Methicillin-resistant Staphylococcus aureus screening by online immunometric monitoring of bacterial growth under selective pressure.
Rapid, high-throughput screening tools are needed to contain the spread of hospital-acquired methicillin (meticillin)-resistant Staphylococcus aureus (MRSA) strains. Most techniques used in current clinical practice still require time-consuming culture for primary isolation of the microbe. We present a new phenotypic assay for MRSA screening. The technique employs a two-photon excited fluorescence (TPX) detection technology with S. aureus-specific antibodies that allows the online monitoring of bacterial growth in a single separation-free process. Different progressions of fluorescence signals are recorded for methicillin-susceptible and -resistant strains when the growth of S. aureus is monitored in the presence of cefoxitin. The performance of the new technique was evaluated with 20 MRSA strains, 6 methicillin-susceptible S. aureus strains, and 7 coagulase-negative staphylococcal strains and two different monoclonal S. aureus-specific antibodies. When either of these antibodies was used, the sensitivity and the specificity of the TPX assay were 100%. All strains were correctly classified within 8 to 12 h, and up to 70 samples were simultaneously analyzed on a single 96-well microtiter plate. As a phenotypic method, the TPX assay is suited for screening purposes. The final definition of methicillin resistance in any S. aureus strain should be based on the presence of the mecA gene. The main benefit afforded by the initial use of the TPX methodology lies in its low cost and applicability to high-throughput analysis.
Stenholm
Teppo
T
Department of Medicine, Turku University Hospital, University of Turku, Turku, Finland. teppo.stenholm@utu.fi
Hakanen
Antti J
AJ
Vaarno
Jonne
J
Pihlasalo
Sari
S
Terho
Perttu
P
Hänninen
Pekka E
PE
Vuopio-Varkila
Jaana
J
Huovinen
Pentti
P
Kotilainen
Pirkko
P
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015373:Bacterial Typing Techniques / Q000379:methods*; D002440:Cefoxitin / Q000494:pharmacology; D007118:Immunoassay / Q000379:methods*; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects / Q000254:growth & development* / Q000302:isolation & purification*
D000900:Anti-Bacterial Agents; D002440:Cefoxitin