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Tissue optical clearing (TOC) and fluorescence imaging have significantly advanced 3D imaging of biological samples. However, few studies have applied these techniques in infection biology although they provide valuable information that can enhance our understanding of spatiotemporal dynamics in infections and disease progression. This thesis addresses this gap by utilizing these techniques to analyze optically cleared tissue samples from animals experimentally infected with rabies virus (RABV) or severe acute respiratory syndrome corona virus 2 (SARS-CoV-2), in order to gain deeper insights into virus replication and pathogenicity in the brain and lung. To achieve this, image segmentation and quantification pipelines, including the use of AI, for the extraction of comparable data on cell tropism and the spatiotemporal development of infection processes were developed. Previous work demonstrated that intramuscular (i.m) inoculation of mice with highly virulent field RABV strains led to the infection of both neurons and astrocytes in the brain, of which the latter may significantly shape the host response to RABV infection. Accordingly, the first aim was to investigate whether astrocyte tropism of field viruses is a late phase phenotype or whether it already occurs in early phases before the onset of clinical signs. Immunofluorescence imaging of optically cleared mouse brain sections confirmed astrocyte infection at early time points of brain invasion and indicates that astrocyte tropism of RABV indeed could shape host reactions to infection already at those early time points of infection. The second aim was to investigate whether resistance of rRABV DogA to interferon (IFN) response, mediated by the phosphoprotein (P), is a prerequisite for CNS infection from peripheral sites and astrocyte tropism in the brain. Using a recombinant virus mutant, rRABV DogA-IFNmut, which has impaired type I interferon antagonistic activity, age-dependent courses of brain infection following i.m. inoculation were observed. In 4-week-old mice, the infection proved lethal, while older mice recovered despite showing clinical signs. While these data demonstrated insufficient block of the virus by the interferon system in peripheral tissues, they also suggest a significant role of the type I interferon response in restricting virus spread and astrocyte tropism in the brain. The third aim was to explore the effect of increased glycoprotein (G) cell surface expression that was previously observed in cell culture on the virulence of field RABV DogA. Based on the hypothesis that de-regulated G transport to non-synaptic plasma membrane sites results in the release of extracellular virus from neurons and hence recognition by the immune system, the mutant rRABV DogA-NTN, that has three cell culture-adaptive mutations in G was used. Although delayed pathogenesis and requirement of higher viral loads to induce clinical signs were observed, no detectable increase in surface expression of the protein was found in neurons, despite visualized changes in intra-neuronal G distribution. These results indicate that differences in surface transport regulation between standard cell lines and neurons in vivo exist. Involved mechanisms, however, remain unclear.
Finally, to effectively manage SARS CoV-2 infection, a thorough understanding of the immune dynamics within the lungs is needed. Consequently, TOC combined with 3D imaging techniques was used to investigate viral infection and the corresponding immune response in the lungs of experimentally infected hamsters over a 7-day period. This approach enabled high-resolution visualization of cellular interactions, revealing a spatially localized antiviral response. Infected epithelial cells displayed an upregulation of major histocompatibility complex II (MHC II), highlighting immune system activation. Monocyte-derived macrophages played a crucial role in viral clearance; however, their presence was also associated with postinfection endothelial damage, illustrating their dual role in both defending against the virus and contributing to tissue injury. Furthermore, early signs of tissue repair were observed alongside inflammatory and necrotizing processes, potentially laying the groundwork for longterm alterations in lung structure and function. These findings provide valuable insights into the complexities of immune responses in viral infections and may inform future therapeutic strategies.
The rewetting of formerly drained peatlands can help to counteract climate change through the reduction of CO2 emissions. However, this can lead to resuming CH4 emissions due to changes in the microbiome, favoring CH4-producing archaea. How plants, hydrology and microbiomes interact as ultimate determinants of CH4 dynamics is still poorly understood. Using a mesocosm approach, we studied peat microbiomes, below-ground root biomass and CH4 fluxes with three different water level regimes (stable high, stable low and fluctuating) and four different plant communities (bare peat, Carex rostrata, Juncus inflexus and their mixture) over the course of one growing season. A significant difference in microbiome composition was found between mesocosms with and without plants, while the difference between plant species identity or water regimes was rather weak. A significant difference was also found between the upper and lower peat, with the difference increasing as plants grew. By the end of the growing season, the methanogen relative abundance was higher in the sub-soil layer, as well as in the bare peat and C. rostrata pots, as compared to J. inflexus or mixture pots. This was inversely linked to the larger root area of J. inflexus. The root area also negatively correlated with CH4 fluxes which positively correlated with the relative abundance of methanogens. Despite the absence or low abundance of methanotrophs in many samples, the integration of methanotroph abundance improved the quality of the correlation with CH4 fluxes, and methanogens and methanotrophs together determined CH4 fluxes in a structural equation model. However, water regime showed no significant impact on plant roots and methanogens, and consequently, on CH4 fluxes. This study showed that plant roots determined the microbiome composition and, in particular, the relative abundance of methanogens and methanotrophs, which, in interaction, drove the CH4 fluxes.
Charakterisierung von Membranvesikeln aus Gram-negativen und Gram-positiven pathogenen Bakterien
(2024)
Bakterielle Pathogene können in ihren Wirten Infektionen auslösen, die gesundheitsgefährdend sind oder sogar zum Tod führen können. Zu diesen Pathogenen zählen auch das Gram-negative Bakterium Aeromonas salmonicida (A. salmonicida) und die Gram-positiven Bakterien Renibacterium salmoninarum (R. salmoninarum) und Streptococcus pneumoniae (S. pneumoniae). Während R. salmoninarum und A. salmonicida Fischpathogene sind, die in Salmoniden die bakterielle Nierenkrankheit (bacterial kidney disease) (BKD) (R. salmoninarum) bzw. die Fischfurunkulose (A. salmonicida) auslösen können, ist S. pneumoniae ein humaner Pathogen, der den oberen Atemwegstrakt und andere Gewebe kolonisieren und Krankheiten wie Sepsis, Meningitidis und Pneumonie auslösen kann. Gegen alle diese Pathogene existieren Vakzin-Strategien bestehend aus Kapselpolysacchariden oder inaktivierten Bakterien, um einer Infektion und deren Folgen vorzubeugen. Allerdings wirken diese Vakzine oft nur gegen bestimmte Serotypen (S. pneumoniae) oder sind mit hohem Stress und Nebenwirkungen für den Wirt verbunden (A. salmonicida, R. salmoninarum). Aus diesem Grund wird aktiv an alternativen Strategien zur Vakzinierung geforscht. Eine mögliche Route sind bakterielle Membranvesikel (MV), die sich von der bakteriellen Oberfläche Gram-positiver und Gram-negativer Bakterien abschnüren und viele potenziell immunogene Proteine auf ihrer Oberfläche tragen.
In dieser Arbeit wurde das Proteinrepertoire der bakteriellen MV von R. salmoninarum, A. salmonicida und S. pneumoniae mittels Massenspektrometrie untersucht. In der aufgereinigten MV Fraktion von R. salmoninarum wurden die immunsupprimierenden Proteine P57/Msa und P22, die wichtige Virulenzfaktoren von R. salmoninarum sind, in hoher Abundanz detektiert. Weiterhin wurden neues Lipoprotein C/Protein von 60 kDa (NlpC/P60) Hydrolasen und weitere Zellwand-modifizierende Proteine in der MV Fraktion gefunden, was einen Hinweis darauf darstellen könnte, dass diese Proteinklasse eine Relevanz bei der Entstehung von MV in Gram-positiven Bakterien hat. Bei der Untersuchung der MV von A. salmonicida wurde analysiert, inwieweit sich das MV Proteinrepertoire durch Kultivierungsbedingungen beeinflussen lässt. Die MV, die unter Bedingungen mit geringer Eisenverfügbarkeit aufgereinigt wurden, hatten eine ähnliche Größe und Konzentration im Vergleich zu der Kontrollbedingung. Allerdings wurden zahlreiche Proteine in der MV Fraktion detektiert, die bei geringer Eisenverfügbarkeit signifikant in ihrer Abundanz erhöht waren. Hierzu zählten vor allem TonB-abhängige Eisen- und Siderophoretransporter, aber auch Hemolysin und Lipasen. Eine erhöhte Kultivierungstemperatur resultierte in einer geringeren Vesikelkonzentration verglichen mit der Kontrollbedingung. Allerdings führte die erhöhte Kultivierungstemperatur zu einer signifikant gesteigerten Hemolysin Abundanz. Bei der Kultivierung von A. salmonicida mit dem Antibiotika Florfenicol waren die MV in ihrer Größe deutlich verringert. Weiterhin wurden viele ribosomale Proteine in der MV Fraktion gefunden, was auf bakterielle Lyse hinweisen könnte. Der Vergleich zwischen dem Erntezeitpunkt der MV in der stationären Wachstumsphase und der Sterbephase von S. pneumoniae zeigte, dass Vesikel aus der Sterbephase im Durchschnitt leicht vergrößert und in der Konzentration 10-fach erhöht waren. Zusätzlich war Autolysin in den MV, die in der Sterbephase geerntet wurden, signifikant in der Abundanz erhöht.
Zusammenfassend konnte gezeigt werden, dass sich das Proteinrepertoire der MV durch Faktoren wie die Wachstumsphase oder Kultivierungsbedingungen drastisch beeinflussen lässt. Es wurden putativ immunogene Proteine (Eisentransporter, Lipoproteine) in den MV aller untersuchten Pathogenen gefunden, was das Potenzial der MV als Vakzin-Plattform zeigt. Besonders eine Kultivierung bei Bedingungen mit geringer Eisenverfügbarkeit könnte durch die hohe Anzahl an Eisen-regulierten Membranproteinen bei einer Vakzinentwicklung von Vorteil sein.
Investigating protective immune responses against SARS-CoV-2 infections in small animal models
(2025)
During the SARS-CoV-2 pandemic, different viral variants emerged with distinct
transmissibility, pathogenicity, and immune evasion features. Although these traits
were reflected in the pandemic course, knowledge about immunity against the
emerging variants remained limited and lagged behind. Information about early
antiviral immunity mainly came from blood or tissue samples of individuals who died
from COVID-19, which cannot fully reflect local protective immune responses during
acute SARS-CoV-2 infections. This thesis utilized small animals to model COVID-19 in
humans and used it afterward to test mRNA vaccine efficacy. The aim was to gain
insights into early immune responses after SARS-CoV-2 infection and mRNA-induced
protective immunity in various tissues, which is challenging to accomplish in humans.
Publication I demonstrates that infection of K18-hACE2 mice with ancestral SARS-CoV-
2, Beta, or Delta VOC results in unique patterns of viral dissemination into the lungs
and distinct inflammatory responses within the first 5 days post-infection (DPI). The
Beta variant spread to the lungs earlier and caused higher levels of inflammatory
cytokines and more significant infiltration of innate immune cells than the ancestral
SARS-CoV-2. Adaptive immune responses were detected within 7 DPI and were
associated with lower viral burden, suggesting a potential role in clearing the infection.
The depletion of B and T cells allowed correlating viral clearance with virus-specific
antibody levels and T cell responses. While innate immune responses differed among
SARS-CoV-2 variants, adaptive responses developed similarly, suggesting a potential
target for pan-variant countermeasures. Publication II indicates that low-dose mixed
mRNA vaccines, containing half the amount of each monovalent vaccine, produce
similar levels of neutralizing antibodies and virus-specific T cells in the lungs, protecting
K18-hACE2 mice from lethal disease. The vaccination of Wistar rats showed that
bivalent vaccination can elicit neutralizing antibodies against different variants (e.g.,
BA1 and BA.5) that are not included in the vaccine formulation. The study suggests that
alternative vaccination strategies could generate cross-protective immunity, helping to
overcome immune evasion by SARS-CoV-2 variants. Publication III investigates
protective immunity against SARS-CoV-2 upon mRNA vaccination in a mouse model
with restricted B cell responses, commonly seen in vaccinated individuals with
immunodeficiencies. It demonstrates that T cells can compensate for the lack of serum
antibodies to prevent sublethal disease in mice. Serum antibody levels correlated with
reduced disease severity and mRNA vaccine-induced IgG limited viral replication in the
nasal conchae, indicating a potential role in protecting from virus transmission.
This thesis provides valuable insights into protective immune responses against
infection with various SARS-CoV-2 variants and upon mRNA vaccination and informs
about vaccination strategies to overcome immune escape. The numerous
experimental studies within this thesis are essential for characterizing emerging viral
variants and developing effective vaccines. These findings offer valuable insights that
can guide the development of effective countermeasures to combat immune evasion
by future SARS-CoV-2 variants.
Influence of plasma-treated air on surface microbial communities on freshly harvested lettuce
(2023)
Plant-based foods like lettuce are an important part of the human diet and worldwide industry. On a global scale, the number of food-associated illnesses increased in the last decades. Conventional lettuce sanitation methods include cleaning either with tap or chloritized water. Beside these water-consuming strategies, physical plasma is an innovative and effective possibility for food sanitation. Recent studies with plasma-treated water showed an effective reduction of the microbial load. Plasma-processed air (PPA) is another great opportunity to reduce the microbial load and save water. To test the efficiency of PPA, the surface microbiome of treated lettuce was analyzed via proliferation assays with special agars, live/dead assays and tests for respiratory activity of the microorganisms. PPA showed a reduction of the colony forming units (CFU/mL) on all tested microbial groups (Gram-negative and Gram-positive bacteria, yeasts and molds). These results were supported by the live/dead assay. For further insights, the PPA-ingredients were detected with Fourier Transformation Infrared Spectroscopy (FTIR), which revealed NO2, NO and N2O5 as the main reactive species in the PPA. In the future, PPA could be an outstanding, on-demand sanitation step for higher food safety standards, especially in situations where humidity and high temperature should be avoided.
With 2.56 million deaths worldwide annually, pneumonia is one of the leading causes of death. The most frequent causative pathogens are Streptococcus pneumoniae and influenza A virus. Lately, the interaction between the pathogens, the host, and its microbiome have gained more attention. The microbiome is known to promote the immune response toward pathogens; however, our knowledge on how infections affect the microbiome is still scarce. Here, the impact of colonization and infection with S. pneumoniae and influenza A virus on the structure and function of the respiratory and gastrointestinal microbiomes of mice was investigated. Using a meta-omics approach, we identified specific differences between the bacterial and viral infection. Pneumococcal colonization had minor effects on the taxonomic composition of the respiratory microbiome, while acute infections caused decreased microbial complexity. In contrast, richness was unaffected following H1N1 infection. Within the gastrointestinal microbiome, we found exclusive changes in structure and function, depending on the pathogen. While pneumococcal colonization had no effects on taxonomic composition of the gastrointestinal microbiome, increased abundance of Akkermansiaceae and Spirochaetaceae as well as decreased amounts of Clostridiaceae were exclusively found during invasive S. pneumoniae infection. The presence of Staphylococcaceae was specific for viral pneumonia. Investigation of the intestinal microbiomés functional composition revealed reduced expression of flagellin and rubrerythrin and increased levels of ATPase during pneumococcal infection, while increased amounts of acetyl coenzyme A (acetyl-CoA) acetyltransferase and enoyl-CoA transferase were unique after H1N1 infection. In conclusion, identification of specific taxonomic and functional profiles of the respiratory and gastrointestinal microbiome allowed the discrimination between bacterial and viral pneumonia.
The Lobaria pulmonaria holobiont comprises algal, fungal, cyanobacterial and bacterial components. We investigated L. pulmonaria's bacterial microbiome in the adaptation of this ecologically sensitive lichen species to diverse climatic conditions. Our central hypothesis posited that microbiome composition and functionality aligns with subcontinental‐scale (a stretch of ~1100 km) climatic parameters related to temperature and precipitation. We also tested the impact of short‐term weather dynamics, sampling season and algal/fungal genotypes on microbiome variation. Metaproteomics provided insights into compositional and functional changes within the microbiome. Climatic variables explained 41.64% of microbiome variation, surpassing the combined influence of local weather and sampling season at 31.63%. Notably, annual mean temperature and temperature seasonality emerged as significant climatic drivers. Microbiome composition correlated with algal, not fungal genotype, suggesting similar environmental recruitment for the algal partner and microbiome. Differential abundance analyses revealed distinct protein compositions in Sub‐Atlantic Lowland and Alpine regions, indicating differential microbiome responses to contrasting environmental/climatic conditions. Proteins involved in oxidative and cellular stress were notably different. Our findings highlight microbiome plasticity in adapting to stable climates, with limited responsiveness to short‐term fluctuations, offering new insights into climate adaptation in lichen symbiosis.
Studying the fates of oil components and their interactions with ecological systems is essential for developing comprehensive management strategies and enhancing restoration following oil spill incidents. The potential expansion of Kazakhstan’s role in the global oil market necessitates the existence of land-specific studies that contribute to the field of bioremediation. In this study, a set of experiments was designed to assess the growth and biodegradation capacities of eight fungal strains sourced from Kazakhstan soil when exposed to the hydrocarbon substrates from which they were initially isolated. The strains were identified as Aspergillus sp. SBUG-M1743, Penicillium javanicum SBUG-M1744, SBUG-M1770, Trichoderma harzianum SBUG-M1750 and Fusarium oxysporum SBUG-1746, SBUG-M1748, SBUG-M1768 and SBUG-M1769 using the internal transcribed spacer (ITS) region. Furthermore, microscopic and macroscopic evaluations agreed with the sequence-based identification. Aspergillus sp. SBUG-M1743 and P. javanicum SBUG-M1744 displayed remarkable biodegradation capabilities in the presence of tetradecane with up to a 9-fold biomass increase in the static cultures. T. harzianum SBUG-M1750 exhibited poor growth, which was a consequence of its low efficiency of tetradecane degradation. Monocarboxylic acids were the main degradation products by SBUG-M1743, SBUG-M1744, SBUG-M1750, and SBUG-M1770 indicating the monoterminal degradation pathway through β-oxidation, while the additional detection of dicarboxylic acid in SBUG-M1768 and SBUG-M1769 cultures was indicative of the fungus’ ability to undertake both monoterminal and diterminal degradation pathways. F. oxysporum SBUG-M1746 and SBUG-M1748 in the presence of cyclohexanone showed a doubling of the biomass with the ability to degrade the substrate almost completely in shake cultures. F. oxysporum SBUG-M1746 was also able to degrade cyclohexane completely and excreted all possible metabolites of the degradation pathway. Understanding the degradation potential of these fungal isolates to different hydrocarbon substrates will help in developing effective bioremediation strategies tailored to local conditions.
Marine particle microbiomes during a spring diatom bloom contain active sulfate-reducing bacteria
(2024)
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.
For centuries, peatlands have received little attention or recognition due to their perceived insignificance, invisibility, and negative associations. However, they are now receiving increasing attention due to the ecosystem services they provide as carbon and water reservoirs, as well as their rich biodiversity of rare and endangered animals and plants. In the discussion of greenhouse gas (GHG) emissions and their impact on the climate, peatlands are also recognized as being of significant importance. To comprehend the source of these GHG, it is necessary to examine the microbiome of peatlands and the effects of lowering (draining) and raising (rewetting) the water level. Peatlands store large amounts of soil organic carbon (SOC) in form of peat. Dead plant material is preserved in the water-saturated soils and the carbon dioxide (CO2) absorbed from the atmosphere through photosynthesis is stored as peat. However, when the water table in peatlands is lowered, atmospheric oxygen enters the soil, and the soil microbiome changes, leading to the microbial decomposition of SOC and the release of large quantities of CO2. Rewetting can halt aerobic decomposition processes by blocking oxygen. Anaerobic conditions are not conducive to aerobic organisms. Complex plant polymers, such as lignin, can no longer be broken down, preserving the peat body and allowing the peatland to once again serve as a long-term carbon sink. Under these anaerobic conditions, methanogenic archaea produce methane from metabolic products of fermentation processes. Methane is emitted in much smaller quantities, but has about 30 times the warming potential of CO2, making it the second most important GHG after CO2. To gain a better understanding of the processes involved in methanogenesis, it is essential to conduct a detailed investigation of the individual groups of archaea involved. The metabolic pathways of hydrogenotrophic methanogenesis from CO2 and hydrogen, as well as acetoclastic methanogenesis from acetate cleavage, have already been extensively researched and are considered the most significant pathways of methanogenesis. In recent years, additional organisms have been discovered that carry out methylotrophic methanogenesis from methanol and methylated compounds. The methylotrophic order of Methanomassiliicoccales was first described in 2012, but only one pure culture, Methanomassiliicoccus luminyensis, has been isolated. Further research into Methanomassiliicoccales, commonly found in mires, can enhance our understanding of methanogenesis not only in mires but also in other habitats, such as the gastrointestinal tracts (GIT) of animals or water body sediments. This study investigates the peat soil microbiomes, with a focus on methanogenic Archaea of the order Methanomassiliicoccales, at the six study sites of the WETSCAPES project. The sites are located in the three important fen types of Mecklenburg-Western Pomerania (alder forest, percolation fen, and coastal flooded fen), each in drained and rewetted areas. The first article investigates the microbial composition of prokaryotes and eukaryotes in the sites using Illumina MiSeq Amplicon sequencing of 16S rRNA and 18S rRNA genes. The aerobic microorganisms responsible for decomposing plant biomass are present in drained peatlands. In rewetted sites, the relative abundance of these organisms was significantly lower, while other groups, such as fermenting bacteria and methanogenic archaea, occurred in greater numbers. Quantitative polymerase chain reaction (qPCR) was used to determine the number of methanogenic archaea per gram soil. The results indicate that methanogens are at least 10 times more abundant in rewetted peatlands than in drained sites, suggesting increased methanogenesis in rewetted peatlands. These findings were confirmed by GHG measurements. The second article presents an overview of the interactions between water transport, soil chemistry, primary production, peat formation, material conversion and transport, microorganisms, and GHG exchange. The findings are based on state-of-the-art methods in the relevant research areas and combine initial results from all project partners of the WETSCAPES project. The third article focuses on an application-oriented approach and tests different rewetting strategies. The objective of this study was to decrease methane emissions by removing the topsoil and additional planting of peat mosses. The results of the study, which included measurements of GHG emissions and qPCR analyses of methanogenic archaea, showed a significant reduction in methane emissions after the top 30 cm of soil were removed compared to the untreated area. The fourth article deals with the methylotrophic order Methanomassiliicoccales, which belongs to the group of methanogenic archaea. Over a period of 2.5 years, at 12 time points, at three soil depths, 16S rRNA gene sequencing was used to produce a seasonal and spatial analysis of the Methanomassiliicoccales phylotypes present. The results revealed a distribution of the phylotypes based on soil depth, redox potential, and season, which were related to their physiological characteristics. In the fifth article, the 1.52 Mbp genome of "Ca. Methanogranum gryphiswaldense U3.2.1", a Methanomassiliicoccales strain, enriched to > 80% from peat soil of the rewetted percolation fen, was presented. The sixth article presented the microscopic, physiological, and genomic studies of Ca. Methanogranum gryphiswaldense U3.2.1. This hydrogen-dependent, methylotrophic strain from the family Methanomethylophilaceae, uses methanol and trimethylamine as electron acceptors. Despite various attempts, a pure culture has not yet been achieved. The study demonstrates the impact of rewetting on the soil microbiome, revealing its complexity. The development of the microbiome was influenced by various biotic and abiotic factors, depending on the location and water level. The abundance of methanogenic archaea was significantly higher in the rewetted sites compared to the drained ones. The Methanomassiliicoccales were found to be a significant contributor to the methanogenic archaea in the investigated sites. It remains to be examinated whether they also significantly contribute to methane formation. Enrichment cultures and genome analyses were used to investigate the substrate spectrum of Ca. Methanogranum gryphiswaldense U3.2.1 obtained from peat soil. The extraction of a pure culture from peat soils has not yet been successful and therefore remains a goal of future investigations.