Borderline oxacillin-resistant Staphylococcus aureus (BORSA) exhibit oxacillin 
MIC values of 1-8 microg ml(-1), but lack mecA, which encodes the low-affinity 
penicillin-binding protein (PBP)2a. The relationship of the BORSA phenotype with 
specific genetic backgrounds was assessed, as well as amino acid sequence 
variation in the normal PBP2. Among 38 BORSA, 26 had a common PFGE profile of 
genomic DNA, and were multilocus sequence type (ST)25. The other isolates were 
genetically diverse. Complete pbp2 sequences were determined for three BORSA, 
corresponding to ST25, ST1 and ST47, which were selected on the basis of lacking 
blaZ-encoded beta-lactamase. The essential transpeptidase-domain-encoding 
segment of pbp2 was also sequenced from seven additional ST25 isolates. Amino 
acid substitutions occurred in the transpeptidase domain of all BORSA, 
irrespective of clonal type. A Gln(629)-->Pro substitution was common to all 
ST25 BORSA, but most could be distinguished from one another by additional 
unique substitutions in the transpeptidase domain. The ST1 and ST47 isolates 
also possessed unique substitutions in the transpeptidase domain. 
Plasmid-mediated expression of pbp2 from an ST25 or ST1 isolate in S. aureus 
RN6390 increased its oxacillin MIC from 0.25 to 4 microg ml(-1), while pbp2 from 
a susceptible strain, ATCC 25923, had no effect. Therefore, different amino acid 
substitutions in PBP2 of diverse BORSA lineages contribute to borderline 
resistance. The predominant ST25 lineage was not related to any of the five 
clonal complexes that contain meticillin-resistant S. aureus (MRSA), suggesting 
that ST25 cannot readily acquire mecA-mediated resistance.