- Volume 27, Issue 3, 1977

Examination of 682 strains representing four species of mycobacteria (Mycobacterium phlei, M. smegmatis, M. fortuitum, and M. marinum) and seven species of nocardiae (Nocardia asteroides, N. caviae, N. brasiliensis, N. autotrophica, N. dassonvillei, N. madurae, and N. pelletieri) indicated that, with the exception of N. madurae and N. pelletieri, resistance to rifampin (20 μg/ml) is a useful property for describing these species. Resistance to lysozyme (300 to 500 U/ml) is taxonomically useful for delineating all 11 species.

One hundred and two strains of aerobic, gram-negative bacteria, isolated mainly from drugs and cosmetics, were tested for catabolic products after incubation in liquid media containing single amino acids. Breakdown products from L-arginine, lysine, and ornithine were identified as their N-heptafluoryl butyl esters by gas-liquid chromatographic techniques. A Finnigan model 1015 D gas chromatograph/mass spectrometer/PDP-8E data system was used to identify trace amounts of catabolites after amino acid substrates were inoculated with bacteria. The derivatization reaction was monitored in like manner. The Finnigan data system may be used to identify catabolic compounds rapidly and thus provide a means for bacterial characterization and identification.

The migration of ribosomal ribonucleic acids (rRNA's) from bacteria, bacterial L forms, mycoplasmas, and acholeplasmas in nondenaturing and formamide gels has been examined. Mycoplasmal and acholeplasmal RNAs migrate in a pattern distinct from each other and from bacteria in nondenaturing gels. The differences observed appear to be mainly due to conformational differences in the molecules. When examined under denaturing conditions in formamide gels, the 23S RNAs of all organisms examined comigrated. The 16S RNAs of mycoplasmas and acholeplasmas comigrated and appear to be of a smaller molecular weight than bacterial 16S RNA. It is suggested that the differences observed are indicative of significant phylogenetic divergence between mycoplasmas and bacteria or bacterial L forms.

Hollis et al. (J. Clin. Microbiol. 3:425-431, 1976) recently described an apparently new group of bacteria, with characteristics of the genus Vibrio, which was isolated from human blood, spinal fluid, and local infections of the extremities, and which caused fatal fulminant septicemia in at least 5 of 38 cases. The organisms were morphologically and biochemically similar to, but not identical with, both Vibrio alginolyticus and V. parahaemolyticus; they had different isolation sources from those of the latter two species, and they also fermented lactose and possessed different tolerances for sodium chloride in nutrient broth. This paper reports the results of deoxyribonucleic acid (DNA) reasociation experiments, which showed that DNA fragments from six strains of the newly described organisms, incubated at 60°C with DNA fragments from V. alginolyticus, V. cholerae, and V. parahaemolyticus, resulted in DNA duplex formation showing a level of relatedness consistent with that of different species within a genus. All strains of the newly described vibrio were highly related. Incubation at 75°C produced less than 10% relative duplex formation. When tested for heat stability, these duplexes had low temperatures of dissociation, indicating imperfectly matched DNA molecules. We thus concluded that the newly described vibrio is a species distinct from the other species studied.

Ten obligately anaerobic mycoplasma strains isolated from the rumen of sheep and cattle were grouped serologically by agglutination and gel diffusion precipitin tests. They could be placed into four serovars that fit well with group separations based upon cultural, biochemical, and biophysical properties. Hyperimmune sera prepared against a strain representing each serovar showed no cross-species interaction when tested with antigens of several known aerobic bovine mycoplasma and acholeplasma strains. Growth inhibition tests of Anaeroplasma strains confirmed the grouping by agglutination and gel diffusion precipitin tests.

Taxonomic data, obtained for 141 Enterobacteriaceae strains for which 240 unit characters were recorded, were subjected to numerical taxonomy analysis employing 36 coefficients. Clustering was by unweighted average linkage. From sorted similarity matrices, it was found that 15 coefficients, which included SSM, SH, STD, SJ, SNM, SO, SRT, SSHD, Sin−1(SSM), SP, Sø, SUN1, SUN4, SD, and SK2, provided useful discriminating properties. The coefficients SH and STD were found to provide results indistinguishable from SSM, and the coefficients SO and Sø yielded results very similar to those obtained with SSM coefficient.

Relatedness of pectobacteria pathogenic to corn, grass, and sugar cane, and of isolates of Pectobacterium chrysanthemi pathogenic for chrysanthemum, dahlia, and dieffenbachia was determined by deoxyribonucleic acid (DNA) reassociation. Heterologous DNA preparations were allowed to reassociate. Thermal elution chromatography was used to separate reassociated nucleotide sequences from unreacted sequences and to determine the thermal stability of related sequences. DNA relatedness data indicate that corn stalk rot bacteria and strains pathogenic on grass are highly interrelated. Furthermore, strains of both types are distinct from Pectobacterium carotovorum and are most closely related to P. chrysanthemi. P. chrysanthemi strains fall into two relatedness groups, one of which contains isolates from chrysanthemum and guayule and the other isolates from dahlia or dieffenbachia. A sugar cane pathogen is 65% related to both the corn stalk rot bacteria and to strains of P. chrysanthemi. It is suggested that all of these organisms be considered as P. chrysanthemi and that the corn stalk rot and grass isolates be designated P. chrysanthemi pathovar zeae.

We prepared hybrids between 14C-labeled ribosomal ribonucleic acid (rRNA) from either Agrobacterium tumefaciens ICPB TT111 or A. rhizogenes ICPB TR7, and deoxyribonucleic acid (DNA) from a great variety of reference gramnegative and gram-positive bacteria. Each hybrid was described by (i) its T m(e) the temperature at which 50% of the hybrid was denatured, and (ii) percentage of rRNA binding, i.e., micrograms of 14C-labeled rRNA duplexed per 100 μg of filter-fixed DNA. Each taxon occupied a definite area on the rRNA similarity map. The size and shape of this area depended on the phenotypic and genetic heterogeneity of the taxon. There appeared to be a correlation between Tm(e) of the heterologous hybrids and the overall phenotypic similarities of the organisms and taxa involved. Tm(e) values above 65°C were taxonomically most meaningful. DNA: rRNA hybridizations condensed all strains from a genus in one narrow cluster; the method had little resolution to distinguish species within a genus, but it seemed to be a very useful approach to detect remote relationships at the inter- and suprageneric level, for taxonomic and identification purposes. The hybrid parameters of Azotomonas fluorescens, Mycoplana bullata, Mycoplana dimorpha, Phyllobacterium, two misnamed “Chromobacterium lividum” strains from leaf-nodulating plants, two misnamed agrobacteria from the Baltic Sea, and a few misnamed “Achromobacter” strains were all in the vicinity of Agrobacterium and Rhizobium. We suggest that all of these organisms are remote relatives and belong in the family of the Rhizobiaceae. Azotomonas insolita NCIB 9749 is misnamed; it is an Agrobacterium. Several organisms which had been misnamed Agrobacterium formed DNA: rRNA hybrids with properties outside the Agrobacterium area.

Strains of a new type of group III nonphotochromogenic mycobacteria have been repeatedly isolated from four patients with clinical and roentgenological signs of lung mycobacteriosis. These strains split nicotinamide and pyrazinamide as do those of Mycobacterium avium, but they show no esterase activity. Furthermore, the strains produce thermolabile catalase and hydrolyze Tween 80. They also have a unique lipid pattern and special sensitins and agglutinins. These strains, therefore, are considered as belonging to a new species of pathogenic, nonphotochromogenic mycobacteria, for which we propose the name Mycobacterium malmoense. Strain M-Ö 816 is the type strain, and it has been deposited in the American Type Culture Collection under the number 29571.

A new genus and species of obligately methylotrophic bacteria are described. These bacteria are nonmotile, gram-negative rods occurring singly and in pairs. Only methanol and methylamine can support growth. Formaldehyde fixation occurs mainly via the 3-hexulose phosphate pathway, and cell extracts contain a glutathione-independent, nicotinamide adenine dinucleotide-linked formaldehyde dehydrogenase. The deoxyribonucleic acid base composition is 54.1 mol% guanine plus cytosine. Nitrogen-limited cells accumulate over 5% of their dry weight as a glycogen-like reserve material. This polysaccharide is a homoglucan which is similar to glycogen in its iodine-staining properties and its degree of degradation by phosphorylase a. Some of the glucose molecules are α-1,4 linked, and the presence of other types of bonds in the glucan is implied. The name of the genus proposed for these bacteria is Methylobacillus gen. nov. The name of the type species, Methylobacillus glycogenes sp. nov., refers to the ability of this species to form a glycogen-like reserve material. The type strain of M. glycogenes is T-11 (= ATCC 29475).

The 46 strains of Bacillus firmus and Bacillus lentus in our collection were found to share a group of 17 physiological properties that distinguished them from morphologically similar strains of other species of the genus Bacillus. Thirteen other characteristics separated the 46 strains into one group of 20 similar to the nomenclatural type strain ofB. firmus (ATCC strain 14575) and a second group of eight strains resembling the nomenclatural type strain of B. lentus (ATCC strain 10840). The pattern of these 13 additional characteristics of the remaining 18 of the 46 strains fell between the two groups. Guanine plus cytosine content in the deoxyribonucleic acid of 31 of the 46 strains ranged from 39.0 to 48.3 mol% and did not allow separation of the strains into distinct groups. The 46 strains of our collection could not, therefore, be cleanly divided by the properties tested and appeared as a spectrum, or series, of strains.

Fifty-seven clinical isolates of previously unidentified gram-positive, fermentative, nonsporeforming rods were studied and compared to the type strains of Oerskovia turbata and O. xanthineolytica. Thirty-five of the isolates were identified as Oerskovia species: 9 were identified as O. turbata, and 26 were identified as O. xanthineolytica. The Oerskovia cultures could be differentiated from the other isolates on the basis of the development of filamentous colonies.

The strains CP3 and CP4, isolated from fermenting sugar-cane juice in Brazil, and the strain Ag11, isolated from fermenting agave juice in Mexico, belong in Zymomonas mobilis subsp. mobilis. The existence of a separate Zymomonas mobilis subsp. recifensis for the former two is not justified. The strains NCIB 8777 and NCIB 10565 from sick cider and apple pulp, respectively, in England, belong in Zymomonas mobilis subsp. pomaceae. Protein electropherograms allow a quick and accurate identification in Zymomonas.

The recommendation to redefine the genus Herpetosiphon is disputed.

ATCC 15749, a hydrogen bacterium placed in Hydrogenomonas ruhlandii by Packer and Vishniac (1955) and then transferred to Pseudomonas by Davis et al. (1969), is peritrichous and belongs in the genus Alcaligenes. ATCC 15749 is the type strain of H. ruhlandii Packer and Vishniac, and by reason of its transfer to the genus Alcaligenes, the name of the species becomes Alcaligenes ruhlandii (Packer and Vishniac) comb. nov. It differs from Alcaligenes eutrophus by lacking the ability both to use aromatic compounds and to denitrify and by possessing broad, sheathed flagella.

Twelve Streptomyces species having hairy spores were examined by transmission electron microscopy and scanning electron microscopy. Each species was assigned to one of the three previously proposed subgroups of the hairy-spored group.

Mycobactins and exochelins have been isolated from four strains of Mycobacterium avium indicating this species to be unexceptional in the production of compounds involved in iron assimilation.

Improvements are presented and procedural details are clarified concerning the melanin formation test of streptomycetes of Arai and Mikami (1972).