Genetic characterization of the fusidic acid and cadmium resistance determinants of Staphylococcus aureus plasmid pUB101.

Abstract

We report the cloning of the fusidic acid and cadmium resistance determinants from Staphylococcus aureus plasmid pUB101. The pUB101 fusidic acid resistance determinant was located on a 2.9 kb HindIII fragment. Sequencing of this fragment revealed three putative open reading frames (ORFs) of 213 (far1), 152 (orf152) and 170 amino acids (orf170), which are flanked by the right-hand end of insertion sequence IS431/257 (IS431/257RH) and a partial ORF. Far1 and Orf152 demonstrated homology with a chromosomally encoded fibronectin-binding protein of Listeria monocytogenes and the putative protein YosT, found on the SPβc2 prophage of Bacillus subtilis, respectively. Transformation of S. aureus with a construct containing a 949 bp far1-specific amplicon led to the isolation of a fusidic acid-resistant transformant, thereby identifying the pUB101 fusidic acid resistance structural gene. Between orf152 and far1 we identified a unique 113 bp symmetrical element and other repeat elements that may be involved with the control of orf152 and/or far1 expression. Hybridization of Southern blots revealed that far1 was not located on the chromosome or plasmid content of a limited number of Australian, UK and Hong Kong fusidic acid-resistant isolates. The pUB101 cadmium resistance determinant was located on a 3.6 kb HindIII fragment that carried a cadDX operon, remnants of two putative plasmid replication protein genes and IS431/257RH. Sequence analysis also demonstrated the presence of a single-stranded origin of replication, normally found on rolling circle replicating plasmids, within the putative promoter region of the cadDX operon.