Microbiome engineering aims to manipulate plant and soil microbiomes to optimize agricultural functions like stress tolerance, growth promotion, and phytoremediation. Techniques include transferring native or synthetic microbiomes to seeds, seedlings, or soil. This can improve drought tolerance, disease resistance, and heavy metal accumulation. Challenges include understanding complex microbiome dynamics and interactions under field conditions. Microbiome engineering shows potential to develop sustainable agriculture through balanced, beneficial microbiome compositions.

1. Welcome
1

2. (FAO and world bank, 2007)
World scenario of human
population growth
2

3. Phytoremediation
Disease
resistance
Abiotic stress
tolerance
Crop
improvement
Sustainability in
agriculture
3

4. Microbiome engineering: current
uses and future
Ajayasree. T. S.
2018-21-026
Department of Plant Pathology
4

5. Outline
• Introduction
• Plant and soil microbiome
• Steps in microbiome engineering
• Microbiome engineering techniques
• Advantages of microbiome engineering in agriculture
• Challenges of microbiome engineering
• Future thrust
• Summary
• Conclusion
5

6. Introduction
6
Microbiome - aggregate of all microbes and
their genomes in a particular habitat
(Bulgarelli et al., 2013)

7. Introduction
7
Plant
microbiomes
Protecting the plant from
potential pathogens
Improving
growth
Improving health, adaptive
advantages to plant
Improving
production
(Haney et al., 2015)
…Contd.

8. 8
What is microbiome engineering?
Balanced microbiome
composition
Decreased
diversity Altered
proportion
Increased
diversity
Diseases/
Disorders
Low fitness
Slow
growth
Low
productivity
Low fertility
Optimum
health
High fitness
Fast growth
High
productivity
High fertility
Perturbation
Homeostasis Dyshomeostasis
(Foo et al., 2017)
Microbiome
engineering

9. Why plant microbiome engineering?
9
• Manipulate the microbiome
• Optimize plant functions of interest
• Broad spectrum mechanisms of action
• Improve reliability without genetic engineering
• Sustainable in nature

10. Types of microbiome engineering
10
• Plant microbiome engineering
• Soil microbiome engineering
• Human microbiome engineering
• Animal microbiome engineering
(Foo et al., 2017)

11. Plant microbiome
11
(Haney and Ausubel, 2015)
Microbial communities associated
with the plant which can live,
thrive, and interact with different
tissues such as roots, shoots,
leaves, flowers, and seeds Plant microbiome

12. Microbiome transfer in plants
12
Synthetic root - associated
microbiota transplant
Inhibit plant diseases, resist
environmental stresses,
promote growth
Native root - associated
microbiota transplant
(Foo et al., 2017)

13. Microbiome transfer in plants
13
• Transfer of conducive soil microbiota
• Easy to manipulate
• Limited availability - functional native microbiome
Native root - associated microbiota transplant
(Foo et al., 2017)
…Contd.

14. Microbiome transfer in plants
14
…Contd.
• Customization of microbial composition
• Limited understanding - core microbiome
• Applicable to culturable microbes
Synthetic root - associated microbiota transplant
(Foo et al., 2017)

15. Soil microbiome
15
Soil microbiome refers to microbial
communities in the bulk soil beyond the
rhizosphere and is mainly influenced by
agricultural management practices
(Foo et al., 2017)
Soil Microbiome

16. Overview of micro-organisms
present in the rhizosphere
16
Fungi/Oomycetes
~15500 genes
(18 to 82 Mb)
(105 to 106 per g)
Archaea
~ 1300 genes
(1.6 Mb)
(107 to 108 per g)
Protozoa
~ 14000 genes
(34 Mb)
(103 to 105 per g)
Viruses
~ 45 genes
(4 to 69 Kb)
107 to 109 per g
Algae
~ 13000 genes
(42 to 105 Mb)
(103 to 106 per g)
Nematodes
~ 18000 genes
(54 to 100 Mb)
(101 to 102 per g)
Bacteria
~ 6500 genes
(4 to 9 Mb)
(108 to 109 per g)
(Mendes et al., 2013)

17. 17
Rhizosphere
microbiome
the
GOOD
Nutrient
acquisition
Protection
against
pathogens
Immune
response
Growth and
development
Tolerance
to abiotic
stress
Physiology/
metabolism
the
BAD
Plant
diseases
the
UGLY
Food
contamination
Role of rhizosphere microbiome
17
(Mendes et al., 2013)

18. Engineering of soil microbiome
• Implement organic farming
• Change land utilization
• Tillage
• Cropping systems
• Other agricultural practices
18
(Foo et al., 2017)

19. Steps in microbiome engineering
19
1. Identification and culturing of potential PGPMs
2. Deep analysis/ selection of the various components
Culturing of PGPM Analysis
(Woo and Pepe, 2018)
*PGPM - Plant Growth Promoting Microbes

20. Steps in microbiome engineering
20
3. Evaluate compatibility
4. Effects in the native agroecosystem
5. Develop formulation and distribution technology
6. Technical support to end users
(Woo and Pepe, 2018)
…Contd.

21. Techniques of microbiome
engineering
21
(Keresa et al., 2008)
1. Host-
mediated and
multi-
generation
microbiome
selection
2. Inoculation
into the soil
and
rhizosphere
3. Inoculation
into seeds or
seedlings
4. Tissue
atomisation
5. Direct
injection into
tissues or
wounds

22. 1. Host-mediated and multi-generation
microbiome selection
• Cycle-dependent strategy
• Indirect selection-microbiomes
• Utilizes the host phenotype
22
(Jochum et al., 2019)

23. Concept of host-mediated microbiome
engineering
23
2) Germinate seedlings under well watered conditions
3) Expose plants to water
deficiency
1) Initial
microbiome
inoculation
6) Repeat
steps 2-4
5) Add
selected
rhizospheres
to new
sterile
medium and
reseed
4) Sub-select and harvest,
and amalgamate the
rhizospheres from the most
drought resistant plants
(Jochum et al., 2019)
…Contd.

24. Host-mediated and multi-generation
microbiome selection
24
1 2
6 5 4
3
…Contd.
(Mueller and Sachs, 2015)

25. 2. Inoculation into soil and rhizosphere
25
• Inoculation of external strains
• Agrobacterium sp. 10C2 - Phaseolus vulgaris
• Bacillus licheniformis, Bacillus pumilus,
Paenibacillus koreensis, and the genera
Arthrobacter, Microbacterium, Brevibacterium
(Chihaoui et al., 2015)

26. 3. Inoculation into seeds or seedlings
26
Dendrobium nobile Lindl
(Pavlova et al., 2017)
Inoculation of Pseudomonas
fluorescens + Klebsiella
oxytoca into Dendrobium
nobile Lindl seeds
Growth
capacity
Germination
Increased
Adaptive
capacity

27. Inoculation into seedlings
27
…Contd.
(Rojas-Solís et al., 2018)
Pseudomonas stutzeri E25 and
Stenotrophomonas maltophilia
CR71 into the rhizosphere of
tomato seedlings
Plant growth-promotion,
Management of tomato gray mold
Tomato seedlings

28. 4. Tissue atomisation
28
(Mitter et al., 2017)
• Modify growth characteristics
• Changes endogenous microbiome of seeds - vertical
inheritance
• Decrease in α- and γ-Proteobacteria
• Increase in β-Proteobacteria
Endophytic bacterium
Paraburkholderia phytofirmans
PsJN into wheat and maize
flowers
Wheat

29. Tissue atomisation
29
…Contd.
A. Method of tissue atomisation
B. PsJN starts moving from embryo to germinated parts
A
B
(Mitter et al., 2017)

30. Tissue atomisation
30
…Contd.
• Modified microbiome - inherited
for more than one generation
• Plants in the second generation -
not inherit the PsJN strain
(Mitter et al., 2017)
• Bioengineering plant microbiome without genetic
manipulation

31. 5. Direct injection into tissues or wounds
31
(Wicaksono et al., 2017)

32. Direct injection into tissues or wounds
32
(Wicaksono et al., 2017)
Pseudomonas sp. R4R21AP
Pseudomonas sp. T1R12P
Pseudomonas sp. T1R21
Pseudomonas sp. T4MS32AP and
Pseudomonas T4MS33.
Endophytes
Pseudomonas syringae pv. actinidiae (Psa)
Bacterial canker of kiwifruit
Leptospermum scoparium
resistant to Pseudomonas
syringae pv. actinidiae (Psa)
Isolated
Biocontrol agents
against
…Contd.

33. Assessment of endophytic movement
33…Contd.
Foliar sprays - inoculate endophytic bacteria
(Wicaksono et al., 2017)

34. Inhibition mechanism
34
(Wicaksono et al., 2017)
…Contd.
Pseudomonas sp. T1R12P Pseudomonas sp. T1R21
Dual culture assay

35. Inference of study
35
• Inhibition of Pseudomonas syringae pv.
actinidiae biovar 3 of kiwi plants
• Reduced pathogen population
• Disease management by single
application
…Contd.
(Wicaksono et al., 2017)

36. 36
Microbiomes
Plants
3. Phytoremediation
4. Plant growth enhancement
6. Role of
Signaling
(Tian et al., 2020)
5. Salinity stress tolerance
Advantages of microbiome engineering in
agriculture
1. Drought
stress
tolerance
2. Disease
stress
tolerance

37. 37
1. Drought stress tolerance

38. 38
SI.
No.
Microbes Crop Mechanisms References
I Bacterial - phytohormone modulators
1. Rhizobium leguminosarum
LR-30, Mesorhizobium ciceri
CR-30 and CR-39 &
Rhizobium phaseoli MR-2
Wheat IAA improved
growth,
biomass
Hussain et al.,
2014
II Bacterial - ACC deaminase (ACCd) producers
2. Burkholderia phytofirmans,
Enterobacter sp.
Maize Increases
chlorophyll
content
Naveed et al.,
2014
III Bacterial - exopolysaccharide producers
3. Pseudomonas putida,
Rhizobium sp.
Sunflower EPS Alami et al.,
2000
Mechanisms of drought tolerance

39. Drought tolerance by microbiome
engineering
39
(Jochum et al., 2019)

40. Effect of HMME on drought tolerance
10
11
12
13
14
15
16
Numberofdayswithoutwater
Rounds of selection
40
…Contd.
*HMME: Host-Mediated Microbiome Engineering
(Jochum et al., 2019)

41. Effect of HMME on wheat seedlings
under drought stress
41
(Jochum et al., 2019)
…Contd.
Sterile rhizosphere soil Non-sterile rhizosphere soil

42. Inference of study
42
• Alter rhizosphere microbiome
• Drought stress symptoms onset delayed - 10th to 15th day
• Plant biomass, root dry weight, root length
• Soil aggregation, water holding capacity
• Less per cent water loss
• Reduction in alphaproteobacteria
• Increase of betaproteobacteria
…Contd.
(Jochum et al., 2019)

43. SI.
No.
Plant Microbiome Stress Effects References
1. Arabid
opsis
thalian
a
Xanthomonas sp.
WCS2014-23,
Stenotrophomonas
sp. WCS2014-113,
Microbacterium sp.
WCS2014-259
Hyaloperonos
pora
arabidopsidis
Less fungal
spores, higher
plant fresh
weight
Berendsen
et al., 2018
2. Potato Pseudomonas spp.
R32, R47, R76,
R84, S04, S19, S34,
S35, S49
Phytophthora
infestans
Reduced fungal
sporangiophore
development
Vrieze
et al., 2018
3. Tomato Pseudomonas spp.
CHA0, PF5, Q2-87,
Q8R1-96, 1M1-96,
MVP1-4, F113,
Phl1C2
Ralstonia
solanacearum
Reduced
disease
severity,
pathogen
abundance
Hu et al., 2016
43
2. Disease stress tolerance

44. 3. Phytoremediation
44
Betula celtiberica
54 culturable rhizobacteria and 41 root
endophytes
Metal plant accumulation
Phytoremediation
Birch tree
(Betula celtiberica)
(Mesa et al., 2017)
Against arsenic toxicity

45. Mechanisms of phytoremediation
45
plant growth by bacterial metabolites
• Plant growth by bacterial
metabolites - IAA
• Metal chelation by siderophores
• Organic acid production
• Phosphate solubilisation
• ACCD activity
(Mesa et al., 2017)
…Contd.
Neorhizobium sp.
Rhizobium sp.
Variovorax sp.
Phyllobacterium sp.
Rhodococcus sp.
Aminobacter sp.
Ensifer adhaerens

46. 4. Plant growth enhancement 46
(Kong et al., 2018)
High
quality
crops
High -
throughput
sequencing
Root
associated
microbiome
KOMODO: predict
microbial culture media
Core microbial
taxa and hubs
Network
analysis
Biofertilizer
Microbial
consortium
PGP
activities
Microbial
synergism
Dyanamic
changes
Artificial construction of synthetic microbial consortia
Ecological
evaluation
Efficacy
assessment

47. 5. Salinity stress tolerance
47
• Saline soil - EC ˃ 4 dS m-1
• Deteriorated growth
• Nitrogen content
• Photosynthetic capacity
• Metabolic processes Maize
(Upadhyay et al., 2011)

48. Microbiome against salinity…
48
SI.
No.
Microbiome Crops References
1. Serratia sp.+ Rhizobium sp. Lettuce Han and Lee, 2005
2. Rhizobium tropici (CIAT899) or R.
etli (ISP42) + Ensifer fredii SMH12,
HH103 + Chryseobacterium
balustinum Aur9
Common
bean,
Soybean
Estevezi et al., 2009
3. Pseudomonas sp.+ Rhizobium sp. Maize Bano and Fatima,
2009
4. Bacillus sp. + Burkholderia sp. +
Enterobacter sp. + Microbacterium
sp. + Paenibacillus sp.
Wheat Upadhyay et al., 2012
5. Brachybacterium saurashtrense
(JG-06) + Brevibacterium casei (JG-
08) + Haererohalobacter (JG-11)
Ground
nut
Shukla et al., 2012
…Contd.

49. 6. Role of signaling molecules
49
Administration of root
exudates
e.g. salicylic acid
Resist
environmental
stresses, promote
growth
• Promote - balanced
microbiome
• Limited availability -
signaling molecules
(Foo et al., 2017)

50. 50
Environmental
factors
Plant phenotype
Plant genotype,
Plant age
Factors influencing plant microbiome
engineering
(Compant et al., 2019)
Agricultural management
Soil characteristics
Abundance, diversity, functionality, and colonization of
microorganism in above- and below- ground plant parts

51. Challenges of microbiome engineering
1. Effect of abiotic or environmental factors
2. Deeper understanding of the microbial community structure
over time
3. Limited ability to harness and manipulate the microbiome in
agriculture
4. Nature and mechanisms of microbiota-plant relationship
5. Bridging the lab-field gap
51

52. Summary
52
 Microbiome engineering enhances sustainability in
agriculture
 Steps in microbiome engineering
 Various techniques for microbiome engineering
 Advantages of microbiome engineering
 Factors influencing microbiome engineering
 Challenges of microbiome engineering

53. Future thrusts
• Microbiome manipulation by plant
• Identification of stable, stress tolerant microbiomes
• Develop more microbial consortia
53

54. Conclusion
54
“ I play with microbes. There are of course, many rules
to this play… but when you have acquired knowledge
and experience it is very pleasant to break the rules and
to be able to find something nobody has thought of ˮ
Alexander Fleming

55. Thank you
55