[Show abstract][Hide abstract] ABSTRACT: Three halophilic isolates, strains Halo-G*T, AUS-1 and Naxos II, were compared. Halo-G* was isolated from an evaporitic salt crystal from Baja California, Mexico, whereas AUS-1 and Naxos II were isolated from salt pools in Western Australia and the Greek island of Naxos, respectively. Halo-G*T had been exposed previously to conditions of outer space and survived 2 weeks on the Biopan facility. Chemotaxonomic and molecular comparisons suggested high similarity between the three strains. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the strains clustered with Halorubrum species, showing sequence similarities of 99.2-97.1%. The DNA-DNA hybridization values of strain Halo-G*T and strains AUS-1 and Naxos II are 73 and 75%, respectively, indicating that they constitute a single species. The DNA relatedness between strain Halo-G*T and the type strains of 13 closely related species of the genus Halorubrum ranged from 39 to 2%, suggesting that the three isolates constitute a different genospecies. The G+C content of the DNA of the three strains was 65.5-66.5 mol%. All three strains contained C20C20 derivatives of diethers of phosphatidylglycerol, phosphatidylglyceromethylphosphate and phosphatidylglycerolsulfate, together with a sulfated glycolipid. On the basis of these results, a novel species that includes the three strains is proposed, with the name Halorubrum chaoviator sp. nov. The type strain is strain Halo-G*T (=DSM 19316T=NCIMB 14426T=ATCC BAA-1602T).
Full-text · Article · Jul 2009 · International Journal of Systematic and Evolutionary Microbiology
[Show abstract][Hide abstract] ABSTRACT: The aerobic degradation of light fuel oil in sandy and loamy soils by an environmental bacterial consortium was investigated. Soils were spiked with 1 or 0.1% of oil per dry weight of soil. Acetone extracts of dried soils were analyzed by GC and the overall degradation was calculated by comparison with hydrocarbon recovery from uninoculated soils. In sandy soils, the sum of alkanes n-C(12) to n-C(23) was degraded to about 45% within 6 days at 20 degrees C and to 27-31% within 28 days, provided that moisture and nutrients were replenished. Degradation in loamy soil was about 12% lower. The distribution of recovered alkanes suggested a preferential degradation of shorter chain molecules (n-C(12) to n-C(16)) by the bacterial consortium. Partial 16S rDNA sequences indicated the presence of strains of Pseudomonas aeruginosa, Pseudomonas citronellolis, and Stenotrophomonas maltophilia. Toxicity tests using commercial standard procedures showed a moderate inhibition of bacterial activity. The study showed the applicability of a natural microbial community for the degradation of oil spills into soils at ambient temperatures.
No preview · Article · Dec 2007 · Biotechnology Journal
[Show abstract][Hide abstract] ABSTRACT: Pseudomonas beijerinckii (type strain DSM 7218(T)=ATCC 19372(T)=NCIMB 9041(T)) was isolated from salted beans and was first described by Hof in 1935. 16S rRNA gene sequence comparisons demonstrated its close relatedness (>97-99 %) to species of the genus Chromohalobacter. A recent isolate from salted herrings originating from the Baltic Sea, strain 3b, also clustered phylogenetically within this genus. Phenotypic features, substrate utilization, fatty acid profile, quinone and polar lipid composition and whole-cell protein patterns supported the similarity of strain 3b to P. beijerinckii DSM 7218(T) and confirmed its relatedness to members of the genus Chromohalobacter. The G+C content of the DNA from strain 3b and P. beijerinckii DSM 7218(T) was 60.4 and 60.7 mol%, respectively. DNA-DNA hybridization data showed that the two strains represent the same species, but are separated from Chromohalobacter canadensis, the closest species from a phylogenetic point of view. Therefore, the reclassification of Pseudomonas beijerinckii as Chromohalobacter beijerinckii comb. nov. (type strain DSM 7218(T)=ATCC 19372(T)=NCIMB 9041(T)) is proposed. The species description has been emended considering the new data on both the type strain and strain 3b.
Full-text · Article · Aug 2006 · International Journal of Systematic and Evolutionary Microbiology
[Show abstract][Hide abstract] ABSTRACT: Two rod-shaped haloarchaeal strains, A1 and A2, were isolated from a bore core from a salt mine in Austria. The deposition of the salt is thought to have occurred during the Permian period (225-280 million years ago). The 16S rDNA sequences of the strains were 97.1% similar to that of the type species of the genus Halobacterium, which was also determined in this work. Polar lipids consisted of C20-C20 derivatives of phosphatidylglycerol, methylated phosphatidylglycerol phosphate, phosphatidylglycerol sulfate, triglycosyl diether and sulfated tetraglycosyl diether. Optimal salinity for growth was 15-17.5% NaCl; Mg++ was tolerated up to a concentration of 1 M. The DNA-DNA reassociation value of strain A1T was 25% with H. salinarum DSM 3754T and 41% with Halobacterium sp. NRC-1, respectively. Based on these results and other properties, e.g. whole cell protein patterns, menaquinone content and restriction patterns of DNA, strains A1 and A2 are members of a single species, for which we propose the name H. noricense. The type strain is A1 (DSM 15987T, ATCC BAA-852T, NCIMB 13967T). Since we present evidence that Halobacterium sp. NRC-1 is a member of H. salinarum, an emended description of H. salinarum is provided.
[Show abstract][Hide abstract] ABSTRACT: Several viable extremely halophilic archaebacteria (haloarchaea) were
isolated previously from Permo-Triassic salt sediments. We described two
halococcal strains from rock salt of the salt mine in Bad Ischl,
Austria. Recently, we isolated a rod-shaped haloarchaeal species from a
salt bore core. In addition, sequencing of 16S rRNA genes from dissolved
rock salt indicated the presence of a vast diversity of as yet
uncultured haloarchaea. The results are compatible with the notion that
these haloarchaea might be the remnants of microbial communities which
populated the ancient hypersaline seas. How they survived in the dry
salt evaporites is not known. Extremely dry conditions are
characteristic for the Martian surface and perhaps also subsurface;
since extraterrestrial halite has been discovered in meteorites from
Mars and other celestial bodies, it appears feasible to explore the
response of haloarchaea to simulated Martian environments. Preliminary
results suggested survival rates of haloarchaea, following freezing,
freeze-drying or embedding in salt crystals, respectively, in the range
of 0.1 to 10%. Thus, low water activities and low temperatures appear to
be tolerated well by haloarchaea.
[Show abstract][Hide abstract] ABSTRACT: A coccoid- to ovoid-shaped, Gram-positive bacterial strain, designated C3H-21(T), was isolated from the liver of the laboratory mouse strain C3H/He and characterized by a polyphasic approach. The peptidoglycan type was variation A4gamma with meso-diaminopimelic acid as the diagnostic cell-wall diamino acid and an interpeptide bridge of D-asp-D-Glu. The isolate contained menaquinone MK-7 (88 %) as the major component of the quinone system and minor amounts of menaquinone MK-8 (9 %) and menaquinone MK-6 (3 %). The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylglycerol, unidentified glycolipids and unidentified phospholipids. The fatty acid profile contained predominantly anteiso-C(15 : 0) and significant amounts of iso-C(16 : 0), iso-C(14 : 0,) anteiso-C(17 : 0) and C(19 : 0). The polyamine pattern consisted of spermine and spermidine as the major compounds. Genomic fingerprints clearly distinguished strain C3H-21(T) from other Brachybacterium species. The isolate shared the highest 16S rDNA sequence similarities with members of the genus Brachybacterium, in particular Brachybacterium sacelli LMG 20345(T), Brachybacterium nesterenkovii DSM 9573(T), Brachybacterium rhamnosum LMG 19848(T), Brachybacterium alimentarium CNRZ 925(T) and Brachybacterium fresconis LMG 20336(T) (97.8-97.2 %). The results of biochemical/physiological characterization, chemotaxonomic characteristics and REP-PCR-generated fingerprints demonstrated that the isolate represents a novel species of the genus Brachybacterium, for which the name Brachybacterium muris (type strain C3H-21(T)=DSM 15460(T)=CCM 7047(T)) [corrected] is proposed.
Full-text · Article · Dec 2003 · International Journal of Systematic and Evolutionary Microbiology
[Show abstract][Hide abstract] ABSTRACT: Haloarchaeal strains have been isolated repeatedly from ancient rock salt. Halococcus salifodinae BIp and Halococcus dombrowskii H4 were isolated from different rock salt samples obtained from the salt mine near Bad Ischl, Austria. Based on the geological age of the horizon the salt has been deposited during the Permian period (225 to 280 million years ago). The medium which is commonly used for the cultivation of neutrophilic haloarchaea is called M2 medium. Especially halococci grow very slowly and yield only low densities when cultured in this medium. It was presumed that these microorganisms might grow better using a culture medium with the chemical components of their natural environment. Addition of dissolved and sterile-filtered rock salt to standard M2 medium proved indeed successful. The modified medium (M2S) showed a growth stimulating effect on all tested halococcal strains. Cultures of Halococcus morrhuae DSM 1307, Halococcus salifodinae DSM 8989, Halococcus saccharolyticus DSM 5750 and Halococcus dombrowskii DSM 14522 grown in M2S medium had lower doubling times and reached a greater turbidity compared to cultures grown in M2 medium. In addition representatives of the genera Haloferax, Haloarcula, Halorubrum and Halobacterium, including Halobacterium sp. NRC-1 whose complete genome sequence is known, were cultivated on these media. Halobacterium sp. NRC-1 and Haloarcula hispanica DSM 4426 showed also an increase in growth whereas Haloferax volcanii DSM 3757 and Halorubrum saccharovorum DSM 1137 grew better when cultivated on standard M2 medium. The stimulating effect of the heat stable factors in rock salt on haloarchaea appears correlated with certain proteins that are altered or may be exclusively produced when cells were grown in presence of rock salt as well as changes in morphology and ultrastructure of cells. The establishment of optimized media for haloarchaea is important for further studies concerning insights how haloarchaea are able to survive over long periods embedded in salt crystals.
[Show abstract][Hide abstract] ABSTRACT: Extremely halophilic archaebacteria (halobacteria) are found today in hypersaline surface waters, such as the brines in solar salterns, or the Dead Sea. However, from Alpine rock salt of Permo-Triassic age several species of halobacteria were isolated during the last years (1, 2). Halobacteria are not known to produce spores or dormant forms; thus it remains enigmatic how they survived in the salt sediments. Extraterrestrial halite has been detected in meteorites from Mars and from the asteroids; in addition, the Jovian moon Europa is thought to contain a salty ocean. Therefore halobacteria would be useful model organisms when considering the search for extraterrestrial life. We are developing experimental protocols to evaluate the effects of physico-chemical stress factors on halobacteria, in particular present-day Martian conditions. But the effect of higher temperatures is also of interest, since Mars may have been warmer in the past, and the Alpine salt sediments are known to have experienced local temperature peaks. Cells of Halococcus dombrowskii (2) and, for comparison, of Halobacterium sp. NRC-1 were grown in complex medium, containing up to 4 M NaCl (2). Aliquots of cultures were kept at minus 70oC for several days, or freeze-dried in a lyophilizer, or incubated at temperatures of 50 to 80oC for 24 hours, respectively. In addition, exposure experiments of halobacterial cells in a liquid nitrogen cooled Martian simulation chamber were begun. Survival of cells was evaluated by determining colony-forming units and by examination of cellular morphology by fluorescence microscopy, following staining with the LIVE-DEAD kit. Results indicated that the LIVE-DEAD kit can be successfully used in the presence of 4 M NaCl, although it was developed for tests at low ionic strength. Data will be presented which show that Hc. dombrowskii survived deep freezing, temperatures of up to 80 oC and Martian atmospheric conditions generally better than Halobacterium sp. NRC-1, suggesting possibly a protective action of its cell envelope. (1) Stan-Lotter H, McGenity TJ, Legat A, Denner EBM, Glaser K, Stetter KO, Wanner G (1999) Microbiology 145, 3565-3574. (2) Stan-Lotter H, Pfaffenhuemer M, Legat A, Busse HJ, Radax C, Gruber C (2002) Int. J. System. Evol. Microbiol. 52, 1807-1814.
[Show abstract][Hide abstract] ABSTRACT: Several viable halobacteria were isolated from Alpine rock salt of Permo-Triassic age and described as novel species [see 1]. They have apparently survived in the salt sediments over extremely long periods of time. Halobacteria could therefore be suitable model organisms for exploring the possibility of long-term survival of microorganisms on other planets. In addition, the discovery of extraterrestrial halite makes it plausible to consider a specific search for halophiles, perhaps in the planned sample return missions to Mars. We are developing experimental procedures to test the responses of halobacteria to Martian conditions.
[Show abstract][Hide abstract] ABSTRACT: Halococcus dombrowskii was described as a novel haloarchaeal species isolated from Alpine rock salt which is believed to have been deposited about 250 million years ago . During its characterization, a new medium (M2S) was created by addition of dissolved and sterile-filtered rock salt to standard halophile medium (M2), which showed a pronounced growth stimulating effect on halococcal strains. Cultures grown in M2S had lower doubling times and reached a greater turbidity compared to cultures grown in M2. In addition, changes in cell morphology and significant differences in whole cell protein patterns were observed. It was examined whether this stimulating effect is reproducible within other halobacterial genera. M2S medium appears to approach the composition of the natural habitat of halophilic archaea and is therefore of interest for astrobiological research.
[Show abstract][Hide abstract] ABSTRACT: Several extremely halophilic coccoid archaeal strains were isolated from pieces of dry rock salt that were obtained three days after blasting operations in an Austrian salt mine. The deposition of the salt is thought to have occurred during the Permian period (225-280 million years ago). On the basis of their polar-lipid composition, 16S rRNA gene sequences, cell shape and growth characteristics, the isolates were assigned to the genus Halococcus. The DNA-DNA reassociation values of one isolate, strain H4T, were 35 and 38% with Halococcus salifodinae and Halococcus saccharolyticus, respectively, and 65.8-67.8% with Halococcus morrhuae. The polar lipids of strain H4T were C20-C25 derivatives of phosphatidylglycerol and phosphatidylglycerol phosphate. Whole-cell protein patterns, menaquinone content, enzyme composition, arrangements of cells, usage of carbon and energy sources, and antibiotic susceptibility were sufficiently different between strain H4T and H. morrhuae to warrant designation of strain H4T as a new species within the genus Halococcus. It is proposed that the isolate be named Halococcus dombrowskii, and the type strain is H4T (= DSM 14522T = NCIMB 13803T = ATCC BAA-364T).
Full-text · Article · Oct 2002 · International Journal of Systematic and Evolutionary Microbiology
[Show abstract][Hide abstract] ABSTRACT: Several viable halophilic archaebacteria were isolated previously from rock salt of Permo-Triassic age in an Austrian salt mine; one of these strains was the first to be recognized as a novel species from subterranean halite and was designated Halococcus
salifodinae. The halophilic microorganisms have apparently survived in the salt sediments over extremely long periods of time. Halobacteria could therefore be suitable model organisms for exploring the possibility of long-term survival of microbes on other planets, in particular, since extraterrestrial halite has been detected in meteorites and is assumed to be present in the subsurface ocean on Europa. Our efforts are directed at the identification of the microbial content of ancient rock salt and the development of procedures for the investigation of the halobacterial response to extreme environmental conditions. Using modified culture media, further halophilic strains were isolated from freshly blasted rock salt and bore cores; in addition, growth of several haloarchaea was substantially improved. Molecular methods indicated the presence of at least 12 different 16S rRNA gene species in a sample of Alpine rock salt, but these strains have not been cultured yet. The exploration of Mars is a target of space missions in the 21st century; therefore, testing the survival of haloarchaea under conditions comparable to present-day Mars, using a simulation chamber, was begun. Preliminary results with Halococcus and Halobacterium species suggested at least tenfold higher survival rates when cells were kept in liquid brines than under dry conditions; staining of cells with the LIVE–DEAD kit, which discriminates between damaged and intact membranes, corroborated these data.
[Show abstract][Hide abstract] ABSTRACT: Nine yellow-pigmented, spherical bacterial strains isolated from a medieval wall painting (strain D7), from indoor air (strains 3, 6, 7, 13C2, 38, 83 and 118) and from an activated-sludge plant (strain Ballarat) were classified by a polyphasic approach. Analyses of the 16S rRNA gene sequences of three representatives (strains D7, 118 and Ballarat) indicated that they all belong to the genus Micrococcus. The three isolates shared the highest sequence similarities with Micrococcus luteus DSM 20030T (97.9-98%), Micrococcus antarcticus AS 1.2372T (97.9-98.3%) and Micrococcus lylae DSM 20315T (97.5-97.9%). DNA-DNA reassociation studies clearly demonstrated that all nine isolates belong to the species M. luteus. However, neither their chemotaxonomic features nor their physiological and biochemical properties were consistent with those of M. luteus DSM 20030T. In contrast to M. luteus DSM 20030T, all isolates investigated possessed MK-8(H2) as the major respiratory quinone, and strain Ballarat had an A4alpha peptidoglycan type. On the basis of analyses of their Fourier transform-infrared spectroscopy spectra, isolates D7, 3, 6, 7, 13C2, 38, 83 and 118 could be grouped into a single cluster separate from M. luteus DSM 20030T, strain Ballarat and M. lylae DSM 20315T. In addition, all these isolates could be distinguished from M. luteus DSM 20030T by their ability to assimilate D-maltose, D-trehalose, DL-3-hydroxybutyrate, DL-lactate, pyruvate and L-histidine and to hydrolyse casein. Strains D7, 3, 6, 7, 13C2, 38, 83 and 118 differed from both M. luteus DSM 20030T and strain Ballarat by their ability to assimilate acetate, L-phenylalanine, L-serine and phenylacetate. Furthermore, REP-PCR fingerprinting yielded one common band for these strains, whereas this band was not observed for M. luteus DSM 20030T, strain Ballarat or M. lylae DSM 20315T. On the basis of these data, the species M. luteus can be divided into three biovars that are distinguished by several chemotaxonomic and biochemical traits: biovar I, represented by M. luteus DSM 20030T; biovar II, represented by strains D7 (= DSM 14234 = CCM 4959), 3, 6, 7, 13C2, 38, 83 and 118; and biovar III, represented by strain Ballarat (= DSM 14235 = CCM 4960). On the basis of the results generated in this study, emended descriptions of the genus Micrococcus and the species M. luteus and M. lylae are given.
Full-text · Article · Apr 2002 · International Journal of Systematic and Evolutionary Microbiology
[Show abstract][Hide abstract] ABSTRACT: Prokaryotic diversity in Alpine salt sediments was investigated by polymerase chain reaction (PCR) amplification of 16S rRNA genes, sequencing of cloned products, and comparisons with culturable strains. DNA was extracted from the residue following filtration of dissolved Permo-Triassic rock salt. Fifty-four haloarchaeal sequences were obtained, which could be grouped into at least five distinct clusters. Similarity values of three clusters to known 16S rRNA genes were less than 90%-95%, suggesting the presence of uncultured novel taxa; two clusters were 98% and 99% similar to isolates from Permo-Triassic or Miocene salt from England and Poland, and to Halobacterium salinarum, respectively. Some rock salt samples, including drilling cores, yielded no amplifiable DNA and no cells or only a few culturable cells. This result suggested a variable distribution of haloarchaea within different strata, probably consistent with the known geologic heterogeneity of Alpine salt deposits. We recently reported identical culturable Halococcus salifodinae strains in Permo-Triassic salt sediments from England, Germany, and Austria; together with the data presented here, those results suggest one plausible scenario to be an ancient continuous hypersaline ocean (Zechstein sea) populated by haloarchaea, whose descendants are found today in the salt sediments. The novelty of the sequences also suggested avoidance of haloarchaeal contaminants during our isolation of strains, preparation of DNA, and PCR reactions.