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BackgroundRecognition is growing that many ecological and evolutionary processes in animals are dependent upon microbial symbioses. Although there is much known about the ecology and evolution of spiders, the role of the microbiome in these processes remains mostly unknown. We conducted an exploratory study of the microbiome of a range-expanding spider, comparing between populations, individuals, and tissue types (leg, prosoma, hemolymph, book lungs, ovaries, silk glands, midgut, and fecal pellets). Our study is one of the very first to go beyond targeting known endosymbionts in spiders, and characterizes the total microbiome across different body compartments.nnResultsThe microbiome of the wasp spider Argiope bruennichi is dominated by a novel bacterial symbiont, which is highly abundant in every tissue type in spiders from geographically distinct populations, and also present in offspring. The novel symbiont is affiliated with the Tenericutes, but has low sequence identity (

Increased warming in spring and prolonged summer drought may alter soil microbial denitrification. We measured potential denitrification activity and denitrifier marker gene abundances (nirK, nirS, nosZ) in grasslands soils in three geographic regions characterized by site-specific land-use indices (LUI) after warming in spring, at an intermediate sampling and after summer drought. Potential denitrification was significantly increased by warming, but did not persist over the intermediate sampling. At the intermediate sampling, the relevance of grassland land-use intensity was reflected by increased potential N2O production at sites with higher LUI. Abundances of total bacteria did not respond to experimental warming or drought treatments, displaying resilience to minor and short-term effects of climate change. In contrast, nirS- and nirK-type denitrifiers were more influenced by drought in combination with LUI and pH, while the nosZ abundance responded to the summer drought manipulation. Land-use was a strong driver for potential denitrification as grasslands with higher LUI also had greater potentials for N2O emissions. We conclude that both warming and drought affected the denitrifying communities and the potential denitrification in grassland soils. However, these effects are overruled by regional and site-specific differences in soil chemical and physical properties which are also related to grassland land-use intensity

Complex microbial communities in environmental systems play a key role in the detoxification of chemical contaminants by transforming them into less active metabolites or by complete mineralization. Biotransformation, i.e., transformation by microbes, is well understood for a number of priority pollutants, but a similar level of understanding is lacking for many emerging contaminants encountered at low concentrations and in complex mixtures across natural and engineered systems. Any advanced approaches aiming to reduce environmental exposure to such contaminants (e.g., novel engineered biological water treatment systems, design of readily degradable chemicals, or improved regulatory assessment strategies to determine contaminant persistence a priori) will depend on understanding the causal links among contaminant removal, the key driving agents of biotransformation at low concentrations (i.e., relevant microbes and their metabolic activities), and how their presence and activity depend on environmental conditions. In this Perspective, we present the current understanding and recent methodological advances that can help to identify such links, even in complex environmental microbiomes and for contaminants present at low concentrations in complex chemical mixtures. We discuss the ensuing insights into contaminant biotransformation across varying environments and conditions and ask how much closer we have come to designing improved approaches to reducing environmental exposure to contaminants., ACS ES&T Water, 1 (7), ISSN:2690-0637

The research of anaerobic degradation of non-substituted polycyclic aromatic hydrocarbons is still in its infancy and most processes therein only poorly understood. Due to the poor bacterial degradation capabilities of PAHs, only few cultures exist that can be used to explore the underlying mechanisms. Their growth times are considerably longer than those of compounds with a smaller molecular weight and the production of biomass is substantially lower as shown by strain-specific FISH analyses and flow cytometry. Elucidation of pathways started in enrichment cultures growing on naphthalene as sole carbon and electron source and is at a point where individual steps are fairly well characterized through years of research. The next logical step is to look at compounds with a higher molecular weight to find similarities or dissimilarities in the pathways. The compound of interest in this thesis is phenanthrene, a three-ringed PAH without known degradation steps apart from a carboxylation as initial reaction (Zhang and Young 1997; Davidova et al. 2007). Parallels to anaerobic naphthalene degradation are expected due to similarities in the aromatic ring structure. In this thesis I wanted to find insights into the anaerobic degradation of phenanthrene as the next sized PAH beyond the initial carboxylation reaction. Therefore, we isolated bacteria able to degrade hydrocarbons under anoxic conditions from a naturally contaminated ecosystem, the pitch lake in Trinidad, Trinidad & Tobago. An enrichment culture growing anaerobically under sulfate reducing conditions was set up from sediments from the pitch lake. This culture was able to degrade phenanthrene as sole carbon and energy source. It was used to elucidate the anaerobic phenanthrene degradation pathway. Beyond the carboxylation reaction in the C2 position, which could be shown indirectly within the framework of this thesis by metabolite analysis and biochemical enzyme assays, I was able to show the ligation reaction from 2-phenanthroic acid to 2-phenanthroyl-CoA within this culture. This step is therefore similar to the ligation in anaerobic naphthalene degradation from 2-napthoic acid to 2-naphthoyl-CoA. Further downstream reductions steps could not be shown with enzyme assays yet. Nevertheless, metabolite analysis was able to indicate a stepwise ring reduction, which again would be in accordance with the naphthalene reduction steps (Eberlein, Estelmann, et al. 2013; Eberlein, Johannes, et al. 2013). The main dominating bacterium within the culture belongs to the Desulfobacteraceae family and made up for 60% of the culture as confirmed by flow cytometry and genome-resolved metagenomics. It has a 93% similarity to the known naphthalene degrading, sulfate reducing strain NaphS2, indicating that degradation pathways of both molecules were the result of similar gene clusters or mechanisms. While looking for life in the naturally forming asphalt, minuscule water droplets have been discovered, that contain living bacteria (Meckenstock et al. 2014). As an extreme and seemingly uninhabitable habitat within the naturally formed asphalt, important questions towards the origin of the bacteria within the pitch, their way of coping with the lack of oxygen and their access to nutrients remain to be answered. In this thesis, sequencing of DNA extracted from both pitch and water droplets gave further insights into the community composition in compartments and their possible interactions. The diversity in the pitch was higher than in the water droplets, indicating a more specialized microbiota in the water droplets. Diversity within the droplets might change compared to the original diversity in the source water during upward movement due to a local selection and evolution on the “micro-scale”. The diversity in both compartments was very low compared to other oil sources, with more than 30% of the contained bacteria without any known relatives. In other samples like marine oil sources, there are only 10% of the bacteria without known relatives. The dominant bacterium in all sequenced droplets was Tepidiphilus sp., it belongs to the family Hydrogenophilaceae within the β-Proteobacteria and is able to degrade organic acids as sole carbon source under nitrate reducing conditions. It is therefore a plausible candidate to live in this extreme environment, but it still is not characterized well yet. In summary this thesis advances our understanding of anaerobic phenanthrene degradation and I was able to discover the similarities between naphthalene and phenanthrene degradation on an enzymatic basis. This also allows us to assume similarities for PAHs of an even higher molecular weight than that of phenanthrene. The diversity of the degrader community just opened a small window into the life in oil and the background metagenome from the pitch serves as a template for a deeper look into this extreme habitat. Die anaeroben Abbauwege polyzyklischer aromatischer Kohlenwasserstoffe (PAKs) sind bisher weitestgehend unerforscht geblieben. Aufgrund des langsamen Wachstums und geringer Biomassebildung existieren nur wenige Kulturen, anhand derer die Abbauwege aufgeklärt werden können. Des Weiteren sind deutliche Unterschiede der am Abbau beteiligten bekannten Gene zwischen verschiedenen Spezies aufgedeckt worden, was überdies die Aufklärung durch Sequenzierungsdaten verhindert. Der einzig bislang untersuchte anaerobe Abbauweg eines PAKs ist der von Naphthalin, eines Aromaten mit zwei Ringen und einem Molekulargewicht von 128 g/mol. Für diesen Abbauweg wurden die initialisierende Carboxylierung, die Reduktion der aromatischen Strukturen sowie die letztendliche Ringöffnung bereits beschrieben. Der naheliegend folgende Schritt betrifft die Untersuchung der Abbauwege für Schadstoffe mit höherem Molekulargewicht auf Gemeinsamkeiten und Unterschiede. Das Augenmerk dieser Arbeit liegt hierbei auf dem Schadstoff Phenanthren, einem aus drei aromatischen Ringen aufgebauter Kohlenwasserstoff mit einem Molekulargewicht von 178,23 g/mol. Aufgrund der Ähnlichkeiten in der aromatischen Ringstruktur ist diesbezüglich mit Parallelen zum anaeroben Naphthalinabbau zu rechnen. Auf der Suche nach Spezialisten des Phenanthrenabbaus wurden Bakterien aus einem natürlichen Teersee in der Karibik isoliert und unter anoxischen Bedingungen angereichert. Diese Anreicherungskultur wächst auf Phenanthren als einziger Kohlenstoff- und Energiequelle und unter sulfatreduzierenden Bedingungen. Anhand dieser Kultur konnten weitere – über die initialisierende Carboxylierung hinausgehender Schritte – des anaerobe Kohlenwasserstoffabbaus für Phenanthren untersucht werden. Dabei wurde mittels einer Metabolitenanalyse die Carboxylierung in der C2-Position bestimmt. Die anschließende Ligase-Reaktion konnte in einem Enzym-Assay nachgewiesen werden, wobei nur 2-Phenanthroesäure zu 2-Phenanthroyl-CoA umgesetzt wurde, was die Bestimmung der C2-Position ebenfalls bestätigt. Bis dahin verläuft der Abbau, parallel zum Naphthalin-Abbau, auch hier an der C2-Position. Die weitergehenden Schritte des Abbauprozesses konnte mit Enzym-Assays bislang nicht beschrieben werden. Metabolitenuntersuchungen ergaben jedoch erste Indizien für eine sukzessive Reduktion, ebenfalls vergleichbar mit dem Abbau des Naphthalins (Eberlein, Estelmann, et al. 2013; Eberlein, Johannes, et al. 2013). Mittels Durchflusszytometrie nach einer FISH-Analyse konnte eine Bakterienspezies, in Übereinstimmung mit 60% der Sequenzdaten, als das in der untersuchten Kultur vorherrschenden Bakterium bestimmt werden. Diesem Bakterium wird auch kausal der Phenanthren-Abbau zugeschrieben. Das Bakterium gehört zur Familie der Desulfobacteraceae und zeigte einen mit 93% hohe Verwandtschaftsgrad zu dem bekannten sulfatreduzierendem Naphthalin-Abbauer NaphS2, was auf ähnliche Gen-Cluster oder vergleichbare Abbaumechanismen schließen lässt. Auf der Suche nach Leben in dem natürlich entstehenden Asphalt fanden Meckenstock et al. (2014) außerdem kleine Wassertröpfchen im Teer, in denen wiederum lebende Bakterien entdeckt wurden. Durch weitere Isolierung und Sequenzierung, vor allem mittels Einzelzellanalyse, erwartete man neue Einsichten in eine gerichtete Evolution innerhalb der Tröpfchen, da diese als individuelle Ökosysteme fungieren. Zusammen mit den DNA-Sequenzen, die direkt aus dem Teer isoliert wurden, ergab sich ein Überblick über die Diversität innerhalb des Asphaltsees: Die Bakterien zeigten im Vergleich mit den Tröpfchen eine deutlich höhere Artenvielfalt. Dies lässt auf besser spezialisierte Bakterien in den Tröpfchen sowie auf eine breitere Verteilung von Bakterien im Teer schließen, die sich darin schneller und flexibler an sich ändernde Bedingungen anpassen können. Im Vergleich zu anderen Öl-Proben ergab sich jedoch nur eine sehr geringe Diversität im Teer; und etwa 30% der Bakterien konnten keiner Art zugewiesen werden. In Proben marinen Ursprungs liegt dieser Anteil bei etwa 10%. Mit über 50% Anteil war Tepidiphilus sp. das dominierende Bakterium in den Wassertröpfchen. Es gehört zur Familie der Hydrogenophilaceae innerhalb der β-Proteobakterien. Tepidiphilus sp. ist in der Lage, organische Säuren als einzige Kohlenstoffquelle unter nitrifizierenden Bedingungen abzubauen. Damit ist es ein plausibler Kandidat, um perfekt angepasst innerhalb des „Ökosystems Wassertropfen“ zu existieren. Eine weitere Charakterisierung dieses Bakteriums ist bislang nicht erfolgt. Mit dieser Arbeit konnten Fortschritte in der Aufklärung des anaeroben Phenanthrenabbaus beschrieben werden, die weitere Rückschlüsse auf generelle Abbauwege von hochmolekularen polyzyklischen aromatischen Kohlenwasserstoffen ermöglichen. Zudem wurden die Diversitäten von Bakterien, die in einem extremen Habitat wie dem Teersee überleben können, durch verschiedene Sequenzierungs- und Auswertungsmethoden differenzierter beleuchtet, um einen tieferen Einblick in das „Leben im Öl“ zu ermöglichen.

This thesis addresses fundamental questions towards the specific involvement of soil microbiota in plant-associated soil food webs. Bacterial micropredators and niche differentiation to protozoa were characterised in a maize soil. Specific root-associated bacterial populations were shown to be preferentially mobilised with seepage water after extreme precipitation events. These findings allow for an advanced understanding of links between carbon pools and microbial communities in different soil compartments. Diese Doktorarbeit beschäftigt sich mit grundlegenden Fragen zur Beteiligung von spezifischen Bodenmikrobiota in pflanzenassoziierten Nahrungsnetzen. Bakterielle Mikropräparatoren und die Nischendifferenzierung zu den Protozoen wurden für einen Maisfeldboden untersucht. Es wurde gezeigt, dass spezifische wurzelassoziierte Mikroben nach Starkregenereignissen selektiv mit dem Sickerwasser mobilisiert werden. Diese Ergebnisse erlauben ein verbessertes Verständnis der Verbindungen zwischen Kohlenstoffpools und mikrobiellen Gemeinschaften in Bodenkompartimenten.

The dissertation shows that Pelotomaculum candidate BPL is the first member of the genus that can perform full dissimilatory sulfate reduction, besides its potential to degrade benzene. Results from physiological, genomic, and proteomic analyses are presented indicating that environmental Spirochaetes recycle detrital biomass and convert it into end products or electrons that support growth of associated degraders. Lastly, a novel reverse stable isotope labelling analysis is developed as a sensitive means to monitor biodegradation. Die Dissertation zeigt, dass Pelotomaculum cand. BPL als erstes Mitglied dieser Gattung sowohl Benzol abbauen als auch Sulfat vollständig reduzieren kann. Ergebnisse von physiologischen, genomischen und proteomischen Analysen machen deutlich, dass Umwelt-Spirochäten tote Biomasse wiederverwerten. Dabei entstehen Endprodukte oder Elektronen, die das Wachstum von nahestehenden Bakterien unterstützen. Schließlich wird eine neue stabile Isotopen-Methode entwickelt, mit der man sehr niedrige Mineralisierungsraten von Mikroorganismen messen kann.

Microbial transformation of n-alkanes in anaerobic ecosystems plays a pivotal role in biogeochemical carbon cycling and bioremediation, but the requisite genetic machinery is not well elucidated.Desulfatibacillum alkenivorans AK-01 utilizes n-alkanes (C13 to C18) and contains two genomic loci encoding alkylsuccinate synthase (ASS) gene clusters. ASS catalyzes alkane addition to fumarate to form methylalkylsuccinic acids. We hypothesized that the genes in the two clusters would be differentially expressed depending on the alkane substrate utilized for growth. RT-qPCR was used to investigate ass-gene expression across AK-01's known substrate range, and microarray-based transcriptomic analysis served to investigate whole-cell responses to growth on n-hexadecane versus hexadecanoate. RT-qPCR revealed induction of ass gene cluster 1 during growth on all tested alkane substrates, and the transcriptional start sites in cluster 1 were determined via 5'RACE. Induction of ass gene cluster 2 was not observed under the tested conditions. Transcriptomic analysis indicated that the upregulation of genes potentially involved in methylalkylsuccinate metabolism, including methylmalonyl-CoA mutase and a putative carboxyl transferase. These findings provide new directions for studying the transcriptional regulation of genes involved in alkane addition to fumarate, fumarate recycling and the processing of methylalkylsuccinates with regard to isolates, enrichment cultures and ecological datasets.