A genetically-defined riboflavin auxotroph of Actinobacillus pleuropneumoniae as a live attenuated vaccine

Actinobacillus pleuropneumoniae is a gram negative pleiomorphic rod that is the causative agent of a severe, highly infectious and often fatal pleurop neumonia in swine. We have previously reported the construction of genetica lly-defined stable riboflavin auxotrophs by replacement of a portion of the APP riboflavin biosynthetic operon (ribGBAH) with an antibiotic cassette e ncoding resistance to kanamycin, and have demonstrated that such riboflavin auxotrophs are avirulent. In this study, we evaluated riboflavin auxotroph s of A. pleuropneumoniae for their ability to stimulate protective immunity against pleuropneumonia. An initial challenge experiment demonstrated that intramuscular Vaccination with a live attenuated serotype 1A rib mutant, i n a vaccine formulation that included a limiting amount of exogenous ribofl avin, provided better protection against challenge with virulent A. pleurop neumoniae than either intratracheal immunization or intramuscular immunizat ion with live bacteria in the absence of exogenous riboflavin. Subsequent s tudies in which the vaccine inoculating dose, concentration of exogenous ri boflavin, and serotype of the vaccine strain were varied demonstrated that immunization with live avirulent riboflavin auxotrophs could elicit signifi cant protection against experimental challenge with both homologous and het erologous virulent serotypes of A. pleuropneumoniae. (C) 2000 Elsevier Scie nce Ltd. All rights reserved.