Molecular and Genetic Properties of Plasmid R485 Conferring Resistance to Sulphonamides

Abstract

Plasmid R485 carrying a determinant of resistance to sulfonamides was isolated from rec+ and recA Escherichia coli hosts and its molecular properties were examined and compared with the characteristics of an incompatible plasmid, R6K, a type plasmid of incompatibility group X (IncX). Examination of R485 DNA by EM revealed monomer, dimer and miniplasmid molecules in preparations isolated from the rec+ host, while only monomers were seen in preparations obtained from the recA host. The molecular sizes of the R485 monomers from the rec+ and the recA host were 34.9 .+-. 3.7 and 38.3 .+-. 0.8 megadaltons (Mdal), respectively. The dimer molecules of R485 (59.7 .+-. 2.9 Mdal) probably resulted from the recombination between one monomer molecule of normal size and another from which the miniplasmid (11.2 Mdal) was segregated. The G + C content of R485 DNA was 44.3 mol%. With a minimum copy number of approximately 1/E. coli chromosome equivalent, plasmid R485 differs conspicuously from the multicopy plasmid R6K; this finding indicates that the replication of R485 and R6K is differently controlled. The presence of plasmid R485 in a cell brought about a reduction of the frequency of conjugative transfer of an IncP plasmid, RP1, and an IncN plasmid, N3A, but it did not affect the transfer of IncW plasmid Sa. Plasmid R485 showed integrative suppression of the expression of a chromosomal temperature-sensitive dnaA mutation, indicating that its replication is independent of the dnaA gene product. The simultaneous presence of incompatible plasmids R485 and R6K in a cell resulted in the elimination of R6K in the absence of selection pressure, while under selective conditions only a decrease in the number of R6K copies was found. Inhibition of growth of E. coli E52int6K by plasmid R485 at elevated temperature also indicated the negative effect of R485 on R6K replication.