Development of a multiplex PCR gene fingerprinting method using gyrA and pflA polymorphisms to identify genotypic...

The two genes gyrA and pflA, whose sequence variability had been previously described, were evaluated separately for their potential value in discriminating strains of Campylobacter jejuni. A single method was then developed by which the two loci were simultaneously amplified using a multiplex PCR procedure, and banding patterns were generated using a pre-selected set of restriction endonuclease enzymes. The method was applied to 18 strains of Camp. jejuni from different poultry sources varying in geographical origin and year of isolation. Results were combined and compared by means of numerical analysis with the classification obtained by flaA-typing and macrorestriction SmaI and KpnI. The usefulness of PCR fingerprinting of the gyrA/pflA genes for rapid ordering of strains by genotypic relatedness and providing additional information for estimating the degree of linkage between strains was demonstrated.