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Plasmids having Escherichia coli ribosomal DNA sequences under control of a promoter for T7 RNA polymerase have been constructed. Transcription of the rDNA sequences is dependent on T7 RNA polymerase because the tandem promoters for E. coli RNA polymerase, normally used to direct transcription of these sequences, have been removed. The entire 16S, 23S and 5S coding sequences from the rrnB operon can be efficiently transcribed by T7 RNA polymerase in vitro to yield full‐length 30S precursor RNA. When such plasmids are placed into an E. coli strain containing a chromosomal copy of the gene for T7 RNA polymerase under control of the lac UV5 promoter, high‐level synthesis of rRNAs from the plasmid can be induced by adding IPTG to exponentially growing cells. Subsequent addition of rifampicin to inhibit further initiation of transcription by E. coli RNA polymerase provides a simple method to study the fate of plasmid‐coded rRNAs in the complete absence of host‐coded rRNA synthesis. Gel electrophoretic analysis demonstrated that the rRNAs synthesized by T7 RNA polymerase in the presence of rifampicin are processed to their mature forms and assembled into ribosomal particles for at least 35 min after rifampicin addition. T7 RNA polymerase is also capable of efficient transcription of the entire rrnB operon in the reverse direction.

The folic acid analog trimethoprim has been in clinical use for more than 10 years. The use of it in Sweden has doubled in the last 6 to 7 years, and from the distribution statistics it can be calculated that during 1 year 4 to 5% of the population in Sweden are given this drug. The The folic acid analog trimethoprim has been used as an antimicrobial agent for more than a decade. A few years after the introduction of trimethoprim for clinical use, R plasmid-mediated resistance to this drug was reported (13). This plasmid-borne resistance also demonstrated a new principle regarding bacterial insensitivity, in that a drug-resistant variation of the target enzyme, dihydrofolate reductase, was found to be expressed from the plasmid (1, 22). Several types of plasmid-borne, drug-resistant dihydrofolate reductases have been subsequently discerned, and at least one of them was shown to be located on a transposon, Tn7 (3, 4). In later years, trimethoprim has been used extensively for clinical purposes, and resistance to it seems to have increased in frequency (7,9,25).The use of trimethoprim in Sweden can be defined quantitatively by data from the computerized Swedish Drug Information System (Department of Drugs, National Board of Health and Welfare). From these data it could thus be seen that more than 0.15% of the population receive trimethoprim each day or that during 1 year 4 to 5% of the population are given this drug. It was interesting to investigate the types of bacterial resistance mechanisms that could be found in response to such a selection pressure in a relatively isolated population in northern Sweden (the county of Jamtland), in which one centrally located bacteriological laboratory serves the area. During an 8-month period, trimethoprim-resistant strains were collected from consecutive specimens of bacteria from the urinary tracts of patients. Among highly resistant bacteria, transposon Tn7 was found to be common, either located chromosomally or borne on a conjugative Incl plasmid, 50 kilobases (kb) of size, which seemed to be endemic to the area. Two cases of marked overproduction of chromosomal dihydrofolate reductase which caused high resistance to trimethoprim were also found. * Corresponding author. MATERIALS AND METHODSBacterial strains and plasmids. Laboratory strains of bacteria and plasmids are listed in Table 1. Clinically isolated enterobacterial strains are listed in Table 2, and their collection is described in the text. Testing for antibiotic resistance was performed by the agar diffusion method with antibiotics-containing paper disks from AB Biodisk, Solna, Sweden.The serotyping of Escherichia strains was kindly performed by I. and F. Orskov, International Escherichia and Klebsiella Centre, Copenhagen.Media. The rich medium LB or the mineral salts medium M9, supplemented with Casamino Acids (0.05% [wt/vol]), was used throughout (19).Transfer of R plasmids. The transfer of R plasmids was performed as described earlier (21) and also on agar plates, and recombinants were selected on M9 agar ...

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