Halo(natrono)archaea from hypersaline lakes can utilize sulfoxides other than DMSO as electron acceptors for anaerobic respiration.

Dimethylsulfoxide (DMSO) has long been known to support anaerobic respiration in a few species of basically aerobic extremely halophilic euryarchaea living in hypersaline lakes. Recently, it has also been shown to be utilized as an additional electron acceptor in basically anaerobic sulfur-reducing haloarchaea. Here we investigated whether haloarchaea would be capable of anaerobic respiration with other two sulfoxides, methionine sulfoxide (MSO) and tetramethylene sulfoxide (TMSO). For this, anaerobic enrichment cultures were inoculated with sediments from hypersaline salt and soda lakes in southwestern Siberia and southern Russia. Positive enrichments were obtained for both MSO and TMSO with yeast extract but not with formate or acetate as the electron donor. Two pure cultures obtained from salt lakes, either with MSO or TMSO, were obligate anaerobes closely related to sulfur-reducing Halanaeroarchaeum sulfurireducens, although the type strain of this genus was unable to utilize any sulfoxides. Two pure cultures isolated from soda lakes were facultatively anaerobic alkaliphilic haloarchaea using O2, sulfur and sulfoxides as the electron acceptors. One isolate was identical to the previously described sulfur-reducing Natrarchaeobaculum sulfurireducens, while another one, enriched at lower alkalinity, is forming a new species in the genus Halobiforma. Since all isolates enriched with either MSO or TMSO were able to respire all three sulfoxides including DMSO and the corresponding activities were cross-induced, it suggest that a single enzyme of the DMSO-reductase family with a broad substrate specificity is responsible for various sulfoxide-dependent respiration in haloarchaea.

Syntrophic associations from hypersaline soda lakes converting organic acids and alcohols to methane at extremely haloalkaline conditions.

Until now anaerobic oxidation of VFA at high salt-pH has been demonstrated only at sulfate-reducing conditions. Here, we present results of a microbiological investigation of anaerobic conversion of organic acids and alcohols at methanogenic conditions by syntrophic associations enriched from hypersaline soda lakes in Central Asia. Sediment incubation experiments showed active, albeit very slow, methane formation from acetate, propionate, butyrate and C2 C4 alcohols at pH 10 and various levels of salinity. Enrichments of syntrophic associations using hydrogenotrophic members of the genus Methanocalculus from soda lakes as partners resulted in several highly enriched cultures converting acetate, propionate, butyrate, benzoate and EtOH to methane. Most syntrophs belonged to Firmicutes, while the propionate-oxidizer formed a novel lineage within the family Syntrophobacteraceae in the Deltaproteobacteria. The acetate-oxidizing syntroph was identified as 'Ca. Syntrophonatronum acetioxidans' previously found to oxidize acetate at sulfate-reducing conditions up to salt-saturating concentrations. Butyrate and a benzoate-degrading syntrophs represent novel genus-level lineages in Syntrophomonadales which are proposed as Candidatus taxons 'Syntrophobaca', 'Syntrophocurvum' and 'Syntropholuna'. Overall, despite very slow growth, the results indicated the presence of a functionally competent syntrophic community in hypersaline soda lakes, capable of efficient oxidation of fermentation products to methane at extremely haloalkaline conditions.

Magnetospirillum caucaseum sp. nov., Magnetospirillum marisnigri sp. nov. and Magnetospirillum moscoviense sp. nov., freshwater magnetotactic bacteria isolated from three distinct geographical locations in European Russia.

Three strains of helical, magnetotactic bacteria, SO-1T, SP-1T and BB-1T, were isolated from freshwater sediments collected from three distinct locations in European Russia. Phylogenetic analysis showed that the strains belong to the genus Magnetospirillum. Strains SO-1T and SP-1T showed the highest 16S rRNA gene sequence similarity to Magnetospirillum magnetotacticum MS-1T (99.3 and 98.1 %, respectively), and strain BB-1T with Magnetospirillum gryphiswaldense MSR-1T (97.3 %). The tree based on concatenated deduced amino acid sequences of the MamA, B, K, M, O, P, Q and T proteins, which are involved in magnetosome formation, was congruent with the tree based on 16S rRNA gene sequences. The genomic DNA G+C contents of strains SO-1T, SP-1T and BB-1T were 65.9, 63.0 and 65.2 mol%, respectively. As major fatty acids, C18 : 1ω9, C16 : 1ω7c, C16 : 0 and C18 : 0 were detected. DNA-DNA hybridization values between the novel strains and their closest relatives in the genus Magnetospirillum were less than 51.7 ± 2.3 %. In contrast to M. magnetotacticum MS-1T, the strains could utilize butyrate and propionate; strains SO-1T and BB-1T could also utilize glycerol. Strain SP-1T showed strictly microaerophilic growth, whereas strains SO-1T and BB-1T were more tolerant of oxygen. The results of DNA-DNA hybridization and physiological tests allowed genotypic and phenotypic differentiation of the strains from each other as well as from the two species of Magnetospirillum with validly published names. Therefore, the strains represent novel species, for which we propose the names Magnetospirillum caucaseum sp. nov. (type strain SO-1T = DSM 28995T = VKM B-2936T), Magnetospirillum marisnigri sp. nov. (type strain SP-1T = DSM 29006T = VKM B-2938T) and Magnetospirillum moscoviense sp. nov. (type strain BB-1T = DSM 29455T = VKM B-2939T).

Sulfate-dependent acetate oxidation under extremely natron-alkaline conditions by syntrophic associations from hypersaline soda lakes.

So far, anaerobic sulfate-dependent acetate oxidation at high pH has only been demonstrated for a low-salt-tolerant syntrophic association of a clostridium 'Candidatus Contubernalis alkalaceticum' and its hydrogenotrophic sulfate-reducing partner Desulfonatronum cooperativum. Anaerobic enrichments at pH 10 inoculated with sediments from hypersaline soda lakes of the Kulunda Steppe (Altai, Russia) demonstrated the possibility of sulfate-dependent acetate oxidation at much higher salt concentrations (up to 3.5 M total Na(+)). The most salt-tolerant purified cultures contained two major components apparently working in syntrophy. The primary acetate-fermenting component was identified as a member of the order Clostridiales forming, together with 'Ca. Contubernalis alkalaceticum', an independent branch within the family Syntrophomonadaceae. A provisional name, 'Ca. Syntrophonatronum acetioxidans', is suggested for the novel haloalkaliphilic clostridium. Two phylotypes of extremely haloalkaliphilic sulfate-reducing bacteria of the genus Desulfonatronospira were identified as sulfate-reducing partners in the acetate-oxidizing cultures under extreme salinity. The dominant phylotype differed from the two species of Desulfonatronospira described so far, whilst a minor component belonged to Desulfonatronum thiodismutans. The results proved that, contrary to previous beliefs, sulfate-dependent acetate oxidation is possible, albeit very slowly, in nearly saturated soda brines.

Proteinivorax tanatarense gen. nov., sp. nov., an anaerobic, haloalkaliphilic, proteolytic bacterium isolated from a decaying algal bloom, and proposal of Proteinivoraceae fam. nov.

Two strains of a novel anaerobic, protein- and nucleoside-utilizing bacterium, Z-910(T) and Z-810, were isolated. The strains were spore-forming, mainly nonmotile rods, exhibiting positive Gram reaction with Gram-positive cell wall structure. The strains were mesophilic and haloalkaliphilic. Cultures used proteins and proteinaceous substrates as carbon, nitrogen, and energy sources. Both strains used also ribonucleosides, cellobiose, pyruvate, and glycerol. Ribose and nucleobases did not support growth. The fermentation products from all utilized substrates were identical but varied in content and included straight and branched acids, as well as hydrogen and ammonia. When grown on tryptone, strain Z-910(T) was able to reduce fumarate, dimethyl sulfoxide, thiosulfate, and elemental sulfur. Neither nitrate nor sulfate was reduced. The DNA G + C content of strain Z-910(T) was 32.2 mol%. Phylogenetic analysis based on the 16S rRNA gene sequence similarity revealed that strains Z-910(T) and Z-810 represented a new branch within the order Clostridiales, with 90.2 % similarity to the nearest genus with a validly published name Anaerobranca gottschalkii DSM 13577(T). According to their physiological, chemotaxonomic, and phylogenetic properties, strains Z-910(T) and Z-810 represented a new genus and novel species, for which the name Proteinivorax tanatarense gen. nov., sp. nov. was proposed. Phylogenetic analysis showed that the genera Proteinivorax gen. nov. and Anaerobranca formed a separate cluster within the order Clostridiales. The family Proteinivoraceae fam. nov. comprising the genera Proteinivorax gen. nov. and Anaerobranca was therefore proposed within the order Clostridiales of the phylum Firmicutes with Proteinivorax as a type genus of the new family.

Analysis of community composition of sulfur-oxidizing bacteria in hypersaline and soda lakes using soxB as a functional molecular marker.

The diversity of soxB gene encoding a key enzyme of the Sox pathway sulfate thiohydrolase has been investigated in pure cultures of various halophilic and haloalkaliphilic sulfur-oxidizing bacteria (SOB) and in salt and soda lakes in southwestern Siberia and Egypt. The gene was detected in the majority of strains belonging to eleven SOB genera excluding members of genera Thiohalospira and Thioalkalimicrobium. The uncultured diversity of soxB in salt and soda lakes was low with a majority of detected sequences belonging to autotrophic SOB from the Gammaproteobacteria. In addition, the soxB analysis allowed detection of putative heterotrophic Gamma- and Alphaproteobacterial SOB yet unknown in culture. All clone libraries obtained from soda lakes contained soxB belonging to the genus Thioalkalivibrio in agreement with the cultivation results. Besides, representatives of the genera Halothiobacillus, Marinobacter, and Halochromatium and of the family Rhodobacteraceae have been detected in both type of saline lakes.

Genome sequence and analysis of the tuber crop potato.

Potato (Solanum tuberosum L.) is the world's most important non-grain food crop and is central to global food security. It is clonally propagated, highly heterozygous, autotetraploid, and suffers acute inbreeding depression. Here we use a homozygous doubled-monoploid potato clone to sequence and assemble 86% of the 844-megabase genome. We predict 39,031 protein-coding genes and present evidence for at least two genome duplication events indicative of a palaeopolyploid origin. As the first genome sequence of an asterid, the potato genome reveals 2,642 genes specific to this large angiosperm clade. We also sequenced a heterozygous diploid clone and show that gene presence/absence variants and other potentially deleterious mutations occur frequently and are a likely cause of inbreeding depression. Gene family expansion, tissue-specific expression and recruitment of genes to new pathways contributed to the evolution of tuber development. The potato genome sequence provides a platform for genetic improvement of this vital crop.

Draft genome sequence of the anoxygenic filamentous phototrophic bacterium Oscillochloris trichoides subsp. DG-6.

Oscillochloris trichoides is a mesophilic, filamentous, photoautotrophic, nonsulfur, diazotrophic bacterium which is capable of carbon dioxide fixation via the reductive pentose phosphate cycle and possesses no assimilative sulfate reduction. Here, we present the draft genome sequence of Oscillochloris trichoides subsp. DG-6, the type strain of the species, which has permitted the prediction of genes for carbon and nitrogen metabolism and for the light-harvesting apparatus.

Diversity of RuBisCO and ATP citrate lyase genes in soda lake sediments.

Sediments from six soda lakes of the Kulunda Steppe (Altai, Russia) and from hypersaline alkaline lakes of Wadi Natrun (Egypt) were analyzed for the presence of cbb and aclB genes encoding key enzymes Ci assimilation (RuBisCO in Calvin-Benson and ATP citrate lyase in rTCA cycles, respectively). The cbbL gene (RuBisCO form I) was found in all samples and was most diverse, while the cbbM (RuBisCO form II) and aclB were detected only in few samples and with a much lower diversity. The cbbL libraries from hypersaline lakes were dominated by members of the extremely haloalkaliphilic sulfur-oxidizing Ectothiorhodospiraceae, i.e. the chemolithotrophic Thioalkalivibrio and the phototrophic Halorhodospira. In the less saline soda lakes from the Kulunda Steppe, the cbbL gene comprised up to ten phylotypes with a domination of members of a novel phototrophic Chromatiales lineage. The cbbM clone libraries consisted of two major unidentified lineages probably belonging to chemotrophic sulfur-oxidizing Gammaproteobacteria. One of them, dominating in the haloalkaline lakes from Wadi Natrun, was related to a cbbM phylotype detected previously in a hypersaline lake with a neutral pH, and another, dominating in lakes from the Kulunda Steppe, was only distantly related to the Thiomicrospira cluster. The aclB sequences detected in two samples from the Kulunda Steppe formed a single, deep branch in the Epsilonproteobacteria, distantly related to Arcobacter sulfidicus.

Bacillus alkalidiazotrophicus sp. nov., a diazotrophic, low salt-tolerant alkaliphile isolated from Mongolian soda soil.

Strain MS 6(T) was obtained from a microoxic enrichment with a soda soil sample from north-eastern Mongolia in nitrogen-free alkaline medium at pH 10. The isolate had clostridia-like motile cells and formed ellipsoid endospores. It was able to fix dinitrogen gas growing on nitrogen-free alkaline medium. Strain MS 6(T) was a strictly fermentative bacterium without a respiratory chain, although it had a high catalase activity and tolerated aerobic conditions. It was an obligate alkaliphile with a pH range for growth between 7.5 and 10.6 (optimum at 9.0-9.5). Growth and nitrogen fixation at pH 10 were possible at a total salt content of up to 1.2 M Na(+) (optimum at 0.2-0.3 M). The dominant cellular fatty acids included C(16 : 0), C(16 : 1)omega7, anteiso-C(15 : 0) and C(14 : 0). The dominant isoprenoid quinone was MK-7. The cell-wall peptidoglycan contained meso-diaminopimelic acid as the diagnostic diamino acid. 16S rRNA gene sequencing identified strain MS 6(T) as a member of the genus Bacillus. Its closest relative was Bacillus arseniciselenatis E1H(T). The key functional nitrogenase gene nifH was detected in both strain MS 6(T) and its close relative and these strains formed a novel lineage in the nifH gene family. On the basis of these phenotypic and genetic comparisons, strain MS 6(T) is proposed to represent a novel species of the genus Bacillus, Bacillus alkalidiazotrophicus sp. nov. with the type strain MS 6(T) (=NCCB 100213(T)=UNIQEM U377(T)).

Methylohalomonas lacus gen. nov., sp. nov. and Methylonatrum kenyense gen. nov., sp. nov., methylotrophic gammaproteobacteria from hypersaline lakes.

Aerobic enrichment at 4 M NaCl, pH 7.5, with methanol as carbon and energy source from sediments of hypersaline chloride-sulfate lakes in Kulunda Steppe (Altai, Russia) resulted in the isolation of a moderately halophilic and obligately methylotrophic bacterium, strain HMT 1(T). The bacterium grew with methanol and methylamine within a pH range of 6.8-8.2 with an optimum at pH 7.5 and at NaCl concentrations of 0.5-4 M with an optimum at 2 M. In addition to methanol and methylamine, it can oxidize ethanol, formate, formaldehyde and dimethylamine. Carbon is assimilated via the serine pathway. The main compatible solute is glycine betaine. 16S rRNA gene sequence analysis placed the isolate as a new lineage in the family Ectothiorhodospiraceae (Gammaproteobacteria). It is proposed, therefore, to accommodate this bacterium within a novel genus and species, Methylohalomonas lacus gen. nov., sp. nov., with HMT 1(T) (=DSM 15733(T) =NCCB 100208(T) =UNIQEM U237(T)) as the type strain. Two strains were obtained in pure culture from sediments of soda lake Magadi in Kenya and the Kulunda Steppe (Russia) on a mineral medium at pH 10 containing 0.6 M total Na(+) using methanol as a substrate. Strain AMT 1(T) was enriched with methanol, while strain AMT 3 originated from an enrichment culture with CO. The isolates are restricted facultative methylotrophs, capable of growth with methanol, formate and acetate as carbon and energy sources. With methanol, the strains grew within a broad salinity range from 0.3 to 3.5-4 M total Na(+), with an optimum at 0.5-1 M. The pH range for growth was between 8.3 and 10.5, with an optimum at pH 9.5, which characterized the soda lake isolates as obligate haloalkaliphiles. Carbon is assimilated autotrophically via the Calvin-Benson cycle. Sequence analysis of the gene coding for the key enzyme RuBisCO demonstrated that strain AMT 1(T) possessed a single cbbL gene of the 'green' form I, clustering with members of the family Ectothiorhodospiraceae. Analysis of the 16S rRNA gene sequence showed that strains AMT 1(T) and AMT 3 belong to a single species that forms a separate lineage within the family Ectothiorhodospiraceae. On the basis of phenotypic and genetic data, the novel haloalkaliphilic methylotrophs are described as representing a novel genus and species, Methylonatrum kenyense gen. nov., sp. nov. (type strain AMT 1(T) =DSM 15732(T) =NCCB 100209(T) =UNIQEM U238(T)).

Thiomicrospira halophila sp. nov., a moderately halophilic, obligately chemolithoautotrophic, sulfur-oxidizing bacterium from hypersaline lakes.

Enrichments at 2 M NaCl and pH 7.5-8, with thiosulfate or sulfide as electron donor, inoculated with sediments from hypersaline chloride-sulfate lakes of the Kulunda Steppe (Altai, Russia) resulted in the domination of two different groups of moderately halophilic, chemolithoautotrophic, sulfur-oxidizing bacteria. Under fully aerobic conditions with thiosulfate, bacteria belonging to the genus Halothiobacillus dominated while, under microaerophilic conditions, a highly motile, short vibrio-shaped phenotype outcompeted the halothiobacilli. Three genetically and phenotypically highly similar vibrio-shaped isolates were obtained in pure culture and one of them, strain HL 5(T), was identified as a member of the Thiomicrospira crunogena cluster by 16S rRNA gene sequencing. The new isolates were able to grow with thiosulfate as electron donor within a broad salinity range from 0.5 to 3.5 M NaCl with an optimum at 1.5 M and within a pH range from 6.5 to 8.5 with an optimum at pH 7.5-7.8. Comparative analysis of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) gene sequences demonstrated that strain HL 5(T) possessed two genes, cbbL-1 and cbbL-2, of the form I RuBisCO and a cbbM gene of the form II RuBisCO, similar to the other members of the Thiomicrospira crunogena cluster. On the basis of phenotypic and genetic comparison, the new halophilic isolates are proposed to be placed into a novel species, Thiomicrospira halophila sp. nov. (type strain HL 5(T)=DSM 15072(T)=UNIQEM U 221(T)).

Thermincola carboxydiphila gen. nov., sp. nov., a novel anaerobic, carboxydotrophic, hydrogenogenic bacterium from a hot spring of the Lake Baikal area.

A novel anaerobic, thermophilic, alkalitolerant bacterium, strain 2204(T), was isolated from a hot spring of the Baikal Lake region. The cells of strain 2204(T) were straight rods of variable length, Gram-positive with an S-layer, motile with one to two lateral flagella, and often formed aggregates of 3-15 cells. The isolate was shown to be an obligate anaerobe oxidizing CO and producing equimolar quantities of H(2) and CO(2) according to the equation CO+H(2)O-->CO(2)+H(2). No organic substrates were used as energy sources. For lithotrophic growth on CO, 0.2 g acetate or yeast extract l(-1) was required but did not support growth in the absence of CO. Growth was observed in the temperature range 37-68 degrees C, the optimum being 55 degrees C. The pH range for growth was 6.7-9.5, the optimum pH being 8.0. The generation time under optimal conditions was 1.3 h. The DNA G+C content was 45 mol%. Penicillin, erythromycin, streptomycin, rifampicin, vancomycin and tetracycline completely inhibited both growth and CO utilization by strain 2204(T). Thus, isolate 2204(T) was found to be the first known moderately thermophilic and alkalitolerant H(2)-producing anaerobic carboxydotroph. The novel bacterium fell within the cluster of the family Peptococcaceae within the low-G+C-content Gram-positive bacteria, where it formed a separate branch. On the basis of morphological, physiological and phylogenetic features, strain 2204(T) should be assigned to a novel genus and species, for which the name Thermincola carboxydiphila gen. nov., sp. nov. is proposed. The type strain is strain 2204(T) (=DSM 17129(T)=VKM B-2283(T)=JCM 13258(T)).

Cultivated anaerobic acidophilic/acidotolerant thermophiles from terrestrial and deep-sea hydrothermal habitats.

Metabolic and phylogenetic diversity of cultivated anaerobic microorganisms from acidic continental hot springs and deep-sea hydrothermal vents was studied by molecular and microbiological methods. Anaerobic organotrophic enrichment cultures growing at pH 3.5-4.0 and 60 or 85 degrees C with organic energy sources were obtained from samples of acidic hot springs of Kamchatka Peninsula (Pauzhetka, Moutnovski Volcano, Uzon Caldera) and Kunashir Island (South Kurils) as well as from the samples of chimneys of East Pacific Rise (13 degrees N). The analyses of clone libraries obtained from terrestrial enrichment cultures growing at 60 degrees C revealed the presence of archaea of genus Thermoplasma and bacteria of genus Thermoanaerobacter. Bacterial isolates from these enrichments were shown to belong to genera Thermoanaerobacter and Thermoanaerobacterium, being acidotolerant with the pH optimum for growth at 5.5-6.0 and the pH minimum at 3.0. At 85 degrees C, domination of thermoacidophilic archaea of genus Acidilobus in terrestrial enrichments was found by both molecular and microbiological methods. Five isolates belonging to this genus possessed some phenotypic features that were new for this genus, such as flagellation or the ability to grow on monosaccharides or disaccharides. Analyses of clone libraries from the deep-sea thermoacidophilic enrichment cultures showed that the representatives of the genus Thermococcus were present at both 60 and 85 degrees C. From the 60 degrees C deep-sea enrichment, a strain belonging to Thermoanaerobacter siderophilus was isolated. It grew optimally at pH 6.0 with the minimum pH for growth at 3.0 and with salinity optimum at 0-2.5% NaCl and the maximum at 7%, thus differing significantly from the type strain. These data show that fermentative degradation of organic matter may occur at low pH and wide temperature range in both terrestrial and deep-sea habitats and can be performed by acidophilic or acidotolerant thermophilic prokaryotes.

Methylobacillus pratensis sp. nov., a novel non-pigmented, aerobic, obligately methylotrophic bacterium isolated from meadow grass.

Strain F31T was isolated from meadow grass (Poa trivialis L.) sampled from the city park in Helsinki. Analysis of phenotypic and genotypic properties showed the strain to be related to the group of obligately methylotrophic non-methane utilizing bacteria (methylobacteria) with the ribulose monophosphate pathway of formaldehyde assimilation. Phylogenetic analysis showed the strain to be closely related to the genus Methylobacillus, and analysis of fatty acid composition confirmed this association. Thus, on the basis of its genotypic and phenotypic properties, the isolate is proposed as a novel species of the genus Methylobacillus, Methylobacillus pratensis sp. nov., with F31T as the type strain (= VKM B-2247T = NCIMB 13994T).