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Multiple-drug-resistant strains of Escherichia coli were isolated from the water at an estuarine site. They represented about 8.3% of the total E. coli population. Fifty-five strains, representing each of the 32 resistance patterns identified, were mated with an E. coli K-12 F- strain. Matings were performed on membrane filters, and the cells were washed to remove any colicins produced by the donors. Thirty-one strains, about 5% of the mean E. coli density in the samples, transferred drug resistance and, hence, posessed conjugative R plasmids. Of these, 80% transferred drug resistance at a frequency of about 10(-4) or less. Nine environmental R+ strains were mated with three fecal recipients. The R-plasmid transfer frequencies to the fecal strains from the environmental donors correlated well with those from a derepressed K-12 R+ laboratory donor. The R+ X K-12 F- lac- transconjugants from 16 environmental strains were "backcrossed" to a lac+ K-12 F- strain. All transfer frequencies were higher in the backcrosses than in the original matings from the environmental donor. Furthermore, 7 of 13 different transconjugants, which accepted plasmids at repressed frequencies of less than 10(-3), donated them at frequencies greater than 10(-2). This suggests that these were derepressed plasmids in a repressed host.