New dfr2 gene as a single-gene cassette in a class 1 integron from a trimethoprim-resistant Escherichia coli isolate

Resistance to trimethoprim among Enterobacteriaceae is increasing in spite of a stable or decreasing use of the drug. Integrons are common among these bacteria and many of them contain dfr gene cassettes. A clinical isolate of Escherichia coli from a urine specimen obtained at the Karolinska Hospital in Stockholm was resistant to trimethoprim, ampicillin, sulfonamides, chloramphenicol, streptomycin, and norfloxacin. PCR analysis with primers specific for the 5' and 3' sequences flanking the cassettes in class 1 integrons, detected a very short cassette region of no more than approximately 400 bp. Sequence analysis of the entire integron was performed, revealing a single-gene cassette of 411 bp encoding a new dihydrofolate reductase. The gene cassette comprised all sequences between the flanking conserved sequences and encoded only 78 amino acids. By homology it belongs to the unique group of dfr2 gene cassettes and being the fourth described gene in this group the new gene is called dfr2d. The enzymes encoded by the dfr2 genes are 67% identical and are not related to any other known dfr gene products.