A Novel Approach Based on Real-Time PCR with High-Resolution Melting Analysis for the Simultaneous Identification of Staphylococcus aureus and Staphylococcus argenteus

<i>Staphylococcus</i> (<i>S</i>.) <i>aureus</i> is a pathogenic bacterium able to cause several diseases in humans and animals as well as foodborne intoxications. <i>S. argenteus</i>, being phenotypically and genotypically related to <i>S. aureus</i>, is part of the so-called <i>S. aureus</i> complex and recently recognized as an emerging pathogen able to cause, like <i>S. aureus</i>, several diseases both in humans and animals, and foodborne poisoning outbreaks. However, it has been reported that the widely used conventional PCR of Brakstad et al. [<i>Journal of Clinical Microbiology</i>, <i>30</i>(7), 1654-1660, (1992)] targeting the thermostable nuclease gene may provide false-positive <i>S. aureus</i>, as it is able to amplify also <i>S. argenteus</i>. Here, we developed a novel two-step approach that, following the PCR of Brakstad et al. (1992), discriminates <i>S. aureus</i> from <i>S. argenteus</i> by a real-time PCR with high-resolution melting analysis (rt-PCR-HRM). In particular, targeting a polymorphic 137 bp region of the <i>sodA</i> gene, our developed rt-PCR-HRM method clearly discriminated <i>S. aureus</i> from <i>S. argenteus</i>, showing a remarkable difference in their amplification product melting temperatures (approximately 1.3 °C) as well as distinct melting curve shapes. The good sensitivity, reproducibility, user friendliness, and cost effectiveness of the developed method are advantageous attributes that will allow not only its easy employment to correctly identify misidentified isolates present in various collections of <i>S. aureus,</i> but also expand the still lacking knowledge on the prevalence and distribution of <i>S. argenteus</i>.