Molecular investigation of an epidemic MRSA: A comparative study of its recognition, introduction and spread in the west of Scotland

The incidence of infections caused by methicillin-resistant Staphylococcus aureus (MRSA) has increased world-wide over the past 30 years. A strain that was introduced into Scotland in 1990 via a patient recently returned from Lisbon, Portugal had an unusually resistant phenotype. Ninety- three isolates were selected and investigated by molecular methods. The principal method chosen was agarose gel electrophoresis following digestion of whole cell genomic DNA with the restriction enzymes HhaI and Sau3AI. These enzymes recognise 4-base DNA sequences and produced an analytical window at the top of an agarose gel, which allowed the recognition of plasmid DNA fragments and partial digest products. The final result of electrophoresis by this method was a considerable improvement over previous methods employing enzymes that are 6-base cutters. The strain was studied in parallel with control groups of Staph, aureus that consisted of methicillin-sensitive Staph, aureus, sporadic isolates of MRSA and the epidemic strains EMRSA-1, EMRSA-15 and EMRSA-16. Analysis of the HhaI restriction enzyme fragmentation patterns (REFP) of the "new" strain and control groups by Dice coefficients of similarity validated the technique with respect to discrimination; it was demonstrated that REFP's of epidemiologically um-elated MSSA isolates had low Dice coefficient values (mean So value = 66%) and that REFP's of known epidemiologically related isolates such as EMRSA-15 had high coefficients of similarity (mean SD value = 99%). The technique showed that all isolates of the new strain were clonal in origin (mean SD value = 95%) and in addition, highlighted the existence of a number of clonal variants (subtypes) to the major REFP type. Sixty-eight isolates (73%) gave a genomic fingerprint identical to the index case and were designated HhaI type H1. Twenty-five isolates were variants of this type and were designated type LH2 (7 isolates), LH3 (6 isolates), LH10 (2 isolates) and fifteen of the twenty-five were unique variants designated LH4 - LH9 and LH11 - LH14. Nine isolates of another strain, imported from France and phenotypically similar to the study strain were shown to be genetically closely related to it. Inter-group matching of REFP's showed each control group to be genetically distinct to each other and to the "new" MRSA strain. In a collaborative study, this new strain which has been trivially termed the "Lisbon strain" was shown to be closely related to the now well characterised Iberian clone MRSA. Variants detected using HhaI/Sau3AI typing also showed parallel variation in PFGE. A small number of genomic variants were also found within the EMRSA-1, 15 and 16 control groups, highlighting the capacity of the technique to detect minor genetic change. Restriction enzyme fingerprinting of whole cell genomic DNA using the restriction enzymes HhaI and Sau3AI proved to be a simple, economic and highly discriminatory method of typing Staph. aureus strains requiring no expensive apparatus.