Oral Presentation Australian Society for Microbiology Annual Scientific Meeting 2024

Host adaptation imposes limitations on bacterial pathogens, as they need to survive long-term in a niche in which they also cause disease. Haemophilus influenzae is a human-adapted pathogen and can cause significant tissue damage and inflammation or persist asymptomatically in the human respiratory tract state for extended periods. Infections mediated by this highly successful pathogen cause annual healthcare costs in excess of A$890 million in Australia alone. However, while plenty of evidence exists for the transition of H. influenzae between asymptomatic and virulent stages, molecular details of the interactions between H. influenzae and human epithelial in either stage are scarce.

Using a novel, human primary cell-based infection model that closely resembles human nasal epithelia (NHNE), we report for the first time the molecular adaptations that take place in both H. influenzae and human epithelial cells during a 14-day infection.

Physiological assays combined with dualRNAseq revealed that the only large-scale changes in NTHi gene expression occurred within 24h of infection and were mostly due to metabolic adaptations, including upregulation of purine biosynthesis and pyrimidine uptake. In contrast, NHNE from five healthy donors all responded to H. influenzae infection with an initial, ‘unproductive’ inflammatory response that lasted ~48h and included a strong hypoxia signalling signature. Unexpectedly, this did not lead to significant production of pro-inflammatory cytokines. By 72h post-infection, an apparent tolerance to large burdens of H. influenzae developed, with NHNE transcriptional profiles resembling the pre-infection state. Tolerance development occurred in parallel with the development of intracellular bacterial populations and appears to involve interruption of NFkB signalling. This is the first time that large-scale, persistence-promoting immunomodulatory effects of H. influenzae during infection have been demonstrated. Interestingly, NHNE were able to re-activate pro-inflammatory responses towards the end of the 14-day infection, resulting in release of pro-inflammatory cytokines (IL8, TNFa). Our data also indicate the presence of infection stage-specific gene expression modules, highlighting fundamental similarities between immune responses in NHNE and canonical immune cells, which merit further investigation.