Recovery and Enumeration of Staphylococcus aureus by the Selective Agar Overlay Method

Abstract

Staphylococcus aureus is an example of a commensal bacterium responsible for emesis, acute diarrheal syndrome, and sepsis. S. aureus often must be isolated from patient samples in a clinical setting or from food samples during food processing in an industrial setting, although these bacterial cells may be injured by the human immune system or by food processing measures. Therefore, injured cells may not be fully recovered on media selective for S. aureus and enumeration (e.g., CFU/mL) may not reflect the true concentration of the original sample. The objective of this study was to determine whether the selective agar overlay method of recovery is more sensitive, selective, and time-effective for enumeration of artificially injured S. aureus cultures when compared to more traditional techniques. The selective agar overlay method involves pour plating S. aureus in non-selective medium, allowing the sample to incubate for a four hour recovery period, and then overlaying selective medium over the non-selective medium. Artificial injury of S. aureus cells was accomplished by treatment with carvacrol, an extract from oil of oregano. Our results indicated that carvacrol-injured S. aureus cells were recovered by the selective agar overlay at the same concentration as recovery on non-selective media, and at a significantly higher concentration than recovery on selective media. This method allows for more rapid and accurate diagnoses, and may be more cost-effective due to the reduction or elimination of false negative results