Isolation of rhizobacteria in Southwestern Québec, Canada: An investigation of their impact on the growth and salinity stress alleviation in Arabidopsis thaliana and crop plants Di Fan Department of Plant Science Faculty of Agricultural and Environmental Sciences Macdonald Campus of McGill University 21111 Lakeshore Road, Sainte-Anne-de-Bellevue, Québec H9X 3V9 December 2017 A thesis submitted to McGill University in partial fulfillment of the requirements of the degree of DOCTOR OF PHILOSOPHY © Di Fan, Canada, 2017 Table of contents Abstract ................................................................................................. x Résumé ................................................................................................ xiii Acknowledegments ............................................................................. xv Preface ................................................................................................ xviii Contribution of authors ................................................................................ xviii Chapter 1................................................................................................. 1 Introduction ............................................................................................ 1 Chapter 2................................................................................................. 5 Literature Review ................................................................................... 5 2.1 What are root exudates? .......................................................................... 5 2.2 Roles of root exudates .............................................................................. 5 2.2.1 Manipulating the rhizosphere environment ....................................................... 6 2.2.2 Plant-to-microbe signals ....................................................................................... 7 2.2.2.1 Flavonoids ................................................................................................................. 7 2.2.2.2 Jasmonates ................................................................................................................ 8 2.2.2.3 Other nod gene inducers ........................................................................................... 9 2.2.2.4 Strigolactones............................................................................................................ 9 2.2.3 Quorum sensing ................................................................................................... 10 2.3 Plant growth promotion ........................................................................ 12 2.3.1 Root colonization ................................................................................................. 12 2.3.2 Biofertilization ..................................................................................................... 13 ii 2.3.2.1 Nitrogen fixation ..................................................................................................... 13 2.3.2.2 Phosphate solubilization ......................................................................................... 14 2.3.2.3 Iron availability ....................................................................................................... 15 2.4 Phytoremediation ................................................................................... 16 2.5 Phytostimulation .................................................................................... 17 2.5.1 Indole-3-acetic acid (IAA) .................................................................................. 17 2.5.2 Gibberellins .......................................................................................................... 18 2.5.3 Cytokinins ............................................................................................................ 18 2.5.4 Regulation of Ethylene ........................................................................................ 19 2.5.5 Other plant growth regulators ........................................................................... 19 2.6 Biocontrol ............................................................................................... 20 2.6.1 Antibiotics ............................................................................................................ 21 2.6.2 Bacteriocins .......................................................................................................... 21 2.6.3 Extracellular lytic enzymes ................................................................................ 22 2.6.4 Siderophores ........................................................................................................ 22 2.6.5 Phytoalexins ......................................................................................................... 23 2.6.6 Control of other biotic stresses ........................................................................... 23 2.6.7 Signal interference............................................................................................... 24 2.6.8 Induced systemic resistance................................................................................ 24 2.7 Novel type of signaling ........................................................................... 26 2.7.1 Nod factors ........................................................................................................... 26 2.7.2 Bacteriocins .......................................................................................................... 27 2.8 Old-timer versus rising star .................................................................. 29 iii 2.9 PGPR screening ..................................................................................... 30 Connecting text ..................................................................................... 33 Chapter 3 ............................................................................................. 34 Isolation and diversity of culturable rhizobacteria associated with economically important crops and uncultivated plants in Québec, Canada ................................................................................................. 34 3.1 Abstract .................................................................................................. 34 3.2 Introduction ........................................................................................... 35 3.3 Materials and Methods .......................................................................... 39 3.3.1 Study site and sample collection ........................................................................ 39 3.3.2 Isolation and preservation of cultivable rhizobacteria .................................... 42 3.3.2.1 Isolation of ectophytic bacteria ............................................................................... 42 3.3.2.2 Isolation of cultivable endophytes ........................................................................... 43 3.3.2.3 Isolation of endophytic bacteria from soybean root nodules .................................. 44 3.3.3 Taxonomic identification and diversity indices ................................................ 45 3.3.3.4 Statistical analysis ............................................................................................ 46 3.4 Results .................................................................................................... 46 3.4.1 Enumeration, isolation, and identification of rhizobacteria ........................... 46 3.4.2 Diversity of culturable ectophytic bacterial ...................................................... 51 3.4.3 Culturable endophytic bacterial community survey........................................ 55 3.5 Discussion................................................................................................ 64 3.6 Conclusions ............................................................................................ 69 Connecting text ................................................................................... 123 iv Chapter 4 ........................................................................................... 124 Inoculation of Arabidopsis thaliana by novel rhizobacterial isolates improves plant growth under normal and stress conditions .......... 124 4.1 Abstract .................................................................................................124 4.2 Introduction ..........................................................................................125 4.3 Materials and Methods .........................................................................129 4.3.1 Plant material, bacterial strains, and growth conditions .............................. 129 4.3.2 Rapid bacterial screening assay ....................................................................... 131 4.3.3 Secondary bacterial screening.......................................................................... 131 4.3.4 Petri plate assay for effects of bacterial inoculation on seedling growth ..... 132 4.3.4.1 Germination activities of Arabidopsis seeds after bacterization .......................... 132 4.3.4.2 In Vitro osmotic stress .......................................................................................... 132 4.3.4.3 In Vitro salt stress ................................................................................................. 133 4.3.4.4 Root architecture analysis ...................................................................................