Identification of a novel trimethoprim resistance gene, dfrK, in a methicillin-resistant Staphylococcus aureus ST398 strain and its physical linkage to the tetracycline resistance gene tet(L)

Abstract

A novel trimethoprim resistance gene, designated dfrK, was detected in close proximity to the tetracycline resistance gene tet(L) on the ca. 40-kb plasmid pKKS2187 in a porcine methicillin (meticillin)-resistant Staphylococcus aureus isolate of sequence type 398. The dfrK gene encodes a 163-amino-acid dihydrofolate reductase that differs from all so-far-known dihydrofolate reductases.

FIG. 1.

Comparison of the tet(L)-dfrK segment of plasmid pKKS2187 (accession no. FM207105) with plasmid pBC16 from B. cereus (accession no. AAA84922). The arrows indicate the extents and directions of the transcription of the genes repU (plasmid replication), tet(L) (tetracycline resistance), dfrK (trimethoprim resistance), and pre/mob (plasmid recombination/mobilization). It should be noted that the repU gene of pBC16 is intact and has been displayed only in a disrupted form for better comparison to the pKKS2187 sequence. The IS257 elements are shown as black boxes, with the white arrow indicating the transposase gene tnp. The 8-bp direct repeats at the IS257 integration sites are shown in boxes. The regions of >99% homology between pBC16 and pKKS2187 are marked by gray shading. The sequences at the junctions of pBC16-homologous and -nonhomologous parts in pKKS2187 are shown for comparison to the corresponding pBC16 sequence between the two maps. The BglII cleavage site in the IS257 element is indicated as Bgl. A size scale in kilobase pairs is given below each map.

FIG. 2.

Phylogenetic tree of the staphylococcal DfrS1 (14), DfrD (1, 3), DfrG (17), and DfrK (this study) proteins, which are involved in trimethoprim resistance, and the DfrB (6, 7) and DfrC (2) proteins, which represent trimethoprim-sensitive dihydrofolate reductases. Branch lengths are scaled according to amino acid exchanges observed in a multisequence alignment. The numbers at the major branch points refer to the percentage of times that a particular node was found in 10,000 bootstrap replications.