- Volume 50, Issue 6, 2000

In 1994, the uncultured phloem-restricted bacteria of citrus huanglongbing (ex-greening) disease in Asia and Africa were characterized as 'Candidatus Liberobacter asiaticum' and 'Candidatus Liberobacter africanum', respectively. Following the rules of the International Code of Nomenclature of Bacteria, the two bacterial species have now been renamed 'Candidatus Liberibacter asiaticus' and 'Candidatus Liberibacter africanus'. A third liberibacter was detected by PCR in an ornamental rutaceous tree, Cape chestnut (Calodendrum capense), in South Africa. The new liberibacter was characterized by serology and from the sequences of its 16S rDNA, intergenic 16S/23S rDNA and ribosomal protein genes of the beta operon. Phylogenetic analysis showed that the liberibacter present in C. capense differed from the two previously described liberibacter species from citrus and that it was more closely related to 'Candidatus Liberibacter africanus' than to 'Candidatus Liberibacter asiaticus'. It is proposed that the liberibacter from C capense be assigned a subspecies status, 'Candidatus Liberibacter africanus subsp. capensis'.

Within the framework of the characterization of the microflora of an industrial sourdough fermentation, strains of Lactobacillus amylovorus, Lactobacillus pontis and two other strains were isolated which could not be associated with a valid species. These latter strains were Gram-positive, catalase-negative, non-spore-forming, non-motile rods that could be clearly differentiated from known species by 16S rDNA sequence analysis. For further characterization, the morphological, physiological (sugar fermentation, formation of DL-lactate, hydrolysis of arginine, growth temperature, CO2 production) and chemotaxonomic (G+C content, cell wall composition, SDS-PAGE of whole-cell proteins) properties were determined. Fitting of the complete 16S rDNA sequence into alignments of such sequences, together with the subsequent phylogenetic calculations, allowed the reconstruction of a phylogenetic tree. These data showed that the two strains were phylogenetically related but formed an independent cluster distinct from their closest neighbours, L. pontis, Lactobacillus panis, Lactobacillus oris, Lactobacillus vaginalis and Lactobacillus reuteri. The results of DNA-DNA hybridization experiments indicated that the two isolates represent a new Lactobacillus species, for which the name Lactobacillus frumenti is proposed; the type strain of this species is DSM 13145T (= LMG 19473T).

Comparison of the 16S rDNA sequence of Pseudomonas antimicrobica LMG 18920T with published 16S rDNA sequences from other pseudomonads indicated that Pseudomonas antimicrobica belongs to the genus Burkholderia, with Burkholderia gladioli, Burkholderia glumae and Burkholderia plantarii as its closest neighbours. DNA-DNA hybridizations confirmed that Pseudomonas antimicrobica and Burkholderia gladioli represent the same species. Strain LMG 18920T and other Burkholderia gladioli strains were also indistinguishable by SDS-PAGE of whole-cell proteins and had similar biochemical characteristics. The whole-cell fatty acid composition, however, was different from that of other Burkholderia gladioli strains. It is concluded that Pseudomonas antimicrobica is a later synonym of Burkholderia gladioli. As Burkholderia gladioli is known to cause infections in patients with cystic fibrosis and chronic granulomatous disease, the eventual use of strain LMG 18920T as a biological control agent should be approached with caution.

A new thermophilic, anaerobic glucose-fermenting, Gram-positive, rod-shaped bacterium, designated strain SEBR 7858T, was isolated from an oilfield water sample. Under optimal conditions on a glucose-containing medium (3% NaCl, 65 degrees C and pH 7.5), the generation time was 2.5 h. No growth occurred at 35 or 80 degrees C, nor at pH 5..5 or 9.0. Strain SEBR 7858T possessed lateral flagella. Spores were undetected but heat-resistant forms were present. Strain SEBR 7858T fermented a range of carbohydrates to acetate, L-alanine, lactate, H2 and CO2. The isolate reduced thiosulfate and elemental sulfur, but not sulfate or sulfite to sulfide. In the presence of thiosulfate, the ratio of acetate produced per mole of glucose consumed increased, suggesting a shift in the use of electron acceptors during carbohydrate metabolism. The DNA G+C content was 41 mol%. Based on 16S rRNA gene sequence analysis, the strain was almost equidistantly related to all members of the genus Thermoanaerobacter (mean similarity 92%). Based on phenotypic, genomic and phylogenetic characteristics, strain SEBR 7858T was clearly different from all members of the genus Thermoanaerobacter and was therefore designated as a new species, Thermoanaerobacter subterraneus sp. nov. The type strain is SEBR 7858T (= CNCM 1-2383T, DSM 13054T).

Phenotypic and phylogenetic studies were performed with 10 strains of bacteriochlorophyll-containing bacteria isolated from a variety of marine environments (surface of Rhodophyta, sand and algal sand mat) on the east and west coasts of Australia. The strains were aerobic, chemoheterotrophic, Gram-negative, motile rods with peritrichous flagella. Bacteriochlorophyll a was synthesized under aerobic conditions. Catalase, nitrate reductase, oxidase and phosphatase were produced. ONPG reaction was positive. The strains have been divided into genotype group 1 (seven strains) and genotype group 2 (three strains) according to previously described DNA-DNA hybridization data. Strains OCh 254T and OCh 368T have been included in genotype groups 1 and 2, respectively. The results of 165 rRNA gene sequence comparisons revealed that strains OCh 254T and OCh 368T formed a new cluster within the alpha-2 group of the alpha subclass of the Proteobacteria. The similarity value of the 16S rRNA gene sequences between strain OCh 254T and the most closely related species, Stappia aggregata, was 95.6 %. The sequence similarity value between strains OCh 254T and OCh 368T was 97.1%. It was concluded that these two strains should be placed into a new genus, Roseibium gen. nov., as Roseibium denhamense sp. nov. and Roseibium hamelinense sp. nov. The type species of the genus is Roseibium denhamense. The type strains of Roseibium denhamense and Roseibium hamelinense are OCh 254T (= JCM 10543T) and OCh 368T (= JCM 10544T), respectively.

Four strains of purple sulfur bacteria containing bacteriochlorophyll b were isolated from cyanobacterial mats of soda lakes in the steppe of south-east Siberia, Russia. Cells of all strains were cocci without gas vesicles. Eventually, cells with flagella were seen in the electron microscope, but motile cells were observed very rarely in cultures. Internal photosynthetic membranes were of the tubular type. Photosynthetic pigments were bacteriochlorophyll b and carotenoids with spectral characteristics similar to 3,4,3',4'-tetrahydrospirilloxanthin. The bacteria were obligately phototrophic and strictly anaerobic. Hydrogen sulfide and elemental sulfur were used as photosynthetic electron donors. Thiosulfate was not used. During growth on sulfide, sulfur globules were formed as intermediate oxidation products, deposited inside the cells and centrally located. In the presence of sulfide and sodium bicarbonate, acetate, malate, propionate, pyruvate, succinate, fumarate and yeast extract were photoassimilated. Growth factors were not required. The new bacterium is an obligate alkaliphile growing at pH 8-10 with an optimum at pH 9. It showed good growth up to 6.0% sodium chloride and up to 8.5% sodium carbonates. Phenotypically, it is similar to Thiococcus pfennigii, but different by virtue of its alkaliphily and salt tolerance. The DNA G+C content was 63.6-64.8 mol %, compared to 69.4-69.9 mol % for Thiococcus pfennigii. The 16S rDNA sequence of strain A26T was approximately 92% similar to that of Thiococcus pfennigii DSM 226 and therefore a new genus and species name, Thioalkalicoccus limnaeus gen. nov. and sp. nov., are proposed for the new bacterium.

From sequence divergence of 16S rRNA genes and the internally transcribed spacer (ITS) region it is reported that variation in phylogenetic placement exists among the 17 different serotype strains of Bradyrhizobium that have been isolated from nodules of soybean. Evolutionary relationships among the bradyrhizobia were more resolved using reconstructions derived from ITS than from 16S rRNA gene sequence divergence. Strain USDA 129 was placed together with USDA 62, 110, 122 and 126, but did not cluster with USDA 123 and 127, with which it shares antigenic determinants. The results from the phylogenetic analysis were supported with data from determinations of genetic diversity among additional strains within each of these serogroups using amplified fragment length polymorphism analysis. From these results it was concluded that strains of serogroup 129 were more similar to strains of serogroups 62, 110 and 122 than they were to strains of serogroups 123 and 127. The serotype strain of Bradyrhizobiumjaponicum USDA 135 and the type strain for Bradyrhizobium liaoningense possessed identical 16S rRNA gene and ITS region sequences. Also, the type strain for B. liaoningense cross-reacted with antisera prepared against somatic antigens of USDA 135. Therefore, it was not possible to distinguish B. liaoningense from serogroup 135 in our analysis of B. japonicum and Bradyrhizobium elkanii.

The taxonomic position of a bacterial strain (PDB9T) that is capable of degrading pyridine was clarified by a polyphasic taxonomic approach using phenotypic, chemotaxonomic and genetic methods. The cells, which are rods and branched filaments during the early growth phase, fragment into short rods or cocci, thereby completing the growth cycle. Strain PDB9T was found to have a cell wall of chemotype IV, MK-8(H2) as the predominant menaquinone, mycolic acids with 36-46 carbon atoms and C16:0' C18:1 cis9, 10-methyl-C18:0 (TBSA) as the major fatty acids. The G+C content of the DNA was 66 mol%. The phylogenetic tree showed that strain PDB9T falls within an evolutionary radiation comprising Rhodococcus species and is most closely related to the type strain of Rhodococcus rhodochrous, sharing 99% 16S rDNA similarity. The differences in some phenotypic characteristics and the genetic distinctiveness distinguish strain PDB9T from the Rhodococcus species described previously. Therefore, strain PDB9T should be placed in the genus Rhodococcus as a new species, for which the new name Rhodococcus pyridinivorans sp. nov. is proposed. The type strain of the new species is strain PDB9T (= KCTC 0647BPT = KCCM 80005T).

A novel bacterial strain (171544T) was recently isolated from silage and was classified in the genus Bacillus by 16S rDNA sequence analysis. Additional silage samples have been investigated in the present study and four organisms resembling strain 171544T were isolated. Phenotypic and genotypic characterization of these bacteria showed that they constitute a new species of the genus Bacillus. This taxon was positioned in the family Bacillaceae on the basis of evolutionary distance trees using 16S rDNA sequences. Bacillus circulans, Bacillus firmus and Bacillus benzoevorans were the most closely related species with 165 rDNA similarities of 97.2, 96.3 and 95.9%, respectively. All five silage isolates shared a higher order structural feature in the 3' region of the 16S rRNA gene comprising an extension to helix 49 of 24 bp and highly similar random amplified polymorphic DNA patterns that distinguished them from the type strains of B. circulans and B. firmus. Moreover, they possessed a unique pattern of phenotypic features including subterminally or terminally located endospores which distinctly swelled the sporangium, strictly aerobic metabolism but with the ability to utilize nitrate as a terminal electron acceptor under anaerobic conditions, and hydrolysis of casein but not starch. The name Bacillus siralis is therefore proposed for this new taxon. The type strain of B. siralis is strain 171544T (= NCIMB 13601T = CIP 106295T).

Results of a polyphasic taxonomic study on an unknown Gram-negative, facultatively anaerobic, coccobacillus-shaped organism isolated from an infected human gall bladder are presented. Phenotypic and molecular taxonomic studies revealed the organism to be close to, but distinct from, organisms designated CDC (Centers for Disease Control and Prevention) group DF-3. The unknown bacterium was readily distinguished from reference strains of Bacteroides, Prevotella, Porphyromonas and related taxa by 16S rRNA gene sequencing, biochemical tests, analysis of cellular long-chain fatty acids and electrophoretic analysis of whole-cell proteins. Based on the results of the present study, it is proposed that the unknown bacterium be classified in a new genus, Dysgonomonas, as Dysgonomonas gadei sp. nov. (type strain CCUG 42882T = CIP 106420T). In addition, a new species, Dysgonomonas capnocytophagoides sp. nov., is proposed to accommodate strains previously belonging to CDC group DF-3. The type species of the genus Dysgonomonas is Dysgonomonas gadei.

Numerous micro-organisms have been described as cytoplasmic symbionts of eukaryotes. Many so-called obligate endosymbionts rely exclusively on maternal (vertical or transovarial) transmission to maintain themselves, rendering them dependent on the host sex ratio, which they would tend to manipulate to their own advantage. The latter phenomenon is often associated with the presence of Wolbachia pipientis (alpha-Proteobacteria) in arthropods and nematodes. A potentially similar situation was discovered involving members of a new clade of Verrucomicrobia, another main line of descent in the Bacteria. Nematode species of the Xiphinema americanum group (Nematoda, Longidoridae), viz. Xiphinema americanum, Xiphinema rivesi and Xiphinema brevicollum, each harbour their own specific verrucomicrobial endosymbionts. They are exclusively maternally inherited and their hosts reproduce by thelytokous (mother-to-daughter) parthenogenesis, males being extremely rare. A new genus, 'Candidatus Xiphinematobacter' gen. nov., along with three new candidate verrucomicrobial species, 'Candidatus Xiphinematobacter americani' sp. nov., 'Candidatus Xiphinematobacter rivesi' sp. nov. and 'Candidatus Xiphinematobacter brevicolli' sp. nov., are described on the basis of transmission electron microscopy, scanning electron microscopy, DAPI (4',6-diamidino-2-phenylindole) epifluorescence microscopy and 16S rDNA sequence analysis. These are the first endosymbiotic species described among the Verrucomicrobia. They share a mean 16S rDNA similarity of about 93%, whereas similarity to their closest relative, clone WCHD3-88, is less than 87%. Thus, the endosymbionts form a homogeneous clade for which the new candidate genus 'Candidatus Xiphinematobacter' gen. nov. is proposed. The type species is 'Candidatus Xiphinematobacter brevicolli' sp. nov.

A large collection of bacterial strains, immunotrapped from soil and from the wheat rhizoplane, was subjected to polyphasic taxonomy by examining various pheno- and genotypic parameters. Strains were grouped on (inter) repetitive extragenic palindromic DNA (REP) PCR profiles at the intraspecies level. Pheno- and genotypic characters were assessed for representatives from 13 different REP groups. Strains of nine REP groups constituting two physiological BIOLOG clusters fell in the coherent DNA-DNA reassociation group of Ochrobactrum anthropi. Strains of two REP groups constituting a separate BIOLOG cluster fell in the coherent DNA-DNA reassociation group of Ochrobactrum intermedium. Additional phenotypic characters differentiating O. anthropi and O. intermedium were found. REP group K strains constituted a different BIOLOG cluster, a separate DNA-DNA reassociation group and a distinct phylogenetic lineage in 165 rDNA homology analysis, indicating that REP group K strains represent a new species. Diagnostic phenotypic characters were found. Closest relatives were Ochrobactrum species. The name Ochrobactrum grignonense sp. nov. is proposed (type strain OgA9aT = LMG 18954T = DSM 13338T). REP group J strains again constituted a different BIOLOG cluster, a separate DNA-DNA reassociation group and showed, as a biological particularity, a strict preference for the rhizoplane as habitat. Diagnostic phenotypic characters were found. This indicated that REP group J strains represent a further new species, although phylogenetic analyses using 16S rDNA homology were not able to separate the cluster of REP group J sequences significantly from 16S rDNA sequences of Ochrobactrum anthropi. The name Ochrobactrum tritici sp. nov. is proposed (type strain SCII24T = LMG 18957T = DSM 13340T).

Acetobacter oboediens Sokollek et al. 1998 and Acetobacter intermedius Boesch et al. 1998 are transferred to the genus Gluconacetobacter as Gluconacetobacter oboediens comb. nov. and Gluconacetobacter intermedius comb. nov. because, on the basis of their 16S rRNA gene sequences, the type strains of both species are located in the cluster of the genus Gluconacetobacter along with those of Gluconacetobacter xylinus, Gluconacetobacter europaeus, Gluconacetobacter hansenii, Gluconacetobacter liquefaciens (the type species) and Gluconacetobacter diazotrophicus. The significance of growth on mannitol agar and the presence of a ubiquinone isoprenologue composed of Q-10 is discussed for characterization of the genus Gluconacetobacter.

A formal proposal is made to rename the order Haloanaerobiales as Halanaerobiales, the family Haloanaerobiaceae as Halanaerobiaceae, the genera Haloanaerobium and Haloanaerobacter as Halanaerobium and Halanaerobacter, respectively, and to change the specific epithets of Halanaerobium acetoethylicum, Halanaerobacter lacunaris, Halocella cellulolytica and Halanaerobium chitinovorans to acetethylicum, lacunarum, cellulosilytica and chitinivorans, respectively. These changes are required in view of Rule 61, Appendix 9, of the Bacteriological Code.

The International Committee of Systematic Bacteriology Subcommittee on the taxonomy of Campylobacter and related bacteria has agreed in principle on minimum requirements for the description of new species of the genus Helicobacter. These requirements include the recommendation that the description of a putative new species or subspecies be based on examination of at least five strains in order to provide a measure of phenotypic and genotypic variation. Recommendations for required phenotypic and molecular data are presented. The requirements are consistent with the polyphasic approach to bacterial systematics.