ABSTRACT
A DNA clone from Rhodococcus equi conferring low-level rifampin resistance through the ability to inactivate this antibiotic via its decomposition was identified. The iri (inactivation of rifampin) gene consisted of an open reading frame of 1,437 bp encoding a 479-amino-acid sequence strongly resembling those of monooxygenases acting upon phenolic compounds or involved in polyketide antibiotic synthesis. When expressed in Escherichia coli, the gene conferred resistance to a > 50-micrograms/ml concentration of the drug.
Subject(s)
Antibiotics, Antitubercular/pharmacology , Genes, Bacterial/genetics , Mixed Function Oxygenases/genetics , Rhodococcus equi/genetics , Rifampin/pharmacology , Amino Acid Sequence , Base Sequence , DNA, Bacterial/genetics , Drug Resistance, Microbial , Escherichia coli/drug effects , Escherichia coli/genetics , Molecular Sequence Data , Rhodococcus equi/drug effects , Sequence Homology, Amino AcidABSTRACT
Genes essential for Rhodococcus chromosomal conjugation were found to be located on an unstable genetic element in one of the strains investigated. From this element two segments of DNA could be identified which were involved in conjugation of Rhodococcus strains. One region, spread over 8 kb, was involved in the property of self-incompatibility. A second region, of about 3 kb, was essential for conjugation between ATCC 12674 and related nocardioforms.
Subject(s)
Chromosomes, Bacterial/physiology , Conjugation, Genetic/physiology , DNA, Bacterial/genetics , Plasmids/genetics , Rhodococcus/genetics , Drug Resistance, Microbial , Genetic Markers , Mutagenesis , Restriction MappingABSTRACT
Azo dyes are recalcitrant pollutants. Two sulfonated azo dyes, Orange II and Amido black, are effectively decolorized by certain nocardioform strains of the genus Rhodococcus. A mutant of one of these strains was isolated which had lost azo-dye decolorizing ability and the strain was used to clone DNA conferring this ability, by screening a BclI library constructed from DNA of a decolorizing strain. The relevant genetic information was located on a 6.3-kb fragment of DNA.
Subject(s)
Amido Black/metabolism , Azo Compounds/metabolism , Benzenesulfonates/metabolism , DNA, Bacterial/genetics , Rhodococcus/genetics , Rhodococcus/metabolism , Biodegradation, Environmental , Cloning, Molecular , Coloring Agents/metabolism , Mutagenesis , NADH, NADPH Oxidoreductases/metabolism , Nitroreductases , Restriction MappingABSTRACT
One of a number of large nocardioform plasmids previously obtained by a primarily genetic approach was reduced in size to about approximately 11 kb. This smaller plasmid possessed determinants for resistance to sodium arsenate and sodium arsenite, as well as immunity to nocardiophage Q4. It was joined to an Escherichia coli-positive selection vector constructed by M. Zabeau and colleagues, which had the EcoR1 endonuclease gene placed under the control of the PR promoter of lambda as well as a bla determinant. The resulting shuttle vector of about 14.6 kb was maintained in E. coli and in several strains of Rhodococcus. The vector was efficient in cloning DNA without prior alkaline phosphatase treatment, as a result of the presence of the positive selection function. This function was not significantly expressed in Rhodococcus, and the presence of the nocardioform resistance determinants led to no increase in arsenate or arsenite resistance in E. coli. The presence of the bla gene resulted in an increase of about threefold in ampicillin resistance in Rhodococcus strains.
