T. N. Zhilina

Desulfonatronovibrio hydrogenovorans gen. nov., sp. nov., an alkaliphilic, sulfate-reducing bacterium.

A new alkaliphilic, sulfate-reducing bacterium, strain Z-7935T (T = type strain), was isolated from a soda-depositing lake, Lake Magadi in Kenya. This organism is a motile vibrio which utilizes only hydrogen and formate as electron donors and sulfate, sulfite, and thiosulfate, but not sulfur, as electron acceptors. Thiosulfate is dismutated. Strain Z-7935T is an obligately sodium-dependent alkaliphile which grows in sodium carbonate medium and does not grow at pH 7; the maximum pH for growth is more than pH 10, and the optimum pH is 9.5 to 9.7. The optimum NaCl concentration for growth is 3% (wt/vol). The optimum temperature for growth is 37 degrees C. The G + C content of the DNA is 48.6 mol%. 16S ribosomal DNA sequence analysis revealed that strain Z-7935T represents a new lineage with genus status in the delta subclass of the Proteobacteria. The name Desulfonatronovibrio hydrogenovorans gen. nov., sp. nov. is proposed for this organism; the type strain of D. hydrogenovorans is strain Z-7935 (= DSM 9292).

Spirochaeta alkalica sp. nov., Spirochaeta africana sp. nov., and Spirochaeta asiatica sp. nov., Alkaliphilic Anaerobes from the Continental Soda Lakes in Central Asia and the East African Rift

During a study of microbial communities in athalassic bodies of water, three new species within the genus Spirochaeta were described. These are alkaliphilic Spirochaeta alkalica sp. nov. Z-7491 (DSM 8900) and halophilic S. africana sp. nov. Z-7692 (DSM 8902) from the soda-depositing Lake Magadi in Central Africa and haloalkaliphilic S. asiatica sp. nov. Z-7591 (DSM 8901) from Lake Khatyn, Central Asia. These mesophilic spirochetes develop at pHs of > 9 as anaerobic saccharolytic dissipotrophs. The DNA base compositions (moles percent G+C) of the strains were as follows: S. alkalica Z-7491, 57.1; S. africana Z-7692, 56.1; and S. asiatica Z-7591, 49.2. The optimum growth parameters (temperature, pH, and NaCl concentration [percent, wt/vol], respectively) were as follows: for S. alkalica Z-7491, 35 degrees C, 9.2, and 5 to 7%; for S. africana Z-7692, 35 degrees C, 9.3, and 5 to 7%; and for S. asiatica Z-7591, 35 degrees C, 8.9, and 3 to 6%. The products of glucose fermentation were acetate, hydrogen, ethanol, and lactate, in different proportions, for S. alkalica and S. africana; for S. asiatica, they were acetate, ethanol, and lactate. S. asiatica is strictly anaerobic, while S. alkalica and S. africana are rather aerotolerant. All three species group within the radiation of the majority of the species of the genus Spirochaeta. Studies of the genes encoding 16S rRNA indicate a possible fanning out of the phylogenetic tree of spirochetes.

New species of psychrophilic acetogens: Acetobacterium bakii sp. nov., A. paludosum sp. nov., A. fimetarium sp. nov.

Three strains of new acetogenic bacteria were isolated from several low temperature environments. Cells were gram-positive, oval-shaped flagellated rods. The organisms fermented H2/CO2, CO, formate, lactate, and several sugars to acetate. Strains Z-4391 and Z-4092 grew in the temperature range from 1 to 30°C with an optimum at 20°C; strain Z-4290 grew in the range from 1 to 35°C with an optimum at 30°C. The DNA G+C content of strains Z-4391, Z-4092, and Z-4290 was 42.1, 41.7, and 45.8 mol% respectively.

Clostridium alkalicellum sp. nov., an Obligately Alkaliphilic Cellulolytic Bacterium from a Soda Lake in the Baikal Region

The first anaerobic alkaliphilic cellulolytic microorganism has been isolated from the Verkhnee Beloe soda lake (Buryatiya, Russia) with pH 10.2 and a salt content of up to 24 g/l. Five strains were characterized. Strain Z-7026 was chosen as the type strain. The cells of the isolate are gram-positive spore-forming rods. A mucous external capsule is produced. The microorganism is obligately alkaliphilic, growing in a pH range of 8.0–10.2, with an optimum at pH 9.0. Sodium ions and, in carbonate-buffered media, sodium chloride are obligately required. The microorganism is slightly halophilic; it grows at 0.017–0.4 M Na+ with an optimum at 0.15–0.3 M Na+. The metabolism is fermentative and strictly anaerobic. Cellulose, cellobiose, and xylan can be used as growth substrates. Plant and algal debris can be fermented. Lactate, ethanol, acetate, hydrogen, and traces of formate are produced during cellulose or cellobiose fermentation. Yeast extract or vitamins are required for anabolic purposes. The microorganism fixes dinitrogen and is nitrogenase-positive. It is tolerant to up to 48 mM Na2S. Growth is not inhibited by kanamycin or neomycin. Chloramphenicol, streptomycin, penicillin, ampicillin, ampiox, bacillin, novobiocin, and bacitracin suppress growth. The DNA G+C content is 29.9 mol %. According to the nucleotide sequence of its 16S rRNA gene, strain Z-7026 is phylogenetically close to the neutrophilic cellulolytic bacteria Clostridium thermocellum (95.5%), C. aldrichii (94.9%), and Acetivibrio cellulolyticus (94.8%). It is proposed as a new species: Clostridium alkalicellum sp. nov.

Thermanaerovibrio velox sp. nov., a new anaerobic, thermophilic, organotrophic bacterium that reduces elemental sulfur, and emended description of the genus Thermanaerovibrio

A moderately thermophilic, organotrophic bacterium with vibrioid cells was isolated from a sample of a cyanobacterial mat from caldera Uzon, Kamchatka, Russia, and designated strain Z-9701T. Cells of strain Z-9701T were curved, Gram-negative rods, 0.5-0.7 x 2.5-5.0 microm in size, with tapering ends and with fast, wavy movement by means of lateral flagella located on the concave side of the cell. Colonies were small, white, irregular or round, 0.2 mm in diameter, and with even edges. Strain Z-9701T was an obligate anaerobe with a temperature optimum at 60-65 degrees C and a pH optimum at 7.3. It fermented glucose, fructose, mannose, N-acetyl-D-glucosamine, adonite, arginine, serine, peptone, yeast extract and Casamino acids. The fermentation products formed during growth on glucose were acetate, lactate, H2, CO2 and ethanol. Strain Z-9701T reduced elemental sulfur to H2S during organotrophic growth with glucose or peptides as energy and carbon sources. In the presence of S0, strain Z-9701T was capable of lithotrophic growth with molecular hydrogen as energy substrate and 0.1 g yeast extract l(-1) as carbon source. Sulfate, thiosulfate, nitrate, Fe(III) and sulfite were not reduced and did not stimulate growth. The G+C content of strain Z-9701T DNA was 54.6 mol%. The results of 16S rDNA sequence analyses revealed that strain Z-9701T belongs to the cluster within the Clostridium group formed by Thermanaerovibrio acidaminovorans, Dethiosulfovibrio peptidovorans, Anaerobaculum thermoterrenum and Aminobacterium colombiense, but the level of sequence similarity with the members of this cluster was not very high (87.6-92.2%). Among these organisms, Thermanaerovibrio acidaminovorans is phenotypically close to strain Z-9701T. However, the two organisms showed a relatively low level of similarity of their 16S rRNA sequences (92.2%) and of DNA-DNA hybridization (15 +/- 1%). Nevertheless, on the basis of the similar morphology and physiology of the new isolate and Thermanaerovibrio acidaminovorans, strain Z-9701T was placed in the genus Thermanaerovibrio and a new species, Thermanaerovibrio velox, proposed for it. The type strain is Z-9701T (= DSM 12556T).

Amphibacillus fermentum sp. nov. and Amphibacillus tropicussp. nov., New Alkaliphilic, Facultatively Anaerobic, Saccharolytic Bacilli from Lake Magadi

New alkaliphilic, saccharolytic, rod-shaped, gram-positive bacteria resistant to heating and drying and phylogenetically affiliated to the Bacilluslineage were isolated under strictly anaerobic conditions from sediments of the alkaline and highly mineralized Lake Magadi. Strain Z-7792 forms endospores; in strain Z-7984, endospore formation was not revealed. The strains are capable of both anaerobic growth (at the expense of fermentation of glucose and certain mono- and disaccharides with the formation of formate, ethanol, and acetate) and aerobic growth. Among polysaccharides, the strains hydrolyze starch, glycogen, and xylan. Yeast extract or methionine are required for growth. The strains are strict alkaliphiles exhibiting obligate requirement for Na+and carbonate ions, but not for Cl–ions. Growth occurs at a total mineralization as high as 3.3–3.6 M Na+, with an optimum at 1–1.7 M Na+. Strain Z-7792 is an obligate alkaliphile with a pH growth range of 8.5–11.5 and an optimum of 9.5–9.7. Strain Z-7984 grows in a pH range of 7.0–10.5 with an optimum at 8.0–9.5. Both strains are mesophiles having a growth optimum at 37–38°C. The G+C contents of the DNA of strains Z-7792 and Z-7984 are 39.2 and 41.5 mol %, respectively. These isolates of facultatively anaerobic, strictly alkaliphilic, Na+-dependent bacilli can be considered representatives of the ecological group adapted to life at drying-up shoals of soda lakes. Because of their independence of NaCl and lack of obligate dependence on sodium carbonates, the isolates are to be assigned to athalassophilic organisms. According to their physiological and phylogenetic characteristics, they taxonomically belong to group 1 of the species of bacilli with a low G+C content and occupy a position intermediate between the genera Amphibacillusand Gracilibacillus.The isolates are described as new species of Amphibacillus: A. fermentum(type strain, Z-7984T) and A. tropicus(type strain, Z-7792T).

["Candidatus contubernalis alkalaceticum," an obligately syntrophic alkaliphilic bacterium capable of anaerobic acetate oxidation in a coculture with Desulfonatronum cooperativum].

From the silty sediments of the Khadyn soda lake (Tuva), a binary sulfidogenic bacterial association capable of syntrophic acetate oxidation at pH 10.0 was isolated. An obligately syntrophic, gram-positive, spore-forming alkaliphilic rod-shaped bacterium performs acetate oxidation in a syntrophic association with a hydrogenotrophic, alkaliphilic sulfate-reducing bacterium; the latter organism was previously isolated and characterized as the new species Desulfonatronum cooperativum. Other sulfate-reducing bacteria of the genera Desulfonatronum and Desulfonatronovibrio can also act as the hydrogenotrophic partner. Apart from acetate, the syntrophic culture can oxidize ethanol, propanol, isopropanol, serine, fructose, and isobutyric acid. Selective amplification of 16S rRNA gene fragments of the acetate-utilizing syntrophic component of the binary culture was performed; it was found to cluster with clones of uncultured gram-positive bacteria within the family Syntrophomonadaceae. The acetate-oxidizing bacterium is thus the first representative of this cluster obtained in a laboratory culture. Based on its phylogenetic position, the new acetate-oxidizing syntrophic bacterium is proposed in the Candidatus status for a new genus and species: “Candidatus Contubernalis alkalaceticum.”

Alkaliphilus peptidofermentans sp. nov., a New Alkaliphilic Bacterial Soda Lake Isolate Capable of Peptide Fermentation and Fe(III) Reduction

A novel strain, Z-7036, of anaerobic spore-forming bacteria was isolated from a cellulolytic consortium obtained from the bottom sediments of the low-mineralization soda lake Verkhnee Beloe (Buryatia). The cells of the new strain are short motile gram-positive rods, 1.1–3.0 × 0.25–0.4 μm. The organism is an aerotolerant anaerobe and obligate alkaliphile growing within the pH range of 7.5–9.7 with an optimum at pH 9.1. The strain is mesophilic and halotolerant and grows at NaCl concentrations from 0 to 50 g/l with an optimum at 20 g/l. Carbonates are required. The microorganism ferments peptone, yeast extract, trypticase, tryptone, Bacto Soytone, meat extract, Casamino acids, ornithine, arginine, threonine, and tryptophan. The strain hydrolyzes the bacterial preparations “Gaprin” and “Spirulina”. Acetate and formate are the major fermentation products. The strain reduces amorphous ferric hydroxide (AFH), EDTA-Fe(III), anthraquinone-2,6-disulfonate (quinone), S2O32−, fumarate, and crotonate. Major fatty acids are C16:0, C16:1ω7c, iso-C17, iso-C15, and iso-C17:1. The DNA G+C content is 33.8 ± 0.5 mol %. According to the results of the 16S rRNA gene analysis, strain Z-7036 belongs to the genus Alkaliphilus within the cluster XI of low G+C gram-positive bacteria of the family Clostridiaceae. The novel strain is closely related to A. transvaalensis SAGM1T and A. crotonatoxidans B11-2T (93.3 and 93.9% 16S rRNA sequence identities, respectively). On the basis of the existing genotypic and phenotypic differences, we propose that strain Z-7036 should be classified as a novel species Alkaliphilus peptidofermentans sp. nov.

Description of Anaerobacillus alkalilacustre gen. nov., sp. nov.—Strictly anaerobic diazotrophic bacillus isolated from soda lake and transfer of Bacillus arseniciselenatis, Bacillus macyae, and Bacillus alkalidiazotrophicus to Anaerobacillus as the new combinations A. arseniciselenatis comb. nov., A. macyae comb. nov., and A. alkalidiazotrophicus comb. nov.

An anaerobic, spore-forming bacterium (strain Z-0521) was isolated from the iron-reducing microbial community enriched from sample of bottom sediments from low-mineralized soda lake Khadyn, Tuva upper Yenisey region (Russia). Cells of strain Z-0521 are motile straight Gram-positive rods, 0.7–1.1 (µm in diameter and 3.0–7.0 µm length. It is a mesophilic halotolerante obligate alkaliphilic bacterium with a pH range for growth 8.5–10.7 (optimum at 9.6–9.7). Utilizes carbohydrates. Peptides, organic acids or alcohols are not utilized. In the presence of mannite strain Z-0521 reduces AQDS, arsenate, selenate and selenite. It is capable of N2 fixation and has nitrogenase gene nifH. The dominant cellular fatty acids are C16:0, C16:1w7c and Ca15. The G+C content in the DNA is 36.2 mol %. 16S rRNA gene sequencing identified strain Z-0521 as a member of rRNA group 6 of the genus Bacillus. Its closest relatives are B. alkalidiazotrophicus and B. macyae (98.3 and 98.1% sequence similarity). On the basis of physiological properties and genetic analysis, it is proposed that strain Z-0521T should be assigned to a new species of a new genus, Anaerobacillus alkalilacustre gen. nov., sp. nov. It is also proposed that Bacillus arseniciselenatis, Bacillus macyae and Bacillus alkalidiazotrophicus should be transferred to this new genus, with Anaerobacillus arseniciselenatis (formely Bacillus arseniciselenatis) as the type species.

Fuchsiella alkaliacetigena gen. nov., sp. nov., an alkaliphilic, lithoautotrophic homoacetogen from a soda lake.

The first alkaliphilic obligately anaerobic hydrogenotrophic homoacetogenic bacterium, strain Z-7100(T), was isolated from sediments of the soda-depositing soda lake Tanatar III (Altay, Russia). Cells were thin, flexible rods, motile, Gram-negative and spore-forming. The organism was an obligate alkaliphile, growing at pH 8.5 to 10.5, with optimum growth at pH 8.8-9.3, and it grew in soda brines containing 1.9-4.7 M total Na(+) (optimum at 2.8-3.3 M). It exhibited an obligate dependence upon sodium carbonate but not upon chloride ions with an NaCl range for growth of 0-14% (w/v) and an optimum at 7.0-8.5% (w/v). The isolate was mesophilic and grew at temperatures from 25 to 45 °C, with an optimum at 40 °C. An H(2)+CO(2) mixture, ethanol, pyruvate and lactate were utilized with the formation of acetate as the sole metabolic product. Carbohydrates and amino acids did not support growth. The isolate had a respiratory type of metabolism, reducing NO3(-), SeO(4)(2-) or anthraquinone-2,6-disulfonate (as electron acceptors with ethanol as an electron donor). It was able to grow chemolithotrophically on H(2)+CO(2) in medium supplemented with a vitamin solution only. The major cellular fatty acids were the saturated fatty acids anteiso-C(15), C(14:0) and C(16:0) and the aldehydes C(16), C(14) and anteiso-C(15). The DNA G+C content of the isolate was 32.0 mol%. 16S rRNA gene sequence analysis showed that strain Z-7100(T) is a member of the order Halanaerobiales and represents a new branch within the family Halobacteroidaceae, clustering with the type strain of Selenihalanaerobacter shriftii (92.9% gene sequence similarity). On the basis of its physiological characteristics and phylogenetic position, the isolate is considered to represent a novel species in a new genus within the family Halobacteroidaceae. The name Fuchsiella alkaliacetigena gen. nov., sp. nov. is proposed. The type strain of the type species is Z-7100(T) (=DSM 24880(T)=VKM B-2667(T)).