- Volume 55, Issue 6, 2005

A haloalkaliphilic archaeon (strain DS12T) isolated from Lake Zabuye, the Tibetan Plateau, China, was characterized to elucidate its taxonomy. The strain was aerobic, chemo-organotrophic, and grew optimally at 40 °C, pH 9·5–10·0 and 3·4 M NaCl. Cells of strain DS12T were non-motile cocci and stained Gram-variable. The major polar lipids of strain DS12T were diphytanyl and phytanyl-sesterterpanyl diether derivatives of phosphatidylglycerol and phosphatidylglycerol phosphate methyl ester. No glycolipids were detected. Phylogenetic analysis revealed that the strain formed a distinct lineage within the family Halobacteriaceae. The low 16S rRNA gene sequence similarity values to its closest relatives (91·5–92·5 %) and its signature bases both suggest that the strain has no close affinity with any members of the family Halobacteriaceae with validly published names. Therefore, it is proposed that strain DS12T (=AS 1.3240T=JCM 11890T) represents the type strain of a novel species in a new genus, Halalkalicoccus tibetensis gen. nov., sp. nov.

A cell-fusing hyperthermophilic archaeon was isolated from hydrothermal fluid obtained from Suiyo Seamount of the Izu-Bonin Arc. The isolate, TS1T, is an irregular coccus, usually 0·5–2 μm in diameter and motile with a polar tuft of flagella. Cells in the exponential phase of growth fused at room temperature in the presence of DNA-intercalating dye to become as large as 5 μm in diameter. Fused cells showed dark spots that moved along in the cytoplasm. Large cells with a similar appearance were also observed upon culture at 87 °C, suggesting the occurrence of similar cell fusions during growth. Transmission electron microscopy revealed that cells in the exponential phase possessed a thin and electron-lucent cell envelope that could be lost subsequently during culture. The fragile cell envelope must be related to cell fusion. The cells grew at 57–90 °C, pH 5·2–8·7 and at NaCl concentrations of 1·5–4·5 %, with the optima being 87 °C, pH 6·5 and 2·5 % NaCl. The isolate was an anaerobic chemo-organotroph that grew on either yeast extract or tryptone as the sole growth substrate. The genomic DNA G+C content was 53·9 mol%. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that the isolate was closely related to Thermococcus species. However, no significant DNA–DNA hybridization was observed between genomic DNA of strain TS1T and phylogenetically related Thermococcus species. We propose that isolate TS1T represents a novel species, Thermococcus coalescens sp. nov., with the name reflecting the cell fusion activity observed in the strain. The type strain is TS1T (=JCM 12540T=DSM 16538T).

Three novel strains of methylotrophic methanogens were isolated from Skan Bay, Alaska, by using anaerobic cultivation techniques. The water was 65 m deep at the sampling site. Strains AK-4 (=OCM 774), AK-5T (=OCM 775T=DSM 17273T) and AK-9 (=OCM 793) were isolated from the sulfate-reducing zone of the sediments. Each of the strains was a non-motile coccus and occurred singly. Cells grew with trimethylamine as a catabolic substrate and strain AK-4 could also catabolize methanol. Yeast extract and trypticase peptones were not required for growth, but their addition to the culture medium slightly stimulated growth. Each of the strains grew at temperatures of 5–28 °C; they were slight halophiles and grew fastest in the neutral pH range. Analysis of the 16S rRNA gene sequences indicated that strain AK-4 was most closely related to Methanosarcina baltica. DNA–DNA hybridization studies showed 88 % relatedness, suggesting that strain AK-4 represents a novel strain within this species. Strains AK-5T and AK-9 had identical 16S rRNA gene sequences that were most closely related to the sequence of Methanococcoides burtonii (99·8 % sequence similarity). DNA–DNA hybridization studies showed that strains AK-5T and AK-9 are members of the same species (88 % relatedness value), but strain AK-5T had a DNA–DNA relatedness value of only 55 % to Methanococcoides burtonii. This indicates that strains AK-5T and AK-9 should be considered as members of a novel species in the genus Methanococcoides. We propose the name Methanococcoides alaskense sp. nov., with strain AK-5T (=OCM 775T=DSM 17273T) as the type strain.

A novel extremely halophilic strain, isolated from Aibi salt lake, Xin-Jiang, China, was subjected to polyphasic taxonomic characterization. This strain, designated AB14T, is neutrophilic, motile and requires at least 10 % (w/v) NaCl for growth. Strain AB14T grows at 24–58 °C, with optimal growth at 42–45 °C. Mg2+ is not required, but growth is observed in MgCl2 concentrations as high as 1·0 M. Strain AB14T possesses the diphytanyl (C20C20) and phytanyl-sesterterpanyl diether (C20C25) derivatives of phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester and mannose-2,6 disulfate 1→2 glucose-glycerol diether. The genomic DNA G+C content is 66·6 mol%. The 16S rRNA gene sequence similarity values of strain AB14T with its nearest phylogenetic neighbours (Haloterrigena thermotolerans and Haloterrigena turkmenica) are 98·6 and 96·0 %, respectively. DNA–DNA hybridization revealed 54 % relatedness between strain AB14T and Haloterrigena thermotolerans JCM 11050T and 21 % between strain AB14T and Haloterrigena turkmenica JCM 9101T. It is therefore proposed that strain AB14T represents a novel species, for which the name Haloterrigena saccharevitans sp. nov. is proposed. The type strain is AB14T (=AS 1.3730T=JCM 12889T).

Two proteolytic, strictly anaerobic bacterial strains (TB107T and TB6-6) were isolated from the granule sludge of an upflow anaerobic sludge blanket reactor treating brewery wastewater. The strains were Gram-negative, non-spore-forming and motile. Cells were rod-shaped (0·6–0·9×1·9–2·2 μm). Growth of the strains was observed at 20–45 °C and pH 6·0–9·7. The strains were proteolytic. Yeast extract, peptone, pyruvate, glycine and l-arginine could be used as carbon and energy sources. Weak growth was also observed with tryptone, l-serine, l-threonine and l-alanine as carbon and energy sources. Both strains did not use any of the tested carbohydrates, alcohols and fatty acids except pyruvate. Acetic acid and NH3 were produced from yeast extract, peptone and l-arginine, and propionic acid was also produced from yeast extract. Pyruvate was converted to acetic acid and CO2. Gelatin was not hydrolysed. Indole and H2S were not produced. The two strains did not grow in medium containing 20 % bile. Addition of strain TB107T to a syntrophic propionate-degrading co-culture accelerated the propionate-degradation rate. The predominant cellular fatty acid was the branched-chain fatty acid anteiso-C15 : 0 (46·21 %). The genomic DNA G+C contents of strains TB107T and TB6-6 were 46·6 and 48·9 mol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the two strains represent a new phyletic sublineage within the Cytophaga–Flavobacterium–Bacteroides (CFB) group, with <91 % 16S rRNA gene sequence similarity to the closest species with validly published names. On the basis of polyphasic evidence from this study, a new genus and species, Proteiniphilum acetatigenes gen. nov., sp. nov., is proposed, with strain TB107T (=JCM 12891T=AS 1.5024T) as the type strain.

A novel thermophilic, sulfur-oxidizing Gram-negative bacterium, designated strain SS-5T, was isolated from the Calcite Hot Springs in Yellowstone National Park, USA. The cells were motile rods (1·2–2·8 μm long and 0·6–0·8 μm wide). The new isolate was a facultative heterotroph capable of using elemental sulfur or thiosulfate as an electron donor and O2 (1–18 %; optimum 6 %, v/v) as an electron acceptor. Hydrogen did not support growth. The isolate grew autotrophically with CO2. In addition, strain SS-5T utilized various organic carbon sources such as yeast extract, tryptone, sugars, amino acids and organic acids. Growth was observed between 55 and 78 °C (optimum 70 °C; 3·5 h doubling time), pH 6·0 and 8·0 (optimum pH 7·5), and 0 and 0·6 % (w/v) NaCl (optimum 0 %). The G+C content of the genomic DNA was 32 mol%. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that the isolate was a member of the genus Sulfurihydrogenibium. On the basis of the physiological and molecular characteristics of the new isolate, we propose the name Sulfurihydrogenibium yellowstonense sp. nov. with SS-5T (=JCM 12773T=OCM 840T) as the type strain. In addition, emended descriptions of the genus Sulfurihydrogenibium, Sulfurihydrogenibium subterraneum and Sulfurihydrogenibium azorense are proposed.

An orange-pigmented, Gram-negative, non-motile, rod-shaped, strictly aerobic bacterium (UST030701-324T) was isolated from a microbial mat in an estuary in the Bahamas. The DNA G+C content was 33·6 mol%. Predominant fatty acids were i15 : 0, i16 : 0, i17 : 0 3-OH, and summed feature 3, comprising i15 : 0 2-OH and/or 16 : 1ω7c. MK-6 was the only respiratory quinone. Flexirubin-type pigments were not produced. Phylogenetic analysis based on 16S rRNA gene sequences placed UST030701-324T in a distinct lineage in the family Flavobacteriaceae with less than 90·7 % sequence similarity to the members of the nearest genus, Psychroflexus. UST030701-324T can be distinguished from other members of Flavobacteriaceae by a number of chemotaxonomic and phenotypic characteristics. It is thus proposed that UST030701-324T represents a novel taxon designated Nonlabens tegetincola gen. nov., sp. nov. The type strain is UST030701-324T (=NRRL B-41136T=JCM 12886T).

A bacterial strain, designated 4M15T, was isolated from cotton-waste composts used as mushroom cultivation in South Korea. Properties of this isolate were studied on the basis of physiological and biochemical characteristics, fatty acid profile, isoprenoid quinone, DNA G+C content and phylogenetic position based on 16S rRNA gene sequence analysis. The strain was found to form a distinct phylogenetic lineage related to the family ‘Flexibacteraceae’ within the phylum ‘Bacteroidetes’. No recognized species showed >85 % 16S rRNA gene sequence similarity to strain 4M15T. The fatty acid profile of strain 4M15T included C16 : 1 ω7c/iso-C15 : 0 2-OH (30·5 %), iso-C15 : 0 (24·2 %), iso-C15 : 0 2-OH/C16 : 1 ω7c (15·9), iso-C17 : 0 3-OH (10·5 %) and C16 : 0 (5·6 %). The major isoprenoid quinone was menaquinone MK-7. The DNA G+C content was 33·0 mol%. Cells were Gram-negative, strictly aerobic, rod-shaped, non-motile, catalase-positive, oxidase-positive and flexirubin-positive. The strain hydrolysed aesculin, gelatin, starch and tyrosine. Several phenotypic tests could be used to differentiate strain 4M15T from other members of the family ‘Flexibacteraceae’. On the basis of the data presented, strain 4M15T should be assigned to the phylum ‘Bacteroidetes’ as a novel genus and species, for which the name Leadbetterella byssophila gen. nov., sp. nov. is proposed. The type strain is 4M15T (=KACC 11308T=DSM 17132T).

A new bacterial strain, designated SHRIMPT, isolated from the haemolymph of the Pacific white shrimp, Penaeus vannamei, was serologically distinct from other spiroplasmas. Cells of this strain were helical in form and variable in length. Examination by electron microscopy revealed wall-less cells delineated by a single cytoplasmic membrane. The organisms grew well in M1D media supplemented with 2 % NaCl. Strain SHRIMPT grew at temperatures of 20–37 °C, with optimum growth occurring at 28 °C. The strain catabolized glucose and hydrolysed arginine, but did not hydrolyse urea. The G+C content of the DNA was 29±1 mol%. Strain SHRIMPT (=ATCC BAA-1082T=CAIM 1252T) is designated the type strain of a novel species, Spiroplasma penaei sp. nov., which represents a new subgroup (I-9) of the group I spiroplasmas.

Two Gram-negative, non-motile, non-spore-forming, slightly halophilic bacterial strains, DSW-1T and DSW-21, were isolated from sea water and subjected to a polyphasic taxonomic study. They grew optimally at 30 °C and in the presence of 2 % (w/v) NaCl. Strains DSW-1T and DSW-21 were characterized chemotaxonomically as containing MK-6 as the predominant menaquinone and iso-C15 : 0, iso-C17 : 0 3-OH and iso-C15 : 1 as the major fatty acids. Their DNA G+C content was 38 mol%. Strains DSW-1T and DSW-21 exhibited four nucleotide differences in their 16S rRNA gene sequences and possessed a mean DNA–DNA relatedness level of 78 %. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strains DSW-1T and DSW-21 formed a distinct lineage within the family Flavobacteriaceae. The 16S rRNA gene sequences of strains DSW-1T and DSW-21 had similarity levels of less than 92·2 % to the sequences of other members of the family Flavobacteriaceae. There were some differences in phenotypic properties between the two strains and Cellulophaga species, the nearest phylogenetic neighbours. On the basis of phenotypic, phylogenetic and genetic data, strains DSW-1T (=KCTC 12391T=DSM 17200T) and DSW-21 were classified in a novel genus and species, Dokdonia donghaensis gen. nov., sp. nov.

The taxonomic position of a novel marine, heterotrophic, aerobic, pigmented, non-motile bacterium isolated from sea water was determined. 16S rRNA gene sequence analysis revealed that strain KMM 6143T is a member of the genus Cyclobacterium. The results of DNA–DNA hybridization experiments supported by phenotypic and chemotaxonomic data showed that the isolate represents a novel species of the genus Cyclobacterium, for which the name Cyclobacterium amurskyense sp. nov. is proposed. The type strain is KMM 6143T (=KCTC 12363T=LMG 23026T).

Two gamma- and UV-radiation-tolerant, Gram-negative, rod-shaped bacterial strains, VCD115T and VCD117, were isolated from a mixture of sand samples collected in the Sahara Desert in Morocco and Tunisia, after exposure of the sand to 15 kGy gamma radiation. Phylogenetic analysis based on 16S rRNA gene sequences and DNA–DNA hybridizations showed that VCD115T and VCD117 are members of a novel species belonging to the genus Deinococcus, with Deinococcus grandis as its closest relative. The DNA G+C contents of VCD115T and VCD117 are 59·8 and 60·6 mol%, respectively. The major fatty acids (straight-chain 15 : 1, 16 : 1, 17 : 1 and 16 : 0), polar lipids (dominated by phosphoglycolipids and glycolipids) and quinone type (MK-8) support the affiliation to the genus Deinococcus. The strains did not grow on rich medium such as trypticase soy broth (TSB), but did grow as whitish colonies on tenfold-diluted TSB. The genotypic and phenotypic properties allowed differentiation of VCD115T and VCD117 from recognized Deinococcus species. Strains VCD115T and VCD117 are therefore identified as representing a novel species, for which the name Deinococcus deserti sp. nov. is proposed, with the type strain VCD115T (=DSM 17065T=LMG 22923T).

Spiroplasma sp. strain SMAT, isolated in Poland from white satin moth larvae, Leucoma salicis L. (Lepidoptera: Lymantriidae), was serologically distinct from other Spiroplasma species, groups or subgroups. Dark-field microscopy of the cells revealed the classical helical shape and subsequent transmission electron microscopy revealed cells surrounded by only a single cell membrane (lacking a cell wall). Growth of strain SMAT occurred in M1D medium at 30 °C. Strain SMAT catabolized both glucose and arginine, but did not hydrolyse urea. The G+C content of the DNA was 24±1 mol% as determined by melting temperature analysis. Serological analysis revealed a very weak cross-reactivity (positive reaction only up to a 1 : 80 dilution) with two Spiroplasma strains, 277F (Spiroplasma sp. group I-4) and LB-12 (Spiroplasma sp. group I-5). Strain SMAT (=ATCC BAA-521T=NBRC 100392T) is designated the type strain of a novel species, Spiroplasma leucomae sp. nov. (class Mollicutes: order Entomoplasmatales: family Spiroplasmataceae).

During 2003, surveys of sugarcane yellow leaf disease and papaya bunchy top-like disease were carried out on plantations in Havana province, Cuba, to determine the roles of weeds and Auchenorrhyncha insects in the epidemiology of these diseases. More than 250 plant and insect samples were collected and indexed by using a nested PCR for phytoplasma 16S rDNA with the generic primer pairs P1/P7 and R16F2n/R16R2. The PCR products were further characterized by restriction fragment length polymorphism using HaeIII, AluI, Sau3AI, Tru9I, HhaI, HpaII and TaqI endonucleases, giving patterns that distinguished them from those of the other reference phytoplasmas analysed. Phylogenetic analysis of 16S rRNA gene sequences identified the phytoplasmas present in sugarcane (Saccharum officinarum L.), Cynodon dactylon L., Conyza canadensis L. Cronq., Sorghum halepense L. Pers., Macroptilium lathyroides L. Urb., Saccharosydne saccharivora (Westwood) and Cedusa spp., and those in papaya (Carica papaya L.) and Empoasca papayae, as two novel provisional phytoplasma species. We propose that these phytoplasmas should be given Candidatus status, as ‘Candidatus Phytoplasma graminis' and ‘Candidatus Phytoplasma caricae’, respectively.

A yellow-pigmented, Gram-negative, slowly gliding, rod-shaped, strictly aerobic bacterium (UST040801-001T) was isolated from marine sediment. The DNA G+C content was 39·9 mol%. The predominant fatty acids were a15 : 0, i15 : 0, i15 : 0 3-OH, i17 : 1ω9c, i17 : 0 3-OH and summed feature 3, comprising i15 : 0 2-OH and/or 16 : 1ω7c (altogether representing 76·2 % of the total). MK-6 was the only respiratory quinone. Flexirubin-type pigments were not produced. Phylogenetic analysis based on 16S rRNA gene sequences indicated that Gramella echinicola KMM 6050T (the only species in the genus) was the closest relative of UST040801-001T, sharing 98·0 % sequence similarity. The DNA–DNA relatedness between UST040801-001T and Gramella echinicola KMM 6050T was 13 %. Strain UST040801-001T can be distinguished from G. echinicola by means of 11 phenotypic traits. The results of molecular and phenotypic analyses suggested that UST040801-001T represents a novel species of Gramella. The name Gramella portivictoriae sp. nov. is proposed for this bacterium, with UST040801-001T (=NRRL 41137T=JCM 13192T) as the type strain.

Two Gram-negative, anaerobic, non-spore-forming, rod-shaped organisms were isolated from a swine-manure storage pit. Based on morphological and biochemical criteria, the strains were tentatively identified as belonging to the genus Bacteroides but they did not appear to correspond to any recognized species of the genus. Comparative 16S rRNA gene sequencing studies showed that the strains were related closely to each other and confirmed their placement in the genus Bacteroides, but sequence divergence values of >10 % from reference Bacteroides species demonstrated that the organisms from manure represent a novel species. Based on biochemical criteria and molecular genetic evidence, it is proposed that the unknown isolates from manure be assigned to a novel species of the genus Bacteroides, as Bacteroides coprosuis sp. nov. The type strain is PC139T (=CCUG 50528T=NRRL B-41113T).