Genus- and species-specific detection ofListeria monocytogenesusing polymerase chain reaction assays targeting the 16S/23S intergenic spacer region of the rRNA operon

Abstract

In this study, the 16S/23S rRNA intergenic spacer (IGS) regions of six Listeria species were examined. DNA bands of 590 and 340 bp were observed following polymerase chain reaction (PCR) amplification of DNA from Listeria monocytogenes, Listeria innocua, Listeria seeligeri, Listeria welshimeri, and Listeria ivanovii strains with generic rRNA IGS oligonucleotide primers. For strains of Listeria grayi subspp. grayi and murrayi, DNA band sizes of 550 and 340 bp were observed with this primer pair. DNA bands of these sizes were not observed for other Gram-negative or- positive bacteria in this PCR assay. Four RsaI digestion profiles were noted for the Listeria PCR products. Listeria monocytogenes strains had one profile; L. innocua strains had a second; L seeligeri, L. welshimeri, and L ivanovii strains had a third; and L. grayi strains had a fourth. The small and large 16S/23S rRNA IGSs of L. monocytogenes ATCC 15313 were identical in the first 58 5' and the last 169 3' nucleotides. However, the large rRNA IGS contained a central 267-bp region with tRNA(Ile) and tRNA(Ala) genes. Large rRNA 16S/23S IGS nucleotide sequence data has not been previously reported. This data was used to develop novel Listeria genus-specific and L.monocytogenes species-specific PCR assays.