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Adaptations of microorganisms to extreme
conditions
Silvano Onofri
and Laura Selbmann, Giuliano Scalzi, Laura Zucconi, Daniela Isola
CAREX Summer School
Pieve Tesino 27/06-04/07/2010
Organisms living in extreme
environments
Daniel Prieur
Laboratoire de Microbiologie des Environnements extrêmes
UMR 6197 (CNRS,IFREMER,UBO)
Université de Bretagne Occidentale, Brest, France
Be prepared to some scale changes
• From AU to micrometers
• From Gy to hours and minutes
• From Jupiter mass to 10-12 g
• No models, but true creatures
Organisms living in extreme
environments
• What is a living organism?
• What is an extreme environment?
Living organisms: diversity of creatures
Deer
Shark
Hibiscu
Heron
Shrimp
Microbe
Living organisms: diversity of sizes
Bacteriophages: 80 nm
Blue Whale: 30 m
Living organisms are distributed
within 3 Domains
All living organisms
• 3 Domains:
– Bacteria
– Archaea
– Eukarya
• 2 cellular Structures:
– Prokarotes
– Eukaryotes
• 1 basic Unit:
– The Cell
Prokaryotic cells
Eukaryotic cells
Same structure for
Deers or Yeasts !
How do cells work?
Environments on Earth
Life is influenced by physiochemical parameters
Pressure
pH
Metabolic ressources
Salinity
Temperature
Ionizing radiations
Toxic compounds
Survival
Death
Resting stages
Death
A
Low values
Extreme
B
C
Conditions
D
High values
Extreme
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Extremophiles and Extremotolerants
Extremophiles are organisms that thrive in or even may require
physical or chemical extreme conditions that are detrimental to
the majority of life on Earth...
Usually extreme conditions are defined extreme from an
anthropocentric point of view.
Actually extremophilic and extremotolerant microorganisms are
able to live in conditions that are extreme from a biocentric point
of view (i.e. close to the edge of life)
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…Many kinds of stresses can define an extreme
environment:
•Dryness
•high or low temperature
•high radiations
•hypersaline environments
•acidic or alkaline environments
•high pressure
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Most of extremophiles and extremotolerant organisms already
known, are Bacteria, Archaea and Cyanobacteria…
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Extreme Prokaryotes
Haloquadratum walsbyi
First discovered in 1980 by A. E. Walsby in the Gavish Sabkha, a coastal
Brine pool on the Sinai Peninsula in Egypt, this archaeon was not
successfully cultured until 2004. While attempting to culture
Haloquadratum walsbyi, a new species, Haloarcula quadra, was found.
Haloquadratum are remarkable for their shape, motility, and relative
abundance in halophilic environments.
The fireball from the abyss
(Pyrococcus abyssi)
Peptide fermenter
Optimum temperature: 96°C
Doubling time: 25 minutes
A Methane-maker Ball from Hell
(Methanococcus infernus)
With hydrogen and
carbon dioxide only,
this organism produces
methane,
optimally at 85°C.
The Smoker ’s fire lobe
Pyrolobus fumarii
World record for high temperature
Optimum 106°C, maximum 113°C
Survives 1 hour in an autoclave at 120°C
e- donnor: H2
e- acceptor: NO3-, S2O32-, O2
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Pyrolobus fumarii
Pyrolobus fumarii is a species of Archaea microbes known for its ability to
survive at extremely high temperatures. It was first discovered in 1997 in a black
smoker hydrothermal vent at the Mid-Atlantic Ridge, setting the upper
temperature threshold for known life to exist at 113 degrees C. Strain 121, a
microbe from the same family found at a vent in the Paceific ocean, survived and
multiplied during a 10-hour blast in a 121°C autoclave. It was finally killed at a
temperature of 130 degrees C.
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Strain 121
Strain 121 is a single-celled microbe, of the domain Archaea. First discovered
200 miles (320 km) off Puget Sound in a hydrothermal vent; it is a
hyperthermophile, able to survive and reproduce at 121 °C (hence its name). It is
the only known form of life that can tolerate such incredibly high temperatures.
130 °C (266 °F) is proven to be only bacteriostatic for Strain 121, meaning that
although growth is halted, the archaeon remains viable, and can resume
reproducing once it has been transferred to a cooler medium.
Kashefi, Kazem, Lovley, Derek R. (2003). "Extending the upper temperature limit for life". Science 301 (5635):
934.
Halobacterium salinarum
• To survive in extremely salty
environments, this archaeon—as with
other halophilic Archaeal species—
utilizes compatible solutes to reduce
osmotic stress. Potassium levels are
not at equilibrium with the environment,
so
H.
salinarum
expresses
multiple active transporters which
pump potassium into the cell
Deinococcus audaxviator
from deep biosphere in South-Africa mines is able
to use radiations from uranite for its metabolic needs
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Nowadays many microbial eukaryotes are known
as very resistant extremophilic or extremotolerant
organisms
Eukaryotic microbial life may be found actively
growing in almost any extreme condition where
there is a sufficient energy source to sustain it,
with the exception of very high temperatures
Possibly resistance to extreme stress conditions in
microbial eukaryotes is underestimated and in
some cases it could overcome limits of prokaryotes
mainly under multiple combined stressing factors
We can find extremophilic and
extremotolerant microbial
eukaryotic organisms in different
groups, e.g.
•Red algae
•Green algae
•Diatoms
•Protozoa
•Fungi
High temperatures molecular
adaptations
No single cellular feature is
responsible for extreme heat
resistance
• Aminoacid composition of proteins (hydrophobic cores,
more ionic interactions, compact protein structure)
•Lipids composition (more stable towards hydrolysis by
heat)
•DNA stability is mediated by enzymes and small DNAbinding proteins.
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Phylum
Order
Galdiera sulphuraria Thermophilic
Rhodophyta
Cyanidiales
The unicellular red micro-alga Galdieria sulphuraria (Cyanidiales) is a
eukaryote that can represent up to 90% of the biomass in extreme
habitats such as hot sulfur springs with pH values of 0 to 4 and
temperatures of up to 56°C.
Molecular adaptations to high
acidity
•Active transport mechanisms that help acidophilic
organisms to regulate their internal pH
•The plasma membrane of these organisms contains a
P-type ATPase
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Cyanidium caldarium
Phylum
Order
Rhodophyta
Cyanidiales
Thermophilic
Acidophilic
The best studied high-temperature eukaryote is
the acidophilic phototroph Cyanidium
caldarium. Brock (1978) carefully examined the
growth and ecology of this organism and
determined its optimal growth temperature was
45ºC and the maximum temperature at which
growth occurred was 57ºC. pH 0
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Phylum: Euglenozoa
Family: Eutreptiaceae
Euglena mutabilis
Acidophilic
The competitive advantage of Euglena mutabilis in AMD, in comparison with
other Euglena species, is well documented (Olaveson and Nalewajko, 2000).
Uptake of metals by algal communities and mineral-algae interactions, in
general, induce more or less discrete modifications in aquatic environments.
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Trichosporon cerebriae, Acremonium spp.,
Acidomyces acidophilus Selbmann et al.
Acidophiles
Two fungi, Acremonium sp., and Trichosporon cerebriae, grow near pH 0.
These polyextremophiles (tolerant to multiple environmental extremes) thrive
in a brew of sulfuric acid and high levels of copper, arsenic, cadmium, and
zinc with only a cell membrane and no cell wall.
Phylum
Family
Basidiomycota
Sporidiobolaceae
Phylum
Family
Ascomycota
Hypocreales
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Genus Klebsormidium
Phylum Streptophyta (Chlorophyta)
Family Klebsormidiaceae
Acidophilic
Polluted water Particularly K. subtile, K.
rivulare and K. fl accidium are often referred
species in AMD (Acid Mine Drainage).These
algae may control acidity and metals in
solution.
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Dunaliella acidophila
phylum Chlorophyta
family Chlamydomonadaceae
Dunaliella acidophila was isolated from acid soils of the Solfatara at Pisciarelli,
Campi Flegrei, Naples, Italy, where pH fluctuate from 0.6 to 1.6. this alga was
found in four other sites in Italy, three of which are sulfure springs (pH 0.8 to
1.5) and one sulfurous exhalation (pH 1.5). This microbial alga thrives only in
acidic environments.
Molecular adaptations to high
pH values (basicity)
Plasma membranes may also maintain pH
homeostasis by using the Na+/H+ antiporter
system
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Neosartorya stramenia
Emericellopsis minima
Melanospora zamiae
alkalophilic
Emericellopsis minima, Neosartorya stramenia and Melanospora zamiae were
isolated at pH 10 to 11.
These species are alkalophilic according to the pH isolation medium.
Taking into account the great myceliar development at pH 11, we can state that this
species preferred alkaline conditions.
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Acremonium
and Chrysosporium spp
alkalophilic
The fungal species isolated from alkaline soils at two limestone caves in
Japan were considerably different from the species reported in alkaline
environments in other countries.
Acremonium sp did not grow at all at pH 5.
Two isolates of Chrysosporium also grew well in alkaline conditions, and
Chrysosporium sp. was designated as an alkalophile
Low temperatures adaptations
• Psychrophilic microorganisms have adapted to cold
environments by producing largely unsaturated fatty
acids for the lipids in their plasma membranes.
• psychrophilic eukaryotes membranes contain
polyunsaturated fatty acids which generally not occur in
provkaryotes
• Enzymes function at a reduced rate at near-freezing
ambient temperatures.
• Proteins are not as rigid as their mesophilic counterparts,
due to high α-helix and random coil contents, and a very
low content of β-sheets.
• Isofunctional enzymes working at low temperatures
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Chlamydomonas raudensis
phylum Chlorophyta
family Chlamydomonadaceae
psychrophilic
A psychrophilic green alga was isolated from
the photic zone at a depth of 17m in the
permanently ice-covered lake, Lake Bonney,
Antarctica. This green alga is obligate
psychrophile because it doesn’t grow at
temperatures above 16° C and the optimum
is 5° C (Morgan et al. 1998)
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Euglenozoa
psychrophilic
Tetreutreptia pomquetensis
Phylum: Euglenozoa
Family: Eutreptiaceae
Growth in culture, was limited at normal seawater
salinities to temperatures between 0° and 7° C, and the
alga could not be maintained at 10° C; thus, it is a strict
psychrophile.
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Fragilariopsis cylindrus
Phylum
Family
Bacillariophyta
Bacillariaceae
psychrophylic
One of the most abundant diatoms, especially in
the southern polar oceans, is Fragilariopsis
cylindrus (Grunow) Krieger (Bacillariophyceae).
The optimum growth temperature of F. cylindrus
is +5°C (Fiala & Oriol, 1990)
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All amoebae were isolated and maintained at temperatures below 4°C.
Growth, rate of locomotion, and general morphology were observed at
an environmentally appropriate temperature (1°C).
Neoparamoeba aestuarina antartica
Phylum Rhizopoda
Family
Dactylopodida
Platyamoeba oblongata
Phylum Rhizopoda
Family
Vannellidae
psychrophilic
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Three isolates were assigned to a psychrophilic subspecies of Neoparamoeba aestuarina, N.
aestuarina antarctica n. subsp., one isolate was assigned to a new species of Platyamoeba, P.
oblongata n. sp., two isolates were also assigned to a new species of Platyamoeba, P. contorta n.
sp., and one isolate was a novel psychrophilic gymnamoeba Vermistella antarctica n. gen. n. sp.
Vermistella antarctica
Phylum Rhizopoda
Family
Incertae sedis
Platyamoeba contorta
Phylum Rhizopoda
Family
Vannellidae
psychrophylic
Molecular properties of
halophilic organisms
• The ability to sense changes in Na+ concentrations in the
environment is pivotal in cell viability.
• Halophilic Cyanobacteria and Bacteria use compatible
solutes such as sucrose, threalose or glycine betaine
• Saccharomyces cerevisiae, as well as xerophilic fungi and
algae (Dunaliella) use glycerol. the main pathway involved in
sensing these changes and in responding to them is known
as the high osmolarity glycerol (HOG) signalling pathway.
• Important new features have been discovered in the fungus
Hortaea werneckii that relate to the HOG signal
transduction pathway; these are involved in the sensing and
responding to increases in NaCl
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Phylum
Alveolata
Fabrea salina
Phylum
Family
Alveolata
Climacostomidae
halophilic
Species richness of phytoplankton, ciliates and zooplankton
greatly decrease above a salinity of 150 and typical halophiles
were found between 150 and 350 salinity. In this environment,
F. salina appeared more adapted than the brine shrimp to
survive during phytoplankton blooms.
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Hortaea werneckii
halophylic
Hortaea werneckii is a black yeastlike hyphomycete associated with the human
superficial infection tinea nigra. This fungus has been reported from several
environments around the world and it has shown a preference to hypersaline habitats.
These strains had the highest frequency of isolation and grew at the maximum NaCl
concentration (25%).
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Eurotium repens
Eurotium rubrum
Aspergillus ustus
xerophylic
Eurotium rubrum, eurotium repens, aspergillus ustus, that are fungi
commonly known as ―moulds‖, can grow in low water availability. This
ability could be harmful because xerophilic fungi can grow on
commercial food stored and kept dry.
Adaptation to high radiations
• Very efficient DNA repair system (enzyme
complex that excise misincorporated
bases in DNA double-strand and replace
them with the correct ones)
• Production of antioxidant pigments ( carotene, melanin, etc.) protect cells
exposed to high UV radiations.
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Dunaliella bardawil
phylum Chlorophyta
family Chlamydomonadaceae
Radiation-resistant
High -carotene containing D. bardawil was shown to be strongly protected
against damage induced by excessive irradiation. This green alga can grow in
presence of high UV-A and UV-B radiations because it is capable to protect
itself producing antioxidant pigments.
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Thalassiosira weissflogii Radiation-resistant
Phylum
Family
Bacillariophyta
Thallassiosiraceae
Unialgal cultures of the marine diatom Thalassiosira
weissflogii (Grunow) were exposed for 40 days to
artificial UV-B and UV-A radiation to examine long-term
response was supplied for 4 h/ day. T. weissflogii is a
relatively UV-tolerant species and that its long-term
response to UV exposure involves an activation of the
xanthophylls cycle.
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Phylum
Ascomycota
Alternaria alternata
Phylum
Family
Ascomycota
Pleosporaceae
Radiation-resistant
The filamentous fungus Alternaria alternata is
known to grow in highly radioactive
environments and may therefore represent a
unique model for investigating the genetic
basis of the phenomenon of radioresistance.
A. alternata is also well known for producing melanin, a black pigment which is produced and accumulates inside
the mycelium (Kimura and Tsuge, 1993). There are some indications that the radioresistance of microorganisms
may result from melanization of their cells (Pointing et al., 1996).
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Aspergillus versicolor and Cladosporium cladosporoides,
Radiation-resistant and radiation eating organism!!!
Sustained exposure of microfungi to radiation appears to have resulted in formerly
unknown adaptive features, such as directed growth of fungi to sources of ionizing
radiation. these isolates showed significant growth stimulation in the presence of ionizing
radiation, a property we call radiostimulation. Mitosporic fungi, isolated from
uncontaminated locations did not exhibit these properties.
Molecular properties of
barophilic microorganisms
• Proteins do not appear to be so greatly affected
by high pressures
• However, when cells were grown at high
pressures, membranes contain a high quantity
of unsaturated fatty acids to avoid membrane
gelling and ensure retainment of fluidity
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Aspergillus sydowii
Magnaporthe grisea
Barophylic
Recently, a filamentous fungus Aspergillus sydowii was isolated from 5000 m depth in the
Central Indian Basin (Raghukumar et al., 2004). The spores of A. sydowii germinated at
500 bar pressure.
The fungus Magnaporthe grisea is capable of producing an internal pressure of 8 MPa
during the process of mechanical penetration of its host plant (de Jong et al., 1997) by the
synthesis of glycerol (cf Dunaliella above).
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Recently…
microcolonial cryptoendolithic fungi,
Cryomyces minteri and Cryomyces
antarcticus, have been discovered in
McMurdo Dry Valleys where they live
and thrive under multiple stresses, in
particular, a continuous exposure to
UV-A and UV-B radiations.
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Conclusions
• There are few studies about adaptations in
extreme conditions of microbial eukaryotes
• Extreme eukaryotes can withstand many
environmental stresses
•Deepening knowledge about extreme microbial
eukaryotes could be useful to understand stress
responses and possible defence mechanisms in
eukaryotic cells including human cells
Suggestions
•Some adaptations to extreme conditions are
common in different groups of eukaryotic and
prokaryotic organisms (e.g. glycerol in cold
adaptaion). This could be a common way to
approach adptation to extremes
•Symbiosis
•Fungi are very interesting as the only
organisms actually adapted to terrestrial
environment
Thanks to the people in my lab
…and thank you for the attention!