Transcript Results

Ground-based research for soybean cultivation in BLSSs:
harmonization of plant structure and physiology
in hydroponics
Crew
Regenerate air
(photosynthesis)
Purified water
(transpiration)
Human wastes
(faeces-urine)
Edible products
(fresh food)
Water, Food,
Oxygen
Wastes
(Urine, Faeces)
CO2
Wastes
(substrate for
microorganisms/plants)
Inedible materials and O 2
Higher plants,
Bacteria
Bacteria
Mineral elements and CO2
Paradiso R., De Micco V., Arena C., Aronne G., Pepe O., De Pascale S.
University of Naples Federico II - Italy
ISLSWG Workshop Bioregenerative Life Support
Turin, 18-19 May 2015
Why soybean?
Nutritional value of seeds
Hoff et al., 1982
NASA Contract Report 166324
SEEDS
Osteoporosis risk
Khalil et al., 2002
J. Nutr. 132: 2605-2608
Washing
Germination
Soaking
SPROUTS
Milling
Paste boiling
Peroxidation products in brain
Chowdhury and Soulsby, 2002
Ann. Cl. Lab. Sci. 32(2):188-192
Filtration (0.2 mm)
Soymilk
Sanitization
(15 min boiling)
Proteolysis of myofibrillar proteins
Tada and Yokogoshi, 2002
J. Nutr. Sci. Vit. 48(2): 115-119
SANITIZED SOYMILK
OKARA
Objectives of research on Soybean within MELiSSA project
FC Phase 1
(Micro Ecological Life Support System Alternative)
I. to develop an objective theoretical procedure to identify candidate cvs
De Micco et al., 2012 - Proceedings 63rd Intl. Astronautical Congress (IAF)
De Micco et al., 2012 - Adv. in Space Res. 49: 1415-21
II. to evaluate plant behavior in hydroponics in controlled environment
Paradiso et al., 2012 - Adv. in Space Res. 50: 1501-11
Paradiso et al., 2014 – Plant biology, 16(suppl. 1): 69-78
III. to compare nutritional composition of seeds in hydroponics/open field
Palermo et al., 2012 - J. Agric. Food Chem., 60: 250-255
Interaction of soybean plants
with favourable microorganisms
C
C uu ll S
S ee ll
FC Phase 2
to evaluate the effects of root symbiosis with nitrogen fixing bacteria on plant
performance in closed-loop hydroponics in controlled environment
IV. to evaluate the effects of beneficial microorganisms on plant performance in
closed-loop hydroponics in controlled environment
Myco Madness microbial mix
(Humboldt nutrients)
Bacteria
Bacillus licheniformis
Bacillus azotoformans
Bacillus megaterium
Bacillus coagulans
Bacillus pumilus
Bacillus thuringiensis
Bacillus stearothermophilus
Paenibacillus polymyxa
Paenibacillus durum
Paenibacillus florescence
Paenibacillus gordonae
Azotobacter polymyxa
Azotobacter chroococcum
Pseudomonas aureofaciens
Yeast
Saccharomyces cerevisiae
Mycorrhiza
Glomus intraradices
Glomus mosseae
Glomus aggregatum
Glomus etunicatum
Glomus clarum
Glomus deserticola
Gigaspora margarita
Gigaspora brasilianum
Gigaspora monosporum
Trichoderma
Trichoderma harzianum
Trichoderma koningii
Root nodulation and seeds yield and quality of
plants inoculated with B. japonicum
in cooperation with School of Environmental Sciences
(University of Guelph)
Aim
 to evaluate the effect of inoculation with Bradyrhizobium
japonicum (soybean bacterial symbiont) in 2 hydroponic
systems (NFT and cultivation on rockwool) with urea as
alternative N-source to nitrate
Results
 Root inoculation did not influence plant performance,
regardless of the N-source
 Cultivation on rockwool positively influenced root
nodulation and plant growth and yield, without affecting
the composition of seeds, compared to NFT
 Urea improved root symbiosis but it reduced plant
growth and yield.
Paradiso et al., 2014 - Adv. in Space Res. 53: 574-84
Hydroponic cultivation of soybean for BLSSs:
the effect of nitrogen source and bacterial root symbiosis
Paradiso et al., Proceedings 63rd Intl. Astronautical Congress (IAF)
Materials & Methods
Growth chamber & environmental control
Lighting
Fog system
Osram PlantaStar ® 600W
Heating
Hydroponics
gullies
Air extraction
200300 cm
T/RH control probe
Materials & Methods
Environmental parameters
PPFD 500 mmol m-2s-1
light/dark 12/12 h
Temperature 26/20 °C
RH 6080%
Fertigation strategy
Modified Hoagland & Arnon ½ strength
N
K
P
Ca
Mg
S
Fe
Mn
Zn
Cu
B
Mo
mM
7.5
3.0
0.5
2.5
1.0
1.0
mM
60.00
7.40
0.96
1.04
7.13
0.01
(Wheeler et al., 2008)
Fresh
solution
0.25
Acid
EC target 1.2 dS m-1 C - 1.4 dS m-1
pH target 5.8
Deionised
water
Submerged pump
Recirculating solution
Materials & Methods
Inoculation protocol
SOYBEAN STERILIZED SEEDS
cultivar ‘Pr91m10’ (Pioneer Intl.)
Myco Madness mix preparation
1/4-strength Ringer’s solution
Cell count (5x105 cells/ml)
Thoma cell counting chamber, Microscope Nikon Eclipse E200
Seeds inoculation (12 h)
Washing in sterilized distilled water
Sowing
22 °C, darkness, 8 days
Seedling roots inoculation
Nutrient solution inoculation
3 times in 3 weeks
(Bashan, 1986. Soil Biol. and Bioch. 18(3): 297-301)
Materials & Methods
Measurements
Gas exchanges & Chlorophyll fluorescence
Infra red gas analyzer Walz HCM-1000
Junior – Pam Walz
Leaf greenness & Chlorophyll content
Hach 4000 Spectrophotometer
Spad Minolta 502
Leaf anatomical features
Rotative mnicrotome
Light & Epi-fluorescence microscopy on leaf thin section
Plant growth and seed yield and quality
Proximate composition of seeds (AOAC, 1995)
S. Pieters & E. Dulière, Institut Paul Lambin , Louvain, Belgium
Rhizosphere microbiome characterization
Microorganisms DNA Sequencing
C. Sheridan & D. Geelen, Dep. Plant Production, Gent University
Results
Photosynthesis and transpiration rate
Plants at reproductive phase,
Temperature 26 °C, RH 70%, PPFD 400 mmol m-2 s-1
( m mol CO 2 m -2 s-1 )
Net photosynthesis
14
12
10
8
6
4
2
0
C
I
C
I
Transpiration rate
-2 -1
(mmol m s )
2.5
2.0
1.5
1.0
0.5
0.0
Measurements of gas exchanges revealed higher CO2 assimilation and transpiration rate
in Inoculated plants compared to Control, starting from the beginning of flowering.
Results
Photosynthesis: Photochemical efficiency
(Plants at reproductive phase, Temperature 26°C, RH 70%, PPFD 400 mmol m-2 s-1)
0.9
F PSII
0.8
b
0.7
a
0.6
0.5
C
I
Similarly, Chl fluorescence measurements demonstrated better photochemical performance in treated
plants in the reproductive phase, with higher values of quantum yield of Photosystem II (FPSII)
Results
Leaf anatomy: stomata size and frequency
C
I
Changes in plant behaviour were also due to modifications in anatomical traits:
Inoculated plants showed higher frequency of smaller stomata than control,
which accounts for a better control of gas exchanges
Results
Leaf anatomy: lamina structure
C
100 micron
I
Moreover, Inoculated plants showed a mesophyll structure
(thicker layer of palisade parenchyma and larger intercellular space)
favouring the control of gas-exchanges
Results
Increase (% of Control)
Plant growth and Seed yield
Plant height (cm)
60
50
50
40
36,9
29,9
30
20
11,9
10
0
Plant height
40
LA max
Seed yield
30
Inoculation determined higher values of plant
20
C
10
I
height and leaf area. Consistently, seed yield was
higher in Inoculated plants compared to Control,
0
20
Vegetative
growth
40
60
80
Days after sowing
Flowering &
Fruit setting
100
Pod
Seed
filling drying
with no change in nutritional quality.
Results
Rhizosphere microbiome analysis
Christopher Sheridan & Danny Geelen
Dep. Plant Production, Gent University
C
I
Root exosphere
Root endosphere
C
I
At the end of the experiment, dominant microbial
composition differed between control and treated
plants in both the regions of rhizosphere:
In the exosphere, Control communities were
dominated by Chitinophagaceae bacteria and
Ascomycetes fungi, while they were much more
heterogeneous in Inoculated plants, with a mix of
Actinobacteria, Bacilli and Betaproteobacteria.
In the endosphere, Control was mainly composed
by Ascomycetes, while treated samples were
dominated by Ochrobactrum, which is a plant
growth-promoting taxon.
CONCLUSIONS
- Root inoculation of soybean with Myco Madness mix enhanced plant growth and
seed productivity, inducing functional anatomical modifications and enhancing
physiological performance compared to control
- Improved mineral nutrition and hormone activity by plant-growth promoting
bacteria (Ochrobactrum spp.) in the endosphere of treated plants may explain these
results
Inoculation with beneficial microorganisms can be considered as a tool to
improve performance of soybean in hydroponics, under controlled environment!