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Bioremediation Of 
Syeda Sakeena Gilani 
Mphil Leading to PhD, 
Microbiology
Lead - An Introduction 
• bluish - gray colored solid, nonessential and toxic heavy metal 
• Primary form galena PbS--0.001-0.002% of the earth's crust 
• with an oxidation state of 0, +2, +4 
• Pb2+most stable form 
• persistent environmental pollutant with half life of approximately 
50,000 years 
• Half-life in blood is 28-36 days
2007 Estimate Of Lead Production
Lead Reserves In Pakistan
Sources Of Lead 
• Natural sources 
▫ windborne dusts 
▫ sea and salt lake aerosols 
▫ forest fires 
▫ volcanic eruptions 
• Anthropogenic sources 
▫ Minning 
▫ Smelting 
▫ Fuel combustion 
▫ paint and coatings 
manufacturers 
▫ battery manufacturing
Lead Compounds 
Inorganic lead Organic lead 
• Lead powder 
• Acetates 
• Carbonates 
• Sulphates 
• Chlorides 
• Phosphates 
• Found in dyes, paints and 
pesticides 
• low soluble in water 
• Tetramethyl lead 
• Tetraethyl lead 
• Used as anti-knocking agents 
in gasoline products 
• Highly soluble in organic 
solvents 
• Can be absorbed by skin 
• Accumulated in fat
Lead Intake And Absorption 
• Inhalation: small sized lead particles~40% 
• Ingestion: absorption through GIT 10-15% for adults, 50% in children 
• Tetraethyl lead, by liver action turns into triethyl leadactive toxic
Permissible limits for lead in drinking water EPA 
0.015 mg/L and for wastewaters is 0.1 mg/L
LEAD DETECTION RECIPE 
• Ingredients 
• 1 Cotton swab 
• 1 Disposable plastic cup 
• 1 Small disposable white 
plastic plate 
• Rubbing alcohol (ethanol) 
• Water (preferably de-ionized) 
• Vinegar 
• Lead Indicator: Sodium 
rhodizonate
Lead Detection Procedure 
• Lightly soak a cotton swab with rubbing alcohol 
and rub against the item you wish to test for about 
1 minute. 
• Let swab air dry to evaporate the alcohol. 
• In a cup mix the small pinch of Lead Indicator with 
a¼cup of water. Label this solution – “Indicator”. 
• On a white plastic plate mix a drop of the Indicator 
solution with a drop of vinegar. The resulting drop 
solution mixture should be colorless to yellow.
Contt.. 
• Dip the dried swab into the drop solution mixture 
and allow it to soak up the liquid. If Lead is 
• present on the swab a pink-red coloration should 
appear on it within 1-10 minutes (depending on 
the Lead concentration). 
• Repeat steps 4 and 5 with a clean (untreated) swab 
which will serve as a reference to help distinguish 
the color-change.
• lead reacts with sodium sulfide to produce a gray 
or black color. Lead reacts with sodium 
rhodizonate to give a pink or red color.
Environmental Performance Index 
According to EPI 2014 
Pakistan ranks 148 out of 178 countries with a 
score of 34.58. 
In air quality, the score is 23.02 ranking 175
Bioremediation of Lead 
• Biosorption/Complexation 
• Precipitation 
• Bioaccumulation 
• Rhizofiltration
Biosorption/Complexation 
• By extracellular PolymerP. marginalis 
• Results in the immobilization and sequestration 
of Pb. 
▫ ion exchange reactions with peptidoglycan and teichoic 
acid 
▫ precipitation through nucleation reactions, 
▫ complexation with nitrogen and oxygen ligands
Advantages of biosorption 
• highly selective, more efficient, easy to operate, 
and 
• hence cost effective for the treatment of large 
volumes of wastewaters containing low metal 
• concentrations
Precipitation 
• Formation of insoluble Pb complexes with 
sulfide, hydroxide, and carbonates. 
• Kiebsiella aerogenes formed galena (PbS) 
• C. metallidurans removed Pb by formation of 
Pb-hydroxide or carbonate complexes.
Rhizofiltration 
• Heavy metals are primarily retained within the roots. 
• contaminants don’t get translocated to the shoots. 
• The terrestrial plants have a fibrous and elongated 
root system. 
• Examples: sunflower, indian mustard, tobacco, rye, 
spinach, and corn 
• sunflower significantly reduced lead concentrations 
with in 1hr after treatment.
Bioaccumulation 
• Metallothioneins - two cysteine residues, achieved a 
33%increase in Pb2+ adsorption 
• Significant intracellular accumulation of lead is due to 
sequestration of lead by SmtA metallothionein 
• B. megaterium accumulated Pb intracellularly as dense 
granules in the cytoplasm. 
• Example: significant intracellular bioaccumulation of lead 19 
and 22 mg/gram dry weight in case of Salmonella choleraesuis 
strain 4A and Proteus penneri strain GM-10 respectively
Interesting Fact-Use of Fish Bones 
• Fish bones are made of the phosphate mineral 
apatite, which readily combines with lead to form 
pyromorphite, a stable crystalline mineral that 
can’t be absorbed by the human digestive system 
• Lead concentrations in soil leachate treated with 
fish bones dropped from 0.28 mg/L to 0.00065 
mg/L within weeks
Pb Resistance Operon 
The plasmid-encoded lead-resistance gene cluster of C. 
metallidurans CH34, pbrTRABCD consists of six genes 
that code for: 
• PbrR, a transcription factor belonging to the MerR family 
• PbrT, a putative Pb2+ uptake Protein 
• PbrA, a P1B-type ATPase 
• PbrB, undecaprenyl pyrophosphate phosphatase 
• PbrC, a signal peptidase (not necessary for resistance) 
• PbrD, a putative intracellular Pb-binding protein
The genes are arranged in two opposing operons, one containing 
the pbrTR genes and the other the pbrABCD genes
A Study Conducted In India 
• Determining the heavy metal lead removing 
ability of microbes isolated from the industry 
effluents 
• Bacillus licheniformis-89.45% 
• Salmonella typhi 92.15% 
• Pseudomonas fluorescence 94.32% 
• Escherichia coli 86.73%
For Further Reading…
Types of lead bioremediation

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Types of lead bioremediation

  • 1. Bioremediation Of Syeda Sakeena Gilani Mphil Leading to PhD, Microbiology
  • 2. Lead - An Introduction • bluish - gray colored solid, nonessential and toxic heavy metal • Primary form galena PbS--0.001-0.002% of the earth's crust • with an oxidation state of 0, +2, +4 • Pb2+most stable form • persistent environmental pollutant with half life of approximately 50,000 years • Half-life in blood is 28-36 days
  • 3. 2007 Estimate Of Lead Production
  • 4. Lead Reserves In Pakistan
  • 5. Sources Of Lead • Natural sources ▫ windborne dusts ▫ sea and salt lake aerosols ▫ forest fires ▫ volcanic eruptions • Anthropogenic sources ▫ Minning ▫ Smelting ▫ Fuel combustion ▫ paint and coatings manufacturers ▫ battery manufacturing
  • 6. Lead Compounds Inorganic lead Organic lead • Lead powder • Acetates • Carbonates • Sulphates • Chlorides • Phosphates • Found in dyes, paints and pesticides • low soluble in water • Tetramethyl lead • Tetraethyl lead • Used as anti-knocking agents in gasoline products • Highly soluble in organic solvents • Can be absorbed by skin • Accumulated in fat
  • 7. Lead Intake And Absorption • Inhalation: small sized lead particles~40% • Ingestion: absorption through GIT 10-15% for adults, 50% in children • Tetraethyl lead, by liver action turns into triethyl leadactive toxic
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  • 13. Permissible limits for lead in drinking water EPA 0.015 mg/L and for wastewaters is 0.1 mg/L
  • 14. LEAD DETECTION RECIPE • Ingredients • 1 Cotton swab • 1 Disposable plastic cup • 1 Small disposable white plastic plate • Rubbing alcohol (ethanol) • Water (preferably de-ionized) • Vinegar • Lead Indicator: Sodium rhodizonate
  • 15. Lead Detection Procedure • Lightly soak a cotton swab with rubbing alcohol and rub against the item you wish to test for about 1 minute. • Let swab air dry to evaporate the alcohol. • In a cup mix the small pinch of Lead Indicator with a¼cup of water. Label this solution – “Indicator”. • On a white plastic plate mix a drop of the Indicator solution with a drop of vinegar. The resulting drop solution mixture should be colorless to yellow.
  • 16. Contt.. • Dip the dried swab into the drop solution mixture and allow it to soak up the liquid. If Lead is • present on the swab a pink-red coloration should appear on it within 1-10 minutes (depending on the Lead concentration). • Repeat steps 4 and 5 with a clean (untreated) swab which will serve as a reference to help distinguish the color-change.
  • 17. • lead reacts with sodium sulfide to produce a gray or black color. Lead reacts with sodium rhodizonate to give a pink or red color.
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  • 19. Environmental Performance Index According to EPI 2014 Pakistan ranks 148 out of 178 countries with a score of 34.58. In air quality, the score is 23.02 ranking 175
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  • 22. Bioremediation of Lead • Biosorption/Complexation • Precipitation • Bioaccumulation • Rhizofiltration
  • 23. Biosorption/Complexation • By extracellular PolymerP. marginalis • Results in the immobilization and sequestration of Pb. ▫ ion exchange reactions with peptidoglycan and teichoic acid ▫ precipitation through nucleation reactions, ▫ complexation with nitrogen and oxygen ligands
  • 24. Advantages of biosorption • highly selective, more efficient, easy to operate, and • hence cost effective for the treatment of large volumes of wastewaters containing low metal • concentrations
  • 25. Precipitation • Formation of insoluble Pb complexes with sulfide, hydroxide, and carbonates. • Kiebsiella aerogenes formed galena (PbS) • C. metallidurans removed Pb by formation of Pb-hydroxide or carbonate complexes.
  • 26. Rhizofiltration • Heavy metals are primarily retained within the roots. • contaminants don’t get translocated to the shoots. • The terrestrial plants have a fibrous and elongated root system. • Examples: sunflower, indian mustard, tobacco, rye, spinach, and corn • sunflower significantly reduced lead concentrations with in 1hr after treatment.
  • 27. Bioaccumulation • Metallothioneins - two cysteine residues, achieved a 33%increase in Pb2+ adsorption • Significant intracellular accumulation of lead is due to sequestration of lead by SmtA metallothionein • B. megaterium accumulated Pb intracellularly as dense granules in the cytoplasm. • Example: significant intracellular bioaccumulation of lead 19 and 22 mg/gram dry weight in case of Salmonella choleraesuis strain 4A and Proteus penneri strain GM-10 respectively
  • 28. Interesting Fact-Use of Fish Bones • Fish bones are made of the phosphate mineral apatite, which readily combines with lead to form pyromorphite, a stable crystalline mineral that can’t be absorbed by the human digestive system • Lead concentrations in soil leachate treated with fish bones dropped from 0.28 mg/L to 0.00065 mg/L within weeks
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  • 31. Pb Resistance Operon The plasmid-encoded lead-resistance gene cluster of C. metallidurans CH34, pbrTRABCD consists of six genes that code for: • PbrR, a transcription factor belonging to the MerR family • PbrT, a putative Pb2+ uptake Protein • PbrA, a P1B-type ATPase • PbrB, undecaprenyl pyrophosphate phosphatase • PbrC, a signal peptidase (not necessary for resistance) • PbrD, a putative intracellular Pb-binding protein
  • 32. The genes are arranged in two opposing operons, one containing the pbrTR genes and the other the pbrABCD genes
  • 33. A Study Conducted In India • Determining the heavy metal lead removing ability of microbes isolated from the industry effluents • Bacillus licheniformis-89.45% • Salmonella typhi 92.15% • Pseudomonas fluorescence 94.32% • Escherichia coli 86.73%

Editor's Notes

  1. Duddar mine- in Balochistan 1.6 million tonnes of lead
  2. lead metal ions present in the textile dye effluent cause toxicity in receiving aquatic environment by interacting with nucleic acids, by binding to essential respiratory proteins (Vallee and Ulmer 1972) and by displacing Ca2+ and Zn2+ in protein
  3. . Longbone Radiograph of Hands - “lead lines” shown as increased density on the metaphysis growth plate of the proximal segments of phalanges and distal segments of ulna and radius in a five-year-old male with radiological growth retardation and blood lead level of 37.7µg/dL.
  4. Three main mechanisms for the binding of metals to bacterial cell walls are known: ion exchange reactions with peptidoglycan and teichoic acid precipitation through nucleation reactions, complexation with nitrogen and oxygen ligands Gram-negative bacterial cell membranes are lower in these components and are poorer metal absorbers Gram-positive species, such as Bacillus and Clostridium, and Lactobacillus have high adsorptive capacity due to high peptidoglycan and teichoic acid content in their cell walls.
  5. Pb2+ is exported from the cytoplasm by PbrA and then sequestered as a phosphate salt with the inorganic phosphate produced by PbrB.