Figure 2.

Model of NTHi Biofilm Strategies and Host-Pathogen Interactions Facilitating Chronic Infection in the COPD Airways. (A) The LuxS-RbsB AI-2 quorum sensing system facilitates coordination of intra-specific and inter-specific gene expression that promotes a transition from the planktonic to biofilm lifestyle in response to the local environmental conditions, leading to the downstream transcription of biofilm-associate genes. NTHi also express a QSeB two-component secondary system that may support biofilm gene expression coordination where the LuxS-RbsB system has been impaired. The quorum sensing system drives dissemination of biofilm bacteria to secondary sites, and formation of oxygen, nutrient and water channels towards the centre of the biofilm. (B) NTHi biofilms produce a thick protein- and eDNA-rich EPS matrix that provides a physical and chemical barrier, slowing penetration against pharmacological and immune system antimicrobial agents by blocking phagocytes and macrophages and chelating peptides and antibiotics such as β-defensin and β-lactams. The EPS matrix also provides structural stability and provides a site for mature biofilm structures to form, whilst also limiting metabolic activity, altering microenvironments, and driving persister cells through the development of gradients towards the centre of the biofilm. (C) NTHi express chemical defence strategies that allow for the evasion of the host’s NETosis response through recruitment of NETs into the EPS and expression of catalase that provides oxidative stress tolerance. Recognition by the immune system is also modulated through the differential expression of LOS and OMP P5 and OMP P2 that recruit neutrophils to form NETs but by evading the phagocytosing and cytotoxic mechanisms. NTHi biofilms further undergo a series of surface modifications that promote irreversible attachment, recruit secondary colonisers and impair recognition and/or clearance by immune system factors. These include sialyation, phosphorycholination, and the expression of TfP, OMPs and HMWs. (D) NTHi biofilms exhibit multidrug resistance due to upregulated carbohydrate metabolism and a preference for glycogen metabolism, as well as expressing antibiotic resistance genes including penicillin-binding protein 3 (PBP3) and β-lactamases. Negatively charged eDNA in the EPS matrix also sequesters positively charged antimicrobials. (E) Biofilms are seldom mono-species and exhibit complex inter-specific interactions with other inhabitants of the microbiome. In particular NTHi interacts with other COPD-associated pathogens including S. pneumoniae and M. catarrhalis that selects for hypoxia tolerant NTHi and damages cilia and mucociliary clearance, respectively, favouring NTHi biofilm formation. Tissue shown is the tracheobronchial respiratory mucosa, a pseudostratified, ciliated, columnar epithelium predominated by ciliated epithelial cells and interspersed by secretory cells (mucus-secreting goblet cells and secretoglobin secreting club cells) and basal cells. Created with BioRender.com.