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Thirteen bacteria have been isolated from nine different soil samples by selective enrichment culture on m-toluate (m-methylbenzoate) minimal medium. Eight of these were classified as Pseudomonas putida, one as a fluorescent Pseudomonas sp., and four as nonfluorescent Pseudomonas sp. All 13 strains appeared to carry TOL plasmids superficially similar to that previously described in P. putida mt-2 in that: (i) all the wild-type strains could utilize toluene, m-xylene, and p-xylene as sole carbon and energy sources, (ii) these growth substrates were metabolized through the corresponding alcohols and aldehydes to benzoate, m-toluate, and p-toluate, respectively, and thence by the divergent meta (or alpha-ketoacid) pathway, and (iii) the isolates could simultaneously and spontaneously lose their ability to utilize the hydrocarbons, alcohols, aldehydes, and acids, particularly during growth on benzoate, giving rise to cured strains which could grow only on benzaldehyde and benzoate of the aromatic substrates by the alternative ortho (or beta-ketoadipate) pathway. Eight of the isolates were able to transfer their TOL plasmids into their own cured strains, but only five were able to transfer them in interstrain conjugation into the cured strains, but only five were able to transfer them in interstrain conjugation into the cured derivative of P. putida mt-2. However, P. putida mt-2 was able to transfer its TOL plasmid into 11 of the cured isolates, and eight of these were able to retransmit this foreign plasmid in intrastrain conjugation with their own cured derivatives. Three of the isolates, MT 14, MT 15, and MT 20, differed significantly from the others in that the wild-type strains dissimilated the p-methyl-substituted substrates poorly, and also, during growth on benzoate, in addition to the cured derivatives, they gave rise to derivatives with a phenotype intermediate between the cured and wild-type strains, the biochemical and genetic nature of which has not been elucidated.

Ubiquity of plasmids in coding for toluene and xylene metabolism in soil bacteria: evidence for the existence of new TOL plasmids