The developmental cycle of Dictyostelium discoideum ensures curing of a mycobacterial infection at both cell-autonomous level and by collaborative exclusion

During its life cycle, the social amoeba alternates between a predatory amoeba and a facultative multicellular form. The single-celled amoeba is a well-established model system to study cell-autonomous mechanisms of phagocytosis and defence against intracellular bacterial pathogens, whereas the multicellular forms are arising as models to study the emergence of innate immune defence strategies. Importantly, during evolution, prokaryotes have also evolved their own strategies to resist predation. Considering these complex ecological relationships, we wondered whether cells infected with intracellular pathogenic mycobacteria would be able to undergo their developmental cycle and what would be the fate of the infection. We show that the combination of cell-autonomous mechanisms and the organisation into a multicellular organism leads to the efficient multistep-curing of a mycobacteria-infected population, thereby ensuring germ-free spores and progeny. Specifically, using a microfluidic device to trap single infected cells, we revealed that in the first curing phase, individual cells rely on three mechanisms to release intracellular bacteria: exocytic release, ejection and lytic release. The second phase occurs at the collective level, when remaining infected cells are excluded from the forming cell aggregates

A microfluidic cell-trapping device for single-cell tracking of host-microbe interactions

The impact of cellular individuality on host-microbe interactions is increasingly appreciated but studying the temporal dynamics of single-cell behavior in this context remains technically challenging. Here we present a microfluidic platform, InfectChip, to trap motile infected cells for high-resolution time-lapse microscopy. This approach allows the direct visualization of all stages of infection, from bacterial uptake to death of the bacterium or host cell, over extended periods of time. We demonstrate the utility of this approach by co-culturing an established host-cell model, Dictyostelium discoideum, with the extracellular pathogen Klebsiella pneumoniae or the intracellular pathogen Mycobacterium marinum. We show that the outcome of such infections is surprisingly heterogeneous, ranging from abortive infection to death of the bacterium or host cell. InfectChip thus provides a simple method to dissect the time-course of host-microbe interactions at the single-cell level, yielding new insights that could not be gleaned from conventional population-based measurements