The candidate phylum Termite Group 1 [Elektronische Ressource] : diversity, distribution, metabolism and evolution of representatives of an unexplored bacterial phylum / vorgelegt von Daniel Philipp Ralf Herlemann

Herlemann

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• Biologie

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• Publié par: philipps-universitat_marburg
• Publié le: 01 janvier 2009
• Nombre de lectures: 48
• Langue: Deutsch
• Poids de l'ouvrage: 4 Mo

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The Candidate phylum "Termite Group 1"
Diversity, distribution, metabolism and evolution of
representatives of an unexplored bacterial phylum
Dissertation
Zur Erlangung des Doktorgrades der
Naturwissenschaften im Fachbereich Biologie der
Philipps-Universität Marburg
Vorgelegt von
Daniel Philipp Ralf Herlemann
aus Offenburg Marburg/Lahn 2009
Die Untersuchungen zur folgenden Arbeit wurden von Juni 2006 bis Juli 2009 am
Max-Planck-Institut für terrestrische Mikrobiologie in Marburg unter Leitung von
Prof. Dr. Andreas Brune durchgeführt.
Vom Fachbereich Biologie der Philipps-Universität Marburg als Dissertation
angenommen am:
Erstgutachter: Prof. Dr. Andreas Brune
Zweitgutachter: Prof. Dr. Uwe Maier The following publications are integrated in this thesis:
Herlemann, D. P. R., Geissinger, O., and A. Brune. 2007. The Termite Group
1 phylum is highly diverse and widespread in the environment. Appl. Environ.
Microbiol. 73, 6682-6685.
Geissinger O., Herlemann, D. P. R., Maier U., and A. Brune. 2009.
Elusimicrobium minutum gen. nov., the first isolate of the Termite Group 1
phylum. Appl. Environ. Microbiol. 75, 2831-2840.
Herlemann, D. P. R., Geissinger, O., Ikeda-Ohtsubo, W., Kunin, V., Sun, H,
Lapidus A., Hugenholtz P., and A. Brune. 2009. Genome analysis of
Elusimicrobium minutum, the first cultivated representative of the Elusimicrobia
phylum (formerly Termite Group 1). Appl. Environ. Microbiol. 75, 2841-2849.Erklärung
Ich versichere, dass ich meine Dissertation
„The Candidate phylum “Termite Group 1” – Diversity, distribution, metabolism
and evolution of representatives of an unexplored bacterial phylum”
selbständig und ohne unerlaubte Hilfe angefertigt habe und mich keiner als der von
mir ausdrücklich bezeichneten Quellen und Hilfen bedient habe. Diese Dissertation
wurde in der jetzigen oder einer ähnlichen Form noch bei keiner anderen
Hochschule eingereicht und hat noch keinen sonstigen Prüfungszwecken gedient.
Marburg, Juli 2009 Danksagung
An erster Stelle möchte ich mich bei meinem Doktorvater Prof. Dr. Andreas Brune
für die Überlassung des Themas sowie die stets offene Tür bei Fragen und
Diskussionen als auch den Freiraum für die Entfaltung eigener Ideen bedanken.
Herrn Prof. Dr. Uwe Maier danke ich für die Übernahme des Zweitgutachtens und
Prof. Dr. Ralf Conrad danke ich für die Möglichkeit in seiner Abteilung zu arbeiten.
Ebenfalls möchte ich dem IMPRS „Thesis advisory comitee“ bestehend aus Prof.
Dr. Michael Friedrich, Dr. Werner Liesack und Prof. Dr. Rolf Thauer für
umfangreiche Diskussionen und Hinweise im Laufe meiner Doktorarbeit danken.
Weiterer Dank gilt allen aktuellen und ehemaligen Mitgliedern der
„Termitengruppe“ für die gute und fröhliche Arbeitsatmosphäre. Im Besonderen
danke ich unserer technischen Assistentin Katja Meuser für die große Hilfe bei
vielen kleinen Dingen ohne die ein Labor nicht funktioniert.
Mein letzter und größter Dank gilt meiner Familie und Inga Krämer für die
Unterstützung und Standhaftigkeit in schwierigen Situationen. Table of Contents
1 Introduction 1
—————————————————————————————
Microbial diversity in the gut of lower termites 1
Candidate phylum "Termite Group 1" 3
Functional characterization of uncultivated microorganisms 3
Endomicrobia in termite guts 4
Chapter outline 6
References 6
2 The Termite group 1 Phylum is highly diverse and widespread
in the environment 11
—————————————————————————————
Summary 11
Introduction 12
Data mining 12
Primer design and PCR 13
Phylogenetic analysis 13
Abundance in the environment 17
Conclusion 17
References 17
3 The ultramicrobacterium "Elusimicrobium minutum" gen.
nov., sp. nov., the first cultivated representative of the Termite
Group 1 Phylum 21
—————————————————————————————
Summary 21
Introduction 22
Material and methods 22
Isolation and morphological characterization 27
Growth and nutrition 29
Chemotaxonomic analysis 33
Phylogenetic analysis 34
Physiology 35
Ecology 37
Size 38
Taxonomy 39
References 42 4 Genomic analysis of Elusimicrobium minutum, the first
cultivated representative of the phylum Elusimicrobia
(formerly Termite Group 1) 51
—————————————————————————————
Summary 51
Introduction 52
Material and methods 53
Genome structure 55
Phylogeny and taxonomy 57
Energy metabolism 58
Anabolism 61
Peptide degradation 63
Secretion 64
Oxygen stress 67
Ecological considerations 68
References 69
5 Parallel genomic evolution of Candidatus "Endomicrobium
trichonymphae" genomes from Trichonympha protists in
termites 77
———————————————————————––––––––––––
Summary 77
Introduction 78
Enrichment of Candidatus "Endomicrobium trichonymphae" 79
Metagenome 80
Parallel evolution 81
Genome rearrangement 84
References 85
6 General discussion 89
—————————————————————————————
The undiscovered diversity of Elusimicrobia 89
Alanine – and unusual fermentation end product for
glycolytic organisms 90
Hydrogenases – key enzymes in the metabolism of E. minutum 91
Orthologous proteins often but not always have the same function 92
Implications for endomicrobia deduced from the E. minutum
genome 93
Does homologous recombination help to escape Muller’s ratchet 94
References 96
Summary 101
Zusammenfassung 105
Curriculum vitae 109
Appendix 110Introduction
1. Introduction
Termites (Insecta, Isoptera) are terrestrial arthropods abundant in tropical habitats and also
present in temperate zones. Termite colonies can exceed 6000 individuals per m², and they have
a high impact on the dynamics of carbon and nitrogen in soil (3, 9). The majority of the termite
species are able to digest lignocellulosic compounds like wood, grass, and plant litter. This
ability has recently made them the subject of biofuel-production studies and has attracted much
attention to the digestive processes within the termite intestinal tracts (28).
Termites are divided into seven families: Mastotermitidae, Kalotermitidae, Hodotermitidae,
Termopsidae, Rhinotermitidae, Serritermitidae, and Termitidae (1). The Termitidae represent
the evolutionarily higher termites, characterized by a nutritionally diverse diet and the lack of
cellulolytic gut flagellate symbionts. All six families of lower termites feed exclusively on wood
and have relatively simple intestinal tracts consisting of three compartments (foregut, midgut,
and hindgut). The midgut contains primarily enzymes provided by the host, whereas the
digestion in the hindgut depends on special single-celled eukaryotes responsible for cellulose
degradation. The microbial activity yields high concentrations of fermentation products that are
absorbed by the termite and further oxidized (6). A major part of this rich reservoir of microbial
diversity in the hindgut has remained elusive for classical cultivation techniques. Owing to the
difficulty in cultivating most of the members from the termite gut microbiota and the complexity
of the community, our understanding of the major bacterial groups in the termite gut is still
poor.
Microbial diversity in the gut of lower termites
Interdisciplinary research, combining classical microbiological techniques with genetic and
biochemical approaches, became one of the most successful strategies for the characterization of
diversity in the termite gut. A standard method for the identification and classification of
bacteria is performed by targeting the small subunit ribosomal RNA (SSU rRNA). The SSU
rRNA is an ideal phylogenetic marker because it is present in all living organisms and has
highly conserved and variable regions (32). Comparative analysis of the SSU rRNA sequence
not only resulted in the separation of the three domains Bacteria, Archaea, and Eukarya (32), but
also transformed microbial taxonomy from an identification system to an evolutionary-based
1Introduction
systematic framework (26, 33). The diversity of 16S rRNA sequences present in a sample is
therefore used to describe the microbial diversity.
Anaerobic flagellate protists are essential for the depolymerization of cellulose and
hemicellulose in lower termites (15, 4). Molecular studies showed that these flagellates belong
to the phyla Parabasalia and Preaxostyla (18, 5) and occur exclusively in the gut of termites and
wood-feeding cockroaches. Other than Trichomitopsis termopsidis (23, 34), termite gut
flagellates have not been cultivated in axenic culture. Flagellates in the hindgut of lower
termites occupy the majority of the volume (19). Their surface is usually colonized by many
prokaryotic ectosymbionts (2). In addition, the cytoplasm provides an important habitat for
many endosymbiotic prokaryotes (7).
The adaptation of termite gut prokaryotes to gut-specific niches is reflected in the phylogeny of
the 16S rRNA sequences. Sequences derived from termites often form unique monophyletic
lineages, indicating that many bacterial speci