The Structure of the Lipid A from the Halophilic Bacterium Spiribacter salinus M19-40T.

The study of the adaptation mechanisms that allow microorganisms to live and proliferate in an extreme habitat is a growing research field. Directly exposed to the external environment, lipopolysaccharides (LPS) from Gram-negative bacteria are of great appeal as they can present particular structural features that may aid the understanding of the adaptation processes. Moreover, through being involved in modulating the mammalian immune system response in a structure-dependent fashion, the elucidation of the LPS structure can also be seen as a fundamental step from a biomedical point of view. In this paper, the lipid A structure of the LPS from Spiribacter salinus M19-40T, a halophilic gamma-proteobacteria, was characterized through chemical analyses and matrix-assisted laser desorption ionization (MALDI) mass spectrometry. This revealed a mixture of mono- and bisphosphorylated penta- to tri-acylated species with the uncommon 2 + 3 symmetry and bearing an unusual 3-oxotetradecaonic acid.

From metagenomics to pure culture: isolation and characterization of the moderately halophilic bacterium Spiribacter salinus gen. nov., sp. nov.

Recent metagenomic studies on saltern ponds with intermediate salinities have determined that their microbial communities are dominated by both Euryarchaeota and halophilic bacteria, with a gammaproteobacterium closely related to the genera Alkalilimnicola and Arhodomonas being one of the most predominant microorganisms, making up to 15% of the total prokaryotic population. Here we used several strategies and culture media in order to isolate this organism in pure culture. We report the isolation and taxonomic characterization of this new, never before cultured microorganism, designated M19-40(T), isolated from a saltern located in Isla Cristina, Spain, using a medium with a mixture of 15% salts, yeast extract, and pyruvic acid as the carbon source. Morphologically small curved cells (young cultures) with a tendency to form long spiral cells in older cultures were observed in pure cultures. The organism is a Gram-negative, nonmotile bacterium that is strictly aerobic, non-endospore forming, heterotrophic, and moderately halophilic, and it is able to grow at 10 to 25% (wt/vol) NaCl, with optimal growth occurring at 15% (wt/vol) NaCl. Phylogenetic analysis based on 16S rRNA gene sequence comparison showed that strain M19-40(T) has a low similarity with other previously described bacteria and shows the closest phylogenetic similarity with species of the genera Alkalilimnicola (94.9 to 94.5%), Alkalispirillum (94.3%), and Arhodomonas (93.9%) within the family Ectothiorhodospiraceae. The phenotypic, genotypic, and chemotaxonomic features of this new bacterium showed that it constitutes a new genus and species, for which the name Spiribacter salinus gen. nov., sp. nov., is proposed, with strain M19-40(T) (= CECT 8282(T) = IBRC-M 10768(T) = LMG 27464(T)) being the type strain.

Maricoccus atlantica gen. nov. sp. nov., isolated from deep sea sediment of the Atlantic Ocean.

A taxonomic study was carried out on strain 22II-S10r2(T), which was isolated from the deep sea sediment of the Atlantic Ocean using oil-degrading enrichment. The bacterium was Gram-negative, oxidase positive and catalase negative, spherical in shape, and motile by polar flagella. Growth was observed at salinities of 0.5-7 % and at temperatures of 10-41 °C. The isolate was capable of aesculin hydrolysis, but unable to reduce nitrate to nitrite or degrade Tween 80 or gelatine. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain 22II-S10r2(T) belonged to the family Ectothiorhodospiraceae, with highest sequence similarity to Thioalkalivibrio sulfidiphilus HL-EbGR7(T) (90.9 % similarity). The principal fatty acids were Sum In Feature 8 (C18:1 ω7c/ω6c (29.9 %), C18:1 ω9c (13.5 %), C16:1 ω5c (12.3 %), C12:03OH (6.8 %), C18:1 ω5c (5.7 %) and C16:0 (5.3 %). The G+C content of the chromosomal DNA was 60.7 mol%. The respiratory quinone was determined to be Q-7 (25 %) and Q-8 (75 %). Phosphatidylethanolamine, phosphatidylglycerol, aminophospholipid, glycolipid, three phospholipids and lipid were present. The strain was aerobic, non-phototrophic and non-chemolithoautotrophic. The combined genotypic and phenotypic data show that strain 22II-S10r2(T) represents a novel species within a novel genus, for which the name Maricoccus atlantica gen. nov. sp. nov. is proposed, with the type strain 22II-S10r2(T) (=CGMCC NO.1.12317(T) = LMG 27155(T) = MCCC 1A09384(T)).

Evaluation of the electrical potential on the membrane of the extremely alkaliphilic bacterium Thioalkalivibrio.

The electrical potential on the membrane was measured in cells of strains AL2 and ALJ15 of the extremely alkaliphilic bacterium Thioalkalivibrio versutus using the penetrating cation tetraphenylphosphonium (TPP(+)) and a TPP(+)-selective electrode. The potentials were -228 ± 5 and -224 ± 5 mV, respectively, i.e. higher than in most alkaliphilic bacteria. Membrane potential in the cells was estimated by measuring the inner cell volume by two independent methods: (1) estimation of total cell volume by light microscopy and (2) estimation of the inner aqueous volume of the cells with allowance for the distribution difference of tritium labeled water penetrating through the membranes and a nonpenetrating colored protein. The inner cell volume was 2.4 ± 0.2 and 2.2 ± 0.1 µl/mg of cell protein by the two methods, respectively. Computer computation was used as an alternative to manual calculation to count the number of cells for estimation of total cell volume.

Electroautotrophy of Thioalkalivibrio nitratireducens.

The electrochemical behavior and electro-autotrophy of a halo-alkaliphilic chemo-autotrophic Ectothiorhodospiraceae isolated from a soda lake, Thioalkalivibrio nitratireducens, were investigated using electrochemical methods and confocal fluorescence microscopy. The electrocatalysis of oxygen reduction was observed at -0.25 V/Ag/AgCl (-0.055 V/ SHE) with bioelectrodes polarized at -0.3 V/Ag/AgCl (-0.105 V/SHE), displaying a maximum catalytic current density of -620 mA m-2 (voltammetry). No catalytic signal toward nitrate reduction was observed under anaerobia. The microscopic observation of the polarized electrodes compared to non-polarized ones, however, revealed a bacterial proliferation both under aerobic and anaerobic conditions, demonstrating the ability of Tv. nitratireducens to grow with the polarized electrode as sole electron source.

Thiohalospira halophila gen. nov., sp. nov. and Thiohalospira alkaliphila sp. nov., novel obligately chemolithoautotrophic, halophilic, sulfur-oxidizing gammaproteobacteria from hypersaline habitats.

A previously unknown ecotype of obligately chemolithoautotrophic, sulfur-oxidizing bacteria was discovered in sediments of various inland hypersaline lakes and a solar saltern. The salt requirements for these bacteria were similar to those of haloarchaea, representing the first example of extreme halophiles occurring among the chemolithoautotrophs. They were enriched and isolated at 4 M NaCl under aerobic conditions with thiosulfate or tetrathionate as the electron donor or under micro-oxic conditions with sulfide. In total, 20 strains were obtained from hypersaline inland lakes in central Asia, central Russia and Crimea and a sea saltern of the Adriatic Sea. The isolates were thin, motile spirilla, some of which possessed a yellow, membrane-bound pigment. They were obligately aerobic, chemolithoautotrophic, sulfur-oxidizing bacteria that used thiosulfate, sulfide, sulfur and tetrathionate as electron donors. The characteristic feature of the group was the production of large amounts of tetrathionate as an intermediate during the oxidation of thiosulfate to sulfate. All but one of the strains grew within the pH range 6.5-8.2 (optimally at pH 7.3-7.8) and at NaCl concentrations from 2.0 to 5 M (optimally at 3.0 M). A single strain, designated ALgr 6sp(T), obtained (by enrichment) from the hypersaline alkaline lakes of the Wadi Natrun valley, was found to be moderately halophilic and facultatively alkaliphilic (capable of growth at pH 10). The predominant cellular fatty acids were quite unusual, with 10-methyl C(16 : 0) and C(16 : 0) predominating. Cells grown at 4 M NaCl accumulated extremely high concentrations of glycine betaine as a compatible solute. The 20 neutrophilic isolates contained three genospecies (on the basis of DNA-DNA relatedness data) but could not be discriminated phenotypically. On the basis of the phenotypic and genotypic analyses, the isolates constitute a novel genus and species, for which the name Thiohalospira halophila gen. nov., sp. nov. is proposed. The type strain of Thiohalospira halophila is HL 3(T) (=DSM 15071(T)=UNIQEM U219(T)). The haloalkaliphilic strain ALgr 6sp(T) represents a second species of the new genus, for which the name Thiohalospira alkaliphila sp. nov. is proposed. The type strain of Thiohalospira alkaliphila is ALgr 6sp(T) (=DSM 17116(T)=UNIQEM U372(T)).