PCR-RFLP analysis of the coagulase gene of Staphylococcus aureus: application to the differentiation of epidemic and sporadic methicillin-resistant strains.
Preventing cross-infection with epidemic strains of methicillin-resistant Staphylococcus aureus (MRSA) requires effective control measures. These call for simple, rapid, discriminatory and reproducible methods for typing this pathogen. In this study 140 isolates/strains from 105 hospitals in England and Wales, representing 72 diverse phage types, were analysed by bacteriophage typing and PCR coagulase (coa) gene restriction fragment length polymorphism (RFLP). Isolates gave a coa gene PCR product that was either 660 base pairs (bp), 603 bp or 547 pb in size. The PCR products were digested with Alu I and Cfo I, and the fragments separated by gel electrophoresis. Eight coa gene RFLP patterns, numbered 1 to 8, were observed. Pattern 3 was most common (N = 25 isolates), followed by patterns 2 and 5 (18 isolates each), pattern 1 (14 isolates), pattern 4 (11 isolates), pattern 7 (10 isolates), pattern 8 (eight isolates) and pattern 6 (six isolates). Isolates of the same phage type often gave different coa gene RFLP patterns, and the patterns within the epidemic types EMRSA-03, EMRSA-15 and EMRSA-16 were heterogeneous. Thus, representatives of EMRSA-03 were subtyped to coa RFLP patterns 1 and 2, those of EMRSA-05 to coa RFLP patterns 1, 2, 7 and 8, and those for EMRSA-16 to coa RFLP patterns 2, 3, 4, 5 and 6. The range of patterns within single phage types of S. aureus could help to discriminate between isolates/strains, and in a hierarchical approach coa gene RFLP could occupy an intermediate position between phage typing and pulsed-field gel electrophoresis (PFGE).
Hookey
J V
JV
Molecular Biology Unit, Virus Reference Laboratory, Central Public Health Service, Colindale, London, UK.
Edwards
V
V
Cookson
B D
BD
Richardson
J F
JF
D001434:Bacteriophage Typing; D002876:Chromosomes, Bacterial / Q000235:genetics; D003030:Coagulase / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D004739:England; D005798:Genes, Bacterial / Q000235:genetics*; D006801:Humans; D016106:Methicillin Resistance / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D013211:Staphylococcus aureus / Q000145:classification / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification; D014852:Wales
D003030:Coagulase; D004269:DNA, Bacterial