Detection of pathogenicYersinia enterocoliticausing the multiplex polymerase chain reaction

Abstract

SummaryA multiplex polymerase chain reaction (PCR) was developed to detect the presence of theail, yst, andvirFgenes ofYersinia enterocoliticasimultaneously, quickly and accurately. The amplified fragment sizes were 356 base-pairs (bp) for theailgene, 134 bp for theystgene, and 231 bp for thevirFgene. The specificity of the amplified products was confirmed by hybridization with digoxigenin-labelled oligonucleotide probes. Amplification was successful whether the template was derived from a single colony of bacteria, aliquots of boiled bacterial suspensions, from DNA extracted from pure or mixed cultures or from stool specimens. Amplification of thevirFgene was also achieved from strains ofY. pseudotuberculosiscarrying the 70 kb plasmid but not with preparations from other relatedYersiniaspecies or from other members of the familyEnterobacteriaceae. The detection limit we established was 5–10 colony forming units per millilitre (cfu/ml) and 1·0 pg of DNA.