Mercury resistance and volatilization by oil utilizing haloarchaea under hypersaline conditions

The hydrocarbon utilizing haloarchaea, Haloferax (two strains), Halobacterium and Halococcus from a hypersaline coastal area of the Arabian Gulf, had the potential for resistance and volatilization of Hg²⁺. Individual haloarchaea resisted up to between 100 and 200 ppm HgCl₂ in hydrocarbon free media with salinities between 1 and 4 M NaCl, but only up to between 20 and 30 ppm in a mineral medium containing 3 M NaCl, with 0.5% (w/v) crude oil, as a sole source of carbon and energy. Halococcus and Halobacterium volatilized more mercury than Haloferax. The individual haloarchaea consumed more crude oil in the presence of 3 M NaCl than in the presence of 2 M NaCl. At both salinities, increasing the HgCl₂ concentration in the medium from 0 to 20 ppm resulted in decreasing the oil consumption values by the individual haloarchaea. However, satisfactory oil consumption still occurred in the presence of 10 ppm HgCl₂. It was concluded that haloarchaea with the combined potential for mercury resistance and volatilization and hydrocarbon consumption could be useful in removing toxic mercury forms effectively from oil free, mercury contaminated, hypersaline environments, and mercury and oil, albeit less effectively, from oily hypersaline environments.     

Isolation of hydrocarbon-degrading extremely halophilic archaea from an uncontaminated hypersaline pond (Camargue, France)

Little information exists about the ability of halophilic archaea present in hypersaline environments to degrade hydrocarbons. In order to identify the potential actors of hydrocarbon degradation in these environments, enrichment cultures were prepared using samples collected from a shallow crystallizer pond with no known contamination history in Camargue, France, with n-alkanes provided as source of carbon and energy. Five alkane-degrading halophilic archaeal strains were isolated: one (strain MSNC 2) was closely related to Haloarcula and three (strains MSNC 4, MSNC 14, and MSNC 16) to Haloferax. Biodegradation assays showed that depending on the strain, 32 to 95% (0.5 g/l) of heptadecane was degraded after 30 days of incubation at 40°C in 225 g/l NaCl artificial medium. One of the strains (MSNC 14) was also able to degrade phenanthrene. This work clearly shows for the first time the potential role of halophilic archaea belonging to the genera Haloarcula and Haloferax in the degradation of hydrocarbons in both pristine and hydrocarbon-contaminated hypersaline environments.     

Oil-bioremediation potential of two hydrocarbonoclastic, diazotrophic Marinobacter strains from hypersaline areas along the Arabian Gulf coasts

Two halophilic, hydrocarbonoclastics bacteria, Marinobacter sedimentarum and M. flavimaris, with diazotrophic potential occured in hypersaline waters and soils in southern and northern coasts of Kuwait. Their numbers were in the magnitude of 10³ colony forming units g^?1. The ambient salinity in the hypersaline environments was between 3.2 and 3.5 M NaCl. The partial 16S rRNA gene sequences of the two strains showed, respectively, 99 and 100 % similarities to the sequences in the GenBank. The two strains failed to grow in the absence of NaCl, exhibited best growth and hydrocarbon biodegradation in the presence of 1 to 1.5 M NaCl, and still grew and maintained their hydrocarbonoclastic activity at salinities up to 5 M NaCl. Both species utilized Tween 80, a wide range of individual aliphatic hydrocarbons (C₉–C₄₀) and the aromatics benzene, biphenyl, phenanthrene, anthracene and naphthalene as sole sources of carbon and energy. Experimental evidence was provided for their nitrogen-fixation potential. The two halophilic Marinobacter strains successfully mineralized crude oil in nutrient media as well as in hypersaline soil and water microcosms without the use of any nitrogen fertilizers.     

Temperature and pH optima of extremely halophilic archaea: a mini-review

Archaeal microorganisms that grow optimally at Na⁺ concentrations of 1.7 M, or the equivalent of 10% (w/v) NaCl, and greater are considered to be extreme halophiles. This review encompasses extremely halophilic archaea and their growth characteristics with respect to the correlation between the extent of alkaline pH and elevated temperature optima and the extent of salt tolerance. The focus is on poly-extremophiles, i.e., taxa growing optimally at a Na⁺ concentration at or above 1.7 M (approximately 10% w/v NaCl); alkaline pH, at or above 8.5; and elevated temperature optima, at or above 50°C. So far, only a very few extreme halophiles that are able to grow optimally under alkaline conditions as well as at elevated temperatures have been isolated. The distribution of extremely halophilic archaea growing optimally at 3.4 M Na⁺ (approximately 20% w/v NaCl) is bifurcated with respect to pH optima, either they are neutrophilic, with a pH_opt of approximately 7, or strongly alkaliphilic, with pH_opt at or above 8.5. Amongst these extreme halophiles which have elevated pH optima, only four taxa have an optimum temperature above 50°C: Haloarcula quadrata (52°C), Haloferax elongans (53°C), Haloferax mediterranei (51°C) and Natronolimnobius ‘aegyptiacus’ (55°C).     

<Emphasis Type="Italic">Desulfonatronovibrio halophilus</Emphasis> sp. nov., a novel moderately halophilic sulfate-reducing bacterium from hypersaline chloride–sulfate lakes in Central Asia

Four strains of lithotrophic sulfate-reducing bacteria (SRB) have been enriched and isolated from anoxic sediments of hypersaline chloride–sulfate lakes in the Kulunda Steppe (Altai, Russia) at 2 M NaCl and pH 7.5. According to the 16S rRNA gene sequence analysis, the isolates were closely related to each other and belonged to the genus Desulfonatronovibrio, which, so far, included only obligately alkaliphilic members found exclusively in soda lakes. The isolates utilized formate, H₂ and pyruvate as electron donors and sulfate, sulfite and thiosulfate as electron acceptors. In contrast to the described species of the genus Desulfonatronovibrio, the salt lake isolates could only tolerate high pH (up to pH 9.4), while they grow optimally at a neutral pH. They belonged to the moderate halophiles growing between 0.2 and 2 M NaCl with an optimum at 0.5 M. On the basis of their distinct phenotype and phylogeny, the described halophilic SRB are proposed to form a novel species within the genus Desulfonatronovibrio, D. halophilus (type strain HTR1^T = DSM24312^T = UNIQEM U802^T).     

Effects of different ion compositions on growth of obligately halophilic protozoan Halocafeteria seosinensis

Substantial halophilic organisms have been found in 100–200‰ salinities. These ranges represent a highly specialized halophilic environment to which only a few halotolerant species have adapted. Recent studies have underlined the existence of diverse obligately halophilic protozoa in the salinity ranges of 100–200‰. The ranges of salinity under which these organisms can grow have been examined to some extent, but the balance of specific ions that will support growth has not been investigated. The heterotrophic nanoflagellate Halocafeteria, the type strain of which grows optimally at 150‰ salinity and 35°C, is a commonly encountered obligate halophile found in very hypersaline environments. These extreme environments can vary in their Mg:Ca ratios (i.e. weight ratios) and sulfate concentrations. To examine growth response of Halocafeteria to the different chemical compositions, densities of Halocafeteria seosinensis strain EHF34 were monitored in seven different ion composition media for 9 days at 1- to 2-day intervals (at 150‰ salinity and 35°C, with no prey limitation). Halocafeteria does not grow at Mg:Ca ratios of 35 and 100 and at high sulfate concentrations of 11.6 and 31.6 g l⁻¹. It grows well in 0.6 g l⁻¹ sulfate media at Mg:Ca ratios of 2, 10 or 35, but not 100. The present study demonstrates that the growth of the obligate halophile Halocafeteria can be affected by different ion compositions in hypersaline environments. Therefore, Halocafeteria may not be ubiquitous in hypersaline environments due to its ionic requirements.     

Isolation and characterisation of Thiobacillus halophilus sp. nov., a sulphur-oxidising autotrophic eubacterium from a Western Australian hypersaline lake

The isolation of a novel obligately chemolithotrophic, halophilic and extremely halotolerant Thiobacillus from a hypersaline lake is described. Attempts to demonstrate sulphur- and ferrous iron-oxidizing chemolithotrophs in neighbouring hypersaline lakes were unsuccessful. The organism isolated differs from any other Thiobacillus species previously described and is formally named as Thiobacillus halophilus. It possesses ribulose bisphosphate carboxylase and grows chemolithoautotrophically on thiosulphate, tetrathionate and sulphur, oxidising them to sulphate. Kinetic constants for oxidation of sulphide, thiosulphate, trithionate and tetrathionate are presented. The organism is obligately halophilic, growing best with 0.8–1.0 M NaCl, and tolerating up to 4 M NaCl. Optimum growth was obtained at about 30° C and pH 7.0–7.3. It contains ubiquinone Q-8 and its DNA contains 45 mol % G+C. Organisms of this type might contribute significantly to the autotrophic fixation of carbon dioxide in some hypersaline extreme environments of the kind described.     

<Emphasis Type="Italic">Marinimicrobium haloxylanilyticum</Emphasis> sp. nov., a new moderately halophilic,polysaccharide-degrading bacterium isolated from Great Salt Lake,Utah

A new moderately halophilic, strictly aerobic, Gram-negative bacterium, strain SX15^T, was isolated from hypersaline surface sediment of the southern arm of Great Salt Lake (Utah, USA). The strain grew on a number of carbohydrates and carbohydrate polymers such as xylan, starch, carboxymethyl cellulose and galactomannan. The strain grew at salinities ranging from 2 to 22% NaCl (w/v). Optimal growth occurred in the presence of 7–11% NaCl (w/v) at a temperature of 35°C and a pH of 6.7–8.2. Major whole-cell fatty acids were C16:0 (30.5%), C18:0 (14.8%), C18:1ω7c (13.1%) and C12:0 (7.8%). The G+C content of the DNA was 60 ± 0.5 mol%. By 16S rRNA gene sequence analysis, strain SX15^T was shown to be affiliated to members of the gammaproteobacterial genus Marinimicrobium with pair wise identity values of 92.9–94.6%. The pheno- and genotypic properties suggest that strain SX15^T represents a novel species of the genus Marinimicrobium for which the name Marinimicrobium haloxylanilyticum is proposed. The type strain is SX15^T (= DSM 23100^T = CCUG 59572^T).     

Identification of several intracellular carbohydrate-degrading activities from the halophilic archaeon Haloferax mediterranei

Three different amylolytic activities, designated AMY1, AMY2, and AMY3 were detected in the cytoplasm of the extreme halophilic archaeon Haloferax mediterranei grown in a starch containing medium. This organism had also been reported to excrete an α-amylase into the external medium in such conditions. The presence of these different enzymes which are also able to degrade starch may be related to the use of the available carbohydrates and maltodextrins, including the products obtained by the action of the extracellular amylase on starch that may be transported to the cytoplasm of the organism. The behavior of these intracellular hydrolytic enzymes on starch is reported here and compared with their extracellular counterpart. Two of these glycosidic activities (AMY1, AMY3) have also been purified and further characterized. As with other halophilic enzymes, they were salt dependent and displayed maximal activity at 3 M NaCl, and 50°C. The purification steps and molecular masses have also been reported. The other activity (AMY2) was also detected in extracts from cells grown in media with glycerol instead of starch and in a yeast extract medium. This enzyme was able to degrade starch yielding small oligosaccharides and displayed similar halophilic behavior with salt requirement in the range 1.5–3 M NaCl. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Methanethiol utilization and sulfur reduction by anaerobic halophilic saccharolytic bacteria

A number of sulfur compounds were tested as sulfur sources for the growth of three strains of anaerobic halophilic saccharolytic bacteria isolated from hypersaline water bodies of the eastern Crimea (USSR). Dithionite and sulfite at 1 mM concentration completely inhibited the growth of all strains. Methanethiol turned out to be the sole sulfur source for growth ofHalobacteroides strains in the defined medium with glucose and leucine. Methanethiol also stimulated growth of cultures in the complex medium with yeast extract. TheHaloincola saccharolytica Z-7787 appeared to be capable of methanethiol formation from methionine. All organisms studied were capable of heterotrophic sulfur reduction, producing up to 13 mM H₂S, but no evidence that they gain energy from the process has been obtained. The extremely halophilicHalobacteroides lacumaris may participate in sulfidogenesis at the high salinity (20–30% NaCl). The ecological position of haloanaerobes in halophilic community is discussed.     

Cold-active halophilic bacteria from the ice-sealed Lake Vida,Antarctica

Lake Vida is a large, permanently ice-covered lake in the Victoria Valley of the McMurdo Dry Valleys, Antarctica and is unique among Dry Valley lakes because it is ice-sealed, with an ice-cover of nearly 19 m. Enrichment cultures of melt-water from Lake Vida 15.9 m ice yielded five pure cultures of aerobic, heterotrophic bacteria. Of these, one strain grew at −8°C and the four others at −4°C. All isolates were either halotolerant or halophilic, with two strains capable of growth at 15% NaCl. Phylogenetic analysis revealed the Lake Vida isolates to be Gammaproteobacteria, related to species of Psychrobacter and Marinobacter. This is the first report of pure cultures of bacteria from Lake Vida, and the isolates displayed a phenotype consistent with life in a cold hypersaline environment.     

Phylogeny and ecophysiological features of prokaryotes isolated from temporary saline tidal pools

Although hypersaline environments have been extensively examined, only a limited number of microbial community studies have been performed in saline tide pools. We have studied a temporary salt-saturated tide pool and isolated prokaryotes from the water. Chlorinity measurements revealed that the tide pool brine could be characterized as one of the most hypersaline ecosystems on earth. Enumeration of microorganisms at different salinities showed that the tide pool was dominated by moderate halophiles. Based on 16S rRNA gene sequence analysis, the prokaryotic strains isolated were related to the bacterial genera Rhodovibrio, Halovibrio, Aquisalimonas, Bacillus and Staphylococcus and to the haloarchaeal species Haloferax alexandrinus. Four bacterial isolates were distantly related to their closest validly described species Aquisalimonas asiatica (96.5 % similarity), representing a novel phylogenetic linkage. Ecophysiological analysis also revealed distinct phenotypic profiles for the prokaryotic strains analyzed. The herbicide 2,4-dichlorophenoxyacetate could be effectively utilized by selected strains as the sole carbon source, but phenolic compounds could not be utilized by any of the halophilic isolates examined. None of the halophilic strains were able to grow without the presence of sea salt or seawater. Based on these results, we conclude that moderate halophilic bacteria rather than extremely halophilic archaea dominate in such a hypersaline environment.     

<Emphasis Type="Italic">Salinarchaeum laminariae</Emphasis> gen. nov., sp. nov.: a new member of the family <Emphasis Type="Italic">Halobacteriaceae</Emphasis> isolated from salted brown alga <Emphasis Type="Italic">Laminaria</Emphasis>

Halophilic archaeal strains R26^T and R22 were isolated from the brown alga Laminaria produced at Dalian, Liaoning Province, China. Cells from the two strains were pleomorphic rods and Gram negative, and colonies were red pigmented. Strains R26^T and R22 were able to grow at 20–50°C (optimum 37°C) in 1.4–5.1 M NaCl (optimum 3.1–4.3 M) at pH 5.5–9.5 (optimum pH 8.0–8.5) and neither strain required Mg²⁺ for growth. Cells lyse in distilled water and the minimum NaCl concentration required to prevent cell lysis was 8% (w/v) for strain R26^T and 12% (w/v) for strain R22. The major polar lipids of the two strains were phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester and minor phosphatidylglycerol sulfate; glycolipids were not detected. Phylogenetic analyses based on 16S rRNA genes and rpoB′ genes revealed that strains R26^T and R22 formed a distinct clade with the closest relative, Natronoarchaeum mannanilyticum. The DNA G+C content of strains R26^T and R22 was 65.8 and 66.4 mol%, respectively. The DNA–DNA hybridization value between strains R26^T and R22 was 89%. The phenotypic, chemotaxonomic and phylogenetic properties suggest that the strains R26^T and R22 represent a novel species in a new genus within the family Halobacteriaceae, for which the name Salinarchaeum laminariae gen. nov., sp. nov. is proposed. The type strain is R26^T (type strain R26^T = CGMCC 1.10590^T = JCM 17267^T, reference strain R22 = CGMCC 1.10589).     

Halorussus rarus gen. nov., sp. nov., a new member of the family Halobacteriaceae isolated from a marine solar saltern

Two halophilic archaeal strains TBN4^T and TBN5 were isolated from Taibei marine solar saltern in Jiangsu, China. Both strains showed light red-pigmented colonies and their cells were rod, motile and Gram-stain-negative. They were able to grow at 25–50°C (optimum 37°C), at 1.4–4.3 M NaCl (optimum 2.1 M NaCl), at 0–1.0 M MgCl₂ (optimum 0.005 M MgCl₂) and at pH 6.0–9.0 (optimum pH 7.0). Their cells lyse in distilled water and minimal NaCl concentration to prevent cell lysis is 8% (w/v). The major polar lipids of the two strains were PG (phosphatidylglycerol), PGP-Me (phosphatidylglycerol phosphate methyl ester), PGS (phosphatidylglycerol sulfate) and five glycolipids chromatographically identical to S-TGD-1 (sulfated galactosyl mannosyl glucosyl diether), S-DGD-1 (sulfated mannosyl glucosyl diether), TGD-1 (galactosyl mannosyl glucosyl diether), DGD-1 (mannosyl glucosyl diether) and DGD-2 (an unknown diglycosyl diether). Phylogenetic analysis revealed that TBN4^T and strain TBN5 formed a distinct clade with genus Haladaptatus (showing 90.0–90.9% 16S rRNA gene similarities). The DNA G + C content of strain TBN4^T and strain TBN5 are 66.1 and 65.4 mol%, respectively. The DNA–DNA hybridization value between strain TBN4^T and strain TBN5 was 94.3%. The phenotypic, chemotaxonomic and phylogenetic properties suggest that strain TBN4^T and strain TBN5 represent a novel species in a new genus within the family Halobacteriaceae, for which the name Halorussus rarus gen. nov., sp. nov. is proposed. The type strain is TBN4^T (=CGMCC 1.10122^T = JCM 16429^T).     

Production,optimization and purification of a novel extracellular protease from the moderately halophilic bacterium <Emphasis Type="Italic">Halobacillus karajensis</Emphasis>

The production of a protease was investigated under conditions of high salinity by the moderately halophilic bacterium Halobacillus karajensis strain MA-2 in a basal medium containing peptone, beef extract, maltose and NaCl when the culture reached the stationary growth phase. Effect of various temperatures, initial pH, salt and different nutrient sources on protease production revealed that the maximum secretion occurred at 34°C, pH 8.0–8.5, and in the presence of gelatin. Replacement of NaCl by various concentrations of sodium nitrate in the basal medium also increased the protease production. The secreted protease was purified 24-fold with 68% recovery by a simple approach including a combination of acetone precipitation and Q-Sepharose ion exchange chromatography. The enzyme revealed a monomeric structure with a relative molecular mass of 36 kDa by running on SDS-PAGE. Maximum caseinolytic activity of the enzyme was observed at 50°C, pH 9.0 and 0.5 M NaCl, although at higher salinities (up to 3 M) activity still remained. The maximum enzyme activity was obtained at a broad pH range of 8.0–10.0, with 55 and 50% activity remaining at pH 6 and 11, respectively. Moreover, the enzyme activity was strongly inhibited by phenylmethylsulfonyl fluoride (PMSF), Pefabloc SC and EDTA; indicating that it probably belongs to the subclass of serine metalloproteases. These findings suggest that the protease secreted by Halobacillus karajensis has a potential for biotechnological applications from its haloalkaline properties point of view.     

Phylogenetic analysis and screening of antimicrobial and cytotoxic activities of moderately halophilic bacteria isolated from the Weihai Solar Saltern (China)

A total of 45 moderately halophilic bacteria was isolated from sediment and saline water collected from the Weihai Solar Saltern (China). The phylogenetic position of all the isolated strains was determined by 16S rRNA sequencing. The halophilic strains were tested for their antimicrobial activity. Cytotoxicity assay was performed to determine which of the halophilic strains could inhibit proliferation of human hepatocellular carcinoma Bel 7402 cells. Our results showed that all of the isolated 45 strains displayed moderately halophilic characteristics. Phylogenetic analysis indicated that 17 of the isolated strains were related to the phylum Firmicutes and belonged to four genera, Bacillus, Halobacillus, Planococcus and Salinicoccus. The other strains identified as genus of Halomonas belonged to phylum γ-Proteobacteria. Most of the halophilic bacterial strains showed potent activities against Gram-positive bacteria, human pathogenic fungi and plant pathogenic fungi. In addition, the crude extracts from 14 halophilic bacterial strains showed cytotoxic activity against tumor cells Bel 7402, and five of them showed remarkable activities with IC₅₀ less than 40 μg ml⁻¹. Our results suggest that the moderately halophilic bacteria may be developed as promising sources for the discovery of novel bioactive substances.     

Purification and biological characterization of a halophilic thermostable protease from <Emphasis Type="Italic">Haloferax lucentensis</Emphasis> VKMM 007

An extremely halophilic archaeon Haloferax lucentensis VKMM 007, isolated from a solar saltern, was found to produce a protease. This extracellular enzyme consisted of a single polypeptide chain of 57.8 kDa as determined by SDS–PAGE and was purified by a combination of ultrafiltration, bacitracin–Sepharose affinity chromatography and Sephadex G-100 gel filtration. The purified protein was stable in a wide range of temperatures (20–70°C), NaCl concentrations (0.85–5.13 M) and pH (5.0–9.0) with maximal activity observed at 60°C, 4.3 M NaCl and pH 8.0. Proteolytic activity was enhanced by Ca²⁺, K⁺, Mg²⁺, Na⁺, and Fe²⁺ ions and the protein was classified as a trypsin-like serine protease. Further assays indicated highest degree of specificity when hemoglobin was used as an enzyme substrate. Most importantly, the proteolytic activity remained stable or only marginally inhibited in the presence of various polar and non-polar solvents, surfactants and reducing agents thus emphasizing the biotechnological potential of this novel halophilic protease.     

Polymer production by two newly isolated extremely halophilic archaea: application of a novel corrosion-resistant bioreactor

A novel corrosion-resistant bioreactor composed of polyetherether ketone (PEEK), tech glass and silicium nitrite ceramics was constructed and applied for the cultivation of two newly isolated, extremely halophilic archaea producing poly(γ-glutamic acid) (PGA), or poly(β-hydroxy butyric acid) (PHB), respectively. These bacteria were isolated from hypersaline soil close to Aswan (Egypt). The isolate strain 40, which is related to the genus Natrialba, produced large amounts of PGA when cultivated on solid medium. Culture conditions were optimised applying the corrosion-resistant bioreactor. PGA production was dependent on NaCl concentration and occurred about at 20% (w/v) NaCl in the medium. A maximum cell density of about 1.6 g cell dry matter/l was obtained when the bioreactor was stirred and aerated in a batch fermentation process using proteose-peptone medium. The supernatant was monitored with respect to PGA formation, and after 90 h a maximum of 470 mg/l culture volume was detected by HPLC analysis. Culture conditions were optimized for the isolate 56, which accumulated PHB as intracellular granules. Batch fermentations in the stirred and aerated bioreactor applying acetate and n-butyric acid as carbon sources led to cell density of 2.28 g cell dry matter/l and a maximum PHB accumulation contributing to about 53% of cellular dry weight. About 4.6 g PHB were isolated from 10.6 g dried cells of strain 56, which exhibited a weight average molar mass of 2.3 × 10⁵ g mol⁻¹ and a polydispersity of about 1.4. Received: 3 December 1999 / Received revision: 22 February 2000 / Accepted: 25 February 2000     

Optimization of culture conditions for the production of halothermophilic protease from halophilic bacterium <Emphasis Type="Italic">Chromohalobacter</Emphasis> sp. TVSP101

An extremely halophilic Chromohalobacter sp. TVSP101 was isolated from solar salterns and screened for the production of extracellular halothermophilic protease. Identification of the bacterium was done based upon biochemical tests and the 16S rRNA sequence. The partially purified enzyme displayed maximum activity at pH 8 and required 4.5 M of NaCl for optimum proteolytic activity. In addition, this enzyme was thermophilic and active in broad range of temperature 60–80°C with 80°C as optimum. The Chromohalobacter sp. required 4 M NaCl for its optimum growth and protease secretion and no growth was observed below 1 M of NaCl. The initial pH of the medium for growth and enzyme production was in the range 7.0–8.0 with optimum at pH 7.2. Various cations at 1 mM concentration in the growth medium had no significant effect in enhancing the growth and enzyme production but 0.5 M MgCl₂ concentration enhanced enzyme production. Casein or skim milk powder 1% (w/v) along with 1% peptone proved to be the best nitrogen sources for maximum biomass and enzyme production. The carbon sources glucose and glycerol repressed the protease secretion. Immobilization of whole cells in absence of NaCl proved to be useful for continuous production of halophilic protease.     

Isolation and characterization of halophilic bacteria from Urmia Lake in Iran

Urmia Lake is one of the most permanent hypersaline lakes in the world which is threatened by hypersalinity and serious dryness. In spite of its importance no paper has been published regarding bacterial community of this lake. Accordingly, the present study aimed to investigate the halophilic bacteria in the aforementioned lake. In so doing, thirty seven strains were isolated on six different culture media. The isolated strains were characterized using phenotypic and genotypic methods. Growth of the strains occurred at 25–35°C, pH 6–9 and 7 to 20% (w/v) NaCl indicating that most of the isolates were moderately halophiles. Catalase, oxidase and urease activities were found to be positive for the majority of the isolates. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolated bacteria belonged to two major taxa: Gammaproteobacteria (92%, including Salicola [46%], Pseudomonas [13.5%], Marinobacter [11%], Idiomarina [11%], and Halomonas [8%]) and Firmicutes (8%, including Bacillus [5%] and Halobacillus [3%]). In addition, a novel bacterium whose 16S rRNA gene sequence showed almost 98% sequence identity with the taxonomically troubled DSM 3050^T, Halovibrio denitrificans HGD 3^T and Halospina denitriflcans HGD 1–3, each, was isolated. 16S rRNA gene similarity levels along with phenotypic characteristics suggest that some of the isolated strains could be regarded as potential type strain for novel species, on which further studies are recommended.