Multicolor time - lapse imaging of immune damage to a fungal pathogen

Candida albicans is an opportunistic fungal pathogen that poses a serious threat to immunocompromised patients. Candida’s greatest mechanism of defense against the host immune system is its cell wall. It has been shown that chitin, a component of Candida’s cell wall, is synthesized during cellular stress. We investigated the temporal dynamics of chitin deposition during immune attack by performing ex vivo time-lapse and time-course experiments where we imaged Candida and PMNs. Concurrently, we measured local damage to the outer cell wall protein layer which allowed us to see that cell wall protein degradation and chitin deposition occurred simultaneously at sites of PMN attack, suggesting that PMNs cause localized cell wall damage that triggers cell wall remodeling. We were also interested in whether or not chitin deposition leads to β-glucan exposure or if the PMNs themselves cause β-glucan exposure. Previous work has shown that β-glucan becomes unmasked during murine infection. Our experiments showed that dead Candida cell segments have increased levels of β-glucan exposure only, suggesting that PMNs are directly causing β-glucan exposure on dead cells by degrading layers of the cell wall. In contrast, live Candida cell segments show co-localized areas of chitin deposition and β-glucan exposure suggesting that β-glucan exposure is a result of cell wall remodeling at sites of PMN attack. We believe that both mechanisms of β-glucan unmasking occur during infection, although further investigation is required to fully elucidate the temporal dynamics of β-glucan exposure and mechanistic aspects of PMN attack.