Pathogenic Mannheimia haemolytica Invades Differentiated Bovine Airway Epithelial Cells

Abstract

The Gram-negative bacterium Mannheimia haemolytica is the primary bacterial species associated with bovine respiratory disease (BRD) and is responsible for significant economic losses to livestock industries worldwide. Healthy cattle are frequently colonized by commensal serotype A2 strains, but disease is usually caused by pathogenic strains of serotype A1. For reasons that are poorly understood, a transition occurs within the respiratory tract and a sudden explosive proliferation of serotype A1 bacteria leads to the onset of pneumonic disease. Very little is known about the interactions of M. haemolytica with airway epithelial cells of the respiratory mucosa which might explain the different abilities of serotype A1 and A2 strains to cause disease. In the present study, host-pathogen interactions in the bovine respiratory tract were mimicked using a novel differentiated bovine bronchial epithelial cell (BBEC) infection model. In this model, differentiated BBECs were inoculated with serotype A1 or A2 strains of M. haemolytica and the course of infection followed over a 5-day period by microscopic assessment and measurement of key proinflammatory mediators. We have demonstrated that serotype A1, but not A2, M. haemolytica invades differentiated BBECs by transcytosis and subsequently undergoes rapid intracellular replication before spreading to adjacent cells and causing extensive cellular damage. Our findings suggest that the explosive proliferation of serotype A1 M. haemolytica that occurs within the bovine respiratory tract prior to the onset of pneumonic disease is potentially due to bacterial invasion of, and rapid proliferation within, the mucosal epithelium. The discovery of this previously unrecognized mechanism of pathogenesis is important because it will allow the serotype A1-specific virulence determinants responsible for invasion to be identified and thereby provide opportunities for the development of new strategies for combatting BRD aimed at preventing early colonization and infection of the bovine respiratory tract.

Differences between predicted outer membrane proteins of genotype 1 and 2 Mannheimia haemolytica

“…It is also not known what the binding targets are for these proteins. Very recently, a report by Cozens et al reported that A1 strains of M. haemolytica, and not A2 strains, invade bovine bronchial epithelial cells through transcytosis, undergo rapid intracellular replication, and spread to adjacent cells, and that this mechanism might explain the proliferation of pathogenic M. haemolytica strains in animals versus commensal strains during times of stress [41]. It will be interesting to learn if the adhesins identified in this study, that are specific to pathogenic genotype 2 M. haemolytica, play a role in this newly identified process.…”

Differentiated ovine tracheal epithelial cells support the colonisation of pathogenic and non-pathogenic strains of Mannheimia haemolytica

“…It is therefore apparent that the host-specificity of M. haemolytica does not arise at the level of colonisation (at least within differentiated OTECs). As such, there appears to be an inherent lack of specificity associated with colonisation of OTECs in direct contrast to the selectivity we have observed for BBECs 28 , highlighting the potential for variation in the outcome of infection experiments performed using tissue culture models of different origins. It has been established that leukotoxin is the primary virulence factor of M. haemolytica contributing greatly to the fibrinous necrotising pneumonia observed in infected animals.…”

“…Thus, in vitro differentiated cattle and sheep airway epithelial cell culture models represent an excellent opportunity to study the molecular basis of such host-specificity. On comparison of M. haemolytica PH2 (a pathogenic bovine serotype A1 isolate) with PH202 (a non-pathogenic serotype A2 isolate) colonisation on differentiated BBECs, it was observed that only PH2 was capable of proliferation beyond 12 h post-infection, whereas PH202 was completely cleared from the majority of BBEC cultures 28 . We hypothesised that this may have been due to susceptibility of PH202 to BBEC-produced antimicrobial compounds such as defensins and cathelicidins, which are well-defined innate defence mechanisms of the respiratory epithelial cells 40,41 .…”

“…These models provide an excellent platform for the study of initial interactions between M. haemolytica and the respiratory epithelium. We have shown that differentiated bovine bronchial epithelial cells (BBECs) support the colonisation of pathogenic M. haemolytica 28 . In this study, we aimed to assess whether differentiated ovine tracheal epithelial cells (OTECs) displayed similar pathogen-specific colonisation.…”

Bovine Respiratory Syncytial Virus Enhances the Adherence of Pasteurella multocida to Bovine Lower Respiratory Tract Epithelial Cells by Upregulating the Platelet-Activating Factor Receptor

“…Notably, BRSV infection appears to facilitate bacterial adherence by modulating the expression of surface proteins on BRECs. To our knowledge, only a few studies have examined viral induction of changes in BRECs, despite the fact that cells of this type are the portals of entry for both viruses (Goris et al, 2009;Eberle et al, 2016) and bacteria (Cozens et al, 2019).…”