The emerging methicillin-resistant Staphylococcus aureus ST398 clone can easily be typed using the Cfr9I SmaI-neoschizomer

Aim: To establish a PFGE protocol using Cfr9I, neoschizomer of SmaI, for typing of Staphylococcus aureus isolates belonging to the emerging MRSA ST398 clone. Methods and Results: Staphylococcus aureus ST398 and non-ST398 isolates were analysed using the PFGE conditions recommended by the HARMONY consensus protocol. Genomic DNA of non-ST398 isolates could be digested with SmaI, XmaI (also a SmaI-neoschizomer) and Cfr9I. The DNA of SmaI-nontypeable ST398 isolates was partially resistant to XmaI, but could be digested with Cfr9I. By PCR-amplification/sequencing, the presence of a novel C5-cytosine methyltransferase gene (sauST398M) was detected in the ST398 isolates. The encoded enzyme, which shows high similarity with C5-cytosine methyltransferases that modify the CCCGGG recognition sequence, could be responsible for the different restriction results. Conclusion: SmaI-PFGE is regarded as the 'gold standard' for typing S. aureus. Because of different susceptibility of the GGGCCC recognition sites of the ST398 DNA against SmaI, XmaI and Cfr9I, the proposed protocol is a valuable tool for ST398 typing. Significance and Impact of the Study: The use of this protocol allows the comparison of results from SmaI-nontypeable isolates with S. aureus SmaI-PFGE databases and can be applied for outbreak investigations and traceability studies of this emerging MRSA clone.