Understanding of plant colonization by endophytic bacteria by tracking them inside plants

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1 Understanding of plant colonization by endophytic bacteria by tracking them inside plants Dr. Stéphane Compant Bioresources Unit, Health & Environment Department, AIT Austrian Institute of Technology GmbH, 3430 Tulln, Austria

2 Introduction Bryophytae, Pteridophytae, Gymnospermae Angiospermae Contain various bacterial taxa Adv. Agronomy in press Endophytes in various plants

3 Introduction Various bacterial taxa analyzed, communities described by cultivable /metagenomic analysis But what can be their niches and ways of colonization? Endophytes analyzed in different organs such as roots, stems, leaves, flowers and fruits and seeds of many plants Adv. Agronomy in press AEM 2005a SBB 2010

4 Rhizoplane Endorhiza Introduction irmicutes Rh Co Native microflora in the rhizosphere and endorhiza c Xy Xy a b d Co cillus spp. Rh f Co h Example with Bacillus spp. in grapevine e g i Rhizosphere colonization and endophytism by bacteria

5 Introduction Native microflora in stem Endophytism in xylem vessels by bacteria

6 Introduction Native microflora in flowers Example with flowers of grapevine Microbial Ecol 2011 Presence in various tissues of flowers

7 Introduction Native microflora in fruits and seeds Microbial Ecol 2011 Presence in fruits and seeds

8 General ways of colonization by endophytes Sbb 2010 From the soil then to the other plant parts

9 General ways of colonization by endophytes There is some microbes inside plants, they derive from the soil, and colonization should be studied 1887 MLV Galippe The 125 th anniversary of the first postulation of the soil origin of endophytic bacteria a tribute to MLV Galippe. Plant and Soil 2012

10 Other ways of colonization by endophytes Endophytes can also derive from the caulosphere, the laimosphere, the anthosphere, the phyllosphere, and the carposphere SBB 2010 Other sources of colonization

11 PGPB effects and endophytic bacteria: niches should be determined Angiospermae Endophytes once applied where can be present the beneficial bacterium once applied? Under greenhouse or field conditions Stem and leaves, flowers, fruits? Should be determined before to study ISR Roots? AEM 2005; SBB 2010 Which tissues? Which organs are colonized? Does the bacterium interact with the native plant microbiome? Persistence?

12 Tools to track microbial colonization Various tools to study the colonization process and behaviour of beneficial bacteria Microscopic tools: Intercaling DNA markers, peptide and lipid markers gfp/dsred/gusa markers, FISH, DOPE-FISH, LNA-FISH, PNA-FISH, card-fish etc It depends of the model used and if field experiments are used or not In order to track the microbes once applied. To know their sites of colonization

13 Colonization behaviour of B. phytofirmans strain PsJN 100 µm 30 µm 75 µm Soil Biology and Biochemistry µm 30 µm PsJN::gfp2x on the rhizoplane of grapevine. 100 µm FEMS Microbiol Ecol 2008 Colonization from the soil

14 Colonization behaviour of B. phytofirmans strain PsJN Secondary root Rhizodermis Exodermis Endodermis Xylem Cortex Exodermis Exodermis Rh Ex Co Co Cortex Root hair Central Hairy root cylinder Endodermis Pericycle a b c En Cc d Xy icial endophytic bacteria. epi- and endophytic microorganisms. Soil Biology and Biochemistry 2010 PsJN::gfp2x inside endorhiza of grapevine. Fems Microbiol Ecol 2008 Colonization from the soil

15 Colonization behaviour of B. phytofirmans strain PsJN Pedicel Berry Vegetative and reproductive organs a Inflorescence stalk 10 µm b Xylem of stalk 10 µm a 10 µm b Fems Microbiol Ecol 2008 Colonization from the soil and then a spread to reproductive organs

16 Colonization of grapevine by NRRL B PGPB Saccharothrix algeriensis NRRL B Colonization? DOPE-FISH microscopy of S. algeriensis NRRL B dpi Soil Biology and Biochemistry 2010 Plant and Soil 2013 colonization from the rhizosphere to rhizoplane at root hair level and secondary root emergence site

17 Colonization of grapevine by NRRL B DOPE-FISH microscopy of S. algeriensis NRRL B dpi PGPB Saccharothrix algeriensis NRRL B Colonization? Soil Biology and Biochemistry 2010 Plant and Soil 2013 colonization from the rhizosphere to rhizoplane at root elongation zone

18 Colonization of grapevine by NRRL B Secondary root DOPE-FISH microscopy of S. algeriensis NRRL B dpi Rhizodermis Exodermis Exodermis Cortex Root hair Central Hairy root cylinder Endodermis Pericycle ficial endophytic bacteria. r epi- and endophytic microorganisms. Soil Biology and Biochemistry 2010 Plant and Soil 2013 colonization of the endorhiza of grapevine plants

19 Wheat colonization of a beneficial bacterium from desert soil by using other tools Collaboration with A. Zitouni and O. Toumatia IBSM alger, Algeria A biocontrol Streptomyces mutabilis strain, isolated from desert soil, applied on wheat. It reduces Fusarium spp. contamination on roots, but where can be the microbe once inoculated?

20 Wheat colonization behaviour of a Streptomyces mutabilis strain in wheat DOPE-FISH Microscopy using HGC69a probe coupled with dylight 594 Streptomyces sp. appeared as orange colored and different forms. My: mycelium, Pmy: pack of mycelium. Al: aleuronic cells, En: endocarp, Endos: endosperm, Enp: endopleura, Ep: epicarp, Me: mesocarp, In preparation Sites of colonization inside the fruit parts of the caryopsis, e.g. up to the endocarp

21 Colonization behaviour of a Streptomyces mutabilis strain in wheat DOPE-FISH Microscopy using HGC69a probe coupled with dylight 594 Streptomyces sp. appeared as orange colored. Gsp: germinated spore, My: mycelium. Ep: epidermis, Pa: parenchyma, Xy: xylem In preparation Better colonization inside leaves. From the phyllosphere to internal tissues but also spread inside xylem vessels

22 General ways of colonization by endophytes Sbb 2010 From the soil then to the other plant parts but it depends of the strain!

23 Conclusions Different tools and process of colonization Knowledge on microhabitats of applied strains and natural microflora Allow to determine if the strain, once applied, can colonize plants and various organs or be restricted Biofertilization and biocontrol properties and studies will depend of these colonizations but if you use a strain in the field how to differenciate it from the native plant microbiome if the species already exist? Microbes can restrict their studies of colonization

24 Bioresources Unit, Tulln, Austria A. Sessitsch Head; Senior scientist G. Brader F. Trognitz B. Mitter E. Hackl S. Compant Project leaders-scientists working on plant-microbe interactions Deciphering the plant microbiome Genetic/omic analysis of beneficial strains Mechanisms of plantmicrobe interactions Biocontrol of plant diseases Biofertilization of crops Biomiming

25 Vielen Dank