ExtremophilesExtremophiles These are organisms that These are organisms that thrive in extreme conditions thrive in extreme conditions

they include:they include: ThermophilesThermophiles

HalophilesHalophiles BarophilesBarophiles AcidophilesAcidophiles XerophilesXerophiles

Many thermophiles inhabit Many thermophiles inhabit areas of multiple extremes and areas of multiple extremes and

some are methanogenic some are methanogenic (produce methane)(produce methane)

Thermophiles and Thermophiles and HyperthermophilesHyperthermophiles

Thermophiles are organisms that have Thermophiles are organisms that have optimum growth temperatures of 45optimum growth temperatures of 45ooC - 80C - 80ooC C

(113(113o o F - 176F - 176o o F) F) Hyperthermophiles are organisms that have an Hyperthermophiles are organisms that have an

optimum growth temperature above 80optimum growth temperature above 80ooC (176C (176o o

F)F)

Thermophilic Thermophilic EnvironmentsEnvironments

Environments that foster Environments that foster the growth of the growth of

thermophiles and thermophiles and hyperthermophiles hyperthermophiles

include hot springs, deep-include hot springs, deep-sea hydrothermal vents, sea hydrothermal vents,

areas associated with areas associated with volcanic phenomena, hot volcanic phenomena, hot water heaters, and even water heaters, and even

surfaces of compost heapssurfaces of compost heaps

Energy GenerationEnergy Generation

These organisms tend to These organisms tend to be either be either

chemoorganotrophs or chemoorganotrophs or chemolithotrophschemolithotrophs

Example of Example of chemolithotrophic energy chemolithotrophic energy generation include iron generation include iron oxidation and elemental oxidation and elemental

sulfur (Ssulfur (S00) reduction) reduction

DNA StabilityDNA Stability Many of these organisms use large concentrations of Many of these organisms use large concentrations of

compatible solutes to increase the temperature at compatible solutes to increase the temperature at which their DNA denatureswhich their DNA denatures

Whereas the DNA of most organisms is negatively Whereas the DNA of most organisms is negatively supercoiled, the DNA of these organisms is positively supercoiled, the DNA of these organisms is positively

supercoiledsupercoiled

Positive SupercoilingPositive Supercoiling

Thermophiles and Thermophiles and Hyperthermophiles use Hyperthermophiles use a unique enzyme called a unique enzyme called

reverse gyrase to reverse gyrase to introduce positive introduce positive

supercoils into their supercoils into their DNADNA

This stabilizes the DNA This stabilizes the DNA and increases its and increases its

melting temperaturemelting temperature

Protein StabilityProtein Stability Thermophiles and Hyperthermophiles use Thermophiles and Hyperthermophiles use

unique heat active chaperonins to refold unique heat active chaperonins to refold partially denatured proteinspartially denatured proteins

Many proteins of these organisms contain Many proteins of these organisms contain a very hydrophobic core and a greater a very hydrophobic core and a greater

extent of disulfide bonds and ionic extent of disulfide bonds and ionic interactions which increases the interactions which increases the

temperature at which the proteins temperature at which the proteins denaturedenature

TaqTaq polymerase is an important enzyme polymerase is an important enzyme from these microbes isolated from from these microbes isolated from

Thermus aquaticusThermus aquaticus

Unusual MembranesUnusual Membranes Archaeal hyperthermophiles have cellular Archaeal hyperthermophiles have cellular

membranes lacking fatty acidsmembranes lacking fatty acids They have a lipid monolayer composed of They have a lipid monolayer composed of

CC4040 (biphytanyl) linked to glycerol by an (biphytanyl) linked to glycerol by an ether, not an ester, linkageether, not an ester, linkage

Extremophile VirusesExtremophile Viruses

Most viruses have classic Most viruses have classic head and tail morphologyhead and tail morphology

Most are grouped under Most are grouped under the Myoviridae and the the Myoviridae and the Siphoviridae families of Siphoviridae families of

viruses (some exceptions)viruses (some exceptions) 12 different types have 12 different types have

been isolated so farbeen isolated so far

Viruses of Halobacteria & Viruses of Halobacteria & MethanobacteriaMethanobacteria

Viruses of Thermophiles: Viruses of Thermophiles: Unusual VirusesUnusual Viruses

The viruses of The viruses of thermophiles and thermophiles and

hyperthermophiles are as hyperthermophiles are as unique as their hostsunique as their hosts

Some of these viruses Some of these viruses possess bottle-shaped or possess bottle-shaped or

lemon-shaped virionslemon-shaped virions Others have the ability to Others have the ability to

undergo certain undergo certain morphological morphological

developments outside of a developments outside of a host cell host cell

A)Lipothrix Virus SIFV B)Lipothrix Virus AFV 1 C)Rudivirus SIRV1 D)Fusellovirus SSV1

E)Guttavirus SNDV

AcidianusAcidianus bottle-shaped bottle-shaped virus (ABV)virus (ABV)

Double stranded DNA virus with a Double stranded DNA virus with a linear genome of 24 kblinear genome of 24 kb

Infects archaea of the genus Infects archaea of the genus AcidianusAcidianus Has a bottle-shaped virion with a Has a bottle-shaped virion with a

funnel-shaped corefunnel-shaped core The narrow end is important for DNA The narrow end is important for DNA

transfer to the host celltransfer to the host cell The wide end possesses a ring of The wide end possesses a ring of filaments the function of which is not filaments the function of which is not

clearly understoodclearly understood

SulfolobusSulfolobus: A : A ThermoacidophileThermoacidophile

Several different viruses infect the Several different viruses infect the thermoacidophile sulfolobus among thermoacidophile sulfolobus among

these are:these are: Fuselloviridae: SSV1 (Japan) SSV2 and Fuselloviridae: SSV1 (Japan) SSV2 and

SSV3 (Iceland)SSV3 (Iceland) Rudiviridae: SIRV1 and SIRV2 (Iceland)Rudiviridae: SIRV1 and SIRV2 (Iceland)

Lipothrixviridae: SIFVLipothrixviridae: SIFV• Alphalipothrixvirus: Thermoproteus virus 1Alphalipothrixvirus: Thermoproteus virus 1• Betalipothrixvirus: Betalipothrixvirus: Sulfolobus mislandicusSulfolobus mislandicus

filamentous virusfilamentous virus• Gammalipothrixvirus: Acidianous Gammalipothrixvirus: Acidianous

filamentous virus 1filamentous virus 1 Guttaviridae: SNDV (Sulfolobus Guttaviridae: SNDV (Sulfolobus

neozealandicus)neozealandicus)

Information on Information on SulfolobusSulfolobus VirusesViruses

FuselloviridaeFuselloviridae-60 x 90 nm spindle -60 x 90 nm spindle

shaped virionsshaped virions

-Circular dsDNA 15.5 -Circular dsDNA 15.5 kbkb

-35 ORF’s-35 ORF’s

-Lysogenic propagation-Lysogenic propagation RudiviridaeRudiviridae-Stiff 23 x 800-900 helical -Stiff 23 x 800-900 helical

rodsrods

-Linear dsDNA with -Linear dsDNA with covalently closed endscovalently closed ends

-45 ORF’s-45 ORF’s

More Sulfolobus VirusesMore Sulfolobus Viruses

LipothrixviridaeLipothrixviridae--24 x 250 nm flexible rods 24 x 250 nm flexible rods

with attachment fiberswith attachment fibers

-Linear dsDNA 43 kb-Linear dsDNA 43 kb

-74 ORF’s-74 ORF’s

GuttaviridGuttaviridaeae

--Droplet Droplet ShapedShaped

-Circular and -Circular and highly highly

modified modified genomegenome

-dsDNA 20 -dsDNA 20 kbkb

Relationship of these Virus Relationship of these Virus IsolatesIsolates

Comparisons of Comparisons of previous and previous and

recent isolates recent isolates in order to in order to determine determine

phylogenetic phylogenetic and evolutionary and evolutionary

relationshipsrelationships

Comparison of Isolate Comparison of Isolate GenomesGenomes

AcidianusAcidianus two-tailed two-tailed virus (ATV)virus (ATV)

This virus also has an This virus also has an interesting virion that is interesting virion that is

lemon-shapedlemon-shaped It infects the archaeon It infects the archaeon

Acidianus convivatorAcidianus convivator This virus is found in hot-This virus is found in hot-

springs which reach springs which reach temperatures between 85-temperatures between 85-9393o o C with a pH of about C with a pH of about

1.51.5

Development Outside of Development Outside of HostsHosts

It has been shown that this virus is able It has been shown that this virus is able to develop one long tail on each end of to develop one long tail on each end of its virion in the absence of host cellsits virion in the absence of host cells

If this virus is incubated at natural If this virus is incubated at natural temperatures in the absence of hosts, temperatures in the absence of hosts, about half the population develops the about half the population develops the

tails within a weektails within a week Since this is one of the few viruses of Since this is one of the few viruses of hyperthermophiles which causes lysis, it hyperthermophiles which causes lysis, it is thought that the development of the is thought that the development of the

tail aids in stabilizing the virion and tail aids in stabilizing the virion and infecting a new host in harsh conditions infecting a new host in harsh conditions

with low host concentrationswith low host concentrations

Extracellular Extracellular DevelopmentDevelopment

From Häring M, Vestergaard G, et. al. 2005. Virology: independent virus development outside a host. Nature 436 (7054): 1101-2.

RM378RM378 Infects Infects Rhodothermus marinusRhodothermus marinus, a type of , a type of

thermoacidophilethermoacidophile Myoviridae, A2 morphology (includes the T4 Myoviridae, A2 morphology (includes the T4

bacteriophage of bacteriophage of E. coliE. coli)) Shows some sequence homology to T4 bacteriophageShows some sequence homology to T4 bacteriophage

Elongated head 95 x 85 nm and a 150 nm long tail with a Elongated head 95 x 85 nm and a 150 nm long tail with a connector connector

Infectious up to 65 C, optimum temperature is 64 CInfectious up to 65 C, optimum temperature is 64 C 130 kb dsDNA with 200+ ORF’s and a 42% G+C content130 kb dsDNA with 200+ ORF’s and a 42% G+C content

Thermostable RNA ligase 1 has been isolatedThermostable RNA ligase 1 has been isolated Advantages: RLM-RACE, other RNA-DNA ligation Advantages: RLM-RACE, other RNA-DNA ligation

applications of biotechnologyapplications of biotechnology

Virus-Like Particles Virus-Like Particles (VLP’s)(VLP’s)

His1, lytic viral isolate His1, lytic viral isolate of of Haloarcula hispanicaHaloarcula hispanica

Linear dsDNALinear dsDNA 14.9 kb14.9 kb

Pav1, a deep-sea isolate of Pav1, a deep-sea isolate of Pyrococcus Pyrococcus AbysiiAbysii

Circular dsDNA 17.5 kbCircular dsDNA 17.5 kb

Was first isolated from Was first isolated from Methanococcus voltaeMethanococcus voltae strain strain

A3A3 23 kb covalently closed 23 kb covalently closed

circular DNAcircular DNA

ReferencesReferences Bath C and Dyall-Smith ML. 1998. His1, an archaeal virus of the Bath C and Dyall-Smith ML. 1998. His1, an archaeal virus of the

FuselloviridaeFuselloviridae family that infect family that infect Haloacula hispanicaHaloacula hispanica. Journal of . Journal of Virology 72(11): 9392-5.Virology 72(11): 9392-5.

Beeby M, O’Connor BD, et al. 2005. The genomics of disulfide Beeby M, O’Connor BD, et al. 2005. The genomics of disulfide bonding and protein stabilization in thermophiles. DOI: bonding and protein stabilization in thermophiles. DOI: 10.1371/journal.pbio..0030309.10.1371/journal.pbio..0030309.

Bettstetter M, Peng X, et. al. 2003. Bettstetter M, Peng X, et. al. 2003. AFV1, a novel virus infecting hyperthermophilic archaea of the geAFV1, a novel virus infecting hyperthermophilic archaea of the genus acidianus.nus acidianus. Virology 215 (1): 68-80. Virology 215 (1): 68-80.

Brock, T.D., M. T. Madigan, J.M. Martinko, and J. Parker. 2006. Brock, T.D., M. T. Madigan, J.M. Martinko, and J. Parker. 2006. Biology of Microorganisms, 11th editionBiology of Microorganisms, 11th edition. Prentice Hall, Upper . Prentice Hall, Upper Saddle River, NJ.Saddle River, NJ.

Geslin C, Romancer M, et. al. 2004. PAV1 the first virus-like Geslin C, Romancer M, et. al. 2004. PAV1 the first virus-like particle isolated from a hyperthermophilic Euryarchaote, particle isolated from a hyperthermophilic Euryarchaote, Pyrococcus abysiiPyrococcus abysii. Journal of Virology (78)4: 1954-61.. Journal of Virology (78)4: 1954-61.

Häring M, Reinhard R, et. al. 2005. Viral diversity in hot springs Häring M, Reinhard R, et. al. 2005. Viral diversity in hot springs of Pozzuoli, Italy, and characterization of a unique archaeal virus, of Pozzuoli, Italy, and characterization of a unique archaeal virus, Acidianus Bottle-Shaped Virus, from a new family, the Acidianus Bottle-Shaped Virus, from a new family, the Ampullaviridae. Journal of Virology 79 (15): 9904-12.Ampullaviridae. Journal of Virology 79 (15): 9904-12.

References ContinuedReferences Continued Häring M, Vestergaard G, et. al. 2005. Virology: independent Häring M, Vestergaard G, et. al. 2005. Virology: independent

virus development outside a host. Nature 436 (7054): 1101-2.virus development outside a host. Nature 436 (7054): 1101-2. Rice G, et al. 2001. The structure of a thermophilic archaeal Rice G, et al. 2001. The structure of a thermophilic archaeal

virus shows a double-stranded DNA viral capsid type that virus shows a double-stranded DNA viral capsid type that spans all domains of life. Proceedings of the National Academy spans all domains of life. Proceedings of the National Academy of Sciences of the United States of America 98(23): 13341-5.of Sciences of the United States of America 98(23): 13341-5.

Robb FT. 2004. Genomics of thermophiles. In: Fraser CM, Robb FT. 2004. Genomics of thermophiles. In: Fraser CM, Read TD, Nelson KE. Microbial Genomes. Totawa, NJ: Humana Read TD, Nelson KE. Microbial Genomes. Totawa, NJ: Humana Press. p. 245-267.Press. p. 245-267.

Staley, J.T., and A. Reysenbach. 2002. Staley, J.T., and A. Reysenbach. 2002. Biodiversity of Microbial Biodiversity of Microbial LifeLife. John Wiley & Sons, Inc., New York.. John Wiley & Sons, Inc., New York.

Thorarinn B, Hjorleifsdottir SH, et. al. 2003. Discovery and Thorarinn B, Hjorleifsdottir SH, et. al. 2003. Discovery and characterization of a thermostable bacteriophage RNA ligase characterization of a thermostable bacteriophage RNA ligase to T4 RNA ligase 1. Nucleic Acids Research 31(24): 7247-54.to T4 RNA ligase 1. Nucleic Acids Research 31(24): 7247-54.

Wiedenheft B, et. al. 2004. Comparative genomic analysis of Wiedenheft B, et. al. 2004. Comparative genomic analysis of hyperthermophilic archaeal hyperthermophilic archaeal FuselloviridaeFuselloviridae viruses.” Journal of viruses.” Journal of Virology 78(4):1954-61.Virology 78(4):1954-61.

Questions???Questions???

?