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Specific identification of Staphylococcus aureus using the extracellular adherence protein gene as a novel target

Abstract number: P1761

Becker K., Hussain M., Sinha B., Joost I., Herrmann M., Peters G., von Eiff C.

Objectives: The widely used diagnostic assays based on biochemical or immunologic reactions to identify the species Staphylococcus aureus present various problems including unsatisfying sensitivity, specificity, and assay time. Here, the cell surface-associated extracellular adherence protein (Eap) – mediating adherence of S. aureus to host extracellular matrix components and inhibiting inflammation, wound healing and angiogenesis – was analysed as target for S. aureus identification purposes. The feasibility of an eap-based approach was systematically analysed on genomic, transcriptional and protein levels using a large collection of S. aureus and non-S. aureus staphylococcal isolates.

Methods: A well-characterized collection of S. aureus (n = 597) and non-S. aureus staphylococcal isolates (n = 216) comprising human and veterinary isolates from different European and non-European countries was tested for the presence of the Eap-encoding gene (eap) by PCR. In addition, Western and Northern blot analyses were performed. Clinical isolates were identified on species level using biochemical test kits. If the identification was ambiguous or categorised as unacceptable, universal target gene sequencing (16S rRNA, rpoB) was performed. PCR assays for detection of S. aureus and S. intermedius specific nuc genes were applied to confirm the identification of both species.

Results: Due to the polymorphism of the eap gene, novel PCR oligonucleotide primers for diagnostic purposes were designed. While all S. aureus isolates were eap-positive using the newly designed primers, this gene was not detected in non-S. aureus staphylococci comprising 47 different species and subspecies of coagulase-negative staphylococci and non-aureus coagulase positive/variable staphylococci. Furthermore, none of the non-S. aureus isolates expressed EAP homologs on transcriptional and protein levels. Based on these data, the sensitivity and specificity of the newly developed PCR targeting the eap gene were both 100%.

Conclusion: Eap was shown to be a virulence factor specifically restricted to S. aureus, occurring in all isolates of this species tested. Thus, Eap seems to be a core trait of S. aureus, offering a promising tool particularly suitable for molecular diagnostics of this pathogen.

Session Details

Date: 19/04/2008
Time: 00:00-00:00
Session name: 18th European Congress of Clinical Microbiology and Infectious Diseases
Subject:
Location: Barcelona, Spain
Presentation type:
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