A rapid differentiation of Listeria monocytogenes by use of PCR-SSCP in the listeriolysin O (hlyA) locus

Single strand conformation polymorphism of PCR-amplified fragments (PCR-SSC P) was employed to develop a typing protocol for Listeria monocytogenes. Tw elve sets of PCR primers were designed to amplify fragments within the codi ng and non-coding region of hlyA locus. In parallel, PFGE analysis of ApaI and AscI digested L. monocytogenes DNA was performed to determine the numbe r of different genotypes and distribution of strains within serovar-subgrou ps. SSCP analysis of PCR generated amplicon K9 derived from the non-coding region of the hlyA gene revealed reproducible and highly polymorphic patter ns whereas other amplicons showed either monomorphic or 2 to 6 different pa tterns. Combining the results of all 12 primer pairs, 25 genotypes were obs erved in 39 strains representing seven serovars. Results were confirmed by PFGE typing, only two differences in the contribution to subgroups in serov ar 3b strains were observed. The data substantiate that the PCR-SSCP analys is is a reliable and highly discriminating method for characterizing L. mon ocytogenes strains on the molecular level. (C) 1999 Elsevier Science B.V. A ll rights reserved.