Molecular phylogeny, taxonomy and evolution of arbuscular

mycorrhizal fungi

DNA-based characterization and identification of Glomeromycota

Kumulative Dissertation

der Fakultät für Biologie

an der Ludwig-Maximilians-Universität München

zur Erlangung des Doktorgrades der Naturwissenschaften (Dr. rer. Nat.)

vorgelegt von

Dipl.-Nat. Manuela Krüger

aus Zwickau

Tag der Einreichung: 1. Februar 2011

2

1. Gutachter: PD Dr. Arthur Schüßler

2. Gutachter: Prof. Dr. Martin Parniske

Tag der mündlichen Prüfung: 6. Mai 2011

Table of contents

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Table of contents

List of Abbreviations (except SI units) ................................................................................................5

1. Abstract .......................................................................................................................................6

2. Zusammenfassung .......................................................................................................................7

3. Introduction .................................................................................................................................8

3.1 Arbuscular mycorrhizal fungi ................................................................................................8

3.2 Evolution of AMF .................................................................................................................9

3.3 Morphological characterization and taxonomy of AMF ........................................................ 10

3.4 Molecular characterization of AMF ..................................................................................... 12

3.5 In-field detection of AMF and community analyses ............................................................. 13

3.6 DNA barcoding ................................................................................................................... 14

3.7 Deep sequencing of AMF communities................................................................................ 14

3.8 Aim of this study ................................................................................................................. 15

4. DNA-based species level detection of Glomeromycota: one PCR primer set for all arbuscular

mycorrhizal fungi ...................................................................................................................... 16

5. DNA barcoding of arbuscular mycorrhizal fungi ........................................................................ 29

6. Acaulospora brasiliensis comb. nov. and Acaulospora alpina (Glomeromycota) from upland

Scotland: morphology, molecular phylogeny and DNA-based detection in roots ........................ 44

Abstract ..................................................................................................................................... 45

Introduction ............................................................................................................................... 45

Materials and Methods .............................................................................................................. 46

Results ...................................................................................................................................... 48

Discussion ................................................................................................................................. 53

Acknowledgements ................................................................................................................... 56

References ................................................................................................................................. 56

Legends to figures ..................................................................................................................... 59

7. Revealing natural relationships among arbuscular mycorrhizal fungi: culture line BEG47

represents Diversispora epigaea, not Glomus versiforme .......................................................... 63

Table of contents

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8. A phylogenetic framework for the natural systematics of arbuscular mycorrhizal fungi: from

phylum to species-level resolution and environmental deep sequencing ..................................... 76

Summary ................................................................................................................................... 77

Introduction ............................................................................................................................... 77

Materials and Methods .............................................................................................................. 79

Results ...................................................................................................................................... 82

Discussion ................................................................................................................................. 89

Acknowledgements ................................................................................................................... 94

References ................................................................................................................................. 95

Figure Legends .......................................................................................................................... 98

9. Discussion ............................................................................................................................... 105

9.1 General discussion ............................................................................................................. 105

9.2 The recent taxonomy of Glomeromycota ............................................................................ 106

9.3 Evolution of Glomeromycota ............................................................................................. 107

9.4 Molecular phylogeny of Glomeromycota ........................................................................... 108

9.5 DNA barcoding of Glomeromycota .................................................................................... 110

10. Outlook ................................................................................................................................... 111

11. References ............................................................................................................................... 112

12. Acknowledgment .................................................................................................................... 120

13. Appendix ................................................................................................................................ 121

13.1 Supplementary data – chapter 5........................................................................................ 121

13.2 Supplementary data – chapter 6........................................................................................ 152

13.3 Supplementary data – chapter 7........................................................................................ 153

13.4 Supplementary data – chapter 8........................................................................................ 159

14. Contribution of the author........................................................................................................ 171

15. Curriculum vita ....................................................................................................................... 172

List of Abbreviations

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List of Abbreviations (except SI units)

~ approximately AFTOL Assembling the Fungal Tree of Life AM arbuscular mycorrhiza AMF arbuscular mycorrhizal fungi approx. approximate(ly) Att attempt BEG International Bank for the Glomeromycota bp base pair(s) BS bootstrap support BSA bovine serum albumin cf. Latin: confer (English: compare) comb. nov. Latin: combinatio nova (English: new combination) CTAB cetyltrimethylammonium bromide DAOM Agriculture and Agri-Food Canada National Mycological Herbarium DMSO dimethyl sulfoxide DNA deoxyribonucleic acid DNase deoxyribonuclease dNTP deoxyribonucleoside triphosphate e.g. Latin: exempli gratia (English: for example) GlGrA Glomus Group A GlGrB Glomus Group B GlGrAa Glomus Group Aa GlGrAb Glomus Group Ab INVAM International Culture Collection of (Vesicular) Arbuscular Mycorrhizal Fungi ITS internal transcribed spacer kb kilo base pair(s) LB lysogeny broth (see Bertani, 2004) LSU large subunit ML maximum likelihood MOTU molecular operational taxonomic unit mt mitochondrial MUCL Mycothèque de l'Universite Catholique de Louvain Mya million years ago OTU operational taxonomic unit PCR polymerase chain reaction rDNA ribosomal DNA RFLP restriction fragment length polymorphism RNA ribonucleic acid ROC root organ culture rRNA ribosomal RNA sensu English: in the sense of SB sodium borate SDS sodium dodecyl sulfate SSU small subunit Taq Thermus aquaticus Tm melting temperature