A culture-independent method for the direct detection in food of Yersinia spp. was developed in this study. It is based on the
amplification of a 359 bp PCR product from the RNA polymerase beta-subunit gene (rpoB) and subsequent analysis by
denaturing gradient gel electrophoresis (DGGE). Direct detection of Yersinia spp. by PCR–DGGE was carried out in ready-toeat
vegetables and the results compared with the results of the traditional, culture-dependent method. The DGGE profiles were
determined to be species-specific. As a matter of fact, Yersinia enterocolitica, Yersinia intermedia, Yersinia frederiskenii and
Yersinia kristensenii showed differential migrations in the gels. Moreover, Y. enterocolitica serotypes O:3, O:5 and O:9 were
distinguishable, as well. Only for a limited number of traditionally isolated strains, the biochemical and molecular identification
agree. In particular, an overestimation of Y. enterocolitica, as determined biochemically, was observed. Finally when the
protocol was applied to 27 food samples, a good correlation was obtained when the results of traditional and direct methods
were analyzed. The molecular method was able to identify Y. enterocolitica, not detected by plating analysis. However, for 4
samples, that, by plating analysis, were determined to contain Yersinia spp., no PCR product could be obtained after
enrichment, probably due to low numbers of target cells, thereby not allowing the possibility to perform DGGE analysis.
The protocol described here represents a reliable tool for the detection of Yersinia spp. in food, which can be used to obtain the
needed results faster than with traditional culturing methods.
