Mtb likely interacts extensively with the alveolar lining fluid (ALF) – the thin layer of surfactant-rich liquid that coats alveolar cells. We are developing capabilities to deliver and monitor pulmonary surfactant trafficking at the air-liquid interface and obtain an integrative real-time perspective of how ALF regulates alveolar immunity and enables disease progression/control. This capability will also help identify how ALF could serve as an effective carrier of therapeutics or antigen cargoes directly to the distal lung, which could optimally stimulate tissue-resident immune responses. The LoC can speedily identify new cell-cell interactions in this open area of research that can then be followed up in animal models. 

 Mycobacterial cords are a unique form of biofilm formed from strong hydrophobic interactions, We want to understand the tribology of the interfaces between bacteria and use cutting-edge imaging to understand how the tight-packing of Mtb and Mycobacterium abscessus (Mabs) in cords alters the structure of the cell wall and spatial localization and function of cell wall proteins. These may uncover the presence of novel interbacterial communication mechanisms, and phenotypic specialization of bacteria in cords with consequent effects on antibiotic uptake and efflux. In addition to direct studies on bacteria, we are also applying techniques from synthetic biology to recreate Mtb lipid interactions  in controlled protein-free conditions outside the BSL-3.

Early experiments in the LoC model show that cords harbour antibiotic tolerant bacteria. We seek to understand the mechanistic basis for this observation by leveraging the LoC model to make precise and high spatiotemporal measurements of bacterial translational activity and growth, and correlating these observations with data from animal models. We aim to measure the local concentrations of antibiotics experienced by individual bacteria and identify whether inhibitors of cord formation are effective in the tissue context in speeding up clearance. 

Our recent work has shown a prominent role for Mtb cord formation in first contact alveolar macrophages and epithelial cells. We aim to identify how cords enable new intracellular interactions of Mtb virulence proteins with host organelle membranes (mitochondria, nucleus) that can influence bacterial control and extracellular interactions with the plasma membrane that cause contact-dependent cell death. This interdisciplinary approach will seamlessly identify how mechanobiology and mechanobiology-enabled new functionality for virulence factors enables evasion from immune sensing. Similar mechanisms are likely to operate in extracellular Mabs biofilms and in intracellular bacterial communities (IBCs) in urinary tract infections. 

Organ-chip models have a long runway ahead for development of complexity, functionality and readout. We aim to combine organoids with organ-chips to generate more mature alveolar epithelial cells in co-culture with functional alveolar-like macrophages, using cells from healthy individuals or persons with co-morbidities that can impact disease. We also seek to incorporate additional tissue-resident immune cells to model later stages of infection, and optics-compatible biophysical measurements of tissue function that are compatible with BSL-3 conditions.