Antifungal heteroresistance causes prophylaxis failure and facilitates breakthrough Candida parapsilosis infections.

Breakthrough fungal infections in patients on antimicrobial prophylaxis during allogeneic hematopoietic cell transplantation (allo-HCT) represent a major and often unexplained cause of morbidity and mortality. Candida parapsilosis is a common cause of invasive candidiasis and has been classified as a high-priority fungal pathogen by the World Health Organization. In high-risk allo-HCT recipients on micafungin prophylaxis, we show that heteroresistance (the presence of a phenotypically unstable, low-frequency subpopulation of resistant cells (~1 in 10,000)) underlies breakthrough bloodstream infections by C. parapsilosis. By analyzing 219 clinical isolates from North America, Europe and Asia, we demonstrate widespread micafungin heteroresistance in C. parapsilosis. Standard antimicrobial susceptibility tests, such as broth microdilution or gradient diffusion assays, which guide drug selection for invasive infections, fail to detect micafungin heteroresistance in C. parapsilosis. To facilitate rapid detection of micafungin heteroresistance in C. parapsilosis, we constructed a predictive machine learning framework that classifies isolates as heteroresistant or susceptible using a maximum of ten genomic features. These results connect heteroresistance to unexplained antifungal prophylaxis failure in allo-HCT recipients and demonstrate a proof-of-principle diagnostic approach with the potential to guide clinical decisions and improve patient care.

Epithelial responses to fungal pathogens.

Epithelial cells orchestrate immune responses against fungal pathogens. This review highlights advances in integrating epithelial cells in immune responses against inhaled molds and dimorphic fungi, and against Candida species that colonize mucosal surfaces. In the lung, epithelial cells respond to interleukin-1 (IL-1) and interferon signaling to regulate effector cell influx and fungal killing. In the alimentary and vulvovaginal tracts, epithelial cells modulate fungal commensalism, invasive growth, and local immune tone, in part by responding to damage caused by candidalysin, a C. albicans peptide toxin, and through IL-17-dependent release of antimicrobial peptides that contribute to Candida colonization resistance. Understanding fungal-epithelial interactions in mammalian models of disease is critical to predict vulnerabilities and to identify opportunities for immune-based strategies to treat fungal infections.

Oral bacteria relative abundance in faeces increases due to gut microbiota depletion and is linked with patient outcomes.

The detection of oral bacteria in faecal samples has been associated with inflammation and intestinal diseases. The increased relative abundance of oral bacteria in faeces has two competing explanations: either oral bacteria invade the gut ecosystem and expand (the 'expansion' hypothesis), or oral bacteria transit through the gut and their relative increase marks the depletion of other gut bacteria (the 'marker' hypothesis). Here we collected oral and faecal samples from mouse models of gut dysbiosis (antibiotic treatment and DSS-induced colitis) and used 16S ribosomal RNA sequencing to determine the abundance dynamics of oral bacteria. We found that the relative, but not absolute, abundance of oral bacteria increases, reflecting the 'marker' hypothesis. Faecal microbiome datasets from diverse patient cohorts, including healthy individuals and patients with allogeneic haematopoietic cell transplantation or inflammatory bowel disease, consistently support the 'marker' hypothesis and explain associations between oral bacterial abundance and patient outcomes consistent with depleted gut microbiota. By distinguishing between the two hypotheses, our study guides the interpretation of microbiome compositional data and could potentially identify cases where therapies are needed to rebuild the resident microbiome rather than protect against invading oral bacteria.

BTK drives neutrophil activation for sterilizing antifungal immunity.

We describe a previously-unappreciated role for Bruton's tyrosine kinase (BTK) in fungal immune surveillance against aspergillosis, an unforeseen complication of BTK inhibitors (BTKi) used for treating B-cell lymphoid malignancies. We studied BTK-dependent fungal responses in neutrophils from diverse populations, including healthy donors, BTKi-treated patients, and X-linked agammaglobulinemia patients. Upon fungal exposure, BTK was activated in human neutrophils in a TLR2-, Dectin-1-, and FcγR-dependent manner, triggering the oxidative burst. BTK inhibition selectively impeded neutrophil-mediated damage to Aspergillus hyphae, primary granule release, and the fungus-induced oxidative burst by abrogating NADPH oxidase subunit p40phox and GTPase RAC2 activation. Moreover, neutrophil-specific Btk deletion in mice enhanced aspergillosis susceptibility by impairing neutrophil function, not recruitment or lifespan. Conversely, GM-CSF partially mitigated these deficits by enhancing p47phox activation. Our findings underline the crucial role of BTK signaling in neutrophils for antifungal immunity and provide a rationale for GM-CSF use to offset these deficits in susceptible patients.

GM-CSF-mediated epithelial-immune cell crosstalk orchestrates pulmonary immunity to Aspergillus fumigatus.

Aspergillus fumigatus causes life-threatening mold pneumonia in immune compromised patients, particularly in those with quantitative or qualitative defects in neutrophils. While innate immune cell crosstalk licenses neutrophil antifungal activity in the lung, the role of epithelial cells in this process is unknown. Here, we find that that surfactant protein C (SPC)-expressing lung epithelial cells integrate infection-induced IL-1 and type III interferon signaling to produce granulocyte-macrophage colony-stimulating factor (GM-CSF) preferentially at local sites of fungal infection and neutrophil influx. Using in vivo models that distinguish the role of GM-CSF during acute infection from its homeostatic function in alveolar macrophage survival and surfactant catabolism, we demonstrate that epithelial-derived GM-CSF increases the accumulation and fungicidal activity of GM-CSF-responsive neutrophils, with the latter being essential for host survival. Our findings establish SPC + epithelial cells as a central player in regulating the quality and strength of neutrophil-dependent immunity against inhaled mold pathogens. HIGHLIGHTS: GM-CSF is essential for host defense against A. fumigatus in the lung IL-1 and IFN-λ promote GM-CSF production by lung epithelial cells in parallelEpithelial cell-derived GM-CSF increases neutrophil accumulation and fungal killing capacityEpithelial cells preferentially upregulate GM-CSF in local sites of inflammation.