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Detection of methicillin-resistant Staphylococcus aureus (MRSA) by single-locus PCR assays that target the extremity of the staphylococcal cassette chromosome-mec (SCCmec) and part of the adjacent S. aureus-specific open reading frame gene (orfX) is a significant diagnostic advancement, since it provides real-time detection directly from screening specimens. However, isolates harboring mecA deletions within SCCmec may result in false-positive identification of MRSA in these assays. We characterized 24 methicillin-susceptible S. aureus (MSSA) isolates that tested positive in one such assay to investigate this phenomenon. Seven isolates resembled USA100 and carried SCCmec II elements with mecA deletions that spanned 20 to 46 kbp. The mecA excisions in USA100-resembling isolates appeared to be linked with IS431 transposable elements present in SCCmec II. For 17 isolates that resembled USA400 and/or MSSA476, the identity and possible excision of SCC elements could not be confirmed. The downstream common sequence (dcs) shared by SCCmec I, II, and IV elements was detected in these isolates. Sequence analysis of the chromosomal regions flanking the missing SCC element revealed an intact SCC integration site, a duplicate dcs, and the enterotoxin gene cluster downstream of orfX. An annealing sequence for one of the SCCmec-specific primers (mecii574) in the single-locus PCR assay was identified in the duplicate dcs. In the absence of SCC, a 176-bp amplicon can be generated from this mecii574 annealing sequence to yield a false-positive result. In conclusion, partial SCCmec II excisions via IS431 elements in strains that resembled USA100 and the presence of a duplicate mecii574 annealing sequence in strains that resembled USA400/MSSA476 were identified as causes for false-positive results in a single-locus PCR assay that targets the SCCmec/orfX junction.

Characterization of Staphylococcus aureus Isolates with a Partial or Complete Absence of Staphylococcal Cassette Chromosome Elements