- Volume 49, Issue 1, 1999

Sequencing of 16S rRNA genes and phylogenetic analysis of Methanogenium tationis DSM 2702T (OCM 43T) (T = type strain) and Methanogenium liminatans GKZPZT ( = DSM 4140T) as well as other members of the family Methanomicrobiaceae revealed that both species belong to a separate line of descent within this family. In addition, a new strain of Methanogenium liminatans, strain BM1 ( DSM 10196), was isolated from a butyrate-degrading, fluidized bed reactor and characterized. Cells of both species are mesophilic, highly irregular cocci that use H2/CO2 and formate for growth and methanogenesis. In addition, Methanogenium liminatans strains GKZPZT and BM1 used 2-propanol/CO2, 2-butanol/CO2 and cyclopentanol/CO2. Both species contained diether and tetraether lipids. The polar lipids comprised amino-phosphopentanetetrol derivatives, which appear to be characteristic lipids within the family Methanomicrobiaceae. The pattern of glycolipids, phosphoglycolipids and amino-phosphoglycolipids was consistent with the assignment of these two species to a taxon within the family Methanomicrobiaceae, but also permitted them to be distinguished from other higher taxa within this family. The G+C contents of the DNA of Methanogenium tationis and Methanogenium liminatans were 54 and 60 mol% (T m and HPLC), respectively. On the basis of the data presented, the transfer of Methanogenium tationis and Methanogenium liminatans to the genus Methanofollis gen. nov. as Methanofollis tationis comb. nov. and Methanofollis liminatans comb, nov., respectively, is proposed, with Methanofollis tationis as the type species.

Two haloalkaliphilic archaea were isolated from a soda lake in Tibet. The two strains, designated A33T and GA33T, were Gram-negative, pleomorphic, flat, non-motile and strictly aerobic. Growth required at least 12% NaCI. Growth was between pH 8·0 and pH 11 with an optimum at pH 9·0–9·5. Cells were chemo-organotrophic. Polar lipids were C20–C25 derivatives of phosphatidylglycerol and phosphatidylglycerol phosphate. The nucleotide sequences of the 16S rRNA genes from the two strains were obtained by the analysis of the cloned rDNAs. On 16S rRNA phylogenetic trees, the two strains formed a monophyletic cluster. They differed from their closest neighbours, Halobacterium trapanicum and Natrialba asiatica, in polar lipid composition, as well as physiological and phenotypic characteristics. DNA-DNA hybridization indicated that the two strains belonged to different species of the same genus. The results indicated that the strains A33T and GA33T should be classified in a new genus Natronorubrum gen. nov. as Natronorubrum bangense sp. nov. (strain A33T) and Natronorubrum tibetense sp. nov. (strain GA33T).

Thirteen Gram-negative bacterial isolates were recovered from diseased pigeons and were tentatively classified as Riemerella anatipestifer-like strains based on conventional phenotypic features and disease symptoms. Phenotypic characteristics that differentiated the pigeon isolates from R. anatipestifer included their greyish-white to beige pigment formation on Columbia blood agar and the hydrolysis of aesculin. Furthermore, R. anatipestifer strains have thus far not been reported in pigeons. The phenotypic differences together with the unique host range of the new isolates have prompted the inclusion of these strains in a polyphasic taxonomic study. Extensive phenotypic examination, PAGE of total proteins and GC analysis of fatty acid contents revealed that the pigeon isolates constitute a homogeneous cluster, distinct from the R. anatipestifer reference strains. The phylogenetic position of representative strains was examined by using DNA–rRNA hybridizations and indicated that this taxon belongs to the genus Riemerella. Finally, DNA-binding values confirmed that the strains constitute a separate species for which the name Riemerella columbina sp. nov. is proposed. Strain LMG 11607T was selected as the type strain. Clinical observations suggest that these organisms are involved in pathogenesis of respiratory diseases, similar to those associated with R. anatipestifer infections. However, the role of co-factors and the interaction with other agents are unknown.

Reference strains of Burkholderia cocovenenans and Burkholderia vandii were compared with strains of other Burkholderia species using SDS-PAGE of whole cell proteins, DNA-DNA hybridization and extensive biochemical characterization. Burkholderia gladioli and B. cocovenenans were indistinguishable in the chemotaxonomic and biochemical analyses. Burkholderia plantarii and B. vandii had indistinguishable whole-cell protein patterns but the B. vandii type strain differed from B. plantarii strains in several biochemical tests. The DNA-DNA binding levels (higher than 70%) indicated that (i) B. gladioli and B. cocovenenans, and (ii) B. plantarii and B. vandii each represent a single species. It is concluded that B. cocovenenans and B. vandii are junior synonyms of B. gladioli and B. plantarii. respectively.

Fifty-five Chinese isolates from nodules of Amorpha fruticosa were characterized and compared with the type strains of the species and genera of bacteria which form nitrogen-fixing symbioses with leguminous host plants. A polyphasic approach, which included RFLP of PCR-amplified 16S rRNA genes, multilocus enzyme electrophoresis (MLEE), DNA-DNA hybridization, 16S rRNA gene sequencing, electrophoretic plasmid profiles, cross-nodulation and a phenotypic study, was used in the comparative analysis. The isolates originated from several different sites in China and they varied in their phenotypic and genetic characteristics. The majority of the isolates had moderate to slow growth rates, produced acid on YMA and harboured a 930 kb symbiotic plasmid (pSym). Five different RFLP patterns were identified among the 16S rRNA genes of all the isolates. Isolates grouped by PCR-RFLP of the 16S rRNA genes were also separated into groups by variation in MLEE profiles and by DNA-DNA hybridization. A representative isolate from each of these DNA homology groups had a separate position in a phylogenetic tree as determined from sequencing analysis of the 16S rRNA genes. A new species, Mesorhizobium amorphae, is proposed for the majority of the isolates, which belonged to a moderately slow- to slow-growing, acid-producing group based upon their distinct phylogenetic position, their unique electrophoretic type, their low DNA homology with reference strains representing the species within the genus Mesorhizobium and their distinct phenotypic features. Strain ACCC 19665 was chosen as the type strain for M. amorphae sp. nov.

The present paper presents the conclusions of a polyphasic investigation of the taxonomy of the trehalose-negative [Pasteurella] haemolytica complex. Clusters previously identified by ribotyping and multilocus enzyme electrophoresis (MEE) have been evaluated by 16S rRNA sequencing and DNA–DNA hybridizations. Results obtained by the different techniques were highly related and indicated that the [P.] haemolytica complex contains distinct genetic and phenotypic groups. At least seven species were outlined, five of which were named. We refrained in formal naming of more groups until additional strains are characterized. Five 16S rRNA clusters were identified corresponding to distinct lineages previously outlined by MEE. Within 16S rRNA cluster I two distinct genotypic groups have been outlined in addition to [P.] haemolytica sensu stricto (biogroup 1). Each of the clusters II, III, IV and V represent at least one new species. The investigations underline that [P.] haemolytica sensu stricto only contains strains that do not ferment l-arabinose even though they are referred to as ‘biotype A’ of [P.] haemolytica. The five 16S rRNA clusters identified had a common root relative to the other species within the family Pasteurellaceae, and the overall sequence similarity among these five clusters was higher than what is observed within the existing genera of the family. The allocation of the trehalose-negative [P.] haemolytica complex to a new genus seems to be indicated. Based on the polyphasic investigation performed a new genus Mannheimia is proposed for the trehalose-negative [P.] haemolytica complex. At the present stage two previously named species are transferred to this new genus and three new species are described. [P.] haemolytica is reclassified as Mannheimia haemolytica comb. nov., whereas Pasteurella granulomatis, Bisgaard taxon 20 and [P.] haemolytica biovar 3J are reclassified and combined in the species Mannheimia granulomatis comb. nov. Mannheimia glucosida sp. nov. corresponds to [P.] haemolytica biogroups 3A-3H and the β-glucosidase and meso-inositol-positive strains of [P.] haemolytica biogroup 9. All typable strains within M. glucosida belong to serotype 11. Mannheimia ruminalis sp. nov. consists of strains previously classified as Bisgaard taxon 18 and [P.] haemolytica biogroup 8D. Finally, Mannheimia varigena sp. nov. includes [P.] haemolytica biogroup 6 as well as Bisgaard taxon 15 and Bisgaard taxon 36. The type strains are NCTC 9380T (M. haemolytica), ATCC 49244T (M. granulomatis), CCUG 38457T = P925T (M. glucosida). CCUG 38470T = HPA92T (M. ruminalis) and CCUG 38462T = 177T (M. varigena).

Two strains of a hitherto undescribed Gram-positive catalase-negative, facultatively anaerobic coccus isolated from human sources were characterizec by phenotypic and molecular taxonomic methods. Comparative 16S rRNA gene sequencing studies demonstrated the unknown strains were genealogically identical, and constitute a new line close to, but distinct from, the genera Facklamia and Globicatella. The unknown bacterium was readily distinguished from Facklamia species and Globicatella sanguinus by biochemical tests and electrophoretic analysis of whole-cell proteins. Based on phylogenetic and phenotypic evidence it is proposed that the unknown bacterium be classifiec as lgnavigranum ruoffiae gen. nov., sp. nov. The type strain of lgnavigranum ruoffiae is CCUG 37658T.

A novel halophilic fermentative bacterium has been isolated from the black sediment below a gypsum crust and a microbial mat in hypersaline ponds of Mediterranean salterns. Morphologically, physiologically and genetically this organism belongs to the genus Haloanaerobacter. Haloanaerobacter strain SG 3903T (T = type strain) is composed of non-sporulating long flexible rods with peritrichous flagella, able to grow in the salinity range of 5–30% NaCI, with an optimum at 14–15%. The strain grows by fermenting carbohydrates or by using the Stickland reaction with either serine or H2 as electron donors and glycine-betaine as acceptor, which is reduced to trimethylamine. The two species described so far in the genus Haloanaerobacter are not capable of Stickland reaction with glycine-betaine + serine; however, Haloanaerobacter chitinovorans can use glycine-betaine with H2 as electron donor. Strain SG 3903T thus represents the first described strain in the genus Haloanaerobacter capable of the Stickland reaction with two amino acids. Although strain SG 3903T showed 67% DNA-DNA relatedness to H. chitinovorans, it is physiologically sufficiently different from the two described species to be considered as a new species which has been named Haloanaerobacter salinarius sp. nov.

Eight Gram-negative, aerobic, pointed and budding bacteria were isolated from various depths of the hypersaline, heliothermal and meromictic Ekho Lake (Vestfold Hills, East Antarctica). The cells contained storage granules and daughter cells could be motile. Bacteriochlorophyll a was sometimes produced, but production was repressed by constant dim light. The strains tolerated a wide range of temperature, pH, concentrations of artificial seawater and NaCI, but had an absolute requirement for sodium ions. Glutamate was metabolized with and without an additional source of combined nitrogen. The dominant fatty acid was C18:1; other characteristic fatty acids were C18:2, C12:0 2-OH, C12:1 3-OH, C16:1 C16:0 and C18:0. The main polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine. The DNA G+C base composition was 62–64 mol%. 16S rRNA gene sequence comparisons showed that the isolates were phylogenetically close to the genera Antarctobacter, ‘Marinosulfonomonas’, Octadecabacter, Sagittula, Sulfitobacter and Roseobacter. Morphological, physiological and genotypic differences to these previously described and distinct genera support the description of a new genus and a new species, Roseovarius tolerans gen. nov., sp. nov. The type strain is EL-172T ( = DSM 11457T).

A new halophilic sulfate-reducing bacterium, strain GSL-But2T, was isolated from surface sediment of the Southern arm of the Great Salt Lake, UT, USA. The organism grew with a number of straight-chain fatty acids (C4–C16), 2-methylbutyrate, l-alanine and pyruvate as electron donors. Butyrate was oxidized incompletely to acetate. Sulfate, but not sulfite or thiosulfate, served as an electron acceptor. Growth was observed between 2 and 19% (w/v) NaCl with an optimum at 4–5% (w/v) NaCl. The optimal temperature and pH for growth were around 34 °C and pH 6·5–7·3, respectively. The generation time under optimal conditions in defined medium was around 28 h, compared to 20 h in complex medium containing yeast extract. The G+C content was 35·0 mol%. 16S rRNA gene sequence analysis revealed that strain GSL-But2T belongs to the family Desulfobacteriaceae within the delta-subclass of the Proteobacteria and suggested that strain GSL-But2T represents a member of a new genus. The name Desulfocella halophila gen. nov., sp. nov. is proposed for this organism. The type strain of D. halophila is strain GSL-But2T (= DSM 11763T = ATCC 700426T).

Phenotypic and phylogenetic studies were performed with two strains (OCh 317Tand OCh 318; T = type strain) of aerobic chemoheterotrophic bacteriochlorophyll-containing bacteria isolated from water of a saline lake located on the west coast of Australia. Both strains were Gram-negative, short rods and were motile by means of polar flagella. Catalase, oxidase, nitrate reductase, phosphatase and urease were produced. The cells utilized d-glucose, citrate, glycolate, pyruvate and ethanol. Acids were produced from l-arabinose, d-fructose, d-galactose, d-glucose, d-ribose and d-xylose. The strains could grow in media containing 0·5–7·5% NaCl. Bacteriochlorophyll a was synthesized under aerobic conditions. The results of 16S rRNA gene sequence comparisons revealed that strain OCh 317Trepresented a new lineage in the α-3 group of the class Proteobacteria. Strains OCh 317Tand OCh 318 were identified as strains of the same species because of their very similar phenotypic characteristics and their previously described high DNA-DNA homology. Therefore, it was concluded that the two strains should be assigned to a new genus and species, for which the name Rubrimonas cliftonensis is proposed. The type strain is OCh 317T(= JCM 10189T).

Strain 130ZT was isolated from the bovine rumen. It is a facultatively anaerobic, pleomorphic, Gram-negative rod. It exhibits a ‘Morse code’ form of morphology, which is characteristic of the genus Actinobacillus. Strain 130ZT is a capnophilic, osmotolerant succinogen that utilizes a broad range of sugars. It accumulates high concentrations of succinic acid (> 70gl-1). Strain 130ZT is positive for catalase, oxidase, alkaline phosphatase and β-galactosidase, but does not produce indole or urease. Acid but no gas is produced from d-glucose and d-fructose. 16S rRNA sequence analysis places strain 130ZT within the family Pasteurellaceae; the most closely related members of the family Pasteurellaceae have 16S rRNA similarities of 95.5% or less with strain 130ZT. Strain 130ZTwas compared with Actinobacillus lignieresii and the related Bisgaard Taxa 6 and 10. Based upon morphological and biochemical properties, strain 130ZT is most similar to members of the genus Actinobacillus within the family Pasteurellaceae. It is proposed that strain 130ZT be classified as a new species, Actinobacillus succinogenes. The type strain of Actinobacillus succinogenes sp. nov. is ATCC 55618T.

The phylogenetic position of Vibrio iliopiscarius was inferred by the maximum-likelihood, maximum-parsimony and neighbour-joining methods on the basis of almost complete 16S rRNA gene sequences. The results showed that this species falls into the same cluster as Photobacterium species and is clearly distinct from other Vibrio species. Its nearest phylogenetic neighbour is Photobacterium phosphoreum. From these results, it is concluded that V. iliopiscarius should be reclassified as Photobacterium iliopiscarium comb. nov. the type strain of which is PS1T(= ATCC 51760T).

A secondary intracellular symbiotic bacterium was isolated from the haemolymph of the tsetse fly Glossina morsitans morsitans and cultured in Aedes albopictus cell line C6/36. Pure-culture isolation of this bacterium was achieved through the use of solid-phase culture under a microaerobic atmosphere. After isolation of strain M1T, a range of tests was performed to determine the phenotypic properties of this bacterium. Considering the results of these tests, along with the phylogenetic position of this micro-organism, it is proposed that this intracellular symbiont from G. m. morsitans should be classified in a new genus Sodalis gen. nov., as Sodalis glossinidius gen. nov., sp. nov. Strain M1Tis the type strain for this new species.

The strain E-396T, isolated from soil, was Gram-negative, aerobic, orangepigmented, rod-shaped, motile by peritrichous flagella and astaxanthin-producing. This organism produced carotenoids, mainly astaxanthin, and did not produce bacteriochlorophyll. The ubiquinone system was Q-10. Analysis of the 16S rRNA sequence of strain E-396T showed it to be a member of the α-3 subclass of the Proteobacteria, forming a cluster with the species of the genus Paracoccus. On the basis of the production of orange pigments and motility by peritrichous flagella, together with DNA-DNA reassociation data, it is concluded that the new isolate should be classified into a new species of the genus Paracoccus, Paracoccus carotinifaciens sp. nov. The type strain is E-396T (= IFO 16121T).