Identification of Staphylococcus aureus enterotoxins A and B genes by PCR-ELISA

We developed PCR-enzyme linked immunosorbent (ELISA) assays to detect Staph ylococcus aureus enterotoxins A and B genes. The assays use internal biotin -labelled oligonucleotides as capture probes for immobilizing and subsequen tly detecting target sequences on microtiter plates. The detection limits o f the PCR-ELISAs were approximately 250 gene copies, versus 2500 gene copie s by agarose gel analysis. The sensitivity of the assays, as determined fro m a reference panel of 46 coded samples that included DNA purified from 31 different bacterial species and strains, SEA and SEB plasmid controls, and no-template controls was 100%. No cross-reactivity was observed with DNA fr om non-staphylococcal species, Using 27 clinical isolates of S. aureus, the SEA PCR-ELISA identified the enterotoxin A (sea) gene in 26 samples, and t he SEB PCR-ELISA identified the enterotoxin B (seb) gene in all 27 samples. Compared with conventional antigen capture ELISAs for SEA and SEB toxins, the PCR-ELISAs showed overall superior detection limits. The sensitivity an d specificity levels of the SEA PCR-ELISA and the SEA toxin ELISA were comp arable within their respective detection thresholds, but the sensitivity an d specificity of the SEB PCR-ELISA was much greater than that of SEB toxin ELISA. (C) 2000 Academic Press.