- Volume 46, Issue 4, 1996
Volume 46, Issue 4, 1996
- Original Papers Relating To Systematic Bacteriology
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Novel Psychrobacter Species from Antarctic Ornithogenic Soils
More LessOrnithogenic soil is derived from the deposition of the fecal matter of various species of birds and is a major source of nutrient input in the Antarctic marine ecosystem. A significant proportion of the microbiota of ornithogenic soil collected from an Adélie penguin colony in eastern Antarctica (Vestfold Hills ice-free zone) consisted of gram-negative, coccoid bacteria identified on the basis of their phospholipid ester-linked fatty acid and lipid class profiles as Psychrobacter strains. Phenotypic, genotypic, and 16S ribosomal DNA phylogenetic analyses revealed that the Antarctic psychrobacters belonged to three distinct groups. Comparisons with Psychrobacter immobilis and Moraxella phenylpyruvica reference cultures isolated from fish, seawater, poultry, and human clinical specimens revealed the relationships of these groups within the genus Psychrobacter. Two of the groups represent the following two novel species: Psychrobacter urativorans sp. nov. (type strain, strain ACAM 534) and Psychrobacter frigidicola sp. nov. (type strain, strain ACAM 304). The third group of strains included members of the previously described species P. immobilis (Juni and Heym 1986). In addition, M. phenylpyruvica (Bøvre and Henriksen 1967) is renamed Psychrobacter phenylpyruvicus comb. nov. (type strain, strain ACAM 535) on the basis of 16S ribosomal DNA phylogenetic data. In general, the genus Psychrobacter could be differentiated from the related genera Moraxella and Acinetobacter by the fact that the members of the genus Psychrobacter are psychrotolerant or psychrophilic and halotolerant, which reflects the ubiquitous distribution of the genus in both marine and terrestrial environments. On the basis of the results of this and previous studies, the genus Psychrobacter is the predominant genus in ornithogenic soils in Antarctica and is diverse.
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Bordetella trematum sp. nov., Isolated from Wounds and Ear Infections in Humans, and Reassessment of Alcaligenes denitrificans Rüger and Tan 1983
Ten strains recognized on the basis of a computer-assisted numerical comparison of whole-cell protein patterns as members of a novel species belonging to the family Alcaligenaceae were examined by using an integrated phenotypic and genotypic approach. This species, for which we propose the name Bordetella trematum sp. nov., was more closely related to the type species of the genus Bordetella (Bordetella pertussis) than to the type species of the genus Alcaligenes (Alcaligenes faecalis) and had the general characteristics of members of this family (i.e., a DNA base ratio in the range from 57 to 70 mol%, a fatty acid profile characterized by high percentages of 16:0, 17:0 cyclo, and 14:0 30H, nonsaccharolytic metabolism, and several classical biochemical characteristics, including aerobic and microaerobic growth, catalase activity, assimilation of citrate, an absence of anaerobic growth, and an absence of acetylmethylcarbinol and indole production, gelatin liquefaction, and esculin hydrolysis). A reevaluation of the criteria used to classify Alcaligenes denitrificans Rüger and Tan 1983 and Achromobacter xylosoxidans Yabuuchi and Ohyama 1971 as subspecies of Alcaligenes xylosoxidans and additional evidence provided in recent studies revealed that, consistent with present standards, it is appropriate to consider these two taxa distinct species of the genus Alcaligenes.
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Phylogenesis of Relapsing Fever Borrelia spp.
More LessThe phylogenetic relationships of 20 relapsing fever (RF) Borrelia spp. were estimated on the basis of the sequences of rrs genes. Complete sequences were aligned and compared with previously published sequences, and the similarity values were found to be 97.7 to 99.9%. Phylogenetic trees were constructed by using the three neighbor-joining, maximum-parsimony, and maximum-likelihood methods. The results of the comparative phylogenetic analysis divided the RF Borrelia spp. into three major clusters. One cluster included Borrelia crocidurae, Borrelia duttonii, Borrelia recurrentis, and Borrelia hispanica. Another cluster comprised two main branches with Borrelia coriaceae, Borrelia lonestari, and Borrelia miyamotoi on one side and Borrelia parkeri, Borrelia turicatae, and Borrelia hermsii on the other side. Borrelia anserina constituted the third cluster. The phylogenetic position of Borrelia persica was more uncertain. These results suggested that the taxonomy of these spirochetes should be revised. To overcome the problems of culturing the spirochetes, RF Borrelia primers were defined. Following PCR amplification of the rrs gene, restriction length fragment polymorphism could be used to distinguish between RF Borrelia strains.
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Emendation of the Genus Planococcus and Transfer of Flavobacterium okeanokoites Zobell and Upham 1944 to the Genus Planococcus as Planococcus okeanokoites comb. nov.
More LessThe taxonomic position of Flavobacterium okeanokoites IFO 12536T (T = type strain) was determined by 16S rRNA gene sequencing and chemotaxonomic methods. Phylogenetic evidence derived from a 16S rRNA sequence analysis indicated that F. okeanokoites, which forms rod-shaped cells, belongs to the genus Planococcus, which forms spherical cells. A phylogenetically close relationship was supported by chemotaxonomic characteristics, such as the presence of menaquinone 7 and menaquinone 8 as major isoprenoid quinones, the presence of phosphatidylglycerol, bisphosphatydylglycerol, and phosphatidylethanolamine as cellular polar lipids, and the G+C content of the DNA (46.3 mol%). These data suggest that whether a cell is a rod or a coccus is not a generic criterion. Accordingly, we propose that F. okeanokoites should be transferred to the genus Planococcus and that the description of the genus Planococcus should be emended.
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Anaerofilum pentosovorans gen. nov., sp. nov., and Anaerofilum agile sp. nov., Two New, Strictly Anaerobic, Mesophilic, Acidogenic Bacteria from Anaerobic Bioreactors †
More LessStrictly anaerobic, gram-positive, nonsporing, thin rod-shaped organisms whose cells were 0.2 to 0.6 by 3 to 6 μm were isolated from a Hoechst Biohochreaktor (strain FaeT [T = type strain]) and from the biofilm population of a fixed-film reactor treating sour whey (strain FT). Strain FT was vigorously motile during early logarithmic growth by means of peritrichously inserted flagella, while strain FaeT was seldom motile and usually possessed no flagella. During the stationary growth phase both strains formed spheroplasts. The temperature optimum was close to 37°C (temperature range for growth, ≥17 to <45°C) and the pH optimum was 7.0 to 7.4 (pH range, 6.5 to 8.0) for both strains. The two organisms grew chemoorganotrophically on a number of mono- and disaccharides, including glucose and xylose; yeast extract was required for growth. The principal fermentation products from glucose included lactate, acetate, ethanol, formate, and CO2. Hydrogen was not generated. The G+C contents of the DNAs of strains FaeT and FT were 55 and 54.5 mol%, respectively. The cell wall architecture was typical of gram-positive bacteria; the cells had an extraordinarily thin type A3α peptidoglycan layer containing muramic acid. Analysis of 16S ribosomal DNA sequences of the two new isolates demonstrated that they represent members of a new genus of bacteria in Clostridium cluster IV of the domain Bacteria and that the misclassified organism Fusobacterium prausnitzii and Clostridium leptum are among their closest relatives. The names Anaerofilum pentosovorans gen. nov., sp. nov. (type strain, strain Fae [= DSM 7168]) and Anaerofilum agile sp. nov. (type strain, strain F [= DSM 4272]) are proposed.
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Gordona hirsuta sp. nov.
More LessBacterial strain K718aT (T = type strain), which was isolated from the packing material of a biofilter used for deodorization of animal-rendering plant emissions, was subjected to a polyphasic taxonomic study in which physiological, chemotaxonomic, and genomic methods were used. On the basis of the chemotaxonomic and physiological properties found and the results of 16S ribosomal DNA sequence comparisons, it was evident that strain K718aT belongs to a new species in the genus Gordona. We propose that strain K718a should be the type strain of the new species Gordona hirsuta.
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Reduction of Benzyl Viologen Distinguishes Genera of the Class Mollicutes
We tested the ability of 62 growing strains belonging to the class Mollicutes to reduce the redox indicator and free-radical generator 1,1’-dibenzyl-4,4’-bipyridinium dichloride (benzyl viologen [BV]) to a blue-violet-purple color. BV was reduced by 12 Acholeplasma species but not by Acholeplasma multiforme PN525T (T = type strain). BV was also reduced by five of nine Mesoplasma species and by four of six Entomoplasma species. BV was not reduced by 19 Mycoplasma species, six Spiroplasma species, five unnamed Spiroplasma strains belonging to different serogroups, three Ureaplasma species, and one unnamed Ureaplasma strain. The BV-reducing ability was localized in the membrane of Acholeplasma laidlawii B-PG9 and was dependent on NADH. Reduction of BV could be expressed in mixed cultures, and this activity may be useful for recognizing the contaminating presence of an Acholeplasma species. The reductive BV response may have phylogenetic value. We believe that the test described in this paper readily distinguishes all Acholeplasma species and some Mesoplasma and Entomoplasma species from all Mycoplasma, Spiroplasma, and Ureaplasma species tested.
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Comparative Metabolism of Mesoplasma, Entomoplasma, Mycoplasma, and Acholeplasma
Cytoplasmic fractions from species of the Mollicutes genera Entomoplasma, Mesoplasma, Mycoplasma, and Acholeplasma were assayed for NADH oxidase (NADH ox), ATP- and PPi-dependent phosphofructokinase (PFK), ATP- and PPi-dependent deoxyguanosine kinase (dGUOK), thymidine kinase (TK), TMP kinase (TMPK), glucose-6-phosphate dehydrogenase (G6Pde), lactate dehydrogenase (LDH), malate dehydrogenase (MDH), phosphoenolpyruvate carboxylase, hypoxanthine-guanine phosphoribosyl transferase, dUTPase, and uracil-DNA glycosylase (UNG) activities. Membrane fractions were also examined for NADH ox activity. These activities were used as indicators of the presence and relative activities of major Mollicutes metabolic and DNA repair pathways. This was the first study to determine the presence of these enzymes in members of the genera Entomoplasma and Mesoplasma. Using the data obtained, we constructed a preliminary scheme for distinguishing genera of the class Mollicutes on the basis of the results of signature functional enzyme assays. This scheme includes phylogenetic relationships deduced from rRNA analyses, but is more informative with respect to metabolic potential. The criteria used include the presence of PPi-dependent PFK, urease, dUTPase, and dGUOK activities. Entomoplasma ellychniae ELCN-1T (T = type strain), Entomoplasma melaleucae M-1T, Mesoplasma seiffertii F7T, Mesoplasma entomophilum TACT, Mesoplasma florum L1T, Mycoplasma fermentans PG18T, and Acholeplasma multilocale PN525T were similar in most respects. NADH ox activity was localized in the cytoplasm of these organisms. These strains had ATP-dependent PFK, MDH, LDH, ATP- and PPi-dependent dGUOK, and UNG activities, but not dUTPase or G6Pde activities. In contrast, Acholeplasma equifetale C112T, Acholeplasma oculi 19LT, Acholeplasma hippikon C1T, Acholeplasma modicum PG49T, and Acholeplasma morum 72-043T had membrane-localized NADH ox activity, PPi-dependent PFK, G6Pde, and dUTPase activities, and significantly lower MDH and LDH activities and exhibited a faster rate with PPi than with ATP in the dGUOK reaction. All of the members of the Mollicutes tested had hypoxanthine-guanine phosphoribosyl transferase, phosphoenolpyruvate carboxylase, and (except for Mesoplasma entomophilum TACf) UNG activities. All of the Acholeplasma strains except Acholeplasma multilocale PN525T had TK, TMPK, and UNG activities. Mesoplasma entomophilum TACT was distinguished by having no detectable dUTPase, UNG, TK, and TMPK activities, indicating that there is a severe restriction in or an absence of a synthetic route to dTTP. Our data also suggest that A. multilocale PN525T is a member of an unrecognized metabolic subgroup of the genus Acholeplasma or is not an Acholeplasma strain.
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Comparison of Partial Citrate Synthase Gene (gltA) Sequences for Phylogenetic Analysis of Bartonella Species
More LessNucleotide base sequence data were obtained for a 940-bp fragment of the citrate synthase-encoding gene (gltA) of representatives of the eight validly described Bartonella species and seven uncharacterized Bartonella strains obtained from small mammals. Complete 16S rRNA gene sequences were also determined for the uncharacterized strains, and these sequences revealed that each strain had a unique sequence which was very similar to the sequences of the previously recognized Bartonella species. A comparison of the gltA sequences of the different Bartonella species revealed that the levels of similarity between sequences were 83.8 to 93.5%, whereas comparisons of sequences obtained from different strains of the same species revealed that the levels of similarity were more than 99.8%. One of the uncharacterized strains had a gltA sequence that matched the sequence of Bartonella elizabethae, three uncharacterized strains had sequences which were more than 99.6% similar to each other (but less than 93.5% similar to any other sequence), and the remaining three uncharacterized strains each exhibited less than 93.5% sequence similarity to other Bartonella species or isolates. Phylogenetic trees were inferred from multiple alignments of both gltA and 16S ribosomal DNA (rDNA) sequences. Whereas the proposed intra-Bartonella architecture of trees inferred from 16S rDNA sequence data by using both distance matrix and parsimony methods had virtually no statistical support, the trees inferred from the gltA sequence data contained four well-supported lineages in the genus. The gltA-derived phylogeny appears to be more useful than the phylogeny derived from 16S rDNA sequence data for investigating the evolutionary relationships of Bartonella species, and the validity of the lineages identified by the gltA analysis is discussed in this paper.
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Phylogenetic Analysis of Borrelia Species Based on Flagellin Gene Sequences and Its Application for Molecular Typing of Lyme Disease Borreliae
More LessWe determined almost complete flagellin gene sequences of various Borrelia species and aligned them with previously published sequences. A neighbor-joining phylogenetic analysis showed that the genus Borrelia was divided into the following three major clusters: New World relapsing fever borreliae (Borrelia turicatae, Borrelia parkeri and Borrelia hermsii), Old World relapsing fever borreliae (Borrelia crocidurae, Borrelia duttonii, and Borrelia hispanica), and Lyme disease borreliae (Borrelia burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii). Agents of animal spirochetosis (Borrelia coriaceae and Borrelia anserina) and species of unknown pathogenicity (Borrelia miyamotoi and Borrelia lonestari) were related to relapsing fever borreliae. Although the Lyme disease borreliae, two related species (Borrelia japonica and Borrelia andersonii), and some newly described genomic groups (groups PotiB2, VS116, DN127, Hk501, and Ya501) were closely related to each other, each taxon formed an independent branch on the phylogenetic tree. The data obtained in this study indicate that the flagellin genes are useful in Borrelia taxonomy. To distinguish the Lyme disease borreliae from related organisms easily, we designed an oligonucleotide primer set for the flagellin gene and performed a PCR-restriction fragment length polymorphism (PCR-RFLP) analysis. The primer set amplified an approximately 580-bp DNA fragment that included species-specific restriction sites, and Hapll, Hhal, CelII, HincII, or Ddel digestion of the product resulted in distinctively different PCR-RFLP patterns. The PCR-RFLP typing method which we developed should facilitate rapid identification of Lyme disease borreliae and related organisms obtained from biological and clinical specimens.
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Spiroplasma leptinotarsae sp. nov., a Mollicute Uniquely Adapted to Its Host, the Colorado Potato Beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae)
Spiroplasma strain LD-1T (T = type strain), which was isolated from the gut of a Colorado potato beetle (Leptinotarsa decemlineata) larva collected in Maryland, was serologically distinct from other spiroplasmas. Similar isolates were obtained from other L. decemlineata specimens collected in various parts of North America, in Poland, and in other eastern European countries and from Leptinotarsa texana specimens collected in Texas. Cells of strain LD-1T, which in early passages were spiral, exhibited exceptionally rapid translational motility. This rapid motility and the spiral shape were lost after extended passage in culture. The organism required serum for growth. Originally isolated in coculture with insect cells in DCCM medium, strain LD-1T adapted to several media in the absence of cocultured cells. Use of anaerobic conditions allowed primary isolation in a variety of media. The organism did not grow in serum-free media containing 2% serum fraction. Optimal growth in M1D medium occurred at 30 to 37°C (doubling time, 7.2 h). On solid M1D medium containing 2.0% Noble agar (pH 6.25) at 30°C, strain LD-1T produced discrete colonies with numerous satellites. Strain LD-1T hydrolyzed arginine, but did not utilize urea; there was evidence of weak fermentation of glucose. The guanine-plus-cytosine content of the DNA was determined to be 25 ± 1 mol%, and the genome size was 1,085 kb. The results of extensive studies of the ecology of this spiroplasma suggest that it is host specific for Leptinotarsa beetles. Strain LD-1 (= ATCC 43213) is designated the type strain of a new species, Spiroplasma leptinotarsae.
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Amended Data on Arginine Utilization by Spiroplasma Species
More LessHydrolysis of arginine is a classical diagnostic test for species in the mollicute order Entomoplasmatales. In this paper we report data for arginine utilization by spiroplasmas, as determined by standard methods. In addition, modified methods were developed for fastidious spiroplasmas, such as strain LD-1T (T = type strain), the Colorado potato beetle spiroplasma. Twenty-one spiroplasma strains representing 13 groups or subgroups and eight ungrouped spiroplasmas (seven of which represent putative groups) were studied. The arginine reactions of eight strains were the same as the reactions reported previously, but previously reported positive tests for spiroplasma subgroups I-5 and I-6 (Spiroplasma insolitum) could not be repeated, and the data for the latter taxa are corrected. Although other workers have reported that addition of carbohydrate to media may be necessary for the utilization of arginine, the presence of glucose tended to obscure arginine hydrolysis in our studies.
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Helicobacter trogontum sp. nov., Isolated from the Rat Intestine
A new Helicobacter species that colonizes the colonic mucosa of Wistar and Holtzman rats was isolated and characterized. This bacterium was gram negative, its cells were rod shaped with pointed ends, and its protoplasmic cylinder was entwined with periplasmic fibers. It was catalase and oxidase positive, rapidly hydrolyzed urea, and was susceptible to metronidazole and resistant to cephalothin and nalidixic acid. The new organism was microaerophilic and grew at 42°C, a feature that differentiates it from two other murine intestine colonizers, Helicobacter hepaticus and Helicobacter muridarum. On the basis of 16S rRNA sequence analysis data, the new organism was identified as a Helicobacter species that is most closely related to H. hepaticus. This bacterium is named Helicobacter trogontum. The type strain is strain LRB 8581 (= ATCC 700114).
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High Degree of Similarity between Chromatium vinosum and Chromatium minutissimum as Revealed by Riboprinting
More LessThe riboprinting technique (restriction fragment length polymorphism [RFLP] analysis of PCR-amplified ribosomal DNA) was used to study five strains representing three species of the genus Chromatium. An RFLP analysis following digestion of the amplified small-subunit ribosomal DNA with 25 restriction enzymes revealed that the patterns obtained for all strains of Chromatium vinosum were identical. Chromatium gracile was different from C. vinosum with seven enzymes. On the other hand, Chromatium minutissimum produced the same patterns as C. vinosum with all enzymes, indicating that these organisms have a high degree of similarity. An RFLP analysis of the PCR-amplified spacer sequence between the 16S and 23S ribosomal DNAs gave similar results except that there was a larger number of differences between C. gracile and the other organisms examined.
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Reclassification of Flavobacterium odoratum (Stutzer 1929) Strains to a New Genus, Myroides, as Myroides odoratus comb. nov. and Myroides odoratimimus sp. nov.
More LessOn the basis of genotypic, chemotaxonomic, and phenotypic data, Flavobacterium odoratum was excluded from the emended genus Flavobacterium. In the present study, the known heterogeneity within this species was examined by a polyphasic approach that included DNA-rRNA hybridizations, a determination of DNA base ratios, DNA-DNA hybridizations, a numerical analysis of whole-cell protein patterns, a determination of cellular fatty acid compositions, and a phenotypic analysis. All of the methods revealed the presence of two distinct species. We propose that F. odoratum strains should be placed in a new genus, the genus Myroides, which comprises two species, Myroides odoratus comb. nov. and Myroides odoratimimus sp. nov.
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A Proposal To Transfer Microbispora bispora (Lechevalier 1965) to a New Genus, Thermobispora gen. nov., as Thermobispora bispora comb. nov.
More LessWe determined almost complete 16S rRNA gene sequences of two Microbispora bispora (Lechevalier 1965) strains, ATCC 19993T (T = type strain) and JCM 3082. The two sequences were 99% similar to each other but exhibited only 81 to 87.8% similarity with the 16S rRNA gene sequences of seven other Microbispora strains. A phylogenetic analysis revealed that the two sequences clustered not only distantly from other Microbispora strains, but also outside the cluster containing members of the family Streptosporangiaceae. On the basis of the results of our phylogenetic analysis and the results of a comprehensive review of the genus Microbispora by Miyadoh et al. (S. Miyadoh, S. Amano, H. Tohyama, and T. Shomura, J. Gen. Microbiol. 136:1905-1913, 1990) in which chemotaxonomic and DNA-DNA hybridization analyses were performed, we propose that Microbispora bispora should be transferred to a new genus, Thermobispora gen. nov., as Thermobispora bispora comb. nov.
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Proposal for Two New Genera, Brevibacillus gen. nov. and Aneurinibacillus gen. nov.
More Less16S rRNA gene sequences of the type strains of 11 species belonging to the Bacillus brevis and Bacillus aneurinolyticus groups were determined. On the basis of the results of gene sequence analyses, these species were separated into two clusters. The B. brevis cluster included 10 species, namely, Bacillus brevis, Bacillus agri, Bacillus centrosporus, Bacillus choshinensis, Bacillus parabrevis, Bacillus reuszeri, Bacillus formosus, Bacillus borstelensis, Bacillus laterosporus, and Bacillus thermoruber. Bacillus aneurinolyticus and Bacillus migulanus belonged to the B. aneurinolyticus cluster. Moreover, the two clusters were phylogenetically distinct from other Bacillus, Amphibacillus, Sporolactobacillus, Paenibacillus, and Alicyclobacillus species. On the basis of our data, we propose reclassification of the B. brevis cluster as Brevibacillus gen. nov. and reclassification of the B. aneurinolyticus cluster as Aneurinibacillus gen. nov. By using 16S rRNA gene sequence alignments, two specific PCR amplification primers were designed for differentiating the two new genera from each other and from other aerobic, endospore-forming organisms.
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Spiroplasma corruscae sp. nov., from a Firefly Beetle (Coleoptera: Lampyridae) and Tabanid Flies (Diptera: Tabanidae)
Spiroplasma strain EC-1T (T = type strain), which was isolated from the gut of a lampyrid beetle (Ellychnia corrusca) in Maryland, was serologically distinct from other spiroplasma species and groups. Similar strains were obtained from other E. corrusca specimens, and, later, numerous isolates of similar or partially related strains were obtained from several species of tabanid flies. Cells of strain EC-1T were helical, motile filaments that were bound by a single cytoplasmic membrane, and there was no evidence of a cell wall. The cells were filterable through 220-nm-pore-size membrane filters but not through 100-nm-pore-size membrane filters. The organism was absolutely resistant to penicillin (1,000 U/ml) and required sterol for growth. Strain EC-1T grew well in MID and SP-4 liquid media and could be cultivated in the Edward formulation of conventional mycoplasma medium and in 1% serum fraction medium. Optimal growth occurred at 32°C (doubling time, 1.5 h). Strain EC-1T multiplied at 10 to 41°C, but not at 5 or 43°C. This organism produced acid from glucose, but did not hydrolyze arginine or utilize urea. The guanine-plus-cytosine content of the DNA was determined to be 26.3 mol% by the melting temperature method and 27.0 mol% by the buoyant density method. As a result of our studies, strain EC-1 (= ATCC 43212) is designated the type strain of a new species, Spiroplasma corruscae.
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Actinobacillus minor sp. nov., Actinobacillus porcinus sp. nov., and Actinobacillus indolicus sp. nov., Three New V Factor-Dependent Species from the Respiratory Tract of Pigs
More LessThe results of DNA-DNA relatedness experiments and comparisons of sequences of genes coding for 16S rRNA were used to determine the genetic relationships of selected V factor-dependent species belonging to the family Pasteurellaceae and obtained from the porcine respiratory tract. These results showed that the Minor group and taxa C, D plus E, and F are distinct phylogenetic groups that are separate from each other and from other members of the family Pasteurellaceae. On the basis of these results, three new species, corresponding to the Minor group, taxa D plus E, and taxon F, are proposed; the names of these new species are Actinobacillus minor (type strain, NM305), Actinobacillus porcinus (type strain, NM319), and Actinobacillus indolicus (type strain, 46KC2), respectively.
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Phylogeny of Oral Asaccharolytic Eubacterium Species Determined by 16S Ribosomal DNA Sequence Comparison and Proposal of Eubacterium infirmum sp. nov. and Eubacterium tardum sp. nov.
More Less16S rRNA gene sequences of Eubacterium brachy, Eubacterium nodatum, Eubacterium saphenum, Eubacterium timidum, and two previously unnamed taxa were determined. The results of a phylogenetic analysis indicated that all of the strains sequenced belonged to a deep branch of the low-G + C-content gram-positive group. The levels of 16S ribosomal DNA sequence similarity between species were low, suggesting that a number of genera may be represented in this group. The representatives of the two unnamed taxa, which were isolated from patients with periodontitis, were clearly distinct from the previously described species, and, therefore, the following two new species are proposed: Eubacterium infirmum (type strain, NCTC 12940) and Eubacterium tardum (type strain, NCTC 12941).
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Phylogeny of Prosthecobacter, the Fusiform Caulobacters: Members of a Recently Discovered Division of the Bacteria
More LessProsthecobacter fusiformis is morphologically similar to caulobacters; however, it lacks a dimorphic life cycle. To determine the relatedness of the genus Prosthecobacter to dimorphic caulobacters and other prosthecate members of the α subgroup of the Proteobacteria (α-Proteobacteria), we isolated and sequenced 16S rRNA genes from four Prosthecobacter strains. Surprisingly, the results of phylogenetic analyses placed the fusiform caulobacters in a deeply rooted division of the Bacteria that was most closely affiliated with the Planctomyces-Chlamydia group and only distantly related to the α-Proteobacteria. The genus Prosthecobacter shares a common lineage in this division with Verrucomicrobium spinosum, a polyprosthecate, heterotrophic bacterium. Consistent with this phylogenetic placement, menaquinones were isolated from Prosthecobacter strains and menaquinones have been isolated from Verrucomicrobium strains and planctomycetes but not from members of the α-Proteobacteria. Thus, the genus Prosthecobacter is a second genus in the recently described order Verrucomicrobiales. Members of the genus Prosthecobacter are susceptible to β-lactam antibiotics and contain meso-diaminopimelic acid, indicating that they, unlike members of the Planctomycetales or Chlamydiales, have peptidoglycan cell walls. This major phenotypic difference, together with the phylogenetic independence of the verrucomicrobia, indicates that these bacteria and the sources of related 16S ribosomal DNAs obtained from soils, freshwater, and the marine pelagic environment represent an unrecognized division of the Bacteria.
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Leucobacter komagatae gen. nov., sp. nov., a New Aerobic Gram-Positive, Nonsporulating Rod with 2,4-Diaminobutyric Acid in the Cell Wall
More LessA new aerobic, gram-positive, nonsporulating rod-shaped organism is described. Strain IFO 15245T (T = type strain) has the following characteristics: The menaquinone contains a side chain with 11 isoprenyl units; the guanine-plus-cytosine content of the DNA is 66.2 moI%; 2,4-diaminobutyric acid, glutamic acid, alanine, glycine, and γ -aminobutyric acid are present in the cell wall at a molar ratio of ca. 1:1:2:1:1; and glucose and galactose are also present in the cell wall. A comparison of partial 16S rRNA sequences revealed that IFO 15245T represents a distinct line of descent within the gram-positive bacteria with high guanine-plus-cytosine contents. The taxonomic characteristics of this organism are different from those of previously described aerobic, gram-positive, nonsporulating, rod-shaped bacteria. The name Leucobacter komagatae gen. nov., sp. nov., is proposed for this organism. The type strain is strain IFO 15245.
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Sinorhizobium medicae sp. nov., Isolated from Annual Medicago spp.
More LessThe taxonomic position of isolates of a new genomic species (designated genomic species 2) obtained from several annual Medicago species and originating from different geographical locations was established through the results of phenotypic tests (including the of auxanographic and biochemical tests and symbiotic properties) and 16S rRNA phylogenetic inferences. A comparison of the complete 16S rRNA sequence of a representative of genomic species 2 (strain A 321T [T = type strain]) with the 16S rRNA sequences of other members of the Rhizobiaceae and closely related taxa showed that genomic species 2 was phylogenetically related to Sinorhizobium meliloti, Sinorhizobium fredii, Sinorhizobium saheli, and Sinorhizobium teranga. The levels of sequence similarity and observed numbers of nucleotide substitutions in Sinorhizobium strains indicated that A 321T and S. meliloti exhibited the highest level of sequence similarity (99.7%), with four nucleotide substitutions and one deletion. The results of a numerical analysis based on data from 63 auxanographic and biochemical tests clearly separated genomic species 2 isolates from S. meliloti. Genomic species 2 isolates nodulated and fixed nitrogen with Medicago polymorpha, whereas S. meliloti isolates were ineffective and formed rudimentary nodules on this host plant. On the basis of phenotypic and 16S sequence analysis data, genomic species 2 isolates cannot be assigned to a previously described species. We propose that these isolates belong to a new species, Sinorhizobium medicae.
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Oxidation of Thiosulfate by a New Bacterium, Bosea thiooxidans. (strain BI-42) gen. nov., sp. nov.: Analysis of Phylogeny Based on Chemotaxonomy and 16S Ribosomal DNA Sequencing
More LessA gram-negative bacterium which was capable of oxidizing reduced inorganic sulfur compounds was isolated from agricultural soil and designated BI-42. This new isolate grew on a wide range of organic substrates but was not able to grow autotrophically and lacked ribulose 1,5-bisphosphate carboxylase, a key enzyme of carbon dioxide fixation. These results suggested that strain BI-42 was a chemolithoheterotroph. Ammonia and nitrate were not used as sole nitrogen sources for growth, and strain BI-42 lacked glutamate synthase activity, which resulted in glutamate auxotrophy. The glutamate dehydrogenase activity of this organism was apparently insufficient for ammonia assimilation. On the basis of the results of additional biochemical tests, the G+C content of the DNA, the results of a respiratory ubiquinone analysis, the results of a 16S ribosomal DNA sequence analysis, the fatty acid composition, and the results of a membrane lipid analysis, strain BI-42 was identified as a phylogenetically and physiologically distinct taxon belonging to the alpha subclass of the Proteobacteria. Bosea thiooxidans gen. nov., sp. nov. is the name proposed for this taxon.
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A Polyphasic Reassessment of the Genus Paenibacillus, Reclassification of Bacillus lautus (Nakamura 1984) as Paenibacillus lautus comb. nov. and of Bacillus peoriae (Montefusco et al. 1993) as Paenibacillus peoriae comb. nov., and Emended Descriptions of P. lautus and of P. peoriae
Seventy-seven strains representing 10 species in the Paenibacillus polymyxa 16S rRNA group and 3 other species that exhibit phenetic relatedness to members of this group, Bacillus lautus, “Bacillus longisporus,” and Bacillus peoriae, were characterized genotypically and phenotypically by performing an amplified ribosomal DNA restriction analysis, a randomly amplified polymorphic DNA analysis, a fatty acid methyl ester analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole-cell proteins, pyrolysis mass spectrometry, and API and other routine phenotypic tests. These analyses revealed distinct clusters representing Paenibacillus alvei, Paenibacillus amylolyticus, Paenibacillus azotofixans, Paenibacillus durum, Paenibacillus larvae subsp. larvae. Paenibacillus larvae subsp. pulvifaciens, B. lautus, Paenibacillus macerans, Paenibacillus macquariensis, B. peoriae, P. polymyxa, and Paenibacillus validus, which confirmed the distinctness of these species, but appreciable within-species heterogeneity was observed in P. alvei, B. lautus, P. macerans, P. polymyxa, and P. validus. The type strain of Paenibacillus pabuli did not cluster with other strains of this species, and in several analyses a relationship between strains of P. pabuli and “B. longisporus” was observed. As the analyses showed that B. lautus and B. peoriae are closely related to the genus Paenibacillus, these species are reclassified as members of this genus.
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Analysis of the β’ Subunit of DNA-Dependent RNA Polymerase Does Not Support the Hypothesis Inferred from 16S rRNA Analysis that Oenococcus oeni (Formerly Leuconostoc oenos) Is a Tachytelic (Fast-Evolving) Bacterium
More LessrRNA sequencing has shown that Leuconostocs comprise three distinct phylogenetic lineages which have been designated separate genera (viz., the genera Leuconostoc sensu stricto, Oenococcus, and Weissella). In addition, the 16S rRNA line formed by Oenococcus oeni (formerly Leuconostoc oenos) is exceptionally long: This fact, together with variations in the compositions of conserved positions in the 16S rRNA, has led to the hypothesis (D. Yang and C. R. Woese, Syst. Appl. Microbiol. 12:145-149, 1989) that this organism is a fast-evolving bacterium. Previous evidence that the leuconostocs should be divided into three genera and that O. oeni is an example of tachytelic evolution has come solely from rRNA analyses. In this study we sequenced the rpoC gene encoding the β’ subunit of DNA-dependent RNA polymerase of leuconostocs and performed a comparative phylogenetic analysis. The subdivision of the leuconostocs into three distinct lineages was confirmed by the rpoC gene data, but no evidence that indicated that O. oeni is evolving at an extraordinary rate was found. If O. oeni is truly tachytelic, then fast-evolving phenomena would be expected to occur throughout the whole genome, including this independent molecular chronometer.
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Isolation and Characterization of Desulfitobacterium frappieri sp. nov., an Anaerobic Bacterium Which Reductively Dechlorinates Pentachlorophenol to 3-Chlorophenol
More LessAn anaerobic bacterium, strain PCP-1T (T = type strain), which dechlorinates pentachlorophenol (PCP) to 3-chlorophenol, was isolated from a methanogenic consortium. This organism is a spore-forming rod-shaped bacterium that is nonmotile, asaccharolytic, and Gram stain negative but Gram type positive as determined by electron microscopic observations. Inorganic electron acceptors, such as sulfite, thiosulfate, and nitrate (but not sulfate), stimulate growth in the presence of pyruvate and yeast extract. The optimum pH and optimum temperature for growth are 7.5 and 38°C, respectively. The dechlorination pathway is: PCP → 2,3,4,5-tetra-chlorophenol → 3,4,5-trichlorophenol → 3,5-dichlorophenol → 3-chlorophenol. This bacterium dechlorinates several different chlorophenols at ortho, meta, and para positions; exceptions to this are 2,3-dichlorophenol, 2,5-dichlorophenol, 3,4-dichlorophenol, and the monochlorophenols. The time course of PCP dechlorination suggests that two enzyme systems are involved in dehalogenation in strain PCP-1T. One system is inducible for ortho dechlorination, and the second system is inducible for meta and para dechlorinations. A 16S rRNA analysis revealed that strain PCP-1T exhibits 95% homology with Desulfitobacterium dehalogenans JW/IU-DC1, an anaerobic bacterium which can dehalogenate chlorophenols only in ortho positions. These results suggest that strain PCP-1T is a member of a new species and belongs to the recently proposed genus Desulfitobacterium. Strain PCP-1T differs from D. dehalogenans JW/IU-DC1 by its broader range of chlorophenol dechlorination. Strain PCP-1 is the type strain of the new species, Desulfitobacterium frappieri.
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Classification of Bacteria Nodulating Lathyrus japonicus and Lathyrus pratensis in Northern Quebec as Strains of Rhizobium leguminosarum biovar viciae †
More LessThe diversity of two populations of rhizobia isolated from Lathyrus japonicus (30 strains) and Lathyrus pratensis (49 strains) growing in northern regions of Quebec, Canada, was determined on the basis of phenotypic characteristics, multilocus enzyme electrophoresis, DNA-DNA homology, and 16S ribosomal DNA sequencing. According to numerical analysis of phenotypic characteristics, strains were divided into four groups. Strains isolated from L. pratensis fell in groups I to III; the latter included reference strains of Rhizobium leguminosarum. All strains isolated from L. japonicus were included in group IV. All strains had nodulation characteristics similar to those of R. leguminosarum bv. viciae. Strains isolated from L. japonicus originating from an arctic region were usually able to grow at 5°C and were more likely to be tolerant to copper (CuCI2 · H2O, 100 μg/ml) and lead [Pb(CH3COO)2, 500 μg/ml] than strains isolated from L. pratensis from a boreal zone. However, both populations of Lathyrus strains were adapted to the cold in comparison to reference strains from temperate regions. Each population had similar genetic diversity (H = 0.45), determined by multilocus enzyme electrophoresis of the loci encoding eight enzymes, but the diversity obtained by analyzing all strains including the reference strains (H = 0.58) was higher. Representative strains of both populations showed high levels of DNA homology among themselves and with R. leguminosarum. Partial sequences of the 16S ribosomal RNA genes were similar to those reported for R. leguminosarum bv. viciae. We conclude that the strains isolated from L. japonicus and L. pratensis belong to R. leguminosarum bv. viciae but are distinguishable by growth at 5°C, which is a characteristic related to their geographic origin.
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Thermobrachium celere gen. nov., sp. nov., a Rapidly Growing Thermophilic, Alkalitolerant, and Proteolytic Obligate Anaerobe
More than 40 isolates of a novel, ubiquitous, proteolytic, moderately alkaliphilic, thermophilic obligate anaerobe were obtained from geothermally and anthropogenically heated environments and mesobiotic environments located on three continents. Whole-cell protein sodium dodecyl sulfate gel electrophoresis revealed that most of these organisms are very similar. Eight of the isolates were characterized in detail; this analysis included 16S ribosomal DNA sequence analysis. The cells of those organisms are (depending on the isolate) 0.5 to 0.8 μm in diameter and 1.5 to 13 μm long, exhibit tumbling motility, and have a positive Gram stain reaction. The temperature range for growth is 43° to 75°C (optimum temperature, 66°C), and the pH range for growth is 5.4 to 9.5 (optimum pH, 8.2); the shortest doubling time is around 10 min. Yeast extract is required for growth, and (depending on the strain) glucose, sucrose, fructose, galactose, and ribose are utilized. The fermentation products from glucose in the presence of yeast extract are CO2, H2, acetate, formate, and ethanol. The G+C content is 30 to 31 mol%. On the basis of these properties, which differentiate these strains from all alkalitolerant thermophiles described previously, and the results of a comparison of the 16S ribosomal DNA sequences of these organisms with previously described sequences, we propose that our isolates be placed in a single species of the new genus Thermobrachium; strain JW/YL-NZ35 is the type strain of the the type species, Thermobrachium celere.
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Phenotypic and DNA Relatedness between Nematode Symbionts and Clinical Strains of the Genus Photorhabdus (Enterobacteriaceae) †
More LessBacterial strains isolated from wide ranges of nematode hosts and geographic sources and strains isolated from human clinical specimens were used to assess the taxonomic structure of the genus Photorhabdus. The following two methods were used: DNA relatedness and phenotypic characterization. Analysis of the DNA relatedness data revealed that all of the strains studied were congeneric and that the genus Photorhabdus is, on the basis of DNA relatedness data, more homogeneous than the other genus of nematode-symbiotic bacteria, the genus Xenorhabdus. In contrast to previous reports, only two DNA relatedness groups were identified in the genus Photorhabdus. These groups corresponded to the symbiotic strains and the clinical strains. There appeared to be some subgroups within the symbiotic strain group on the basis of the interactions of the strains with nematodes, which corresponded to some extent with the DNA relatedness data. However, there were significant ambiguities in the DNA relatedness data, and this group could not be subdivided on the basis of DNA relatedness data or phenotypic data. The distinct functional differences within and between the DNA relatedness groups of symbiotic Photorhabdus strains indicated that there are biologically significant subgroups within the genus Photorhabdus that cannot be defined at this time. Further investigation of the taxonomy of Photorhabdus by using different approaches and a suitably wide range of strains is recommended. However, it is clear that the clinical strains form a recognizable subgroup within the genus even though no formal subtaxon can be defined at this time.
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Description of Chlorophenol-Degrading Pseudomonas sp. Strains KF1T, KF3, and NKF1 as a New Species of the Genus Sphingomonas, Sphingomonas subarctica sp. nov.
Gram-negative polychlorophenol-degrading bacterial strains KF1T (T = type strain), KF3, and NKF1, which were described previously as Pseudomonas saccharophila strains, were studied by chemotaxonomic, genetic, and physiological methods and by electron microscopy and compared with selected xenobiotic compound-degrading bacteria. These strains contained sphingolipids with d-18:0, d-20:1, and d-21:1 as the main dihydrosphingosines, ubiquinone 10 as the main respiratory quinone, and spermidine as the major polyamine, and the DNA G+C content was 66 mol%. The cellular fatty acids included about 60% octadecenoic acid, 9% 2-hydroxymy-ristic acid, 14% cis-9-hexadecenoic acid, and 10% hexadecanoic acid. These strains exhibited less than 97% 16S ribosomal DNA sequence similarity to all of the other taxa studied. In the DNA-DNA reassociation studies the highest levels of reassociation between these strains and previously described species were less than 40%. Thin sections of cells of strains KF1T, KF3, and NKF1 were examined by electron microscopy, and the results showed that the cells had peculiar concentrically arranged layered membranous blebs that extruded from the outer membrane, especially at the cell division points. On the basis of the results of this study, polychlorophenol-degrading strains KF1T, KF3, and NKF1 are considered members of a new species of the genus Sphingomonas, Sphingomonas subarctica. The polycyclic aromatic hydrocarbon-degrading organism Sphingomonas paucimobilis EPA 505 was closely related to Sphingomonas chlorophenolica as determined by chemotaxonomic, phylogenetic, and physiological criteria. The xenobiotic compound degraders Alcaligenes sp. strain A175 and Pseudomonas sp. strain BN6 were identified as members of species of the genus Sphingomonas.
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Sulfobacillus disulfidooxidans sp. nov., a New Acidophilic, Disulfide-Oxidizing, Gram-Positive, Spore-Forming Bacterium
More LessAn acidophilic, disulfide-oxidizing, mesophilic, aerobic bacterium was isolated from wastewater sludge. The new organism is a gram-positive sporulated rod. It can use elemental sulfur and pyrite as sole energy sources and grows on organic substrates such as glutamate and glucose. It also grows on the following organic sulfur substrates: Oxidized and reduced glutathione, cysteine, cystine, and dithio(bis)benzothiazole and clearly shows a preference for disulfide bond-containing substrates. The optimal pH of growth is between 1.5 and 2.5, depending on the substrate used, and the growth temperature range varies from 4 to 40°C, with an optimal value at 35°C. The G+C chromosomal DNA content was measured at 53 ± 1 mol%. Phylogenetic analysis of 16S genes coding for rRNA sequences places the new isolate in the genus Sulfobacillus. In addition, unique phenotypic and physiologic characteristics and DNA homology values assign the isolate to a new species in the genus. Therefore, this new isolate has been named Sulfobacillus disulfidooxidans and has been assigned ATCC number 51911.
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Phylogenetic Positions of Desulfofustis glycolicus gen. nov., sp. nov. and Syntrophobotulus glycolicus gen. nov., sp. nov., Two New Strict Anaerobes Growing with Glycolic Acid
More LessThe glycolate-oxidizing, sulfate-reducing bacterium strain PerGlyS and the syntrophically glycolate-oxidizing bacterium strain FIGIyR were studied with respect to their phylogenetic relationships on the basis of in vitro amplification and direct sequencing of 16S rRNA-encoding DNA. Strain PerGlyS clustered with representatives of the δ subclass of the class Proteobacteria, close to “Desulforhopalus vacuolatus” but sufficiently distinct to preclude its assignment to this genus. These organisms, together with Desulfobulbus propionicus, represent a phylogenetic subgroup among members of the δ subclass of Proteobacteria. Strain FIGIyR was found to cluster with the gram-positive bacteria with low-G+C DNA, and Desulfitobacterium dehalogenans and Desulfotomaculum orientis are its closest relatives. Other species of the genus Desulfotomaculum are phyloge-netically only moderately closely related to these organisms. These results necessitate the establishment of new genera and species for these two strains. Strain PerGlyS was designated the type strain of Desulfofustis glycolicus gen. nov., sp. nov., and strain FIGIyR was designated the type strain of Syntrophobotulus glycolicus gen. nov., sp. nov.
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Aeropyrum pernix gen. nov., sp. nov., a Novel Aerobic Hyperthermophilic Archaeon Growing at Temperatures up to 100°C
A novel aerobic hyperthermophilic archaeon was isolated from a coastal solfataric vent at Kodakara-Jima Island, Japan. The new isolate, strain K1, is the first strictly aerobic organism growing at temperatures up to 100°C. It grows optimally at 90 to 95°C, pH 7.0, and a salinity of 3.5%. The cells are spherical shaped and 0.8 to 1.2 μm in diameter. Various proteinaceous complex compounds served as substrates during aerobic growth. Thiosulfate stimulates growth without producing H2S. The core lipids consist solely of C25-isopranyl archaeol(glycerol diether). The G+C content of the genomic DNA is 67 mol%. Phylogenetic analysis based on 16S rRNA sequence indicates that strain K1 is a new member of Crenarchaeota. On the basis of our results, the name Aeropyrum pernix gen. nov., sp. nov. is proposed (type strain: K1; JCM 9820).
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Reassessment of the Phylogenetic Position of the Bacterium Associated with Whipple’s Disease and Determination of the 16S-23S Ribosomal Intergenic Spacer Sequence
More LessWhipple’s disease is a rare chronic illness associated with an unculturable bacterium that is constantly present in affected tissues. This bacterium was previously characterized at the molecular level by PCR and sequencing of the 16S rRNA gene. On the basis of 1,321 nucleotides of the sequence of its gene coding for 16S rRNA (16S rDNA), a phylogenetic relationship to the actinomycetes was established. In this study, we determined an almost complete 16S rDNA sequence (1,495 nucleotides), the 16S-23S ribosomal intergenic spacer sequence, and 200 nucleotides of the 23S rRNA gene. The 16S rDNA sequence was compared with the large number of actinomycete sequences that have been added to the database since the original study. Phylogenetic analysis revealed a branching position as the deepest branch of the cluster comprising the actinomycetes with group B peptidoglycan between this group and the family Cellulomonadaceae. This provides additional information on the phylogenetic position of this bacterium and some clues as to its characteristics. The spacer region between the 16S and 23S rRNA genes is 294 nucleotides long and does not contain tRNA genes. As has been shown in other instances, the increased variability of the ribosomal intergenic spacer compared with the 108 rRNA gene makes it a potential target for use in the differentiation of strains of the bacterium associated with Whipple’s disease.
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Phylogenetic Relationships of the Genera Acetobacterium and Eubacterium Sensu Stricto and Reclassification of Eubacterium alactolyticum as Pseudoramibacter alactolyticus gen. nov., comb. nov.
More Less16S rRNA gene sequences of the type strains of the seven previously described Acetobacterium species were determined. The Acetobacterium species were found to form a tight phylogenetic cluster within the Clostridium subphylum of the gram-positive bacteria. Within this subphylum these organisms belong to cluster XV as defined by Collins et al. (M. D. Collins, P. A. Lawson, A. Willems, J. J. Cordoba, J. Fernandez-Garayzabal, P. Garcia, J. Cai, H. Hippe, and J. A. E. Farrow, Int. J. Syst. Bacteriol. 44:812-826, 1994) together with Eubacterium alactolyticum, Eubacterium barkeri, Eubacterium callanderi, and Eubacterium limosum. Our data indicate that Clostridium cluster XV consists of at least the following three genera: The genus Acetobacterium, the genus Eubacterium sensu stricto (comprising E. limosum, E. barkeri, and E. callanderi), and the genus Pseudoramibacter gen. nov., which is created for E. alactolyticum, which we reclassify as Pseudoramibacter alactolyticus comb. nov.
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The Genus Nocardiopsis Represents a Phylogenetically Coherent Taxon and a Distinct Actinomycete Lineage: Proposal of Nocardiopsaceae fam. nov.
More LessThe genus Nocardiopsis was shown to be phylogenetically coherent and to represent a distinct lineage within the radiation of the order Actinomycetales. The closest relatives of the genus Nocardiopsis are members of the genera Actinomadura, Thermomonospora, Streptosporangium, and Microtetraspora. The intrageneric structure of the genus Nocardiopsis is shown to consist of a highly related species group containing Nocardiopsis dassonvillei, Nocardiopsis alborubida, and Nocardiopsis antarctica and a second group of less highly related species comprising Nocardiopsis alba subsp. alba, Nocardiopsis alba subsp. prasina, and Nocardiopsis listeri. Nocardiopsis lucentensis occupies a position intermediate between the two species groups. The results of a 16S ribosomal DNA sequence analysis are generally consistent with the available chemotaxonomic, phenotypic, and DNA-DNA hybridization data. The phylogenetic position and the morpho- and chemotaxonomic properties of Nocardiopsis species support the creation of a family for the genus Nocardiopsis, Nocardiopsaceae fam. nov.
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Phylogeny of Some Mycoplasmas from Ruminants Based on 16S rRNA Sequences and Definition of a New Cluster within the Hominis Group
More LessAlmost complete (>96%) 16S rRNA sequences from nine ruminant mycoplasmas have been determined by solid-phase DNA sequencing. Polymorphisms were found in four of the 16S rRNA sequences, which indicated the existence of two different rRNA operons. Seven polymorphisms were found in Mycoplasma agalactiae, three were found in Mycoplasma bovis, one was found in Mycoplasma alkalescens, and one was found in Mycoplasma bovirhinis. The sequence data were used for construction of phylogenetic trees. All but one of the ruminant mycoplasmas sequenced in this work clustered in the hominis group. A close relationship was found between M. agalactiae and M. bovis, with a 99% nucleotide similarity between their 16S rRNA sequences. They were also found to be members of the Mycoplasma lipophilum cluster of the hominis group. Furthermore, the 16S rRNA comparisons showed that Mycoplasma alkalescens and Mycoplasma canadense are closely related (>98.5%), and these species were found to cluster in the Mycoplasma hominis cluster of the hominis group. Interestingly, M. bovirhinis grouped in a new phylogenetic cluster of the hominis group. The new cluster, which was supported by bootstrap percentage values, signature nucleotide analysis, and higher-order structural elements, was named the Mycoplasma synoviae cluster. Mycoplasma bovoculi, Mycoplasma conjunctivae, and Mycoplasma ovipneumoniae clustered in the Mycoplasma neurolyticum cluster of the hominis group. Mycoplasma alvi clustered with Mycoplasma pirum in the M. pneumoniae cluster of the pneumoniae group.
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Rhodothermus obamensis sp. nov., a Modern Lineage of Extremely Thermophilic Marine Bacteria
More LessA novel extremely thermophilic bacterium was isolated from a shallow marine hydrothermal vent environment (depth, 22 m) in Tachibana Bay, Nagasaki Prefecture, Japan. The cells of this organism were gramnegative rods. Growth occurred at temperatures between 50 and 85°C (optimum temperature, 80°C; doubling time at optimum temperature, 90 min), at pH 5.5 and 9.0 (optimum pH, 7.0), and in the presence of 1 and 5% NaCl (optimum NaCl concentration, 3%). The new isolate was an aerobic heterotroph which utilized the following compounds as sole energy and carbon sources: Yeast extract, peptone, starch, casein, Casamino Acids, a variety of sugars, some carboxylic acids, and amino acids. As determined by a sequence analysis of the 16S rRNA, the new isolate belongs to the genus Rhodothermus and represents a modern lineage of extreme thermophiles within the domain Bacteria. On the basis of the physiological and molecular properties of the new isolate, we describe a new species, Rhodothermus obamensis. The type strain of R. obamensis is strain OKD7 (= JCM 9785).
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Organization and Phylogenetic Interrelationships of Genes Encoding Components of the Botulinum Toxin Complex in Proteolytic Clostridium botulinum Types A, B, and F: Evidence of Chimeric Sequences in the Gene Encoding the Nontoxic Nonhemagglutinin Component
More LessThe cluster of genes encoding components of the botulinum neurotoxin (BoNT) complex was mapped in proteolytic (group I) Clostridium botulinum strains encoding BoNT types A, B, and F. Two different arrangements of genes were found: Type A strain 62A and type B strain NCTC 7273 have similar organizations of genes encoding BoNT, the nontoxic nonhemagglutinin component (NTNH), hemagglutinin components, and P-21; type F strain Langeland has genes encoding BoNT, NTNH, and P-21, and a previously unidentified open reading frame encoding a protein of 416 amino acids. A group of type A strains typified by infant strain Kyoto-F, which is unlike type A strain 62A, lacks genes for hemagglutinin components and exhibits an organization similar to that of type F. Sequencing and pairwise analysis revealed the presence of possible chimeric sequences in some NTNH genes of proteolytic C. botulinum. Discordance in genealogical trees derived from different regions of the NTNH genes was observed which could be symptomatic of recombination and which may indicate that the NTNH gene represents a hot spot for such events within the cluster of genes encoding the BoNT complex. It is also evident that the phylogenetics of the NTNH gene, which is linked to the gene encoding BoNT, does not mirror the evolutionary history of the BoNT, upon which the C. botulinum species complex is defined and subdivided.
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Thermococcus fumicolans sp. nov., a New Hyperthermophilic Archaeon Isolated from a Deep-Sea Hydrothermal Vent in the North Fiji Basin
An extremely thermophilic archaeon, strain ST557T (T = type strain), was isolated from a deep-sea hydrothermal vent in the North Fiji Basin. This strain is a strictly anaerobic coccus whose cells are about 0.8 to 2 μm in diameter. The optimum temperature, pH and sea salt concentration for growth are 85°C, 8.5, and 20 to 40 g/liter, respectively. Strain ST557T grows preferentially in the presence of elemental sulfur on proteinaceous substrates and on a mixture of 20 amino acids. It grows slowly on pyruvate and maltose. Growth is inhibited by rifampin. The DNA G+C content is 54 to 55 mol%. Sequencing of the 16S rRNA gene revealed that strain ST557T belongs to the genus Thermococcus. We propose that this organism should be placed in a new species, Thermococcus fumicolans.
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Eubacterium exiguum sp. nov., Isolated from Human Oral Lesions
More LessEubacterium exiguum sp. nov. is the name proposed for organisms formerly described as Eubacterium group S strains and similar bacteria isolated from various types of oral lesions. This new species was established on the basis of the results of DNA-DNA hybridization experiments and DNA base composition determinations (G+C contents, 60 to 64 mol%). The results of an API ZYM analysis, Western blotting (immunoblotting) reactions, and phenotypic tests are also given. The type strain of E. exiguum is strain S-7.
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Isolation and Characterization of a New Gram-Negative, Acetone-Degrading, Nitrate-Reducing Bacterium from Soil, Paracoccus solventivorans sp. nov.
More LessAn acetone-degrading, nitrate-reducing, coccoid to rod-shaped bacterium, strain L1, was isolated from soil on the site of a natural gas company. Cells of the logarithmic growth phase reacted gram positive, while those of the stationary growth phase were gram negative. Single organisms were 0.4 to 0.5 by 0.9 to 1.5 μm in size, nonmotile, and non-spore forming and had poly-β-hydroxybutyrate inclusions. The doubling time of strain L1 on acetone-CO2-nitrate at the optimal pH of 7 to 8 and the optimal temperature of 30 to 37°C was 12 h. More than 0.2% NaCl or 10 mM thiosulfate inhibited growth. For oxygen or nitrate respiration, acetone and a few organic chemicals were utilized as carbon sources whereas many others could not be used (for details, see Results). Bicarbonate (or CO2) was essential for growth on acetone but not for growth on acetoacetate. The growth yields for acetone-CO2 and acetoacetate were 28.3 and 27.3 g/mol, respectively. With acetone as the carbon source, poly-β-hydroxybutyrate accounted for up to 40% of the cellular dry weight The DNA of strain L1 had a G+C content of 68.5 mol% (as determined by high-performance liquid chromatography of nucleotides) or 70 mol% (as determined by the Tm method). The sequence of the gene coding for the 16S rRNA led to the classification of strain L1 in the paracoccus group of the alpha subclass of the Proteobacteria. The new isolate is named Paracoccus solventivorans sp. nov. DSM 6637.
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Thermosyntropha lipolytica gen. nov., sp. nov., a Lipolytic, Anaerobic, Alkalitolerant, Thermophilic Bacterium Utilizing Short- and Long-Chain Fatty Acids in Syntrophic Coculture with a Methanogenic Archaeum
More LessThree strains of an anaerobic thermophilic organoheterotrophic lipolytic alkalitolerant bacterium, Thermosyntropha lipolytica gen. nov., sp. nov. (type strain JW/VS-265T; DSM 11003), were isolated from alkaline hot springs of Lake Bogoria (Kenya). The cells were nonmotile, non-spore forming, straight or slightly curved rods. At 60°C the pH range for growth determined at 25°C [pH25°C] was 7.15 to 9.5, with an optimum between 8.1 and 8.9 (pH60°C of 7.6 and 8.1). At a pH25°C of 8.5 the temperature range for growth was from 52 to 70°C, with an optimum between 60 and 66°C. The shortest doubling time was around 1 h. In pure culture the bacterium grew in a mineral base medium supplemented with yeast extract, tryptone, Casamino Acids, betaine, and crotonate as carbon sources, producing acetate as a major product and constitutively a lipase. During growth in the presence of olive oil, free long-chain fatty acids were accumulated in the medium but the pure culture could not utilize olive oil, triacylglycerols, short- and long-chain fatty acids, and glycerol for growth. In syntrophic coculture (Methanobacterium strain JW/VS-M29) the lipolytic bacteria grew on triacylglycerols and linear saturated and unsaturated fatty acids with 4 to 18 carbon atoms, but glycerol was not utilized. Fatty acids with even numbers of carbon atoms were degraded to acetate and methane, while from odd-numbered fatty acids 1 mol of propionate per mol of fatty acid was additionally formed. 16S rDNA sequence analysis identified Syntrophospora and Syntrophomonas spp. as closest phylogenetic neighbors.
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DNA Relatedness among Pseudomonas Strains Isolated from Natural Mineral Waters and Proposal of Pseudomonas veronii sp. nov.
More LessThe taxonomic position of eight strains isolated from mineral water and previously grouped in the authentic pseudomonads on the basis of a phenotypic analysis (cluster Ib of M. Elomari, L. Coroler, D. Izard, and H. Leclerc [J. Appl. Bacteriol. 78:71-81, 1995]) has been further studied by DNA-DNA hybridizations. Using the S1 nuclease method at 60°C and labeled reference DNA from a representative strain, CFML 92-134, we showed that members of cluster Ib constituted a homogeneous group with a relative binding ratio of greater than 80% and changes in melting temperature of less than 1°C. With a total of 67 strains representing known or partially characterized species of the genus Pseudomonas, only 4 to 47% DNA hybridization and changes in melting temperature of between 8 and 20°C were found, the highest hybridization values being measured with members of the saprophytic fluorescent pseudomonads. Since cluster Ib could also be clearly differentiated from members of the latter group and from other phenotypic clusters containing isolates from mineral water, we designated the Ib strains members of a new Pseudomonas species for which the name Pseudomonas veronii sp. nov. has been proposed. Members of this species grew on α-aminobutyrate, sucrose, butyrate, isobutyrate, erythritol, l-tryptophan, and trigonelline as sole sources of carbon and energy. The average G+C content of the DNA of the eight strains of P. veronii was 61.5 ± 0.5 mol%. The type strain is CFML 92-134T (CIP 104663T), with a G+C content of 61 mol%. The clinical significance of P. veronii is unknown.
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Clostridium ultunense sp. nov., a Mesophilic Bacterium Oxidizing Acetate in Syntrophic Association with a Hydrogenotrophic Methanogenic Bacterium
More LessA syntrophic acetate-oxidizing bacterium, strain BST (T = type strain), was isolated from a previously described mesophilic triculture that was able to syntrophically oxidize acetate and form methane in stoichiometric amounts. Strain BST was isolated with substrates typically utilized by homoacetogenic bacteria. Strain BST was a spore-forming, gram-positive, rod-shaped organism which utilized formate, glucose, ethylene glycol, cysteine, betaine, and pyruvate. Acetate and sometimes formate were the main fermentation products. Small amounts of alanine were also produced from glucose, betaine, and cysteine. Strain BST grew optimally at 37°C and pH 7. The G+C content of the DNA of strain BST was 32 mol%. A 16S rRNA sequence analysis revealed that strain BST was a member of a new species of the genus Clostridium. We propose the name Clostridium ultunense for this organism; strain BS is the type strain of C. ultunense.
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Chrysiogenes arsenatis gen. nov., sp. nov., a New Arsenate-Respiring Bacterium Isolated from Gold Mine Wastewater
A new strictly anaerobic bacterium (strain BAL-1T) has been isolated from a reed bed at Ballarat Goldfields in Australia. The organism grew by reducing arsenate [As(V)] to arsenite [As(III)], using acetate as electron donor and carbon source; acetate alone did not support growth. When BAL-1T was grown with arsenate as the terminal electron acceptor, acetate could be replaced by pyruvate, l- and d-lactate, succinate, malate, and fumarate but not by H2, formate, citrate, glutamate, other amino acids, sugars, or benzoate. With acetate was the electron donor, arsenate could be replaced by nitrate or nitrite but not by sulfate, thiosulfate, or iron oxide. Nitrate was reduced to ammonia via nitrite. The doubling time for growth on acetate (5 mM) plus arsenate (5 mM) or nitrate (5 mM) was 4 h. The G+C content of the DNA is 49 mol%. The 16S rRNA sequence data for the organism support the hypothesis that this organism is phylogenetically unique and at present is the first representative of a new deeply branching lineage of the Bacteria. This organism is described as Chrysiogenes arsenatis gen. nov., sp. nov.
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Lactobacillus curvatus subsp. curvatus subsp. nov. and Lactobacillus curvatus subsp. melibiosus subsp. nov. and Lactobacillus sake subsp. sake subsp. nov. and Lactobacillus sake subsp. carnosus subsp. nov., New Subspecies of Lactobacillus curvatus Abo-Elnaga and Kandler 1965 and Lactobacillus sake Katagiri, Kitahara, and Fukami 1934 (Klein et al. 1996, Emended Descriptions), Respectively
More LessLactobacillus curvatus and Lactobacillus sake are each genetically homogeneous species, as indicated by the high levels of DNA homology (≥76%) exhibited by strains of these taxa. However, the results of a numerical analysis of total soluble cell protein patterns and biochemical test data revealed that there are two phenotypic subgroups within L. curvatus and two phenotypic subgroups within L. sake. The overall randomly amplified polymorphic DNA (RAPD)-PCR band patterns obtained for the majority of L. curvatus strains corresponded well to the pattern obtained for the type strain of L. curvatus (strain DSM 20019). However, six strains of L. curvatus had different, but similar, RAPD-PCR profiles and grouped in a separate genetic cluster, which was linked to one of the clusters of L. sake strains. On the basis of these results, differences in biochemical and physiological characteristics, and total soluble cell protein profiles, we describe the subspecies L. curvatus subsp. curvatus subsp. nov. and L. curvatus subsp. melibiosus subsp. nov. for L. curvatus Abo-Elnaga and Kandler 1965 (Klein et al. 1996, emended description). Strains of L. sake grouped in two RAPD-PCR clusters, which was consistent with previous reports of phenotypic heterogeneity. Strains of Lactobacillus bavaricus, including type strain LMG 9844, clustered with the type strain of L. sake (strain NCFB 2714), indicating that these organisms belong to the same genetic group. We propose that strains of L. sake Katagiri, Kitahara, and Fukami 1934 (Klein et al. 1996, emended description) should be reclassified as members of L. sake subsp. sake subsp. nov. and L. sake subsp. carnosus subsp. nov. Strains of L. bavaricus are reclassified as members of L. sake subsp. sake, and the name L. bavaricus Stetter and Stetter 1980 is rejected.
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Determination of Mycobacterial Phylogeny on the Basis of Immunological Relatedness of Superoxide Dismutases
Sixteen strains of cultivable mycobacteria were grown in Sauton’s medium, and Mycobacterium leprae was purified from armadillo liver. Cell extracts were prepared from log-phase growths of each of the cultivable mycobacterial strains. Superoxide dismutase (SOD) enzyme was purified from all cultivable mycobacterial strains included in the study, and antibodies against purified SOD enzyme were raised in rabbits. Immunological distances (ImDs) between these anti-SOD antibodies and SOD antigens were determined by a previously described immunoprecipitation method and by a recently developed enzyme-linked immunosorbent assay (ELISA) technique. The reciprocal ImDs among mycobacterial strains were constant, reproducible and consistent by these two methods. An evolutionary tree was constructed on the basis of estimated ImDs. Except for M. duvalii and M. terrae, slowly and rapidly growing mycobacterial species appeared to be separately grouped by this analysis. Rapid growers clustered into a group which is near that of some slow-growing mycobacteria. M. avium falls almost in the middle of the evolutionary tree and the position of M. leprae was found to be between those of M. avium and M. bovis BCG. Measurement of immunological relatedness of SODs provides an alternative system with which to study the taxonomical relatedness among mycobacteria.
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NOTES: Phylogeny of Methanopyrus kandleri Based on Methyl Coenzyme M Reductase Operons
The mcrBDCGA operon that encodes methyl coenzyme M reductase (MR) in the hyperthermophile Methanopyrus kandleri was cloned and sequenced. The results of a phylogenetic analysis of the nine MR sequences now available support the position that M. kandleri is a separate methanogen lineage. As in other methanogens, the M. kandleri mcr operon is located immediately upstream of the mtrE gene, the promoter-proximal gene in an operon that encodes the N 5-methyltetrahydromethanopterin:coenzyme M methyltransferase that catalyzes the step preceding the MR-catalyzed reaction in methanogenesis. In contrast to other methanogens and hyperthermophilic members of the Archaea. CG dinucleotides and CG-containing codons occur frequently in M. kandleri DNA. The MR subunit-encoding genes are preceded by sequences consistent with ribosome binding sites, indicating that mRNA-rRNA base pairing can still direct translation initiation in cells growing at temperatures above 100°C.
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