Bovine pneumonic pasteurellosis is an economically significant infection of cattle caused by the Gram-negative bacterium Mannheimia haemolytica. Healthy cattle are often colonised by commensal serotype A2 strains but, for reasons which are not completely understood, disease is usually caused by a sudden explosive proliferation of serotype A1 strains within the upper respiratory tract (URT). In the present study, we have used a previously-optimised differentiated bronchial epithelial cell (DBEC) model of the bovine respiratory tract to compare the host-pathogen interactions of serotype A1 and A2 strains. The infection process was followed at various time-points for five days by bacterial enumeration, immunofluorescence and scanning electron microscopy of cultures, and light microscopy of histological sections. The innate pro-inflammatory immune response of the epithelial cells was also assessed by measuring the production of IL-1b, IL-6, CXCL8 (Il-8) and TNFa. The serotype A1 and A2 isolates behaved very differently and there was a strong correlation between events in vitro and typical behaviour in vivo. Bacterial numbers of the serotype A1 isolate increased significantly over the first 24 h of infection and microscopy demonstrated that this was due to invasion, rapid intracellular replication and subsequent spread of bacteria through the epithelial layer; tissue damage increased significantly from 24 h onwards. In contrast, bacterial numbers of the serotype A2 isolate did not increase and there were no signs of invasion; indeed, the epithelium remained healthy at day 5 and bacteria were mostly completely eliminated. Isolates of both serotypes stimulated the production of all four cytokines/chemokines in a manner that mimicked published in vivo data; notably, production of IL-1b and TNFa varied for the two strains. In conclusion, use of our DBEC model has identified a previously unknown invasion mechanism of M. haemolytica serotype A1 which provides a possible explanation for its sudden proliferation within the bovine URT.