Showing papers in "International Journal of Systematic and Evolutionary Microbiology in 2009"

An update of the structure and 16S rRNA gene sequence-based definition of higher ranks of the class Actinobacteria, with the proposal of two new suborders and four new families and emended descriptions of the existing higher taxa.

Abstract: The higher ranks of the class Actinobacteria were proposed and described in 1997. At each rank, the taxa were delineated from each other solely on the basis of 16S rRNA gene sequence phylogenetic clustering and taxon-specific 16S rRNA signature nucleotides. In the past 10 years, many novel members have been assigned to this class while, at the same time, some members have been reclassified. The new 16S rRNA gene sequence information and the changes in phylogenetic positions of some taxa influence decisions about which 16S rRNA nucleotides to define as taxon-specific. As a consequence, the phylogenetic relationships of Actinobacteria at higher levels may need to be reconstructed. Here, we present new 16S rRNA signature nucleotide patterns of taxa above the family level and indicate the affiliation of genera to families. These sets replace the signatures published in 1997. In addition, Actinopolysporineae subord. nov. and Actinopolysporaceae fam. nov. are proposed to accommodate the genus Actinopolyspora, Kineosporiineae subord. nov. and Kineosporiaceae fam. nov. are proposed to accommodate the genera Kineococcus, Kineosporia and Quadrisphaera, Beutenbergiaceae fam. nov. is proposed to accommodate the genera Beutenbergia, Georgenia and Salana and Cryptosporangiaceae fam. nov. is proposed to accommodate the genus Cryptosporangium. The families Nocardiaceae and Gordoniaceae are proposed to be combined in an emended family Nocardiaceae. Emended descriptions are also proposed for most of the other higher taxa.

Construction of an interactive online phytoplasma classification tool, iPhyClassifier, and its application in analysis of the peach X-disease phytoplasma group (16SrIII).

Abstract: Phytoplasmas, the causal agents of numerous plant diseases, are insect-vector-transmitted, cell-wall-less bacteria descended from ancestral low-G+C-content Gram-positive bacteria in the Bacillus–Clostridium group. Despite their monophyletic origin, widely divergent phytoplasma lineages have evolved in adaptation to specific ecological niches. Classification and taxonomic assignment of phytoplasmas have been based primarily on molecular analysis of 16S rRNA gene sequences because of the inaccessibility of measurable phenotypic characters suitable for conventional microbial characterization. In the present study, an interactive online tool, iPhyClassifier, was developed to expand the efficacy and capacity of the current 16S rRNA gene sequence-based phytoplasma classification system. iPhyClassifier performs sequence similarity analysis, simulates laboratory restriction enzyme digestions and subsequent gel electrophoresis and generates virtual restriction fragment length polymorphism (RFLP) profiles. Based on calculated RFLP pattern similarity coefficients and overall sequence similarity scores, iPhyClassifier makes instant suggestions on tentative phytoplasma 16Sr group/subgroup classification status and ‘Candidatus Phytoplasma’ species assignment. Using iPhyClassifier, we revised and updated the classification of strains affiliated with the peach X-disease phytoplasma group. The online tool can be accessed at http://www.ba.ars.usda.gov/data/mppl/iPhyClassifier.html.

Proposed minimal standards for describing new taxa of aerobic, endospore-forming bacteria.

Abstract: Minimal standards for describing new taxa within the aerobic endospore-forming bacteria are proposed, following Recommendation 30b of the Bacteriological Code (1990 Revision). These minimal standards are recommended as guidelines to assist authors in the preparation of descriptions for novel taxa. They encourage broad polyphasic characterization and the construction of descriptions that are practically useful in routine diagnostic laboratories. The proposals have been endorsed by the Subcommittee on the Taxonomy of the Genus Bacillus and Related Organisms of the International Committee on Systematics of Prokaryotes.

Taxon K, a complex within the Burkholderia cepacia complex, comprises at least two novel species, Burkholderia contaminans sp. nov. and Burkholderia lata sp. nov.

Abstract: The aim of the present study was to re-examine the taxonomic position and structure of taxon K (also known as group K) within the Burkholderia cepacia complex (Bcc). For this purpose, a representative set of strains was examined by a traditional polyphasic taxonomic approach, by multilocus sequence typing (MLST) analysis and by analysis of available whole-genome sequences. Analysis of the recA gene sequence revealed three different lineages, designated recA-I recA-II and recA-III. DNA-DNA hybridization experiments demonstrated that recA-I and recA-II isolates each represented a single novel species. However, DNA-DNA hybridization values of recA-II strains towards recA-III strains and among recA-III strains were at the threshold level for species delineation. By MLST, recA-I isolates were clearly distinguished from the others and represented a distinct lineage referred to as MLST-I, whereas recA-II and recA-III isolates formed a second MLST lineage referred to as MLST-II. A divergence value of 3.5% was obtained when MLST-I was compared with MLST-II. The internal level of concatenated sequence divergence within MLST-I and MLST-II was 1.4 and 2.7%, respectively; by comparison with the level of concatenated sequence divergence in established Bcc species, these data demonstrate that the MLST-I and MLST-II lineages represent two distinct species within the Bcc. The latter conclusion was supported by comparison of the whole-genome average nucleotide identity (ANI) level of MLST-I and MLST-II strains with strains of established Bcc species and by a whole-genome-based phylogenetic analysis. We formally propose to classify taxon K bacteria from the MLST-I and MLST-II lineages as Burkholderia contaminans sp. nov. (with strain J2956(T) =LMG 23361(T) =CCUG 55526(T) as the type strain) and Burkholderia lata sp. nov. (with strain 383(T) =ATCC 17760(T) =LMG 22485(T) =CCUG 55525(T) as the type strain), respectively. The MLST approach was confirmed as a valuable instrument in polyphasic taxonomic studies; more importantly, the cumulative data for about 1000 Bcc isolates analysed demonstrate that the 3% concatenated sequence divergence level correlates with the 70% DNA-DNA hybridization or 95% whole-genome ANI threshold levels for species delineation.

Zhihengliuella alba sp. nov., and emended description of the genus Zhihengliuella.

Abstract: A Gram-positive, non-motile, white-pigmented, short rod actinobacterium, designated YIM 90734T, was isolated from a saline soil sample collected from Ganjiahu Suosuo Forest National Nature Reserve in Xinjiang province, north-west China, and its taxonomic position was investigated by using a polyphasic approach. Strain YIM 90734T grew optimally at 28–37 °C and pH 6.0–8.0 and in 5 % (w/v) NaCl. The peptidoglycan type was A4α, l-Lys–l-Ala–l-Glu and tyvelose and mannose were the major cell-wall sugars. The predominant menaquinones were MK-10 and MK-9. Major cellular fatty acids (>10 % of total) were anteiso-C15 : 0 and anteiso-C17 : 0. The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, one unknown phospholipid and two unknown glycolipids. The DNA G+C content was 70.3 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain YIM 90734T belonged to the genus Zhihengliuella. The 16S rRNA gene sequence similarity between strain YIM 90734T and the type strain of the only recognized Zhihengliuella species, Zhihengliuella halotolerans, was 97.7 %. However, the level of DNA–DNA relatedness of the two strains was 41.4 %. The DNA–DNA relatedness data and differential phenotypic properties, together with the phylogenetic distinctiveness, demonstrated that strain YIM 90734T could be differentiated from Z. halotolerans. On the basis of the data presented, strain YIM 90734T is considered to represent a novel species of the genus Zhihengliuella, for which the name Zhihengliuella alba sp. nov. is proposed. The type strain is YIM 90734T (=KCTC 19375T=DSM 21143T). The description of the genus Zhihengliuella has also been emended.

Phylogeny and molecular taxonomy of the Bacillus subtilis species complex and description of Bacillus subtilis subsp. inaquosorum subsp. nov.

Abstract: The Bacillus subtilis species complex is a tight assemblage of closely related species. For many years, it has been recognized that these species cannot be differentiated on the basis of phenotypic characteristics. Recently, it has been shown that phylogenetic analysis of the 16S rRNA gene also fails to differentiate species within the complex due to the highly conserved nature of the gene, yet DNA-DNA hybridization values fall well below 70 % for the same species comparisons. As a complementary approach, we propose that phylogenetic analysis of multiple protein-coding loci can be used as a means to detect and differentiate novel Bacillus taxa. Indeed, our phylogenetic analyses revealed the existence of a previously unknown group of strains closely related to, but distinct from, Bacillus subtilis subsp. spizizenii. Results of matrix-assisted laser desorption ionization-time of flight mass spectrometry analyses revealed that the group produces a novel surfactin-like lipopeptide with mass m/z 1120.8 that is not produced by the other currently recognized subspecies. In addition, the group displayed differences in the total cellular content of the fatty acids C(16 : 0) and iso-C(17 : 1)omega10c that distinguish it from the closely related B. subtilis subsp. spizizenii. Consequently, the correlation of these novel phenotypic traits with the phylogenetic distinctiveness of this previously unknown subspecies group showed that phylogenetic analysis of multiple protein-coding loci can be used as a means to detect and differentiate novel Bacillus taxa. Therefore, we propose that this new group should be recognized as representing a novel taxon, Bacillus subtilis subsp. inaquosorum subsp. nov., with the type strain NRRL B-23052(T) (=KCTC 13429(T)=BGSC 3A28(T)).

Phylogenetic structure of Xanthomonas determined by comparison of gyrB sequences.

Abstract: Previously, we have produced a phylogeny of species type strains from the plant-pathogenic genus Xanthomonas based on gyrB sequences. To evaluate this locus further for species and infraspecies identification, we sequenced an additional 203 strains comprising all the pathovar reference strains (which have defined plant hosts), 67 poorly characterized pathovars, currently classified as Xanthomonas campestris, and 59 unidentified xanthomonads. The well-characterized pathovars grouped either in clades containing their respective species type strain or in clades containing species related to Xanthomonas axonopodis. The Xanthomonas euvesicatoria, Xanthomonas perforans and Xanthomonas alfalfae species complex, Xanthomonas fuscans and Xanthomonas citri were discriminated as X. axonopodis-related clades and comprised a large proportion of unidentified strains as well as 80 pathovars representing all the X. axonopodis pathovars and many poorly characterized pathovars, greatly increasing the plant host ranges of the constituent species. Most xanthomonads from these three large clades were isolated from a taxonomically diverse range of plant hosts, including many weed species, from field systems in India, suggesting that these lineages became established and diversified in agricultural areas in this region. The majority of these xanthomonads had minimal sequence diversity, consistent with rapid and highly extensive pathovar diversification that has occurred in relatively recent times. Low-intensity farming practices may have provided conditions conducive to pathovar development, and evidence for pathovar diversification within other regional angiosperm floras is discussed. The gyrB locus was sufficiently discriminating to identify diversity within many species. Seven branches or clades were sufficiently distinct to be considered as potential novel species. This study has provided a comprehensive xanthomonad classification framework and has firmly established gyrB sequencing as a rapid and efficient identification tool.

Dehalogenimonas lykanthroporepellens gen. nov., sp. nov., a reductively dehalogenating bacterium isolated from chlorinated solvent-contaminated groundwater

Abstract: Two recently reported bacterial strains that are able to reductively dehalogenate polychlorinated aliphatic alkanes, including 1,2,3-trichloropropane, 1,2-dichloropropane, 1,1,2,2-tetrachloroethane, 1,1,2-trichloroethane and 1,2-dichloroethane, were further characterized to clarify their taxonomic position. The two strains, designated BL-DC-8 and BL-DC-9T, were mesophilic, non-spore-forming, non-motile, Gram-negative staining and strictly anaerobic. Cells were irregular cocci, 0.3–0.6 μm in diameter. The two strains were resistant to ampicillin and vancomycin. Hydrogen was utilized as an electron donor. The genomic DNA G+C content of strains BL-DC-8 and BL-DC-9T was 54.0 and 53.8 mol%, respectively. The major cellular fatty acids were C18 : 1 ω9c, C16 : 1 ω9c, C16 : 0 and C14 : 0. Phylogenetic analyses based on 16S rRNA gene sequences indicated that the strains cluster within the phylum Chloroflexi, but are related only distantly to all recognized taxa in the phylum. Morphological, physiological and chemotaxonomic traits as well as phylogenetic analysis support the conclusion that these two strains represent a novel species of a new genus in the phylum Chloroflexi, for which the name Dehalogenimonas lykanthroporepellens gen. nov., sp. nov. is proposed. The type strain of Dehalogenimonas lykanthroporepellens is BL-DC-9T (=ATCC BAA-1523T =JCM 15061T).

'Candidatus Liberibacter solanacearum', associated with plants in the family Solanaceae

Abstract: A liberibacter (isolate NZ082226) was detected in a symptomatic tomato plant and subsequently in five other members of the family Solanaceae: capsicum, potato, tamarillo, cape gooseberry and chilli. Phylogenetic analyses of the 16S rRNA gene sequence, the deduced amino acid sequence of the rplJ gene and a partial nucleotide sequence of the β operon indicated that isolate NZ082226 represents a novel candidate species of ‘Candidatus Liberibacter’, for which the name ‘Candidatus Liberibacter solanacearum’ is proposed.

Janibacter hoylei sp. nov., Bacillus isronensis sp. nov. and Bacillus aryabhattai sp. nov., isolated from cryotubes used for collecting air from the upper atmosphere

Abstract: Three novel bacterial strains, PVAS-1(T), B3W22(T) and B8W22(T), were isolated from cryotubes used to collect air samples at altitudes of between 27 and 41 km. Based on phenotypic characteristics, chemotaxonomic features, DNA-DNA hybridization with the nearest phylogenetic neighbours and phylogenetic analysis based on partial 16S rRNA gene sequences (PVAS-1(T), 1196 nt; B3W22(T), 1541 nt; B8W22(T), 1533 nt), the three strains were identified as representing novel species, and the names proposed are Janibacter hoylei sp. nov. (type strain PVAS-1(T) =MTCC 8307(T) =DSM 21601(T) =CCUG 56714(T)), Bacillus isronensis sp. nov. (type strain B3W22(T) =MTCC 7902(T) =JCM 13838(T)) and Bacillus aryabhattai sp. nov. (type strain B8W22(T) =MTCC 7755(T) =JCM 13839(T)).

Phylogeny and taxonomy of a diverse collection of Bradyrhizobium strains based on multilocus sequence analysis of the 16S rRNA gene, ITS region and glnII, recA, atpD and dnaK genes.

Abstract: The genus Bradyrhizobium encompasses a variety of bacteria that can live in symbiotic and endophytic associations with legumes and non-legumes, and are characterized by physiological and symbiotic versatility and broad geographical distribution. However, despite indications of great genetic variability within the genus, only eight species have been described, mainly because of the highly conserved nature of the 16S rRNA gene. In this study, 169 strains isolated from 43 different legumes were analysed by rep-PCR with the BOX primer, by sequence analysis of the 16S rRNA gene and the 16S-23S rRNA intergenic transcribed spacer (ITS) and by multilocus sequence analysis (MLSA) of four housekeeping genes, glnII, recA, atpD and dnaK. Considering a cut-off at a level of 70 % similarity, 80 rep-PCR profiles were distinguished, which, together with type strains, were clustered at a very low level of similarity (24 %). In both single and concatenated analyses of the 16S rRNA gene and ITS sequences, two large groups were formed, with bootstrap support of 99 % in the concatenated analysis. The first group included the type and/or reference strains of Bradyrhizobium japonicum, B. betae, B. liaoningense, B. canariense and B. yuanmingense and B. japonicum USDA 110, and the second group included strains related to Bradyrhizobium elkanii USDA 76(T), B. pachyrhizi PAC48(T) and B. jicamae PAC68(T). Similar results were obtained with MLSA of glnII, recA, atpD and dnaK. Greatest variability was observed when the atpD gene was amplified, and five strains related to B. elkanii revealed a level of variability never reported before. Another important observation was that a group composed of strains USDA 110, SEMIA 5080 and SEMIA 6059, all isolated from soybean, clustered in all six trees with high bootstrap support and were quite distinct from the clusters that included B. japonicum USDA 6(T). The results confirm that MLSA is a rapid and reliable way of providing information on phylogenetic relationships and of identifying rhizobial strains potentially representative of novel species.

Ferrimicrobium acidiphilum gen. nov., sp. nov. and Ferrithrix thermotolerans gen. nov., sp. nov.: heterotrophic, iron-oxidizing, extremely acidophilic actinobacteria.

Abstract: Two novel extremely acidophilic, iron-oxidizing actinobacteria were isolated, one from a mine site in North Wales, UK (isolate T23T), and the other from a geothermal site in Yellowstone National Park, Wyoming, USA (Y005T). These new actinobacteria belong to the subclass Acidimicrobidae, and in contrast to the only other classified member of the subclass (Acidimicrobium ferrooxidans), both isolates were obligate heterotrophs. The mine site isolate was mesophilic and grew as small rods, while the Yellowstone isolate was a moderate thermophile and grew as long filaments, forming macroscopic flocs in liquid media. Both isolates accelerated the oxidative dissolution of pyrite in yeast extract-amended cultures, but neither was able to oxidize reduced forms of sulfur. Ferrous iron oxidation enhanced growth yields of the novel mesophilic actinobacterium T23T, though this was not confirmed for the Yellowstone isolate. Both isolates catalysed the dissimilatory reduction of ferric iron, using glycerol as electron donor, in oxygen-free medium. Based on comparative analyses of base compositions of their chromosomal DNA and of their 16S rRNA gene sequences, the isolates are both distinct from each other and from Acidimicrobium ferrooxidans, and are representatives of two novel genera. The names Ferrimicrobium acidiphilum gen. nov., sp. nov. and Ferrithrix thermotolerans gen. nov., sp. nov. are proposed for the mesophilic and moderately thermophilic isolates, respectively, with the respective type strains T23T (=DSM 19497T=ATCC BAA-1647T) and Y005T (=DSM 19514T=ATCC BAA-1645T).

Acinetobacter beijerinckii sp. nov. and Acinetobacter gyllenbergii sp. nov., haemolytic organisms isolated from humans.

Abstract: The taxonomic status of 24 haemolytic, non-glucose acidifying Acinetobacter strains that did not belong to any previously described species was investigated by means of a polyphasic approach. Using AFLP fingerprinting, amplified rDNA restriction analysis and phenotypic characterization, the strains were classified into two phenetically coherent groups (comprising 15 and 9 strains) that were distinct from each other and from all known Acinetobacter species. Confirmation that these groups formed two separate lineages within the genus Acinetobacter was obtained from comparative analysis of partial sequences of the gene encoding the β-subunit of RNA polymerase in all strains and also from 16S rRNA gene sequence analysis of representative strains. Previously published DNA–DNA reassociation data for some of the strains used also supported the species rank for both groups, for which the names Acinetobacter beijerinckii sp. nov. and Acinetobacter gyllenbergii sp. nov. are proposed. The strains of A. beijerinckii sp. nov. originated from human and animal specimens and from various environmental sources, whereas those of A. gyllenbergii sp. nov. were isolated exclusively from human clinical specimens. The phenotypic characteristics most useful for the differentiation of these species from other Acinetobacter species that comprise haemolytic strains were the inability of A. beijerinckii sp. nov. to grow on l-arginine and the ability of A. gyllenbergii sp. nov. to grow on azelate. The type strain of A. beijerinckii sp. nov. is NIPH 838T (=LUH 4759T=CCUG 51249T=CCM 7266T=58aT) and the type strain of A. gyllenbergii sp. nov. is NIPH 2150T (=RUH 422T=CCUG 51248T=CCM 7267T=1271T).

Description of ‘Synergistetes’ phyl. nov. and emended description of the phylum ‘Deferribacteres’ and of the family Syntrophomonadaceae, phylum ‘Firmicutes’

Abstract: The number of bacterial phyla has greatly increased in the past decade. Among them, a candidate division named ‘Synergistes’ was proposed in a phylogenetic study on the global diversity of bacteria. We previously described the genus Jonquetella and suggested that it belonged to this not yet well-delineated candidate phylum. 16S rRNA gene based-phylogeny studies were conducted using four reconstruction methods and 599 sequences forming five datasets were used in an alternative treeing approach. These analyses indicated that the genera Aminiphilus, Aminobacterium, Aminomonas, Anaerobaculum, Dethiosulfovibrio, Jonquetella, Synergistes, Thermanaerovibrio and Thermovirga should be grouped in the same high-level taxon. This taxon was shown to be a phylum-rank lineage in the domain Bacteria and, because of the prior use of the name Synergistes for a genus, the name ‘Synergistetes’ is proposed for this candidate phylum. We also propose an emended delineation of the phylum ‘Deferribacteres’, which is now only represented by the family Deferribacteriaceae. The emended family Syntrophomonadaceae is limited to the genera Pelospora, Syntrophomonas, Syntrophothermus and Thermosyntropha.

Mycoplasma leachii sp. nov. as a new species designation for Mycoplasma sp. bovine group 7 of Leach, and reclassification of Mycoplasma mycoides subsp. mycoides LC as a serovar of Mycoplasma mycoides subsp. capri.

Abstract: The Mycoplasma mycoides cluster consists of six pathogenic mycoplasmas causing disease in ruminants, which share many genotypic and phenotypic traits. The M. mycoides cluster comprises five recognized taxa: Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC), M. mycoides subsp. mycoides Large Colony (MmmLC), M. mycoides subsp. capri (Mmc), Mycoplasma capricolum subsp. capricolum (Mcc) and M. capricolum subsp. capripneumoniae (Mccp). The group of strains known as Mycoplasma sp. bovine group 7 of Leach (MBG7) has remained unassigned, due to conflicting data obtained by different classification methods. In the present paper, all available data, including recent phylogenetic analyses, have been reviewed, resulting in a proposal for an emended taxonomy of this cluster: (i) the MBG7 strains, although related phylogenetically to M. capricolum, hold sufficient characteristic traits to be assigned as a separate species, i.e. Mycoplasma leachii sp. nov. (type strain, PG50(T) = N29(T) = NCTC 10133(T) = DSM 21131(T)); (ii) MmmLC and Mmc, which can only be distinguished by serological methods and are related more distantly to MmmSC, should be combined into a single subspecies, i.e. Mycoplasma mycoides subsp. capri, leaving M. mycoides subsp. mycoides (MmmSC) as the exclusive designation for the agent of contagious bovine pleuropneumonia. A taxonomic description of M. leachii sp. nov. and emended descriptions of M. mycoides subsp. mycoides and M. mycoides subsp. capri are presented. As a result of these emendments, the M. Mycoides cluster will hereafter be composed of five taxa comprising three subclusters, which correspond to the M. mycoides subspecies, the M. capricolum subspecies and the novel species M. leachii

Kosmotoga olearia gen. nov., sp. nov., a thermophilic, anaerobic heterotroph isolated from an oil production fluid.

Abstract: A novel thermophilic, heterotrophic bacterium, strain TBF 19.5.1(T), was isolated from oil production fluid at the Troll B oil platform in the North Sea. Cells of strain TBF 19.5.1(T) were non-motile rods with a sheath-like structure, or toga. The strain was Gram-negative and grew at 20-80 degrees C (optimum 65 degrees C), pH 5.5-8.0 (optimum pH 6.8) and NaCl concentrations of 10-60 g l(-1) (optimum 25-30 g l(-1)). For a member of the order Thermotogales, the novel isolate is capable of unprecedented growth at low temperatures, with an optimal doubling time of 175 min (specific growth rate 0.24 h(-1)) and a final optical density of >1.4 when grown on pyruvate at 37 degrees C. Various carbohydrates, proteinaceous compounds and pyruvate served as growth substrates. Thiosulfate, but not elemental sulfur, enhanced growth of the isolate. Sulfate also enhanced growth, but sulfide was not produced. The strain grew in the presence of up to approximately 15 % oxygen, but only if cysteine was included in the medium. Growth of the isolate was inhibited by acetate, lactate and propionate, while butanol and malate prevented growth. The major fermentation products formed on maltose were hydrogen, carbon dioxide and acetic acid, with traces of ethanol and propionic acid. The G+C content of the genomic DNA was 42.5 mol%. Phylogenetic analyses of the 16S and 23S rRNA gene sequences as well as 29 protein-coding ORFs placed the strain within the bacterial order Thermotogales. Based on the phylogenetic analyses and the possession of a variety of physiological characteristics not previously found in any species of this order, it is proposed that the strain represents a novel species of a new genus within the family Thermotogaceae, order Thermotogales. The name Kosmotoga olearia gen. nov., sp. nov. is proposed. The type strain of Kosmotoga olearia is TBF 19.5.1(T) (=DSM 21960(T) =ATCC BAA-1733(T)).

Bradyrhizobium pachyrhizi sp. nov. and Bradyrhizobium jicamae sp. nov., isolated from effective nodules of Pachyrhizus erosus.

Abstract: Several strains isolated from the legume Pachyrhizus erosus were characterized on the basis of diverse genetic, phenotypic and symbiotic approaches. These novel strains formed two groups closely related to Bradyrhizobium elkanii according to their 16S rRNA gene sequences. Strains PAC48T and PAC68T, designated as the type strains of these two groups, presented 99.8 and 99.1 % similarity, respectively, in their 16S rRNA gene sequences with respect to B. elkanii USDA 76T. In spite of these high similarity values, the analysis of additional phylogenetic markers such as atpD and glnII genes and the 16S–23S intergenic spacer (ITS) showed that strains PAC48T and PAC68T represented two separate novel species of the genus Bradyrhizobium with B. elkanii as their closest relative. Phenotypic differences among the novel strains isolated from Pachyrhizus and B. elkanii were found regarding the assimilation of carbon sources and antibiotic resistance. All these differences were congruent with DNA–DNA hybridization analysis which revealed 21 % genetic relatedness between strains PAC48T and PAC68T and 46 % and 25 %, respectively, between these strains and B. elkanii LMG 6134T. The nodD and nifH genes of strains PAC48T and PAC68T were phylogenetically divergent from those of bradyrhizobia species that nodulate soybean. Soybean was not nodulated by the novel Pachyrhizus isolates. Based on the genotypic and phenotypic data obtained in this study, the new strains represent two novel species for which the names Bradyrhizobium pachyrhizi sp. nov. (type strain PAC48T=LMG 24246T=CECT 7396T) and Bradyrhizobium jicamae sp. nov. (type strain PAC68T=LMG 24556T=CECT 7395T) are proposed.

Butyricimonas synergistica gen. nov., sp. nov. and Butyricimonas virosa sp. nov., butyric acid-producing bacteria in the family 'Porphyromonadaceae' isolated from rat faeces.

Abstract: Two bacterial strains, designated MT01T and MT12T, isolated from rat faeces were characterized by using a polyphasic taxonomic approach that included analysis of their phenotypic and biochemical features, cellular fatty acid profiles, menaquinone profiles and phylogeny based on 16S rRNA gene sequences. The 16S rRNA gene sequence analysis showed that these strains were members of the family ‘Porphyromonadaceae’. The strains shared 94 % 16S rRNA gene sequence similarity with each other and were related to Odoribacter splanchnicus NCTC 10825T (86–87 % sequence similarity). The strains consisted of obligately anaerobic, non-pigmented, non-spore-forming, non-motile, Gram-negative rods. Growth of the strains was inhibited on medium containing 20 % bile. The two strains produced significant levels of butyric and isobutyric acids as end products from glucose. Although the major cellular fatty acid of these two strains and O. splanchnicus JCM 15291T was iso-C15 : 0, strains MT01T and MT12T showed a higher level of iso-C15 : 0 (66 and 74 %, respectively) than did O. splanchnicus JCM 15291T (48 %). In addition, the ratios of iso-C15 : 0 to anteiso-C15 : 0 in whole-cell methanolysates of the two isolates were very much higher than that of O. splanchnicus JCM 15291T. The major menaquinone of the isolates was MK-10. This menaquinone composition was different from those of other genera of the family ‘Porphyromonadaceae’, such as Barnesiella (predominant menaquinones: MK-11 and MK-12), Odoribacter (MK-9), Paludibacter (MK-8), Parabacteroides (MK-9 and MK-10), Porphyromonas (MK-9 and MK-10) and Tannerella (MK-10 and MK-11). Menaquinone composition is therefore an important chemotaxonomic characteristic of these micro-organisms. Strains MT01T and MT12T have DNA G+C contents of 46 mol%. On the basis of these data, strains MT01T and MT12T represent two novel species of a novel genus, for which the names Butyricimonas synergistica gen. nov., sp. nov. and Butyricimonas virosa sp. nov., respectively, are proposed. The type strains of B. synergistica and B. virosa are MT01T (=JCM 15148T =CCUG 56610T) and MT12T (=JCM 15149T=CCUG 56611T), respectively.

Caldisericum exile gen. nov., sp. nov., an anaerobic, thermophilic, filamentous bacterium of a novel bacterial phylum, Caldiserica phyl. nov., originally called the candidate phylum OP5, and description of Caldisericaceae fam. nov., Caldisericales ord. nov. and Caldisericia classis nov.

Abstract: An anaerobic, thermophilic, thiosulfate-reducing bacterium, strain AZM16c01T, isolated from a hot spring in Japan [Mori, K., Sunamura, M., Yanagawa, K., Ishibashi, J., Miyoshi, Y., Iino, T., Suzuki, K. & Urabe, T. (2008). Appl Environ Microbiol 74, 6223–6229] was characterized in detail. The 16S rRNA gene sequence analysis had revealed that strain AZM16c01T was the first cultivated representative of the candidate phylum OP5. The cells were multicellular filaments with a single polar flagellum. The strain contained iso-C17 : 0 as the major fatty acid and menaquinone-8(H6), menaquinone-8(H8) and menaquinone-8(H10) as the respiratory quinones. The G+C content of the genomic DNA of strain AZM16c01T was 34.6 mol%. Optimum growth was obtained at 65 °C, pH 6.5 and in the absence of NaCl, with a doubling time of 10.6 h. On the basis of the results of phylogenetic analysis based on the 16S rRNA gene sequence and the characterization of the strain in this study, we propose the name Caldisericum exile gen. nov., sp. nov. for strain AZM16c01T (=NBRC 104410T=DSM 21853T). In addition, we propose the new phylum name Caldiserica phyl. nov. for the candidate phylum OP5 represented by C. exile gen. nov., sp. nov., and Caldisericaceae fam. nov., Caldisericales ord. nov. and Caldisericia classis nov.

Providencia sneebia sp. nov. and Providencia burhodogranariea sp. nov., isolated from wild Drosophila melanogaster.

Abstract: Multiple isolates of the genus Providencia were obtained from the haemolymph of wild-caught Drosophila melanogaster fruit flies. Sixteen isolates were distinguished from the six previously described species based on 16S rRNA gene sequences. These isolates belonged to two distinct groups, which we propose each comprise previously undescribed species. Two isolates, designated A T and B T , were characterized by DNA sequences of the fusA, lepA, leuS, gyrB and ileS housekeeping genes, whole-genome DNA–DNA hybridizations with their nearest relatives and utilization of substrates for metabolism. The closest phylogenetic relatives of strain A T are strain B T (86.9% identity for the housekeeping genes) and Providencia stuartii DSM 4539 T (86.0% identity). The closest phylogenetic relatives of strain B T are strain A T (86.9% identity) and P. stuartii DSM 4539 T (86.6% identity). The type strains of described species in this genus shared between 84.1 and 90.1% identity for these sequences. DNA–DNA hybridization between the strain pairs A T –B T ,A T –P. stuartii DSM 4539 T and B T –P. stuartii DSM 4539 T all resulted in less than 25% relatedness. In addition, patterns of utilization of amygdalin, arbutin, aesculin, salicin, D-sorbitol, trehalose, inositol, D-adonitol and D-galactose distinguish strains A T and B T from other members of this genus. Strains A T and B T therefore represent novel species, for which the names Providencia sneebia sp. nov. (type strain A T 5DSM 19967 T 5ATCC BAA-1589 T ) and Providencia burhodogranariea sp. nov. (type strain B T 5DSM 19968 T 5ATCC BAA-1590 T ) are

Emended descriptions of genera of the family Halobacteriaceae.

Abstract: The family Halobacteriaceae currently contains 96 species whose names have been validly published, classified in 27 genera (as of September 2008) In recent years, many novel species have been added to the established genera but, in many cases, one or more properties of the novel species do not agree with the published descriptions of the genera Authors have often failed to provide emended genus descriptions when necessary Following discussions of the International Committee on Systematics of Prokaryotes Subcommittee on the Taxonomy of Halobacteriaceae, we here propose emended descriptions of the genera Halobacterium, Haloarcula, Halococcus, Haloferax, Halorubrum, Haloterrigena, Natrialba, Halobiforma and Natronorubrum

Arcobacter mytili sp. nov., an indoxyl acetate-hydrolysis-negative bacterium isolated from mussels.

Abstract: Three Arcobacter isolates, recovered from mussels (genus Mytilus), and one isolate from brackish water in Catalonia (north-east Spain) showed a novel pattern using a recently described identification method for members of the genus Arcobacter, 16S rRNA gene RFLP. Enterobacterial repetitive intergenic consensus PCR fingerprinting demonstrated that the three isolates from mussels belonged to two genotypes and that the fourth isolate from water belonged to a third genotype. Analysis of the 16S rRNA and rpoB gene sequences showed that the new isolates formed a separate lineage within the genus Arcobacter. This was also confirmed by the low DNA-DNA relatedness values (16-30 %) of the isolates with the type strains of recognized Arcobacter species. Hydrolysis of indoxyl acetate, a characteristic trait for all species of the genus Arcobacter, was negative for the novel isolates. The susceptibility of the novel isolates to cefoperazone, together with the lack of urease production and nitrate reduction, further enabled them to be differentiated from recognized Arcobacter species based on physiological characteristics. Genotypic and phenotypic characteristics indicated that the new isolates represent a novel species of the genus Arcobacter, for which the name Arcobacter mytili sp. nov. is proposed, with the type strain F2075(T) (=CECT 7386(T) =LMG 24559(T)). The DNA G+C content of strain F2075(T) was 26.9 mol%.

Clostridium clariflavum sp. nov. and Clostridium caenicola sp. nov., moderately thermophilic, cellulose-/cellobiose-digesting bacteria isolated from methanogenic sludge.

Abstract: Two novel anaerobic, moderately thermophilic and cellulose-/cellobiose-digesting bacteria, EBR45T and EBR596T, were isolated from anaerobic sludge of a cellulose-degrading methanogenic bioreactor. Phylogenetic analysis based on 16S rRNA gene sequences indicated that these strains belonged to cluster III within the low-G+C-content Gram-positive bacteria. The close relatives of EBR45T were Clostridium straminisolvens DSM 16021T (sequence identity, 94.6 %) and Clostridium thermocellum DSM 1237T (93.4 %). The closest relative of EBR596T was Clostridium stercorarium DSM 8532T (95.9 %). Both isolates were rod-shaped sporulators, growing optimally at 60 °C. EBR45T was Gram-staining-reaction-variable and non-motile, formed bright-yellow colonies on solid media, and grew on a relatively narrow range of carbohydrates including cellulose and cellobiose. EBR596T was Gram-staining-reaction-negative and motile, formed glossy white colonies and grew on cellobiose and various carbohydrates except cellulose. Major fatty acid compositions were 16 : 0 iso, 16 : 0 and 16 : 0 dimethylacetal (strain EBR45T) and 15 : 0 iso, 16 : 0 iso, 15 : 0 anteiso and 17 : 0 anteiso (strain EBR596T). The DNA G+C contents were 36.9 mol% (EBR45T) and 51.1 mol% (EBR596T). Based on the phenotypic and phylogenetic data and genomic distinctiveness, strains EBR45T and EBR596T represent two novel species, for which the names Clostridium clariflavum sp. nov. (type strain EBR45T =DSM 19732T =NBRC 101661T) and Clostridium caenicola sp. nov. (type strain EBR596T =DSM 19027T =NBRC 102590T) are proposed.

Description of seven candidate species affiliated with the phylum Actinobacteria, representing planktonic freshwater bacteria.

Abstract: Actinobacteria comprise a substantial fraction of the bacterioplankton in freshwater lakes and streams. Numerous cultivation-independent investigations have retrieved actinobacterial 16S rRNA gene sequences from such habitats. The taxa detected in freshwater habitats are usually absent from terrestrial and marine systems. So far, none of the indigenous freshwater lineages is represented by a taxon with a validly published name. The seven organisms for which Candidatus status is described here were isolated from freshwater lakes and ponds located in tropical, subtropical and temperate climatic zones. Phylogenetic analyses demonstrated that they are affiliated with one of the actinobacterial lineages indigenous to freshwater bacterioplankton. The seven novel taxa could only be cultivated to date as mixed cultures that also contain non-actinobacterial strains. Due to the lack of pure cultures, I propose to establish the candidate species ‘Candidatus Planktoluna difficilis’, ‘Candidatus Aquiluna rubra’, ‘Candidatus Flaviluna lacus’, ‘Candidatus Rhodoluna limnophila’, ‘Candidatus Rhodoluna planktonica’, ‘Candidatus Rhodoluna lacicola’ and ‘Candidatus Limnoluna rubra’ for these taxa.

Ferruginibacter alkalilentus gen. nov., sp. nov. and Ferruginibacter lapsinanis sp. nov., novel members of the family 'Chitinophagaceae' in the phylum Bacteroidetes, isolated from freshwater sediment.

Abstract: Six Gram-negative, non-gliding, rod-shaped bacterial strains isolated from freshwater sediment were subjected to polyphasic analyses to determine their taxonomic positions. Analysis of the 16S rRNA gene sequences of the six strains revealed that they represent two separate genomic species in a new lineage within the phylum Bacteroidetes, related to members of the family 'Chitinophagaceae', in which they were most closely related to members of the genus Terrimonas (93.4-90.5 %). Two of the isolates, HU1-GD23(T) and HU1-HG42(T), had a sequence similarity of 96.2 %, with DNA G+C contents of 39.4 and 38.5 mol%, respectively. They possessed MK-7 as the predominant respiratory quinone and contained high amounts of iso-pentadecanoic acid and 3-hydroxy-iso-heptadecanoic acid in their cell envelopes, properties shared by members of the family 'Chitinophagaceae'. They were well differentiated from other members of the family 'Chitinophagaceae' by additional physiological and biochemical characteristics. In conclusion, strains HU1-GD23(T) (=KCTC 22306(T)=LMG 24312(T)) and HU1-HG42(T) (=KCTC 22305(T)=LMG 24324(T)) are considered to represent two novel species of a novel genus, Ferruginibacter gen. nov., for which the names Ferruginibacter alkalilentus sp. nov. and Ferruginibacter lapsinanis sp. nov., are proposed, respectively.

Phylogenomics and protein signatures elucidating the evolutionary relationships among the Gammaproteobacteria

Abstract: The class Gammaproteobacteria, which forms one of the largest groups within bacteria, is currently distinguished from other bacteria solely on the basis of its branching in phylogenetic trees. No molecular or biochemical characteristic is known that is unique to the class Gammaproteobacteria or its different subgroups (orders). The relationship among different orders of gammaproteobacteria is also not clear. In this study, we present detailed phylogenomic and comparative genomic analyses on gammaproteobacteria that clarify some of these issues. Phylogenetic trees based on concatenated sequences for 13 and 36 universally distributed proteins were constructed for 45 members of the class Gammaproteobacteria covering 13 of its 14 orders. In these trees, species from a number of the subgroups formed distinct clades and their relative branching order was indicated as follows (from the most recent to the earliest diverging): Enterobacteriales >Pasteurellales >Vibrionales, Aeromonadales >Alteromonadales >Oceanospirillales, Pseudomonadales >Chromatiales, Legionellales, Methylococcales, Xanthomonadales, Cardiobacteriales, Thiotrichales. Four conserved indels in four widely distributed proteins that are specific for gammaproteobacteria are also described. A 2 aa deletion in 5′-phosphoribosyl-5-aminoimidazole-4-carboxamide transformylase (AICAR transformylase; PurH) was a distinctive characteristic of all gammaproteobacteria (except Francisella tularensis). Two other conserved indels (a 4 aa deletion in RNA polymerase β-subunit and a 1 aa deletion in ribosomal protein L16) were found uniquely in various species of the orders Enterobacteriales, Pasteurellales, Vibrionales, Aeromonadales and Alteromonadales, but were not found in other gammaproteobacteria. Lastly, a 2 aa deletion in leucyl-tRNA synthetase was commonly present in the above orders of the class Gammaproteobacteria and also in some members of the order Oceanospirillales. The presence of the conserved indels in these gammaproteobacterial orders indicates that species from these orders shared a common ancestor that was separate from other bacteria, a suggestion that is supported by phylogenetic studies. Systematic blastp searches were also conducted on various open reading frames (ORFs) in the genome of Escherichia coli K-12. These analyses identified 75 proteins that were unique to most members of the class Gammaproteobacteria or were restricted to species from some of its main orders (Enterobacteriales; Enterobacteriales and Pasteurellales; Enterobacteriales, Pasteurellales, Vibrionales, Aeromonadales and Alteromonadales; and the Enterobacteriales, Pasteurellales, Vibrionales, Aeromonadales, Alteromonadales, Oceanospirillales and Pseudomonadales etc.). The genes for these proteins have evolved at various stages during the evolution of gammaproteobacteria and their species distribution pattern, in conjunction with other results presented here, provide valuable information regarding the evolutionary relationships among these bacteria.

Pantoea vagans sp. nov., Pantoea eucalypti sp. nov., Pantoea deleyi sp. nov. and Pantoea anthophila sp. nov.

Abstract: Bacteria isolated from eucalyptus leaves and shoots showing symptoms of blight and die-back collected in Uganda, Uruguay and Argentina and from maize displaying brown stalk rot symptoms in South Africa were tentatively placed in the genus Pantoea on the basis of phenotypic and biochemical tests. These isolates, together with two strains (LMG 2558 and LMG 2560) previously assigned to Pantoea agglomerans based on protein electrophoregrams but later excluded from this species, were further investigated using molecular techniques. 16S rRNA gene sequencing and multilocus sequence analyses (MLSA) revealed that the strains were phylogenetically closely related to Pantoea agglomerans, Pantoea stewartii and Pantoea ananatis. MLSA and amplified fragment length polymorphism analysis placed the strains into four separate clusters, not containing any of the type strains of species of the genus Pantoea. DNA–DNA hybridization confirmed the classification of the isolates into four novel species, for which the names Pantoea vagans sp. nov. (type strain R-21566T =LMG 24199T =BCC 105T =BD 765T), Pantoea eucalypti sp. nov. (type strain R-25678T =LMG 24197T =BCC 076T =BD 769T), Pantoea deleyi sp. nov. (type strain R-31523T =LMG 24200T =BCC 109T =BD 767T) and Pantoea anthophila sp. nov. (type strain LMG 2558T =BD 871T =NCPPB 1682T) are proposed.

Paraprevotella clara gen. nov., sp. nov. and Paraprevotella xylaniphila sp. nov., members of the family 'Prevotellaceae' isolated from human faeces.

Abstract: Two anaerobic, non-spore-forming, pleomorphic, Gram-negative rods, designated YIT 11840T and YIT 11841T, were isolated from human faeces. The organisms were catalase-negative, produced succinic and acetic acids as end products of glucose metabolism and had DNA G+C contents of approximately 48–49 mol%. Although the phenotypic characteristics of these two strains were very similar, analysis of their 16S rRNA gene sequences showed that they are only distantly related (93.8 %), indicating that they represent two different species. A comparative sequence analysis revealed that these two species are members of the family ‘Prevotellaceae’ but are phylogenetically distant (<88 % sequence similarity) from the known genera belonging to this family, including Prevotella, Hallela and Xylanibacter. On the basis of the phylogenetic analysis and physiological tests, strains YIT 11840T and YIT 11841T represent two novel species of a new genus, for which the names Paraprevotella clara gen. nov., sp. nov. (type strain YIT 11840T =JCM 14859T =DSM 19731T), the type species, and Paraprevotella xylaniphila sp. nov. (type strain YIT 11841T =JCM 14860T =DSM 19681T) are proposed.

Mesorhizobium metallidurans sp. nov., a metal-resistant symbiont of Anthyllis vulneraria growing on metallicolous soil in Languedoc, France.

Abstract: A polyphasic taxonomic approach was used to characterize 31 rhizobial isolates obtained from Anthyllis vulneraria, a metallicolous legume species, growing close to a zinc mine in the south of France (Saint Laurent le Minier). Comparative analysis of nearly full-length 16S rRNA gene sequences showed that these Gram-negative bacteria belonged to the genus Mesorhizobium and that they were related most closely to Mesorhizobium tianshanense ORS 2640(T). The phylogenetic relationships of these isolates with other Mesorhizobium species were confirmed by sequencing and analysis of the recA and atpD genes, which were used as alternative chromosomal markers. These novel mesorhizobial strains tolerated high concentrations of heavy metals: 16-32 mM Zn and 0.3-0.5 mM Cd. DNA-DNA hybridizations revealed >73 % relatedness between the strains isolated from A. vulneraria, but only 19-33 % relatedness between these and the type strains of M. tianshanense and Mesorhizobium mediterraneum. These results, together with other phenotypic characteristics, support the conclusion that these isolates represent a single, novel species of the genus Mesorhizobium, for which the name Mesorhizobium metallidurans sp. nov. is proposed. The type strain is STM 2683(T) (=CFBP 7147(T)=LMG 24485(T)).

Partial sequence comparison of the rpoB, sodA, groEL and gyrB genes within the genus Streptococcus.

Abstract: Phylogenetic analysis and species identification of members of the genus Streptococcus were carried out using partial sequence comparison of the 16S rRNA gene (1468–1478 bp), rpoB, encoding the β subunit of RNA polymerase (659–680 bp), sodA, encoding the manganese-dependent superoxide dismutase (435–462 bp), groEL, encoding the 60 kDa heat-shock protein (757 bp), and gyrB, encoding the Β subunit of DNA gyrase (458–461 bp). For the first time, most species within the genus Streptococcus were represented in the study (65 strains, representing 58 species and nine subspecies). Phylogenies inferred from rpoB, sodA, gyrB and groEL sequence comparisons were more discriminative than those inferred from 16S rRNA gene sequence comparison, and showed common clusters. The minimal interspecies divergence was 0.3, 2.7, 0, 2.5 and 3.4 % for the 16S rRNA gene, rpoB, sodA, gyrB and groEL, respectively. In general, groEL partial gene sequence comparison represented the best tool for identifying species and subspecies and for phylogenetic analysis.