Clostridiisalibacter paucivorans gen. nov., sp. nov., a novel moderately halophilic bacterium isolated from olive mill wastewater.

A moderately halophilic, strictly anaerobic bacterium, designated 37HS60(T), was isolated from an olive mill wastewater in southern Morocco (Marrakesh). The cells were straight, motile and stained Gram-positive, forming spherical and terminal spores and with an atypical thick and stratified multilayered cell wall. Major fatty acid components were iso-C17:1omega10c or anteiso-C17:1omega3c (19.3%), C14:0 (14.3%), C16:1omega7c (9.9%), C16:1omega7c DMA (8.5%) and C16:0 (7.6%). Strain 37HS60(T) grew from 20 to 50 degrees C with an optimum at 40-45 degrees C, and growth was observed from pH 5.5 to 8.5 with an optimum of 6.8. The salinity range for growth was 10-100 g NaCl l(-1) with an optimum at 50 g NaCl l(-1). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain 37HS60(T) fell within the evolutionary radiation enclosed by cluster XII of the Clostridium subphylum. Strain 37HS60(T) exhibited highest 16S rRNA gene sequence similarity of 92.0% with Caloranaerobacter azorensis and 90.6% with Clostridium purinilyticum. Moreover, 37HS60(T) did not grow on basal medium with hexose or pentose sugars as carbon and energy sources. Pyruvate, fumarate and succinate were the best substrates for 37HS60(T) growth with 1.0 g yeast extract l(-1). The DNA G+C content was 33.0 mol%. Due to its phenotypic and genotypic characteristics, isolate 37HS60(T) is proposed as a novel species of a new genus, Clostridiisalibacter paucivorans gen. nov., sp. nov. The type strain is 37HS60(T) (=JCM 14354(T)=CCUG 53849(T)).

[Phenotypic properties of Sulfobacillus thermotolerans: comparative aspects].

The phenotypic characteristics of the species Sulfobacillus thermotolerans Kr1(T), as dependent on the cultivation conditions, are described in detail. High growth rates (0.22-0.30 h(-1)) and high oxidative activity were recorded under optimum mixotrophic conditions at 40 degrees C on medium with inorganic (Fe(II), S(0), or pyrite-arsenopyrite concentrate) and organic (glucose and/or yeast extract) substrates. In cells grown under optimum conditions on medium with iron, hemes a, b, and, most probably, c were present, indicating the presence of the corresponding cytochromes. Peculiar extended structures in the form of cylindrical cords, never observed previously, were revealed; a mucous matrix, likely of polysaccharide nature, occurred around the cells. In the cells of sulfobacilli grown litho-, organo-, and mixotrophically at 40 degrees C, the enzymes of the three main pathways of carbon utilization and some enzymes of the TCA cycle were revealed. The enzyme activity was maximum under mixotrophic growth conditions. The growth rate in the regions of limiting temperatures (55 degrees C and 12-14 degrees C) decreased two- and tenfold, respectively; no activity of 6-phosphogluconate dehydrogenase, one of the key enzymes of the oxidative pentose phosphate pathway, could be revealed; and a decrease in the activity of almost all enzymes of glucose metabolism and of the TCA cycle was observed. The rate of 14CO2 fixation by cells under auto-, mixo-, and heterotrophic conditions constituted 31.8, 23.3, and 10.3 nmol/(h mg protein), respectively. The activities of RuBP carboxylase (it peaked during lithotrophic growth) and of carboxylases of heterotrophic carbon dioxide fixation were recorded. The physiological and biochemical peculiarities of the thermotolerant sulfobacillus are compared versus moderately thermophilic sulfobacilli.

Proposal of Viridibacillus gen. nov. and reclassification of Bacillus arvi, Bacillus arenosi and Bacillus neidei as Viridibacillus arvi gen. nov., comb. nov., Viridibacillus arenosi comb. nov. and Viridibacillus neidei comb. nov.

A polyphasic study was undertaken to clarify the taxonomic position of endospore-forming strains 433-D9, 433-E17 and 121-X1. BOX-PCR-generated fingerprints indicated that they may be members of a single species. 16S rRNA gene sequence similarity demonstrated that a representative of this group, 433-D9, is affiliated closely with Bacillus arvi DSM 16317(T) (100 %), Bacillus arenosi DSM 16319(T) (99.8 %) and Bacillus neidei NRRL BD-87(T) (97.1 %). Sequence similarities revealed Bacillus pycnus NRRL NRS-1691(T) and several Kurthia species as the next nearest relatives. DNA-DNA hybridization results showed that strain 433-D9 is a member of B. arvi. Detection of l-Lys-d-Asp-based peptidoglycan in strain 433-D9, B. arvi DSM 16317(T) and B. arenosi DSM 16319(T) was in agreement with their close relationship, but differentiated these strains from B. neidei NRRL BD-87(T) and B. pycnus NRRL NRS-1691(T), for which l-Lys-d-Glu was reported. A similar quinone system was detected in strains 433-D9, 433-E17, 121-X1, B. arvi DSM 16317(T), B. arenosi DSM 16319(T) and B. neidei NRRL BD-87(T). This system, unusual for bacilli, consisted of the major compound menaquinone MK-8 (69-80 %) and moderate amounts of MK-7 (19-30 %). This observation was in contrast to the predominance of MK-7 of the closest relative B. pycnus NRRL NRS-1691(T), as also reported for representatives of the closely related non-endospore-forming genus Kurthia. Strains 433-D9, B. arvi DSM 16317(T) and B. arenosi DSM 16319(T) exhibited homogeneous and discriminative polar lipid profiles and fatty acid profiles consisting of major acids i-C(15 : 0) and ai-C(15 : 0) and moderate amounts of i-C(17 : 1)omega10c and i-C(17 : 1) I/ai-C(17 : 1) B that discriminated them from closely related strains such as B. neidei NRRL BD-87(T). On the basis of clear-cut discriminative chemotaxonomic markers, we propose strains 433-D9, 433-E17 and 121-X1, B. arvi DSM 16317(T), B. arenosi DSM 16319(T) and B. neidei NRRL BD-87(T) to be reclassified within a separate genus. For this new taxon, we propose the name Viridibacillus gen. nov., and we propose the reclassification of Bacillus arvi, Bacillus arenosi and Bacillus neidei as Viridibacillus arvi gen. nov., comb. nov. (the type species of Viridibacillus, with the type strain DSM 16317(T) =LMG 22165(T)), Viridibacillus arenosi comb. nov. (type strain DSM 16319(T) =LMG 22166(T)) and Viridibacillus neidei comb. nov. (type strain NRRL BD-87(T) =DSM 15031(T) =JCM 11077(T)).

Cohnella laeviribosi sp. nov., isolated from a volcanic pond.

A novel thermophilic and endospore-forming Gram-positive bacterium capable of assimilating and isomerizing l-ribose was isolated from a volcanic area in Likupang, Indonesia. The isolate, RI-39(T), was able to grow at high temperatures (37-60 degrees C); optimum growth was observed at pH 6.5 and 45 degrees C. The strain contained MK-7 (87 %) as the main respiratory quinone and had a DNA G+C content of 51 mol%. The major cellular fatty acids of the isolate were iso-C(16 : 0) and anteiso-C(15 : 0) and the predominant polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and lysyl-phosphatidylglycerol. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the isolate represents an evolutionary lineage that is distinct from those of other Cohnella species. Based on morphological, physiological and chemotaxonomic characteristics and 16S rRNA gene sequence comparisons, it is proposed that strain RI-39(T) represents a novel species, Cohnella laeviribosi sp. nov. The type strain is RI-39(T) (=KCTC 3987(T) =KCCM 10653P(T) =CCUG 52217(T)).

Proteome analysis of Paenibacillus polymyxa E681 affected by barley.

Paenibacillus polymyxa E681 is known to be able to suppress plant diseases by producing antimicrobial compounds and to promote plant growth by producing phytohormones, and secreting diverse degrading enzymes. In spite of these capabilities, little is known regarding the flow of information from the bacterial strain to the barley roots. In an attempt to determine the flow of information from the bacterial strain to barley roots, the train was grown in the presence and absence of barley, and two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and MALDI-TOF mass spectrometry were used. 2D-PAGE detected approximately 1000 spots in the cell and 1100 spots in the supernatant at a pH 4-10 gradient. Interestingly, about 80 spots from each sample showed quantitative variations. Fifty-three spots from these were analyzed by MALDI-TOF mass spectrometry and 28 proteins were identified. Most of the cytosolic proteins expressed at higher levels were found in P. polymyxa E681 cells grown in the presence of barley rather than in the absence of barley. Proteins detected at a lower level in the surpernatant of P. polymyxa E681 cells grown in the presence of barley were lipoprotein, glucose-6-phosphate 1-dehydrogenase, heat-shock protein HtpG spermidine synthase, OrfZ, ribonuclease PH, and coenzyme PQQ synthesis protein, and flagellar hook-associated protein 2 whereas proteins detected at a higher level in the surpernatant of P. polymyxa E681 cells grown in the presence of barley included D-alanyl-D-alanine ligase A, isopentenyldiphosphate delta-isomerase, ABC transporter ATP-binding protein Uup, lipase. Many of the proteins belonging to plant-induced stimulons are associated with biosynthetic metabolism and metabolites of proteins and transport. Some of these proteins would be expected to be induced by environmental changes resulting from the accumulation of plant-secreted substances.

Paenibacillus humicus sp. nov., isolated from poultry litter compost.

Two bacterial strains, PC-142 and PC-147(T), isolated from poultry litter compost, were characterized with respect to their phenetic and phylogenetic characteristics. The isolates were endospore-forming rods that were reddish in colour after Gram staining. They were catalase- and oxidase-positive, were able to degrade starch and gelatin and grew at 15-40 degrees C and pH 5.5-10.0. The predominant fatty acids were anteiso-C(15 : 0), iso-C(15 : 0) and iso-C(16 : 0), the major respiratory quinone was menaquinone MK-7, the cell-wall peptidoglycan was of the A1gamma type and the G+C content of the DNA was 58 mol%. The 16S rRNA gene sequence analysis and phenetic characterization indicated that these organisms belong to the genus Paenibacillus, with Paenibacillus pasadenensis SAFN-007(T) as the closest phylogenetic neighbour (97.5 %). Strains PC-142, PC-147(T) and P. pasadenensis SAFN-007(T) represent a novel lineage within the genus Paenibacillus, characterized by a high DNA G+C content (58-63 mol%). The low levels of 16S rRNA gene sequence similarity with respect to other taxa with validly published names and the identification of distinctive phenetic features in the two isolates indicate that strains PC-142 and PC-147(T) represent a novel species of the genus Paenibacillus, for which the name Paenibacillus humicus sp. nov. is proposed. The type strain is PC-147(T) (=DSM 18784(T) =NBRC 102415(T) =LMG 23886(T)).

Desulfovirgula thermocuniculi gen. nov., sp. nov., a thermophilic sulfate-reducer isolated from a geothermal underground mine in Japan.

A thermophilic, Gram-positive, endospore-forming, sulfate-reducing bacterial strain, designated RL80JIV(T), was isolated from a geothermally active underground mine in Japan. Cells were rod-shaped and motile. The temperature and pH ranges for growth were 61-80 degrees C (optimum at 69-72 degrees C) and pH 6.4-7.9 (optimum at pH 6.8-7.3), and the strain tolerated up to 0.5 % NaCl. Strain RL80JIV(T) utilized sulfate, sulfite, thiosulfate and elemental sulfur as electron acceptors. Electron donors utilized were H(2) in the presence of CO(2), and carboxylic acids. Fermentative growth occurred on lactate and pyruvate. The cell wall contained meso-diaminopimelic acid and the major respiratory isoprenoid quinone was menaquinone MK-7. Major whole-cell fatty acids were iso-C(15 : 0), iso-C(17 : 0) and C(16 : 0). Strain RL80JIV(T) was found to be affiliated with the thiosulfate-reducer Thermanaeromonas toyohensis DSM 14490(T) (90.9 % 16S rRNA gene sequence similarity) and with the sulfate-reducer Desulfotomaculum thermocisternum DSM 10259(T) (90.0 % similarity). Strain RL80JIV(T) is therefore considered to represent a novel species of a new genus, for which the name Desulfovirgula thermocuniculi gen. nov., sp. nov. is proposed. The type strain of Desulfovirgula thermocuniculi is RL80JIV(T) (=DSM 16036(T)=JCM 13928(T)).

Paenibacillus ginsengisoli sp. nov., a novel bacterium isolated from soil of a ginseng field in Pocheon Province, South Korea.

A Gram-positive, aerobic or facultative anaerobic, motile, spore-forming bacterial strain, designated Gsoil 1638T, was isolated from a soil sample of a ginseng field in Pocheon province (South Korea), and was characterized taxonomically by using a polyphasic approach. It grew well on nutrient agar medium, utilized a fairly narrow spectrum of carbon sources and tolerated 10% NaCl. The isolate was positive for catalase and oxidase tests but negative for the degradation of macromolecules such as casein, collagen, starch, chitin, CM-cellulose, xylan and DNA. The G + C content of the genomic DNA was 50.7 mol%. The predominant isoprenoid quinone was menaquinone 7 (MK-7). The major fatty acids were anteiso-C15:0 (44%) and C16:0 (25%). Comparative 16S rRNA gene sequence analysis showed that strain Gsoil 1638T fell within the radiation of the cluster comprising Paenibacillus species and joined Paenibacillus anaericanus DSM 15890T with a bootstrap value of 100%. These two strains shared 99.5% 16S rRNA gene sequence similarity with each other. The phylogenetic distance from any other validly described species within the genus Paenibacillus was less than 96.2%. DNA-DNA relatedness value between strain Gsoil 1638T and its closest phylogenetic neighbor, Paenibacillus anaericanus, was 62%. On the basis of its phenotypic properties and phylogenetic distinctiveness, strain Gsoil 1638T (= KCTC 13931T = LMG 23406T = CCUG 52472T) was classified in the genus Paenibacillus as the type strain of a novel species, for which the name Paenibacillus ginsengisoli sp. nov. is proposed.

Virgibacillus koreensis sp. nov., a novel bacterium from a salt field, and transfer of Virgibacillus picturae to the genus Oceanobacillus as Oceanobacillus picturae comb. nov. with emended descriptions.

A novel Virgibacillus strain, BH30097T, was isolated from a salt field near Taean-Gun on the Yellow Sea in Korea. Cells were Gram-positive rods and bore ellipsoidal endospores in terminal positions. The optimum pH and temperature for growth of this organism were pH 7 and 25 degrees C, respectively. The main respiratory quinone was MK-7 and the major cellular fatty acids were anteiso-C(15 : 0) and iso-C(16 : 0). Analysis based on 16S rRNA gene sequence data revealed that the isolate formed an evolutionary lineage distinct from other Virgibacillus species. Levels of sequence similarity between the isolate and other Virgibacillus species ranged from 93.8 to 96.7 %. DNA-DNA relatedness values between strain BH30097T and a phylogenetically closely related strain, Virgibacillus halodenitrificans KCTC 3790T, were less than 24 %. On the basis of morphological, physiological and chemotaxonomic characteristics, 16S rRNA gene sequence comparison and DNA-DNA hybridization, a novel species, Virgibacillus koreensis sp. nov., is proposed, with the type strain BH30097T (= KCTC 3823T = JCM 12387T). It is also proposed that Virgibacillus picturae be transferred to the genus Oceanobacillus as Oceanobacillus picturae comb. nov. based on its 16S rRNA gene sequences and other taxonomic characteristics.

A novel Acetivibrio cellulolyticus anchoring scaffoldin that bears divergent cohesins.

Sequencing of a cellulosome-integrating gene cluster in Acetivibrio cellulolyticus was completed. The cluster contains four tandem scaffoldin genes (scaA, scaB, scaC, and scaD) bounded upstream and downstream, respectively, by a presumed cellobiose phosphorylase and a nucleotide methylase. The sequences and properties of scaA, scaB, and scaC were reported previously, and those of scaD are reported here. The scaD gene encodes an 852-residue polypeptide that includes a signal peptide, three cohesins, and a C-terminal S-layer homology (SLH) module. The calculated molecular weight of the mature ScaD is 88,960; a 67-residue linker segment separates cohesins 1 and 2, and two approximately 30-residue linkers separate cohesin 2 from 3 and cohesin 3 from the SLH module. The presence of an SLH module in ScaD indicates its role as an anchoring protein. The first two ScaD cohesins can be classified as type II, similar to the four cohesins of ScaB. Surprisingly, the third ScaD cohesin belongs to the type I cohesins, like the seven ScaA cohesins. ScaD is the first scaffoldin to be described that contains divergent types of cohesins as integral parts of the polypeptide chain. The recognition properties among selected recombinant cohesins and dockerins from the different scaffoldins of the gene cluster were investigated by affinity blotting. The results indicated that the divergent types of ScaD cohesins also differ in their preference of dockerins. ScaD thus plays a dual role, both as a primary scaffoldin, capable of direct incorporation of a single dockerin-borne enzyme, and as a secondary scaffoldin that anchors the major primary scaffoldin, ScaA and its complement of enzymes to the cell surface.

Structure, diversity, and evolution of protein toxins from spore-forming entomopathogenic bacteria.

Gram-positive spore-forming entomopathogenic bacteria can utilize a large variety of protein toxins to help them invade, infect, and finally kill their hosts, through their action on the insect midgut. These toxins belong to a number of homology groups containing a diversity of protein structures and modes of action. In many cases, the toxins consist of unique folds or novel combinations of domains having known protein folds. Some of the toxins display a similar structure and mode of action to certain toxins of mammalian pathogens, suggesting a common evolutionary origin. Most of these toxins are produced in large amounts during sporulation and have the remarkable feature that they are localized in parasporal crystals. Localization of multiple toxin-encoding genes on plasmids together with mobilizable elements enables bacteria to shuffle their armory of toxins. Recombination between toxin genes and sequence divergence has resulted in a wide range of host specificities.

Bacillus salexigens sp. nov., a new moderately halophilic Bacillus species.

Bacillus salexigens sp. nov. is proposed based on the characteristics of six moderately halophilic, grampositive, rod-shaped strains isolated from salterns and hypersaline soils located in different geographical areas of Spain. These strains were motile, formed endospores, were strictly aerobic, were catalase and oxidase positive, and contained peptidoglycan of the meso-diamlnopimelic acid type in their vegetative cell walls. The DNA base compositions of these strains ranged from 36.3 to 39.5 mol%, and these organisms constitute a homology group with levels of DNA-DNA homology ranging from 73 to 100%. The 16S rRNA sequence of strain C-20MoT, which was used as the representative strain of these isolates, groups with the 16S rRNA sequences of members of the genus Bacillus, and the highest level of similarity is 95.4%. The type strain is strain C-20Mo (= ATCC 700290 = DSM 11483 = CCM 4646).