- Volume 53, Issue 4, 2003

A new taxonomy of the green sulfur bacteria is proposed, based on phylogenetic relationships determined using the sequences of the independent 16S rRNA and fmo (Fenna–Matthews–Olson protein) genes, and supported by the DNA G+C content and sequence signatures. Comparison of the traditional classification system for these bacteria with their phylogenetic relationship yielded a confusing picture, because properties used for classification (such as cell morphology, photosynthetic pigments and substrate utilization) do not concur with their phylogeny. Using the genetic information available, strains and species assigned to the genera Chlorobium, Pelodictyon and Prosthecochloris are considered, and the following changes are proposed. Pelodictyon luteolum is transferred to the genus Chlorobium as Chlorobium luteolum comb. nov. Pelodictyon clathratiforme and Pelodictyon phaeoclathratiforme are transferred to the genus Chlorobium and combined into one species, Chlorobium clathratiforme comb. nov. The name Pelodictyon will become a synonym of Chlorobium. Strains known as Chlorobium limicola subsp. thiosulfatophilum that have a low DNA G+C content (52–52·5 mol%) are treated as strains of Chlorobium limicola; those with a high DNA G+C content (58·1 mol%) are transferred to Chlorobaculum gen. nov., as Chlorobaculum thiosulfatiphilum sp. nov. Chlorobium tepidum is transferred to Chlorobaculum tepidum comb. nov., and defined as the type species of the genus Chlorobaculum. Strains assigned to Chlorobium phaeobacteroides, but phylogenetically distant from the type strain of this species, are assigned to Chlorobium limicola and to Chlorobaculum limnaeum sp. nov. Strains known as Chlorobium vibrioforme subsp. thiosulfatophilum are transferred to Chlorobaculum parvum sp. nov. Chlorobium chlorovibrioides is transferred to ‘Chlorobaculum chlorovibrioides’ comb. nov. The type strain of Chlorobium vibrioforme is phylogenetically related to Prosthecochloris, and is therefore transferred to this genus as Prosthecochloris vibrioformis comb. nov. Consequently, the name Chlorobium vibrioforme will become a synonym of Prosthecochloris vibrioformis, and other strains that were assigned to this species are now considered to belong to Chlorobium luteolum, Chlorobium phaeovibrioides and ‘Chlorobaculum chlorovibrioides’, according to their phylogenetic relatedness.

Phylogenetic relationships of five jujube witches'-broom (JWB) phytoplasma isolates from four different districts, and other phytoplasmas, were investigated by 16S rDNA PCR amplification and sequence analysis. The 16S rDNA sequences of any pair of the five isolates of JWB phytoplasmas were >99·5 % similar. The JWB phytoplasma 16S rDNA sequences were most closely related to that of the elm yellows (EY) phytoplasma in 16S-group VIII. Phylogenetic analysis of the 16S rDNA sequences from the JWB phytoplasma isolates, together with sequences from most of the phytoplasmas archived in GenBank, produced a tree in which the JWB isolates clustered as a discrete subgroup. The uniqueness of the JWB phytoplasma appears to be correlated with a specific insect vector (Hishimonus sellatus) and the host plant (Zizyphus jujuba), or with a specific geographical distribution. The unique properties of the JWB phytoplasma sequences clearly indicate that it represents a novel taxon, ‘Candidatus Phytoplasma ziziphi’.

A novel thermophilic, microaerophilic, facultatively chemolithoheterotrophic bacterium designated strain TRT was isolated from a sample of a deep-sea hydrothermal chimney collected at the Rainbow vent field on the Mid-Atlantic Ridge (36°14′N). Gram-negative, non-spore-forming, non-motile rods occurred singly or in pairs. The organism grew in the temperature range 37–80 °C with an optimum at 70 °C and at pH 5·5–8·4 with an optimum around 6·7. The NaCl range for growth was 10–50 g l−1 with an optimum of 30 g l−1. Strain TRT grew chemoorganoheterotrophically with carbohydrates, proteinaceous substrates, organic acids and alcohols using oxygen or nitrate as electron acceptors. The isolate was able to grow at oxygen concentrations from 0·5 to 21 %. Oxygen concentrations that promoted fastest growth ranged from 4 to 8 % under agitation. The novel isolate was able to grow lithoheterotrophically with molecular hydrogen as the energy source. The G+C content of the genomic DNA was 68·4 mol%. Phylogenetic analysis of the 16S rDNA sequence placed strain TRT within the phylum Deinococcus–Thermus of the Bacteria. On the basis of phenotypic and phylogenetic data, it is proposed that this isolate should be described as a member of a novel species of a new genus as Vulcanithermus mediatlanticus gen. nov., sp. nov. The type strain is TRT (=DSM 14978T =VKM B-2292T =JCM 11956T).

A phylogenetically novel aerobic bacterium was isolated from an anaerobic–aerobic sequential batch reactor operated under enhanced biological phosphorus removal conditions for wastewater treatment. The isolation strategy used targeted slowly growing polyphosphate-accumulating bacteria by combining low-speed centrifugations and prolonged incubation on a low-nutrient medium. The isolate, designated strain T-27T, was a Gram-negative, rod-shaped aerobe. Cells often appeared to divide by budding replication. Strain T-27T grew at 25–35 °C with an optimum growth temperature of 30 °C, whilst no growth was observed below 20 °C or above 37 °C within 20 days incubation. The pH range for growth was 6·5–9·5, with an optimum at pH 7·0. Strain T-27T was able to utilize a limited range of substrates, such as yeast extract, polypepton, succinate, acetate, gelatin and benzoate. Neisser staining was positive and 4,6-diamidino-2-phenylindole-stained cells displayed a yellow fluorescence, indicative of polyphosphate inclusions. Menaquinone 9 was the major respiratory quinone. The cellular fatty acids of the strain were mainly composed of iso-C15 : 0, C16 : 1 and C14 : 0. The G+C content of the genomic DNA was 66 mol%. Comparative analyses of 16S rRNA gene sequences indicated that strain T-27T belongs to candidate division BD (also called KS-B), a phylum-level lineage in the bacterial domain, to date comprised exclusively of environmental 16S rDNA clone sequences. Here, a new genus and species are proposed, Gemmatimonas aurantiaca (type strain T-27T=JCM 11422T=DSM 14586T) gen. nov., sp. nov., the first cultivated representative of the Gemmatimonadetes phyl. nov. Environmental sequence data indicate that this phylum is widespread in nature and has a phylogenetic breadth (19 % 16S rDNA sequence divergence) that is greater than well-known phyla such as the Actinobacteria (18 % divergence).

Thirteen isolates of Acinetobacter were obtained from activated sludge plants in Victoria, Australia. Earlier 16S–23S rDNA genomic fingerprinting and partial 16S rDNA sequence data had suggested that these isolates might contain previously undescribed species. This view was confirmed here. A polyphasic taxonomic approach involving phenotypic characterization, near-complete 16S rDNA sequence data and DNA–DNA hybridization analyses support the view that seven novel genomic species can be differentiated in this group of isolates. However, when fluorescence in situ hybridization (FISH) studies were performed with a 16S-rRNA-targeted probe specific for the genus Acinetobacter, used to identify Acinetobacter in activated sludge plants, all these strains responded positively. This suggests that these isolates would not have been missed in earlier FISH studies where their role as polyphosphate-accumulating bacteria has been questioned. This report describes these isolates and proposes that they be named Acinetobacter baylyi (type strain B2T=DSM 14961T =CIP 107474T), Acinetobacter bouvetii (type strain 4B02T=DSM 14964T =CIP 107468T), Acinetobacter grimontii (type strain 17A04T=DSM 14968T =CIP 107470T), Acinetobacter tjernbergiae (type strain 7N16T=DSM 14971T =CIP 107465T), Acinetobacter towneri (type strain AB1110T=DSM 14962T =CIP 107472T), Acinetobacter tandoii (type strain 4N13T=DSM 14670T =CIP 107469T) and Acinetobacter gerneri (type strain 9A01T=DSM 14967T =CIP 107464T).

Two red pigment-producing bacterial strains with a metallic green sheen were isolated from a sediment sample of getbol, the Korean tidal flat. Phylogenetic analysis based on 16S rDNA sequences showed that these isolates represent a phyletic lineage within the γ-Proteobacteria that is distantly related to the genus Hahella. No bacterial species with validly published names showed ⩾92 % 16S rRNA similarity with the getbol isolates. The strains were Gram-negative, chemo-organotrophic, aerobic and required NaCl (1–7 %) for growth. They produced pigments with maximum absorption at 540 nm, which indicated the presence of prodigiosin, a well-known red pigment previously detected in Serratia marcescens. The major isoprenoid quinone was ubiquinone-9. The predominant cellular fatty acids were saturated and monounsaturated straight-chain fatty acids. The DNA G+C contents ranged from 40 to 42 mol%. The combination of physiological, biochemical and chemotaxonomic data clearly separated the test strains from other phylogenetically related genera in the γ-Proteobacteria. On the basis of polyphasic evidence from this study, it is proposed that the two getbol isolates should be classified in a novel genus, Zooshikella gen. nov., as Zooshikella ganghwensis sp. nov.

Two bacterial strains, HTCC2503T and HTCC2517, were isolated from the Bermuda Atlantic Time Series Station in the western Sargasso Sea, Atlantic Ocean, by new high-throughput culture methods that rely on dilution to extinction in very-low-nutrient media. Characterization of the two strains by polyphasic approaches revealed that they belonged to the same species. These isolates are Gram-negative, strictly aerobic, chemoheterotrophic, slightly motile short rods with a single flagellum. The temperature, pH and NaCl concentration ranges for growth were 10–37 °C, 6·0–9·0 and 0·75–20 % (w/v), respectively. Colonies on marine agar were very small (0·3–0·8 mm in diameter), yellowish-brown and very hard. Carotenoid pigments were synthesized but bacteriochlorophyll a was not. Several kinds of pentose, hexose, sugar alcohol, oligosaccharide and amino acid were utilized as sole carbon sources. Oxidase was produced, but catalase was not. All cellular fatty acids were even-numbered monounsaturated or saturated fatty acids and the major fatty acid was cis-7-octadecenoic acid (73·3 %). The DNA G+C content of strain HTCC2503T was 60·8 mol%. Phylogenetic analyses of 16S rRNA gene sequences clearly indicated that the strains formed a distinct lineage, allied with activated sludge environmental clone H9, in the α-Proteobacteria. The clade containing strains HTCC2503T and HTCC2517 and clone H9 could not be phylogenetically associated with any of the six known orders of the α-Proteobacteria. From this polyphasic evidence, it is proposed that the novel strains should be classified as Parvularcula bermudensis gen. nov., sp. nov. The type strain is HTCC2503T (=ATCC BAA-594T =KCTC 12087T) and the reference strain is HTCC2517.

Thirteen Ralstonia pickettii-like isolates from the environment (water, soil and activated sludge) and human clinical samples (including respiratory secretions of cystic fibrosis patients) were investigated in a polyphasic taxonomic study that employed 16S rDNA sequence analysis, DNA–DNA hybridization, determination of DNA base composition, whole-cell protein analysis, biochemical characterization and PCR-based assays. All isolates were classified as a novel Ralstonia species, for which the name Ralstonia insidiosa sp. nov. is proposed. The type strain, LMG 21421T (=CCUG 46789T), was isolated from the sputum of a patient with acute lymphoblastic leukaemia. R. insidiosa can be differentiated from other species of the genus Ralstonia and phenotypically similar species (including the Burkholderia cepacia complex and Achromobacter xylosoxidans) by a variety of biochemical tests, whole-cell protein analysis and several PCR-based assays. Some outstanding issues in the taxonomy of the genus Ralstonia are also discussed.

A bacterial strain (Chol-1ST) that is able to oxidize cholesterol to CO2 and reduce nitrate to dinitrogen was enriched and isolated from an upflow sludge bed (USB) anoxic reactor that treats sanitary landfill leachate from the city of Montevideo, Uruguay. Cells of strain Chol-1ST were Gram-negative, rod-shaped to slightly curved, measured 0·5–0·6×1·0-1·3 μm and were motile by a single polar flagellum. Strain Chol-1ST grew optimally at 30–32 °C and pH 7·0, with a doubling time of 44–46 h when cholesterol was used as the sole carbon and energy source. The metabolism of strain Chol-1ST was strictly respiratory, with oxygen or nitrate as the terminal electron acceptor. The presence of ubiquinone Q-8 as the sole respiratory lipoquinone indicated that strain Chol-1ST belonged to the β-subclass of the Proteobacteria. Phosphatidylethanolamine was the predominant polar lipid and the G+C content of the DNA was 65·3 mol%. The fatty acid profile of strain Chol-1ST, cultivated under denitrifying conditions by using a defined mineral medium supplemented with cholesterol, was characterized by the following major components: summed feature 4 (C16 : 1 ω7c and/or iso C15 : 0 2-OH), C16 : 0, C18 : 1 ω7c and hydroxy acid C10 : 0 3-OH. Minor components included C10 : 0, C11 : 0, C12 : 0, C14 : 0, C15 : 0, C19 : 0, C19 : 0 10-methyl and hydroxylated acids C8 : 0 3-OH and C16 : 0 3-OH. Analysis of the 16S rDNA sequence showed that strain Chol-1ST represents a separate lineage within the Thauera, Azoarcus, Zoogloea and Rhodocyclus assemblage of the β-Proteobacteria. Strain Chol-1ST had highest sequence similarity (96·5 %) with strain 72Chol, a denitrifying β-Proteobacterium. On the basis of polyphasic evidence, strain Chol-1ST (=DSM 13999T =ATCC BAA-354T) is proposed as the type strain of Sterolibacterium denitrificans gen. nov., sp. nov.

Eleven psychrophilic bacteria isolated from Antarctic coastal marine environments were subjected to a polyphasic taxonomic study. The isolates were oxidase-positive, halotolerant, Gram-negative, non-motile coccobacilli with a strictly oxidative metabolism. The DNA G+C content ranged from 44 to 47 mol%. DNA–DNA hybridization experiments showed six homology groups, two of them related at the species level to the type strain of Psychrobacter immobilis, LMG 7203T (70–83 %). The highest DNA relatedness of two other groups to known Psychrobacter species was found to the type strain of Psychrobacter glacincola, LMG 21282T (51–57 %), and no significant similarity was found between Psychrobacter type strains and the last two groups. The predominant cellular fatty acids detected were typical of the genus Psychrobacter and included 18 : 1ω9c, 16 : 1ω7c and 17 : 1ω8c. 16S rRNA gene sequence analysis confirmed that the strains isolated belonged to the genus Psychrobacter. The results of the study assigned five isolates to P. immobilis, three isolates to P. glacincola and three isolates to novel Psychrobacter species. The names Psychrobacter luti sp. nov. (type strain NF11T=LMG 21276T= CECT 5885T) and Psychrobacter fozii sp. nov. (type strain NF23T=LMG 21280T =CECT 5889T) are proposed for these organisms.

The taxonomic position of the marine bacterium ‘Alteromonas marinoglutinosa’ NCIMB 1770 was investigated in a polyphasic study. Analysis of 16S rDNA sequence and DNA–DNA reassociation values confirmed the phylogenetic position of strain NCIMB 1770 within the genus Pseudoalteromonas as a separate species, distinct from all Pseudoalteromonas species with validly described names. On the basis of physiological and molecular properties, it is proposed that strain NCIMB 1770 is classified as Pseudoalteromonas mariniglutinosa sp. nov., nom. rev., comb. nov., with the type strain NCIMB 1770T (=KMM 3635T).

A novel bacterial species, Rhodocista pekingensis sp. nov., was isolated from a municipal wastewater treatment plant and characterized by polyphasic taxonomy. Cells of R. pekingensis were Gram-negative, motile by a single polar flagellum, vibrioid to spiral, 0·6–0·8 μm in width and 0·8–1·5 μm in length. R-bodies were not observed. Phototrophically grown cells contained lamellar photosynthetic membranes and bacteriochlorophyll a. Cell growth was anaerobically phototrophic or aerobically chemoheterotrophic. Anaerobically grown cultures were pink-reddish. Thiamin and vitamin B12, but not biotin, were required for growth and 0·05 % yeast extract stimulated growth. Acetate, lactate, pyruvate and succinate supported growth. Cysts were formed when butyrate was used as the sole carbon source. Molecular hydrogen (H2), but not sulfide or thiosulfate, was used as an electron donor. The major cellular quinone was Q-9. The DNA G+C content of cells was 68·8 mol%. The type strain of Rhodocista pekingensis is 3-pT (=AS 1.2194T =JCM 11669T).

A bacterium previously isolated from a diseased colony of the scleractinian coral Dichocoenia stokesi (common name elliptical star coral) was subjected to a detailed polyphasic taxonomic characterization. The isolate, designated WP1T, was halophilic and strictly aerobic and formed golden-orange-pigmented colonies after prolonged incubation. Cells of WP1T were Gram-negative, rod-shaped and showed a characteristic branching rod morphology. Chemotaxonomically, WP1T was characterized by having Q-10 as the major respiratory lipoquinone and sym-homospermidine as the main component of the cellular polyamine content. The predominant constituent in the cellular fatty acid profile was C18 : 1 ω7c, along with C19 : 0 cyclo ω8c and C16 : 0. Other fatty acids present in smaller amounts were C17 : 0, C18 : 0, C16 : 1 ω7c, C20 : 1 ω7c and C18 : 1 2-OH. The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine. Minor amounts of diphosphatidylglycerol, phosphatidylmonomethylethanolamine and phosphatidyldimethylethanolamine were present. The G+C content of the genomic DNA was 66·3 mol%. Phylogenetic analysis of the 16S rRNA gene sequence showed that WP1T represents a separate subline of descent within the order ‘Rhizobiales’ of the ‘Alphaproteobacteria’. The new line of descent falls within the group of families that includes the Rhizobiaceae, Bartonellaceae, Brucellaceae and ‘Phyllobacteriaceae’, with no particular relative within this group. The 16S rRNA gene sequence similarity to all established taxa within this group was not higher than 92·0 % (to Mesorhizobium mediterraneum). To accommodate this emerging coral pathogen, the creation of a new genus and species is proposed, Aurantimonas coralicida gen. nov., sp. nov. (type strain WP1T=CIP 107386T =DSM 14790T).

Phylogenetic, chemotaxonomic and metabolic data obtained for Desulfobacterium macestii indicate that this species is not a member of the genus Desulfobacterium, but of the genus Desulfomicrobium. Phylogenetically, it is closely related to Desulfomicrobium baculatum and Desulfomicrobium norvegicum, but it can be differentiated from these species by its metabolic properties. It is therefore proposed to reclassify Desulfobacterium macestii as Desulfomicrobium macestii comb. nov.

In order to clarify the taxonomic position of Dechlorosoma suillum, which shares 99·9 % 16S rDNA sequence identity (1433 of 1435 bp) with Azospira oryzae, we compared the two species in a polyphasic taxonomic approach. Results of 122 physiological and biochemical tests for D. suillum DSM 13638T and Azospira oryzae 6a3T were identical, except for the lack of growth of Azospira oryzae 6a3T with perchlorate as the terminal electron acceptor. Presence of a nifH gene and nitrogenase activity, a key feature of Azospira, were also detected in D. suillum by Southern hybridization and by the acetylene reduction assay, respectively. Whole-cell SDS-PAGE profiles of SDS-soluble proteins of strains DSM 13638T and 6a3T were almost identical. DNA–DNA hybridization studies showed more than 90 % binding between D. suillum and two strains of Azospira oryzae. These data provide evidence that the two bacteria belong to the same species and that D. suillum is a later subjective synonym of Azospira oryzae.

A novel thermophilic bacterium, strain K14T, capable of degrading poly(3-hydroxybutyrate) as well as copolymers containing 3-hydroxybutyrate and 3-mercaptopropionate linked by thioester bonds, was isolated. 16S rDNA sequence analysis showed that strain DhA-71, a dehydroabietic acid-degrading bacterium, was the nearest phylogenetic neighbour and that both strains should be placed as members of a newly created genus, Schlegelella gen. nov., in the Rubrivivax subgroup of the β-Proteobacteria. Strain K14T (=LMG 21644T=DSM 15344T) is proposed as the type strain of Schlegelella thermodepolymerans gen. nov., sp. nov. Its phylogenetic, morphological, biochemical and chemotaxonomic characteristics are described in detail.

Two Gram-negative, motile, non-spore-forming, yellow-pigmented and slightly halophilic strains (SW-46T and SW-52) were isolated from sea water of the East Sea, Korea, and subjected to a polyphasic taxonomic study. Strains SW-46T and SW-52 were characterized chemotaxonomically by having ubiquinone-10 (Q-10) as the predominant respiratory lipoquinone and C18 : 1 ω7c as the major fatty acid. Their DNA G+C content was 64·0–64·1 mol%. Strains SW-46T and SW-52 showed 1 bp difference in their 16S rDNA sequences and a mean DNA–DNA relatedness level of 94·4 %. Phylogenetic analysis based on 16S rDNA sequences showed that strains SW-46T and SW-52 fall within the α-subclass of the Proteobacteria and form a coherent cluster with Erythrobacter longus, Erythrobacter litoralis and Erythrobacter citreus. Levels of 16S rDNA similarity between strains SW-46T and SW-52 and the type strains of these three Erythrobacter species were 96·5–97·9 %. Levels of DNA–DNA relatedness between strains SW-46T and SW-52 and the type strains of E. longus, E. litoralis and E. citreus were 3·6–14·7 %. Therefore, on the basis of phenotypic properties, phylogeny and genomic data, strains SW-46T and SW-52 should be placed in the genus Erythrobacter as a novel species, for which the name Erythrobacter flavus sp. nov. is proposed. The type strain is SW-46T (=KCCM 41642T =JCM 11808T).