Journal
DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE
Volume 61, Issue 4, Pages 446-452Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.diagmicrobio.2008.03.012
Keywords
blood culture; DNA extraction methods; real-time (RT)-PCR MRSA
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Although the introduction of automated blood culture systems has dramatically reduced laboratory personnel bench time, 48 to 72 h is still required for the identification of pathogens such as methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) and the accurate determination of antimicrobial sensitivities for prompt optimal patient therapy and infection control initiatives. The following 4 DNA blood culture extraction methods were compared: (a) organic, (b) differential centrifugation and lysis, (c) alkali wash/lysis, and (d) Qiagen lysis/filtration (QIAGEN, West Sussex, UK). The benzyl alcohol extraction method (a) was found to be the most optimal method having a reasonable extraction time of 1.8h and 100% correlation with the gold standard laboratory culture. A dual locus real-time polymerase chain reaction (PCR) targeting the S. aureus-specific thermonuclease nuc gene and the staphylococcal methicillin resistance determinant mecA gene were used as a reliable indicator of the presence of MRSA. In conjunction with the DNA extraction method (a), detection time for MRSA/MSSA isolates from positive blood cultures was dramatically reduced from at least 24 to 48 It to approximately 3 h. (c) 2008 Elsevier Inc. All rights reserved.
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