title: Evaluation of a duplex real-time PCR assay to detect MRSA from broth culture, human sera seeded with MRSA and from patient's serum.
abstract: The need for rapid methods in order to precisely detect methicillin-resistant Staphylococcus aureus (MRSA) is extensively acknowledged. This study evaluated a quantitative real-time PCR assay targeting mecA (encoding high level resistance to methicillin) and femB (a specific genomic marker for S. aureus) genes to detect MRSA from broth culture, from serum seeded with MRSA and straight from the patient's serum. One hundred and thirty-five clinical isolates of MRSA strains and different species were utilised in this study. In addition, a pilot study with 9 patients' serum samples was performed. The sensitivity and specificity values for this assay were 99% and 100% respectively. The detection limit for this method was 1.23×10(2) CFU/ml from the serum seeded with MRSA cells and the limiting concentration of DNA for detection was 18 fg, which equates to 5.14 genomic DNA copies. In addition, this assay detected MRSA from patient's serum (7 out of 9) with sensitivity of 77.8%. Overall, the assay was rapid, efficient, sensitive and easy to perform.
authors.affiliation: University Hospital of South Manchester NHS Foundation Trust, Wythenshawe Hospital, Southmoor Road, Manchester, M23 9LT.
authors.forename: Sawsan
authors.initials: S
authors.lastname: Awad
authors.forename: Issam
authors.initials: I
authors.lastname: Alshami
authors.forename: Ahmed Eid
authors.initials: AE
authors.lastname: Alharbi
keywords: Duplex; Evaluation; MRSA; Sera; femB; mecA; real-time PCR