Staphylococcus aureus resistance to mupirocin is often caused by acquisition of a novel isoleucyl-tRNA synthetase encoded on the plasmid gene mupA. We tested S. aureus isolates from children at Texas Children's Hospital with recurrent skin and soft tissue infections for mupirocin resistance and mupA. Of 136 isolates, 20 were resistant to mupirocin (14.7%). Fifteen isolates (11%) carried mupA, and the gene was more common in methicillin-susceptible S. aureus (21.4%) than methicillin-resistant S. aureus (8.3%; P=0.03). Seven of 20 mupirocin-resistant isolates displayed clindamycin resistance.