The use of a three-dimensional cell culture model to investigate host-pathogen interactions of Francisella tularensis in human lung epithelial cells.

Francisella tularensis inhalation results in bacterial interaction with numerous lung cell types, including those of the epithelium. This work investigates a three-dimensional cell-culture system to characterise the epithelial response to F. tularensis. Immortalised human pneumocytes (A549s) grown in rotating-wall vessel (RWV) bioreactors display an in vivo-like phenotype, which has been confirmed to be driven by specific transcriptional events (8454 genes, p ≤ 0.05). These data support the RWV model as a more in vivo-like culture system to investigate the lung epithelium, compared to monolayer counterparts. RWV-cultured A549s were infected with F. tularensis SchuS4 and LVS and intracellular replication mapped over 22 h compared to monolayer cells. The RWV-cultured A549s were more resistant to SchuS4 and LVS infection (p ≤ 0.05). Transcriptomics identified 2086 genes (p ≤ 0.05) as candidates for host-pathogen interactions which result in the observed increase in resistance of the RWV-cultured A549 cells. Gene and pathway analysis identified processes involved in MMP modulation, endocytosis, mucin production and the complement pathway amongst others. The role of these pathways during infection was further characterised using chemical inhibitors. This work has revealed several new hypotheses worthy of further testing in order to understand the epithelial host response to F. tularensis infection.