- Volume 34, Issue 2, 1984

Sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis was used to compare soluble protein fractions from 36 strains of Rothia dentocariosa and related bacteria. The resulting protein patterns were examined to identify the bands of major and minor frequency within the genus. A cluster analysis revealed three major Rothia groups and a fourth group representing the genus Actinomyces. Our data demonstrated good correlations (>70%) with existing biotyping and serotyping schemes of these isolates. We suggest that electrophoretic group 2 represents a new Rothia species.

A total of 23 strains of coryneform bacteria that cause ratoon stunting disease of sugarcane and Bermudagrass stunting disease were examined. These included 17 sugarcane strains from Florida, Louisiana, South Africa, Brazil, and Japan and six Bermudagrass strains from Florida and Taiwan. The sugarcane and Bermudagrass strains contained 2,4-diaminobutyric acid, rhamnose, and fucose in their cell walls, suggesting a relationship to some other phytopathogenic coryneform bacteria and related organisms with similar cell wall compositions. The cellular protein patterns of the sugarcane and Bermudagrass strains after polyacrylamide gel electrophoresis resembled the patterns of Corynebacterium michiganense subspecies more than the patterns of some other diaminobutyric acid-containing taxa. The protein patterns of the sugarcane strains were distinct from the patterns of the Bermudagrass strains examined. The guanine-plus-cytosine contents of the deoxyribonucleic acids of two sugarcane strains and one Bermudagrass strain were 66 mol%. The sugarcane and Bermudagrass strains formed a homogeneous group based on morphological characteristics and most biochemical characteristics. All of the sugarcane strains were nonpigmented (white) and grew more slowly in culture than the yellow-pigmented Bermudagrass strains. The Bermudagrass strains, unlike the sugarcane strains, hydrolyzed starch and utilized citrate and malate. Although the phytopathogenic coryneform bacteria with 2,4-diaminobutyric acid in the peptidoglycans of their cell walls are presently classified in the genus Corynebacterium, recent studies have indicated that these phytopathogens should be reclassified. These bacteria form a distinct group with characteristics different from those of all existing genera as presently circumscribed; therefore, we propose that a new genus, Clavibacter, be established for these and similar bacteria. We propose reclassification of Corynebacterium michiganense (including Corynebacterium michiganense subsp. michiganense, Corynebacterium michiganense subsp. insidiosum, Corynebacterium michiganense subsp. sepedonicum, Corynebacterium michiganense subsp. nebraskense, and Corynebacterium michiganense subsp. tessellarius), Corynebacterium iranicum, Corynebacterium tritici, and Corynebacterium rathayi in the new genus Clavibacter. We further propose that Clavibacter xyli be recognized as a new species with the following subspecies: Clavibacter xyli subsp. xyli sp. nov., subsp. nov., the species and subspecies type strain of which is sugarcane strain L1A (= ATCC 33974 = NCPPB 3152 = PDDC 7127), and Clavibacter xyli subsp. cynodontis subsp. nov., the subspecies type strain of which is Bermudagrass strain TB1A (= ATCC 33973).

A new species, Bacteroides endodontalis, is proposed for black-pigmented asaccharolytic Bacteroides strains that have negligible deoxyribonucleic acid homology with either Bacteroides gingivalis or Bacteroides asaccharolyticus. Strain HG370 (= ATCC 35406) is the type strain. Unlike B. gingivalis, B. endodontalis does not agglutinate sheep erythrocytes or produce phenylacetic acid. B. endodontalis resembles B. asaccharolyticus physiologically but can be distinguished by its lower deoxyribonucleic acid guanine-plus-cytosine content (50 versus 53 to 54 mol%), by its serological reactions, by its electrophoretic patterns of proteins, and possibly by its menadione requirement.

Presumably because the name “Bacillus psychrophilus” Larkin and Stokes 1967 was considered to be a subjective synonym of Bacillus globisporus, it was not included on the Approved Lists of Bacterial Names and, therefore, has no standing in bacteriological nomenclature. The low deoxyribonucleic acid relatedness values of 8 to 36% found in the present study between “B. psychrophilus” and B. globisporus strains suggested that these two organisms are not closely related genetically. Moreover, the ability of “B. psychrophilus” strains to ferment d-mannitol, d-ribose, trehalose, and d-xylose, to reduce nitrate to nitrite, and to grow at 30°C and in 3% NaCl distinguished them phenotypically from B. globisporus strains. Based on the results of this study, revival of the name Bacillus psychrophilus is proposed. The type strain is strain NRRL NRS-1530.

Previous studies of Acholeplasma species indicated that this genus could be separated from Mycoplasma on the basis of the ability to synthesize lipids from acetate. We report that recently described strains of Acholeplasma florum (strain L1 isolated from a lemon flower and strain GF-1 isolated from a grapefruit flower) and two unclassified Acholeplasma strains (strain J233 from coconut palm and Acholeplasma sp. strain PS-1 from an insect) convert little if any acetate carbon to lipid, in contrast to Acholeplasma sp. strain 0502-CL1 isolated from broccoli and six other established Acholeplasma species of animal origin. Thus, the ability to synthesize lipid from acetate in Edward test medium containing horse serum is not characteristic of all members of the genus Acholeplasma (class Mollicutes), but appears to be a property only of a subgroup in the genus containing the species that are most frequently isolated from animals (Acholeplasma laidlawii, Acholeplasma granularum BTS39, Acholeplasma oculi 19L, Acholeplasma equifetale N93, and probably Acholeplasma axanthum S743 and Acholeplasma modicum PG49). Like other Mycoplasma species which we studied, Mycoplasma bovigenitalium PG11, Mycoplasma hominis PG27, Mycoplasma arginini G230, and four sterol-requiring mycoplasmas of insect or plant origin (Mycoplasma sp. strain MQ3 from an insect, Mycoplasma sp. strain pommier from an apple, Mycoplasma sp. strain melaleuca from a Melaleuca flower, and Mycoplasma sp. strain 831-C4 from lettuce) convert little acetate to lipid. Similarly, eight serovars of Ureaplasma urealyticum convert little if any acetate to lipid.

Two groups of nocardioform organisms isolated from marine sediments were studied, along with representative strains of the genera Rhodococcus, Nocardia, and Mycobacterium. Strains of the red- or orange-colored group showed no marine affinity and exhibited a close relationship to Rhodococcus rhodochrous. The 17 marine isolates of the cream-colored group formed a homogeneous well-separated cluster and were particularly characterized as moderately halophilic and psychrotrophic organisms. These isolates were assigned to a new species within the genus Rhodococcus. We propose the name Rhodococcus marinonascens, referring to the exclusive occurrence of this taxon in the sea, and describe the properties of the species and of type strain 3438W (= DSM 43752).

The physiological characteristics of three facultatively chemolithotrophic Thiobacillus species with ubiquinone 8 (group II) (Thiobacillus delicatus THI 091T [T = type strain] Thiobacillus perometabolis THI 023 , and Thiobacillus intermedius THI 101T) were compared. These organisms differ from other facultatively chemolithotrophic Thiobacillus strains with ubiquinone 10 (group I) in requiring thiosulfate or elemental sulfur for optimum growth. T. delicatus THI 091T, T. perometabolis THI 023 , and T. intermedius THI 101T are regarded as three distinct species on the basis of their physiological characteristics. T. delicatus THI 091T differs from T. perometabolis THI 023T and T. intermedius THI 101T in its lack of motility, its ability to grow anaerobically with nitrate, and its intermediate formation of polythionate during thiosulfate oxidation. T. perometabolis THI 023T is distinguished from T. intermedius THI 101T by its ability to grow heterotrophically with a single carbon source. T. delicatus is not on the Approved Lists of Bacterial Names; therefore, this name is revived. In addition, the description of this organism is emended, considering its inability to grow heterotrophically with a single carbon source.

On the basis of phenotypic properties and the guanine-plus-cytosine content of the deoxyribonucleic acid, as well as the immunological relationship of the glutamine synthetase and the iron-containing superoxide dismutase, strain 217T (= ATCC 19375) (T = type strain), which was previously designated “Pseudomonas nigrifaciens,” was shown to be a member of a distinct species in the genus Alteromonas. Since this species designation was not included on the Approved Lists of Bacterial Names, it has been revived as Alteromonas nigrifaciens sp. nov., nom. rev., with type strain 217 (= ATCC 19375). The phenotypic properties of strains of Alteromonas macleodii and Alteromonas haloplanktis are also described.

A group of organisms recovered from human intestinal tracts, sewage, and river water were shown in a previous study to be related to members of the family Vibrionaceae. On the basis of additional comparative studies of the biochemical and metabolic properties of these organisms and their distinction from other members of the Vibrionaceae by deoxyribonucleic acid hybridization tests, we hereby propose that this group of strains be designated Allomonas enterica gen. nov., sp. nov. Strain 40 (= BKM B1485) is the type strain.

A new cellulolytic Clostridium species that was isolated from decayed grass is described. The colonies produced by this organism on cellulose agar are circular, translucent, and unpigmented and have undulate margins. The cells of this bacterium are gram-positive straight to slightly curved rods 3 to 6 μm long by 0.6 to 1 μm wide and are motile; they form round, terminal spores 1.5 μm in diameter. A variety of carbohydrates are fermented by this mesophilic anaerobe. The major fermentation products from cellulose are carbon dioxide, hydrogen, ethanol, acetate, lactate, and formate. The deoxyribonucleic acid base composition is 41 mol% guanine plus cytosine. The type strain of Clostridium cellulolyticum sp. nov. is strain H10 (= ATCC 35319).

The cellular lipids of 11 strains of Treponema hyodysenteriae and 10 strains of non-enteropathogenic, weakly beta-hemolytic spirochetes classified as Treponema innocens were extracted with chloroform-methanol and analyzed by thin-layer and gas-liquid chromatography for fatty acids, alkenyl moieties, alkenyl ether lipid compositions, monoglycosyldiglyceride, acylmonoglycosyldiglyceride, and the types of sugars in the glycolipids. Most strains were distinguishable at the species level on the basis of the ratio of 14:0 to iso-15:0 alkenyl moieties; 14:0 moieties predominated in most T. hyodysenteriae strains, whereas most strains classified as T. innocens contained more iso-15:0 moieties. The other aspects of lipid composition were not useful for differentiation. All of the strains had similar cellular fatty acid profiles and contained monoglycosyldiglyceride. The mean alkenyl ether lipid compositions were 48 and 32% of the total lipids of strains of T. hyodysenteriae and T. innocens, respectively. Galactose was the predominant sugar in the glycolipids of 18 strains, 17 of which also contained acylmonoglycosyldiglyceride. Three strains classified as T. innocens contained primarily glucose in their glycolipids and did not contain acylmonoglycosyldiglyceride. Gas-liquid chromatographic determinations of the alkenyl chain profiles of the extractable cellular lipids provided useful information for differentiation of the etiological agent of swine dysentery from morphologically similar, nonpathogenic spirochetes.

Previously, a group of 40 Yersinia enterocolitica-like strains that were isolated from water and fish were called group X2. These strains produced acid from l-rhamnose, did not ferment sorbose, cellobiose, melibiose, or raffinose, and rarely fermented sucrose (5% in 48 h, 10% in 7 days). This pattern of reactions separated group X2 strains from Yersinia enterocolitica, Yersinia intermedia, Yersinia frederiksenii, and Yersinia kristensenii. Positive reactions for acetoin production (Voges-Proskauer test), ornithine decarboxylase, and lack of acid production from melibiose distinguished group X2 strains from both Yersinia pseudotuberculosis and Yersinia pestis. Group X2 strains exhibited variable reactions only in tests for citrate utilization, hydrolysis of Tween 80, and acid production from maltose. Genetically, group X2 strains formed a single deoxyribonucleic acid hybridization group with an average level of relatedness of 86% or more (86% as determined by the S1 method at 60°C or by the hydroxyapatite method at 75°C and 92% as determined by the hydroxyapatite method at 60°C). The level of divergence among related sequences in 60°C reactions was 0.5%, as determined by the hydroxyapatite method. The relatedness of group X2 strains to other Yersinia species was 42 to 73% in 60°C reactions. The divergence in these reactions was 11.0 to 15.5%, and the relatedness to other yersiniae in 75°C reactions was 21 to 38%. Group X2 strains were 11 to 32% related to 67 species of the Enterobacteriaceae that belonged to genera other than Yersinia. On the basis of these biochemical and genetic data, we believe that group X2 represents a single new species in the genus Yersinia. The name Yersinia aldovae sp. nov. is proposed for this species. The type strain of Y. aldovae is strain CNY 6005 (= CDC 669-83 = ATCC 35236).

Spiroplasma citri strains Maroc R8A2T (T = type strain), C189, and C3B, corn stunt Spiroplasma sp. strain E275, and honeybee Spiroplasma sp. strain BC3 all cross-reacted in reciprocal serological tests when an agar double-diffusion system was used. These strains, which are members of Spiroplasma group I, did not show any cross-reactions with flower Spiroplasma strains BNR1 and PPS1, (Spiroplasma floricola and “Spiroplasma apis”) or with the tick-derived organism Spiroplasma mirum. Tick Spiroplasma sp. strain 277F and honeybee Spiroplasma sp. strain AS576 reacted only with antisera prepared against other members of group I. Group III flower spiroplasmas S. floricola 23-6T and BNR1 and group IV strains PPS1 and SR3 did not react with antisera prepared against isolates outside their own groups. The members of each major group showed close antigenic relatedness to each other but not to S. mirum strain SMCAT, which showed minimal one-way cross-reactions with members of group I. Previous serogrouping results for the Spiroplasma strains tested were confirmed by the serological reactions of the organisms in the double-diffusion system.

A total of 22 strains of curved, anaerobic bacteria from the vaginae of women with bacterial vaginosis were found to be morphologically, biochemically, and genetically distinct from species in previously described genera. We placed these organisms in the genus Mobiluncus gen. nov., which includes curved, anaerobic, motile, rod-shaped bacteria that have multiple subpolar flagella and multilayered gram-variable cell walls, produce succinate and acetate, are stimulated by rabbit serum, and have guanine-plus-cytosine contents of 49 to 52 mol%. Mobiluncus curtisii is the type species (type strain, ATCC 35241). Mobiluncus curtisii strains are small (length, 1.7 μm) and gram variable, are stimulated by arginine, and produce ornithine, citrulline, and ammonia from arginine. Six of these strains migrate through soft (0.25%) agar and are nitrate negative, and these organisms are designated Mobiluncus curtisii subsp. curtisii. Six strains that do not migrate through soft agar and are nitrate positive are designated Mobiluncus curtisii subsp. holmesii (type strain, ATCC 35242). A total of 10 strains are designated Mobiluncus mulieris (type strain, ATCC 35243). These organisms are large (length, 2.9 μm) and gram negative, produce acid (pH <5.5) from glycogen, and are CAMP test positive. The levels of deoxyribonucleic acid homology were >75% within and ≤25% between strains of the two species.

A new mesophilic, cellulolytic species of Bacteroides was isolated from a methanogenic cellulose enrichment culture of municipal sewage sludge and is described. This species ferments only cellulose and cellobiose. The fermentation products are acetic acid, CO2, H2, ethanol, and small amounts of lactic acid. The deoxyribonucleic acid base composition is 43 mol% guanine plus cytosine. The name Bacteroides cellulosolvens is proposed. Type strain WM2 is deposited in the National Research Council of Canada culture collection as strain NRCC 2944.

A new genus and species of facultatively methylotrophic bacteria are described. These bacteria are polarly flagellated, gram-negative, nonsporeforming, rod-shaped organisms that occur singly and in pairs. Carotenoid pigment and bacteriochlorophyll are formed in the cells. The deoxyribonucleic acid base composition is 65 to 67 mol% guanine plus cytosine. The cellular fatty acids consist of a large amount of straight-chain unsaturated C18:1 acid and small amounts of straight-chain saturated C16:1 acid, C19:0 cyclopropane acid, and 3-OH-C14:0 hydroxy acid. The major ubiquinone is Q-10. Ubiquinones Q-8, Q-9, and Q-11 are present as minor components. A new genus, Protomonas, is proposed to include this group of methylotropic bacteria. The type species of the genus Protomonas is Protomonas extorquens comb. nov., with type strain TK 0001 (=DSM 1337 = NCIB 9399).

We compared the protein and glycoprotein patterns of 39 Bacillus stearothermophilus strains of various origins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The ultrastructures of the cell walls of intact cells were examined by freeze-etching, negative-staining, and thin-sectioning techniques. A total of 30 strains possessed a paracrystalline protein or glycoprotein surface layer (S-layer) which resembled the highest-molecular-weight band(s) on the electrophoretograms. We found a remarkable heterogeneity in the molecular weights of the S-layer subunits and the geometry and constants of the S-layer lattices, even among closely related strains. All of the strains examined were equally susceptible to lysozyme, indicating that the S-layers have pores at least 3.5 nm wide.

The nonmotile, vibrioid, curly or ring-forming bacteria were examined to assess their systematic relationships through determination of the guanine-plus-cytosine contents of their deoxyribonucleic acids and the degrees of homology among the deoxyribonucleic acids of the various genera and species. Our results verify that Runella, Flectobacillus, Spirosoma, and Ancylobacter (“Microcyclus”) are separate genera. However, our results also indicate that (i) “Ancylobacter eburneus” is not a separate species but is in fact a strain of Ancylobacter aquaticus, (ii) Flectobacillus marinus should not be used to create the new genus “Cyclobacterium” and the latter name should be discarded, and (iii) “Renobacter” is very similar to Ancylobacter, but these two genera exhibit very low levels of deoxyribonucleic acid-deoxyribonucleic acid homology and should probably remain separate at least until additional information is available.

Syntrophus buswellii, which was isolated from anaerobic digestor sludge and is present in aquatic sediments, is a motile, gram-negative, anaerobic rod-shaped organism that requires coculture with an appropriate hydrogenotroph for growth. This bacterium produces acetate, CO2, and H2 (or formate) from benzoate and, possibly, hydrocinnamate (phenyl-3-propionate). A detailed description is given.