Four copies of the insertion sequence IS257 are found in the mec region of the chromosome of the Australian methicillin resistant Staphylococcus aureus (MRSA) strain ANS46, two flanking a merAmerB sequence (encoding resistance to mercurial compounds), the other two flanking an integrated copy of the plasmid pT181 (tetracycline resistance). The termini of the integrated copy of the plasmid pT181 carry a direct repeat of 8 bp of plasmid sequence, but otherwise there are no similarities in the 8 bp sequences flanking the four copies of IS257 in this strain. Integrated copies of pT181 in strains R35 (a New Jersey MRSA) and GH32 (MRSA of Greek origin) have the same terminal repeat as in ANS46, suggesting either a specific site of insertion of IS257 into the free plasmid before integration into the chromosome, or a common evolutionary lineage for these geographically diverse isolates. A different 8 bp terminal repeat of plasmid sequence is found in the chromosomally integrated copy of pUB110 (flanked by a pair of IS257s) in R155, another New Jersey MRSA. This 8 bp repeat differs from that reported previously for pUB110/IS257 inserted into the plasmid pSK41, indicating insertion of IS257 into different sites of pUB110 before integration into the chromosome or into pSK41. In the plasmid pSK1, the two outer copies of IS257 of the three associated with Tn4003 (trimethoprim resistance) are also flanked by 8 bp repeats.(ABSTRACT TRUNCATED AT 250 WORDS)