The class Deltaproteobacteria comprises an ecologically and metabolically diverse group of bacteria best known for dissimilatory sulphate reduction and predatory behaviour. Although this lineage is the fourth described class of the phylum Proteobacteria , it rarely affiliates with other proteobacterial classes and is frequently not recovered as a monophyletic unit in phylogenetic analyses. Indeed, one branch of the class Deltaproteobacteria encompassing Bdellovibrio-like predators was recently reclassified into a separate proteobacterial class, the Oligoflexia . Here we systematically explore the phylogeny of taxa currently assigned to these classes using 120 conserved single-copy marker genes as well as rRNA genes. The overwhelming majority of markers reject the inclusion of the classes Deltaproteobacteria and Oligoflexia in the phylum Proteobacteria . Instead, the great majority of currently recognized members of the class Deltaproteobacteria are better classified into four novel phylum-level lineages. We propose the names Desulfobacterota phyl. nov. and Myxococcota phyl. nov. for two of these phyla, based on the oldest validly published names in each lineage, and retain the placeholder name SAR324 for the third phylum pending formal description of type material. Members of the class Oligoflexia represent a separate phylum for which we propose the name Bdellovibrionota phyl. nov. based on priority in the literature and general recognition of the genus Bdellovibrio. Desulfobacterota phyl. nov. includes the taxa previously classified in the phylum Thermodesulfobacteria , and these reclassifications imply that the ability of sulphate reduction was vertically inherited in the Thermodesulfobacteria rather than laterally acquired as previously inferred. Our analysis also indicates the independent acquisition of predatory behaviour in the phyla Myxococcota and Bdellovibrionota, which is consistent with their distinct modes of action. This work represents a stable reclassification of one of the most taxonomically challenging areas of the bacterial tree and provides a robust framework for future ecological and systematic studies.

Ultra-small microorganisms are ubiquitous in Earth's environments. Ultramicrobacteria, which are defined as having a cell volume of <0.1 μm 3 , are often numerically dominant in aqueous environments. Cultivated representatives among these bacteria, such as members of the marine SAR11 clade (e.g., "Candidatus Pelagibacter ubique") and freshwater Actinobacteria and Betaproteobacteria, possess highly streamlined, small genomes and unique ecophysiological traits. Many ultramicrobacteria may pass through a 0.2-μm-pore-sized filter, which is commonly used for filter sterilization in various fields and processes. Cultivation efforts focusing on filterable small microorganisms revealed that filtered fractions contained not only ultramicrocells (i.e., miniaturized cells because of external factors) and ultramicrobacteria, but also slender filamentous bacteria sometimes with pleomorphic cells, including a special reference to members of Oligoflexia, the eighth class of the phylum Proteobacteria. Furthermore, the advent of culture-independent "omics" approaches to filterable microorganisms yielded the existence of candidate phyla radiation (CPR) bacteria (also referred to as "Ca. Patescibacteria") and ultra-small members of DPANN (an acronym of the names of the first phyla included in this superphyla) archaea. Notably, certain groups in CPR and DPANN are predicted to have minimal or few biosynthetic capacities, as reflected by their extremely small genome sizes, or possess no known function. Therefore, filtered fractions contain a greater variety and complexity of microorganisms than previously expected. This review summarizes the broad diversity of overlooked filterable agents remaining in "sterile" (<0.2-μm filtered) environmental samples.

The unusual chemo-organoheterotrophic proteobacterial strain MWH-Nonnen-W8redT was isolated from a lake located in the Black Forest (Schwarzwald), Germany, by using the filtration-acclimatization method. Phylogenetic analyses based on the 16S rRNA gene sequence of the strain could not provide clear hints on classification of the strain in one of the current classes of the phylum Proteobacteria. Whole-genome sequencing resulted in a genome size of 3.5 Mbp and revealed a quite low DNA G+C content of 32.6 mol%. In-depth phylogenetic analyses based on alignments of 74 protein sequences of a phylogenetically broad range of taxa suggested assignment of the strain to a new order of the class Oligoflexia. These analyses also suggested that the order Bdellovibrionales should be transferred from the class Deltaproteobacteria to the class Oligoflexia, that this order should be split into two orders, and that the family Pseudobacteriovoracaceae should be transferred from the order Bdellovibrionales to the order Oligoflexales. We propose to establish for strain MWH-Nonnen-W8redT (=DSM 23856T=CCUG 58639T) the novel species and genus Silvanigrella aquatica gen. nov., sp. nov. to be placed in the new family Silvanigrellaceae fam. nov. of the new order Silvanigrellales ord. nov.

A phylogenetically novel proteobacterium, strain Shr3T, was isolated from sand gravels collected from the eastern margin of the Sahara Desert. The isolation strategy targeted bacteria filterable through 0.2-µm-pore-size filters. Strain Shr3T was determined to be a Gram-negative, aerobic, non-motile, filamentous bacterium. Oxidase and catalase reactions were positive. Strain Shr3T showed growth on R2A medium, but poor or no growth on nutrient agar, trypticase soy agar and standard method agar. The major isoprenoid quinone was menaquinone-7. The dominant cellular fatty acids detected were C16 : 1ω5c and C16 : 0, and the primary hydroxy acid present was C12 : 0 3-OH. The DNA G+C content was 54.0 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain Shr3T was affiliated with an uncultivated lineage of the phylum Proteobacteria; the nearest known type strain, with 83 % sequence similarity, was Desulfomicrobium orale DSM 12838T in the class Deltaproteobacteria. The isolate and closely related environmental clones formed a novel class-level clade in the phylum Proteobacteria with high bootstrap support (96–99 %). Based on these results, the novel class Oligoflexia classis nov. in the phylum Proteobacteria and the novel genus and species Oligoflexus tunisiensis gen. nov., sp. nov. are proposed for strain Shr3T, the first cultivated representative of the Oligoflexia. The type strain of Oligoflexus tunisiensis is Shr3T ( = JCM 16864T = NCIMB 14846T). We also propose the subordinate taxa Oligoflexales ord. nov. and Oligoflexaceae fam. nov. in the class Oligoflexia.

Aquatic mosses in the genera Bryum and Leptobryum form unique tower-like "moss pillars" underwater in some Antarctic lakes, in association with algae and cyanobacteria. These are communities with a two-layer structure comprising an oxidative exterior and reductive interior. Although habitats and photosynthetic properties of moss pillars have been reported, microXoral composition of the two-layer structure has not been described. Here we report fatty acid analysis of one moss pillar and molecular phylogenetic analysis, based on the 16S rRNA gene, of this and one other moss pillar. Cluster analysis of the phospholipid fatty acid composition showed three groups corresponding to the exterior, upper interior, and lower interior of the pillar. This suggested that species composition diVered by section, with the exterior dominated by photosynthetic organisms such as mosses, algae, and cyanobacteria, the upper interior primarily containing gram-positive bacteria and anaerobic sulfate-reducing bacteria, and the lower interior dominated by gram-negative bacteria. Molecular phylogenetic analysis revealed that Proteobacteria dominate the moss pillar as a whole; cyanobacteria were found on the exterior and the gram-positive obligate anaerobe Clostridium in the interior, while gram-positive sulfatereducing bacteria were present in the lowest part of the interior. Nitrogen-Wxing bacteria and denitrifying bacteria were found in all sections. Thus, fatty acid analysis and genetic analysis showed similar patterns. These Wndings suggest that microorganisms of diVerent phylogenetic groups inhabit diVerent sections of a single moss pillar and form a microbial community that performs biogeochemical cycling to establish and maintain a structure in an oxidation-reduction gradient between exterior and interior.

Aurantimicrobium minutum gen. nov., sp. nov., a novel ultramicrobacterium of the family Microbacteriaceae, isolated from river water Graduate School of Biosphere Science, Hiroshima University, 1-4-4 Kagamiyama, Higashihiroshima, Hiroshima 739-8528, Japan A Gram-stain-positive, aerobic, non-motile, curved (selenoid), rod-shaped actinobacterium, designated KNC T , was isolated from the 0.2 mm-filtrate of river water in western Japan. Cells of strain KNC T were ultramicrosized (0.04-0.05 mm 3 ). The strain grew at 15-37 8C, with no observable growth at 10 8C or 40 8C. The pH range for growth was 7-9, with weaker growth at pH 10. Growth was impeded by the presence of NaCl at concentrations greater than 1 %. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain KNC T showed relatively high sequence similarity (97.2 %) to Alpinimonas psychrophila Cr8-25 T in the family Microbacteriaceae. However, strain KNC T formed an independent cluster with cultured, but as-yetunidentified, species and environmental clones on the phylogenetic tree. The major cellular fatty acids were anteiso-C 15 : 0 (41.0 %), iso-C 16 : 0 (21.8 %), C 16 : 0 (18.0 %) and anteiso-C 17 : 0 (12.9 %), and the major menaquinones were MK-11 (71.3 %) and MK-12 (13.6 %). The major polar lipids were phosphatidylglycerol and two unknown glycolipids. The cell-wall muramic acid acyl type was acetyl. The peptidoglycan was B-type, and contained 3-hydroxyglutamic acid, glutamic acid, aspartic acid, glycine, alanine and lysine, with the latter being the diagnostic diamino acid. The G+C content of the genome was unusually low for actinobacteria (52.1 mol%), compared with other genera in the family Microbacteriaceae. Based on the phenotypic characteristics and phylogenetic evidence, strain KNC T represents a novel species of a new genus within the family Microbacteriaceae, for which the name Aurantimicrobium minutum gen. nov., sp. nov. is proposed. The type strain of the type species is KNC T (5NBRC 105389 T 5NCIMB 14875 T ).The family Microbacteriaceae consists of predominantly aerobic Gram-positive actinobacteria that are differentiated from other families by a combination of their unusual cellwall peptidoglycan, unsaturated respiratory menaquinones and irregular cell shapes (Park et al., 1993;Evtushenko & Takeuchi, 2006). At the time of writing, the family comprised 44 genera with validly published names, including the recently described genera Glaciihabitans (Li et al., 2014) and Rhodoluna (Hahn et al., 2014).During the course of a study of ultramicrosized bacteria that could pass through 0.2 mm pore-size filters, which are generally used for sterile filtration to remove microorganisms from fluids, we isolated strains belonging to several novel taxa from various environments (Nakai et al., 2013(Nakai et al., , 2014. Here, one isolate, designated KNC T was characterized using a polyphasic approach. The results showed that strain KNC T represents a novel ultramicrobacterial lineage of the family Microbacteriaceae.Strain KNC T was originally i...

Although the lower size limit of microorganisms was previously believed to be c. 0.2 mm, there is evidence for the existence of microorganisms that can pass through 0.2 mm-pore-size filters called ultramicrobacteria or nanobacteria. However, information on the phylogeny and biogeography of these bacteria is limited. We obtained 53 isolates of 0.2 mm-passable bacteria from 31 samples collected at 26 locations worldwide, including the Arctic Svalbard Islands, deserts, and Maritime Antarctica. Phylogenetic analysis of near full-length 16S rRNA gene sequences revealed that 18 of the 53 isolates were , 97% homologous with previously cultured isolates, representing potentially novel species. Two isolates (order Rhizobiales) (100% identical) collected from Byers Peninsula, Livingston Island in Maritime Antarctica, were closely related (99.8% similarity) to an isolate collected from intertidal sediments in East Antarctica. In addition, the sequence of this Antarctic isolate showed $ 97% similarity to 901 sequences derived from known isolates and samples collected at geographically disparate locations under various environmental conditions. Interestingly, among 13 sequences showing $ 99% similarity, ten were isolated from cryospheric habitats such as Arctic, Antarctic, and alpine environments. This implies that such Rhizobiales strains occur in the cryospheric regions, however, their abundance and biomass may be scarce depending on the geographic location.

One highly diverse phylogenetic group of Bacteria, Ca . Patescibacteria, remains poorly understood, but, from the few cultured representatives and metagenomic investigations, they are thought to live symbiotically or parasitically with other bacteria or even with eukarya.