Showing papers in "International Journal of Systematic and Evolutionary Microbiology in 2014"

Towards a taxonomic coherence between average nucleotide identity and 16S rRNA gene sequence similarity for species demarcation of prokaryotes

Abstract: Among available genome relatedness indices, average nucleotide identity (ANI) is one of the most robust measurements of genomic relatedness between strains, and has great potential in the taxonomy of bacteria and archaea as a substitute for the labour-intensive DNA–DNA hybridization (DDH) technique. An ANI threshold range (95–96 %) for species demarcation had previously been suggested based on comparative investigation between DDH and ANI values, albeit with rather limited datasets. Furthermore, its generality was not tested on all lineages of prokaryotes. Here, we investigated the overall distribution of ANI values generated by pairwise comparison of 6787 genomes of prokaryotes belonging to 22 phyla to see whether the suggested range can be applied to all species. There was an apparent distinction in the overall ANI distribution between intra- and interspecies relationships at around 95–96 % ANI. We went on to determine which level of 16S rRNA gene sequence similarity corresponds to the currently accepted ANI threshold for species demarcation using over one million comparisons. A twofold cross-validation statistical test revealed that 98.65 % 16S rRNA gene sequence similarity can be used as the threshold for differentiating two species, which is consistent with previous suggestions (98.2–99.0 %) derived from comparative studies between DDH and 16S rRNA gene sequence similarity. Our findings should be useful in accelerating the use of genomic sequence data in the taxonomy of bacteria and archaea.

List of new names and new combinations previously effectively, but not validly, published.

Abstract: The purpose of this announcement is to effect the valid publication of the following effectively published new names and new combinations under the procedure described in the Bacteriological Code (1990 Revision). Authors and other individuals wishing to have new names and/or combinations included in future lists should send three copies of the pertinent reprint or photocopies thereof, or an electronic copy of the published paper to the IJSEM Editorial Office for confirmation that all of the other requirements for valid publication have been met. It is also a requirement of IJSEM and the ICSP that authors of new species, new subspecies and new combinations provide evidence that types are deposited in two recognized culture collections in two different countries. It should be noted that the date of valid publication of these new names and combinations is the date of publication of this list, not the date of the original publication of the names and combinations. The authors of the new names and combinations are as given below. Inclusion of a name on these lists validates the publication of the name and thereby makes it available in the nomenclature of prokaryotes. The inclusion of a name on this list is not to be construed as taxonomic acceptance of the taxon to which the name is applied. Indeed, some of these names may, in time, be shown to be synonyms, or the organisms may be transferred to another genus, thus necessitating the creation of a new combination.

Taxonomic use of DNA G+C content and DNA-DNA hybridization in the genomic age

Abstract: The G+C content of a genome is frequently used in taxonomic descriptions of species and genera. In the past it has been determined using conventional, indirect methods, but it is nowadays reasonable to calculate the DNA G+C content directly from the increasingly available and affordable genome sequences. The expected increase in accuracy, however, might alter the way in which the G+C content is used for drawing taxonomic conclusions. We here re-estimate the literature assumption that the G+C content can vary up to 3–5 % within species using genomic datasets. The resulting G+C content differences are compared with DNA–DNA hybridization (DDH) similarities calculated in silico using the GGDC web server, with 70 % similarity as the gold standard threshold for species boundaries. The results indicate that the G+C content, if computed from genome sequences, varies no more than 1 % within species. Statistical models based on larger differences alone can reject the hypothesis that two strains belong to the same species. Because DDH similarities between two non-type strains occur in the genomic datasets, we also examine to what extent and under which conditions such a similarity could be <70 % even though the similarity of either strain to a type strain was ≥70 %. In theory, their similarity could be as low as 50 %, whereas empirical data suggest a boundary closer (but not identical) to 70 %. However, it is shown that using a 50 % boundary would not affect the conclusions regarding the DNA G+C content. Hence, we suggest that discrepancies between G+C content data provided in species descriptions on the one hand and those recalculated after genome sequencing on the other hand ≥1 % are due to significant inaccuracies of the applied conventional methods and accordingly call for emendations of species descriptions.

Integrating genomics into the taxonomy and systematics of the Bacteria and Archaea.

Abstract: The polyphasic approach used today in the taxonomy and systematics of the Bacteria and Archaea includes the use of phenotypic, chemotaxonomic and genotypic data. The use of 16S rRNA gene sequence data has revolutionized our understanding of the microbial world and led to a rapid increase in the number of descriptions of novel taxa, especially at the species level. It has allowed in many cases for the demarcation of taxa into distinct species, but its limitations in a number of groups have resulted in the continued use of DNA-DNA hybridization. As technology has improved, next-generation sequencing (NGS) has provided a rapid and cost-effective approach to obtaining whole-genome sequences of microbial strains. Although some 12,000 bacterial or archaeal genome sequences are available for comparison, only 1725 of these are of actual type strains, limiting the use of genomic data in comparative taxonomic studies when there are nearly 11,000 type strains. Efforts to obtain complete genome sequences of all type strains are critical to the future of microbial systematics. The incorporation of genomics into the taxonomy and systematics of the Bacteria and Archaea coupled with computational advances will boost the credibility of taxonomy in the genomic era. This special issue of International Journal of Systematic and Evolutionary Microbiology contains both original research and review articles covering the use of genomic sequence data in microbial taxonomy and systematics. It includes contributions on specific taxa as well as outlines of approaches for incorporating genomics into new strain isolation to new taxon description workflows.

A polyphasic strategy incorporating genomic data for the taxonomic description of novel bacterial species

Abstract: Currently, bacterial taxonomy relies on a polyphasic approach based on the combination of phenotypic and genotypic characteristics. However, the current situation is paradoxical in that the genetic criteria that are used, including DNA–DNA hybridization, 16S rRNA gene sequence nucleotide similarity and phylogeny, and DNA G+C content, have significant limitations, but genome sequences that contain the whole genetic information of bacterial strains are not used for taxonomic purposes, despite the decreasing costs of sequencing and the increasing number of available genomes. Recently, we diversified bacterial culture conditions with the aim of isolating uncultivated bacteria. To classify the putative novel species that we cultivated, we used a polyphasic strategy that included phenotypic as well as genomic criteria (genome characteristics as well as genomic sequence similarity). Herein, we review the pros and cons of genome sequencing for taxonomy and propose that the incorporation of genome sequences in taxonomic studies has the advantage of using reliable and reproducible data. This strategy, which we name taxono-genomics, may contribute to the taxonomic classification of bacteria.

Polyphasic taxonomic revision of the Ralstonia solanacearum species complex: proposal to emend the descriptions of Ralstonia solanacearum and Ralstonia syzygii and reclassify current R. syzygii strains as Ralstonia syzygii subsp. syzygii subsp. nov., R. solanacearum phylotype IV strains as Ralstonia syzygii subsp. indonesiensis subsp. nov., banana blood disease bacterium strains as Ralstonia syzygii subsp. celebesensis subsp. nov. and R. solanacearum phylotype I and III strains as Ralstonia pseudosolanacearum sp. nov.

Abstract: The Ralstonia solanacearum species complex has long been recognized as a group of phenotypically diverse strains that can be subdivided into four phylotypes. Using a polyphasic taxonomic approach on an extensive set of strains, this study provides evidence for a taxonomic and nomenclatural revision of members of this complex. Data obtained from phylogenetic analysis of 16S-23S rRNA ITS gene sequences, 16S-23S rRNA intergenic spacer (ITS) region sequences and partial endoglucanase (egl) gene sequences and DNA-DNA hybridizations demonstrate that the R. solanacearum species complex comprises three genospecies. One of these includes the type strain of Ralstonia solanacearum and consists of strains of R. solanacearum phylotype II only. The second genospecies includes the type strain of Ralstonia syzygii and contains only phylotype IV strains. This genospecies is subdivided into three distinct groups, namely R. syzygii, the causal agent of Sumatra disease on clove trees in Indonesia, R. solanacearum phylotype IV strains isolated from different host plants mostly from Indonesia, and strains of the blood disease bacterium (BDB), the causal agent of the banana blood disease, a bacterial wilt disease in Indonesia that affects bananas and plantains. The last genospecies is composed of R. solanacearum strains that belong to phylotypes I and III. As these genospecies are also supported by phenotypic data that allow the differentiation of the three genospecies, the following taxonomic proposals are made: emendation of the descriptions of Ralstonia solanacearum and Ralstonia syzygii and descriptions of Ralstonia syzygii subsp. nov. (type strain R 001(T) = LMG 10661(T) = DSM 7385(T)) for the current R. syzygii strains, Ralstonia syzygii subsp. indonesiensis subsp. nov. (type strain UQRS 464(T) = LMG 27703(T) = DSM 27478(T)) for the current R. solanacearum phylotype IV strains, Ralstonia syzygii subsp. celebesensis subsp. nov. (type strain UQRS 627(T) = LMG 27706(T) = DSM 27477(T)) for the BDB strains and Ralstonia pseudosolanacearum sp. nov. (type strain UQRS 461(T) = LMG 9673(T) = NCPPB 1029(T)) for the strains of R. solanacearum phylotypes I and III.

Nitrososphaera viennensis gen. nov., sp. nov., an aerobic and mesophilic, ammonia-oxidizing archaeon from soil and a member of the archaeal phylum Thaumarchaeota.

Abstract: A mesophilic, neutrophilic and aerobic, ammonia-oxidizing archaeon, strain EN76(T), was isolated from garden soil in Vienna (Austria). Cells were irregular cocci with a diameter of 0.6-0.9 µm and possessed archaella and archaeal pili as cell appendages. Electron microscopy also indicated clearly discernible areas of high and low electron density, as well as tubule-like structures. Strain EN76(T) had an S-layer with p3 symmetry, so far only reported for members of the Sulfolobales. Crenarchaeol was the major core lipid. The organism gained energy by oxidizing ammonia to nitrite aerobically, thereby fixing CO2, but growth depended on the addition of small amounts of organic acids. The optimal growth temperature was 42 °C and the optimal pH was 7.5, with ammonium and pyruvate concentrations of 2.6 and 1 mM, respectively. The genome of strain EN76(T) had a DNA G+C content of 52.7 mol%. Phylogenetic analyses of 16S rRNA genes showed that strain EN76(T) is affiliated with the recently proposed phylum Thaumarchaeota, sharing 85% 16S rRNA gene sequence identity with the closest cultivated relative 'Candidatus Nitrosopumilus maritimus' SCM1, a marine ammonia-oxidizing archaeon, and a maximum of 81% 16S rRNA gene sequence identity with members of the phyla Crenarchaeota and Euryarchaeota and any of the other recently proposed phyla (e.g. 'Korarchaeota' and 'Aigarchaeota'). We propose the name Nitrososphaera viennensis gen. nov., sp. nov. to accommodate strain EN76(T). The type strain of Nitrososphaera viennensis is strain EN76(T) ( = DSM 26422(T) = JMC 19564(T)). Additionally, we propose the family Nitrososphaeraceae fam. nov., the order Nitrososphaerales ord. nov. and the class Nitrososphaeria classis nov.

An update of ‘basic light and scanning electron microscopic methods for taxonomic studies of ciliated protozoa’

Abstract: This is an update of the review by Foissner (1991). Since then, I have improved some methods considerably. The following methods are described in detail: live observation, supravital staining with methyl green-pyronin, dry silver nitrate impregnation, wet silver nitrate impregnation, silver carbonate impregnation, protargol impregnation (three procedures), scanning electron microscopy, and deciliation. Familiarity with these methods (or modifications) is a prerequisite for successful taxonomic work. No staining method is equally appropriate to all kinds of ciliates. A table is provided which indicates those procedures which work best for certain groups of ciliates. A second table relates to the structures revealed by the procedures. Good descriptions usually demand at least live observation, silver nitrate and protargol or silver carbonate impregnation. Some instructions are provided for distinguishing mono- and dikinetids as well as ciliated and non-ciliated basal bodies in silvered ciliates. Furthermore, I added a section on ‘Deposition and Labeling of Preparations’. All methods work not only with ciliates but also with many other heterotrophic and autotrophic flagellated and amoeboid protists. The brilliancy of silver preparations has tempted some taxonomists to neglect live observation. However, many important species characteristics cannot be seen or are changed in silvered specimens. I thus consider all species descriptions based exclusively on silver slides as incomplete and of doubtful value for both α-taxonomists and ecologists.

Dickeya solani sp. nov., a pectinolytic plant pathogenic bacterium isolated from potato (Solanum tuberosum)

Abstract: Pectinolytic bacteria have been recently isolated from diseased potato plants exhibiting blackleg and slow wilt symptoms found in a number of European countries and Israel. These Gram-reaction-negative, motile, rods were identified as belonging to the genus Dickeya, previously the Pectobacterium chrysanthemi complex (Erwinia chrysanthemi, on the basis of production of a PCR product with the pelADE primers, 16S rRNA gene sequence analysis, fatty acid methyl esterase analysis, the production of phosphatases and the ability to produce indole and acids from alpha-methylglucoside. Differential physiological assays used previously to differentiate between strains of E chrysanthemi, showed that these isolates belonged to biovar 3. Eight of the isolates, seven from potato and one from hyacinth, were analysed together with 21 reference strains representing all currently recognized taxa within the genus Dickeya. The novel isolates formed a distinct genetic clade in multilocus sequence analysis (MLSA) using concatenated sequences of the intergenic spacer (IGS), as well as dnaX, recA, dnaN, fusA, gapA, purA, rpIB, rpoS and gyrA. Characterization by whole-cell MALDI-TOF mass spectrometry, pulsed field gel electrophoresis after digestion of whole-genome DNA with rare-cutting restriction enzymes, average nucleotide identity analysis and DNA DNA hybridization studies, showed that although related to Dickeya dadantii, these isolates represent a novel species within the genus Dickeya, for which the name Dickeya solani sp. nov. (type strain IPO 2222(T)=LMG25993(T)=NCPP134479(T)) is proposed.

Brucella papionis sp. nov., isolated from baboons (Papio spp.).

Abstract: Two Gram-negative, non-motile, non-spore-forming coccoid bacteria (strains F8/08-60(T) and F8/08-61) isolated from clinical specimens obtained from baboons (Papio spp.) that had delivered stillborn offspring were subjected to a polyphasic taxonomic study. On the basis of 16S rRNA gene sequence similarities, both strains, which possessed identical sequences, were assigned to the genus Brucella. This placement was confirmed by extended multilocus sequence analysis (MLSA), where both strains possessed identical sequences, and whole-genome sequencing of a representative isolate. All of the above analyses suggested that the two strains represent a novel lineage within the genus Brucella. The strains also possessed a unique profile when subjected to the phenotyping approach classically used to separate species of the genus Brucella, reacting only with Brucella A monospecific antiserum, being sensitive to the dyes thionin and fuchsin, being lysed by bacteriophage Wb, Bk2 and Fi phage at routine test dilution (RTD) but only partially sensitive to bacteriophage Tb, and with no requirement for CO2 and no production of H2S but strong urease activity. Biochemical profiling revealed a pattern of enzyme activity and metabolic capabilities distinct from existing species of the genus Brucella. Molecular analysis of the omp2 locus genes showed that both strains had a novel combination of two highly similar omp2b gene copies. The two strains shared a unique fingerprint profile of the multiple-copy Brucella-specific element IS711. Like MLSA, a multilocus variable number of tandem repeat analysis (MLVA) showed that the isolates clustered together very closely, but represent a distinct group within the genus Brucella. Isolates F8/08-60(T) and F8/08-61 could be distinguished clearly from all known species of the genus Brucella and their biovars by both phenotypic and molecular properties. Therefore, by applying the species concept for the genus Brucella suggested by the ICSP Subcommittee on the Taxonomy of Brucella, they represent a novel species within the genus Brucella, for which the name Brucella papionis sp. nov. is proposed, with the type strain F8/08-60(T) ( = NCTC 13660(T) = CIRMBP 0958(T)).

Characterization of Romboutsia ilealis gen. nov., sp. nov., isolated from the gastro-intestinal tract of a rat, and proposal for the reclassification of five closely related members of the genus Clostridium into the genera Romboutsia gen. nov., Intestinibacter gen. nov., Terrisporobacter gen. nov. and Asaccharospora gen. nov.

Abstract: A Gram-positive staining, rod-shaped, non-motile, spore-forming obligately anaerobic bacterium, designated CRIBT, was isolated from the gastro-intestinal tract of a rat and characterized. The major cellular fatty acids of strain CRIBT were saturated and unsaturated straight-chain C12-C19 fatty acids, with C16:0 being the predominant fatty acid. The polar lipid profile comprised six glycolipids, four phospholipids and one lipid that did not stain with any of the specific spray reagents used. The only quinone was MK-6. The predominating cell-wall sugars were glucose and galactose. The peptidoglycan type of strain CRIBT was A1σ lanthionine-direct. The genomic DNA G+C content of strain CRIBT was 28.1 mol%. On the basis of 16S rRNA gene sequence similarity, strain CRIBT was most closely related to a number of species of the genus Clostridium, including Clostridium lituseburense (97.2%), Clostridium glycolicum (96.2%), Clostridium mayombei (96.2%), Clostridium bartlettii (96.0%) and Clostridium irregulare (95.5%). All these species show very low 16S rRNA gene sequence similarity (<85%) to the type strain of Clostridium butyricum, the type species of the genus Clostridium. DNA-DNA hybridization with closely related reference strains indicated reassociation values below 32%. On the basis of phenotypic and genetic studies, a novel genus, Romboutsia gen. nov., is proposed. The novel isolate CRIBT (=DSM 25109T=NIZO 4048T) is proposed as the type strain of the type species, Romboutsia ilealis gen. nov., sp. nov., of the proposed novel genus. It is proposed that C. lituseburense is transferred to this genus as Romboutsia lituseburensis comb. nov. Furthermore, the reclassification into novel genera is proposed for C. bartlettii, as Intestinibacter bartlettii gen. nov., comb. nov. (type species of the genus), C. glycolicum, as Terrisporobacter glycolicus gen. nov., comb. nov. (type species of the genus), C. mayombei, as Terrisporobacter mayombei gen. nov., comb. nov., and C. irregulare, as Asaccharospora irregularis gen. nov., comb. nov. (type species of the genus), on the basis of additional data collected in this study. In addition, an emendation of the species Peptostreptococcus anaerobius and the order Eubacteriales is provided.

Acetobacteroides hydrogenigenes gen. nov., sp. nov., an anaerobic hydrogen-producing bacterium in the family Rikenellaceae isolated from a reed swamp.

Abstract: A strictly anaerobic, mesophilic, carbohydrate-fermenting, hydrogen-producing bacterium, designated strain RL-C-T, was isolated from a reed swamp in China. Cells were Gram-stain-negative, catalase-negative, non-spore-forming, non-motile rods measuring 0.7-1.0 mu m in width and 3.0-8.0 mu m in length. The optimum temperature for growth of strain RL-C-T was 37 degrees C (range 25-40 degrees C) and pH 7.0-7.5 (range pH 5.7-8.0). The strain could grow fermentatively on yeast extract, tryptone, arabinose, glucose, galactose, mannose, maltose, lactose, glycogen, pectin and starch. The main end products of glucose fermentation were acetate, H-2 and CO2. Organic acids, alcohols and amino acids were not utilized for growth. Yeast extract was not required for growth; however, it stimulated growth slightly. Nitrate, sulfate, sulfite, thiosulfate, elemental sulfur and Fe(III) nitrilotriacetate were not reduced as terminal electron acceptors. Aesculin was hydrolysed but not gelatin. Indole and H2S were produced from yeast extract. The G+C content of the genomic DNA was 51.2 mol%. The major cellular fatty acids were iso-C-15:0, anteiso-C-(15:0) and C-16:0. The most abundant polar lipid of strain RL-C-T was phosphatidylethanolamine. 16S rRNA gene sequence analysis revealed that the isolate belongs to the uncultured Blvii28 wastewater-sludge group (http://www.arb-silva.de/) in the family Rikenellaceae of the phylum Bacteroidetes, and shared low sequence similarities with the related species Alistipes shahii WAL 8301(T) (81.8 %), Rikenella microfusus ATCC 29728(T) (81.7%) and Anaerocella delicata WN081(T) (80.9 %). On the basis of these data, a novel species in a new genus of the family Rikenellaceae is proposed, Acetobacteroides hydrogenigenes gen. nov., sp. nov. The type strain of the type species is RL-C-T (=JCM 17603(T)=DSM 24657(T)=CGMCC 1.5173(T)).

Description of Klebsiella quasipneumoniae sp. nov., isolated from human infections, with two subspecies, Klebsiella quasipneumoniae subsp. quasipneumoniae subsp. nov. and Klebsiella quasipneumoniae subsp. similipneumoniae subsp. nov., and demonstration that Klebsiella singaporensis is a junior heterotypic synonym of Klebsiella variicola.

Abstract: Strains previously classified as members of Klebsiella pneumoniae phylogroups KpI, KpII-A, KpII-B and KpIII were characterized by 16S rRNA (rrs) gene sequencing, multilocus sequence analysis based on rpoB, fusA, gapA, gyrA and leuS genes, average nucleotide identity and biochemical characteristics. Phylogenetic analysis demonstrated that KpI and KpIII corresponded to K. pneumoniae and Klebsiella variicola, respectively, whereas KpII-A and KpII-B formed two well-demarcated sequence clusters distinct from other members of the genus Klebsiella. Average nucleotide identity between KpII-A and KpII-B was 96.4 %, whereas values lower than 94 % were obtained for both groups when compared with K. pneumoniae and K. variicola. Biochemical properties differentiated KpII-A, KpII-B, K. pneumoniae and K. variicola, with acid production from adonitol and l-sorbose and ability to use 3-phenylproprionate, 5-keto-d-gluconate and tricarballylic acid as sole carbon sources being particularly useful. Based on their genetic and phenotypic characteristics, we propose the names Klebsiella quasipneumoniae subsp. quasipneumoniae subsp. nov. and K. quasipneumoniae subsp. similipneumoniae subsp. nov. for strains of KpII-A and KpII-B, respectively. The type strain of K. quasipneumoniae sp. nov. and of K. quasipneumoniae subsp. quasipneumoniae subsp. nov. is 01A030(T) ( = SB11(T) = CIP 110771(T) = DSM 28211(T)). The type strain of K. quasipneumoniae subsp. similipneumoniae subsp. nov. is 07A044(T) ( = SB30(T) = CIP 110770(T) = DSM 28212(T)). Both strains were isolated from human blood cultures. This work also showed that Klebsiella singaporensis is a junior heterotypic synonym of K. variicola.

Lactobacillus apinorum sp. nov., Lactobacillus mellifer sp. nov., Lactobacillus mellis sp. nov., Lactobacillus melliventris sp. nov., Lactobacillus kimbladii sp. nov., Lactobacillus helsingborgensis sp. nov. and Lactobacillus kullabergensis sp. nov., isolated from the honey stomach of the honeybee Apis mellifera

Abstract: We discovered a symbiotic lactic acid bacterial (LAB) microbiota in the honey stomach of the honeybee Apis mellifera. The microbiota was composed of several phylotypes of Bifidobacterium and Lactobacillus. 16S ribosomal ribonucleic acid (rRNA) gene analyses and phenotypic and genetic characteristics revealed that the Lactobacillus phylotypes isolated represent seven novel species. One is grouped with Lactobacillus kunkeei and the others belong to the Lactobacillus buchneri and Lactobacillus delbrueckii subgroups of Lactobacillus. We propose the names Lactobacillus apinorum sp. nov., Lactobacillus mellifer sp. nov., Lactobacillus mellis sp. nov., Lactobacillus melliventris sp. nov., Lactobacillus kimbladii sp. nov., Lactobacillus helsingborgensis sp. nov., and Lactobacillus kullabergensis sp. nov., with the respective type strains being Fhon13NT ( = DSM 26257T = CCUG 63287T), Bin4NT ( = DSM 26254T = CCUG 63291T), Hon2NT ( = DSM 26255T = CCUG 63289T), Hma8NT ( = DSM 26256T = CCUG 63629T), Hma2NT ( = DSM 26263T = CCUG 63633T), Bma5NT ( = DSM 26265T = CCUG 63301T) and Biut2NT ( = DSM 26262T = CCUG 63631T). (Less)

Draconibacterium orientale gen. nov., sp. nov., isolated from two distinct marine environments, and proposal of Draconibacteriaceae fam. nov.

Abstract: The taxonomic characteristics of two bacterial strains, FH5T and SS4, isolated from enrichment cultures obtained from two distinct marine environments, were determined. These bacteria were Gram-stain-negative, facultatively anaerobic rods. Growth occurred at 20–40 °C (optimum, 28–32 °C), pH 5.5–9.0 (optimum, pH 7.0–7.5) and in the presence of 1–7 % NaCl (optimum, 2–4 %). The major cellular fatty acids were anteiso-C15 : 0 and iso-C15 : 0. Menaquinone 7 (MK-7) was the sole respiratory quinone. The major polar lipids were phosphatidylethanolamine, an unkown phospholipid and an unknown lipid. The DNA G+C contents of strains FH5T and SS4 were both determined to be 42.0 mol%. The results of DNA–DNA hybridization studies indicated that the FH5T and SS4 genomes share greater than 95 % relatedness. The strains formed a distinct phyletic line within the class Bacteroidia , with less than 89.4 % sequence similarity to their closest relatives with validly published names. On the basis of physiological and biochemical characteristics, 16S rRNA gene sequences and chemical properties, a novel genus and species, Draconibacterium orientale gen. nov., sp. nov., within the class Bacteroidia , are proposed, with strain FH5T ( = DSM 25947T = CICC 10585T) as the type strain. In addition, a new family, Draconibacteriaceae fam. nov., is proposed to accommodate Draconibacterium gen. nov.

Saccharicrinis carchari sp. nov., isolated from a shark, and emended descriptions of the genus Saccharicrinis and Saccharicrinis fermentans.

Abstract: A Gram-stain-negative, facultatively anaerobic, gliding, yellow-pigmented bacterium, designated SS12T, was isolated from shark gill homogenate and characterized using a polyphasic approach. The strain was catalase-positive and oxidase-negative. Optimal growth occurred at 28–30 °C, pH 7.0–7.5 and in the presence of 2–4 % (w/v) NaCl. The DNA G+C content was 40.0 mol%. The strain contained MK-7 as the prevailing menaquinone; iso-C15 : 0 and anteiso-C15 : 0 as the major cellular fatty acids; and phosphatidylethanolamine and an unknown lipid as the predominant polar lipids. Comparative analysis of 16S rRNA gene sequences demonstrated that the novel isolate showed the highest sequence similarity (94.68 %) to Saccharicrinis fermentans DSM 9555T and the sequence similarities among the type strains of all other species studied were less than 92 %. A phylogenetic tree, based on 16S rRNA gene sequences, showed that strain SS12T and Saccharicrinis fermentans DSM 9555T formed a distinct cluster within the family Marinilabiliaceae . On the basis of its phylogenetic position and phenotypic traits, strain SS12T represents a novel species of genus Saccharicrinis , for which the name Saccharicrinis carchari sp. nov. is proposed. The type strain is SS12T ( = CICC 10590T = DSM 27040T). Emended descriptions of the genus Saccharicrinis and Saccharicrinis fermentans are also provided.

EzEditor: a versatile sequence alignment editor for both rRNA- and protein-coding genes.

Abstract: EzEditor is a Java-based molecular sequence editor allowing manipulation of both DNA and protein sequence alignments for phylogenetic analysis. It has multiple features optimized to connect initial computer-generated multiple alignment and subsequent phylogenetic analysis by providing manual editing with reference to biological information specific to the genes under consideration. It provides various functionalities for editing rRNA alignments using secondary structure information. In addition, it supports simultaneous editing of both DNA sequences and their translated protein sequences for protein-coding genes. EzEditor is, to our knowledge, the first sequence editing software designed for both rRNA- and protein-coding genes with the visualization of biologically relevant information and should be useful in molecular phylogenetic studies. EzEditor is based on Java, can be run on all major computer operating systems and is freely available from http://sw.ezbiocloud.net/ezeditor/.

Listeria floridensis sp. nov., Listeria aquatica sp. nov., Listeria cornellensis sp. nov., Listeria riparia sp. nov. and Listeria grandensis sp. nov., from agricultural and natural environments

Abstract: Sampling of agricultural and natural environments in two US states (Colorado and Florida) yielded 18 Listeria-like isolates that could not be assigned to previously described species using traditional methods. Using whole-genome sequencing and traditional phenotypic methods, we identified five novel species, each with a genome-wide average blast nucleotide identity (ANIb) of less than 85 % to currently described species. Phylogenetic analysis based on 16S rRNA gene sequences and amino acid sequences of 31 conserved loci showed the existence of four well-supported clades within the genus Listeria ; (i) a clade representing Listeria monocytogenes , L. marthii , L. innocua , L. welshimeri , L. seeligeri and L. ivanovii , which we refer to as Listeria sensu stricto, (ii) a clade consisting of Listeria fleischmannii and two newly described species, Listeria aquatica sp. nov. (type strain FSL S10-1188T = DSM 26686T = LMG 28120T = BEI NR-42633T) and Listeria floridensis sp. nov. (type strain FSL S10-1187T = DSM 26687T = LMG 28121T = BEI NR-42632T), (iii) a clade consisting of Listeria rocourtiae , L. weihenstephanensis and three novel species, Listeria cornellensis sp. nov. (type strain TTU A1-0210T = FSL F6-0969T = DSM 26689T = LMG 28123T = BEI NR-42630T), Listeria grandensis sp. nov. (type strain TTU A1-0212T = FSL F6-0971T = DSM 26688T = LMG 28122T = BEI NR-42631T) and Listeria riparia sp. nov. (type strain FSL S10-1204T = DSM 26685T = LMG 28119T = BEI NR- 42634T) and (iv) a clade containing Listeria grayi . Genomic and phenotypic data suggest that the novel species are non-pathogenic.

Gordonibacter urolithinfaciens sp. nov., a urolithin-producing bacterium isolated from the human gut.

Abstract: Urolithins are dibenzopyranone metabolites that exert anti-inflammatory activity in vivo and are produced by the gut microbiota from the dietary polyphenols ellagic acid (EA) and ellagitannins. However, the bacteria involved in this process remain unknown. We report here a novel bacterium, strain CEBAS 1/15PT, capable of metabolizing EA to urolithins, that was isolated from healthy human faeces and characterized by determining phenotypic, biochemical and molecular methods. The strain was related to Gordonibacter pamelaeae 7-10-1-bT, the type and only reported strain of the only species of the genus Gordonibacter, with about 97 % 16S rRNA gene sequence similarity; they were both obligately anaerobic, non-spore-forming, Gram-stain-positive, short-rods/coccobacilli and metabolized only small numbers of carbon sources. l-Fucose, d-fructose, turanose, d-galacturonic acid and α-ketobutyric acid were metabolized by strain CEBAS 1/15PT, while G. pamelaeae was negative for metabolism of these compounds. The whole-cell fatty acids consisted predominantly of saturated fatty acids (70 %); strain CEBAS 1/15PT differed significantly from G. pamelaeae in the major fatty acid, which was C18 : 1ω9c, while anteiso-C15 : 0 was the major component for G. pamelaeae . The presence of a number of different fatty acid peaks, especially C19 : 0 cyclo and C18 : 1ω6c, was also indicative of distinct species. Six glycolipids (GL1–6) were recognized, while, in G. pamelaeae, only four glycolipids were described. On the basis of these data, the novel species Gordonibacter urolithinfaciens sp. nov. is described, with strain CEBAS 1/15PT ( = DSM 27213T = CCUG 64261T) as the type strain.

Leptospira mayottensis sp. nov., a pathogenic species of the genus Leptospira isolated from humans.

Abstract: A group of strains representing species of the genus Leptospira, isolated from patients with leptospirosis in Mayotte (Indian Ocean), were previously found to be considerably divergent from other known species of the genus Leptospira This was inferred from sequence analysis of rrs (16S rRNA) and other genetic loci and suggests that they belong to a novel species Two strains from each serogroup currently identified within this novel species were studied Spirochaete, aerobic, motile, helix-shaped strains grew well at 30–37 °C, but not at 13 °C or in the presence of 8-azaguanine Draft genomes of the strains were also analysed to study the DNA relatedness with other species of the genus Leptospira The new isolates formed a distinct clade, which was most closely related to Leptospira borgpetersenii , in multilocus sequence analysis using concatenated sequences of the genes rpoB, recA, fusA, gyrB, leuS and sucA Analysis of average nucleotide identity and genome-to-genome distances, which have recently been proposed as reliable substitutes for classical DNA–DNA hybridization, further confirmed that these isolates should be classified as representatives of a novel species The G+C content of the genomic DNA was 395 mol% These isolates are considered to represent a novel species, for which the name Leptospira mayottensis sp nov is proposed, with 200901116T ( = CIP 110703T = DSM 28999T) as the type strain

Lactobacillus apis sp. nov., from the stomach of honeybees (Apis mellifera), having an in vitro inhibitory effect on the causative agents of American and European foulbrood.

Abstract: A taxonomic study was performed on Gram-stain-positive, catalase-negative and regular rod-shaped bacterial strains R4BT and R4C, isolated from the stomachs of honeybees. 16S rRNA gene sequence analysis revealed that the phylogenetic position of the novel strains was within the genus Lactobacillus; the highest sequence similarity to R4BT was shown by Lactobacillus acidophilus BCRC 10695T. Lower sequence similarities were found to other obligately homofermentative lactobacilli. A PCR–DGGE method could detect the sequence of the 16S rRNA gene of strain R4BT at different developmental stages of honeybees occurring in two different locations in the Czech Republic. The distinctiveness of the strains from other lactobacilli was also confirmed by analysis of sequences of other phylogenetic markers applicable to the taxonomy of the genus Lactobacillus, ribotyping and rep-PCR analysis. The DNA G+C content of strain R4BT was 41.3% mol. The major polar lipids of strain R4BT were glycolipids, lipids and phospholipids. Phenotypic and phylogenetic characteristics also confirmed the independent status of the strains at the species level. Interestingly, strain R4BT was able to inhibit growth in vitro of Paenibacillus larvae subsp. larvae (causal agent of American foulbrood in honeybees) and Melissococcus plutonius (causal agent of European foulbrood). The name Lactobacillus apis sp. nov. is proposed for this novel taxon.

Microvirga vignae sp. nov., a root nodule symbiotic bacterium isolated from cowpea grown in semi-arid Brazil.

Abstract: 16S rRNA gene sequence analysis of eight strains (BR 3299T, BR 3296, BR 10192, BR 10193, BR 10194, BR 10195, BR 10196 and BR 10197) isolated from nodules of cowpea collected from a semi-arid region of Brazil showed 97 % similarity to sequences of recently described rhizobial species of the genus Microvirga . Phylogenetic analyses of four housekeeping genes (gyrB, recA, dnaK and rpoB), DNA–DNA relatedness and AFLP further indicated that these strains belong to a novel species within the genus Microvirga . Our data support the hypothesis that genes related to nitrogen fixation were obtained via horizontal gene transfer, as sequences of nifH genes were very similar to those found in members of the genera Rhizobium and Mesorhizobium , which are not immediate relatives of the genus Microvirga , as shown by 16S rRNA gene sequence analysis. Phenotypic traits, such as host range and carbon utilization, differentiate the novel strains from the most closely related species, Microvirga lotononidis , Microvirga zambiensis and Microvirga lupini . Therefore, these symbiotic nitrogen-fixing bacteria are proposed to be representatives of a novel species, for which the name Microvirga vignae sp. nov. is suggested. The type strain is BR3299T ( = HAMBI 3457T).

Phylogeny of the class Actinobacteria revisited in the light of complete genomes. The orders 'Frankiales' and Micrococcales should be split into coherent entities: proposal of Frankiales ord. nov., Geodermatophilales ord. nov., Acidothermales ord. nov. and Nakamurellales ord. nov.

Abstract: The phylogeny of the class Actinobacteria remains controversial, essentially because it is very sensitive to the choice of dataset and phylogenetic methods. We used a test proposed recently, based on complete genome data, which chooses among candidate species phylogenies based on the number of lateral gene transfers (LGT) needed to explain the diversity of histories among gene trees for a set of genomes. We used 100 completely sequenced genomes representing 35 families and 17 orders of the class Actinobacteria and evaluated eight different hypotheses for their phylogeny, including one based on a concatenate of 54 conserved proteins present in single copy in all these genomes, trees based on 16S and 23S rRNA gene sequences or their concatenation, and a tree based on the concatenation of MLSA genes (encoding AtpI, GyrA, FtsZ, SecA and DnaK). We used Prunier to infer the number of LGT in 579 proteins (different from those used to build the concatenated tree) present in at least 70 species, using the different hypothetical species trees as references. The best tree, with the lowest number of lateral transfers, was the one based on the concatenation of 54 proteins. In that tree, the orders Bifidobacteriales, Coriobacteriales, 'Corynebacteriales', 'Micromonosporales', 'Propionibacteriales', 'Pseudonocardiales', Streptomycetales and 'Streptosporangiales' were recovered while the orders 'Frankiales' and Micrococcales were not. It is thus proposed that the order 'Frankiales', which has an effectively but not validly published name, be split into Frankiales ord. nov. (type family Frankiaceae), Geodermatophilales ord. nov. (Geodermatophilaceae), Acidothermales ord. nov. (Acidothermaceae) and Nakamurellales ord. nov. (Nakamurellaceae). The order Micrococcales should also be split into Micrococcales (genera Kocuria, Rothia, Micrococcus, Arthrobacter, Tropheryma, Microbacterium, Leifsonia and Clavibacter), Cellulomonales (Beutenbergia, Cellulomonas, Xylanimonas, Jonesia and Sanguibacter) and Brachybacteriales (Brachybacterium) but the formal proposal for this will have to wait until more genomes become available for a significant proportion of strains in this order.

Geobacter anodireducens sp. nov., an exoelectrogenic microbe in bioelectrochemical systems

Abstract: A previously isolated exoelectrogenic bacterium, strain SD-1(T), was further characterized and identified as a representative of a novel species of the genus Geobacter. Strain SD-1(T) was Gram-negative, aerotolerant, anaerobic, non-spore-forming, non-fermentative and non-motile. Cells were short, curved rods (0.8-1.3 µm long and 0.3 µm in diameter). Growth of strain SD-1(T) was observed at 15-42 °C and pH 6.0-8.5, with optimal growth at 30-35 °C and pH 7. Analysis of 16S rRNA gene sequences indicated that the isolate was a member of the genus Geobacter, with the closest known relative being Geobacter sulfurreducens PCA(T) (98% similarity). Similar to other members of the genus Geobacter, strain SD-1(T) used soluble or insoluble Fe(III) as the sole electron acceptor coupled with the oxidation of acetate. However, SD-1(T) could not reduce fumarate as an electron acceptor with acetate oxidization, which is an important physiological trait for G. sulfurreducens. Moreover, SD-1(T) could grow in media containing as much as 3% NaCl, while G. sulfurreducens PCA(T) can tolerate just half this concentration, and this difference in salt tolerance was even more obvious when cultivated in bioelectrochemical systems. DNA-DNA hybridization analysis of strain SD-1(T) and its closest relative, G. sulfurreducens ATCC 51573(T), showed a relatedness of 61.6%. The DNA G+C content of strain SD-1(T) was 58.9 mol%. Thus, on the basis of these characteristics, strain SD-1(T) was not assigned to G. sulfurreducens, and was instead classified in the genus Geobacter as a representative of a novel species. The name Geobacter anodireducens sp. nov. is proposed, with the type strain SD-1(T) ( = CGMCC 1.12536(T) = KCTC 4672(T)).

Re-examination of the taxonomic status of Enterobacter helveticus, Enterobacter pulveris and Enterobacter turicensis as members of the genus Cronobacter and their reclassification in the genera Franconibacter gen. nov. and Siccibacter gen. nov. as Franconibacter helveticus comb. nov., Franconibacter pulveris comb. nov. and Siccibacter turicensis comb. nov., respectively

Abstract: Recently, a taxonomical re-evaluation of the genus Enterobacter, based on multi-locus sequence typing (MLST) analysis, has led to the proposal that the species Enterobacter pulveris, Enterobacter helveticus and Enterobacter turicensis should be reclassified as novel species of the genus Cronobacter. In the present work, new genome-scale analyses, including average nucleotide identity, genome-scale phylogeny and k-mer analysis, coupled with previously reported DNA-DNA hybridization values and biochemical characterization strongly indicate that these three species of the genus Enterobacter are not members of the genus Cronobacter, nor do they belong to the re-evaluated genus Enterobacter. Furthermore, data from this polyphasic study indicated that all three species constitute two new genera. We propose reclassifying Enterobacter pulveris and Enterobacter helveticus in the genus Franconibacter gen. nov. as Franconibacter pulveris comb. nov. (type strain 601/05(T) = LMG 24057(T) = DSM 19144(T)) and Franconibacter helveticus comb. nov. (type strain 513/05(T) = LMG 23732(T) = DSM 18396(T)), respectively, and Enterobacter turicensis in the genus Siccibacter gen. nov. as Siccibacter turicensis comb. nov. (type strain 508/05(T) = LMG 23730(T) = DSM 18397(T)).

Sulfurovum aggregans sp. nov., a hydrogen-oxidizing, thiosulfate-reducing chemolithoautotroph within the Epsilonproteobacteria isolated from a deep-sea hydrothermal vent chimney, and an emended description of the genus Sulfurovum

Abstract: A novel mesophilic, strictly hydrogen-oxidizing, sulfur-, nitrate- and thiosulfate-reducing bacterium, designated strain Monchim33T, was isolated from a deep-sea hydrothermal vent chimney at the Central Indian Ridge. The non-motile, rod-shaped cells were Gram-stain-negative and non-sporulating. Growth was observed between 15 and 37 °C (optimum 33 °C; 3.2 h doubling time) and between pH 5.4 and 8.6 (optimum pH 6.0). The isolate was a strictly anaerobic chemolithoautotroph capable of using molecular hydrogen as the sole energy source and carbon dioxide as the sole carbon source. The G+C content of the genomic DNA was 42.6 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the novel isolate belonged to the genus Sulfurovum and was closely related to Sulfurovum sp. NBC37-1 and Sulfurovum lithotrophicum 42BKT (95.6 and 95.4 % similarity, respectively). DNA–DNA hybridization demonstrated that the novel isolate could be differentiated genotypically from Sulfurovum sp. NBC37-1 and Sulfurovum lithotrophicum . On the basis of the molecular and physiological traits of the new isolate, the name Sulfurovum aggregans sp. nov. is proposed, with the type strain Monchim33T ( = JCM 19824T = DSM 27205T).

Rhizobium laguerreae sp. nov. nodulates Vicia faba on several continents.

Abstract: Several fast-growing strains nodulating Vicia faba in Peru, Spain and Tunisia formed a cluster related to Rhizobium leguminosarum . The 16S rRNA gene sequences were identical to that of R. leguminosarum USDA 2370T, whereas rpoB, recA and atpD gene sequences were phylogenetically distant, with sequence similarities of less than 96 %, 97 % and 94 %, respectively. DNA–DNA hybridization analysis showed a mean relatedness value of 43 % between strain FB206T and R. leguminosarum USDA 2370T. Phenotypic characteristics of the novel strains also differed from those of the closest related species of the genus Rhizobium . Therefore, based on genotypic and phenotypic data obtained in this study, we propose to classify this group of strains nodulating Vicia faba as a novel species of the genus Rhizobium named Rhizobium laguerreae sp. nov. The type strain is FB206T ( = LMG 27434T = CECT 8280T).

Recommended minimal standards for description of new taxa of the genera Bifidobacterium, Lactobacillus and related genera.

Abstract: Minimal standards for the description of new cultivable strains that represent novel genera and species belonging to the genera Bifidobacterium, Lactobacillus and related genera are proposed in accordance with Recommendation 30b of the Bacteriological Code (1990 Revision): the description of novel species should be based on phenotypic, genotypic and ecological characteristics to ensure a rich polyphasic characterization. Concerning genotypic characterization, in addition to DNA G+C content (mol%) data, the description should be based on DNA-DNA hybridization (DDH), 16S rRNA gene sequence similarities and at least two housekeeping gene (e.g. hsp60 and recA) sequence similarities. DDH might not be needed if the 16S rRNA gene sequence similarity to the closest known species is lower than 97 %. This proposal has been endorsed by members of the Subcommittee on the Taxonomy of Bifidobacterium, Lactobacillus and related organisms of the International Committee on the Systematics of Prokaryotes.

Hydrogenispora ethanolica gen. nov., sp. nov., an anaerobic carbohydrate-fermenting bacterium from anaerobic sludge.

Abstract: An anaerobic, spore-forming, ethanol-hydrogen-coproducing bacterium, designated LX-BT, was isolated from an anaerobic sludge treating herbicide wastewater. Cells of strain LX-BT were non-motile rods (0.3-0.5×3.0-18.0 µm). Spores were terminal with a bulged sporangium. Growth occurred at 20-50 °C (optimum 37-45 °C), pH 5.0-8.0 (optimum pH 6.0-7.7) and 0-2.5% (w/v) NaCl. The strain could grow fermentatively on glucose, maltose, arabinose, fructose, xylose, ribose, galactose, mannose, raffinose, sucrose, pectin, starch, glycerol, fumarate, tryptone and yeast extract. The major end-products of glucose fermentation were acetate, ethanol and hydrogen. Yeast extract was not required but stimulated growth. Nitrate, sulfate, thiosulfate, elemental sulfur, sulfite, anthraquinone-2,6-disulfonate, fumarate and Fe (III) nitrilotriacetate were not used as terminal electron acceptors. The G+C content of the genomic DNA was 56.1 mol%. The major cellular fatty acids were anteiso-C15:0, iso-C14:0 and C16:0. The most abundant polar lipids of strain LX-BT were diphosphatidylglycerol and phosphatidylglycerol. 16S rRNA gene sequence analysis revealed that it belongs to an as-yet-unidentified taxon at the order- or class-level (OPB54) within the phylum Firmicutes, showing 86.5% sequence similarity to previously described species of the Desulfotomaculum cluster. The name Hydrogenispora ethanolica gen. nov., sp. nov. is proposed to accommodate strain LX-BT (=DSM 25471T=JCM 18117T=CGMCC 1.5175T) as the type strain.

Dickeya aquatica sp. nov., isolated from waterways.

Abstract: Pectinolytic Gram-negative bacteria were isolated from different waterways in the UK and Finland. Three strains (174/2T, 181/2 and Dw054) had the same 16S rRNA gene sequences which shared 99 % sequence similarity to species of the genus Dickeya , and a phylogeny of related genera confirmed attribution to this genus. Fatty acid profile analysis of all three strains found a high proportion of C16 : 1ω7c/C16 : 1ω7c and C16 : 0 fatty acids, and library profile searches found closest matches to Dickeya chrysanthemi . Production of a concatenated phylogeny using six loci, recA, gapA, atpD, gyrB, infB and rpoB, provided a high-resolution phylogeny which placed strains 174/2T and 181/2 as a distinct clade, separated from the other species of the genus Dickeya by a relatively long branch-length. DNA–DNA hybridization analysis with a limited number of reference species also supported the distinctiveness of strains 174/2T and 181/2 within the genus Dickeya . All three strains could be phenotypically distinguished from other species of the genus by fermentation of melibiose and raffinose but not d-arabinose or mannitol. The name Dickeya aquatica sp. nov. is proposed for the new taxon; the type strain is 174/2T ( = NCPPB 4580T = LMG 27354T).