Halophiles

Shiladitya DasSarma, University of Maryland, Baltimore, Maryland, USA Priya DasSarma, University of Maryland, Baltimore, Maryland, USA

Advanced article
Article Contents
. Introduction . Hypersaline Environments . Prokaryotic Halophiles . Halophilic Archaea . Eukaryotic Halophiles . Biotechnology . Conclusions and Future Prospects

Online posting date: 15th March 2012

Halophiles are salt-loving organisms that flourish in saline environments and can be classified as slightly, moderately or extremely halophilic, depending on their requirement for sodium chloride. Although most marine organisms are slight halophiles, moderate and extreme halophiles are generally more specialised microbes, which inhabit hypersaline environments with salinity higher than in the sea. Hypersaline environments are found all over the world, in arid, coastal, and deep-sea locations, underground salt mines, and artificial salterns. Halophilic microorganisms include a variety of heterotrophic, phototrophic, and methanogenic archaea, photosynthetic, lithotrophic, and heterotrophic bacteria, and photosynthetic and heterotrophic eukaryotes. Examples of well-adapted and widely distributed extremely halophilic microorganisms include archaea for example, Halobacterium sp. NRC-1, cyanobacteria such as Aphanothece halophytica, and the green alga Dunaliella salina. Multicellular halophilic eukaryotic organisms include brine shrimp and the larvae of brine flies. Halophilic organisms either accumulate internal organic compatible solutes to balance the osmotic stress of the environment or produce acidic proteins to increase solvation and improve function in high salinity.

Introduction
Although salts are required for all life forms, halophiles (from the Greek, hal, meaning sea or salt, and philos, meaning loving) are distinguished by their requirement of high salinity conditions for growth. They may be classied according to the degree of their salt requirement: slight
eLS subject area: Microbiology How to cite: DasSarma, Shiladitya; and DasSarma, Priya (March 2012) Halophiles. In: eLS. John Wiley & Sons, Ltd: Chichester. DOI: 10.1002/9780470015902.a0000394.pub3

halophiles grow optimally at 0.20.85 M (15%) sodium chloride (NaCl); moderate halophiles grow optimally at 0.853.4 M (520%) NaCl; and extreme halophiles grow optimally at 3.45.1 M (2030%) NaCl. In contrast, nonhalophiles grow optimally in less than 0.2 M NaCl concentrations. Halotolerant organisms can grow in either high salinity or in the absence of a high concentration of salt. Many halophiles and halotolerant microorganisms can grow over a wide range of salt concentrations, with requirement or tolerance for salts sometimes dependent on environmental and nutritional factors. High osmolarity in hypersaline conditions is deleterious to most cells since water is lost to the external medium. To prevent loss of cellular water, halophiles generally accumulate high solute concentrations within the cytoplasm (Roberts, 2005; Yancey, 2005). When an isoosmotic balance with the medium is achieved, cell volume is maintained. The compatible solutes or osmolytes which accumulate in halophiles are generally amino acids, sugars and polyols, which do not interfere with intracellular processes and have no net charge at physiological pH. Halotolerant yeasts and green algae accumulate polyols, whereas most halophilic and halotolerant bacteria accumulate zwitterionic species (containing both positive and negative charges), such as glycine betaine and ectoine. Compatible solute accumulation may occur by biosynthesis, de novo or from storage material, or by direct uptake from the medium. A major exception is found among the haloarchaea and some extremely halophilic bacteria, which accumulate potassium chloride (KCl) equal to the external concentration of NaCl. These organisms produce acidic proteins that can function in high salinity by remaining solvated and reducing aggregation, precipitation and denaturation (Madern et al., 2000).

Hypersaline Environments
Though the oceans are, by far, the largest body of saline water, constituting approximately 99% of the biosphere, hypersaline environments are generally dened as those containing salt concentrations in excess of seawater (3.5% total dissolved salts). Most hypersaline bodies are thalassic
1

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

(from the Greek, thalassa, sea), as they derive from the evaporation of seawater and retain the relative proportion of salts in the sea. A great diversity of microbial life is observed in thalassic brine upward from marine salinity (  0.6 M) to approximately 33.5 M NaCl, at which point only a few extreme halophiles can survive, for example Halobacterium, Dunaliella, and a few bacterial species. Athalassic waters are those in which the salts are in nonmarine proportions, and are less common. Two large and well-studied hypersaline lakes are the Great Salt Lake (Figure 1), in the western United States, and the Dead Sea, in the Middle East. The Great Salt Lake is larger (  4000 km2) and shallower (on average 10 m deep), and is thalassic. The Dead Sea is smaller (400 km2) and deeper (300 m), and is athalassic, with a relatively high concentration of magnesium salts. Both of these lakes are close to neutral pH, although the former is slightly alkaline whereas the latter is slightly acidic. Compared to smaller hypersaline ponds, the composition of larger lakes remain fairly constant as a result of their size; recently, however, human activities have had signicant eects on the chemistry and biology of many. For example, a railroad causeway built in 1959 divided the Great Salt Lake into northern and southern sections, leading to dilution of the southern section, which receives the greatest inow of freshwater from streams, and the concentration of the northern section to nearly saturating salinity (Gwynn, 2002). Diversion of incoming freshwater streams for irrigation in the Dead Sea basin in recent years has also had a signicant impact on its size and salinity, and has resulted in concern about the lakes long-term viability (Abu Ghazleh et al., 2011). Many small evaporation ponds or sabkhas are found near coastal areas, where seawater penetrates through seepage or via narrow inlets from the sea. Notable among these are Solar Lake and other ponds near the Red Sea coast, Guerrero Negro on the Baja California coast, Lake Sivash near the Black Sea and Shark Bay in Western Australia. Hypersaline evaporation ponds have also been found in Antarctica (e.g. Deep Lake, Organic Lake and Lake Suribati), and also in the Atacama Desert region of

Figure 1 Bloom of halophilic microorganisms. Dense growth of halophilic microorganisms in hypersaline environments leads to reddening of the brine. Photo Dr. S. DasSarma.

Chile, which are located in some of the driest areas of the world (Javor, 1989). A number of alkaline hypersaline soda brines exist, including the Wadi Natrun lakes of Egypt, Lake Magadi in Kenya, and the Great Basin lakes of the western United States (Mono Lake, Owens Lake, Searles Lake and Big Soda Lake), several of which are intermittently dry (Javor, 1989). Soda brines lack magnesium and calcium divalent cations because of their low solubility at alkaline pH. Many smaller hypersaline pools represent especially dynamic environments, experiencing signicant seasonal variations in size, salinity and temperature. In addition to natural hypersaline lakes, numerous articial solar salterns have been constructed for the production of sea salts. These usually consist of a series of shallow evaporation ponds connected by pipes and canals, with brine being directed into smaller ponds as salinity increases through evaporation. During the process of evaporation, sequential precipitation of salts, calcium carbonate, calcium sulfate (gypsum) and NaCl (halite) occurs. After precipitation of NaCl, the potassium and magnesium chloride and sulfate may be harvested, or the remaining brines (called bitterns) returned to the sea. Hypersaline environments also occur in subterranean evaporite deposits and deep-sea basins created by salt domes. Deep-sea brines are relatively stable as a result of their higher density than surrounding seawater and they have been found in the Red Sea, Gulf of Mexico and Mediterranean Sea. Recent studies have shown evidence for microbial activity in some deep-sea hypersaline basins, including one containing 5 M magnesium chloride (MgCl2), and in subsurface halite deposits over 10 million years old (Antunes et al., 2011; van der Wielen et al., 2005; Fish et al., 2002). Sulfur cycling and methanogenesis were shown to drive microbial colonisation in a Mediterranean deep hypersaline basin (Borin et al., 2009). Recent evidence for subterranean brines has also been discovered on Mars, where seasonal ows from the sides of craters were detected by an orbiting satellite (McEwen et al., 2011). Evaporation of hypersaline brines is frequently observed, leading to a gradient of salinity, which in turn leads to sequential blooms of diverse microbial species adapted to dierent ranges of salinity (Figure 2). In solar salterns, as brine is concentrated from 1 M NaCl to approximately 3.5 M, dense algal populations are supported, on which brine shrimp and larvae of brine ies feed. Protozoa are also found, as are yeasts and fungi. Microbial mats, containing predominantly photosynthetic unicellular and lamentous cyanobacteria, purple and green sulfur and nonsulfur bacteria cover the bottom of many hypersaline ponds (Caumette, 1993; Figure 3). In the anoxic zones of the mats and in the sediment below, a variety of sulfur oxidising, sulfate reducing, homoacetogenic, methanogenic, and heterotrophic bacteria and archaea occur. From approximately 4 M NaCl to saturation (45.1 M NaCl), halophilic archaea dominate the brine pools and most other microbial activity ceases.

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

100

PR-6

Fs

Ds

Ah

Growth rate (%)

Sea water

Hatch range

0 Salinity (%) 0 1) NaCl (mol L 0

10 1 2 3

20 4

30 Saturation 5

Figure 2 Salt-tolerance of halophilic organisms. Relative growth rate is plotted against both per cent salinity and NaCl concentration. The five microorganisms depicted are Synechococcus sp. PCC7002 (PR-6), a slightly halotolerant cyanobacterium, Fabrea salina (Fs), a moderately halophilic protozoan, Dunaliella salina (Ds), a halophilic green algae, Aphanothece halophytica (Ah), an extremely halophilic cyanobacterium and Halobacterium sp. (H), an extremely halophilic archaeon. The salinity of seawater and the hatch range for brine shrimp are noted.

0
Unicellular cyanobacteria layer

3
Gypsum crystal layer
Depth (mm)

6 9 12

Filamentous cyanobacteria layer Purple bacteria layer Anaerobic bacteria and archaea layer in black sediment (FeS)

15
Figure 3 Structure of a hypersaline microbial mat. Adapted from Caumette (1993), with permission from Springer.

Prokaryotic Halophiles
A great diversity of prokaryotic halophiles exists in nature and they have been studied by both culturing and nonculturing techniques, with sequence-based approaches becoming increasingly popular for phylogenetic and taxonomic classication. An approach used for classifying cultured halophiles is by analysis of the nucleotide sequence of multiple genes, a method called multilocus sequence typing (MLST) (Papke et al., 2011; de la Haba et al., 2011). For noncultured halophiles, environmental metagenomic studies using sequencing and phylogenetic analysis of 16S ribosomal ribonucleic acid (rRNA) genes are beginning to expand the breadth of diversity known among halophiles.

Halophilic Archaea
These extreme halophiles grow best at the highest salinities (3.45.1 M NaCl) (Figure 2), forming dense blooms (up to

108 cells mL2l), and resulting in the red colour of many brines (Figure 1). Common species of halophilic archaea, also called haloarchaea, are rods, cocci or disk-shaped, although triangular and even square-shaped species exist. Many are pleiomorphic, especially when the ionic conditions of the media are altered, and most lyse below 11.5 M NaCl. Haloarchaea are members of the archaeal domain (they are also called halophilic archaea, and formerly halobacteria). They have recently been proposed to be composed of two families (Minegishi et al., 2011), and include about three dozen genera. A recent metagenomic study of a solar saltern showed the occurrence of a major new phylotype, called nanohaloarchaea, with small cells (50.8 um) (Narasingarao et al., 2011). Other nonculture based studies have suggested that novel species similar to haloarchaea may occur in the human gastrointestinal tract (Oxley et al., 2010). The genomes of haloarchaea have been of signicant interest due to their evolutionary position and physiological capability to grow at the most extreme hypersaline conditions. The rst haloarchaeal genome completed was
3

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

for Halobacterium sp. NRC-1, which has a 2.57 Mb GCrich genome (DasSarma, 2004). The genomes of over a dozen haloarchaeal species have now been completely sequenced and analysed using bioinformatic tools, showing both the conserved information transfer genes and diverse metabolic genes (Capes et al., 2011; Anderson et al., 2011). Early microbiological analysis was conducted on several closely related Halobacterium strains isolated in the midtwentieth century from salted sh and meat from Europe and North America (DasSarma and DasSarma, 2008; DasSarma et al., 2010b, a). These are amino acid-utilising facultative aerobes, which require a number of growth factors and slightly elevated temperatures (38458C) for optimal growth. Most have distinctive features such as gas vesicles, purple membrane and red-orange carotenoids. Many are facultative anaerobes and grow utilising respiration of dimethoxylsulfoxide and trimethylamine N-oxide (a saltwater sh osmolyte) (Yancey, 2005), fermentation of dierent sugars, breakdown of arginine and light energy, mediated by retinal pigments. Some haloarchaeal species utilise carbohydrates, for example Haloarcula marismortui, Haloarcula vallismortis and Haloferax volcanii from the Dead Sea, Haloferax mediterranei and Halorubrum saccharovorum from salterns, and Halorubrum lacusprofundi, a psychrotrophic species from Deep Lake, Antarctica. Glucose is oxidised by a modied EntnerDoudoro pathway and the resulting pyruvate is further oxidised by pyruvate oxidoreductase and the tricarboxylic acid cycle. H. marismortui and H. volcanii have been reported to use a novel pathway for central metabolism, where acetyl-CoA is oxidised to glyoxylate via the key intermediate methylaspartate (Khomyakova et al., 2011). Several haloarchaea are capable of growth on single carbon sources such as sugars, glycerol and acetate, and recent genomic studies are providing additional insights into metabolic pathways (Sato and Atomi, 2011). Some haloarchaea, such as H. mediterranei accumulate polyhydroxyalkanoates, when grown in carbon excess conditions, or respire anaerobically via nitrate reductase (Han et al., 2010; Bonete et al., 2008). Some halophilic species are polyextremophilic and are tolerant of additional extremes (Bowers et al., 2006). For example, the Antarctic species, H. lacusprofundi, is coldadapted and can grow down to 228C, and the Red Sea hot brine species, Halorhabdus tiamatea, is thermotolerant and can tolerate up to 55608C. The alkaliphiles, Natronomonas pharaonis from Wadi Natrun and Natronococcus occultus from Lake Magadi, have pH optima in the 9.510 range and do not grow below pH 8.5. Slight acidophiles, such as Halarchaeum acidiphilum is able to grow at 4.06.0. The intracellular salt concentration of haloarchaea has been found to be extremely high, with K+ ions accumulated internally to near 5 M (Roberts, 2005). The concentration of Na+ ions appears to be in the molar range, with the K+/ Na+ ratio in the cytoplasm over 100. The K+ gradient is maintained by a combination of an electrogenic Na+/H+ antiporter and two K+ uniporters (Figure 4). Amino acid
4

uptake is carried out by a Na+/amino acid symporter. See also: Sodium, Calcium and Potassium Channels Proteins of haloarchaea are either resistant to high salt concentrations or require salt for activity. Both genome sequencing and proteome analysis have shown that they contain an excess proportion of acidic to basic amino acids, a feature likely to be required for protein activity at high salinity (Joo and Kim, 2005; DasSarma, 2004) (Figure 5). This characteristic is shared with proteins from some halophilic bacteria, for example Salinibacter ruber (which also grows in crystalliser ponds). Surface negative charges are thought to be important for solvation of halophilic proteins, and to prevent the denaturation, aggregation and precipitation, which usually result when nonhalophilic proteins are exposed to high salt concentrations. The structures of haloarchaeal glucose dehydrogenase, malate dehydrogenase, dihydrofolate reductase and large ribosomal subunit have been determined by X-ray crystallography (Britton et al., 2006; Ban et al., 2000). See also: Protein Stability A well-studied feature of haloarchaea is the purple membrane, found in specialised regions of the cell membrane, which contains a two-dimensional crystalline lattice of a single chromoprotein, bacteriorhodopsin. Bacteriorhodopsin contains a protein moiety, bacterio-opsin, and a covalently bound chromophore, retinal, and acts as a light-dependent transmembrane proton pump (Lanyi, 2004) (Figure 4). The membrane potential generated can be used to drive adenosine triphosphate (ATP) synthesis and support a period of phototrophic growth. Bacteriorhodopsin is induced by low oxygen tension and high light intensity and can cover more than 50% of the surface of cells. The structures of bacteriorhodopsin and sensory rhodopsin have been solved through electron and X-ray diraction. Although originally thought to be unique to haloarchaea, similar photosensory receptors and lightdriven pumps have been discovered in a wide range of microorganisms, including both bacteria and fungi. See also: Archaeal Cells; Rhodopsin Haloarchaea produce large quantities of red-orange carotenoids. Carotenoids have been shown to be necessary for stimulating an active photorepair system for repair of thymine dimers resulting from ultraviolet radiation (Crowley et al., 2006). The most abundant carotenoids are C-50 bacterioruberins, although smaller amounts of biosynthetic intermediates such as b-carotene and lycopene are also present. Retinal is produced by oxidative cleavage of b-carotene, a step which requires molecular oxygen. Several retinal proteins, in addition to bacteriorhodopsin, are also produced by haloarchaea, including halorhodopsin, which is an inwardly directed light-driven chloride pump, and two sensory rhodopsins, which mediate the phototactic response (swimming toward green light and away from blue and ultraviolet light) (DasSarma et al., 2001). Haloarchaea often produce buoyant gas vesicles, which are hollow, rigid proteinaceous subcellular structures surrounding a gas-lled space. The function of gas vesicles for

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

h
OxlT Formate LctP RbsAC UgpABCE H+

h
SRI/HtrI SRII/HtrII (Ser)

ATP synthase

BR H+

HR

Respiratory chain
Oxalate Lactate PotABCD Spermidine putrescine Ribose Glycerol 3-phosphate

Ret

Ret

Ret

Ret

Ret

Cl
ATP H + ADP, P i

Htr X (HemAT)(O2) Htr IX, XII, XIII XVII, XVIII (?) CheAW

Htr IV -VII Htr XIV -XVI (?) HtrVIII (O2) HtrIII (BasT) (Cys, lle, Leu, Met, Val)

Nutrient uptake

Energy production

Signal transduction
Sensor kinases CheY Flagellar Motor

DppABCDF

Dipeptides

Semi-phosphorylated EntnerDoudoroff
Glucose Gluconate 2-keto-3-deoxygluconate

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

AppBCF

Oligopeptides

2-keto-3-deoxy-6p-gluconate
PutP Proline

Response regulators

Sensor kinases

Glyceraldehyde 3-P

Pyruvate

Cl

Fatty acid oxidation

Cat Cationic amino acids

Chloride channel CysAT

Pyruvate
Amino acid ABC transporter Amino acids

Acetyl Co-A

Acetate Arginine

2 PO 4

3 PO 4

PstABC Phosphate permease ArsAB

Protein translocation
SecDEFY SR K+ 7S RNA SRP54 SRP19 K+ H+

Aspartate

Citric acid cycle

Anion transport
Glutamate

3 PO 4 3 AsO4 1 AsO2

Cation transport
Co 2+ Cu 2+ Fe 3+ Fe 3+

ArsC Zn2+

AsO 1

Na+ TrkAH KdpABC NhaC

Cd 2+

Cd 2+ CbiNOQ NosFY Iron permease HemUV ZurMA

ZntX Cd efflux ATPase

Figure 4 Integrated view of the biology of the extremely halophilic archaeon Halobacterium derived from its genome sequence. Many informational and operational processes revealed from the genome sequence are shown. Transporters in the membrane are highlighted, including light-driven proton and chloride pumps, bacteriorhodopsin (BR) and halorhodopsin (HR), and the sodium/proton antiporter (NhaC), potassium uniporter (TrkAH and KdpABC), dipeptide and amino acid transporters and anion transporters (Ng et al., 2000. & 2000 National Academy of Sciences, USA.)

Halophiles

Halophiles

(a)

(b)

Figure 5 Extremely halophilic archaeal (a) and human (b) protein-DNA complexes. Models of similar transcription initiation complexes, showing the protein surface charges (red for acidic or negative and blue for basic or positive), surrounding the DNA double helix. The haloarchaeal proteins are acidic whereas the human proteins are basic. Adapted from DasSarma et al. (2006) & S. DasSarma.

haloarchaea, organisms whose primary metabolism is aerobic, and that live in environments in which the solubility of molecular oxygen is low (due to high salinity and elevated temperatures), is to enable the cells to oat to the more oxygenated surface layers. This also increases the availability of light for purple membrane-mediated photophosphorylation. See also: Bacterial and Archaeal Inclusions Haloarchaeal mutants, which have lost the ability to produce purple membrane, gas vesicles, or carotenoids, occur spontaneously at a high-frequency. Genetic analysis has shown that recombinational events promoted by transposable insertion sequences are responsible for the observed genetic instabilities (DasSarma, 1989). Extrachromosomal replicons, the structure and function of which have been revealed through genome sequencing, are commonly found to be reservoirs of insertion sequences (Ng et al., 1998). Several of these replicons contain essential genes and have been proposed to function as required minichromosomes rather than dispensable plasmids. The nding of plasmids and insertion sequences has aided the development of genetic tools for analysis and manipulation of these organisms. Haloarchaea are members of the archaeal domain and are phylogenetically distinct from both bacteria and eukaryotes. As such, they exhibit characteristic features, including eukaryotic-like deoxyribonucleic acid (DNA) replication, transcription, and translation machinery, branched-chain, ether-linked archaeal lipids, and a cell wall S-layer composed of a glycoprotein, similar to that found in some bacteria. See also: Archaeal Cell Walls

(Figure 3). This species can grow over a wide range of salt concentrations, from 2 to 5 M NaCl, is an extreme halophile with a salt optimum of 3.5 M, which lyses in distilled water. It uses glycine betaine as its major compatible solute (Figure 6), which it can either take up from the medium or synthesise from choline. A. halophytica and similar unicellular cyanobacteria have been described from Great Salt Lake, Dead Sea, Solar Lake and articial solar ponds. Another planktonic cyanobacterium reported from Great Salt Lake is Dactylococcopsis salina. A thermophilic species has been identied in a geothermal seawater lagoon in southwest Iceland (Banerjee et al., 2009). A variety of lamentous cyanobacteria, for example in the order Oscillatoriales, such as Oscillatoria neglecta, Oscillatoria limnetica and Phormidium ambiguum, develop in the green second layer of mats in hypersaline lakes (Fourc ans et al., 2004; Figure 3). These are more moderate halophiles, usually growing optimally at 12.5 M NaCl, which form heterocysts and x nitrogen. Another common species in the same family is Coleofasciculus chthonoplastes (formerly Microcoleus chthonoplastes). See also: Cyanobacteria

Other phototrophic bacteria

Some phototrophic bacteria occur beneath the cyanobacterial layers in anaerobic, but lighted zones in hypersaline microbial mats (Fourc ans et al., 2004; Figure 3). They usually grow anaerobically by anoxygenic photosynthesis, although many also have the capacity to grow aerobically as heterotrophs. They can use reduced sulfur (hydrogen sulde, elemental sulfur), organic compounds, or hydrogen as electron donors. They include green and purple sulfur and nonsulfur bacteria and are characterised by their use of bacteriochlorophyll pigments. The green sulfur bacteria, such as the slight to moderately halophilic Chlorobium limicola and Chlorobium phaeobacteroides, deposit elemental sulfur granules outside their cells and are capable of nitrogen xation. C. limicola can take up glycine betaine

Cyanobacteria
Cyanobacteria dominate the planktonic biomass and form microbial mats in many hypersaline lakes. The top brown layer of saline microbial mats contains a common unicellular cyanobacterial species, Aphanothece halophytica
6

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

O O O O

Anaerobic bacteria and archaea

Glycine betaine

NH2 OH

Ectoine

HO

OH

Glycerol

Figure 6 Structure of three common compatible solutes in halophiles. Zwitterionic forms of glycine betaine and ectoine, and the neutral glycerol are commonly found in halophilic microorganisms and help to balance the osmotic stress of the environment.

from the environment and synthesises trehalose for use as osmolytes. The moderately halophilic, anoxygenic, phototrophic, lamentous green nonsulfur bacteria such as Chloroexus aurantiacus can also be slightly thermophilic. See also: Green Sulfur Bacteria Halophilic purple sulfur bacteria include Halochromatium glycolicum, which grows photoorganotrophically using glycolate and glycerol, Halochromatium salexigens, and Thiocystis violascens. They synthesise N-acetylglutaminylglutamine amide as a minor component of their compatible solute and use sucrose and glycine betaine from their environment. Thiocapsa roseopersicina and Thiohalocapsa halophila, from Guerrero Negro both synthesise sucrose and take up glycine betaine from the environment. T. halophila also synthesises glycine betaine and N-acetylglutaminylglutamine amide for osmoprotection. The purple nonsulfur bacteria Rhodothalassium salexigens from evaporated seawater pools and Rhodovibrio salinarum from a saltern both use glycine betaine and ectoine as osmolytes (Figure 6). The purple sulfur bacteria, such as Ectothiorhodospira species, dominate alkaline soda lakes (Ollivier et al., 1994). The moderate halophile Ectothiorhodospira mobilis is a strict anaerobe and uses carboxamines as compatible solutes and the osmolyte N-a-carbamoyl-L-glutamine1-amide to counter osmotic stress. The extreme halophile, Halorhodospira halochloris, isolated from Wadi Natrun, was the rst bacterium shown to synthesise and accumulate ectoine, a cyclic amino acid, which it uses along with glycine betaine and trehalose as a compatible solute.

Sulfur oxidising bacteria

Below the cyanobacteria and the phototrophic bacteria in microbial mats, halophilic, lamentous, carbon dioxidexing bacteria oxidise hydrogen sulde (and elemental sulfur) to sulfate (Figure 3). Examples include lamentous Beggiatoa alba from Guerrero Negro and Beggiatoa leptiformis from Solar Lake, and the unicellular proteobacterium Halothiobacillus halophilus from Lake OGrady in Western Australia.

A large variety of facultative and strictly anaerobic bacteria and archaea inhabit the bottom layers of microbial mat communities and sediment in hypersaline lakes (Ollivier et al., 1994, Figure 3). These include fermentative bacteria, homoacetogenic bacteria, sulfate-reducing bacteria and methanogenic archaea. Fermentative anaerobic bacteria, which grow at saturated NaCl concentrations have been described. One example is Halanaerobacter chitinivorans, isolated from a saltern, which is capable of fermenting chitin from brine shrimp and brine ies. Another more moderate halophilic isolate, Halocella cellulolsilytica, ferments carbohydrates including cellulose. Sporohalobacter lortetii and Orenia marismortui are sporogenous and ferment carbohydrates. Several homoacetogens, strict anaerobes which produce acetate from oxidation of sugars or amines, have been described. For example, Halanaerobium saccharolyticum ferments carbohydrates and N-acetylglucosamine and can grow at a wide range of NaCl concentrations. The extremely halophilic Acetohalobium arabaticum (with a salt tolerance of 14.5 M NaCl), isolated from Lake Sivash, grows on glycine betaine and trimethylamine and can reduce carbon dioxide to acetate. It is a likely competitor of sulfate-reducing bacteria for hydrogen. Sulfate-reducing bacteria use sulfate as the terminal electron acceptor, although many can also utilise other sulfur compounds, nitrate and fumarate. They dier in their ability to oxidise dierent compounds, though most use low-molecular weight organic compounds like lactate, pyruvate, ethanol, and volatile fatty acids, or hydrogen as electron donors. A few can use carbon dioxide as their sole carbon source. Although many slightly halophilic sulfate reducers have been isolated, mostly from marine environments, relatively few of these can survive at an extremely high salinity. Desulfohalobium retbaense, isolated from Lake Retba, Senegal, and Desulfovibrio halophilus, from Solar Lake, are two moderately halophilic species which can grow at up to 4 M NaCl, but only relatively slowly. Another isolate, from the deep-sea hypersaline pools in the Red Sea, is similar to D. halophilus. The osmoregulation of sulfate-reducing bacteria likely occurs by accumulation of salts internally. Among anaerobic archaea, halophilic methanogens are generally methylotrophic and are strict anaerobes. Several, mostly moderate halophilic methanogens have been identied, including Methanohalophilus halophilus from a microbial mat, Methanohalophilus mahii from Great Salt Lake and Methanohalophilus portucalensis from a Portuguese saltern. The slight halophile, Methanosalsum zhilinae, is also an alkaliphile and a slight thermophile. The extremely halophilic methanogen, Methanohalobium evestigatum, with a NaCl optimum of 4.5 M, is also a thermophile with a temperature optimum of 508C. Methanogenesis has been reported from moderately to extremely high salinity deep-sea brine pools in the Gulf of Mexico and Mediterranean Sea. Methanogens use
7

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

b-amino acids (b-glutamine, N-a-acetyl-b-lysine) as compatible solutes and play an important role in the anaerobic degradation of glycine betaine in their environments. Their intracellular salt concentration is somewhat higher than in most bacteria (approximately 0.6 M KCl) but is signicantly lower than that found in extremely halophilic archaea. Recently, a community of the halophilic archaea has been reported from sulde and sulfur-rich springs, including one described species, Haloferax sulfurifontis (Elshahed et al., 2004). See also: Euryarchaeota; Methanogenesis Biochemistry; Methanogenesis: Ecology

Aerobic and facultative anaerobic Gram-negative bacteria

Many moderately halophilic, heterotrophic Gramnegative bacteria belonging to the Halomonas and Chromohalobacter genera have been described. Other genera with halophilic representatives include Salinivibrio, Arhodomonas, Pseudomonas, Flavobacterium, Alcaligenes, Alteromonas, Acinetobacter and Spirochaeta. Most of these are heterotrophs, and include the nitrate reducing Arhodomonas aquaeolei, isolated from subterranean brine associated with an oil eld, and Chromohalobacter marismortui, from the Dead Sea. Others include Chromohalobacter beijerinckii, from salted beans preserved in brine, Pseudomonas halophila, from Great Salt Lake and Salinivibrio costicola, originally isolated from Australian bacon. Several Halomonas species are capable of nitrate reduction, including Halomonas elongata, isolated from a solar saltern, and Halomonas halodenitricans, isolated from meat curing brines. Others include Halomonas eurihalina, isolated from saline soil, which produces an extracellular polysaccharide, Halomonas halodurans, from estuarine waters, which is capable of degrading aromatic compounds, Halomonas pantelleriensis, which grows at a pH optimum of 9, and Halomonas subglaciescola, from beneath the ice of Organic Lake in Antarctica. These organisms primarily use glycine betaine or ectoine as compatible solutes. Genes for uptake of glycine betaine from the medium and its synthesis from choline, and synthesis of ectoine have been cloned from Halomonas species novas and some other halophiles (Bestvater et al., 2008; Ca et al., 2000). Among spirochetes, the moderate halophile, Spirochaeta halophila, found in Solar Lake, is a chemolithotroph capable of iron and manganese oxidisation. The avobacteria, Psychroexus gondwanensis and Salegentibacter salegens, are psychrotolerant halophiles isolated from Antarctic Lakes. See also: Spirochaetes

pigments, Nesterenkonia lacusekhoensis, from the hypersaline Ekho Lake in East Antarctica, Tetragenococcus halophilus, from fermented soy sauces, squid liver sauce and brine for curing anchovies, which is capable of lactic acid fermentation, and several Salinicoccus species from salterns. Other examples include Gracilibacillus dipsosauri, from the nasal cavity of a desert iguana, Bacillus haloalkaliphilus, from Wadi Natrun and Virgibacillus halodenitricans, from a solar saltern in southern France. Halobacillus litoralis and Halobacillus trueperi were isolated from Great Salt Lake. Halobacillus halophila is an endospore forming bacterium, from which the compatible solute N-e-acetyllysine was originally isolated. Many of these organisms use proline, ectoine, or N-acetylated diamino acids, which they are capable of synthesising, as compatible solutes. See also: Gram-type Positive Bacteria Halophilic actinomycetes from saline soils include Actinopolyspora halophila, which grows best at moderate NaCl concentrations, and is one of the few heterotrophic bacteria which can synthesise the compatible solute glycine betaine.

Eukaryotic Halophiles
Algae
At moderately high salinities (13.5 M NaCl) dense populations (4105 per mL) of green algae are supported (Javor, 1989). Dunaliella species, for example Dunaliella salina, Dunaliella parva and Dunaliella viridis, are ubiquitous and are the main source of food for brine shrimp and the larvae of brine ies. Most species of green algae are moderate halophiles, with only a few extremely halophilic species, for example Dunaliella salina and Asteromonas gracilis, which are capable of slow growth at up to saturated NaCl concentrations (Figure 2). The algae predominantly use polyols as compatible solutes. Dunaliella salina synthesises glycerol in response to osmotic stress (Chen and Jiang, 2009) (Figure 6). The cytoplasmic concentration of glycerol can reach 7 M when grown in medium containing 5 M NaCl and can constitute over 50% of the dry weight of cells. However, the intracellular sodium concentration has been reported to be less than 100 mM over a wide range of external salt concentrations. During moderate dilution stress glycerol does not leak out of cells, but is instead metabolised and transformed into osmotically inactive reserve material. Diatoms are algae surrounded by silica cell walls and are commonly found, but rarely abundant in hypersaline environments. A variety of species have been found at approximately 2 M NaCl, and the upper limit for diatom growth is reported to be approximately 3 M NaCl. Examples of common diatoms include Amphora coeaeformis and Nitzschia and Navicula species. Though osmoregulation has not been extensively studied in diatoms, accumulation of proline and oligosaccharides has been reported in some species. See also: Diatoms

Gram-positive bacteria
This group includes moderately halophilic species of the genera Nesterenkonia, Tetragenococcus, Salinicoccus, Bacillus, Halobacillus and Marinococcus (Ventosa et al., 1998). They include cocci such as Nesterenkonia halobia, isolated from salterns, which produce yellow-red carotenoid
8

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

Protozoa
A large variety of protozoa exist in hypersaline environments, but few have been extensively described. One moderately halophilic ciliate, Fabrea salina, has been isolated from several saline lakes from Africa to Australia. Although in freshwater, protozoa are known to regulate osmotic pressure with contractile vacuoles that expel water, their mechanism of osmoregulation in hypersaline brine has not yet been investigated. See also: Protozoan Diversity and Biogeography; Protozoan Ecology

Fungi
Yeasts and lamentous fungi are well adapted to tolerate hypersaline environments. They grow best under aerobic conditions on carbohydrates at moderate temperatures and acidic to neutral pH. Debaryomyces hansenii, a halotolerant yeast, isolated from seawater can grow aerobically in salinities of up to 4.5 M NaCl, and has been studied extensively by genome sequencing and transcriptomic analysis (Gonzalez et al., 2009). It produces glycerol as a compatible solute during logarithmic phase and arabitol in stationary phase. Hortaea werneckii, a melanised fungus, was isolated from hypersaline waters of solar salterns and its osmoresponsive genes have been identied by transcriptomic methods (Lenassi et al., 2007). It is one of the most salt tolerant fungi known, growing at a wide range of salinity up to saturating NaCl concentration. Xerophilic, halophilic fungi, for example Polypaecilum pisce have also been isolated from salted sh. See also: Fungal Ecology

Multicellular eukaryotes
There are a surprising number of invertebrates that can survive in hypersaline environments (Javor, 1989). Insects successful in hypersaline environments include brine ies, such as Ephydra cinerea, and brine shrimp, Artemia franciscana and related species. Other invertebrates include rotifers such as Keratella quadrata, at worms such as Macrostomum species, copepods such as Robertsonia salsa, and ostracods such as Cyprideis torosa. A variety of obligate and facultative halophytic plants, for example Atriplex halimus, Mesembryanthemum crystallinum and Laguncularia mangrove species can also survive in soils with moderately high salinity. Few vertebrates can survive hypersaline conditions, with Tilapia species observed in moderately high salinity. Many hypersaline environments support a wide diversity of birds, one of the most spectacular of which is the pink amingo. This bird is born white and obtains its colour from the pigments of halophilic microorganisms in its food.

recent trends are to exploit the many novel and unique molecules (e.g. enzymes, bacteriorhodopsins, etc.) in these organisms, for molecular biotechnological applications (DasSarma et al., 2010a; Oren, 2010). A most useful halophilic biomolecule type is their retinal-containing chromoproteins, which are being used for diverse applications, such as in biocomputing (Birge, 1995), as a lightsensitive neurological probe (Gradinaru et al., 2009), and for treatment of blindness from retinitis pigmentosa (Busskamp et al., 2010). Halophilic microorganisms also produce many stable enzymes (including hydrolytic enzymes such as DNases, lipases, amylases, gelatinases and proteases) capable of functioning under conditions, which would lead to precipitation or denaturation of most other proteins. Compatible solutes of halophilic bacteria are used in cosmetics and improving hydration properties generally (Bestvater et al., 2008). Halophilic proteins also compete eectively with salts for hydration, a property which may result in their functioning in low water activity environments, including organic solvents. Biodegradable polymers such as polyhydroxyalkanoates are produced in large quantities by some halophilic microbes (Han et al., 2010), and have both medical and other specialised applications. A variety of other novel halophilic biomolecules also have been targeted for commercial applications, for example, gas vesicles for bioengineering oating particles, pigments for food colouring and metabolites as stress protectants. Halophiles are becoming increasingly valuable for bioremediation since many industrial processes release salts as contaminated brine euent into the environment. Another growing area for applications of halophiles is in the development of sustainable bioenergy technology to address concerns about petroleum shortage and global warming. See also: Bioremediation

Conclusions and Future Prospects

Halophiles are an interesting class of extremophilic organisms which have adapted to harsh, hypersaline conditions. They can compete successfully for water and resist the denaturing eects of salts. A wide variety of microorganisms are halophilic, including extremely halophilic and methanogenic archaea, cyanobacteria, and green and purple bacteria, sulfur oxidising bacteria, anaerobic fermentative, homoacetogenic, sulfate reducing bacteria, and Gram-negative and Gram-positive heterotrophic bacteria. In addition, some algae, protozoa, fungi and a few higher organisms have adapted to life in high salinity. Adaptation to hypersaline conditions is interesting from an evolutionary standpoint. The earliest prokaryotic fossils found in ancient stromatolites (which may be more than 3.5 billion years old), are very similar in appearance to the microbial mats found in modern hypersaline ponds. The halophilic and methanogenic archaea are both members of the archaeal branch of the phylogenetic tree which likely appeared very early in evolution. These ndings, and the
9

Biotechnology
Although traditional commercial uses of halophiles have been in the food and nutraceutical industries (e.g. fermentation of soy and sh sauces, b-carotene production),

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

likelihood for concentration of brines during prebiotic evolution, suggest that adaptation to salts may have been among the earliest evolutionary inventions (Dundas, 1998). Future studies will provide greater insights into these fascinating evolutionary questions. The diversity of microorganisms in hypersaline environments is also of growing interest in research and education. Few hypersaline environments have been carefully surveyed using molecular methods. Recent ndings of bacterial and archaeal metabolic activities suggest that these environments harbour diverse communities of microbes, many of which have not been cultured. Metagenomic studies of hypersaline environments are beginning to yield interesting results, for example the discovery of a tiny nano species that is smaller than most free-living species. The possibility that extreme halophiles may survive on other planets, such as some recently reported saline environment on Mars, is intriguing. Because of such novel characteristics and their relative safety, some halophiles have become popular for educational uses in schools and colleges (DasSarma, 2006, 2007). The molecular mechanisms used for adaptation to hypersaline conditions are now being investigated using modern technologies. Powerful tools for genetic and genomic analysis of halophiles, including DNA microarrays, gene knockouts, and proteomics, are now routinely used (e.g. see DasSarma et al., 2006). Halophilic organisms can exclude salts by synthesis of an equally high concentration of uncharged compatible solutes or osmolytes, and contain stable macromolecules which can withstand the denaturing eects of salts. The genes and proteins involved in synthesis and accumulation of compatible solutes and their regulation are being explored. For archaea, recent structure-function studies are aimed at determining how their highly acidic proteins can function in high salinity (Bergqvist et al., 2003; Britton et al., 2006). Finally, as a result of natural and man-made environmental changes, desiccated and saline environments are on the increase globally, and are likely to have signicant eects on the biosphere in the twenty-rst century. As a result, a better understanding of halophiles is needed to address future challenges for conservation of ecologically sensitive areas, as well as to engineer plants and sh to grow in more saline environments, and exploit halophilic microbes for many other opportunities in biotechnology.

References
Abu Ghazleh S, Abed AM and Kempe S (2011) The dramatic drop of the dead sea: background, rates, impacts and solutions In: Badescu V and Cathcart RB (eds) Macro-engineering Seawater in Unique Environments, pp. 77105. Springer. Anderson I, Scheuner C, Go ker M et al. (2011) Novel insights into the diversity of catabolic metabolism from ten haloarchaeal genomes. PLoS One 6: e20237. Antunes A, Ngugi DK and Stingl U (2011) Microbiology of the Red Sea (and other) deep-sea anoxic brine lakes. Environmental Microbiology Reports 3: 4.

Ban N, Nissen P, Hansen J, Moore PB and Steitz TA (2000) The complete atomic structure of the large ribosomal subunit at resolution. Science 289: 905920. 2.4 A Banerjee M, Everroad RC and Castenholz RW (2009) An unusual cyanobacterium from saline thermal waters with relatives from unexpected habitats. Extremophiles 13: 707716. Bergqvist S, Williams MA, OBrien R and Ladbury JE (2003) Halophilic adaptation of proteinDNA interactions. Biochemical Society Transactions 31: 677680. Bestvater T, Louis P and Galinski EA (2008) Heterologous ectoine production in Escherichia coli: by-passing the metabolic bottle-neck. Saline Systems 4: 12. Birge RR (1995) Protein-based computers. Scientic American 272: 6671. Bonete MJ, Mart nez-Espinosa RM, Pire C, Zafrilla B and Richardson DJ (2008) Nitrogen metabolism in haloarchaea. Saline Systems 4: 9. Borin S, Brusetti L, Mapelli F et al. (2009) Sulfur cycling and methanogenesis primarily drive microbial colonization of the highly suldic Urania deep hypersaline basin. Proceedings of the National Academy of Sciences of the USA 106: 91519156. Bowers KJ, Mesbah NM and Wiegel J (2006) Biodiversity of polyextremophilic Bacteria: does combining the extremes of high salt, alkaline pH and elevated temperature approach a physicochemical boundary for life? Saline Systems 5: 9. Britton KL, Baker PJ, Fisher M et al. (2006) Analysis of protein solvent interactions in glucose dehydrogenase from the extreme halophile Haloferax mediterranei. Proceedings of the National Academy of Sciences of the USA 103: 48464851. Busskamp V, Duebel J, Balya D et al. (2010) Genetic reactivation of cone photoreceptors restores visual responses in retinitis pigmentosa. Science 329: 413417. novas D, Vargas C, Kneip S et al. (2000) Genes for the synthesis Ca of the osmoprotectant glycine betaine from choline in the moderately halophilic bacterium Halomonas elongata DSM 3043, USA. Microbiology 146: 455463. Capes MD, Coker JA, Gessler R et al. (2011) The information transfer system of halophilic archaea. Plasmid 65: 77101. Caumette P (1993) Ecology and physiology of phototrophic bacteria and sulphate-reducing bacteria in marine salterns. Experientia 49: 473481. Chen H and Jiang JG (2009) Osmotic responses of Dunaliella to the changes of salinity. Journal of Cellular Physiology 219: 251258. Crowley DJ, Boubriak I, Berquist BR et al. (2006) The uvrA, uvrB and uvrC genes are required for repair of ultraviolet light induced DNA photoproducts in Halobacterium sp. NRC-1. Saline Systems 2: 11. DasSarma S (1989) Mechanisms of genetic variability in Halobacterium halobium: the purple membrane and gas vesicle mutations. Canadian Journal of Microbiology 35: 6572. DasSarma S (2004) Genome sequence of an extremely halophilic archaeon. In: Fraser C, Read T and Nelson KE (eds) Microbial Genomes, pp. 383399. Totowa, NJ: C.M. Humana Press, Inc. DasSarma S (2006) Extreme halophiles are models for astrobiology. Microbe 1: 120127. DasSarma S (2007) Extreme microbes. American Scientist 95: 224231. DasSarma P and DasSarma S (2008) On the origin of prokaryotic species: the taxonomy of halophilic Archaea. Saline Systems 4: 5.

10

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

Halophiles

DasSarma S, Kennedy SP, Berquist BR et al. (2001) Genomic perspective on the photobiology of Halobacterium species NRC-1, a phototrophic, phototactic, and UV-tolerant haloarchaeon. Photosynthesis Research 70: 317. DasSarma S, Berquist BR, Coker JA, DasSarma P and Mu ller JA (2006) Post-genomics of the model haloarchaeon Halobacterium sp. NRC-1. Saline Systems 2: 3. DasSarma P, Coker JA, Huse V and DasSarma S (2010a) Halophiles, Biotechnology. In: Flickinger MC (ed.) Encyclopedia of Industrial Biotechnology, Bioprocess, Bioseparation, and Cell Technology, pp. 27692777. John Wiley & Sons Ltd. DasSarma P, Klebahn G and Klebahn H (2010b) Translation of Henrich Klebahns Damaging agents of the klippsh a contribution to the knowledge of the salt-loving organisms. Saline Systems 6: 7. Dundas I (1998) Was the environment for primordial life hypersaline? Extremophiles 2: 375377. Elshahed MS, Najar FZ, Roe BA et al. (2004) Survey of archaeal diversity reveals an abundance of halophilic Archaea in a lowsalt, sulde- and sulfur-rich spring. Applied and Environmental Microbiology 70: 22302239. Fish SA, Shepherd TJ, McGenity TJ and Grant WD (2002) Recovery of 16S ribosomal RNA gene fragments from ancient halite. Nature 417: 432436. Erratum in: Nature (2002) 420:202. Fourc ans A, de Oteyza TG, Wieland A et al. (2004) Characterization of functional bacterial groups in a hypersaline microbial mat community (Salins-de-Giraud, Camargue, France). FEMS Microbiology Ecology 51: 5570. zquez A, Ortiz Zuazaga HG et al. (2009) GenGonzalez NA, Va ome-wide expression proling of the osmoadaptation response of Debaryomyces hansenii. Yeast 26: 111124. Gradinaru V, Mogri M, Thompson KR, Henderson JM and Deisseroth K (2009) Optical deconstruction of parkinsonian neural circuitry. Science 324: 354359. Gwynn JW (2002) Great Salt Lake: An Overview of Change. Utah: Utah Department of Natural Resources. de la Haba RR, del Carmen Marquez M, Papke RT and Ventosa A (2011) Multilocus sequence analysis (MLSA) of the family Halomonadaceae. International Journal of Systematic and Evolutionary Microbiology doi: 10.1099/ijs.0.032938-0 IJSEM. Han J, Li M, Hou J et al. (2010) Comparison of four phaC genes from Haloferax mediterranei and their function in dierent PHBV copolymer biosyntheses in Haloarcula hispanica. Saline Systems 6: 9. Javor B (1989) Hypersaline Environments, Microbiology and Biogeochemistry. Berlin: Springer-Verlag. Joo WA and Kim CW (2005) Proteomics of Halophilic archaea. Journal of Chromatography B. Analytical Technologies in the Biomedical and Life Sciences 815: 237250. , Thomas LK, Erb TJ and Berg IA Khomyakova M, Bu kmez O (2011) A methylaspartate cycle in haloarchaea. Science 331: 334337. Lanyi JK (2004) Bacteriorhodopsin. Annual Review of Physiology 66: 665688. Lenassi M, Vaupotic T, Gunde-Cimerman N and Plemenitas A (2007) The MAP kinase HwHog1 from the halophilic black

yeast Hortaea werneckii: coping with stresses in solar salterns. Saline Systems 3: 3. Madern D, Ebel C and Zaccai G (2000) Halophilic adaptation of enzymes. Extremophiles 4: 9198. McEwen AS, Ojha L, Dundas CM et al. (2011) Seasonal Flows on Warm Martian Slopes. Science 333: 740743. Minegishi H, Kamekura M, Kitajima-Ihara T et al. (2011) Gene orders in the upstream of 16S rRNA genes divide genera of the family Halobacteriaceae into two groups. International Journal of Systematic and Evolutionary Microbiology doi:ijs.0.031708-0. Narasingarao P, Podell S, Ugalde JA et al. (2011) De novo metagenomic assembly reveals abundant novel major lineage of Archaea in hypersaline microbial communities. The ISME Journal. doi:10.1038/ismej.2011.78. Ng WV, Ciufo SA, Smith TM et al. (1998) Snapshot of a large dynamic replicon in a halophilic archaeon: megaplasmid or minichromosome? Genome Research 8: 11311141. Ng WV, Kennedy SP, Mahairas GG et al. (2000) Genome sequence of Halobacterium species NRC-1. Proceedings of the National Academy of Sciences of the USA 97: 1217612181. Ollivier B, Caumette P, Garcia JL and Mah RA (1994) Anaerobic bacteria from hypersaline environments. Microbiology Reviews 58: 2738. Oren A (2010) Industrial and environmental applications of halophilic microorganisms. Environmental Technology 31: 825834. Oxley APA, Lanfranconi MP, Wu rdemann D et al. (2010) Halophilic archaea in the human intestinal mucosa. Environmental Microbiology 12: 23982410. Papke RT, White E, Reddy P et al. (2011) A Multilocus Sequence Analysis (MLSA) approach to Halobacteriales phylogeny and taxonomy. International Journal of Systematic and Evolutionary Microbiology 61: 29842995. Roberts MF (2005) Organic compatible solutes of halotolerant and halophilic microorganisms. Saline Systems 1: 5. Sato T and Atomi H (2011) Novel metabolic pathways in Archaea. Current Opinion in Microbiology 14: 307314. rquez MC, Garabito MJ and Arahal DR (1998) Ventosa A, Ma Moderately halophilic gram-positive bacterial diversity in hypersaline environments. Extremophiles 2: 297304. van der Wielen PW, Bolhuis H, Borin S et al. (2005) The enigma of prokaryotic life in deep hypersaline anoxic basins. Science 307: 121123. Yancey PH (2005) Organic osmolytes as compatible, metabolic and counteracting cytoprotectants in high osmolarity and other stresses. Journal of Experimental Biology 208: 28192830.

Further Reading
DasSarma S, Fleischmann EM, Robb FT et al. (eds) (1995) Archaea, A Laboratory Manual Halophiles. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press. Gunde-Cimerman N, Oren A and Plemenitas A (eds) (2005) Adaptation to Life at High Salt Concentrations in Archaea, Bacteria, and Eukarya. Dordrecht, Netherlands: Springer. Vreeland RH and Hochstein LI (eds) (1993) The Biology of Halophilic Bacteria. Boca Raton, FL: CRC Press, Inc.

eLS & 2012, John Wiley & Sons, Ltd. www.els.net

11