in addition to a previously identified thermophilic strain in subdivision 4. This large-scale acidobacteria genome analysis reveals traits that provide genomic, physiological and metabolic versatility, presumably allowing flexibility and versatility in the challenging and fluctuating soil environment.
Bacterial interactions during sequential degradation of cyanobacterial necromass in a sulfidic arctic marine sediment
Müller AL, Pelikan C, de Rezende JR, Wasmund K, Putz M, Glombitza C, Kjeldsen KU, Jørgensen BB, Loy A
2018 - Environ Microbiol, In press
Seafloor microorganisms impact global carbon cycling by mineralizing vast quantities of organic matter (OM) from pelagic primary production, which is predicted to increase in the Arctic because of diminishing sea ice cover. We studied microbial interspecies-carbon-flow during anaerobic OM degradation in arctic marine sediment using stable isotope probing. We supplemented sediment incubations with 13C-labeled cyanobacterial necromass (spirulina), mimicking fresh OM input, or acetate, an important OM degradation intermediate, and monitored sulfate reduction rates and concentrations of volatile fatty acids (VFAs) during substrate degradation. Sequential 16S rRNA gene and transcript amplicon sequencing and fluorescence in situ hybridization combined with Raman microspectroscopy revealed that only few bacterial species were the main degraders of 13C-spirulina necromass. Psychrilyobacter, Psychromonas, Marinifilum, Colwellia, Marinilabiaceae and Clostridiales species were likely involved in the primary hydrolysis and fermentation of spirulina. VFAs, mainly acetate, produced from spirulina degradation were mineralized by sulfate-reducing bacteria and an Arcobacter species. Cellular activity of Desulfobacteraceae and Desulfobulbaceae species during acetoclastic sulfate reduction was largely decoupled from relative 16S rRNA gene abundance shifts. Our findings provide new insights into the identities and physiological constraints that determine the population dynamics of key microorganisms during complex OM degradation in arctic marine sediments.
Application of stable-isotope labelling techniques for the detection of active diazotrophs
Angel R, Panhölzl C, Gabriel R, Herbold C, Wanek W, Richter A, Eichorst SA, Woebken D
2018 - Environmental Microbiology, 20: 44-61
Investigating active participants in the fixation of dinitrogen gas is vital as N is often a limiting factor for primary production. Biological nitrogen fixation (BNF) is performed by a diverse guild of bacteria and archaea (diazotrophs), which can be free-living or symbionts. Free-living diazotrophs are widely distributed in the environment, yet our knowledge about their identity and ecophysiology is still limited. A major challenge in investigating this guild is inferring activity from genetic data as this process is highly regulated. To address this challenge, we evaluated and improved several 15N-based methods for detecting N2 fixation activity (with a focus on soil samples) and studying active diazotrophs. We compared the acetylene reduction assay and the 15N2 tracer method and demonstrated that the latter is more sensitive in samples with low activity. Additionally, tracing 15N into microbial RNA provides much higher sensitivity compared to bulk soil analysis. Active soil diazotrophs were identified with a 15N-RNA-SIP approach optimized for environmental samples and benchmarked to 15N-DNA-SIP. Lastly, we investigated the feasibility of using SIP-Raman microspectroscopy for detecting 15N-labelled cells. Taken together, these tools allow identifying and investigating active free-living diazotrophs in a highly sensitive manner in diverse environments, from bulk to the single-cell level.
Biodegradability standards for carrier bags and plastic films in aquatic environments: a critical review
Harrison JP, Boardman C, O'Callaghan K, Delort A, Song J
2018 - Royal Society Open Science, in press
Plastic litter is encountered in aquatic ecosystems across the globe, including polar environments and the deep sea. To mitigate the adverse societal and ecological impacts of this waste, there has been debate on whether ‘biodegradable' materials should be granted exemptions from plastic bag bans and levies. However, great care must be exercised when attempting to define this term, due to the broad and complex range of physical and chemical conditions encountered within natural ecosystems. Here, we review existing international industry standards and regional test methods for evaluating the biodegradability of plastics within aquatic environments (wastewater, unmanaged freshwater and marine habitats). We argue that current standards and test methods are insufficient in their ability to realistically predict the biodegradability of carrier bags in these environments, due to several shortcomings in experimental procedures and a paucity of information in the scientific literature. Moreover, existing biodegradability standards and test methods for aquatic environments do not involve toxicity testing or account for the potentially adverse ecological impacts of carrier bags, plastic additives, polymer degradation products or small (microscopic) plastic particles that can arise via fragmentation. Successfully addressing these knowledge gaps is a key requirement for developing new biodegradability standard(s) for lightweight carrier bags.
Fluorinated Gold Nanoparticles for Nanostructure Imaging Mass Spectrometry.
Palermo A, Forsberg EM, Warth B, Aisporna AE, Billings E, Kuang E, Benton HP, Berry D, Siuzdak G
2018 - ACS Nano, in press
Nanostructure imaging mass spectrometry (NIMS) with fluorinated gold nanoparticles (f-AuNPs) is a nanoparticle assisted laser desorption/ionization approach that requires low laser energy and has demonstrated high sensitivity. Here we describe NIMS with f-AuNPs for the comprehensive analysis of metabolites in biological tissues. F-AuNPs assist in desorption/ionization by laser-induced release of the fluorocarbon chains with minimal background noise. Since the energy barrier required to release the fluorocarbons from the AuNPs is minimal, the energy of the laser is maintained in the low μJ/pulse range, thus limiting metabolite in-source fragmentation. Electron microscopy analysis of tissue samples after f-AuNP NIMS shows a distinct "raising" of the surface as compared to matrix assisted laser desorption ionization ablation, indicative of a gentle desorption mechanism aiding in the generation of intact molecular ions. Moreover, the use of perfluorohexane to distribute the f-AuNPs on the tissue creates a hydrophobic environment minimizing metabolite solubilization and spatial dislocation. The transfer of the energy from the incident laser to the analytes through the release of the fluorocarbon chains similarly enhances the desorption/ionization of metabolites of different chemical nature, resulting in heterogeneous metabolome coverage. We performed the approach in a comparative study of the colon of mice exposed to three different diets. F-AuNP NIMS allows the direct detection of carbohydrates, lipids, bile acids, sulfur metabolites, amino acids, nucleotide precursors as well as other small molecules of varied biological origins. Ultimately, the diversified molecular coverage obtained provides a broad picture of a tissue's metabolic organization.
Signatures of ecological processes in microbial community time series.
Faust K, Bauchinger F, Laroche B, de Buyl S, Lahti L, Washburne AD, Gonze D, Widder S
2018 - Microbiome, 1: 120
Growth rates, interactions between community members, stochasticity, and immigration are important drivers of microbial community dynamics. In sequencing data analysis, such as network construction and community model parameterization, we make implicit assumptions about the nature of these drivers and thereby restrict model outcome. Despite apparent risk of methodological bias, the validity of the assumptions is rarely tested, as comprehensive procedures are lacking. Here, we propose a classification scheme to determine the processes that gave rise to the observed time series and to enable better model selection.
We implemented a three-step classification scheme in R that first determines whether dependence between successive time steps (temporal structure) is present in the time series and then assesses with a recently developed neutrality test whether interactions between species are required for the dynamics. If the first and second tests confirm the presence of temporal structure and interactions, then parameters for interaction models are estimated. To quantify the importance of temporal structure, we compute the noise-type profile of the community, which ranges from black in case of strong dependency to white in the absence of any dependency. We applied this scheme to simulated time series generated with the Dirichlet-multinomial (DM) distribution, Hubbell's neutral model, the generalized Lotka-Volterra model and its discrete variant (the Ricker model), and a self-organized instability model, as well as to human stool microbiota time series. The noise-type profiles for all but DM data clearly indicated distinctive structures. The neutrality test correctly classified all but DM and neutral time series as non-neutral. The procedure reliably identified time series for which interaction inference was suitable. Both tests were required, as we demonstrated that all structured time series, including those generated with the neutral model, achieved a moderate to high goodness of fit to the Ricker model.
We present a fast and robust scheme to classify community structure and to assess the prevalence of interactions directly from microbial time series data. The procedure not only serves to determine ecological drivers of microbial dynamics, but also to guide selection of appropriate community models for prediction and follow-up analysis.
Detection of Chlamydiaceae and Chlamydia-like organisms on the ocular surface of children and adults from a trachoma-endemic region
Ghasemian E, Inic-Kanada A, Collingro A, Tagini F, Stein E, Alchalabi H, Schuerer N, Keše D, Babiker BE, Borel N, Greub G, Barisani-Asenbauer T
2018 - Sci Rep, 1: 7432
Trachoma, the leading infectious cause of blindness, is caused by Chlamydia trachomatis (Ct), a bacterium of the phylum Chlamydiae. Recent investigations revealed the existence of additional families within the phylum Chlamydiae, also termed Chlamydia-like organisms (CLOs). In this study, the frequency of Ct and CLOs was examined in the eyes of healthy Sudanese (control) participants and those with trachoma (case). We tested 96 children (54 cases and 42 controls) and 93 adults (51 cases and 42 controls) using broad-range Chlamydiae and Ct-specific (omcB) real-time PCR. Samples positive by broad-range Chlamydiae testing were subjected to DNA sequencing. Overall Chlamydiae prevalence was 36%. Sequences corresponded to unclassified and classified Chlamydiae. Ct infection rate was significantly higher in children (31.5%) compared to adults (0%) with trachoma (p < 0.0001). In general, 21.5% of adults and 4.2% of children tested positive for CLOs (p = 0.0003). Our findings are consistent with previous investigations describing the central role of Ct in trachoma among children. This is the first study examining human eyes for the presence of CLOs. We found an age-dependent distribution of CLO DNA in human eyes with significantly higher positivity in adults. Further studies are needed to understand the impact of CLOs in trachoma pathogenicity and/or protection.
Electron transport within living cells is essential for energy conservation in all respiring and photosynthetic organisms. While a few bacteria transport electrons over micrometer distances to their surroundings, filaments of cable bacteria are hypothesized to conduct electric currents over centimeter distances. We used resonance Raman microscopy to analyze cytochrome redox states in living cable bacteria. Cable-bacteria filaments were placed in microscope chambers with sulfide as electron source and oxygen as electron sink at opposite ends. Along individual filaments a gradient in cytochrome redox potential was detected, which immediately broke down upon removal of oxygen or laser cutting of the filaments. Without access to oxygen, a rapid shift toward more reduced cytochromes was observed, as electrons were no longer drained from the filament but accumulated in the cellular cytochromes. These results provide direct evidence for long-distance electron transport in living multicellular bacteria.
Animals evolved in a world teeming with microbes, which play pivotal roles in their health, development, and evolution. Although the overwhelming majority of living animals are invertebrates, the minority of "microbiome" studies focus on this group. Interest in invertebrate-microbe interactions is 2-fold-a range of immune components are conserved across almost all animal (including human) life, and their functional roles may be conserved. Thus, understanding cross talk between microbes and invertebrate animals can lead to insights of broader relevance. Invertebrates offer unique opportunities to "eavesdrop" on intricate host-microbe conversations because they tend to associate with fewer microbes. On the other hand, considering the vast diversity of form and function that has evolved in the invertebrates, they likely evolved an equally diverse range of ways to interact with beneficial microbes. We have investigated only a few of these interactions in detail; thus, there is still great potential for fundamentally new discoveries.
Sulfur-cycling microorganisms impact organic matter decomposition in wetlands and consequently greenhouse gas emissions from these globally relevant environments. However, their identities and physiological properties are largely unknown. By applying a functional metagenomics approach to an acidic peatland, we recovered draft genomes of seven novel Acidobacteria species with the potential for dissimilatory sulfite (dsrAB, dsrC, dsrD, dsrN, dsrT, dsrMKJOP) or sulfate respiration (sat, aprBA, qmoABC plus dsr genes). Surprisingly, the genomes also encoded DsrL, which so far was only found in sulfur-oxidizing microorganisms. Metatranscriptome analysis demonstrated expression of acidobacterial sulfur-metabolism genes in native peat soil and their upregulation in diverse anoxic microcosms. This indicated an active sulfate respiration pathway, which, however, might also operate in reverse for dissimilatory sulfur oxidation or disproportionation as proposed for the sulfur-oxidizing Desulfurivibrio alkaliphilus. Acidobacteria that only harbored genes for sulfite reduction additionally encoded enzymes that liberate sulfite from organosulfonates, which suggested organic sulfur compounds as complementary energy sources. Further metabolic potentials included polysaccharide hydrolysis and sugar utilization, aerobic respiration, several fermentative capabilities, and hydrogen oxidation. Our findings extend both, the known physiological and genetic properties of Acidobacteria and the known taxonomic diversity of microorganisms with a DsrAB-based sulfur metabolism, and highlight new fundamental niches for facultative anaerobic Acidobacteria in wetlands based on exploitation of inorganic and organic sulfur molecules for energy conservation.
Expanded diversity of microbial groups that shape the dissimilatory sulfur cycle
Anantharaman K, Hausmann B, Jungbluth SP, Kantor RS, Lavy A, Warren LA, Rappé MS, Pester M, Loy A, Thomas BC, Banfield JF
2018 - ISME J, 12: 1715-1728
A critical step in the biogeochemical cycle of sulfur on Earth is microbial sulfate reduction, yet organisms from relatively few lineages have been implicated in this process. Previous studies using functional marker genes have detected abundant, novel dissimilatory sulfite reductases (DsrAB) that could confer the capacity for microbial sulfite/sulfate reduction but were not affiliated with known organisms. Thus, the identity of a significant fraction of sulfate/sulfite-reducing microbes has remained elusive. Here we report the discovery of the capacity for sulfate/sulfite reduction in the genomes of organisms from thirteen bacterial and archaeal phyla, thereby more than doubling the number of microbial phyla associated with this process. Eight of the thirteen newly identified groups are candidate phyla that lack isolated representatives, a finding only possible given genomes from metagenomes. Organisms from Verrucomicrobia and two candidate phyla, Candidatus Rokubacteria and Candidatus Hydrothermarchaeota, contain some of the earliest evolved dsrAB genes. The capacity for sulfite reduction has been laterally transferred in multiple events within some phyla, and a key gene potentially capable of modulating sulfur metabolism in associated cells has been acquired by putatively symbiotic bacteria. We conclude that current functional predictions based on phylogeny significantly underestimate the extent of sulfate/sulfite reduction across Earth’s ecosystems. Understanding the prevalence of this capacity is integral to interpreting the carbon cycle because sulfate reduction is often coupled to turnover of buried organic carbon. Our findings expand the diversity of microbial groups associated with sulfur transformations in the environment and motivate revision of biogeochemical process models based on microbial community composition.
Ecology and Biotechnological Potential of Bacteria Belonging to the Genus Pseudovibrio.
2018 - Appl. Environ. Microbiol., 8: in press
Members of the genus have been isolated worldwide from a great variety of marine sources as both free-living and host-associated bacteria. So far, the available data depict a group of alphaproteobacteria characterized by a versatile metabolism, which allows them to use a variety of substrates to meet their carbon, nitrogen, sulfur, and phosphorous requirements. Additionally, -related bacteria have been shown to proliferate under extreme oligotrophic conditions, tolerate high heavy-metal concentrations, and metabolize potentially toxic compounds. Considering this versatility, it is not surprising that they have been detected from temperate to tropical regions and are often the most abundant isolates obtained from marine invertebrates. Such an association is particularly recurrent with marine sponges and corals, animals that play a key role in benthic marine systems. The data so far available indicate that these bacteria are mainly beneficial to the host, and besides being involved in major nutrient cycles, they could provide the host with both vitamins/cofactors and protection from potential pathogens via the synthesis of antimicrobial secondary metabolites. In fact, the biosynthetic abilities of spp. have been emerging in recent years, and both genomic and analytic studies have underlined how these organisms promise novel natural products of biotechnological value.
The giant colonial ciliate Zoothamnium niveum harbors a monolayer of the gammaproteobacteria Cand. Thiobios zoothamnicoli on its outer surface. Cultivation experiments revealed maximal growth and survival under steady flow of high oxygen and low sulfide concentrations. We aimed at directly demonstrating the sulfur-oxidizing, chemoautotrophic nature of the symbionts and at investigating putative carbon transfer from the symbiont to the ciliate host. We performed pulse-chase incubations with C- and C-labeled bicarbonate under varying environmental conditions. A combination of tissue autoradiography and nanoscale secondary ion mass spectrometry coupled with transmission electron microscopy was used to follow the fate of the radioactive and stable isotopes of carbon, respectively. We show that symbiont cells fix substantial amounts of inorganic carbon in the presence of sulfide, but also (to a lesser degree) in the absence of sulfide by utilizing internally stored sulfur. Isotope labeling patterns point to translocation of organic carbon to the host through both release of these compounds and digestion of symbiont cells. The latter mechanism is also supported by ultracytochemical detection of acid phosphatase in lysosomes and in food vacuoles of ciliate cells. Fluorescence in situ hybridization of freshly collected ciliates revealed that the vast majority of ingested microbial cells were ectosymbionts.
Coexistence of novel gammaproteobacterial and Arsenophonus symbionts in the scale insect Greenisca brachypodii (Hemiptera, Coccomorpha: Eriococcidae)
Michalik A, Schulz F, Michalik K, Wascher F, Horn M, Szklarzewicz T
2018 - Environ. Microbiol., in press
Scale insects are commonly associated with obligate, intracellular microorganisms which play important roles in complementing their hosts with essential nutrients. Here we characterized the symbiotic system of Greenisca brachypodii, a member of the family Eriococcidae. Histological and ultrastructural analyses have indicated that G. brachypodii is stably associated with coccoid and rod-shaped bacteria. Phylogenetic analyses have revealed that the coccoid bacteria represent a sister group to the secondary symbiont of the mealybug Melanococcus albizziae, whereas the rod-shaped symbionts are close relatives of Arsenophonus symbionts in insects - to our knowledge, this is the first report of the presence of Arsenophonus bacterium in scale insects. As a comparison of 16S and 23S rRNA genes sequences of the G. brachypodii coccoid symbiont with other gammaprotebacterial sequences showed only low similarity (∼90%), we propose the name 'Candidatus Kotejella greeniscae' for its tentative classification. Both symbionts are transovarially transmitted from one generation to the next. The infection takes place in the neck region of the ovariole. The bacteria migrate between follicular cells, as well as through the cytoplasm of those cells to the perivitelline space, where they form a characteristic 'symbiont ball'. Our findings provide evidence for a polyphyletic origin of symbionts of Eriococcidae. This article is protected by copyright. All rights reserved.
Molecular hydrogen (H2 ) is the key intermediate in the anaerobic degradation of organic matter. Its removal by H2 -oxidizing microorganisms is essential to keep anaerobic degradation energetically favorable. Sulfate-reducing microorganisms (SRM) are known as the main H2 scavengers in anoxic marine sediments. Although the community of marine SRM has been extensively studied, those consuming H2 in situ are completely unknown. We combined metagenomics, PCR-based clone libraries, single-amplified genomes (SAGs) and metatranscriptomics to identify potentially H2 -consuming SRM in anoxic coastal sediments. The vast majority of SRM-related H2 ase sequences were assigned to group 1b and 1c [NiFe]-H2 ases of the deltaproteobacterial order Desulfobacterales. Surprisingly, the same sequence types were similarly highly expressed in spring and summer, suggesting that these are stable and integral members of the H2 -consuming community. Notably, one sequence cluster from the SRM group 1 consistently accounted for around half of all [NiFe]-H2 ase transcripts. Using SAGs, we could link this cluster with the 16S rRNA genes of the uncultured Sva0081-group of the family Desulfobacteraceae. Sequencing of 16S rRNA gene amplicons and H2 ase gene libraries suggested consistently high in situ abundance of the Sva0081 group also in other marine sediments. Together with other Desulfobacterales these likely are important H2 -scavengers in marine sediments. This article is protected by copyright. All rights reserved.
Microplastics (<5 mm particles) occur within both engineered and natural freshwater ecosystems, including wastewater treatment plants, lakes, rivers, and estuaries. While a significant proportion of microplastic pollution is likely sequestered within freshwater environments, these habitats also constitute an important conduit of microscopic polymer particles to oceans worldwide. The quantity of aquatic microplastic waste is predicted to dramatically increase over the next decade, but the fate and biological implications of this pollution are still poorly understood. A growing body of research has aimed to characterize the formation, composition, and spatiotemporal distribution of microplastic-associated (“plastisphere”) microbial biofilms. Plastisphere microorganisms have been suggested to play significant roles in pathogen transfer, modulation of particle buoyancy, and biodegradation of plastic polymers and co-contaminants, yet investigation of these topics within freshwater environments is at a very early stage. Here, what is known about marine plastisphere assemblages is systematically compared with up-to-date findings from freshwater habitats. Through analysis of key differences and likely commonalities between environments, we discuss how an integrated view of these fields of research will enhance our knowledge of the complex behavior and ecological impacts of microplastic pollutants.
Stickstoffkreisläufe in der Abwasserreinigung - neue und bewährte Wege
2018 - 31-46. in Wiener Mitteilungen Wasser-Abwasser-Gewässer, vol. 247. (Krampe, J; Kreuzinger, N)
Der klassische Weg zur Stickstoff-Eliminierung in Kläranlagen beruht auf der Kombination von Nitrifikation und Denitrifikation. In den letzten Jahren haben molekularbiologische Methoden eine Vielzahl neuer Einblicke in die Biologie der Nitrifikanten (Ammoniak- und Nitritoxidierer) ergeben. Diese Erkenntnisse beinhalten eine unerwartet hohe Diversität dieser Bakterien, alternative Stoffwechselwege sowie komplett allein nitrifizierende Mikroben (Comammox-Organismen). Das resultierende neue Bild der Nitrifikation weicht stark vom etablierten Lehrbuchwissen ab. Ein kosten- und energieeffizienterer Weg zur Stickstoff-Eliminierung nutzt die anaerobe Ammoniumoxidation (Anammox-Prozess). Dieser Ansatz nutzt extrem langsam wachsende Bakterien und erfordert die selektive Unterdrückung bestimmter Organismen (Nitritoxidierer). Aus diesen Gründen ist der Anammox-Prozess zwar vielversprechend, die praktische Implementierung ist jedoch Gegenstand aktiver Forschungs- und Optimierungsarbeiten. Ein weiterer neuer Prozess, die nitrit-abhängige anaerobe Methanoxidation (n-damo), wird großtechnisch noch nicht eingesetzt. Insbesondere in Kombination mit Anammox könnte der n-damo Prozess zur gleichzeitigen Eliminierung von restlichem Methan und von Stickstoff interessant werden.
Draft genome sequence of Telmatospirillum siberiense 26-4b1T, an acidotolerant peatland alphaproteobacterium potentially involved in sulfur cycling
The facultative anaerobic chemoorganoheterotrophic alphaproteobacterium Telmatospirillum siberiense 26-4b1T was isolated from a Siberian peatland. We report on a 6.20 Mbp near complete, high quality draft genome of T. siberiense that reveals expected and novel metabolic potential for the genus Telmatospirillum, including genes for sulfur oxidation.
In this infographic, the key metabolic functions of Nitrospira and the role that these bacteria play in nitrification and other processes in the environment is shown. Nitrospira plays pivotal roles in nitrification as an aerobic chemolithoautotrophic nitrite-oxidizing bacterium. These bacteria often occur in close association with ammonia-oxidizing bacteria or archaea that convert ammonia to nitrite, which is further oxidized to nitrate by Nitrospira. However, in 'reciprocal feeding' interactions, Nitrospira can also provide ammonia oxidizers with ammonia released from urea or cyanate, which is further nitrified as described above. Recently discovered Nitrospira members even catalyze both nitrification steps alone and are therefore called complete ammonia oxidizers or 'comammox' organisms. Some strains of Nitrospira utilize alternative substrates, such as H and formate, using oxygen or nitrate as terminal electron acceptor, and can exploit these energy sources concurrently with aerobic nitrite oxidation. This metabolic versatility enables Nitrospira to colonize a broad range of habitats and to sustain shifts in environmental conditions such as changing oxygen concentrations.