On the basis of ribosomal ribonucleic acid homologies, the genus Pseudomonas can be divided into at least five distinct groups, some of which are as distantly related t o each other as they are to Escherichia coli. One of these groups contains members of the genus Xanthomonas. The data presented support and extend the previous grouping based on deoxyribonucleic acid homologies and support the current view that the portion of the genome coding for ribosomal ribonucleic acid is more conserved in the course of evolution than the bulk of the genome.For about nine years our laboratory has been largely involved in taxonomic studies of the genus Pseudomonas. Our first approach was the phenotypic characterization of a large number of strains using mainly nutritional characters. On the basis of phenotypic resemblance, we recognized several "species groups," comprised of what we regarded as recognizable species. Below the specific rank, we further recognized a number of "biotypes," some of which had been previously described as species, and many of which may deserve specific rank (14). Deoxyribonucleic acid (DNA)-DNA hybridization experiments generally supported our conclusions based on phenotypic data but showed that some of the species and biotypes were quite heterogeneous with respect to DNA homology. It was also found that some species groups were sufficiently closely related to each other t o be united in a larger DNA homology complex. For example, the "P. fluorescens complex" was found to contain not only the "fluorescent group" but also the nonfluorescent pseudomonads that had been assigned to the "alcaligenes" and "stutzeri" groups ( 10, 14). Our studies have been reviewed recently (9).To pursue further our studies on bacterial phylogeny and speciation, we have resorted to ribosomal ribonucleic acid (rRNA)-DNA hybridizations among selected members of the various DNA homology groups, and included three species of Xanthomonas in the studies. The results are reported in the present paper. MATERIALS AND METHODSBacterial strains. The bacterial strains used in the presently described experiments are numbered as in our previous publications (1,2,4,7,8,(10)(11)(12)14). Reparation of rRNA. For the preparation of unlabeled rRNA, most strains were grown in a medium containing 0.033 M K-Na phosphate buffer, pH 6.8; (NH,), SO,, 0.1%; asparagine, 0.2%; yeast extract, 0.5%; Hutner mineral base (3), 10 ml per liter. For the growth of P. rnaltophilia, DL-sodium lactate was substituted for asparagine. P. saccharophila was grown in a medium containing 0.033 M K-Na phosphate buffer, pH 6.8; NH,Cl, 0.1%; MgSO, 07 H,O, 0.05%; ferric ammonium citrate, 0.005%; CaCl, , 0.0005%; and sodium succinate, 0.2%. For the growth of P. diminuta, the same basal medium was used as for P. saccharophila except that succinate was replaced by asparagine and the essential growth factors (1) were added. The cultures were grown at 30 C on a rotary shaker. The cells were suspended in 0.02 M tris-(hydroxymethy1)aminomethane (Tris)-hydrochloride buffer, pH 7....

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Cells of Saccharomyces cerevsiae with mutations in the RAD52 gene have previously been shown to be defective in meiotic and mitotic recombination, in sporulation, and in repair of radiation-induced damage to DNA. In this study we show that diploid cells homozygous for rad52 lose chromosomes at high frequencies and that these frequencies of loss can be increased dramatically by exposure of these cells to x-rays. Genetic analyses of survivors of x-ray treatment demonstrate that chromosome loss events result in the conversion of diploid cells to cells with nearhaploid chromosome numbers.The life cycle ofheterothallic strains ofthe yeast Saccharomyces cerevisiae normally alternates between haplophase and diplophase. Cells in these euploid states contain one or two sets, respectively, of the 17 chromosomes that make up the yeast genome. However, departures from euploidy (i.e., aneuploidy) are found in both haploid and diploid cultures. In this article we describe another mutation, rad52-1, that causes chromosome loss or nondisjunction events at very high frequencies. This is particularly interesting because the rad52 mutation has also been shown to cause cells to be sensitive to ionizing radiation and to block mitotic and meiotic recombination (7-9), and thus presumably to affect DNA directly. MATERIALS AND METHODSThe genotypes of the five diploid yeast strains used in these experiments are described in Table 1. These diploids are all closely related; three (XS118, XS170, and STX135) have a common parent and the other two (STX140 and STX141) were the products of crosses between meiotic descendants of STX135. XS118, STX140, and STX141 are homozygous for radS2. STX135 is heterozygous at this locus and XS170 is homozygous for the wild-type allele of this gene. All five diploids are heterozygous at several other loci and all carry at least one chromosome with markers in coupling on opposite sides of the centromere. All strains were derived from strains available in the Yeast Genetic Stock Center, University ofCalifornia, Berkeley, except for strain 260-1-3, MATa met2, which was kindly supplied by Michael Unger.Standard procedures for the construction and analysis of the genetic crosses and mitotic variants were followed (11, 12). The recipes used for the various media have been described (13); the x-ray source has also been described (14). RESULTSExpression of Recessive Phenotypes in Strain XS118. The diploid strain XS118 was constructed as one ofthe control strains in our studies involving the cloning of the RAD52 gene (unpublished). We observed that the plating efficiency of this strain, which is homozygous for the rad52 mutation, was low and that the colonies grew slowly and were irregular in shape and size. These irregularities were observed to be more pronounced in colonies that had developed from cells exposed to x-rays. In addition, several of the colonies that had developed from both unirradiated and irradiated cells were wholly or partly red, indicating expression of the recessive ade2 mutation on chrom...

The relationships among 93 strains of Pseudomonas fluorescens were investigated by (1) a numerical taxonomic analysis on the results of 150 phenotypic tests, (2) DNA hybridization studies using 16 reference strains, (3) quantitative microcomplement fixation studies using six reference strains with antibodies directed against the protein azurin. In general, the strains fell into distinct clusters. Assignment to these clusters on the basis of azurin immunological similarity showed 98% agreement with assignment based on DNA homology, suggesting that many genes will follow the same pattern. Of the strains that clustered on the basis of genotype (DNA, azurin) 88% also clustered on the basis of phenotype. The occasional noncongruency observed between the genotypic and phenotypic data may be due to the variable rates of phenotypic evolution. These results provide a perspective on the roles of horizontal and vertical transfer of genes in the evolution of this bacterial group.

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In studies reported previously (Doudoroff Palleroni, MacGee, and Ohara, 1956) it has been shown that cell-free preparations of a mutant

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