Site-specific recombination promotes linkage between trimethoprim- and sulfonamide resistance genes. Sequence characterization of dhfrV and sulI and a recombination active locus of Tn21

Abstract

A new gene for trimethoprim resistance, dhfrV, found in several plasmid isolates with different characteristics, was sequenced and found to correspond to a peptide of 157 amino acids showing 75% similarity with the previously characterized, drug resistant dihydrofolate reductase of type I. The sequenced surroundings of dhfrV in plasmid pLMO20, were found to be almost identical with genetic areas surrounding resistance genes in transposon Tn21 and in R plasmid R388. The trimethoprim resistance genes of pLMO20 and R388 and the spectinomycin resistance gene of Tn21 could be regarded as having been inserted, by recombination, into an evolutionary older structure containing the sulfonamide resistance gene, sulI. The latter gene was sequenced and found to correspond to a peptide of 279 amino acids and with a molecular weight of 30,126 daltons. The inserted genes were found to be governed by a promoter situated in the highly conserved structure and also controlling expression of sulI. The insertion points of the different resistance genes were precisely defined, and at the 3' ends of the inserted genes inverted repeats allowing the formation of stem and loop structures were found. Similar structures were found at the 3' ends of the antibiotic resistance genes in Tn7, which could indicate similar recombination mechanisms to be effective in the evolutionary construction of all these different resistance elements.

Non‐palindromic attI sites of integrons are capable of site‐specific recombination with one another and with secondary targets

“…Integrons of three classes, each producing its own integrase, have now been defined by sequence (Arakawa et al, 1995;Sundströ m et al, 1988;Sundströ m and Skö ld, 1990;Recchia et al, 1995a). It is notable that the three integron types carry members of the same population of cassettes, which indicates cross-specificity.…”

“…In order to study recombination involving attI only, we modified the integron of plasmid R388, mediating trimethoprim and sulphonamide resistance (Swift et al, 1981;Sundströ m et al, 1988) to carry a single attI-type recombination site (see Experimental procedures and Fig. 1B).…”

“…Integrons are recombination systems, which capture and rearrange single genes borne on mobile sequences called cassettes (Cameron et al, 1986;Sundströ m et al, 1988;Recchia and Hall, 1995a). Most cassettes described in the literature mediate antibiotic resistance.…”

“…1A) is organized into a variable region composed of an arbitrary selection of cassettes and a conserved sequence (CS), which encodes a site-specific recombinase, IntI, of the lambda integrase family (Argos et al, 1986;Sundströ m et al, 1988). The IntI gene and the cassette-borne genes are divergently transcribed from opposite DNA strands.…”

“…Top: the integron in Tn21 with the aadA1 cassette mediating spectinomycin resistance. Middle: the integron in R388wt with the two mobile cassettes carrying dfr2b, mediating trimethoprim resistance and ORFA respectively (Sundströ m et al, 1988;Recchia and Hall, 1995a). Both integrons in Tn21 and R388 carry the immobilized cassette encoding an efflux protein, QacE, to the right.…”