Genetic Diversity of Actinobacillus pleuropneumoniae Assessed by Amplified Fragment Length Polymorphism Analysis

Abstract

ABSTRACT Amplified fragment length polymorphism (AFLP) was evaluated as a method for genotypic characterization and subtyping within the bacterial species Actinobacillus pleuropneumoniae . A total of 155 isolates of A. pleuropneumoniae , representing the serotypic variation described to occur within this species, were analyzed. In order to elucidate the species boundaries, six strains of the phylogenetically closely related species Actinobacillus lignieresii were also included. Furthermore, the ability of AFLP to subtype was studied using 42 isolates of serovar 2 and the performance compared to that obtained by pulsed-field gel electrophoresis (PFGE). AFLP analysis provided a clear separation of A. lignieresii and A. pleuropneumoniae and divided the isolates of A. pleuropneumoniae into 20 clusters. Most of the serovars of A. pleuropneumoniae were represented by single and quite homogeneous clusters. The exceptions were serovars 10, K2:O7, and K1:O7, which were represented by two clusters each. In the cases where the serovars were represented by more than one cluster, the existence of these clusters was supported by additional phenotypic or genotypic properties. Furthermore, AFLP typing was able to allocate serologically nontypeable isolates to appropriate genetic groups within the species. Further investigations are needed to determine whether some of the clusters revealed through AFLP analysis represent additional serovars. When evaluated as a method for subtyping within serovar 2 of A. pleuropneumoniae , AFLP was found to achieve a degree of separation among isolates superior to that obtained by PFGE. However, a higher degree of separation between serovar 2 isolates was obtained by a combination of the two methods.