Deoxyribonucleic Acid Homologies Among Some Pseudomonas Species

Phylogenetic relationships among a number of strains belonging to the genus Pseudomonas were explored by the use of in vitro deoxyribonucleic acid (DNA) hybridization. The fluorescent nomenspecies (P. fluorescens, P. putida, P. aeruginosa, P. cichorii, P. syringae, and related species), as well as the nonfluorescent species P. stutzeri, P. mendocina, P. alcaligenes, and P. pseudoalcaligenes, were shown to belong to a single DNA homology complex which is isolated from other Pseudomonas species that have been studied [P. cepacia (= P. multivorans), P. caryophylli, P. marginata (= P. alliicola), P. pseudomallei, P. acidovorans, P. testosteroni, P. solanacearum, P. diminuta, P. facilis, P. delafieldii, P. saccharophila, P. palleronii]. A limited numerical analysis of the phenotypic properties of the examined strains supported, with some exceptions, their previous allocation to nomenspecies and biotypes. The internal structure and nomenclature of the "P. fluorescens homology complex" are discussed.     

Deoxyribonucleic Acid Homologies Among Species of the Genus Neisseria

Eleven aerobic species of Neisseria, a Mima sp., and a Herellea sp. were tested for deoxyribonucleic acid (DNA) homology in direct hybridization experiments. DNA labeled with either (14)C or (32)P was prepared from five species of Neisseria. Unlabeled DNA from the various microorganisms was immobilized on membrane filters, which, after pretreatment, were incubated with labeled DNA (4,000 counts per min per filter) for 14 hr at 67 C. The measure of relatedness was expressed as the relative percentage of direct binding compared to that obtained with homologous DNA. All serological types of N. meningitidis, including the newly proposed types, were homologous to the standard strain of N. meningitidis with one possible exception, type Z. The genus Neisseria is heterogeneous in nature, forming at least three distinct groups: first, N. meningitidis and N. gonorrhoeae; second, N. perflava, N. subflava, N. sicca, N. flavescens, and N. flava; third, N. catarrhalis and N. caviae. Mima and Herellea species show no significant homology with the Neisseria.     

Deoxyribonucleic Acid Base Sequence Homologies of Some Budding and Prosthecate Bacteria

The genetic relatedness of a number of budding and prosthecate bacteria was determined by deoxyribonucleic acid (DNA) homology experiments of the direct binding type. Strains of Hyphomicrobium sp. isolated from aquatic habitats were found to have relatedness values ranging from 9 to 70% with strain "EA-617," a subculture of the Hyphomicrobium isolated by Mevius from river water. Strains obtained from soil enrichments had lower values with EA-617, ranging from 3 to 5%. Very little or no homology was detected between the amino acid-utilizing strain Hyphomicrobium neptunium and other Hyphomicrobium strains, although significant homology was observed with the two Hyphomonas strains examined. No homology could be detected between prosthecate bacteria of the genera Rhodomicrobium, Prosthecomicrobium, Ancalomicrobium, or Caulobacter, and Hyphomicrobium strain EA-617 or H. neptunium LE-670. The grouping of Hyphomicrobium strains by their relatedness values agrees well with a grouping according to the base composition of their DNA species. It is concluded that bacteria possessing cellular extensions represent a widely diverse group of organisms.     

Phenotypic Characterization and Deoxyribonucleic Acid Homologies of Pseudomonas solanacearum

Twenty-six strains and colony variants of Pseudomonas solanacearum belonging to four described biotypes were characterized, by using 169 phenotypic characters previously found useful in distinguishing among strains of other Pseudomonas species. Deoxyribonucleic acid (DNA) hybridization (intra- and interspecific DNA-DNA hybridizations) was performed by using the in vitro "DNA competition" technique. P. solanacearum appears to be a moderately homogeneous species, which is, at most, only remotely related to all other species of the genus studied to date. The four biotypes are not clearly distinct from one another with respect to nutritional characters or DNA homologies. Discrepancies between acid production and growth with some carbohydrates were noted. Difficulties were encountered in certain DNA competition experiments and some problems of the methodology are discussed.     

Bacterial Genomes: Habitat Specificity and Uncharted Organisms

The capability and speed in generating genomic data have increased profoundly since the release of the draft human genome in 2000. Additionally, sequencing costs have continued to plummet as the next generation of highly efficient sequencing technologies (next-generation sequencing) became available and commercial facilities promote market competition. However, new challenges have emerged as researchers attempt to efficiently process the massive amounts of sequence data being generated. First, the described genome sequences are unequally distributed among the branches of bacterial life and, second, bacterial pan-genomes are often not considered when setting aims for sequencing projects. Here, we propose that scientists should be concerned with attaining an improved equal representation of most of the bacterial tree of life organisms, at the genomic level. Moreover, they should take into account the natural variation that is often observed within bacterial species and the role of the often changing surrounding environment and natural selection pressures, which is central to bacterial speciation and genome evolution. Not only will such efforts contribute to our overall understanding of the microbial diversity extant in ecosystems as well as the structuring of the extant genomes, but they will also facilitate the development of better methods for (meta)genome annotation.     

Nucleic Acid Homologies Among Species of Saccharomyces

Evolutionary divergence among species of the yeast genus Saccharomyces was estimated from measurements of deoxyribonucleic acid (DNA)/DNA and ribosomal ribonucleic acid (RNA)/DNA homology. Much diversity was found in the DNA base sequences with several species showing little or no homology to the three reference species, S. cerevisiae, S. lactis, and S. fragilis. These three reference species also showed little or no homology to each other. On the other hand the diversity among ribosomal RNA base sequences was small since most species showed a high degree of homology to the reference species. The arrangement of species based on ribosomal RNA homologies agrees in most cases with current taxonomic groupings. A yeast hybrid (S. fragilis x S. lactis) was shown to contain two nonhomologous genomes. A minimum genome size of 9.2 x 10(9) daltons for S. cerevisiae was calculated from the rate of DNA renaturation.     

Nucleic Acid Homologies Among Oxidase-Negative Moraxella Species

The deoxyribonucleic acid (DNA) base composition and DNA homologies of more than 40 strains of oxidase-negative Moraxella species were determined. These bacteria have also been identified as belonging to the Mima-Herellea-Acinetobacter group and the Bacterium anitratum group, as well as to several other genera including Achromobacter and Alcaligenes. The DNA base content of these strains ranged from 40 to 46% guanine plus cytosine. DNA-DNA competition experiments distinguished five groups whose members were determined by showing 50% or more homology to one of the reference strains: B. anitratum type B5W, Achromobacter haemolyticus var. haemolyticus, Alcaligenes haemolysans, Achromobacter metalcaligenes, and Moraxella lwoffi. A sixth group comprised those strains showing less than 50% homology to any of the reference strains. Negligible homology was found between strains of oxidase-negative and oxidase-positive Moraxella species in DNA-DNA competition experiments. However, evidence of a distant relationship between the two groups was obtained in competition experiments by using ribosomal ribonucleic acid.     

Homologies of Deoxyribonucleic Acids from Brucella ovis, Canine Abortion Organisms, and other Brucella Species

The bacterium that causes canine abortion has polynucleotide sequences similar, in deoxyribonucleic acid (DNA)-DNA homology studies, to those of Brucella suis and, by inference from previous data, those of B. abortus and B. melitensis as well as B. neotomae. Therefore, the organism causing canine abortion appears to be a member of the genus Brucella. DNA preparations from Serratia marcescens, Alcaligenes faecalis, and Bordetella bronchiseptica, 58, 62, and 66 mole% guanine plus cytosine, respectively, do not have detectable polynucleotide sequence homologies with B. suis DNA which is 56 mole% guanine plus cytosine. B. ovis DNA lacks some of the polynucleotide sequences present in B. suis DNA and appears to be a deletion mutant. However, a large proportion of B. ovis polynucleotides are similar to those of other Brucella species, which supports the inclusion of B. ovis in the genus.     

Deoxyribonucleic Acid Homology Among Lactic Streptococci

A comparison was made by deoxyribonucleic acid homology of 45 strains of lactic streptococci, using two strains of Streptococcus cremoris and three strains of Streptococcus lactis as reference strains. All S. cremoris strains were grouped together by deoxyribonucleic acid homology. S. lactis strains formed a second group, except that three strains of S. lactis showed a high degree of homology with S. cremoris strains. The three Streptococcus diacetylactis strains could not be differentiated from S. lactis strains. In spite of these differences between S. lactis and S. cremoris strains, the majority of S. cremoris, S. lactis, and S. diacetylactis strains studied had at least 50% of their base sequences in common. In contrast, Streptococcus thermophilus strains generally showed little relationship with the other strains of lactic streptococci. The relevance of these findings to the selection of starter strains for cheese making is discussed.     

Polynucleotide Homologies of Brucella Deoxyribonucleic Acids

Deoxyribonucleic acids (DNA's) extracted from organisms presently placed in the genus Brucella (B. abortus, B. melitensis, B. neotomae, and B. suis) possessed very similar polynucleotide sequences. Unlabeled, single-stranded DNA fragments from B. abortus, B. melitensis, B. neotomae, and B. suis were equally effective in competing with the interaction of corresponding radiolabeled, single-stranded DNA fragments with their homologous DNA-agars. Unlabeled fragments of B. ovis, however, did not compete as effectively as the homologous, unlabeled DNA's, and this organism, therefore, had a detectably different polynucleotide composition. The mole percentages of guanine plus cytosine in Brucella DNA's (56 to 58%) were also similar. DNA's from Francisella tularensis, Escherichia coli, and the slow loris did not compete.     

Deoxyribonucleic Acid Relationships Among Marine Vibrios

The deoxyribonucleic acid (DNA) relationships of 80 strains identified either as Vibrio parahaemolyticus, V. alginolyticus, and V. anguillarum, or as allied marine vibrios were delineated by DNA-DNA competition experiments as well as by measuring the thermal stabilities of the DNA-DNA duplexes formed in direct binding studies. The tested strains included isolates from Japan, Europe, and the United States. The V. parahaemolyticus and V. alginolyticus groups showed an average of 67% homology to one another and 30% to strains of V. anguillarum. Significantly, a number of the isolates from the Pacific Northwest which had been previously identified as V. parahaemolyticus based on morphological, biochemical, and serological evidence were shown either to be strains of V. anguillarum or to belong to as yet unnamed groups. Most strains isolated from diseased salmon in the Pacific Northwest proved to be virtually identical with V. anguillarum type C by DNA homology experiments, thereby differentiating them from similar strains isolated from diseased herring and occasionally from salmon. The latter Pacific Northwest isolates fell into two distinct genotypic groups. A plot of the per cent homology by competition versus the difference in the thermal stabilities of heterologous and homologous duplexes (DeltaT(m,e)) between the same DNA species shows a linear decline in homology of 4.25% per degree of DeltaT(m,e). The use of this relationship for estimating the percentage of the mispaired bases distinguishing DNA preparations directly from competition experiments is discussed.     

Serological Relatedness of Bacterial Deoxyribonucleic Acid Polymerases

A number of bacterial species have been surveyed for serological activities with antiserum to Escherichia coli B deoxyribonucleic acid (DNA) polymerase I (EC 2.7.7.7.). The degree of serological cross-reaction is taken as a measure of relatedness of both the enzyme molecules from various species and the bacterial species themselves. Extracts were assayed by complement fixation only after treatment with deoxyribonuclease, since DNA bound to DNA polymerase alters the serological activity of the enzyme. Antiserum to E. coli DNA polymerase I did not react with either purified E. coli DNA polymerase II or the phage T4-induced DNA polymerase.     

Deoxyribonucleic Acid Homology Among the Fruiting Myxobacteria

Deoxyribonucleic acid similarities were determined by competition experiments. The several strains of fruiting myxobacteria tested showed from 23 to 89% homology with the reference strains Myxococcus xanthus (FB) and M. fulvus (M6).     

Inhibition of Bacterial Conjugation by Ribonucleic Acid and Deoxyribonucleic Acid Male-Specific Bacteriophages

Both deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) male-specific phages, with an F-specific host range, inhibited the bacterial mating process of Escherichia coli. DNA phages prevented the formation of mating pairs but had no effect on mating pairs once they were formed. A step in RNA phage infection, prior to RNA penetration, prevented the formation of mating pairs and, in addition, prevented a fraction of existing mating pairs from completing the mating process. These findings are compatible with the hypothesis that donor cells have a single surface structure involved in both conjugation and male-phage adsorption and that this element is the F pilus.     

Association of Replicative T4 Deoxyribonucleic Acid and Bacterial Membranes

Experiments utilizing CsCl density gradient analysis and radioactive labels specific for bacteriophage T4 deoxyribonucleic acid (DNA) and membranes have shown that replicative T4 DNA is associated with host membranes. The association is inhibited by chloramphenicol and takes place just prior to semi-conservative replication of the phage DNA.     

CISA: Contig Integrator for Sequence Assembly of Bacterial Genomes

A plethora of algorithmic assemblers have been proposed for the de novo assembly of genomes, however, no individual assembler guarantees the optimal assembly for diverse species. Optimizing various parameters in an assembler is often performed in order to generate the most optimal assembly. However, few efforts have been pursued to take advantage of multiple assemblies to yield an assembly of high accuracy. In this study, we employ various state-of-the-art assemblers to generate different sets of contigs for bacterial genomes. A tool, named CISA, has been developed to integrate the assemblies into a hybrid set of contigs, resulting in assemblies of superior contiguity and accuracy, compared with the assemblies generated by the state-of-the-art assemblers and the hybrid assemblies merged by existing tools. This tool is implemented in Python and requires MUMmer and BLAST+ to be installed on the local machine. The source code of CISA and examples of its use are available at http://sb.nhri.org.tw/CISA/.     

Deoxyribonucleic Acid Homologies of Some So-Called “Hydrogenomonas” Species

Evidence based on deoxyribonucleic acid homology supports the abandonment of the genus Hydrogenomonas. Pseudomonas facilis (formerly Hydrogenomonas facilis) is closely related to the nonautotrophic species P. delafieldii. P. facilis and Alcaligenes eutrophus (often called H. eutropha) are not related to each other or to other hydrogen bacteria and pseudomonads studied.     

Deoxyribonucleic Acid Synthesis and Deoxynucleotide Metabolism During Bacterial Spore Germination

Deoxyribonucleic acid (DNA) synthesis during germination of Bacillus megaterium spores takes place in two stages. In stage I (0-55 min) DNA synthesis is slow and there is no detectable net synthesis, whereas in stage II (from 55 min on) the rate of synthesis is much faster and net DNA synthesis occurs. Deoxyribonucleotide pool sizes match the rates of DNA synthesis in stages I and II. The level of deoxyribonucleotide triphosphates is not correlated with the level of deoxyribonucleotide kinases, but rather with that of ribonucleotide reductase activity.     

Relationships Among Mycobacteria and Nocardiae Based upon Deoxyribonucleic Acid Reassociation

Chorismate mutase from Streptomyces aureofaciens was purified 12-fold. This enzyme preparation did not show any activity when tested for anthranilate synthetase, prephenate dehydrogenase, or prephenate dehydratase. The catalytic activity of chorismate mutase has a broad optimum between pH 7 and 8. The initial velocity data followed regular Michaelis-Menten kinetics with a K(m) of 5.3 x 10(-4) M, and the molecular weight of the enzyme was determined by sucrose gradient centrifugation to be 50,000. Heat inactivation of chorismate mutase, which occurs above temperatures of 60 C, is reversible. The enzyme activity can be restored even when chorismate mutase is treated at the temperature of a boiling-water bath for 15 min. Heat-denatured and renatured enzymes showed the same Michaelis constant and the same molecular weight as the native enzyme. l-Phenylalanine, l-tyrosine, l-tryptophan, and metabolites of the aromatic amino acid pathway were tested as potential modifiers of chorismate mutase activity. The activity of the enzyme was inhibited by none of these substances. Chorismate mutase of S. aureofaciens was not repressed in cells grown in minimal medium supplemented with l-phenylalanine, l-tyrosine, or l-tryptophan.     

Sequence Heterogeneity in Closed Simian Virus 40 Deoxyribonucleic Acid

The heteroduplex molecules formed by self-annealing of denatured, singly nicked simian virus 40 (SV40) deoxyribonucleic acid (DNA) prepared from closed viral DNA were examined by formamide-protein film electron microscopy to test the DNA for sequence homogeneity. Sequence inhomogeneity appears in the heteroduplexes as single-strand loops. These result from sequence deletion or from sequence substitution, if regions greater than 50 nucleotides are involved. The undenatured DNA from viruses passaged twice at multiplicities of infection much less than 1 plaque-forming unit (PFU) per cell appeared to be homogeneous in size. The heteroduplexes formed by this DNA indicated that approximately 2% of the molecules carried deletions, but that substitutions were below the level of detection. In contrast, undenatured DNA from viruses grown by passaging undiluted lysates seven times or by infection with stock virus at a multiplicity of infection of 5 PFU per cell contained a large frequency of molecules shorter than the full length. The heteroduplex samples indicated that 12 and 7% of the undenatured material contained base substitutions, and 13 and 11% contained deletions. The deletions and substitutions appear to occur in separate molecules. Length measurements on heteroduplexes displaying the loop characteristic of substitutions have established that these molecules are from true sequence substitutions, and not from adjacent or overlapping deletions. More than 80% of the molecules carrying substitutions are shorter than the native SV40 length. On the average, the substituted sequence is about 20% of the length of SV40, but it replaces a sequence about 30% of the native length. The substituted sequences may be host cell nuclear DNA, possibly arising from integration of SV40 into the chromosome followed by excision of the SV40 DNA together with chromosomal DNA.