The role of the "Thiodendron" consortium in postulating the karyomastigont chimaera of the endosymbiosis theory by Lynn Margulis.

The endosymbiosis theory of the origin of eukaryotic cell was first proposed more than a hundred years ago. In the second half of the 20th century, Lynn Margulis suggested a new interpretation of the origin of the nucleus in modern eukaryotes. The background was the study of the consortium "Thiodendron", a symbiotic bacterial community, which includes anaerobic aerotolerant motile spirochaetes and sulfidogenic bacteria (sulfidogens) of vibrioid form with a fermentation type of metabolism. Spirochaetes supply sulfidogens with metabolites (pyruvate and, probably, organic nitrogenous products of cell lysis) and get hydrogen sulfide from sulfidogens that helps to maintain a low redox potential. At low oxygen concentrations, spirochaetes are able to assimilate glucose more efficiently. Margulis hypothesized about the symbiotic origin of the nucleus by adding the bacterium Spirochaeta to the Thermoplasma-like archaea. She considered the "Thiodendron"-like consortium to be an intermediate stage in evolution. According to Margulis, the conversion of carbohydrates and the oxidation of Н2S to S0 by the bacterium provided the archaea with electron acceptors for anaerobic respiration, as shown for modern thermoplasmas and products saturated with carbon. The use of carbon sources increased by attaching the floating bacterium to the archaea. More efficient microaerobic oxidation of glucose pre-adapted the spirochaetes for association with Thermoplasma. However, modern "Thiodendron"-like consortia are not in stable symbiosis and a sulfidogenic component of the consortium is capable for fermentation, rather than anaerobic respiration, which makes the theory by Margulis disputable.

Description of Oceanispirochaeta crateris sp. nov. and reclassification of Spirochaeta perfilievii as Thiospirochaeta perfilievii gen. nov., comb. nov.

A novel obligately anaerobic spirochete strain K2T was isolated from bottom marine sediments at Crater Bay of Yankicha Island (Kuril Islands, Russia). Strain K2T had helical shape and Gram-negatively stained. The optimal growth conditions were as follows: the optimum temperature was 28-30 °C with range 5-34 °C; optimal pH at 7.0-7.5 with range of 6.8-8.5; NaCl optimum at 3-3.5 % (w/v) and range of 1-7 % (w/v). Strain K2T was catalase- and oxidase-negative. Glucose fermentation products were acetate, lactate, ethanol, CO2, H2. The major fatty acids were C14 : 0, iso-C13 : 0, iso-C15:0, C14 : 0 DMA, iso-C15 : 0 DMA. The G+C content of genomic DNA was 43.2 mol%. Phylogenetic analyses of 16S rRNA genes showed that strain K2T belonged to the genus Oceanispirochaeta of the family Spirochaetaceae. The 16S rRNA gene sequence similarity of strain K2T and O. litoralis DSM 2029T and O. sediminicola DSM 104770T was 96 and 94 %, respectively. Based on the results of our study, we propose the name Oceanispirochaeta crateris sp. nov.; type strain K2T (=DSM 16308T=VKM B-3266T). Also, the taxonomic status of Spirochaeta perfilevii was revised: 16S rRNA genes sequence showed less than 89 % similarity to nearest phylogenetic neighbours. Therefore, we proposed to separate this species into a novel genus Thiospirochaeta - T. perfilievii gen. nov., comb. nov.

Complete Genome Sequences and Methylome Analysis of Two Environmental Spirochaetes.

Here, we report the finished closed genomes of two environmental bacteria, Oceanispirochaeta crateria K2 and Thiospirochaeta perfilievii P (formally known as Spirochaeta perfilievii P). In addition, we provide methylation data and the associated enzymes predicted and confirmed to be responsible for each modified motif.

Complete Genome Sequence and Methylome Analysis of Sphaerotilus natans subsp. sulfidivorans D-507.

Sphaerotilus natans subsp. sulfidivorans D-507 is an environmental isolate from a sulfate spring in the northern Caucasus region of Russia. This heterotrophic bacterium is involved in the oxidation of reduced sulfur derivatives. This report includes the finished genome of this strain. In addition, we provide methylation data and the associated enzymes predicted to be responsible for each modified motif.

Genomics and Biochemistry of Metabolic Pathways for the C1 Compounds Utilization in Colorless Sulfur Bacterium Beggiatoa leptomitoformis D-402.

The metabolic pathways of one-carbon compounds utilized by colorless sulfur bacterium Beggiatoa leptomitoformis D-402 were revealed based on comprehensive analysis of its genomic organization, together with physiological, biochemical and molecular biological approaches. Strain D-402 was capable of aerobic methylotrophic growth with methanol as a sole source of carbon and energy and was not capable of methanotrophic growth because of the absence of genes of methane monooxygenases. It was established that methanol can be oxidized to CO2 in three consecutive stages. On the first stage methanol was oxidized to formaldehyde by the two PQQ (pyrroloquinolinequinone)-dependent methanol dehydrogenases (MDH): XoxF and Mdh2. Formaldehyde was further oxidized to formate via the tetrahydromethanopterin (H4MPT) pathway. And on the third stage formate was converted to CO2 by NAD+-dependent formate dehydrogenase Fdh2. Finally, it was established that endogenous CO2, formed as a result of methanol oxidation, was subsequently assimilated for anabolism through the Calvin-Benson-Bassham cycle. The similar way of one-carbon compounds utilization also exists in representatives of another freshwater Beggiatoa species-B. alba.

Complete Genome Sequence of the Freshwater Bacterium Beggiatoa leptomitoformis Strain D-401.

Here, we report the complete closed genome sequence and methylome analysis of Beggiatoa leptomitoformis strain D-401 (DSM 14945, UNIQEMU 779), which is quite different from the previously described Beggiatoa leptomitoformis neotype strain D-402T (DSM 14946, UNIQEM U 779) with regard to morphology and lithotrophic growth in the presence of thiosulfate.

Whole-Genome Sequence and Methylome Analysis of the Freshwater Colorless Sulfur Bacterium Thioflexothrix psekupsii D3.

In this report, we announce the availability of a whole-genome sequence and methylome analysis of Thioflexothrix psekupsii strain D3.

Beggiatoa leptomitoformis sp. nov., the first freshwater member of the genus capable of chemolithoautotrophic growth.

A strain of filamentous sulfur bacteria was isolated from freshwater spring contaminated with residential and agricultural wastewater in Moscow region, Russia. According to the results of phylogenetic analysis, strain D-402T belonged to the genus Beggiatoa within the family Beggiatoaceae of the class Gammaproteobacteria. Within the genus Beggiatoa, strain D-402T was most closely related to Beggiatoa alba strains. Strain D-402T had a DNA G+C content 42.1 mol%. The DNA-DNA hybridization value between strain D-402T and Beggiatoa alba strain B15LD was 33 %. Predominant fatty acids were C18 : 1 (46.1 and 53.3 %), C16 : 0 (15.5 and 16.2 %) and C16 : 1 (32.9 and 25.4 %) for strains D-402T and B15LD, respectively. In contrast to known representatives of Beggiatoa, strain D-402T was capable of chemolithoautotrophic growth with sulfide and thiosulfate as electron donors. Oxidation of sulfide and thiosulfate was accompanied by deposition of sulfur globules within the cells. Strain D-402T was capable of heterotrophic growth. The strain was capable of using different organic compounds, sulfur compounds and hydrogen as electron donors. Based on these observations, strain D-402T is considered as a representative of a species Beggiatoa leptomitoformis sp. nov. of the genus Beggiatoa. The type strain is D-402T (=DSM 14946T=UNIQEM U 779T).

Genomic insights into metabolic versatility of a lithotrophic sulfur-oxidizing diazotrophic Alphaproteobacterium Azospirillum thiophilum.

Diazotrophic Alphaproteobacteria of the genus Azospirillum are usually organotrophs, although some strains of Azospirillum lipoferum are capable of hydrogen-dependent autotrophic growth. Azospirillum thiophilum strain was isolated from a mineral sulfide spring, a biotope highly unusual for azospirilla. Here, the metabolic pathways utilized by A. thiophilum were revealed based on comprehensive analysis of its genomic organization, together with physiological and biochemical approaches. The A. thiophilum genome contained all the genes encoding the enzymes of carbon metabolism via glycolysis, tricarboxylic acid cycle and glyoxylate cycle. Genes for a complete set of enzymes responsible for autotrophic growth, with an active Calvin-Benson-Bassham cycle, were also revealed, and activity of the key enzymes was determined. Microaerobic chemolithoautotrophic growth of A. thiophilum was detected in the presence of thiosulfate and molecular hydrogen, being in line with the discovery of the genes encoding the two enzymes involved in dissimilatory thiosulfate oxidation, the Sox-complex and thiosulfate dehydrogenase and Ni-Fe hydrogenases. Azospirillum thiophilum utilizes methanol and formate, producing CO2 that can further be metabolized via the Calvin cycle. Finally, it is capable of anaerobic respiration, using tetrathionate as a terminal electron acceptor. Such metabolic versatility is of great importance for adaptation of A. thiophilum to constantly changing physicochemical environment.

Complete Genome Sequence of a Strain of Azospirillum thiophilum Isolated from a Sulfide Spring.

We report the complete, closed genome sequence and complete methylome of Azospirillum thiophilum strain BV-S(T).

Complete Genome Sequence of the Freshwater Colorless Sulfur Bacterium Beggiatoa leptomitoformis [corrected] Neotype Strain D-402T.

In this report, we announce the availability of a complete closed genome sequence and methylome analysis of Beggiatoa leptomitiformis neotype strain D-402(T) (DSM 14946, UNIQEM U 779).

Spirochaeta sinaica sp. nov., a halophilic spirochaete isolated from a cyanobacterial mat.

A strain of free-living obligately anaerobic, halophilic spirochaete, SLT, was isolated from a sample of a cyanobacterial mat of the hypersaline Solar Lake, Sinai shore. The strain had motile helical cells, 0.35-0.40 × 6-10 µm. Strain SLT exhibited high resistance to NaCl among known halophilic spirochaetes growing at NaCl concentrations from 2 to 12% (optimum growth at 7%). The strain grew at temperatures from 10 to 32 °C (optimum at 28 °C) and pH from 6 to 8.5 (optimum at pH 7.0-7.5). Carbohydrates, but not alcohols, organic acids or nitrogenous compounds (peptone, yeast extract and amino acids), were used as energy substrates for growth. Ethanol, acetate, lactate, H2 and CO2 were the products of glucose fermentation. Sulfide was produced in the presence of S0 or thiosulfate in the medium. The DNA G+C content was 44.7 mol%. Based on 16S rRNA gene sequence analysis, strain SLT clustered within the genus Spirochaeta, exhibiting 94.2 and 93.7% similarity with its closest relatives, Spirochaeta bajacaliforniensis DSM 160554T and Spirochaeta smaragdinae DSM 11293T, respectively; similarity with other species did not exceed 86%. The phenotypic and chemotaxonomic characteristics of the strain, as well as the results of phylogenetic analysis support the classification of strain SLT as representing a novel species of the genus Spirochaeta, for which the name Spirochaeta sinaica sp. nov. is proposed. The type strain is SLT ( = DSM 14994 = NIQEM U 783).

Expansion of ability of denitrification within the filamentous colorless sulfur bacteria of the genus Thiothrix.

Filamentous sulfur bacteria of the genus Thiothrix are able to respire nitrate (NO3-→NO2-) under anaerobic growth. Here, Thiothrix caldifontis (G1(T), G3), Thiothrix unzii (A1(T), TN) and Thiothrix lacustris AS were shown to be capable of further reduction of nitrite and/or nitrous oxides (denitrification). In particular, in the genomes of these strains, excluding T. unzii TN, the nirS gene encoding periplasmic respiratory nitrite reductase was detected, and for T. lacustris AS the nirS expression was confirmed during anaerobic growth. The nirK gene, coding for an alternative nitrite reductase, and the nrfA gene, encoding nitrite reduction to ammonia, were not found in any investigated strains. All Thiothrix species capable of denitrification possess the cnorB gene encoding cytochrome c-dependent NO reductase but not the qnorB gene coding for quinol-dependent NO reductase. Denitrifying capacity ('full' or 'truncated') can vary between strains belonging to the same species and correlates with physical-chemical parameters of the environment such as nitrate, hydrogen sulfide and oxygen concentrations. Phylogenetic analysis revealed the absence of recent horizontal transfer events for narG and nirS; however, cnorB was subjected to gene transfer before the separation of modern species from a last common ancestor of the Thiothrix species.

Ferrovibrio denitrificans gen. nov., sp. nov., a novel neutrophilic facultative anaerobic Fe(II)-oxidizing bacterium.

A neutrophilic Fe(II)-oxidizing bacterium was isolated from the redox zone of a low-salinity spring in Krasnodar krai (Russia), at the FeS-Fe(OH)(3) interface deposited at the sediment surface. The cells of strain Sp-1 were short, thin motile vibrioids with one polar flagellum dividing by binary fission. The optimal values and ranges for pH and temperature were pH 6.2 (5.5-8) and 35 °C (5-45 °C), respectively. The organism was a facultative anaerobe. Strain Sp-1 was capable of organotrophic, lithoheterotrophic and mixotrophic growth with Fe(II) as an electron donor. The denitrification chain was 'disrupted'. Oxidation of Fe(II) was coupled to reduction of NO3 - to NO2 - or of N(2) O to N(2) , as well as under microaerobic conditions, with O(2) as an electron acceptor. The DNA G+C content was 64.2 mol%. According to the results of phylogenetic analysis, the strain was 10.6-12% remote from the closest relatives, members of the genera Sneathiella, Inquilinus, Oceanibaculum and Phaeospirillum within the Alphaproteobacteria. Based on its morphological, physiological and taxonomic characteristics, together with the results of phylogenetic analysis, strain Sp-1 is described as a member of a new genus Ferrovibrio gen. nov., with the type species Ferrovibrio denitrificans sp. nov. and the type strain Sp-1(T) (= LMG 25817(T)  = VKM B-2673(T) ).

Taxonomic investigation of representatives of the genus Sphaerotilus: descriptions of Sphaerotilus montanus sp. nov., Sphaerotilus hippei sp. nov., Sphaerotilus natans subsp. natans subsp. nov. and Sphaerotilus natans subsp. sulfidivorans subsp. nov., and an emended description of the genus Sphaerotilus.

Seven strains of the genus Sphaerotilus were obtained from natural thermal sulfide (strains D-501(T), D-502, D-504, D-505 and D-507) and low-temperature ferrous (strain HS(T)) springs and from an activated sludge system (strain D-380). These Sphaerotilus isolates and strains of Sphaerotilus natans obtained from the DSMZ (S. natans DSM 6575(T), DSM 565 and DSM 566) were studied using a polyphasic taxonomic approach. All strains had Q-8 as the major quinone and C(16 : 1)ω7, C(16 : 0) and C(18 : 1)ω7 as the major fatty acids. The DNA-DNA hybridization results and 16S rRNA, hsp60 and gyrB gene sequencing experiments showed that isolates D-501(T), D-502, D-504, D-505, D-507 and D-380 were closely related to the type strain of S. natans DSM 6575(T). However, strains D-501(T), D-502, D-504, D-505 and D-507 significantly differed from the heterotrophic strain S. natans DSM 6575(T) by their capability for lithotrophic growth with reduced sulfur compounds as an electron donor for energy conservation and some other phenotypic features. For this reason, strains D-501(T), D-502, D-504, D-505 and D-507 merit a separate taxonomic classification at the subspecies level. The name Sphaerotilus natans subsp. sulfidivorans subsp. nov. (type strain D-501(T) = DSM 22545(T) = VKM B-2573(T)) is proposed. The subspecies Sphaerotilus natans subsp. natans subsp. nov. is automatically created as a result of this proposal. Strain D-380 was phenotypically closely related to S. natans DSM 6575(T). Strains D-380 and S. natans DSM 6575(T) were assigned to the subspecies Sphaerotilus natans subsp. natans subsp. nov. (type strain DSM 6575(T) = ATCC 13338(T)). The 16S rRNA, hsp60 and gyrB gene sequences obtained for strains HS(T) and DSM 565 showed very low sequence similarity values of 97.3 %, 89.7 % and 88.4 %, respectively, with S. natans DSM 6575(T). Strain HS(T) shared 99 % DNA-DNA relatedness with strain.

Spirochaeta perfilievii sp. nov., an oxygen-tolerant, sulfide-oxidizing, sulfur- and thiosulfate-reducing spirochaete isolated from a saline spring.

A novel strain of fermenting, aerotolerant, chemo-organoheterotrophic spirochaete designated P(T) was isolated from a sulfur 'Thiodendron' mat in a saline spring at the Staraya Russa resort (Novgorod Region, Russia). Cells of strain P(T) exhibited a helical shape. The spirochaete required sulfide in the growth medium and was able to oxidize it non-enzymically to elemental sulfur via the interaction of H(2)O(2) with sulfide and deposit it in the periplasmic space. Growth occurred at 4-32 °C (optimum at 28-30 °C), pH 6.0-8.5 (optimum pH 7.0-7.5), and in 0.1-1 M NaCl (optimum 0.35 M). The isolate used several sugars and polysaccharides as carbon or energy sources but did not use peptides, amino acids, organic acids or alcohols. The products of glucose fermentation were formate, acetate, ethanol, pyruvate, CO(2) and H(2). The genomic DNA G+C content was 41.7 mol%. 16S rRNA gene sequence analysis showed that strain P(T) fell within a group of species in the genus Spirochaeta, including Spirochaeta litoralis, S. isovalerica and S. cellobiosiphila, with which it shared less then 89 % sequence similarity. On the basis of its morphology, physiology and other phenotypic properties, as well as its phylogenetic position, the new isolate is considered to represent a novel species of the genus Spirochaeta, for which the name Spirochaeta perfilievii sp. nov. is proposed. The type strain is P(T) (=DSM 19205(T) =VKM B-2514(T)).

Azospirillum thiophilum sp. nov., a diazotrophic bacterium isolated from a sulfide spring.

A novel nitrogen-fixing strain, designated BV-S(T), was isolated from a sulfur bacterial mat collected from a sulfide spring of the Stavropol Krai, North Caucasus, Russia. Strain BV-S(T) grew optimally at pH 7.5 and 37°C. According to the results of phylogenetic analysis, strain BV-S(T) belonged to the genus Azospirillum within the family Rhodospirillaceae of the class Alphaproteobacteria. Within the genus Azospirillum, strain BV-S(T) was most closely related to Azospirillum doebereinerae GSF71(T), A. picis IMMIB TAR-3(T) and A. lipoferum ATCC 29707(T) (97.7, 97.7 and 97.4 % 16S rRNA gene sequence similarity, respectively). DNA-DNA relatedness between strain BV-S(T) and A. doebereinerae DSM 13131(T), A. picis DSM 19922(T) and A. lipoferum ATCC 29707(T) was 38, 55 and 42 %, respectively. Similarities between nifH sequences of strain BV-S(T) and members of the genus Azospirillum ranged from 94.5 to 96.8 %. Chemotaxonomic characteristics (quinone Q-10, major fatty acid C(18 : 1)ω7c and G+C content 67 mol%) were similar to those of members of the genus Azospirillum. In contrast to known Azospirillum species, strain BV-S(T) was capable of mixotrophic growth under microaerobic conditions with simultaneous utilization of organic substrates and thiosulfate as electron donors for energy conservation. Oxidation of sulfide was accompanied by deposits of sulfur globules within the cells. Based on these observations, strain BV-S(T) is considered as a representative of a novel species of the genus Azospirillum, for which the name Azospirillum thiophilum sp. nov. is proposed. The type strain is BV-S(T) (=DSM 21654(T) =VKM B-2513(T)).

Thiothrix caldifontis sp. nov. and Thiothrix lacustris sp. nov., gammaproteobacteria isolated from sulfide springs.

Five strains of filamentous, sulfur-oxidizing bacteria were isolated from sulfur mats of different sulfide springs from various regions of the Northern Caucasus, Russia. A phylogenetic analysis based on 16S rRNA gene sequence comparison showed that all of the isolates are affiliated with the filamentous, colourless, sulfur-oxidizing bacteria of the genus Thiothrix within the Gammaproteobacteria and are closely related to Thiothrix fructosivorans. All strains are capable of growing heterotrophically, lithoautotrophically with thiosulfate or sulfide as the sole energy source and mixotrophically. Strains G1(T), G2, P and K2 are able to fix molecular nitrogen, but strain BL(T) is not. Randomly amplified polymorphic DNA (RAPD)-PCR analysis was used to assess the level of genetic relationships among the Thiothrix isolates. The Nei and Li similarity index revealed high genetic similarity among strains G1(T), G2, P and K2 (above 75 %), indicating that they are closely related. In combination with physiological and morphological data, strains G1(T), G2, P and K2 can be considered as members of the same species. The lowest genetic similarity (approx. 20 %) was reached between strain BL(T) and the other isolated Thiothrix strains. Strains BL(T) and G1(T) shared 35 % DNA-DNA relatedness and showed 51 and 53 % relatedness, respectively, to Thiothrix fructosivorans ATCC 49749. On the basis of this polyphasic analysis, strains G1(T), G2, P and K2 represent a novel species within the genus Thiothrix, for which the name Thiothrix caldifontis sp. nov. is proposed, with strain G1(T) (=DSM 21228(T) =VKM B-2520(T)) as the type strain. In addition, strain BL(T) represents a second novel species, Thiothrix lacustris sp. nov., with strain BL(T) (=DSM 21227(T) =VKM B-2521(T)) as the type strain.

Proposal of Spirillum winogradskyi sp. nov., a novel microaerophilic species, an emended description of the genus Spirillum and Request for an Opinion regarding the status of the species Spirillum volutans Ehrenberg 1832.

A novel obligately organotrophic, facultatively microaerophilic spirillum, designated strain D-427(T), was isolated from sulfidic sludge of a municipal wastewater-treatment plant. Cells were Gram-negative, large and highly motile due to bipolar tufts of flagella covered with mucous sheaths. Coccoid cells were sometimes formed. Strain D-427(T) grew optimally at pH 7.5-7.8 and 28 degrees C in the presence of 2 % O(2) in the gas phase. The organism showed oxidase and very low catalase activity. The isolate grew chemo-organotrophically with a limited number of organic acids as substrates. The DNA G+C content was 38.0 mol% (T(m)). Phylogenetic analysis of the 16S rRNA gene sequence placed strain D-427(T) in the genus Spirillum within the class Betaproteobacteria. The 16S rRNA gene sequence similarity between strain D-427(T) and Spirillum volutans ATCC 19554(T), the type strain of the single species of the genus, was 98.6 %. The low level of DNA-DNA hybridization and different phenotypic properties indicate that strain D-427(T) is clearly distinguishable from Spirillum volutans. No strain of S. volutans is available from any established culture collection or from the authors who described this species. Therefore, on the basis of phenotypic and genotypic data and the fact that the type and single species of the genus Spirillum cannot be included in any scientific study, since the type strain has been lost, we propose to assign strain D-427(T) as a novel species of the genus Spirillum, Spirillum winogradskyi sp. nov. (type strain D-427(T) =DSM 12756(T) =VKM B-2518(T)), and we request that the Judicial Commission place the name Spirillum volutans on the list of rejected names if a suitable type strain is not found or a neotype is not proposed within 2 years following the publication of this paper. An emended description of the genus Spirillum is also provided.