Antioxidants keep the potentially probiotic but highly oxygen-sensitive human gut bacterium Faecalibacterium prausnitzii alive at ambient air.

The beneficial human gut microbe Faecalibacterium prausnitzii is a 'probiotic of the future' since it produces high amounts of butyrate and anti-inflammatory compounds. However, this bacterium is highly oxygen-senstive, making it notoriously difficult to cultivate and preserve. This has so far precluded its clinical application in the treatment of patients with inflammatory bowel diseases. The present studies were therefore aimed at developing a strategy to keep F. prausnitzii alive at ambient air. Our previous research showed that F. prausnitzii can survive in moderately oxygenized environments like the gut mucosa by transfer of electrons to oxygen. For this purpose, the bacterium exploits extracellular antioxidants, such as riboflavin and cysteine, that are abundantly present in the gut. We therefore tested to what extent these antioxidants can sustain the viability of F. prausnitzii at ambient air. The present results show that cysteine can facilitate the survival of F. prausnitzii upon exposure to air, and that this effect is significantly enhanced the by addition of riboflavin and the cryoprotectant inulin. The highly oxygen-sensitive gut bacterium F. prausnitzii can be kept alive at ambient air for 24 h when formulated with the antioxidants cysteine and riboflavin plus the cryoprotectant inulin. Improved formulations were obtained by addition of the bulking agents corn starch and wheat bran. Our present findings pave the way towards the biomedical exploitation of F. prausnitzii in redox-based therapeutics for treatment of dysbiosis-related inflammatory disorders of the human gut.

Genome-guided analysis of physiological and morphological traits of the fermentative acetate oxidizer Thermacetogenium phaeum.

BACKGROUND: Thermacetogenium phaeum is a thermophilic strictly anaerobic bacterium oxidizing acetate to CO(2) in syntrophic association with a methanogenic partner. It can also grow in pure culture, e.g., by fermentation of methanol to acetate. The key enzymes of homoacetate fermentation (Wood-Ljungdahl pathway) are used both in acetate oxidation and acetate formation. The obvious reversibility of this pathway in this organism is of specific interest since syntrophic acetate oxidation operates close to the energetic limitations of microbial life. RESULTS: The genome of Th. phaeum is organized on a single circular chromosome and has a total size of 2,939,057 bp. It comprises 3.215 open reading frames of which 75% could be assigned to a gene function. The G+C content is 53.88 mol%. Many CRISPR sequences were found, indicating heavy phage attack in the past. A complete gene set for a phage was found in the genome, and indications of phage action could also be observed in culture. The genome contained all genes required for CO(2) reduction through the Wood-Ljungdahl pathway, including two formyl tetrahydrofolate ligases, three carbon monoxide dehydrogenases, one formate hydrogenlyase complex, three further formate dehydrogenases, and three further hydrogenases. The bacterium contains a menaquinone MQ-7. No indications of cytochromes or Rnf complexes could be found in the genome. CONCLUSIONS: The information obtained from the genome sequence indicates that Th. phaeum differs basically from the three homoacetogenic bacteria sequenced so far, i.e., the sodium ion-dependent Acetobacterium woodii, the ethanol-producing Clostridium ljungdahlii, and the cytochrome-containing Moorella thermoacetica. The specific enzyme outfit of Th. phaeum obviously allows ATP formation both in acetate formation and acetate oxidation.

Amino acid-assimilating phototrophic heliobacteria from soda lake environments: Heliorestis acidaminivorans sp. nov. and 'Candidatus Heliomonas lunata'.

Two novel taxa of heliobacteria, Heliorestis acidaminivorans sp. nov. strain HR10B(T) and 'Candidatus Heliomonas lunata' strain SLH, were cultured from shoreline sediments/soil of Lake El Hamra (Egypt) and lake water/benthic sediments of Soap Lake (USA), respectively; both are highly alkaline soda lakes. Cells of strain HR10B were straight rods, while cells of strain SLH were curved rods. Both organisms were obligate anaerobes, produced bacteriochlorophyll g, and lacked intracytoplasmic photosynthetic membrane systems. Although the absorption spectrum of strain HR10B was typical of other heliobacteria, that of strain SLH showed unusually strong absorbance of the OH-chlorophyll a component. Major carotenoids of both organisms were OH-diaponeurosporene glucosyl esters, as in other alkaliphilic heliobacteria, and both displayed an alkaliphilic and mesophilic phenotype. Strain HR10B was remarkable among heliobacteria in its capacity to photoassimilate a number of carbon sources, including several amino acids. Nitrogenase activity was observed in strain HR10B, but not in strain SLH. The 16S ribosomal RNA gene tree placed strain HR10B within the genus Heliorestis, but distinct from other described species. By contrast, strain SLH was phylogenetically more closely related to neutrophilic heliobacteria and is the first alkaliphilic heliobacterium known outside of the genus Heliorestis.

Psychrobacillus gen. nov. and proposal for reclassification of Bacillus insolitus Larkin & Stokes, 1967, B. psychrotolerans Abd-El Rahman et al., 2002 and B. psychrodurans Abd-El Rahman et al., 2002 as Psychrobacillus insolitus comb. nov., Psychrobacillus psychrotolerans comb. nov. and Psychrobacillus psychrodurans comb. nov.

The taxonomic status of three Bacillus species, Bacillus insolitus, B. psychrodurans and B. psychrotolerans was reexamined using a polyphasic approach. In our analysis, these three Bacillus species formed a cluster separate from other members of Bacillus rRNA group 2 [5] and from Bacillus sensu stricto. These three species shared high 16S rRNA gene sequence similarities between them (97.8-99.7%) and showed closest sequence similarity (95.3-96.3%) to Paenisporosarcina quisquiliarum gen. nov., sp. nov. [18]. Sequence similarities with other related genera ranged between 90.9% and 94.5%. Phylogenetic coherence of the three species was supported by phenotypic characteristics, such as growth at low temperatures, negative oxidation and assimilation of many carbohydrates, MK8 as the major isoprenoid quinine and broadly similar polar lipid profiles. All three species had a similar peptidoglycan type of the variation A4ß and similar genomic G+C contents (35.7-36.6 mol% [1]). Genomic relatedness among them was shown to be less than 70% and justified their separate species status [1]. These three species could be differentiated from each other and from related taxa on the basis of phenotypic, including chemotaxonomic, characteristics and ribotype patterns. On the basis of our analysis, we propose a new genus Psychrobacillus gen. nov. and to transfer B. insolitus, B. psychrodurans and B. psychrotolerans to the new genus as Psychrobacillus insolitus comb. nov. (type species of the genus; type strain W16B(T)=DSM 5(T)), P. psychrodurans comb. nov. (type strain 68E3(T)=DSM 11713(T)) and P. psychrotolerans comb. nov. (type strain 3H1(T)=DSM 11706(T)).

Tuberibacillus calidus gen. nov., sp. nov., isolated from a compost pile and reclassification of Bacillus naganoensis Tomimura et al. 1990 as Pullulanibacillus naganoensis gen. nov., comb. nov. and Bacillus laevolacticus Andersch et al. 1994 as Sporolactobacillus laevolacticus comb. nov.

Two thermophilic strains, designated 607T and 606b, were isolated from a compost pile in Japan. The novel strains were Gram-positive, aerobic, spore-forming rods. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strains 607T and 606b were closely related to Bacillus naganoensis (94.0-94.1% similarity) and separated from clusters of the related genera Bacillus (<91.9%) and Sporolactobacillus (91.0-92.5%). In addition, some chemotaxonomic and physiological characteristics of strains 607T and 606b differed from those of B. naganoensis and the two related genera. Several differences in physiological characteristics and 16S-23S rRNA gene internal transcribed spacer region nucleotide sequences were observed between strains 607T and 606b; however, DNA-DNA hybridization indicated that these two strains belonged to the same species. From these results, it is proposed that strains 607T and 606b represent the type species of a new genus, Tuberibacillus calidus gen. nov., sp. nov., with strain 607T (=JCM 13397T=DSM 17572T) as the type strain. In addition, the results of phylogenetic analyses, as well as chemotaxonomic and physiological characterization, indicated that B. naganoensis and Bacillus laevolacticus did not belong to the genus Bacillus. Based on these results, it is proposed that B. naganoensis and B. laevolacticus should be transferred to Pullulanibacillus naganoensis gen. nov., comb. nov. and Sporolactobacillus laevolacticus comb. nov., respectively.

Paenibacillus xinjiangensis sp. nov., isolated from Xinjiang province in China.

Strain B538T is a Gram-positive, motile, rod-shaped bacterium, which was isolated from Xinjiang province in China. This organism grew optimally at 30-35 degrees C and pH 8.0-8.5. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain B538T belonged to the genus Paenibacillus and chemotaxonomic data (DNA G+C content, 47.0 mol%; major isoprenoid quinone, MK-7; cell wall type, A1gamma meso-diaminopimelic acid; major fatty acids, anteiso-C15:0 and C16:0) supported affiliation of the isolate with the genus Paenibacillus. Comparative 16S rRNA gene sequence analyses showed that the isolate was most closely related to Paenibacillus glycanilyticus DS-1T, with 16S rRNA gene sequence similarity of 98.1%; sequence similarities to other members of the genus Paenibacillus used in the phylogenetic tree were less than 96.5%. The DNA-DNA relatedness between strain B538T and P. glycanilyticus DS-1T was about 8.0%. On the basis of physiological and molecular properties, strain B538T (=KCTC 3952T=DSM 16970T) is proposed as the type strain of a novel species within the genus Paenibacillus, for which the name Paenibacillus xinjiangensis sp. nov. is proposed.

Anaerovirgula multivorans gen. nov., sp. nov., a novel spore-forming, alkaliphilic anaerobe isolated from Owens Lake, California, USA.

A novel, alkaliphilic, obligately anaerobic bacterium, strain SCAT, was isolated from mud sediments of a soda lake in California, USA. The rod-shaped cells were motile, Gram-positive, formed spores and were 0.4-0.5x2.5-5.0 microm in size. Growth occurred within the pH range 6.7-10.0 and was optimal at pH 8.5. The temperature range for growth was 10-45 degrees C, with optimal growth at 35 degrees C. NaCl was required for growth. Growth occurred at 0.5-9.0% (w/v) NaCl and was optimal at 1-2% (w/v). The novel isolate was a catalase-negative chemo-organoheterotroph that fermented sugars, proteolysis products, some organic and amino acids, glycerol, d-cellobiose and cellulose. It was also capable of growth by the Stickland reaction. Strain SCAT was sensitive to tetracycline, chloramphenicol, rifampicin and gentamicin, but it was resistant to ampicillin and kanamycin. The G+C content of the genomic DNA was 34.2 mol%. Major fatty acid components were C14:0, iso-C15:0, C16:1omega9c and C16:0. 16S rRNA gene sequence analysis of strain SCAT showed a similarity of approximately 97% with the type strains of Clostridium formicaceticum and Clostridium aceticum in clostridial cluster XI and a similarity of less than 94.2% to any other recognized Clostridium species and those of related genera in this cluster. Strain SCAT was clearly differentiated from C. formicaceticum and C. aceticum based on comparison of their phenotypic properties and fatty acid profiles, as well as low levels of DNA-DNA relatedness between strain SCAT and the type strains of these two species. Therefore, strain SCAT is considered to represent a novel species of a new genus, Anaerovirgula multivorans gen. nov., sp. nov., in clostridial cluster XI. The type strain is SCAT (=ATCC BAA-1084T=JCM 12857T=DSM 17722T=CIP 107910T).

Sulfobacillus thermotolerans sp. nov., a thermotolerant, chemolithotrophic bacterium.

A thermotolerant, Gram-positive, aerobic, endospore-forming, acidophilic bacterium (strain Kr1T) was isolated from the pulp of a gold-containing sulfide concentrate processed at 40 degrees C in a gold-recovery plant (Siberia). Cells of strain Kr1(T) were straight to slightly curved rods, 0.8-1.2 microm in diameter and 1.5-4.5 microm in length. Strain Kr1T formed spherical and oval, refractile, subterminally located endospores. The temperature range for growth was 20-60 degrees C, with an optimum at 40 degrees C. The pH range for growth on medium containing ferrous iron was 1.2-2.4, with an optimum at pH 2.0; the pH range for growth on medium containing S0 was 2.0-5.0, with an optimum at pH 2.5. Strain Kr1T was mixotrophic, oxidizing ferrous iron, S0, tetrathionate or sulfide minerals as energy sources in the presence of 0.02 % yeast extract or other organic substrates. The G+C content of the DNA of strain Kr1T was 48.2+/-0.5 mol%. Strain Kr1T showed a low level of DNA-DNA reassociation with the known Sulfobacillus species (11-44 %). 16S rRNA gene sequence analysis revealed that Kr1T formed a separate phylogenetic group with a high degree of similarity between the nucleotide sequences (98.3-99.6 %) and 100 % bootstrap support within the phylogenetic Sulfobacillus cluster. On the basis of its physiological properties and the results of phylogenetic analyses, strain Kr1T can be affiliated to a novel species of the genus Sulfobacillus, for which the name Sulfobacillus thermotolerans sp. nov. is proposed. The type strain is Kr1T (=VKM B-2339T=DSM 17362T).

Sporacetigenium mesophilum gen. nov., sp. nov., isolated from an anaerobic digester treating municipal solid waste and sewage.

Two mesophilic, anaerobic bacterial strains (ZLJ115T and L4-2) were isolated from the sludge of an anaerobic digester treating municipal solid waste and sewage in Fujian province, China. The strains were Gram-positive, spore-forming, motile rods (0.9-1.0 x 3.6-7.3 microm). Growth of the strains was observed at 20-42 degrees C and pH 6.0-9.5. Both strains fermented several mono- and disaccharides. The main fermentation products from glucose were acetate, ethanol, hydrogen and carbon dioxide. Optimal hydrogen production by the new isolates was observed at pH 8.8 and 39 degrees C, and 1.4 mol H2 was detected from fermentation of 1 mol glucose. The DNA G+C contents of strains ZLJ115T and L4-2 were 53.9 and 54.3 mol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the isolates represented a novel phyletic sublineage within cluster XI of the clostridia, clustering with four thermophilic species, with <93.8 % 16S rRNA gene sequence similarity to previously described species. Phenotypically, the new isolates were distinguished from their phylogenetic relatives by growing mesophilically and by fermenting a variety of pentoses, as well as their higher genome DNA G+C content. On the basis of polyphasic evidence from this study, a novel genus and species are proposed, Sporacetigenium mesophilum gen. nov., sp. nov.; strain ZLJ115T (=DSM 16796T = AS 1.5019T) is the type strain of Sporacetigenium mesophilum.

Syntrophomonas erecta subsp. sporosyntropha subsp. nov., a spore-forming bacterium that degrades short chain fatty acids in co-culture with methanogens.

Two obligate anaerobic bacterial strains (5-3-Z(T) and Y4-1) were isolated from river sediment and rice field mud, respectively. They degraded straight-chain fatty acids with 4-8 carbon atoms in syntrophic association with methanogens, however, neither tested branch-chain fatty acids nor could benzoate be degraded. The strains formed spores when cocultured with methanogens on butyrate, or when grew on butyrate plus dimethyl sulfoxide (DMSO) in pure culture. The cells were slightly curved rods with Gram-negative cell wall structure, and contained small amount of poly beta-hydroxyalkanoate. The strains could not degrade butyrate alone, nor could use fumarate, sulfate, thiosulfate, sulfur or nitrate as electron acceptors except DMSO for butyrate degradation. The generation time of strain 5-3-Z(T) was about 12h when growing on crotonate at 37 degrees C. The growth of the new strains occurred in the range of pH 5.5-8.4, and of temperature 20-48 degrees C, and at NaCl concentration of 0-700 mM. The G+C content of the genomic DNA of strain 5-3-Z(T) was 40.6mol%. Phylogenetic analysis based on 16S rRNA gene similarity showed the two strains to be a member of species Syntrophomonas erecta (98.4-98.9% sequence similarity), however they differed from the existing strains in both phenotypic and genetic characteristics. Therefore, a new subspecies of S. erecta, S. erecta subsp. sporosyntropha was proposed. The type strain was 5-3-Z(T) (=CGMCC1.5032(T)=JCM13344(T)).

Paenibacillus assamensis sp. nov., a novel bacterium isolated from a warm spring in Assam, India.

A polyphasic approach was used to characterize a bacterium, GPTSA 11(T), isolated from a warm spring located in a reserve forest in Assam, India. The cells are Gram-variable, strictly aerobic, sporulating motile rods. The major fatty acids of the strain are C(15 : 0) anteiso (48.42 %), C(16 : 0) iso (11.59 %), C(16 : 1)omega11c (6.16 %), C(15 : 0) iso (6.03 %), C(17 : 0) anteiso (5.68 %) and C(16 : 1)omega7c alcohol (5.01 %). The presence of the fatty acid C(16 : 1)omega7c alcohol distinguishes this strain from other closely related species of the genus Paenibacillus. The strain contains MK-7 as the diagnostic menaquinone. The G+C content of the genomic DNA is 41.2 mol%. Analysis of the 16S rRNA gene sequence (1466 nt) revealed the presence of signature sequences PAEN 515F (5'-GAGTAACTGCTCTCGGAATGACGGTACTTGAGAAGAAAGCCCC-3') and PAEN 862F (5'-TCGATACCCTTGGTGCCGAAGT-3'), which were found in the species of the genus Paenibacillus surveyed by Shida et al. [Shida, O., Takagi, H., Kadowaki, K., Nakamura, L. K. & Komagata, K. (1997). Int J Syst Bacteriol 47, 289-298]. The sequence shows closest similarity (95.85 %) to that of Paenibacillus apiarius, followed by Paenibacillus alvei (94.34 %), Paenibacillus cineris (93.87 %), Paenibacillus favisporus (93.80 %), Paenibacillus chibensis (93.47 %) and Paenibacillus azoreducens (93.40 %). Biochemical, physiological, chemotaxonomic and phylogenetic analyses justify placement of the strain in the genus Paenibacillus but not within any existing species. It should, therefore, be considered as representing a novel species, for which the name Paenibacillus assamensis sp. nov. is proposed. The type strain is GPTSA 11T (=MTCC 6934T=JCM 13186T).

Brevibacillus levickii sp. nov. and Aneurinibacillus terranovensis sp. nov., two novel thermoacidophiles isolated from geothermal soils of northern Victoria Land, Antarctica.

Thirteen strains of endospore-forming bacteria were isolated from geothermal soils at Cryptogam Ridge, the north-west slope of Mt Melbourne, and at the vents and summit of Mt Rittmann in northern Victoria Land, Antarctica. 16S rRNA gene sequencing, SDS-PAGE and routine phenotypic characterization tests indicated that the seven isolates from the north-west slope of Mt Melbourne represent a novel species of Brevibacillus and that the six isolates from Cryptogam Ridge and the vents and summit of Mt Rittmann represent a novel species of Aneurinibacillus. Brevibacillus strains were not isolated from the sites at Mt Rittmann or Cryptogam Ridge and Aneurinibacillus strains were not isolated from the north-west slope of Mt Melbourne. Preliminary metabolic studies revealed that L-glutamic acid, although not essential for growth, was utilized by both species. The Brevibacillus species possessed an uptake system specific for L-glutamic acid, whereas the Aneurinibacillus species possessed a more general uptake system capable of transporting other related amino acids. Both species utilized a K(+) antiport system and similar energy systems for the uptake of l-glutamic acid. The rate of uptake by the Brevibacillus species type strain was 20-fold greater than that shown by the Aneurinibacillus species type strain. The names Brevibacillus levickii sp. nov. and Aneurinibacillus terranovensis sp. nov. are proposed for the novel taxa; the type strains are Logan B-1657(T) (= LMG 22481(T) = CIP 108307(T)) and Logan B-1599(T) (LMG 22483(T) = CIP 108308(T)), respectively.

Paenibacillus rhizosphaerae sp. nov., isolated from the rhizosphere of Cicer arietinum.

Two sporulating bacterial strains designated CECAP06(T) and CECAP16 were isolated from the rhizosphere of the legume Cicer arietinum in Argentina. Almost-complete 16S rRNA gene sequences identified the isolates as a Paenibacillus species. It was most closely related to Paenibacillus cineris LMG 18439(T) (99.6 % sequence similarity), Paenibacillus favisporus LMG 20987(T) (99.4 % sequence similarity) and Paenibacillus azoreducens DSM 13822(T) (97.7 % sequence similarity). The cells of this novel species were motile, sporulating, rod-shaped, Gram-positive and strictly aerobic. The predominant fatty acids were anteiso-C(15 : 0), C(16 : 0) and iso-C(16 : 0). The DNA G+C content of strains CECAP06(T) and CECAP16 was 51.3 and 50.9 mol%, respectively. Growth was observed from many carbohydrates, but gas production was not observed from glucose. Catalase and oxidase activities were present. The isolates produced beta-galactosidase and hydrolysed aesculin. Gelatinase, caseinase and urease were not produced. The results of DNA-DNA hybridization showed that the strains from this study constitute a novel species of the genus Paenibacillus, for which the name Paenibacillus rhizosphaerae sp. nov. is proposed. The type strain is CECAP06(T) (=LMG 21955(T) = CECT 5831(T)).

Mahella australiensis gen. nov., sp. nov., a moderately thermophilic anaerobic bacterium isolated from an Australian oil well.

A novel Gram-positive, anaerobic and moderately thermophilic bacterium, strain 50-1 BON(T), was isolated from an Australian terrestrial oil reservoir. Cells were spore-forming straight rods, motile by peritrichous flagella. The optimum growth conditions were 50 degrees C, pH 7.5 and 0.1 % NaCl. Strain 50-1 BON(T) fermented arabinose, cellobiose, fructose, galactose, glucose, mannose, sucrose, xylose and yeast extract. Glucose was fermented mainly into lactate, formate, hydrogen and CO(2). The major end product of pyruvate fermentation was acetate together with H(2) and CO(2). Thiosulfate, sulfate, elemental sulfur and nitrate were not used as terminal electron acceptors. The DNA G+C content was 55.5 mol%. The closest phylogenetic relative of strain 50-1 BON(T) was Thermoanaerobacterium thermosulfurigenes (16S rRNA gene sequence similarity of 85.7 %). As strain 50-1 BON(T) was physiologically and phylogenetically different from members of the order 'Thermoanaerobacteriales', it is proposed that strain 50-1 BON(T) (=DSM 15567(T)=CIP 107919(T)) be classified as the type strain of a novel species of a new genus, Mahella australiensis gen. nov., sp. nov.

Tindallia californiensis sp. nov., a new anaerobic, haloalkaliphilic, spore-forming acetogen isolated from Mono Lake in California.

A novel extremely haloalkaliphilic, strictly anaerobic, acetogenic bacterium strain APO was isolated from sediments of the athalassic, meromictic, alkaline Mono Lake in California. The Gram-positive, spore-forming, slightly curved rods with sizes 0.55-0.7x1.7-3.0 microm were motile by a single laterally attached flagellum. Strain APO was mesophilic (range 10-48 degrees C, optimum of 37 degrees C); halophilic (NaCl range 1-20% (w/v) with optimum of 3-5% (w/v), and alkaliphilic (pH range 8.0-10.5, optimum 9.5). The novel isolate required sodium ions in the medium. Strain APO was an organotroph with a fermentative type of metabolism and used the substrates peptone, bacto-tryptone, casamino acid, yeast extract, l-serine, l-lysine, l-histidine, l-arginine, and pyruvate. The new isolate performed the Stickland reaction with the following amino acid pairs: proline + alanine, glycine + alanine, and tryptophan + valine. The main end product of growth was acetate. High activity of CO dehydrogenase and hydrogenase indicated the presence of a homoacetogenic, non-cycling acetyl-CoA pathway. Strain APO was resistant to kanamycin but sensitive to chloramphenicol, tetracycline, and gentamycin. The G+C content of the genomic DNA was 44.4 mol% (by HPLC method). The sequence of the 16S rRNA gene of strain APO possessed 98.2% similarity with the sequence from Tindallia magadiensis Z-7934, but the DNA-DNA hybridization value between these organisms was only 55%. On the basis of these physiological and molecular properties, strain APO is proposed to be a novel species of the genus Tindallia with the name Tindallia californiensis sp. nov., (type strain APO = ATCC BAA-393 = DSM 14871).

Desulfotomaculum alkaliphilum sp. nov., a new alkaliphilic, moderately thermophilic, sulfate-reducing bacterium.

A new moderately thermophilic, alkaliphilic, sulfate-reducing, chemolithoheterotrophic bacterium, strain S1T, was isolated from a mixed cow/pig manure with neutral pH. The bacterium is an obligately anaerobic, non-motile, Gram-positive, spore-forming curved rod growing within a pH range of 8.0-9.15 (optimal growth at pH 8.6-8.7) and temperature range of 30-58 degrees C (optimal growth at 50-55 degrees C). The optimum NaCl concentration for growth is 0.1%. Strain S1T is an obligately carbonate-dependent alkaliphile. The G+C content of the DNA is 40.9 mol%. A limited number of compounds are utilized as electron donors, including H2+acetate, formate, ethanol, lactate and pyruvate. Sulfate, sulfite and thiosulfate, but not sulfur or nitrate, can be used as electron acceptors. Strain S1T is able to utilize acetate or yeast extract as sources of carbon. Analysis of the 16S rDNA sequence allowed strain S1T (= DSM 12257T) to be classified as a representative of a new species of the genus Desulfotomaculum, Desulfotomaculum alkaliphilum sp. nov.

Saccharococcus caldoxylosilyticus sp. nov., an obligately thermophilic, xylose-utilizing, endospore-forming bacterium.

Several closely related, xylanolytic, thermophilic bacilli were isolated from local soils on xylose-containing minimal medium. On the basis of morphology and biochemical characteristics, one of the isolates, designated strain S1812T (T = type strain), was studied further. Strain S1812T was a xylanolytic, sporulating, Gram-positive, rod-shaped bacterium. Its Gram-positive nature was confirmed by electron microscopic examination of thin sections of the cells. The isolate was a thermophilic (optimum temperature for growth, 65 degrees C), facultative anaerobe that grew on a wide range of carbon sources including glucose, lactose, starch and xylose. It expressed high levels of both xylose isomerase and xylulokinase on xylose and also on glucose. The DNA G + C content was 44 mol%. rRNA gene sequence analysis placed strain S1812T in Bacillus cluster 5; it was more closely related to Saccharococcus thermophilus than to thermophilic Bacillus species. DNA-DNA hybridization also indicated its close relationship to S. thermophilus. Based on the evidence presented, it is proposed that strain S1812T be designated Saccharococcus caldoxylosilyticus sp. nov. Strain S1812T is the type strain (= ATCC 700356T = DSM 97-987T).

Sporobacterium olearium gen. nov., sp. nov., a new methanethiol-producing bacterium that degrades aromatic compounds, isolated from an olive mill wastewater treatment digester.

A strictly chemo-organotrophic, anaerobic bacterium was isolated from an olive mill wastewater treatment digester on syringate and designated strain SR1T. The cells were slightly curved rods, stained Gram-positive and possessed terminal spores. Strain SR1T utilized crotonate, methanol and a wide range of aromatic compounds including 3,4,5-trimethoxybenzoate (TMB), 3,4,5-trimethoxycinnamate (TMC), syringate, 3,4,5-trimethoxyphenylacetate (TMPA), 3,4,5-trimethoxyphenylpropionate (TMPP), ferulate, sinapate, vanillate, 3,4-dimethoxybenzoate, 2,3-dimethoxybenzoate, gallate, 2,4,6-trihydroxybenzoate (THB), pyrogallol, phloroglucinol and quercetin as carbon and energy sources. Acetate and butyrate were produced from aromatic compounds, methanol and crotonate whereas methanethiol (MT) was produced from methoxylated aromatic compounds and methanol. Strain SR1T had a G + C content of 38 mol% and grew optimally between 37 and 40 degrees C at pH 7.2 on a crotonate-containing medium. Phylogenetically, strain SR1T was a member of cluster XIVa of the Clostridiales group and shared a sequence similarity of 90% with Clostridum aminovalericum and Eubacterium fissicatena. Consequently, its precise neighbourliness to any one of them depended on the selection of strains of the cluster. On the basis of the phylogenetic and phenotypic evidence presented in this paper, the designation of strain SR1T as Sporobacterium olearium gen. nov., sp. nov. is proposed. The type strain is SR1T (= DSM 12504T).

Phylogenetic status of Anaerobacter polyendosporus, an anaerobic, polysporogenic bacterium.

The almost complete sequence of the 16S rRNA gene of the Gram-positive polysporogenic bacterium Anaerobacter polyendosporus was determined. This allowed phylogenetic analysis of A. polyendosporus by comparing sequences of the 16S rRNA gene of this bacterium to similar genes of other Gram-positive bacteria. It was shown that this polysporogenic bacterium belongs to the Clostridium cluster I, subcluster A. Phylogenetically, A. polyendosporus is distantly related to another polysporogenic, but non-cultivatable, bacterium, 'Metabacterium polyspora' and can be satisfactorily clustered within the saccharolytic clostridia with a low DNA G+C content grouped in subcluster A. A. polyendosporus was most closely related to Clostridium intestinale (94.8% identity of 16S rRNA genes) and Clostridium fallax (93.1%). Like other members of the Clostridium cluster I, subcluster A, A. polyendosporus possesses such common phenotypic features as a Gram-positive cell wall structure, anaerobiosis, derivation of energy from carbohydrate fermentation yielding butyric acid among other organic acids and the capacity for endogenous spore-formation. However, the scale of evolutionary change in the 16S rRNA gene between A. polyendosporus and phylogenetically related Clostridium species does not correspond to the profound changes in the phenotype of A. polyendosporus. Distinctive phenotypic features of the latter are large cell size, polysporogenesis (up to seven spores per cell), alternative modes of development and an unusual membrane ultrastructure.