Staphylococcus aureus, particularly methicillin-resistant strains (MRSA), are likely targets for molecular development, particularly in nares and blood cultures. As more institutions implement patient screening protocols using nares specimens for MRSA, replacing routine culture methods with molecular assays has gained increasing attention.
Peptide Nucleic Acid (PNA)-Fluorescence In Situ Hybridization (FISH) assays definitively identify Staphylococcus aureus from positive blood culture vials. PNA-FISH is a relatively straightforward procedure that does not involve amplification and requires limited equipment. It is easy to perform procedurally with minimal hands-on time.
PNA is a synthetic imitator of a nucleic acid sequence whose backbone is a pseudopeptide rather than a sugar. PNA behaves similarly to DNA and will bind to complementary nucleic acid strands. A PNA probe is constructed, utilizing a complementary, hybridizing sequence for a known nucleic acid target sequence. The probe is typically bound to a fluorescent protein as a means of visualizing/detecting the target.
In one commercially available method, once a blood culture vial demonstrates gram-positive cocci in clusters, a drop of the blood culture broth is added to the fixation solution on a slide. Heat or methanol is used to fix the smear. After fixation, the probe that targets species-specific ribosomal RNA is added to the smear, which is then cover-slipped.
Slides are then incubated at 55°C. After incubation, they are immersed in a preheated wash solution, and coverslips are gently removed. After incubation in the wash solution, smears are air-dried; a drop of mounting medium is added, and the slide is cover-slipped again.
The slides are examined with a fluorescent microscope utilizing specific filters. Green fluorescing cocci in clusters are identified as Staphylococcus aureus. Depending on the routine identification system, this identification would be available 24 hours earlier than the norm.
Many blood cultures demonstrating gram-positive cocci in clusters yield coagulase-negative staphylococci (CNS), representing potential contaminants rather than significant infection. What is the significance of differentiating blood cultures containing S. aureus from those growing CNS much earlier?
Studies have shown that if the differentiation of CNS from S. aureus is effectively communicated to clinicians and pharmacy/antimicrobial stewardship teams, active assessment can occur utilizing defined exclusion criteria for those patients whose cultures yielded CNS rather than S. aureus. In scenarios where contamination rather than infection is indicated, vancomycin can be discontinued earlier, and the hospital stay is shortened. Potential benefits include reduced antibiotic exposure, decreased risk of developing resistance, and reduced cost.
