- Volume 50, Issue 1, 2000

Methanosarcina semesiae MD1T (T = type strain), a novel obligately methylotrophic methanogenic archaeon is described. Strain MD1T was isolated from an enrichment on dimethylsulfide inoculated with mangrove sediment. The cells were irregularly coccoid, non-motile, 1.4+/-0.2 microm in diameter and stained Gram-positive. The catabolic substrates used included dimethylsulfide, methanethiol, methanol and methylated amines, but not acetate, formate, H2/CO2 or a combination of these substrates. When cells grown on dimethylsulfide were transferred to trimethylamine or methanol and vice versa, a lag phase was observed. The same lag phase occurred when cells grown on trimethylamine were transferred to methanol and vice versa, indicating that for each substrate different enzymes were induced. Fastest growth occurred within a temperature range of 30-35 degrees C and a pH of 6.5-7.5. Both Na+ and Mg2+ were required for growth, with maximum growth rates at 200-600 mM Na+ and 20-100 mM Mg2+. The cells exhibited specific growth rates (h-1) of 0.07+/-0.02, 0.15+/-0.04 and 0.18-/+0.05 on dimethylsulfide, methanol and trimethylamine, respectively. Analysis of the 16S rRNA gene sequence showed that strain MD1T was phylogenetically closely related to members of the genus Methanosarcina, but clearly differed from all described species of this genus (94-97% sequence similarity).

An unusual catalase-positive pleomorphic Gram-positive rod isolated from a human clinical specimen was subjected to a polyphasic taxonomic analysis. Comparative 16S rRNA gene sequence analysis revealed the unknown bacterium was a member of the high G+C branch of the Gram-positive bacteria (Actinobacteria), and was phylogenetically a member of the family Propionibacteriaceae, with Luteococcus japonicus as its nearest relative. Based on both phenotypic and phylogenetic evidence, it is proposed that the unknown bacterium be classified in the genus Luteococcus, as Luteococcus peritonei sp. nov. The type strain of Luteococcus peritonei is CCUG 38120T.

Strain AX-2T (T = type strain) was isolated from sediment of Great Salt Lake, Utah, USA. Optimal salinity for growth was 27% (w/v) NaCl and only a few carbohydrates supported growth of the strain. Strain AX-2T did not grow on complex substrates such as yeast extract or peptone. 16S rRNA analysis revealed that strain AX-2T was a member of the phyletic group defined by the family Halobacteriaceae, but there was a low degree of similarity to other members of this family. The polar lipid composition comprising phosphatidyl glycerol, the methylated derivative of diphosphatidyl glycerol, triglycosyl diethers and sulfated triglycosyl diethers, but not phosphatidyl glycerosulfate, was not identical to that of any other aerobic, halophilic species. On the basis of the data presented, it is proposed that strain AX-2T should be placed in a new taxon, for which the name Halorhabdus utahensis is appropriate. The type strain is strain AX-2T (= DSM 12940T).

An aerobic, nocardioform actinomycete, named LM 161T, was isolated from a soil sample obtained from a gold mine in Kongiu, Republic of Korea. This organism formed well-differentiated aerial and substrate mycelia and produced branched hyphae that fragmented into short or elongated rods. The cell wall contains major amounts of LL-diaminopimelic acid, alanine, glycine, glutamic acid, mannose, glucose, galactose, ribose and acetyl muramic acid. The major phospholipids of this isolate are phosphatidylcholine, diphosphatidylglycerol, phosphatidylglycerol and phosphatidylinositol, and the major isoprenologue is a tetrahydrogenated menaquinone with nine isoprene units. The whole-cell hydrolysate of strain LM 161T contains 12-methyltetradecanoic and 14-methylpentadecanoic acids as the predominant fatty acids, but does not contain mycolic acids. The G+C content of the DNA is 71.3 mol%. The phylogenetic position of the test strain was investigated using an almost complete 16S rDNA sequence. The isolate formed the deepest branch in the clade encompassing the members of the suborder Propionibacterineae Rainey et al. 1997. On the basis of chemical, phenotypic and genealogical data, it is proposed that this isolate be classified within a new genus as Hongia koreensis gen. nov., sp. nov. in the order Actinomycetales. The type strain is LM 161T (= IMSNU 50530T).

A new Gram-positive non-spore-forming bacterium, strain Lg2T, was isolated from an activated sludge reactor showing enhanced biological phosphorus removal activity. The new isolate was a slowly growing organism and was capable of accumulating large amounts of intracellular glycogen from substrate taken up. Both oxidase and catalase were produced. The new isolate contained meso-diaminopimelic acid (DAP) in the cell wall. Complex fatty acid patterns with iso-C14:0, anteiso-C15:0, C16:0, iso-C16:0 and four other minor saturated or unsaturated straight-chain fatty acids were detected. The isolate contained a high genomic G+C content (70.5 mol%). Phylogenetic analysis based on the 16S rRNA gene sequence placed the isolate in the high G+C Gram-positive group with Microlunatus phosphovorus and Friedmanniella antarctica as the closest relatives (sequence similarities are 93 and 92 %, respectively). These three organisms shared common features in morphology, but strain Lg2T could be differentiated from the other species by its peptidoglycan type (meso-DAP), fatty acid composition, carbon source utilization profile and G+C content. On the basis of these findings, it is proposed that a new genus and species, Micropruina glycogenica, should be created for the new isolate; the type strain is strain Lg2T (= JCM 10248T).

Several yellow-pigmented isolates, with optimum growth temperatures of about 65-70 degrees C, were recovered from hot springs in Iceland. Phylogenetic analysis of the 16S rDNA and DNA-DNA reassociation values showed that these organisms represented two new species of the genus Thermus. Strains RF-4T and HN1-8 had maximum temperatures for growth below 80 degrees C, while strains HN3-7T and HN2-7, unlike all other strains of the species of the genus Thermus except those belonging to Thermus thermophilus, grew at 80 degrees C. The new isolates from Iceland could not be distinguished easily from each other or from other strains of the species of the genus Thermus by biochemical characteristics; however, strains RF-4T and HN1-8 assimilated ribitol, a characteristic which was not detected in any of the other strains examined. Moreover, the species represented by strains RF-4T and HN1-8 and the species represented by strains HN3-7T and HN2-7 could be distinguished clearly from the other species of Thermus by their fatty acid composition. Strains RF-4T and HN1-8 have the highest combined levels of iso-15:0 and iso-17:0 and the lowest levels of iso-16:0 of any of the strains of the species of Thermus, while strains HN3-7T and HN2-7 are characterized by a very low iso-15:0/iso-17:0 ratio. On the basis of the phylogenetic analysis, DNA-DNA reassociation values, physiological and biochemical characteristics and fatty acid composition, the name Thermus igniterrae sp. nov. is proposed for the species represented by strains RF-4T and HN1-8 and the name Thermus antranikianii sp. nov. is proposed for the species represented by strains HN3-7T and HN2-7.

Bdellovibrios are unique bacteria with the ability to prey upon a wide variety of susceptible Gram-negative bacteria. Micro-organisms exhibiting this trait have been included in the genus Bdellovibrio despite their isolation from diverse habitats and relatively unstudied taxonomic relatedness. In this study, 16S rDNA sequences were compared from known terrestrial Bdellovibrio species, Bdellovibrio bacteriovorus 100T, Bdellovibrio stolpii Uki2T and Bdellovibrio starrii A3.12T in order to study their phylogenetic relationship. The two sequences from B. stolpii Uki2T and B. starrii A3.12T were 90.0% similar to each other but exhibited only 81.7% and 81.2% similarity, respectively to B. bacteriovorus 100T. Phylogenetic analysis indicated that B. bacteriovorus 100T clustered in a separate clade from B. starrii A3.12T and B. stolpii Uki2T, demonstrating only a distant relationship between B. bacteriovorus 100T and the other two recognized type species. DNA-DNA hybridization experiments also demonstrated <4% hybridization between these three species. On the basis of the results obtained from the phylogenetic analysis and DNA-DNA hybridization studies, it is proposed that B. stolpii Uki2T and B. starrii A3.12T should be transferred to a new genus, Bacteriovorax gen. nov. as Bacteriovorax stolpii comb. nov. and Bacteriovorax starrii comb. nov., respectively. It is also proposed that the type species for the new genus Bacteriovorax should be Bacteriovorax stolpii comb. nov.

The newly developed cultivars of soybean in Africa, known as Tropical Glycine cross (TGx), are nodulated by bradyrhizobia indigenous to African soils, here designated Bradyrhizobium spp. (TGx). Isolates of Bradyrhizobium spp. (TGx) obtained from nodules of TGx soybeans that were inoculated with soils from 65 locations in six African countries were characterized and grouped into 11 phylogenetic clusters on the basis of RFLP of the 16S rRNA gene. Five restriction enzymes (RsaI, HinfI, MspI, CfoI and HaeIII) established RFLP groups within these Bradyrhizobium spp. (TGx) isolates, which were used to construct a phylogenetic tree showing their genetic relationship with other Bradyrhizobium species. RFLP analysis indicated that Bradyrhizobium spp. (TGx) is a heterogeneous group with some isolates related to Bradyrhizobium japonicum and Bradyrhizobium elkanii strains and some to Bradyrhizobium spp. (misc.) reference strains isolated from a variety of tropical legumes. The heterogeneity within the large phylogenetic clusters was further examined through analysis of randomly amplified polymorphic DNA (RAPD) using GC-rich PCR primers. The RAPD analysis showed additional heterogeneity in the Bradyrhizobium spp. (TGx) phylogenetic clusters, which was not revealed by separations based on RFLP analysis. The Bradyrhizobium spp. (TGx) isolates were classified into effective and ineffective types based on their symbiotic performance on TGx soybean. The isolates were randomly distributed throughout the phylogenetic clusters regardless of their symbiotic effectiveness on TGx soybean.

The bacterial leaf-spot of anthurium emerged during the 1980s, in the French West Indies and Trinidad. This new bacterial disease is presently wide spread and constitutes a serious limiting factor for commercial anthurium production. Twenty-nine strains isolated from leaf-spots of naturally infected anthurium were characterized and compared with reference strains belonging to the Comamonadaceae family, the genera Ralstonia and Burkholderia, and representative fluorescent pseudomonads. From artificial inoculations 25 out of 29 strains were pathogenic on anthurium. Biochemical and physiological tests, fatty acid analysis, DNA-DNA hybridization, 16S rRNA gene sequence analysis, DNA-16S RNA hybridization were performed. The 25 pathogenic strains on anthurium were clustered in one phenon closely related to phytopathogenic strains of the genus Acidovorax. Anthurium strains were 79-99% (deltaTm range 0.2-1.6) related to the strain CFBP 3232 and constituted a discrete DNA homology group indicating that they belong to the same species. DNA-rRNA hybridization, 16S rRNA sequence and fatty acid analysis confirmed that this new species belongs to the beta-subclass of Proteobacteria and to rRNA superfamily III, to the family of Comamonadaceae and to the genus Acidovorax. The name Acidovorax anthurii is proposed for this new phytopathogenic bacterium. The type strain has been deposited in the Collection Française des Bactéries Phytopathogènes as CFBP 3232T.

DNA-DNA reassociation studies were performed on previously described 'CI strains', which form an unusual 16S rRNA population within the 'anginosus' group of Streptococcus. The CI strains displayed reassociation values of >70% with the Streptococcus constellatus NCDO 2226T strain, with Tm values <1 degrees C, indicating phylogenetic species identity.

A new Lactobacillus species from pig small intestine has been identified. In an attempt to isolate Lactobacillus reuteri strains carrying the putative colonization-factor gene (mub, for mucus binding) a mub-derived gene probe was used to screen pig intestinal material. A number of isolates were obtained and primary characterization showed that they were Gram-positive, catalase-negative, non-spore-forming, non-motile rods. Growth occurred at 45 degrees C but not at 15 degrees C and the DNA G+C content was 46 mol%. Cell wall analysis together with DNA-DNA hybridization and analysis of the 16S rRNA sequence revealed that the new isolates represent a previously undescribed Lactobacillus species closely related to L. reuteri, Lactobacillus fermentum and Lactobacillus pontis. The name Lactobacillus mucosae is proposed for this species and the type strain is S32T.

A novel, curved (0.3 x 4.0-5.0 microm), Gram-negative, non-sporulating, mesophilic bacterium, designated strain ILE-3T (T = type strain), was isolated from an anaerobic lagoon in a dairy wastewater treatment plant. Optimal growth occurred at 37 degrees C and pH 7.4 on a medium containing serine as an energy source and yeast extract. The strain was motile by means of one or two lateral flagella. It required yeast extract for growth on serine, glycine, threonine and pyruvate. Poor growth was obtained on cysteine, Casamino acids, biotrypcase, peptone and 2-oxoglutarate. In the presence of Methanobacterium formicicum, strain ILE-3T oxidized alanine, glutamate, leucine, isoleucine, valine and aspartate to a minor extent. The G+C content of the DNA was 44 mol%. Phylogenetic analysis of the 16S rRNA gene of strain ILE-3T indicated that it was related to Aminobacterium colombiense (95% similarity value). On the basis of the phenotypic and phylogenetic characteristics, strain ILE-3T is designated as a new species of the genus Aminobacterium, namely Aminobacterium mobile sp. nov. (= DSM 12262T).

A marine bacterium, Alteromonas elyakovii KMM 162T, which was described recently, and five strains isolated from spot-wounded fronds of Laminaria japonica have been subjected to phylogenetic analysis, and geno- and phenotypic characterization. The phenotypic features of Pseudoalteromonas elyakovii strains were closely related to that of Pseudoalteromonas espejiana IAM 12640T, but utilization of three carbon compounds (D-mannose, L-tyrosine and trehalose) distinguished both species. The G+C content of Pseudoalteromonas elyakovii was between 38.5 and 38.9 mol%. Pseudoalteromonas elyakovii KMM 162T and the five Laminaria isolates constitute a single species different from any other Alteromonas and Pseudoalteromonas species as revealed by DNA-DNA hybridization data, especially Pseudoalteromonas distincta KMM 638T (52.4%), Pseudoalteromonas citrea KMM 216 (49.5%), Pseudoalteromonas carrageenovora NCIMB 302T (46.9%) and Pseudoalteromonas espejiana IAM 12640T (29.9%). All the data indicated that Alteromonas elyakovii KMM 162T should be reclassified as Pseudoalteromonas elyakovii and five strains isolated from Laminaria japonica have to be included in the species. Pseudoalteromonas elyakovii comb. nov. (type strain, KMM 162T = ATCC 700519T) is proposed and a set of phenotypic features which differentiate the Pseudoalteromonas species is described.

A group of yellow-pigmented isolates from ammonia-supplied biofilters showed an unusual denitrification reaction. All strains reduced nitrite but not nitrate without production of nitrogen (N2). The only product found was nitrous oxide (N2O). The strains were divided into two clusters and one separate strain by their fatty acid profiles, which were similar to the fatty acid profiles of the genera Xanthomonas and Stenotrophomonas. Analyses of the 165 rDNA sequences showed that these clusters and the separate strain form three independent lines within the Xanthomonas branch of the Proteobacteria. The evolutionary distances of the isolates to members of the related genera Xanthomonas, Stenotrophomonas and Xylella calculated by the 16S rDNA sequences led to the proposal of two new genera and three new species, Stenotrophomonas nitritireducens sp. nov., Luteimonas mephitis gen. nov., sp. nov. and Pseudoxanthomonas broegbernensis gen. nov., sp. nov. The type strains are Stenotrophomonas nitritireducens L2T (= DSM 12575T), Luteimonas mephitis B1953/27.1T (= DSM 12574T) and Pseudoxanthomonas broegbernensis B1616/1T (= DSM 12573T).

Three scotochromogenic Mycobacterium xenopi-like organisms were isolated from stream waters in Finland. These strains grew at 36-50 degrees C but not at 30 degrees C. One of the three strains was fully compatible with the M. xenopi type strain according to GLC-MS, biochemical tests, and 16S rDNA and 16S-23S rDNA internal transcribed spacer (ITS) sequencing. Two of the strains closely resembled M. xenopi in lipid analyses and biochemical tests, but analysis by GLC-MS verified the presence of two new marker fatty acids (2,4,6,x-tetramethyl-eicosanoic acid and 2,4,6,x,x-pentamethyl-docosanoic acid). The 16S rDNA and ITS region sequences of these two strains differed from those of M. xenopi and other previously described mycobacterial sequences. Therefore, the strains are regarded as new species of slow-growing mycobacteria, for which the name Mycobacterium botniense sp. nov. is proposed. The chemical, physical and microbiological quality of the water reservoirs of M. xenopi and M. botniense are described. As far as is known, this is the first time that M. xenopi has been isolated from natural waters. The strains of M. botniense sp. nov. (E347T and E43) have been deposited in the ATCC as strains 700701T and 700702, respectively.

The fastidious nature of the mollicutes (mycoplasmas), their lack of a classic bacterial cell wall, and their very small genome, make phylogenetic placements of new species in this enlarging group of prokaryotes an important and valuable aid in their classification. In this report we have determined the phylogeny of the Mycoplasma hominis cluster of the hominis group. The 16S rDNA sequences from several previously described Mycoplasma species were determined and ten species were found to belong to the M. hominis cluster. With almost complete sequences available, the phylogenetic analysis revealed that the M. hominis cluster currently comprises 19 species, forming a distinct clade as judged from branch lengths, bootstrap percentage values, nucleotide signature analysis, and structural elements in the 16S rRNA molecule. The 16S rRNA gene sequences of species in the M. hominis cluster were found to be > or = 94% similar and the range within which similarities can be used in the classification of new species is discussed. Members of the M. hominis cluster all share a major biochemical property of M. hominis, in that they hydrolyse arginine and are incapable of fermenting glucose. This consistency in phenotypic pattern has not been found in any of the other phylogenetic clusters of the hominis group. Two species, the non-cultivable agent of Grey Lung disease in rodents (tentatively named 'Candidatus Mycoplasma ravipulmonis') and the avian species Mycoplasma gypis strain B1/T1T, were regarded as close relatives to the M. hominis cluster, but are clearly separated from the species of this cluster. Both species formed early branches of the M. hominis cluster and should be regarded as individual lines containing one species.

Two Gram-negative, aerobic, pointed and budding bacteria were isolated from various depths of hypersaline, heliothermal and meromictic Ekho Lake (Vestfold Hills, East Antarctica). 16S rRNA gene sequence comparisons show the isolates to be phylogenetically close to the genera Sulfitobacter and Roseobacter. Cells can be motile and contain storage granules. Sulfite addition does not stimulate growth. Isolate EL-38T can produce bacteriochlorophyll a and has a weak requirement for sodium ions; polar lipids include phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine and an unidentified amino lipid, but not diphosphatidylgycerol. The dominant fatty acid is 18:1omega7c; other characteristic fatty acids are 3-OH 10:0, 3-OH 14:1, 16:0, 18:0, 18:2 and 19:1. The DNA base composition is 55.0-56.3 mol% G+C. Isolate EL-162T has an absolute requirement for sodium ions. Diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine and an unidentified amino lipid are present in the polar lipids. Dominant fatty acids of this isolate are 18:1omega7c and 18:1omega9c as well as 18:2 which is present as two isomers. Other characteristic fatty acids are 3-OH 10:0, 3-OH 14:1, 16:0 and 18:0. The G+C content is 57.9-58.1 mol%. Morphological, physiological and genotypic differences from related, thus far known genera support the description of Staleya guttiformis gen. nov. and sp. nov. with EL-38T (= DSM 11458T) as the type strain and of Sulfitobacter brevis sp. nov. with the type strain EL-162T (= DSM 11443T).

An aerobic, thermophilic, xylanolytic, spore-forming bacterium, XETP (T = type strain; P = patent strain), has been isolated from farm soil situated underneath a manure heap in northern France. Strain XETP, which stained negative in the Gram test, occurs as short rods which sometimes form chains. Its spores are ellipsoidal, central to subterminal and occur in swollen sporangia. It grows at temperatures up to 63 degrees C and in the pH range 6.5-8.5. When grown on glucose in optimal conditions, its doubling time was found to be 33 min. CO2 was observed to have a growth-stimulating effect at the start of the culture. In addition to glucose, the isolate utilizes xylose, arabinose, mannose, cellobiose, galactose, maltose, sucrose, xylan and starch. Growth is inhibited by 5% NaCl. The G+C content of strain XETP is 57.5 mol%. The 16S rDNA sequence analysis indicated that strain XETP falls into the radiation of the Bacillus-Lactobacillus-Streptococcus subdivision of the Gram-positive phylum. Its three closest phylogenetic relatives are 'Bacillus viscosus', Paenibacillus curdlanolyticus and Bacillus popilliae with identity values of 91.15, 90.94 and 90.92%, respectively. The major cellular fatty acids are 14-methyl pentadecanoic acid (16:0 iso), hexadecanoic acid (16:0) and 14-methyl hexadecanoic acid (17:0 anteiso). On the basis of 16S rRNA sequence and chemotaxonomic characteristics, the isolate is different enough for it to be considered as a member of a new genus. It is therefore proposed that this isolate represents a new genus and species: Thermobacillus xylanilyticus. Strain XETP, the type strain of Thermobacillus xylanilyticus, has been deposited in the Collection Nationale de Cultures Microbiennes (CNCM I-1017) as a patent strain.

A mesophilic, facultatively anaerobic, marine bacterium, designated strain FG1T, was isolated from a seagrass bed sediment sample collected from Nanwan Bay, Kenting National Park, Taiwan. Cells grown in broth cultures were motile, Gram-negative rods; motility was normally achieved by two sheathed flagella at one pole of the cell. Strain FG1T required Na+ for growth, and exhibited optimal growth at 30-35 degrees C, pH 6-7 and about 4% NaCl. It grew anaerobically by fermenting glucose and other carbohydrates with production of various organic acids, including acetate, lactate, formate, malate, oxaloacetate, propionate, pyruvate and succinate, and the gases CO2 and H2. The strain did not require either vitamins or other organic growth factors for growth. Its DNA G+C content was 45.9 mol%. It contained C12:0 as the most abundant cellular fatty acid. Characterization data, together with the results of a 16S rDNA-based phylogenetic analysis, indicate that strain FG1T represents a new species of the genus Vibrio. Thus, the name Vibrio aerogenes sp. nov. is proposed for this new bacterium. The type strain is FG1T (= ATCC 700797T = CCRC 17041T).