- Volume 39, Issue 1, 1989

Reverse transcriptase sequencing of 16S ribosomal ribonucleic acid of Caseobacter polymorphus was performed in order to determine the relationship of this organism to other mycolic acid-containing actinomycetes and coryneform bacteria. The sequence data clearly demonstrate that Caseobacter polymorphus is a member of the genus Corynebacterium. Furthermore, Caseobacter polymorphus showed 100% sequence homology in a large stretch of 1,458 nucleotides with Corynebacterium variabilis. On the basis of the present and previous data we formally propose that Caseobacter polymorphus be considered a subjective synonym of Corynebacterium variabilis.

A new species of coccoid methanogen was isolated from the surface sediments of Tar Pit Lake at the LaBrea Tar Pits in Los Angeles, Calif. Surface colonies of strain ZT (= OGC 1T) (T = type strain) were tan, circular, clear, and convex with entire edges. The cells were irregular, nonmotile, and coccoid (diameter, 0.4 to 2.0 µm) and grew by producing methane from H2-CO2 or formate but not from methanol, methylamines, or acetate. They exhibited a negative Gram reaction, had protein cell walls, and were susceptible to lysis by sodium dodecyl sulfate. They grew over a narrow pH range (pH 6.5 to 7.5), with fastest growth near pH 7, and they were mesophilic (growth was most rapid at 37°C). Cells grew fastest with 15 g or less of NaCl per liter of medium. We propose that this isolate be named as a new species of methanogen, Methanocorpusculum labreanum, with strain Z (= Oregon Graduate Center Collection strain OGC 1 = DSM 4855) as the type strain.

A recent epiphytotic disease on citrus in Florida nurseries was caused by strains of Xanthomonas campestris with different host specificity and lower pathogenic capacities than those of previously described strains of X. campestris pv. citri. The new strains were classified as X. campestris pv. citri because they were isolated from rutaceous hosts and despite the fact that they caused a different disease than strains previously described in that pathovar. Restriction fragment length polymorphism analyses revealed that the Florida strains comprised a heterogeneous (E) group, interrelated with X. campestris pv. alfalfae, X. campestris pv. cyamopsidis, and X. campestris pv. dieffenbachiae. In contrast, the previously described strains of X. campestris pv. citri formed two highly distinct, homogeneous (A and B/C/D) groups. Furthermore, the strains of X. campestris pv. campestris, X. campestris pv. glycines, X. campestris pv. malvacearum, X. campestris pv. phaseoli, X. campestris pv. pisi, and X. campestris pv. vignicola tested were also distinctive and appeared to be only distantly related to one another and to all X. campestris pv. citri strains. We concluded (i) that some pathovars are sufficiently distinct from the type strain of the species X. campestris to be considered as separate species, (ii) that X. campestris pv. citri group A and X. campestris pv. phaseoli (not including X. campestris pv. phaseoli var. fuscans) represent distinctly separate subbranches of the genus and should be respectively reinstated to species as X. citri (ex Hasse) nom. rev. 3213 and X. phaseoli (ex Smith) nom. rev. G27, and (iii) that the X. campestris pv. citri B/C/D strains and the heterogeneous E strains should be respectively renamed X. campestris pv. aurantifolii pv. nov. and X. campestris pv. citrumelo pv. nov.

A comprehensive study of the relationships of Oceanospirillum species by a polyphasic approach is described. Hybridizations were performed between labeled ribosomal ribonucleic acids (rRNAs) from Oceanospirillum linum, Oceanospirillum jannaschii, Oceanospirillum kriegii, and several other organisms and deoxyribonucleic acids (DNAs) from representative Oceanospirillum strains and many other gram-negative organisms. Five Oceanospirillum species, including the type species, constitute a separate rRNA branch in rRNA superfamily II, to which the genus Oceanospirillum should be restricted. The finer relationships among the different members of the Oceanospirillum rRNA branch were further defined by DNA:DNA hybridizations and comparative gel electrophoresis of whole-cell proteins. After comparison of our results with the previously available phenotypic data, we redefine the genus Oceanospirillum to contain Oceanospirillum linum, Oceanospirillum maris, Oceanospirillum beijerinckii, Oceanospirillum multiglobuliferum, and Oceanospirillum japonicum; we also create Oceanospirillum maris subsp. hiroshimense comb. nov. and Oceanospirillum beijerinckii subsp. pelagicum comb. nov. for the former Oceanospirillum hiroshimense and Oceanospirillum pelagicum, respectively. The following taxa are generically misnamed: Oceanospirillum jannaschii, Oceanospirillum kriegii, and Oceanospirillum minutulum. Together with Cellvibrio mixtus and some misnamed Pseudomonas strains, these three taxa constitute at least three separate rRNA branches in rRNA superfamily II. Oceanospirillum pusillum is also misnamed as it belongs in rRNA superfamily IV. Oceanospirillum vagum and Oceanospirillum commune should be relegated to their real generic positions, Marinomonas vaga and Marinomonas communis, respectively.

The deoxyribonucleic acid (DNA):ribosomal ribonucleic acid (rRNA) hybridization technique was used to reveal the relationships and taxonomic positions of an additional 83 strains belonging to 43 saprophytic or pathogenic Pseudomonas species and 29 named and unnamed Pseudomonas-like strains. The DNA:rRNA hybrids were characterized by the following two parameters: (i) the temperature at which one-half of the hybrid was eluted and (ii) the percentage of rRNA binding (the amount of rRNA bound per 100 pg of filter-fixed DNA). We also used, for a limited number of strains, numerical analysis of carbon assimilation tests to delineate the finer taxonomic relationships of organisms. Of the 83 strains examined, 78 could be definitely assigned either to an rRNA branch or to an rRNA superfamily within the Proteobacteria. Only 25 of our strains belong in the genus Pseudomonas sensu stricto (our Pseudomonas fluorescens rRNA branch). In general, about two-thirds of the named Pseudomonas species have been misclassified and are distributed over at least seven genera all through the Proteobacteria. These organisms need to be reclassified and generically renamed according to their phylogenetic relationships. However, more detailed phenotypic and genotypic studies are necessary before definite nomenclatural proposals can be made. A comprehensive list of the phylogenetic affiliations of the Pseudomonas species is included.

A new genus of acidophilic, facultatively methylotrophic bacteria is described. These organisms are gram-negative, nonsporeforming, nonmotile, and rod shaped and grow at pH 2.0 to 5.5. These characteristics are unique among the methanol-utilizing bacteria. The deoxyribonucleic acid base composition is 63 to 65 mol% guanine plus cytosine. Acetobacter methanolicus TK 0705T (T = type strain) is a typical strain in this group. These bacteria are distinguished from type and representative strains of Acetobacter, Gluconobacter, Acidiphilium, and Thiobacillus on the basis of deoxyribonucleic acid-deoxyribonucleic acid homology. A new genus, Acidomonas, is proposed to include this group.of methylotrophic bacteria. The type species of the genus Acidomonas is Acidomonas methanolica comb, nov., with type strain TK 0705 (= IMET 10945).

3-0-methylrhamnose was found in cell walls of Catellatospora ferruginea. This sugar was purified and identified by proton and 13C nuclear magnetic resonance and mass spectroscopy, using the acetylation method. It is localized in the cell wall fraction and occurs at a constant ratio among the cell wall sugars when various carbon sources are used for growth. 3-O-methylrhamnose was found in nine strains of Catellatospora species containing MK-10 as the major menaquinone, but not in nine strains containing MK-9. Among 22 type strains of genera related to Catellatospora (Actinoplanes, Amorphosphorangium, Ampullariella, Dactylosporangium, Micromonospora, Glycomyces, and “Catenuloplanes”), 4 strains contained only trace quantities, and the remainder contained none. Thus, we propose that 3-O-methylrhamnose is a key sugar in the chemotaxonomy of the genes Catellatospora.

Tests allowing the screening of large numbers of enterobacterial strains for the presence of the glucose oxidation pathway (glucose, gluconate, and 2-ketogluconate dehydrogenases) were devised or adapted. A total of 506 strains representing 111 taxa (named species or subspecies and unnamed genomic groups) were studied. The members of the genera Budvicia, Edwardsiella, Leminorella, Providencia, and Xenorhabdus and the species Citrobacter freundii, Erwinia carnegeana, Erwinia carotovora, Erwinia chrysanthemi, Erwinia nigrifiuens, Erwinia Salicis, Moellerella wisconsensis, Proteus penneri, Proteus vulgaris, Yersinia intermedia, Yersinia pestis, Yersinia pseudotuberculosis, and Yersinia ruckeri were negative in all tests. Five species, Erwinia cypripedii, Ewingella americana, Rahnella aquatilis, Serratia marcescens (at 20°C), and Tatumella ptyseos produced 2,5-diketogluconate from glucose without a requirement for pyrroloquinoline quinone (PQQ). When PQQ was provided (required for glucose oxidation), Serratia grimesii and Serratia liquefaciens produced 2,5-diketogluconate from glucose at 20°C. Escherichia blattae had gluconate- and 2-ketogluconate dehydrogenases without glucose dehydrogenase. The members of the genera Hafnia, Obesumbacterium, and Pragia had only gluconate dehydrogenase. Other species had glucose dehydrogenase (with or without a requirement for PQQ) with or without gluconate dehydrogenase. Classification and identification may take advantage of tests exploring the glucose oxidation pathway.

A new anaerobic mesophilic cellulolytic Clostridium isolated from a methanogenic cellulose-enriched culture is described. Optimal growth occurs at 35°C and pH 7.0. Fructose, maltose, rhamnose, lactose, glucose, xylose, ribose, mannose, raffinose, cellobiose, arabinose, galactose, mannitol, adonitol, esculin, salicin, and trehalose serve as substrates for growth. The major fermentation products from cellulose or cellobiose are ethanol, acetate, formate, butyrate, isobutyrate, isovalerate, caproate, lactate, succinate, CO2, and H2. The deoxyribonucleic acid base composition is 38 mol% guanine plus cytosine. The type strain is 18A (= CECT 954).

A new species of the genus Amycolata, Amycolata alni, is proposed for strains which were mostly isolated from root nodules and rhizospheres of alder trees and were formerly assigned to the species Amycolata autotrophica. Strains of this new species could be differentiated from representatives of A. autotrophica by deoxyribonucleic acid homology data (17 to 29% relatedness), as well as by their ability to grow on salicin and D-gluconate, their failure to produce acid from cellobiose and meso-inositol, their resistance to penicillin and carbenicillin (10 µg/ml), and other characteristics. Phenotypic features and low levels of deoxyribonucleic acid homology distinguish A. alni from two other species of the genus Amycolata, Amycolata saturnea and Amycolata hydrocarbonoxydans. The type strain of A. alni is strain 3LS (= VKM Ac-901).

The 5S ribosomal ribonucleic acid (rRNA) sequences were determined for Bacteroides fragilis, Bacteroides thetaiotaomicron, Bacteroides capillosus, Bacteroides veroralis, Porphyromonas gingivalis, Anaerorhabdus furcosus, Fusobacterium nucleatum, Fusobacterium mortiferum, and Fusobacterium varium. A dendrogram constructed by a clustering algorithm from these sequences, which were aligned with all other hitherto known eubacterial 5S rRNA sequences, showed differences as well as similarities with respect to results derived from 16S rRNA analyses. In the 5S rRNA dendrogram, Bacteroides clustered together with Cytophaga and Fusobacterium, as in 16S rRNA analyses. Intraphylum relationships deduced from 5S rRNAs suggested that Bacteroides is specifically related to Cytophaga rather than to Fusobacterium, as was suggested by 16S rRNA analyses. Previous taxonomic considerations concerning the genus Bacteroides, based on biochemical and physiological data, were confirmed by the 5S rRNA sequence analysis.

The genus Bacteroides (Castellani and Chalmers) as presently constituted is phenotypically and phylogenetically extremely heterogeneous. However, biochemical, chemical, and molecular biological data indicate that Bacteroides fragilis (the type species) and related species form a coherent taxonomic group. Therefore, we formally propose that the genus Bacteroides be restricted to the B. fragilis group of species, and the description of the genus is emended accordingly.

A third species, Chlamydia pneumoniae, is proposed for the genus Chlamydia. This bacterium is a human respiratory pathogen, which has been referred to as the TWAR strain of Chlamydia. Species identification is based on ultrastructural differences in the elementary bodies, deoxyribonucleic acid analysis, and serology.

Several methanogenic bacteria that utilize methanol, acetate, or me thy lamines have been isolated from marine habitats, including Methanococcus frisius. However, the 1984 description by Mah and Kuhn of the genus Methanococcus excludes methylotrophic methanebacteria such as Methanococcus frisius. Therefore, we propose to transfer this species to the genus Methanosarcina, naming it Methanosarcina frisia comb. nov. (type strain, DSM 3318 [= ATCC 43330]). Other species having similar properties (e.g., Methanococcus halophilus) should be assignable to Methanosarcina.