Cloning and Nucleotide Sequence Determination of the Entire mec DNA of Pre-Methicillin-Resistant Staphylococcus aureus N315

Abstract

ABSTRACT In methicillin-resistant Staphylococcus aureus , the methicillin resistance gene mecA is localized within a large chromosomal region which is absent in the methicillin-susceptible S. aureus chromosome. The region, designated mec DNA, is speculated to have originated from the genome of another bacterial species and become integrated into the chromosome of the S. aureus cell in the past. We report here cloning and determination of the structure of the entire mec DNA sequence from a Japanese S. aureus strain, N315. The mec DNA was found to be 51,669 bp long, including terminal inverted repeats of 27 bp and a characteristic pair of direct repeat sequences of 15 bp each: one is situated in the right extremity of mec DNA, and the other is situated outside the mec DNA and abuts the left boundary of mec DNA. The integration site of mec DNA was found to be located in an open reading frame (ORF) of unknown function, designated orfX . Clusters of antibiotic resistance genes were noted in mec DNA carried by transposon Tn 554 and an integrated copy of plasmid pUB110. Both the transposon and plasmid were integrated in the proximity of the mecA gene, the latter being flanked by a pair of insertion sequence IS 431 elements. Many ORFs other than those encoding antibiotic resistance were considered nonfunctional because of the acquired mutations or partial deletions found in the ORFs. Two ORFs potentially encoding novel site-specific recombinases were found in mec DNA. However, there was no ORF that might encode mec DNA-specific transposase or integrase proteins, indicating that the mec DNA is not a transposon or a bacteriophage in nature.