The taxonomic position of Peptococcus saccharo-l~epticus is'uncertain [1 ]. The exclusion from the genus tococcus, due to its capacity to ferment sugars [1 ], was justified by results of Schleifer and Nimmermann [2] who showed that the cell wall composition of P. saceharolyticus was characteristic of staphylococci [3,4] ; a finding that may indicate a certain relationship between these two groups of bacteria. The aim of the present study was to determine the genetic relationship ofP. saccharolyticus to other anaerobic and facultative anaerobic Gram.positive cocci usi/ag DNA-rRNA and DNA-DNA hybridization techniques. It could be dearly demonstrated that P. saccharolyticus is genetically related to staphylococci.
Materials and methods
Organisms and growth conditionsThe strains used are listed in Table 1. Apart from anaerobic and facultative anaerobic cocci, Lactobacillus acidophilus was included as an unrelated reference strain with a similar DNA base composition. Staphylococci were cultivated aerobically in peptone-yeast extract-glucose-NaC1 bouillon [5]. All the other strains, except for L. acidophilus and Pediococcus pentosaceus, which were grown in MRS medium [6], were cultivated anaerobically in a simplified PYG medium [7]. Glucose in this medium was replaced by glycine or glutamine for cultivation ofP. variabilis and P. aerogenes. The incubation temperature was 37°C. Ceils were harvested in the late logarithmic growth phase. In order to label DNA or rRNA, the bacteria were grown in broth, in which yeast-extract was replaced by 2/aCi/ml of [6-3H] thymidine and [5,6-3H]uracil, respectively. Specific activities obtained by this method were in the range of 1 • 104-7 • l0 s for [3H]DNA and of 1 • 104-1 • l0 s for [3H]-rRNA.
DNA isolation and re-associationExtraction of labelled and unlabelled DNA was carried out using Marmur's method [8], with modifications described by Meyer and Sehleifer [9,10]. Fixation of unlabelled DNA on nitrocellulose filters was performed according to the method of Gillespie and Spiegelmann [11 ]. DNA-DNA hybridization experi. ments were accomplished following the procedure described by McConaughy et al. [12] in 3 X SSC (Standard Saline Citrate: 0.15 M NaC1, 0.015 M sodium citrate, pH 7.0) containing 10% formamide at 58°C for 30 h. At an average G + C-content of 34 mol%, these conditions correspond to a temperature of 25°C below the thermal melting point of the DNA [13]. The ratio of filter-bound DNA to labelled, frag. mented, DNA in solution was approx. 75 : 1.The percentage of radio-activity bound in the heterologous reaction was normalized to the percentage of radio-activity bound in the homologous reaction and expressed as an homology value.
rRNA isolationUptake of [5,6-all]uracil into cells was followed by measuring the decrease of radio-activity in the