Comparison of primers for the detection of Salmonella enterica serovars using real-time PCR.
OBJECTIVE
To evaluate the specificity and sensitivity of PCR primers for the detection of Salmonella enterica in a real-time PCR assay using pure cultures.


RESULTS
Unenriched whole cells in sterile water were used as template for each PCR. SYBR Green dye was used for the nonspecific detection of dsDNA. The real-time PCR detection limits of five previously published primer sets used in conventional PCR applications were not below 3 x 10(3) CFU per reaction (rxn). A new primer set, Sen, was designed, which detected Salm. enterica Newport down to 6 CFU rxn(-1) in one case, and gave an average detection limit of 35 CFU rxn(-1) over three separate runs.


CONCLUSIONS
Primers originally designed for end-point PCR did not have adequate specificity or sensitivity compared with those specifically designed for real-time PCR.


CONCLUSIONS
This study emphasizes the importance of evaluating real-time PCR primer sets in pure cultures prior to testing in field samples. This study will benefit other researchers in selecting an appropriate primer set for real-time PCR detection of Salm. enterica.
Csordas
A T
AT
Department of Biological and Agricultural Engineering, University of California at Davis, Davis, CA 95616, USA.
Barak
J D
JD
Delwiche
M J
MJ
D001426:Bacterial Proteins / Q000235:genetics; D015373:Bacterial Typing Techniques / Q000379:methods*; D017931:DNA Primers; D016133:Polymerase Chain Reaction / Q000379:methods*; D019779:Salmonella enterica / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification
D001426:Bacterial Proteins; D017931:DNA Primers; C075518:invA protein, Bacteria