Microbial Bioremediation of Radio-nuclides; different aspects/modes/kinds of microbial bioremediation that are direct enzymatic reduction, indirect enzymatic reduction, bio-sorption, bio-accumulation, bio-mineralisation, bio-stimulation and effective biological chelation; role of Genetic Engineering; essential genes in any Genetically Modified Organism (GMO) for this purpose

1. MICROBIAL
BIOREMEDIATION
OF RADIONUCLIDES

2. RADIONUCLEI: THE 21 st CENTURY NEMESIS

3. MAJOR RADIONUCLIDE CONTAMINANTSIN SOIL AND WATER

5. WHY CHANGE FROM
CONVENTIONAL
MANAGEMENT SYSTEM?
• NO EFFECT ON ACTIVITY AND HALF-LIFE
• UNACCEPTABLE COST OF MORE THAN A
TRILLION IN USA AND 50 MILLION
POUND IN UK.

6. MICROBIAL BIOREMEDIATION
• The physico –chemical transformation of radionuclides into stable
non radioactive isotopes by microbial species through the process of
microbial energy transfer
• Has effects on the action and halflige of radionuclides by influencing
their solubility , bioavailability and mobility through various metabolic
reactions.

7. DIFFERENT ASPECTS OF
MICROBIAL
BIOREMEDIATION
1. DIRECT ENZYMATIC REDUCTION
2. INDIRECT NON ENZYMATIC REDUCTION
3. BIOSORPTION
4. BIOACCUMULATION
5. BIOMINERALISATION
6. BIOSTIMULATION
7. EFFECTIVE BIOLOGICAL CHELATION

9. DIRECT ENZYMATIC
REDUCTION
• From high valency soluble high
radioactive state to insoluble lesser
valency with minimal or no radioactivity.
• Radionuclides act as final electron
acceptor in the ETS of anaerobic
respiratory pathway.
• Formate, acetate, lactate and H2 as
electron donor.
• Shewanella putrefaciens; Desulfovibrio
desulfuricans; Geobacter sulfurreducens;
Clostridium sp
• Periplasmic cytochromes play a vital role

10. INDIRECT NON-
ENZYMATIC
REDUCTION
• Coupled reactions between oxidation of
organic acids produced diring excretion of
microbes with reduction of radionuclides
• Precipitate out as oxide or hydroxide
minerals
• Sulfate refucing and dissimilatory metal
reducing bacterias like Desulfobacterales,
Desulfovibrionales

11. BIOSORPTION AND
BIOACCUMULATION
• Biosorption is the sequestration of
positively charged radionuclides to the
cell membrane.
• Involves transport across cell membrane,
complexation,ion exchange precipitation
and physical adsorption
• Functional groups charecterised in the
cell wall is responsible for metal biomass
binding like carboxyl, imidazole,amino,
phenol, hydroxyl,etc.
• Bioaccumulation is the uptake of
radionuclides into the cell where they are
retained by complexations with
negatively charged intracellular
components

12. BIOMINERALISATION/
BIOPRECIPITATION
• Precipitation of radionuclides as stable
biogenic minerals through complexations
with microbial ligands which provide a
nucleation site
• Microbes hydrolyse organophosphate in
the presence of uranium to perform
biomineralisation
• Serratia and Citrobacter genera does
cleavage of glycerol phosphate to
precipitate with uranyl ion as
Chernikovite
• Ex-situ researches has shown to
precipitate 73-95% total uranium in 120
hours;max activity at pH 5.0 and 7.0

13. BIOSTIMULATION AND MICROBIAL
CHELATION
• Novel methods to increase the
bioremediation by providing specific
stimulants
• Addition of ethanol helps the bioreduction of
nitrate for aerobic denitrifying bacteria
• Hydrolic recirculation systems to prevent the
clogging of nitrogen gas, aluminium,
calcification precipitation and biomass.
• U (VI)and Nitrate conc to 5 mm and 0.5mm in
an year and to.126mmin 2years which is
below the EPS’s recommended level.
• Microbial ligands, sequestering agents and
siderophores play vital role in solubility and
bioavailability of radionuclides.
• Siderophore enterobactin effectively
solubilized hydrous PuO2
• EDTA and citric acid reduced radionuclides to
Pu(IV) and Th(IV) before formation.
• Citric acid forms complexes with almostany
heavy metals
• Uranyl citrate – recalcitrant to aerobic
bacteria; required photodegradation to
precipitate stable UO3.xH2O

14. WHY ‘GE’ and ‘GMO’?
• Direct enzymatic reduction of radionuclides
are inhibited in in-situ due to the presence of
heavy metals which inhibit nitrate reduction.
• Species poor radioresistance
• Endogenous Resistance genes
• Biosorption and bioaccumulatiin are
saturation depended and their proliferation
and biofilm formation gets inhibited due to
high radiation
• Bioprecipitation eventhough saturation
independent requires increased no:of
complexes responsible for transport of
radionuclides inside the cell.
• Microorganisms are not always competent to
radionuclides limiting their bioavailability to a
great extent. Uranium hasbgotno metabolic
function and hence its entry is pirely due to
radiotoxicity increasing membrane
permeability and some radionuclides have
resemblance to dietary elements which are
absorbed accidentally
• Citric acid, the most effective biological
chelator ;uranyl citrate complex is recacitrant
along with metal uranyl complexes
• Competition from anions,toxic metals, organic
compounds and other chelating agents in the
environment.

15. ADVANCEMENTS IN GENETIC ENGINEERING
• Incresed rate of biosorption by equipping
bacterial cell surface with metal binding
polypeptides and enhanced bioaccumulation
through introducing special metal
transporters to the cytoplasm
• A recombinant strain of E.Coli was generated
with 5 times sorption ability forU(VI) by
altering the transporter genes
nixA(Helicobacterpylori) and merTP(Serratia
marcescens).
• The gene dcuB from G.sulfurreducens,which
encodes a fumerate transporter was
engineered in G.mettallireducens tofunction
as terminal electron acceptor
• NiCoT gene from Rhodopseudomonas
palustriis removed 85% cobalt with
recombinant E.coli.
• Recombinant E.coli precipitated more than
90%uranyl carbonate when phoK gene was
introduced from Spingomonsp
• merA gene incorporated from E.coli to D.R
provided the GMO to utilise carbon and the
energy from catabolism of toluene and Hg for
biotransformation of Hg(II) to less toxic Hg(I)

16. ESSENTIALS IN THE GENOME OF A GMO
PROTOTYPE FOR IN-SITU OPERATION
1. A Host with high radioresistance
2. Degradation genes
3. Reporter gene
4. Suicide genes
5. Bioluminescence gene

17. DEINOCOCCUS
RADIODURANS
• Most radioresistant organism in the
planet
• 5000Gy-no loss of viability; 15000Gy with
37% viability
• Rapid DNA repair and high proliferation
rate
• Natural ability to detoxify Cr(VI);U(VI)and
Tc(VI)

18. DEGRADATION GENES
The genes that code for enzymes,carrier
proteins, chelators, siderophores,etc all that
functions for bioremediation of radionuclides
are degradation genes.
It is the positive regulation of these genes
introduced in the GMO that does the
bioremediation.

19. THANK YOU...