Defective phagocyte association during infection of Galleria mellonella with Yersinia pseudotuberculosis is detrimental to both insect host and microbe.

Adhesins facilitate bacterial colonization and invasion of host tissues and are considered virulence factors, but their impact on immune-mediated damage as a driver of pathogenesis remains unclear. Yersinia pseudotuberculosis encodes for a multivalent adhesion molecule (MAM), a mammalian cell entry (MCE) family protein and adhesin. MAMs are widespread in Gram-negative bacteria and enable enteric bacteria to colonize epithelial tissues. Their role in bacterial interactions with the host innate immune system and contribution to pathogenicity remains unclear. Here, we investigated howY. pseudotuberculosis MAM contributes to pathogenesis during infection of the Galleria mellonella insect model. We show that Y. pseudotuberculosis MAM is required for efficient bacterial binding and uptake by hemocytes, the host phagocytes. Y. pseudotuberculosis interactions with insect and mammalian phagocytes are determined by bacterial and host factors. Loss of MAM, and deficient microbe-phagocyte interaction, increased pathogenesis in G. mellonella. Diminished phagocyte association also led to increased bacterial clearance. Furthermore, Y. pseudotuberculosis that failed to engage phagocytes hyperactivated humoral immune responses, most notably melanin production. Despite clearing the pathogen, excessive melanization also increased phagocyte death and host mortality. Our findings provide a basis for further studies investigating how microbe- and host-factors integrate to drive pathogenesis in a tractable experimental system.

Characterizing the Mechanisms of Nonopsonic Uptake of Cryptococci by Macrophages.

The pathogenic fungus Cryptococcus enters the human host via inhalation into the lung and is able to reside in a niche environment that is serum- (opsonin) limiting. Little is known about the mechanism by which nonopsonic phagocytosis occurs via phagocytes in such situations. Using a combination of soluble inhibitors of phagocytic receptors and macrophages derived from knockout mice and human volunteers, we show that uptake of nonopsonized Cryptococcus neoformans and C. gattii via the mannose receptor is dependent on macrophage activation by cytokines. However, although uptake of C. neoformans is via both dectin-1 and dectin-2, C. gattii uptake occurs largely via dectin-1. Interestingly, dectin inhibitors also blocked phagocytosis of unopsonized Cryptococci in wax moth (Galleria mellonella) larvae and partially protected the larvae from infection by both fungi, supporting a key role for host phagocytes in augmenting early disease establishment. Finally, we demonstrated that internalization of nonopsonized Cryptococci is not accompanied by the nuclear translocation of NF-κB or its concomitant production of proinflammatory cytokines such as TNF-α. Thus, nonopsonized Cryptococci are recognized by mammalian phagocytes in a manner that minimizes proinflammatory cytokine production and potentially facilitates fungal pathogenesis.