Genetic map of the Actinobacillus pleuropneumoniae RTX-toxin (Apx) operons: Characterization of the ApxIII operons

Abstract

Actinobacillus pleuropneumoniae RTX-toxin III (ApxIII) is implicated as an important virulence factor of A. pleuropneumoniae, the causative agent of porcine pleuropneumonia. Recently, the genes coding for ApxIII (apxIIICA) of serotype 8 were cloned and characterized. The toxin appeared to be a member of the RTX-toxin family, as are the other two secreted toxins of A. pleuropneumoniae, i.e., ApxI and ApxII. In this report, we describe the cloning and sequencing of the remaining part of the ApxIII operon of serotype 8. This sequence coded for the RTX secretion proteins ApxIIIB and ApxIIID, which showed 86 and 63% similarity to ApxIB and ApxID, respectively, and 83 and 63% similarity to HlyB and HlyD of Escherichia coli, respectively. Potential functional domains, such as eight transmembrane regions and an ATP- binding cassette, were present in ApxIIIB. We examined the presence of apxIIICABD sequences in the 12 serotypes of A. pleuropneumoniae and found that these sequences were present only in serotypes 2, 3, 4, 6, and 8, the serotypes that secrete ApxIII. Comparison of the apxIIICABD gene sequences of the serotypes revealed very few serotype-specific differences. Only the C terminus of ApxIIIA of serotype 2 differed from ApxIIIA of the other serotypes. The differences were located between the glycine-rich repeats and the secretion signal. The analysis of the apxIIICABD genes completed our efforts to characterize the ApxI, ApxII, and ApxIII operons of the reference strains of the 12 serotypes of A. pleuropneumoniae. We present a complete map of the ApxI, ApxII, and ApxIII operons and discuss this in terms of gene expression and complementation and the role of the toxins in pathogenesis.