Specific and sensitive two-step polymerase chain reaction assay for the detection of Salmonella species

Abstract

A polymerase chain reaction (PCR) assay was applied for the selective amplification of a characteristic sequence within a Salmonella-specific chromosomal fragment. A two-temperature PCR cycle enhanced both the speed and overall sensitivity of the amplification procedure. Twenty-one well-characterized Salmonella strains and a number of non-Salmonella strains were tested. With the exception of the rarely isolated Salmonella arizonae strain, the PCR-based approach enabled the specific identification of Salmonella with a detection limit of 10(3) organisms. In combination with a nested PCR assay, as few as ten organisms were detectable. Specificity was demonstrated as no distinct amplification products were detectable with any of the tested non-Salmonella strains. With a pre-enrichment step using paramagnetic anti-Salmonella beads, an increase in sensitivity was observed in the case of clinical samples while the amplification process was not influenced.