The impact of an active soil microbial community on greenhouse gas emissions in Arctic cryosols. By Ianina Altshuler Department of Natural Resource Sciences Faculty of Agriculture and Environmental Sciences McGill University April, 2019 A thesis submitted to McGill University in partial fulfillment of the requirements of the degree of Doctor of Philosophy © Ianina Altshuler 2019 Petit à petit, l'oiseau fait son nid… - Old French Proverb ii Table of Contents Table of Contents ........................................................................................................................... iii List of Tables ................................................................................................................................ vii List of Figures .............................................................................................................................. viii Acknowledgments............................................................................................................................x Abstract .......................................................................................................................................... xi Résumé ......................................................................................................................................... xiii Contributions to knowledge ......................................................................................................... xvi List of Abbreviations .................................................................................................................. xvii Introduction and Literature Review: Microbial Life in Permafrost Soils ......................1 Introduction ..........................................................................................................................1 Permafrost environments .....................................................................................................2 Microbial diversity and abundance in permafrost ...............................................................6 Are they livin’ or just chillin’?...........................................................................................11 1.4.1 Soil respiration and laboratory incorporation studies ............................................12 1.4.2 Stable isotope probing in permafrost soils .............................................................13 1.4.3 RNA/DNA ratios ...................................................................................................14 1.4.4 Subzero growth of permafrost isolates ..................................................................15 1.4.5 DNA repair.............................................................................................................17 Live microbes or ancient DNA ..........................................................................................19 1.5.1 Strategies for differentiating between old biomarkers and an active microbial community .............................................................................................................21 Warming climate and permafrost .......................................................................................22 1.6.1 Methane dynamics in permafrost affected cryosols...............................................26 iii Objectives of this thesis .....................................................................................................29 Connecting Text: ............................................................................................................................34 Species interactions and distinct microbial communities in high Arctic permafrost affected cryosols are associated with the CH4 and CO2 gas fluxes...........................................35 Abstract ..............................................................................................................................35 Introduction ........................................................................................................................37 Materials and Methods .......................................................................................................41 2.3.1 Site and sample collection .....................................................................................41 2.3.2 Gas flux measurements ..........................................................................................41 2.3.3 DNA extraction and sequencing ............................................................................42 2.3.4 Isolation of IWP bacteria .......................................................................................43 2.3.5 Bioinformatics........................................................................................................43 2.3.6 Hybrid network analysis ........................................................................................45 Results ................................................................................................................................47 2.4.1 Gas flux ..................................................................................................................47 2.4.2 IWP terrain microbial community diversity ..........................................................48 2.4.3 Hybrid network analysis of the microbial community ..........................................48 2.4.4 PmoA diversity at the IWP terrain .........................................................................49 2.4.5 Bacterial isolates from the IWP terrain ..................................................................50 Discussion ..........................................................................................................................51 Conclusions ........................................................................................................................58 Supplementary Materials ...................................................................................................70 Connecting Text: ............................................................................................................................77 Denitrifiers, Nitrogen-fixing bacteria, and N2O ...........................................................78 soil gas flux in high Arctic ice-wedge polygon cryosols. ..............................................................78 Abstract ..............................................................................................................................79 iv Introduction ........................................................................................................................80 Material and Methods ........................................................................................................83 3.3.1 Gas flux measurements, flux calculation, and statistics ........................................83 3.3.2 Site and sample collection .....................................................................................84 - - + 3.3.3 NO3 & NO2 , NH4 , and soil moisture measurements ..........................................85 3.3.4 DNA and RNA isolation and sequencing ..............................................................85 3.3.5 qPCR analysis and statistics...................................................................................86 3.3.6 Bioinformatics and statistical analysis of sequencing data ....................................87 3.3.7 Phylogenetic analysis .............................................................................................88 Results and Discussion ......................................................................................................90 - - + 3.4.1 N2O Gas Flux, Soil NO3 & NO2 , NH4 , and Moisture Content ...........................90 3.4.2 Quantitative PCR ...................................................................................................91 3.4.3 Community structure of potential denitrifiers and nitrogen fixing bacteria ..........92 3.4.4 Denitrifying community.........................................................................................92 3.4.5 Nitrogen-fixing community ...................................................................................94 3.4.6 Metatranscriptomics ...............................................................................................96 Conclusions ........................................................................................................................98 Supplementary Materials: ................................................................................................109 Connecting Text: ..........................................................................................................................114 In situ DNA-SIP enrichment in concert with genome binning of a high Arctic methanotrophic and methanotroph-associated community. ....................................................115 Abstract ............................................................................................................................116 Introduction ......................................................................................................................117 Methods............................................................................................................................120 Results and Discussion ....................................................................................................123 Conclusion .......................................................................................................................128