title: Immunoquantitative real-time PCR for detection and quantification of Staphylococcus aureus enterotoxin B in foods.
abstract: A real-time immunoquantitative PCR (iqPCR) method for detection of Staphylococcus aureus enterotoxin B (SEB) was developed and evaluated using both pure cultures and foods. The assay consisted of immunocapture of SEB and real-time PCR amplification of the DNA probe linked to the detection antibody. iqPCR was compared to an in-house enzyme-linked immunosorbent assay (ELISA) using the same couple of capture-detection antibodies and to commercial kits for detection of S. aureus enterotoxins (SE). The iqPCR was approximately 1,000 times more sensitive (<10 pg ml(-1)) than the in-house ELISA and had a dynamic range of approximately 10 pg ml(-1) to approximately 30,000 pg ml(-1). iqPCR was not inhibited by any of the foods tested and was able to detect SEB present in these foods. No cross-reactivity with SE other than SEB was observed. Application of iqPCR for detection of SEB in cultures of S. aureus revealed the onset of SEB production after 4 h of incubation at 22, 37, and 42 degrees C, which was in the first half of the exponential growth phase. The total amounts of SEB produced by the two strains tested were larger at 42 degrees C than at 37 degrees C and were strain dependent.
authors.affiliation: Laboratory of Food Microbiology and Food Preservation, Department of Food Safety and Food Quality, Faculty of Bioscience Engineering, Ghent University, Ghent, Belgium.
authors.forename: Andreja
authors.initials: A
authors.lastname: Rajkovic
authors.forename: Benaissa
authors.initials: B
authors.lastname: El-Moualij
authors.forename: Mieke
authors.initials: M
authors.lastname: Uyttendaele
authors.forename: Philippe
authors.initials: P
authors.lastname: Brolet
authors.forename: Willy
authors.initials: W
authors.lastname: Zorzi
authors.forename: Ernst
authors.initials: E
authors.lastname: Heinen
authors.forename: Ellen
authors.initials: E
authors.lastname: Foubert
authors.forename: Johan
authors.initials: J
authors.lastname: Debevere
mesh_terms: D001427:Bacterial Toxins / Q000032:analysis; D017931:DNA Primers; D015342:DNA Probes; D004768:Enterotoxins / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay; D005504:Food Analysis; D005516:Food Microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013202:Staphylococcal Food Poisoning; D013211:Staphylococcus aureus / Q000737:chemistry* / Q000378:metabolism
chemical_list: D001427:Bacterial Toxins; D017931:DNA Primers; D015342:DNA Probes; D004768:Enterotoxins