Elioraea rosea sp. nov., a plant promoting bacterium isolated from floodwater of a paddy field.

A Gram-stain-negative bacterium, designated strain PF-30T, was isolated from floodwater of a paddy field in South Korea. Strain PF-30T was found to be a strictly aerobic, motile and pink-pigmented rods which can grow at 25-40 °C (optimum, 28 °C), at pH 5.0-9.0 (optimum pH 7.0) and at salinities of 0.5-3.0 % NaCl (optimum 0.5 % NaCl). Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain PF-30T belongs to the genus Elioraea, showing highest sequence similarity to Elioraea tepidiphila TU-7T (97.1%) and less than 91.3 % similarity with other members of the family Acetobacteraceae. The average nucleotide identity (ANI) and DNA-DNA relatedness between the strain PF-30T and E. tepidiphila TU-7T yielded an ANI value of 75.1 % and DNA-DNA relatedness of 11.7±0.7 %, respectively. The major fatty acids were identified as C18 : 0 and C18 : 1 ω7c. The predominant respiratory quinone was identified as Q-10. The DNA G+C content was determined to be 69.9 mol%. The strain PF-30T was observed to produce plant-growth-promoting materials such as indole-3-acetic acid (IAA), siderophore and phytase. On the basis of the results from phylogenetic, chemotaxonomic and phenotypic data, we concluded that strain PF-30T represents a novel species of the genus Elioraea, for which the name Elioraea rosea sp. nov. is proposed. The type strain is PF-30T (=KACC 19985T=NBRC 113984T).

Reyranella terrae sp. nov., isolated from an agricultural soil, and emended description of the genus Reyranella.

A novel strain designated 11G32T was isolated from an agricultural soil cultivated with Chinese cabbage in Korea. The cells were Gram-stain-negative, aerobic, non-motile and rod-shaped. The strain grew at 15-28 °C (optimum, 20 °C), pH 5.0-7.0 (optimum, pH 5.0-6.0) and without NaCl. Phylogenetically, the strain was found to be closely related to members of the genus Reyranella and showed 16S rRNA gene sequence similarities of 97.18, 96.76 and 95.99 % with Reyranella graminifolii Wo-34T, Reyranella massiliensis 521T and Reyranella soli KIS14-15T, respectively. The major fatty acids were C19 : 0 cyclo ω8c, C16 : 0, summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) and 11-methyl C18 : 1ω7c. The predominant ubiquinone was Q-10. The polar lipids profile revealed the presence of phosphatidylmonomethylethanolamine, phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, unidentified aminolipids, unidentified phospholipids and unidentified lipids. On the basis of data presented, strain 11G32T is considered to represent a novel species of the genus Reyranella, for which the name Reyranella terrae sp. nov. is proposed. The type strain is 11G32T (=KACC 18486T=NBRC 111476T).

Multisubstrate isotope labeling and metagenomic analysis of active soil bacterial communities.

Soil microbial diversity represents the largest global reservoir of novel microorganisms and enzymes. In this study, we coupled functional metagenomics and DNA stable-isotope probing (DNA-SIP) using multiple plant-derived carbon substrates and diverse soils to characterize active soil bacterial communities and their glycoside hydrolase genes, which have value for industrial applications. We incubated samples from three disparate Canadian soils (tundra, temperate rainforest, and agricultural) with five native carbon ((12)C) or stable-isotope-labeled ((13)C) carbohydrates (glucose, cellobiose, xylose, arabinose, and cellulose). Indicator species analysis revealed high specificity and fidelity for many uncultured and unclassified bacterial taxa in the heavy DNA for all soils and substrates. Among characterized taxa, Actinomycetales (Salinibacterium), Rhizobiales (Devosia), Rhodospirillales (Telmatospirillum), and Caulobacterales (Phenylobacterium and Asticcacaulis) were bacterial indicator species for the heavy substrates and soils tested. Both Actinomycetales and Caulobacterales (Phenylobacterium) were associated with metabolism of cellulose, and Alphaproteobacteria were associated with the metabolism of arabinose; members of the order Rhizobiales were strongly associated with the metabolism of xylose. Annotated metagenomic data suggested diverse glycoside hydrolase gene representation within the pooled heavy DNA. By screening 2,876 cloned fragments derived from the (13)C-labeled DNA isolated from soils incubated with cellulose, we demonstrate the power of combining DNA-SIP, multiple-displacement amplification (MDA), and functional metagenomics by efficiently isolating multiple clones with activity on carboxymethyl cellulose and fluorogenic proxy substrates for carbohydrate-active enzymes. Importance: The ability to identify genes based on function, instead of sequence homology, allows the discovery of genes that would not be identified through sequence alone. This is arguably the most powerful application of metagenomics for the recovery of novel genes and a natural partner of the stable-isotope-probing approach for targeting active-yet-uncultured microorganisms. We expanded on previous efforts to combine stable-isotope probing and metagenomics, enriching microorganisms from multiple soils that were active in degrading plant-derived carbohydrates, followed by construction of a cellulose-based metagenomic library and recovery of glycoside hydrolases through functional metagenomics. The major advance of our study was the discovery of active-yet-uncultivated soil microorganisms and enrichment of their glycoside hydrolases. We recovered positive cosmid clones in a higher frequency than would be expected with direct metagenomic analysis of soil DNA. This study has generated an invaluable metagenomic resource that future research will exploit for genetic and enzymatic potential.

tRNA signatures reveal a polyphyletic origin of SAR11 strains among alphaproteobacteria.

Molecular phylogenetics and phylogenomics are subject to noise from horizontal gene transfer (HGT) and bias from convergence in macromolecular compositions. Extensive variation in size, structure and base composition of alphaproteobacterial genomes has complicated their phylogenomics, sparking controversy over the origins and closest relatives of the SAR11 strains. SAR11 are highly abundant, cosmopolitan aquatic Alphaproteobacteria with streamlined, A+T-biased genomes. A dominant view holds that SAR11 are monophyletic and related to both Rickettsiales and the ancestor of mitochondria. Other studies dispute this, finding evidence of a polyphyletic origin of SAR11 with most strains distantly related to Rickettsiales. Although careful evolutionary modeling can reduce bias and noise in phylogenomic inference, entirely different approaches may be useful to extract robust phylogenetic signals from genomes. Here we develop simple phyloclassifiers from bioinformatically derived tRNA Class-Informative Features (CIFs), features predicted to target tRNAs for specific interactions within the tRNA interaction network. Our tRNA CIF-based model robustly and accurately classifies alphaproteobacterial genomes into one of seven undisputed monophyletic orders or families, despite great variability in tRNA gene complement sizes and base compositions. Our model robustly rejects monophyly of SAR11, classifying all but one strain as Rhizobiales with strong statistical support. Yet remarkably, conventional phylogenetic analysis of tRNAs classifies all SAR11 strains identically as Rickettsiales. We attribute this discrepancy to convergence of SAR11 and Rickettsiales tRNA base compositions. Thus, tRNA CIFs appear more robust to compositional convergence than tRNA sequences generally. Our results suggest that tRNA-CIF-based phyloclassification is robust to HGT of components of the tRNA interaction network, such as aminoacyl-tRNA synthetases. We explain why tRNAs are especially advantageous for prediction of traits governing macromolecular interactions from genomic data, and why such traits may be advantageous in the search for robust signals to address difficult problems in classification and phylogeny.

Going deeper: metagenome of a hadopelagic microbial community.

The paucity of sequence data from pelagic deep-ocean microbial assemblages has severely restricted molecular exploration of the largest biome on Earth. In this study, an analysis is presented of a large-scale 454-pyrosequencing metagenomic dataset from a hadopelagic environment from 6,000 m depth within the Puerto Rico Trench (PRT). A total of 145 Mbp of assembled sequence data was generated and compared to two pelagic deep ocean metagenomes and two representative surface seawater datasets from the Sargasso Sea. In a number of instances, all three deep metagenomes displayed similar trends, but were most magnified in the PRT, including enrichment in functions for two-component signal transduction mechanisms and transcriptional regulation. Overrepresented transporters in the PRT metagenome included outer membrane porins, diverse cation transporters, and di- and tri-carboxylate transporters that matched well with the prevailing catabolic processes such as butanoate, glyoxylate and dicarboxylate metabolism. A surprisingly high abundance of sulfatases for the degradation of sulfated polysaccharides were also present in the PRT. The most dramatic adaptational feature of the PRT microbes appears to be heavy metal resistance, as reflected in the large numbers of transporters present for their removal. As a complement to the metagenome approach, single-cell genomic techniques were utilized to generate partial whole-genome sequence data from four uncultivated cells from members of the dominant phyla within the PRT, Alphaproteobacteria, Gammaproteobacteria, Bacteroidetes and Planctomycetes. The single-cell sequence data provided genomic context for many of the highly abundant functional attributes identified from the PRT metagenome, as well as recruiting heavily the PRT metagenomic sequence data compared to 172 available reference marine genomes. Through these multifaceted sequence approaches, new insights have been provided into the unique functional attributes present in microbes residing in a deeper layer of the ocean far removed from the more productive sun-drenched zones above.

Isolation and characterization of Reyranella massiliensis gen. nov., sp. nov. from freshwater samples by using an amoeba co-culture procedure.

The analysis of three water samples from two cooling towers and one river allowed us to isolate three strains of a novel species of the class Alphaproteobacteria which is phylogenetically related to uncultured alphaproteobacteria. Based upon 16S rRNA gene sequence analysis and phenotypic characterization, we propose to name this novel species Reyranella massiliensis gen. nov., sp. nov., type strain 521(T) ( = CSUR P115(T)  = DSM 23428(T)). The most closely related cultivable micro-organism to this novel bacterium is a member of the genus Magnetospirillum.

Reduction of perchlorate and nitrate by microbial communities in vadose soil.

Perchlorate contamination is a concern because of the increasing frequency of its detection in soils and groundwater and its presumed inhibitory effect on human thyroid hormone production. Although significant perchlorate contamination occurs in the vadose (unsaturated) zone, little is known about perchlorate biodegradation potential by indigenous microorganisms in these soils. We measured the effects of electron donor (acetate and hydrogen) and nitrate addition on perchlorate reduction rates and microbial community composition in microcosm incubations of vadose soil. Acetate and hydrogen addition enhanced perchlorate reduction, and a longer lag period was observed for hydrogen (41 days) than for acetate (14 days). Initially, nitrate suppressed perchlorate reduction, but once perchlorate started to be degraded, the process was stimulated by nitrate. Changes in the bacterial community composition were observed in microcosms enriched with perchlorate and either acetate or hydrogen. Denaturing gradient gel electrophoresis analysis and partial sequencing of 16S rRNA genes recovered from these microcosms indicated that formerly reported perchlorate-reducing bacteria were present in the soil and that microbial community compositions were different between acetate- and hydrogen-amended microcosms. These results indicate that there is potential for perchlorate bioremediation by native microbial communities in vadose soil.

Structure, function and organization of antenna polypeptides and antenna complexes from the three families of Rhodospirillaneae.

Comparative primary structural analysis of polypeptides from antenna complexes from species of the three families of Rhodospirillaneae indicates the structural principles responsible for the formation of spectrally distinct light-harvesting complexes. In many of the characterized antenna systems the basic structural minimal unit is an alpha/beta polypeptide pair. Specific clusters of amino acid residues, in particular aromatic residues in the C-terminal domain, identify the antenna polypeptides to specific types of antenna systems, such as B880 (strong circular dichroism (CD)), B870 (weak CD), B800-850 (high), B800-850 (low) or B800-820. The core complex B880 (B1020) of species from Ectothiorhodospiraceae and Chromatiaceae apparently consists of four (alpha 1 alpha 2 beta 1 beta 2) or three (2 alpha beta 1 beta 2) chemically dissimilar antenna polypeptides respectively. There is good evidence that the so-called variable antenna complexes, such as the B800-850 (high), B800-850 (low) or B800-820 of Rp. acidophila, Rp. palustris and Cr. vinosum, are comprised of multiple forms of peripheral light-harvesting polypeptides. Structural similarities between prokaryotic and eukaryotic antenna polypeptides are discussed in terms of similar pigment organization. The structural basis for the strict organization of pigment molecules (bacteriochlorophyll (BChl) cluster) in the antenna system of purple bacteria is the hierarchical organization of the alpha- and beta-antenna polypeptides within and between the antenna complexes. On the basis of the three-domain structure of the antenna polypeptides with the central hydrophobic domain, forming a transmembrane alpha helix, possible arrangements of the antenna polypeptides in the three-dimensional structure of core and peripheral antenna complexes are discussed. Important structural and functional features of these polypeptides and therefore of the BChl cluster are the alpha/beta heterodimers, the alpha 2 beta 2 basic units and cyclic arrangements of these basic units. Equally important for the formation of the antenna complexes or the entire antenna are polypeptide-polypeptide, pigment-pigment and pigment-polypeptide interactions.

Nucleotide sequence of 16S rRNA and phylogenetic position of the green sulfur bacterium Clathrochloris sulfurica.

The almost complete primary structure of the 16S rRNA from the green sulfur bacterium "Clathrochloris sulfurica" was determined by reverse transcriptase sequencing. Comparison of defined invariable parts of the molecule from representatives of 9 major lines of descent from the eubacterial kingdom shows C. sulfurica to be highly related to Chlorobium vibrioforme. The relationship between "Clathrochloris" and Chlorobium is in accord with the present allocation of these two genera into the family Chlorobiaceae.

The amino acid sequences of the cytochromes c-555 from two green sulphur bacteria of the genus Chlorobium.

Amino acid seauences are proposed for the cytochromes c-555 from Chlorobium thiosulphatophilum and from the Chlorobium limicola component of "Chloropseudomonas ethylica 2K". Each is a sincle polypeptide chain, the former of 86, the latter of 99 residues, and, when aligned so as to give the best match, 47 residues are common to the two sequences. The sequences show some resemblance to those of cytochromes c5 and f. The bacteriochlorophyll a-proteins were also isolated and purified, and their amino acid compositions compared (see the Appendix). There are significant differences in the compositions, but not as great as those found for the cytochromes c-555. The significance of these observations for the taxonomy of the Chlorobiaceae and for the further development of the comparative biochemistry of cytochrome c is discussed. Detailed evidence for the sequences of the cytochromes c-555 has been deposited as Supplementary Publication SUP 50073 (36 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1976) 153, 5.