“The halophiles, named after the greek word for "salt-loving", are extremophiles that thrive in high salt concentrations.” Most halophiles are classified into the Archaea domain, Bacterial halophiles Some eukaryota, such as the alga Dunaliella salina or fungus Wallemia ichthyophaga

1. INDUSTRIALAND ENVIRONMENTAL
APPLICATIONS OF HALOPHILIC
MICROORGANISMS

2. HALOPHILES
“The halophiles, named after the greek word for "salt-
loving", are extremophiles that thrive in high salt
concentrations.”
Most halophiles are classified into the
•Archaea domain,
•Bacterial halophiles
•Some eukaryota, such as the alga Dunaliella salina or
fungus Wallemia ichthyophaga

3. HABITAT
• Habitats like soda lakes
• Dead sea,
• Carbonate springs,
• Salt lakes,
• Alkaline soils
• Thalassohaline,
• Athalassohaline,
• Many others favors the existence of halophiles.

4. TYPES
Halophiles are categorized as slight, moderate, or extreme, by the extent of their
halotolerance.
•Slight halophiles prefer 0.3 to 0.8 M (1.7 to 4.8% — seawater is 0.6 M or 3.5%),
E.g, Erythrobacter flavus
•Moderate halophiles 0.8 to 3.4 M (4.7 to 20%), E.g, Desulfohalobium
•Extreme halophiles 3.4 to 5.1 M (20 to 30%) salt content. E.g, Salinibacter ruber

5. ADAPTATIONS OF
HALOPHILES TO
THEIR
ENVIRONMENTAdaptations of halophiles to hyper saline environment
• Equilibration of cellular and environmental salt conc.
Synthesis of compatible organic solutes e.g ecotine.
Cell membrane adaptation like halorhodpsin.
High salt-in strategy like potassium chloride uptake.
Low-salt, organic salute-in strategy like glycin betaine.
Protein adaptations e.g having low hydrophobic amino
acids

6. APPLICATIONS
•Manufacturing of solar salt from seawater
•Production of traditional fermented foods.
•Utilization of enzymes produced by them.
•Use of specific compounds produced by them (ectoine, glycerol and others).
•Applications of unique compounds made by some halophiles example is
bacteriorhodopsin

7. PROCESS INVOLVING
HALOPHILES IN
BIOTECHNOLOGY AND
ENVIRONMENTAL
APPLICATIONS

8. 1. PRODUCTION OF SOLAR SALT
•Salt making by evaporation of seawater in shallow ponds. When salt starts
crystallizing, the brines become coloured red. Three types of halophilic
microorganisms contribute in the peoduction
Halophilic microorganisms Pigments Function
Extremely halophilic Archaea
(family Halobacteriaceae)
Carotenoids,
bacteriorhodopsin
Involve in the crystallization
of halite (salt).
Green flagellate alga
Dunaliella salina
β-carotene Carotenoid pigments absorb
light energy, raise the water
temperature, leading to
increased evaporation rates.
red halophilic bacterium
Salinibacter ruber
C40-carotenoid acyl
glycoside
Help in absorption of light.

9. THE BENTHIC MICROBIAL MATS
•The benthic microbial mats effectively seal off the bottom of the ponds,
preventing leakage of brine. When grown excessively, the cyanobacteria can
produce large amounts of polysaccharide slime, and results in the formation of
poor-quality salt.

10. 2. PRODUCTION OF
FERMENTED
FOODS
• LARGE AMOUNTS OF SALT ARE
USED IN THE PREPARATION OF
CERTAIN TYPES OF
TRADITIONALLY FERMENTED
FOODS.
Food Region Prepared
from
halophiles
jeotgal Korean seafood Halalkalicoccus jeotgali
fugunoko
nukazuke
Japanese Fish ovaries
in rice bran
Halococcus
thailandensis
nam-pla Thai Fish sauce Halobacterium
salinarum
Halococcus
thailandensis
Natrinema gari
kimchi Korean Fermented
vegetables
halophilic Archaea,
lactic acid bacteria

11. 3. ΒETA-CAROTENE
•Dunaliella salina and D. bardawil produces β-carotene.
•First pilot plant for mass culture of Dunaliella was set up in the mid-1960s
in the Ukraine.
•The pigment is found concentrated in small globules between the
thylakoids of the cell’s single chloroplast.
•FUNCTIONS
As an antioxidant,
As a source of pro-vitamin A (retinol)
As a food coloring agent

12. PRODUCTION OF ΒETA-CAROTENE BY DUNALIELLA
Suitable conditions to produce carotene
High light intensities, high salinity and nutrient
limitation; the slower the cells grow in the presence
of high irradiation levels, the more pigment is
formed.
Optimization of carotene production
1) To optimize growth and carotene production,
nutrient levels and pH should be carefully
controlled.
2) Nitrate limitation is induced to stimulate
carotenogenesis.
3) High yields 2% salt; at 15% salt only little
pigment was produced

13. 4. OSMOTIC SOLUTE
Most halophilic and halotolerant microorganisms produce or accumulate small
organic compounds intracellularly to provide osmotic balance with their
hypersaline environment.
For example
Glycine betaine, simple sugars such as sucrose and trehalose, different amino acid
derivatives and ectoine.

14. ECTOINE
It was first discovered in the haloalkaliphilic photosynthetic sulfur bacterium Ectothiorhodospira
halochloris.
Function
1)Protect many unstable enzymes, thereby increasing shelf life and activity of enzyme
preparations
2)Also nucleic acids against the detrimental action of high salinity, thermal denaturation, drying
and freezing.
3)It counteracts effects of ultraviolet UV-a-induced and accelerated skin ageing.
4)Ectoine also inhibits aggregation and neurotoxicity of Alzheimer’s β-amyloid.
5)Test the efficacy in inhalations against bronchial asthma.

15. PRODUCTION OF ECTOINE
Halophilic microorganisms Procedure
Ectoine by Halomonas elongata
Hydroxyectoine by Marinococcus M52
Bacterial milking, osmotic down-shock, bacteria react by secreting most of the
ectoine, collected by crossflow filtration techniques, purified. Salt is then
added to the bacteria to continue ‘milking’procedure.
Non-Halophilic microorganisms
Transgenic Escherichia coli Use of ‘leaky’ mutants that do not efficiently retain the compatible solutes
inside the cell
Plants
Tobacco plants Transformed with the ectoine genes of Halomonas elongata using an
Agrobacterium mediated gene delivery system showed improved root function
and photosynthesis at increased salinity.

16. 5. GLYCEROL PRODUCTION BY DUNALIELLA
Glycerol is produced by the alga Dunaliella. Cells grown in near-saturated NaCl
solutions may contain over 6–7 M intracellular glycerol.
High cost of the harvesting of the cells, no commercially feasible process was
developed.

17. 6.BACTERIORHODOPSIN
• Discovered in the early 1970s during studies of the purple membrane,
found within the cell membrane of Halobacterium salinarum
• It is a 25-kda protein that carries a retinal group bound by means of a
schiff-base (R₁R₂C=NR’.) to lysine-216.
• Structural Stability
 High salt concentrations
 Functions between 0 and 45°C in the pH range 1–11
 Tolerates temperatures of over 80°C in water up to 140°C when dry.
 It is stable to exposure to sunlight for years, and it resists digestion by
most proteases

18. USES ARE BASED ON
Conversion of light energy into chemical energy, Possible applications being ATP generation,
Conversion of sunlight into electricity
Desalination of seawater.
Others exploit the properties of its photocycle, especially the conversion of the B state
(absorbance maximum 570 nm) to the M state (420 nm) and using site-directed mutagenesis,
• The molecule can be optimized for such uses; for example, replacement of ASP by ASN at
position 96 prolongs the lifetime of the M intermediate a thousand-fold.

19. APPLICATIONS
1. Holographic storage
2. Construction of ‘bioelectronic’ elements of computer memories and
information processing units.
3. Ultrafast light detection, construction of artificial retinas, detection of motion
4. Construction of molecular transistors, molecular motors, artificial retinas and
molecular sensors.

20. 7. SALINE WASTEWATERS
TREATMENT
• AEROBIC TREATMENT SYSTEMS
• ANAEROBIC BIODEGRADATION PROCESSES
brine wastewater
10%
Percolators or
rotating discs to
improve aeration
Inoculum of
Halobacterium
salinarum
Utilities the brine
for growth by
osmosis
Pure water
saline and hypersaline
wastewater
Membrane
Bioreactor
Inoculum of
Haloferax
mediterranei
denitrification (
bioremediation
of nitrate and
nitrite)
Pure water

21. 8. ENZYMES FROM HALOPHILIC
MICROORGANISMS
Enzymes Source Species Optimum
condition
Function
Amylase Archaea Haloarcula sp Funnctions optimally at
4.3 M salt at 50°C, and
is stable in benzene,
toluene and chloroform
Degradation of lipid
Lipase Moderately
halophilic Bacteria
Salinivibrio sp active at 50°C Degradation of lipid
Nuclease H Bacteria Micrococcus
varians
at 60°C and12% salt Used in the production
of flavoring agent 5′-
guanylic acid (5′-GMP).
This enzyme degrades
RNA

22. 9. POLY-ΒETA-HYDROXYALKANOAT
•Poly-β-hydroxyalkanoate (PHA), a polymer containing β-hydroxybutyrate and β-
hydroxyvalerate units
•It is used for the production of biodegradable plastics (‘biologyical polyesters’)
• The technology for the manufacture of poly-β-hydroxyal-kanoate-derived
plastics (Biopol®) was developed by ICI in the UK, using polymer produced by
Cupravidus necator

23. POLY-ΒETA-HYDROXYALKANOATE PRODUCTION
BY HALOPHILIC BACTERIA
Halophilic bacteria Process of production of PHA.
H. mediterranei 1. Grows in sugars or starch as cheap carbon sources.
2. Growth is rapid at high salt concentration.
3. Cells lyse in the absence of salt.
4. Releasing the polymer.
5. Downstream processing.
6. Purification of the product.
Halomonas boliviensis 1. Acetate, butyrate or sucrose used as carbon sources.
2. It can accumulate the compound to up to 88% of its dry.

24. 10.EXOPOLYSACCHARIDES FROM HALOPHILES
• Bacterial extracellular polysaccharides have found different applications as gelling agents and
emulsifiers, and they are also used in microbially enhanced oil recovery.
Source Species Function
Archae Haloferax mediterranei 1. Produce sulfated acidic heteropolysaccharide, its
rheological properties are excellent and it is resistant to
extremes of pH and temperature. Its use to enhance oil
recovery from low productivity oil wells.
2. Archaeal membrane lipids may act as surfactants,
improving the oil carrying properties of the water.
Bacteria Halomonas species Producer of large amounts of an extracellular
polyanionic polysaccharide, a potent emulsifying
agent that exhibits a pseudoplastic behaviour.

25. IMMUNOMODULATING PROPERTIES OF THE SULFATED
POLYSACCHARIDE
• Immunomodulating properties of the sulfated polysaccharide of A.
Halophytica is of special interest: when administered orally in mice, it
significantly inhibited pneumonia induced by influenza virus H1N1

26. 11.BIOFUELS FROM
HALOPHILES
•The halophilic alga Dunaliella, considered as the raw material for biofuel
production.
•Catalytic pyrolysis of Dunaliella cell material at 200–240°C produces an oil-like
substance soluble in benzene.
•The overall process proved to be exothermic, so that most of the thermal energy
needed to initiate the reaction may be regained.
•Up to 75% of the cell material in an algae-seawater slurry could be converted to
extractable oil.

27. THANKS