Construction of an isogenic leukotoxin deletion mutant of Pasteurella haemolytica serotype 1: Characterization and virulence

Allelic replacement was used to generate two isogenic IktA deletion mutants of Pasteurella haemolytica serotype 1 that were incapable of synthesizing leukotoxin (Lkt). Southern blot data confirmed that IktA sequences were absent in the two P. haemolytica deletion mutants. Culture supernatants and whole cell lysates from the wild type P. haemolytica, D153 parent strain, but not the IktA deletion mutants, contained immunoreactive and bioactive leukotoxic protein. In addition, only the parent strain was haemolytic when grown on bovine and sheep blood agar plates. Virulence of the IktA deletion mutant, IktA 77, was compared with the parent in an experimentally infected calf model of pneumonic pasteurellosis. Results revealed significant reduction in virulence in the IktA mutant as measured by clinical and lung lesion scores. Notable differences in histological changes such as markedly reduced necrosis and lack of leukocyte degeneration occurred in calves infected with the IktA mutant in comparison with those infected with the parent wild-type strain. Thus, it appears that leukotoxin plays a important role in the pathogenesis of lung injury in bovine pneumonic pasteurellosis.