The mammalian immune system relies on recognition of pathogen surface antigens for targeting and clearance. In the absence of immune evasion strategies, pathogen clearance is rapid. In the case of Aspergillus fumigatus, the successful fungus must avoid phagocytosis in the lung to establish invasive infection. In healthy individuals, fungal spores are cleared by immune cells; however, in immunocompromised patients, clearance mechanisms are impaired. Here, using proteome analyses, we identified CcpA as an important fungal spore protein involved in pathogenesis. A. fumigatus lacking CcpA was more susceptible to immune recognition and prompt eradication and, consequently, exhibited drastically attenuated virulence. In infection studies, CcpA was required for virulence in infected immunocompromised mice, suggesting that it could be used as a possible immunotherapeutic or diagnostic target in the future. In summary, our report adds a protein to the list of those known to be critical to the complex fungal spore surface environment and, more importantly, identifies a protein important for conidial immunogenicity during infection.

Aspergillus (A.) fumigatus is an opportunistic fungal mold inducing invasive aspergillosis (IA) in immunocompromised patients. Although antifungal activity of human natural killer (NK) cells was shown in previous studies, the underlying cellular mechanisms and pathogen recognition receptors (PRRs) are still unknown. Using flow cytometry we were able to show that the fluorescence positivity of the surface receptor CD56 significantly decreased upon fungal contact. To visualize the interaction site of NK cells and A. fumigatus we used SEM, CLSM and dSTORM techniques, which clearly demonstrated that NK cells directly interact with A. fumigatus via CD56 and that CD56 is re-organized and accumulated at this interaction site time-dependently. The inhibition of the cytoskeleton showed that the receptor re-organization was an active process dependent on actin re-arrangements. Furthermore, we could show that CD56 plays a role in the fungus mediated NK cell activation, since blocking of CD56 surface receptor reduced fungal mediated NK cell activation and reduced cytokine secretion. These results confirmed the direct interaction of NK cells and A. fumigatus, leading to the conclusion that CD56 is a pathogen recognition receptor. These findings give new insights into the functional role of CD56 in the pathogen recognition during the innate immune response.Invasive aspergillosis (IA), primarily caused by the mold Aspergillus fumigatus, is a devastating disease in immunocompromised patients suffering from hematological malignancies or undergoing allogeneic hematopoietic stem cell transplantation (HSCT) 1 . The mortality rate of HSCT patients diagnosed with IA ranges from 60-90% 2 and the prognosis for long-term survival is extremely poor 3 . Recently, it was shown that HSCT patients with probable/proven IA had a delayed reconstitution of natural killer (NK) cells for more than a year post HSCT 4 . In addition, patients with severe IA were found to have a lower NK cell count compared to patients with well-controlled IA, suggesting that NK cells play a critical role in immunity to IA.NK cells comprise 5-15% of the peripheral blood mononuclear cells (PBMCs) in healthy individuals and belong to the innate immune system 5 . Upon activation, NK cells release immune regulatory cytokines to stimulate other immune cells and display cytotoxicity directed against tumor or virus-infected cells by granule release 5 . NK cells are defined as CD56 positive and CD3 negative cells and can be distinguished into CD3 − CD56 dim CD16 + and CD3 − CD56 bright CD16 − cells. While CD56 dim cells are more cytotoxic, CD56 bright cells produce high levels of cytokines such as IFNγ and TNFα 6 . The function of NK cells is induced by the interplay of inhibitory and activating receptors 7 , leading to cytotoxicity directed against tumors and virus-infected cells. Besides the recognition of these cells, NK cells also recognize other infectious pathogens, become activated, and as a response induce either lysis of these pathogens or trigger activation of other...

Highlights d Triple RNA-seq measures gene expression of co-infected immune cells d Gene correlation networks reveal different hub gene sets under co-infection d Co-infection expression includes synergies and interferences between host and pathogens d Molecular basis of viral/fungal pulmonary infection has potential for the clinic Authors

Aspergillus fumigatus is an opportunistic fungal pathogen causing detrimental infections in immunocompromised individuals. Dendritic cells (DCs) are potent antigen-presenting cells and recognize the A. fumigatus cell wall component β-1,3 glucan via Dectin-1, followed by DC maturation and cytokine release. Here, we demonstrate that human primary myeloid DCs (mDCs) interact with different morphotypes of A. fumigatus. Dectin-1 is expressed on mDCs and is down-regulated after contact with A. fumigatus, indicating that mDCs recognize A. fumigatus via this receptor. Blocking of Dectin-1, followed by stimulation with depleted zymosan diminished the up-regulation of the T-cell co-stimulatory molecules CD40, CD80, HLA-DR and CCR7 on mDCs and led to decreased release of the cytokines TNF-α, IL-8, IL-1β and IL-10.

Dendritic cells (DCs) are antigen presenting cells which serve as a passage between the innate and the acquired immunity. Aspergillosis is a major lethal condition in immunocompromised patients caused by the adaptable saprophytic fungus Aspergillus fumigatus . The healthy human immune system is capable to ward off A. fumigatus infections however immune-deficient patients are highly vulnerable to invasive aspergillosis. A. fumigatus can persist during infection due to its ability to survive the immune response of human DCs. Therefore, the study of the metabolism specific to the context of infection may allow us to gain insight into the adaptation strategies of both the pathogen and the immune cells. We established a metabolic model of A. fumigatus central metabolism during infection of DCs and calculated the metabolic pathway (elementary modes; EMs). Transcriptome data were used to identify pathways activated when A. fumigatus is challenged with DCs. In particular, amino acid metabolic pathways, alternative carbon metabolic pathways and stress regulating enzymes were found to be active. Metabolic flux modeling identified further active enzymes such as alcohol dehydrogenase, inositol oxygenase and GTP cyclohydrolase participating in different stress responses in A. fumigatus . These were further validated by qRT-PCR from RNA extracted under these different conditions. For DCs, we outlined the activation of metabolic pathways in response to the confrontation with A. fumigatus . We found the fatty acid metabolism plays a crucial role, along with other metabolic changes. The gene expression data and their analysis illuminate additional regulatory pathways activated in the DCs apart from interleukin regulation. In particular, Toll-like receptor signaling, NOD-like receptor signaling and RIG-I-like receptor signaling were active pathways. Moreover, we identified subnetworks and several novel key regulators such as UBC, EGFR, and CUL3 of DCs to be activated in response to A. fumigatus . In conclusion, we analyze the metabolic and regulatory responses of A. fumigatus and DCs when confronted with each other.