Detection of Staphylococci in Mouse Phagocytic Cells by in Situ Hybridization Using Biotinylated Dna Probes

In situ hybridization was used to detect intracellular Staphylococcus aureus and S. epidermidis in mouse phagocytic cells after experimental infection of C3H mice with Staphylococci via abdominal or intravenous injection. Isolated ascites or whole blood were tested by the phagocyte smear technique, using bacteriolytic enzymes to preserve phagocytic cell morphology. The exposed bacterial DNA was visualized as intracellular hybridized signals by use of biotinylated DNA probes and by immunocytochemistry using streptavidin-alkaline phosphatase conjugates as detector molecules. These DNA probes, prepared from randomly cloned genomic DNA fragments of S. aureus and S. epidermidis, were strain-specific and did not cross-hybridize either in situ or on dot-blot hybridization. This technique of in situ hybridization with phagocyte smears is useful for detection and diagnosis of intracellular bacteria regardless of viability.