1. Development of a panel of seven duplex real-time PCR assays for detecting 13 streptococcal superantigens.
- Author
-
Yang P, Peng X, Cui S, Shao J, Zhu X, Zhang D, Liang H, and Wang Q
- Subjects
- Antigens, Bacterial genetics, Base Sequence, DNA Primers genetics, Limit of Detection, Sequence Alignment, Streptococcus pyogenes genetics, Streptococcus pyogenes immunology, Superantigens genetics, Antigens, Bacterial analysis, Real-Time Polymerase Chain Reaction methods, Streptococcus pyogenes isolation & purification, Superantigens analysis
- Abstract
Background: Streptococcal superantigens (SAgs) are the major virulence factors of infection in humans for group A Streptococcus (GAS) bacteria. A panel consisting of seven duplex real-time PCR assays was developed to simultaneously detect 13 streptococcal SAgs and one internal control which may be important in the control of GAS-mediated diseases., Methods: Primer and probe sequences were selected based on the highly conserved region from an alignment of nucleotide sequences of the 13 streptococcal SAgs. The reaction conditions of the duplex real-time PCR were optimized and the specificity of the duplex assays was evaluated using SAg positive strains. The limit of detection of the duplex assays was determined by using 10-fold serial dilutions of the DNA of 13 streptococcal SAgs and compared to a conventional polymerase chain reaction (PCR) method for evaluating the duplex assays sensitivity., Results: Using the duplex assays, we were able to differentiate between 13 SAgs from Streptococcus strains and other non-Streptococcus bacteria without cross-reaction. On the other hand, the limit of detection of the duplex assays was at least one or two log dilutions lower than that of the conventional PCR., Conclusions: The panel was highly specific (100%) and the limit of detection of these duplex groups was at least ten times lower than that obtained by using a conventional PCR method.
- Published
- 2013
- Full Text
- View/download PDF