R factors were detected in 3.3% of 233 hospital isolates of Pseudomonas aeruginosa using P. aeruginosa recipients in conjugations. Transferred markers included streptomycin, tetracycline, and sulfonamide resistance. Gentamicin resistance was transferred from two strains previously shown to acetylate gentamicin. A group of R factors exemplified by R931 were characterized by failure to transfer to Escherichia coli recipients. Such R factors formed a single compatibility group when examined in a P. aeruginosa recipient. Other P. aeruginosa R factors, including RP4, showed stable coexistence with the R931 group. It is proposed that RP4 and similar R factors be members of the P-1 compatibility group and that R931, R3108, R209, and R130 be members of a group termed P-2. The buoyant densities of all R factors examined were similar, about 1.716 to 1.719 g/cm3. The content of R-factor deoxyribonucleic acid (DNA) relative to the total DNA varied among the different R factors, ranging from about 18 ± 2% in logphase cells of 931 (R931) to undetectable for 679 (R679). However, R679, which transferred from strain 679 at extremely low and irregular frequencies to an E. coli host, was shown to represent about 4% R-factor DNA in that host. The relative DNA content of R931 appeared to decline in the stationary growth phase of 931 (R931) or 280 (R931). R931 covalently closed circular DNA was isolated by ethidium bromide-CsCl gradient centrifugation and examined by electron microscopy. Two major molecular distributions existed, having contour lengths of 0.5 and 12.4 ,um. The molecular weights were estimated to be 106 and 25 x 106. Both molecules were under relaxed replication control. R factor R931 exists as a naturally occurring high-frequency transfer system in P. aeruginosa strains 931 and 1310. However, in strain 280 it acts as if subject to fertility repression. Other members of the P-2 compatibility group also are high-frequency transfer systems in the natural host and in strain 1310. RP4 is restricted from recipient strain 1310. Some additional recipient effects were noted in that strains 1310 or 280 sometimes differed in recipient effectiveness with a given donor. Agglutination reactions with absorbed antiserum were able to distinguish between two members of the same R-factor compatibility group, R931 and R3108.Previous evidence (5) strongly suggested that at least some Pseudomonas R factors, including R931, are host-restricted. Such R factors contrast with other Pseudomonas R factors, described by several workers (11,14,17,27,31), which seem to be freely transferable between P. aeruginosa and members of the Enterobacteriaceae. Grinsted et al. (17) and Datta et al. (11) examined some of the properties of the latter R factors, but the former have not been described in detail. The work in this paper describes some of the characteristics of partially or completely host-restricted Pseudomonas R factors. MATERIALS AND METHODSOrganisms. All P. aeruginosa strains were originally obtained from clinical specimens and identified a...