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ENDEGRADEEndophytic bacteria for improving
phytoremediation

• Fiona Porteous Moore,
• Colin Campbell, Edward Moore
• E.U. partners

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ENDEGRADE7 project partners

• SCOTLAND BELGIUM IRELAND DENMARK
  
• Macaulay IT Carlow
• Institute
• LUC VITO DEC-NV
  DTU NERI
• 
  (INDUSTRIAL
• PARTNER)

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• PHYTOREMEDIATION
• Ideally plants must have ? deep roots
• ? perennial
• ? large water use
• Plants ? breakdown/volatilise
  compounds
• ? store them in
  leaves/shoots
• 4 methods ? phytovolatilisation
• ? phytoextraction
• ? rhizofiltration
  

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Problems with some current remediation strategies

• Phytoremediation
• pollutants kill the plants
• volatilisation through leaves
• partial degradation leading to toxic products
• Soil bioremediation
• indigenous population overcomes introduced
  degraders
• nutrient applications often needed
• pollutants can be taken up by plants faster than
  soil microbes can degrade them

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Endegrade - the concept

• enhance plant survival
• enhance degradation
• reduce volatilisation

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Project aims

• Isolate identify bacteria from Willow and
  Poplar
• Screen for natural degradation potential for
  target compounds
• Equip endophytic bacteria with degradation
  plasmids
• Assess re-colonisation and phytoremediation
  efficiency
• Risk assessment for field use

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• What is known about endophytic bacteria?
• neutral or beneficial effects on the plant
• - direct plant growth-promoting
  activity
• - N2-fixation
• - disease suppression
• - enhanced pest control
• exist in all plant species examined
• 103 - 105 cfu/gm plant tissue
• highest numbers observed in the roots
• bacteria remain localised in specific tissues

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• We do not know
• community dynamics diversity
• colonisation potential
• plant specific relationships?

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Pollutants problematic for phytoremediation...
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Bacterial endophyte isolates analysed to date
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Relative frequencies of bacterial isolates,
classified to the genus level, on the basis of
16S rDNA sequence analysis
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Relative frequencies of isolates classified as
Pseudomonas species, based upon 16S rDNA sequence
analysis
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(No Transcript)
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Distribution of endophytic bacterial isolates in
Poplar
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Effect of tree species
Multivariate analysis - presence / absence of
isolates in different species of trees.
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Napthalene degradation in Pea (proof of concept)
Work by Germaine et al, IT Carlow
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Plant protection

• Iris VM1450 - 2,4-D protection (Germaine et al,
  IT Carlow)
• Pea PCB degraders - 4-chlorobiphenyl
  protection (Germaine et al, IT Carlow)
• Lupine VM1330- Toluene protection, reduced
  volatilisation, enhanced growth (Barac et al,
  LUC)
• Toluene TCE degrader constructs from Brassica
  napus - to test in plantae degradation (Borremans
  et al, VITO)

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Re-inoculation

• K.Germain et al, FEMS Microbiol.Ecol, in press.
• 3 endophytic isolates gfp/Kan marked and
  re-inoculated into Poplar (original host)
• All strains colonised roots, and 2 strains
  colonised stems and leaves after 10 weeks

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VM1449 (Ps. veronii) colony on root xylem of
poplar tree 10 weeks after inoculation x1000
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• Poplar used in phytoremediation field
  trial in Belgium

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Current work

• Endophytic construct in Poplar exposed to 2,4-D -
  does in degrade/protect?
• 2,4-D degradation genes colonisation
• Seasonal community dynamics of endophytes -
  affects inoculation time/method
• intra/inter species variation, seasonal dynamics,
  compartmentalisation

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Acknowledgements
Macaulay Institute - Renate Wendler, Duncan
White NERI - Denmark - Uli Karlson DTU -
Denmark - Stefan Trapp VITO - Belgium - Daniel
van der Lelie, Brigitte Borremans LUC-
Belgium - Jaco Vangronsveld, Licy Oeyen,
Tanja Barac IT Carlow-
Ireland - David Dowling, David Ryan,
Keiran Germaine, Elaine
Keogh DEC NV- Belgium - Siegried DHaene,
Gunther de Becker