Microorganisms

involved in bioleaching

Lecture #2

Introduction

• Bioleaching is the biological

conversion

of an insoluble metal

sulfides into a water soluble forms.

• Metal sulfides are oxidized to metal

ions and sulfate by aerobic,
acidophilic

Bacteria

and

Archaea

.

• Bioleaching involves

chemical

and

biological

reactions.

Colony of bacteria on the

pyrite surface

Domains Archeaa and

Bacteria

• Microorganisms can be subdivided into:

Bacteria

Temperature optima

Mesophic

20-40

o

C

Moderate

thermophilic

40-60

o

C

Extremely

thermophilic

above 60

o

C

Moderate termophilic

bacteria

• Most industrial heap and tank

bioleaching operation run below
40

0

C.

• All acidophilic metal sulfide oxidizing

microorganisms oxidize Fe

2+

and sulfur

compounds.

• Acidithiobacillus ferrooxidans
• Acidithiobacillus thiooxidns
• Acidi thiobacillus caldus

Acidithiobacillus

ferrooxidans

magnified 30,000 times

Acidithiobacillus ferroxidans

colony

Moderate thermophilic

bacteria

Acidithiobacillus

ferrooxidans

• Acidithiobacillus ferrooxidans

, formerly

name

Thiobacillus ferrooxidans

, was the

first described by Colmer and Hinkle at
1947year.

• It derives energy from the oxidation of Fe

2+

ions

and various sulfur compounds : S

0,

S

2-

, S

2

O

32-

(thiosulfate).

• Since

A. ferrooxidans

lives in an

environmental with high metal ion
concentrations, it is adapted to many cationic
metals.

Acidithiobacillus

ferrooxidans

• Acidithiobacillus ferrooxidns is a

Gram-negative

, rod-shaped acidiphilic

bacterium.

• It is an obligate

chemolithoautotrophic

bacterium that

use elemental sulfur or reduced sulfur
compounds as well as ferrous ions a
source of energy and carbon dioxide
as a source of carbon.

Oxidation reaction of A.

ferrooxidans

• Acidithiobacillus ferrooxidans

is also

able to directly oxidize:

U

+4

to U

+6

;

Cu

+

to Cu

+2

;

Mo

+5

to Mo

+6

;

As

+3

to As

+5

Acidithiobacillus thiooxidans

• Acidithiobacillus thiooxidans

was

described by Waksman and Joffe in 1922
year.

• It grows

autotrophically

with various sulfur

compounds e.g. elemental sulfur,
thiosulfate, and tetrathionate.

• Acidithiobacillus thiooxidant does not

oxidize pyrite (FeS

2

) and Fe

2+

.

• Bacteria has been isolated from soil, sulfur

deposits, and mine waste.

Sulfur-oxidizing bacteria

• Reduced forms of sulfur, such as sulfide

(S

2-

) are oxidized to sulfate (SO

42-

).

• Elemental sulfur is oxidized to sulfuric

acid.

0.125 S

8

+ 1.5 O

2

+ H

2

O H

2

SO

4

• Ferrous ions (Fe

2+

) are oxidized to ferric

ions (Fe

3+

).

2Fe

2+

+ 0.5 O

2

+ 2H

+ 

2Fe

3+

+ H

2

O

Leptospirillum sp.

(magnification 20 000x)

A typical spiral shape of a strain of

Leptospirllum

from

Fairview mine (South Africa)

Leptospirillum

ferrooxidans

• The mesophilic species was isolated

by Markosyan at 1972 year.

• They grow obligatelly

chemolithoautotrophically

and derive

energy only from the oxidation of
Fe

2+

but not from sulfur compounds.

• CO

2

is fixed by mean of the Benson-

Calvin Cycle.

Extrimely Thermophilic

Archaea

• All extremely thermophilic metal

oxidizing Archaea belong to the family

Sulfolobaceae.

• They have optimum growth

temperatures between 65 and 95

0

C

and pH optima of around pH 2.

• Acidianus brierleyi

,

Sulfolobus

acidocalcaldarius

and

Metallospharra

species are very efficient ore leaching .

Acidianus brierleyi

• This strain was isolated from a hot

springs in Yellowstone National Park,

USA.

• It was originally described as

Sulfolobus

brieleyi

and reclassified as

Acidianus

brieleyi

by Segerer at 1986 year.

• The facultative

anaerobic

, facultative

chemolithoautotrophic

organism uses

metal sulfides, elemental sulfur, and

organic compounds as substrates.

Sulfolobus

• Mesophilic bacteria have been less

successful in oxidizing chalcopyrite
mostly due to passivation of the
mineral surface during its
bioleaching.

• In order to overcome this difficulty,

attention was focused on the
thermophilic bacteria (archaea) (65-
80

o

C).

Bioleaching fungi

• Aspergillus niger
• Spergillus niger
• Penicillum simplicissimum
• Saccharomyces cervisiae
• Yarrowia lipolytica

Citric and oxalic acids are produced by

fungi

Microbial analysis

• Microorganism communities can be

analyzed using:

 Microscopic techniques
 Cultivation technique
 Immunological techniques
 Nucleic-acid based molecular

techniques

Fluoromicroscope

method

• Total cell number can be determined

by counting cell under a fluorescence
microscope after application of
nucleic acid-staining fluorochromes
(acridine orange, DAPI)

Molecular technique

• Over the last years, nucleic-acid base

molecular techniques have been
increasingly used to identify and
quantify microorganisms in the
environment and technical applications.

• It is based on the extraction of DNA from

a culture, followed by the amplification
of DNA using the Polymerase Chain
Reaction (PCR), and finally an analysis of
DNA amplification products.

Growth cycle

The lag phase marks the initial period of time when
the cell adjust to their new surroundings.

The exponential phase for Escherichia coli -20 to
30minutes.

In the stationary phase, there is no net increase or
decrease in cell number.

Substances toxic for

bacteria

Certain substances are potentially toxic to the

bacteria. These include:

• Thiocyanate

and

cyanide

at very low concentrations;

• Bactericides

and

fungicides reagents

that are

normally used for water treatment;

• Oil, grease

and

degreasing

compounds;

• Chloride concentrations

above 7 g/L, which inhibit

bacterial activity;

• Arsenic

at high concentrations (although the culture

is tolerant to arsenic(V) concentrations as high as 20

g/L, a high concentration of

arsenic(III)

can be toxic)

 

Active bacteria

The key factors maintaining an active

bacteria:

1. Adequate nutrient supply
2. A minimum dissolved oxygen

concentration of 1.5 ppm

3. Sufficient supply of CO

2

to generate

biomass

4. Maintenance of operating temperature at

35

o

C (40,50 70

o

C)

5. Operating pH in the range 1.6 to 2.0

Iron(II) ions biooxidation

• The iron(II) ions oxidation is

connected with electron transfer
from the outer cell wall of bacteria to
the cell inside.
The electron pathway has implicated:

Rusticyanin
Cytochrome „c”
Cytochrome „a”

Electron transport to the inside of bacterial cell

Sulfide Mineral

Biooxidation

Summary

• Microorganisms are found everywhere

on the Earth’s surface where water is
at least temporarily available.

• In acid environments a number of

acidophilic

chemolithoautotrophic

bacteria exist. (Acidithiobacillus
ferrooxidans)

• Typical growth curve for a bacteria

population shows

four phases

.

Lab. Monday 8.12. 13

15

C-6

Building C-6 room 127

Glasses and lab coat obligatory