Comparison of pulsed field gel electrophoresis and repetitive sequence polymerase chain reaction as genotyping methods for detection of genetic diversity and inferring transmission of Salmonella

Abstract

Pulsed field gel electrophoresis (PFGE) using restriction enzymes AvrII, SpeI, and XbaI, and repetitive sequence polymerase chain reaction (Rep-PCR) using BOX, ERIC, and REP primers, were compared with respect to their ability to detect genetic differences among 68 Salmonella isolates from nine Illinois swine farms. Both genotyping methods had high reproducibility of fragment numbers (reliability>0.9) and sizes (reliability>0.85), and produced approximately the same number of DNA fragments, but Rep-PCR fragment profiles had considerably greater variation. Genetic distances between isolates were calculated from fragment size matching. There was good agreement between the genetic distance matrices for the composite (3-enzyme and 3-primer) methods (Mantel’s r=0.83). PFGE detected slightly greater variation in genetic distances among isolates, but failed to differentiate seven pairs of isolates, three of which were sampled at least 1 month apart and therefore unlikely to be truly identical genetically. In contrast, Rep-PCR identified no isolates as genetically identical. In cluster analyses based on genetic distances, there were moderate differences between PFGE and Rep-PCR (about 2/3 agreement in tight cluster membership). Both PFGE and Rep-PCR were able to differentiate isolates of the same serotype. However, some serotypes (Agona, Anatum, Derby, Infantis, Worthington) were distributed across clusters. There was less agreement between individual primer/enzyme and composite results for Rep-PCR than for PFGE. This greater independence of results for individual primers for Rep-PCR accounted in part for the greater discriminative ability of the composite method. Both composite methods indicated that most Salmonella transmission occurred within a farm and that there was no preference for transmission between specific ecological compartments. Given the equally high reliability of both genotyping methods, the greater discriminative ability of Rep-PCR recommends it as the preferred method for precise detection of transmission links.