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Titel:Investigation of microbial groups involved in the uptake of atmospheric trace gases in upland soils
Autor:Pratscher, Jennifer
Weitere Beteiligte: Conrad, Ralf (Prof. Dr.)
Veröffentlicht:2011
URI:https://archiv.ub.uni-marburg.de/diss/z2011/0062
URN: urn:nbn:de:hebis:04-z2011-00625
DOI: https://doi.org/10.17192/z2011.0062
DDC: Biowissenschaften, Biologie
Titel(trans.):Untersuchung mikrobieller Gruppen beteiligt an der Aufnahme von atmosphärischen Spurengasen in upland soils
Publikationsdatum:2011-02-18
Lizenz:https://rightsstatements.org/vocab/InC-NC/1.0/

Dokument

Schlagwörter:
Stable isotope probing, Methanstoffwechsel, NiFe-Hydrogenase, Ammonium-Monooxygenase, Methan-Monooxygenase, Thaumarchaeota, Methane-monooxygenase, Stabile Isotopenbeprobung, Ammonium-monooxygenase, CARD-FISH, CARD-FISH, NiFe-Hydrogenase

Summary:
Atmospheric trace gases play a leading role in the changes occurring in the atmosphere at present, including climate change. A significant part of atmospheric trace gas fluxes occurs at the interface between atmosphere and upland soils. Unfortunately, the microorganisms in charge of these dynamics are not fully understood. This thesis therefore focuses on the investigation of microbial groups in terrestrial environments, responsible for or proposed to be involved in the uptake of atmospheric trace gases (CH4, CO2, H2), namely the potential atmospheric methane oxidizer upland soil cluster α (USCα), the autotrophic ammonia oxidizing archaea (AOA), and the hydrogen oxidizing streptomycetes. Several methods were tested to investigate the incorporation of labeled substrate and to monitor the expression of their functional marker genes, pmoA for the high-affinity particulate methane monooxygenase of USCα, amoA for the ammonia monooxygenase of AOA, and hydB for the high-affinity [NiFe]-hydrogenase of Streptomyces sp. PCB7. Although the upland soil cluster α (USCα) in forest soils is assumed to represent methanotrophic bacteria adapted to the trace level of atmospheric methane and to play an essential part in the removal of this greenhouse gas from the atmosphere, so far it is unclear whether these microorganisms are able to obtain all their energy and carbon solely from CH4 or use additional carbon compounds. Stable isotope probing was applied to investigate incorporation of labeled CH4 and acetate into nucleic acids of USCα. The results of this study indicate that USCα might only use atmospheric CH4 as an additional energy source or survival strategy, but utilizes additional carbon compounds, such as acetate, for growth suggesting the USCα represents rather facultative than obligate methanotrophs. Furthermore, CARD-FISH of pmoA transcripts visualized USCα in situ for the first time. These findings promote the knowledge and understanding of upland soils as a sink for atmospheric methane and the microorganisms proposed to be responsible for this process. While for a long time autotrophic bacteria were believed to be solely responsible for the process of ammonia oxidation, there is now increasing evidence that also Archaea are involved. But to date it remained elusive whether ammonia oxidizing archaea in soil can assimilate CO2 and to what extent they are functionally active. Stable isotope probing of nucleic acids using 13CO2 showed that ammonia oxidizing archaea were actively involved in microbial ammonia oxidation in an agricultural soil and did fix CO2 autotrophically, presumably via the hydroxypropionate-hydroxybutyrate cycle. CARD-FISH further demonstrated the numerical importance of the archaeal ammonia oxidizers to the overall archaeal community in this environment. These results give novel evidence that the contribution of nitrifying Archaea to ammonia oxidation and CO2 fixation in terrestrial environments might be substantial. Although hydrogen is considered to be one of the most important future energy carriers, little is known about the global biogeochemical cycle of this trace gas. Previous findings indicate that microorganisms rather than free soil enzymes are responsible for the uptake of atmospheric H2 in soils. In this thesis, CARD-FISH analyses demonstrated that streptomyces spores instead of the mycelia expressed the high-affinity H2 uptake activity. This suggests that H2-oxidizing streptomycetes, or actinobacteria in general, are essential for the uptake of atmospheric H2 in upland soils.

Zusammenfassung:
Atmosphärischen Spurengasen kommt eine Führungsrolle in den derzeitigen klimatischen Veränderungen zu. Ein signifikanter Anteil am Austausch von atmosphärischen Spurengasen findet an der Schnittstelle zwischen Atmosphäre und „upland soils“ statt. Allerdings sind die für diese Dynamiken verantwortlichen Mikroorganismen nicht vollständig identifiziert und erforscht. Diese Arbeit befasst sich deshalb mit der Untersuchung mikrobieller Gruppen in terrestrischen Habitaten, die in die Aufnahme von atmosphärischen Spurengasen (CH4, CO2, H2) involviert sind, und zwar das potentiell atmosphärisches Methan oxidierende „upland soil cluster α“ (USCα), die autotrophen Ammonium-oxidierenden Archaeen (AOA) und die Wasserstoff-oxidierenden Streptomyzeten. Verschiedene Methoden wurden angewandt, um die Aufnahme von markiertem Substrat in diese mikrobiellen Gruppen zu untersuchen und die Expression ihrer funktionellen Marker-Gene zu analysieren. Dabei handelte es sich um pmoA für die hoch-affine membran-gebundene (partikuläre) Methan-Monooxygenase von USCα, amoA für die Ammonium-Monooxygenase der AOA und hydB für die hoch-affine [NiFe]-Hydrogenase von Streptomyces sp. PCB7. Trotz der Annahme, dass es sich bei dem “upland soil cluster α“ (USCα) in Waldböden um methanotrophe Bakterien handelt, die an atmosphärische Methan-konzentrationen angepasst sind und denen demnach eine grundlegende Rolle in der Aufnahme und Beseitigung dieses Treibhausgases aus der Atmosphäre zuteil wird, war noch ungeklärt, ob diese Mikroorganismen ihren gesamten Energie- und Kohlenstoffbedarf mit Methan decken können oder zusätzlich auf andere Kohlenstoffquellen angewiesen sind. Stabile Isotopenbeprobung wurde angewandt, um den Einbau von markiertem CH4 und Acetat in Nukleinsäuren von USCα zu untersuchen. Die Ergebnisse dieser Studie weisen darauf hin, dass USCα atmosphärisches CH4 möglicherweise nur als zusätzliche Energiequelle oder Überlebensstrategie nutzt, und stattdessen andere Kohlenstoffverbindungen, z.B. Acetat, zum Wachstum verwendet. Somit repräsentiert USCα eher fakultative als obligate Methanotrophe. Die Anwendung von CARD-FISH, spezifisch für pmoA Transkripte, ermöglichte zudem die erste Visualisierung von USCα in situ. Diese Resultate erweitern unseren Wissensstand und das Verständnis in Bezug auf „upland soils“ als Senke für atmosphärisches Methan und die Mikroorganismen, die für diese Prozesse verantwortlich sind. Autotrophe Bakterien wurden lange Zeit alleinverantwortlich für die Ammonium-Oxidation gehalten. Doch inzwischen liegen zunehmend Studien vor, die auf eine zusätzliche Beteiligung von Archaea an diesem Prozess hinweisen. Allerdings war bis heute unbekannt, ob Ammonium-oxidierende Archaeen im Boden CO2 assimilieren können und zu welchem Grad sie funktionell aktiv sind. Stabile Isotopenbeprobung von Nukleinsäuren unter Verwendung von 13CO2 demonstrierte eine aktive Beteiligung der Ammonium-oxidierenden Archaeen an der mikrobiellen Ammonium-Oxidation in einem Feldboden, verbunden mit autotropher CO2-Fixierung, vermutlich über den Hydroxypropionat-Hydroxybutyrat-Zyklus. CARD-FISH zeigte weiterhin den hohen Anteil und damit die große Bedeutung der archaeellen Ammonium-Oxidierer in der gesamten archaeellen Gemeinschaft in dieser Umgebung. Diese Resultate liefern neue Belege für die substanzielle Beteilung der nitrifizierenden Archaea an der Ammonium-Oxidation und CO2-Fixierung in terrestrischen Habitaten. Obwohl Wasserstoff als einer der wichtigsten Energieträger der Zukunft gilt, ist der globale biochemische Zyklus dieses Spurengases noch größtenteils unerforscht. Nach neueren Forschungsergebnissen scheinen eher Mikroorganismen als „freie“ Enzyme für die Aufnahme von atmosphärischem H2 im Boden verantwortlich zu sein. Die CARD-FISH Analyse in dieser Arbeit belegte, dass die hoch-affine H2 Aufnahme-Aktivität nicht im Myzelium, sondern in den Sporen von Streptomyzeten exprimiert wird. Dies zeigt die essentielle Bedeutung von H2-oxidierenden Streptomyzeten, oder Aktinobakterien im Allgemeinen, für die Aufnahme von atmosphärischem H2 in „upland soils“.

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