ISOLATION OF PASTEURELLA-HAEMOLYTICA LEUKOTOXIN MUTANTS

Two mutants of Pasteurella haemolytica A1 that do not produce leukotox in were isolated. Following mutagenesis, colonies were screened with a ntiserum by a filter assay for absence of the secreted leukotoxin. The two mutants both appeared to produce normal amounts of other antigens , as judged by reactivity with polyclonal serum from an animal with pa steurellosis, and were not altered in beta-hemolytic activity as seen on blood agar plates. There was no evidence of either cell-associated or secreted leukotoxin protein when Western blots (immunoblots) were c arried out with the polyclonal serum or with a monoclonal antibody dir ected against the leukotoxin. Southern blots revealed that both mutant s show the wild-type restriction pattern at the leukotoxin locus, alth ough the strain with the lkt42 mutation showed differences in other re gions of the chromosome on analysis by pulsed-field gel electrophoresi s. The strain with the lktA2 mutation grew more slowly than did the wi ld-type strain, while the strain with the lktA1 mutation was indisting uishable from the wild-type strain in its growth properties. The strai n with the lktA1 mutation should be valuable in determining the role o f the leukotoxin in virulence as well as in identifying other virulenc e factors of P. haemolytica.