- Volume 52, Issue 4, 2002

A novel acid-tolerant, hydrogenotrophic methanogen, isolate ATMT, was obtained from an enrichment performed at pH 5.0 using slurry from an acidogenic digester running on alcohol distillery waste. The original pH of the slurry was 5.7 and the volatile fatty acid concentration was 9000 p.p.m. Cells of isolate ATMT were Gram-positive, non-motile and 0.3-0.5 microm in size. They did not form spores. The isolate could grow in the pH range 5.0-7.5, with maximum growth at pH 6.0. The optimum temperature for growth was 35 degrees C. Formate, acetate, methanol, trimethylamine, 2-propanol and 2-butanol were not utilized as growth substrates. Rumen fluid and acetate were required for growth on H2/CO2. Coenzyme M and 2-methylbutyric acid were not required in the presence of rumen fluid. 16S rDNA sequence analysis confirmed the signature sequence of the genus Methanobrevibacter. Morphological and biochemical characteristics of the isolate, together with the 16S rDNA sequence analysis, clearly revealed that the isolate could not be accommodated within any of the existing species of the genus Methanobrevibacter. Therefore, it is proposed that a novel species of the genus Methanobrevibacter should be created for this isolate, Methanobrevibacter acididurans sp. nov., and the type strain is

A novel thermophilic, methane-producing archaeon was isolated from a deep-sea hydrothermal vent chimney at the Iheya Ridge, in the Okinawa Trough, Japan. The cells were highly motile, irregular cocci, with a polar bundle of flagella. Growth was observed between 40 and 75 degrees C (optimum 60-65 degrees C; 30 min doubling time) and between pH 4.5 and 8.5 (optimum pH 6.7). The isolate was a strictly anaerobic autotroph capable of using hydrogen and carbon dioxide as sole sources of energy and carbon. Formate can serve as an alternative energy source. The G+C content of the genomic DNA was 33.5 mol%. Phylogenetic analysis based on 16S rDNA sequences and DNA-DNA hybridization analysis indicated that the isolate was closely related to members of the genera Methanococcus and Methanothermococcus. This isolate, however, could be differentiated from the previously described species of these genera on the basis of its physiological and molecular properties. The name Methanothermococcus okinawensis sp. nov is proposed, with the type strain IH1T (=JCM 11175T=DSM 14208T).

Seventeen strains of rod-shaped, heterotrophic, anaerobic, hyperthermophilic crenarchaeotes were isolated from several hot spring areas in eastern Japan, and eight representative strains were characterized further. Cells of these strains were straight to slightly curved rods, 0.4-0.6 microm in width. Occasionally, cells were branched or bore spherical bodies at the poles. They grew optimally at 85-90 degrees C and at pH 4.0-4.5. They utilized yeast extract, peptone, beef extract, Casamino acids, gelatin, starch, maltose and malate as carbon sources and sulfur and thiosulfate as possible electron acceptors. The DNA G+C contents of the novel isolates were 43.9-46.2 mol%. The lipids were mainly cyclic and acyclic tetraether core lipids. Phylogenetic analysis of the 16S rDNA sequences revealed that they represented an independent lineage in the family Thermoproteaceae. Moreover, comparison of the 16S rDNA sequences and a DNA-DNA hybridization study showed that they comprised two species, which could also be differentiated by the maximal growth temperature and degrees of NaCl tolerance. Therefore, a new genus, Vulcanisaeta gen. nov., in the family Thermoproteaceae is proposed to accommodate two novel species, Vulcanisaeta distributa sp. nov. and Vulcanisaeta souniana sp. nov. The type species is V. distributa and the type strains are V. distributa IC 017T (= JCM 11212T = DSM 14429T) and V. souniana IC-059T (= JCM 11219T = DSM 14430T).

Phylogenetic analysis of Promicromonospora enterophila indicates that this taxon clusters with Cellulomonas species, adjacent to Cellulomonas turbata (basonym Oerskovia turbata). 16S rDNA analysis, DNA-DNA reassociation, riboprinting, peptidoglycan analysis and determination of phenotypic properties of various strains of P. enterophila and C turbata reveal that they form a cluster that can be distinguished unambiguously from other Cellulomonas species by morphology, amino acid composition of the cell wall and 16S rDNA signatures. As a result of thispolyphasic study, it appears taxonomically reasonable to re-establish the genus Oerskovia for C turbata and to reclassify P. enterophila as Oerskovia enterophila comb. nov.; two novel species, Oerskovia jenensis sp. nov. (type strain DSM 46000T = CIP 100330T) and Oerskovia paurometabola sp. nov. (type strain DSM 14281T = LMG 20385T), are also proposed.

Analyses of 165 rRNA gene sequences, restriction endonuclease digestion fingerprints of 16S-23S intergenic regions, DNA base compositions, fatty-acid profiles, cell-wall chemistry, cell physiology and fermentation end-product composition, along with other biochemical and phenotypic properties, supported the view that Trichococcus flocculiformis EchtT (DSM 2094T), Lactosphaera pasteurii KoTa2T (DSM 2381T), Ruminococcus palustris Z-7189T (DSM 9172T) and an isolate named 'Carnococcus allantoicus' NDP were all very similar and should be merged into a single genus. Detailed characterization of strains Ben 77, Ben 200 and Ben 201 described previously as 'Nostocoida limicola' I, a filamentous bacterium which causes bulking in activated sludge systems, revealed that these strains also belonged to the same genus as T. flocculiformis EchtT, L. pasteurii KoTa2T, R. palustris Z-7189T and 'C allantoicus' NDP. In fact, their shared properties suggested that these strains all belonged to a single species. However, DNA-DNA hybridization data indicated that T. flocculiformis EchtT, all of the 'N. limicola' I isolates and 'C allantoicus' NDP belonged to the same species, whereas L. pasteurii KoTa2T, R. palustris Z-7189T and two new isolates, 37AN3*T and 45AN2, represented three distinct species within the same genus. The priority of the genus name Trichococcus is established and since its validation predates the description of the genus Lactosphaera this name should take precedence. Under certain culture conditions, all of the strains mentioned above could produce chains of cocci. Furthermore, the morphology of T. flocculiformis EchtT could change to a non-filamentous form on certain media. This study proposes that the above strains be reclassified as members of the genus Trichococcus as four species, namely Trichococcus flocculiformis emend. (type strain EchtT = DSM 2094T), Trichococcus pasteurii comb. nov. (type strain KoTa2T = DSM 2381T = ATCC 35945T), Trichococcus collinsii sp. nov. (type strain 37AN3*T = DSM 14526T = ATCC BAA-296T, and Trichococcus palustris comb. nov. (type strain Z-7189T = DSM 9172T).

Three glutamic-acid-producing coryneform strains were isolated from soil and vegetable samples. Chemotaxonomic investigations indicated that these strains belonged to the genus Corynebacterium. Phylogenetic studies, based on 16S rDNA analysis, demonstrated that the three strains formed a distinct cluster within the genus Corynebacterium and that their nearest relatives were Corynebacterium glutamicum and Corynebacterium callunae, also known as glutamic-acid-producing species. The data from 16S rDNA sequence and DNA-DNA relatedness studies clearly indicated that the three isolates represented a new species within the genus Corynebacterium. All of the isolates could grow at 45 degrees C and produced acid from dextrin; these were the most significant characteristics differentiating the three isolates from their neighbours. On the basis of the data presented here, it is proposed that the three glutamic-acid-producing isolates together be classified as Corynebacterium efficiens sp. nov., the type strain of which is YS-314T (= AJ 12310T = JCM 11189T = DSM 44549T).

A cis-1,4-polyisoprene-degrading bacterium (strain Kb2T) was isolated from foul water taken from the inside of a deteriorated automobile tyre found on a farmer's field in Westfalia, Germany. The strain was aerobic, Gram-positive, exhibited orange smooth and rough colonies on complex nutrient agar, produced elementary branching hyphae that fragmented into rod/coccus-like elements and showed chemotaxonomic markers which were consistent with its classification within the genus Gordonia, i.e. the presence of mesodiaminopimelic acid, arabinose and galactose in whole-cell hydrolysates (cell-wall chemotype IV), N-glycolylmuramic acid in the peptidoglycan wall, a fatty-acid pattern composed of unbranched saturated and monounsaturated fatty acids plus tuberculostearic acid, mycolic acids comprising 56-60 carbon atoms and MK-9(H2) as the only menaquinone. The 16S rDNA sequence of strain Kb2T was found to be most similar to the 16S rDNA sequences of the type strains of Gordonia alkanivorans (DSM 44369T) and Gordonia nitida (KCTC 0605BPT). However, DNA-DNA relatedness data showed that strain Kb2T ( =DSM 44215T NRRL B-24152T) could be distinguished from these two species and represented a new species within the genus Gordonia, for which the name Gordonia westfalica is proposed.

A novel filamentous Bacillus strain, NAF001T, was reported previously that produces endospores and spore-like resting cells; the latter outgrow by budding. Phylogenetic analysis based on 16S rDNA gene sequences reported in the same paper speculated on the proposal of a novel species for this isolate. This communication describes the DNA-DNA relatedness of strain NAF001T to various members of the genus Bacillus and its whole-cell fatty acid and quinone profiles, in order to authenticate the creation of a novel species, for which the name Bacillus funiculus sp. nov. is proposed. The type strain is NAF001T (= JCM 11201T = CIP 107128T). Further, features of the binding points between filaments of strain NAF001T that enable it to form extremely long filaments are captured by electron microscopy.

An aerobic and heterotrophic isolate, designated IFAM EL-30T, was obtained from hypersaline Ekho Lake (Vestfold Hills, East Antarctica). The isolate consisted of Gram-positive cocci or short rods which occasionally exhibited branching. The organism was moderately halotolerant, required thiamin.HCI and was stimulated by biotin and nicotinic acid. It grew well with glucose, acetate, pyruvate, succinate, malate or glutamate, and hydrolysed DNA but not gelatin, starch or Tween 80. Nitrate was aerobically reduced to nitrite. Chemical analysis revealed diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine and an unidentified glycolipid as the major polar lipids. The cellular fatty acids were predominantly of the anteiso and iso methyl-branched types, and the major menaquinone6 were MK-7 and MK-8. The peptidoglycan type was A4alpha, L-Lys-L-Glu. The DNA base ratio was 66.1 mol% G+C. Comparisons of 16S rRNA gene sequences showed that the unidentified organism was phylogenetically closely related to Nesterenkonia halobia, although a sequence divergence value of > 3% demonstrated that the organism represents a different species. On the basis of phenotypic and genotypic evidence, it is proposed that the unknown bacterium be designated as a new species of the genus Nesterenkonia, namely Nesterenkonia lacusekhoensis sp. nov., the type strain being IFAM EL-30T (= DSM 12544T = CIP 107030T). An emended description of the genus Nesterenkonia is given.

Four strains of a hitherto unknown bacterium isolated from vacuum-packaged refrigerated beef were characterized by using phenotypic and phylogenetic methods. The novel strains were Gram-positive, catalase-negative, psychrophilic, rod-shaped bacteria with lactic acid-homofermentative mechanism. Comparative 16S rDNA gene sequencing demonstrated that the unknown strains represent a novel subline within the genus Lactobacillus, close to but distinct from Lactobacillus curvatus and Lactobacillus sakei. The unknown strains were readily distinguished from all currently described members of the genus Lactobacillus by biochemical properties and SDS-PAGE whole-cell protein profiles. Based on phylogenetic and phenotypic evidence, it is proposed that the unknown bacterium be classified as Lactobacillus fuchuensis sp. nov. The type strain is strain B5M10T (= JCM 11249T = DSM 14340T).

An obligately anaerobic, mesophilic, cellulolytic bacterium, strain ISDgT, was isolated from forest soil. Cells of this isolate stained Gram-negative, despite possessing a Gram-positive cell-wall ultrastructure, and were motile, straight rods that formed spherical terminal spores that swelled the sporangium. Cellulose, pectin, polygalacturonic acid, starch, xylan, arabinose, cellobiose, fructose, galactose, gentiobiose, glucose, lactose, maltose, mannose, ribose and xylose supported growth. The major end products of fermentation were ethanol, acetate, CO2 and H2; formate and lactate were minor products. The optimum temperature for growth was 35-37 degrees C. Phylogenetic analyses based on 16S rRNA sequence comparisons showed that strain ISDgT was related to a group of anaerobes that included Clostridium herbivorans, Clostridium polysaccharolyticum and Clostridium populeti. The G+C content of this strain was 35.9 mol%. On the basis of numerous genotypic and phenotypic differences between strain ISDgT and its close relatives, strain ISDgT is proposed as a novel species in the genus Clostridium, for which the name Clostridium phytofermentans sp. nov. is proposed. The type strain is ISDgT (= ATCC 700394T).

Three isolates of an unknown Gram-positive, catalase-negative, chain-forming, coccus-shaped organism isolated from an outbreak of septicaemia in a flock of adult broiler parents were characterized by phenotypic and molecular taxonomic methods. Comparative 16S rRNA gene sequencing studies demonstrated that the bacterium represents a new subline within the genus Streptococcus, related to, albeit distinct from, Streptococcus acidominimus, Streptococcus ovis, Streptococcus suis and close relatives. The unknown bacterium was readily distinguished from all recognized streptococcal species by biochemical tests. Based on phylogenetic and phenotypic evidence, it is proposed that the unknown bacterium from chickens be classified as Streptococcus gallinaceus sp. nov. The type strain is CCUG 42692T (= CIP 107087T).

A lipophilic, coryneform bacterium isolated from a human clinical specimen was characterized by phenotypic and molecular-taxonomic methods. Chemotaxonomic investigations revealed the presence of cell-wall chemotype IV and short-chain mycolic acids consistent with the genus Corynebacterium. The isolate could be distinguished from other members of the genus Corynebacterium by positive urease and catalase tests as well as its failure to produce acid from carbohydrates. Comparative 16S rRNA gene sequencing showed that this isolate constitutes a distinct subline within the genus Corynebacterium, displaying >3.0% sequence divergence from other known Corynebacterium species. Based on both phenotypic and phylogenetic evidence, it is proposed that this isolate be classified as a novel species, Corynebacterium appendicis sp. nov., represented by strain IMMIB R-3491T (= DSM 44531T = NRRL B-24151T).

Four orange-pigmented strains from pond water (L1-L4) have been subjected to polyphasic taxonomic analyses. On the basis of ribotype analysis and Fourier-transform infrared spectroscopy, these strains form a genomically highly related group. 16S rDNA sequence analysis revealed 98.8% similarity between the 16S rDNA sequences of strains L2T and H2T, isolated previously from a microbial mat from Lake Fryxell, Antarctica. DNA-DNA reassociation values indicated the presence of two genomic clusters. While the DNA of strains L2T and L3 showed 100% DNA relatedness, strains L2T and H2T shared only 51% DNA relatedness. These two clusters differed in some phenotypic properties, e.g. utilization of melibiose, D-mannitol, adenosine 5'-monophosphate and uridine 5'-monophosphate, and in their fatty acid compositions. Based on the composition of isoprenoid quinones, peptidoglycan, polar lipids and fatty acids, these organisms are members of the genus Exiguobacterium. This is supported by 16S rDNA analyses, which revealed 97-98% similarity to Exiguobacterium acetylicum DSM 20416T and 93.2-93.8% similarity to Exiguobacterium aurantiacum DSM 6208T. E. acetylicum DSM 20416T, the closest phylogenetic neighbour, shows only 39% DNA similarity to strain L2T and 40% DNA similarity to strain H2T. Based on genomic distinctiveness and the clear differences in chemotaxonomy and physiology, two novel species are proposed, Exiguobacterium undae sp. nov. and Exiguobacterium antarcticum sp. nov.

A moderately thermophilic, anaerobic bacterium, strain JW/MS-VS5T, was isolated from a mixed sediment/water sample of a hot spring at Bagnaccio (near Viterbo, Italy). The cells of this organism were straight to slightly curved rods, 0.4-0.6 x 2.03.0 microm in dimension. Cells occurred singly and stained Gram-positive. The temperature range for growth at pH(25C) 6.0 was 33-64 degrees C, the optimum being 58 degrees C. The pH(25C) range for growth was from 5.0 to 7.8, the optimum being 6.0-6.5. The substrates utilized included glycerol, glucose, fructose, mannose, galactose, sucrose, cellobiose, lactose, starch and yeast extract. Acetate and 1,3-propanediol were the only detectable organic products of glycerol fermentation; significant amounts of H2 were produced during growth. The strain was unable to grow autotrophically in the presence of H2 and CO2. The main products of glucose fermentation were CO2, H2, acetate and ethanol. Single amino acids, including serine, glutamine, threonine, leucine, methionine, aspartate, valine and histidine (but not arginine), served as carbon sources. Growth was completely inhibited by ampicillin, chloramphenicol, erythromycin, rifampicin and kanamycin at 100 microg ml(-1) and was retarded by streptomycin and tetracycline. The G+C content of the DNA was 32 mol% (HPLC). According to 16S rDNA sequence analysis, the isolate is located within the Gram-type positive Bacillus-Clostridium branch of the phylogenetic tree. On the basis of physiological properties and phylogenetic analysis, it is proposed that strain JW/MS-VS5T (the only, and type, strain) (= DSM 13723T = ATCC PTA 584T), constitutes the new species Caloramator viterbensis.

A novel cellulolytic and xylanolytic bacterium, strain MX5T, was isolated from the hindgut contents of the Australian termite Mastotermes darwiniensis (Froggatt). The isolate was a facultative anaerobe and had a Gram-positive cell-wall profile. The rod-shaped bacterium formed irregular coryneform and coccoid cells during growth. Phylogenetic analysis of the 16S rDNA provided evidence that the organism was closely related to the as-yet undescribed cellulolytic strain SR272 and the non-validly described species 'Cellulomonas pachnodae' as well as Promicromonospora citrea and Promicromonospora sukumoe. Strain MX5T was assigned to the genus Cellulosimicrobium on the basis of phylogenetic and chemotaxonomic criteria. The murein of strain MX5T contained the diamino acid lysine. N-Glycolylmuramic acid, mycolic acids and hydroxy fatty acids were absent. The major neutral sugar in the cell wall was galactose and the major quinone was menaquinone MK-9(H4). The predominant fatty acids were ai-C15:0, i-C15:0, i-C16:0 and C16:0. The G+C content of the DNA was in a range 70-72 mol%. On the basis of 16S rDNA sequence similarities and chemotaxonomic features, MX5T was clearly different from Cellulosimicrobium cellulans and other validly described species within this phylogenetic group. For this reason, a novel species is described, for which the name Cellulosimicrobium variabile sp. nov. is proposed.

Four actinomycete strains, isolated from the overheated region of manure compost, were assigned to the genus Thermobifida on the basis of morphological, physiological and biochemical characteristics. All strains produced single, ovoid, heat-sensitive spores on dichotomically branched aerial hyphae. On the basis of chemotaxonomic traits, these isolates showed strong affinity towards members of the genus Thermobifida. Cell-wall analysis revealed the presence of meso-diaminopimelic acid, but no other characteristic amino acids or sugars in the murein (cell wall type III). According to polar lipid analysis, all strains showed PL II-type phospholipid composition; phosphatidylethanolamine and glycolipid were detected together with some unidentified phospholipids. The isoprenoid quinone composition of the new isolates differed slightly from that of the other two Thermobifida species described thus far. The partial 16S rDNA sequence similarity of the four strains reached 99.8-100%, whereas a nearly complete 16S rDNA sequence of TB100T, the representative strain of this collection, showed only 97.4 and 97.8% similarity to the corresponding rDNA sequences of the type strains of Thermobifida fusca and Thermobifida alba, respectively. These four isolates constituted a homogeneous group with levels of DNA-DNA homology ranging from 94.6 to 99.1%. The DNA-DNA relative homology values of strain TB100T to Thermobifida fusca ATCC 27730T and Thermobifida alba DSM 43795T were 48.1 and 57%, respectively. On the basis of phenotypic, chemotaxonomic and genotypic data, the strains are assigned to a new species within the genus Thermobifida under the name Thermobifida cellulolytica sp. nov. The type strain is TB100T (= DSM 44535T = NCAIM B01997T).

A hitherto undescribed Actinomyces-like bacterium was isolated from the vagina of a dog. Biochemical testing and PAGE analysis of whole-cell proteins indicated that the isolate was phenotypically different from previously described Actinomyces species and related taxa. Sequencing of 165 rRNA showed that the unknown bacterium was distinct from all currently known Actinomyces species. Phylogenetically, the unidentified organism displayed a specific association with Actinomyces europaeus, but a sequence divergence of > 5% demonstrated that it represents a distinct species. Based on both phenotypic and 165 rRNA sequence considerations, it is proposed that the unknown strain from a dog be classified as a novel species, Actinomyces coleocanis sp. nov. The type strain is CCUG 41708T (= CIP 106873T).