Abstract
Staphylococcus aureus is the leading cause of skin and soft tissue infections, bacteremia, infective endocarditis, osteoarticular, pleuropulmonary, and device-related infections. Virulence factors secreted by S. aureus, including superantigens and cytotoxins, play significant roles in driving disease. The ability to identify virulence factors present at the site of infection will be an important tool in better identifying and understanding how specific virulence factors contribute to disease. Previously, virulence factor production has been determined by culturing S. aureus isolates and detecting the mRNA of specific virulence factors. We demonstrated for the first time that virulence factors can be directly detected at the protein level from human samples, removing the need to first culture isolated bacteria. Superantigens and cytotoxins were detected and quantified with a Western dot blot assay by using reconstituted skin swabs obtained from patients with atopic dermatitis. This methodology will significantly enhance our ability to investigate the complex host-microbe environment and the effects various therapies have on virulence factor production. Overall, the ability to directly quantify virulence factors present at the site of infection or colonization will enhance our understanding of S. aureus-related diseases and help identify optimal treatments.IMPORTANCE For the first time, we show that secreted staphylococcal virulence factors can be quantified at the protein level directly from skin swabs obtained from the skin of atopic dermatitis patients. This technique eliminates the need to culture Staphylococcus aureus and then test the strain's potential to produce secreted virulence factors. Our methodology shows that secreted virulence factors are present on the skin of atopic patients and provides a more accurate means of evaluating the physiological impact of S. aureus in inflammatory diseases such as atopic dermatitis.
Highlights
Staphylococcus aureus is the leading cause of skin and soft tissue infections, bacteremia, infective endocarditis, osteoarticular, pleuropulmonary, and devicerelated infections
We assumed that the amounts of superantigens and cytotoxins in vivo on human skin may be low, possibly in the microgram to nanogram per milliliter amounts, if present at all
We were able to determine that the skin swab does not interfere with the detection of TSST-1, as the density of the unknowns was 0.12 Ϯ 0.02 g/ml when spiked with 0.1 g/ml of TSST-1 and 0.02 Ϯ 0.01 g/ml when spiked with 0.01 g/ml of TSST-1
Summary
Staphylococcus aureus is the leading cause of skin and soft tissue infections, bacteremia, infective endocarditis, osteoarticular, pleuropulmonary, and devicerelated infections. Secreted virulence factors were indirectly detected by culturing S. aureus isolates from patients and analyzing the culture media for superantigen and cytotoxin mRNA or proteins [22]. We show that secreted staphylococcal virulence factors (superantigens and cytotoxins) can be quantified directly, without culture, at the protein level from swabs obtained from atopic dermatitis patients and healthy control nonpurulent skin.
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