Abstract Rationale Nontuberculous mycobacterial pulmonary disease (NTM-PD) frequently complicates chronic pulmonary aspergillosis (CPA) and shows a poorer prognosis. However, the mechanism by which NTM-PD tends to complicate remains unclear. We hypothesized that NTM and Aspergillus mutually provide favorable environments for each other’s proliferation within the host and for evading the host immune system, and we investigated the underlying mechanism of this interaction. Methods Culture supernatants from Mycobacterium avium (Mav; ATCC700737) and M. abscessus (Mab; ATCC19977) were applied to Af293 conidia to quantify biofilm formation using crystal violet assay, confocal microscopy, and scanning electron microscopy (SEM). To evaluate the effect of NTM infection on immunity to Aspergillus spp., THP-1-derived macrophages (PMA-differentiated) were infected with Mav (MOI 10) and co-cultured with Af conidia to assess phagocytosis (flow cytometry) and expression of pathogen recognition receptors/genes (qPCR, surface Dectin-1). Conversely, to evaluate the effect of Af infection on immunity to NTM, gliotoxin derived from Af on macrophage uptake of fluorescent Mav was tested. As an In vivo model, we used a co-infection model of NTM and Af. BALB/c mice received intratracheal Mav (Day 0 lung Af burden was quantified at 1 day, 1 and 2 weeks after Af infection. Results NTM supernatants significantly increased Af biofilm biomass and thickness with expansion of extracellular matrix; biofilms formed in NTM supernatants showed reduced susceptibility to voriconazole. In contrast, Af supernatant did not alter NTM growth. Prior Mav infection of THP-1 cells increased Af hyphal outgrowth and significantly reduced phagocytosis of Af conidia, accompanied by downregulation of Dectin-1 mRNA and surface expression (cell viability unchanged). Conversely, Gliotoxin inhibited THP-1 phagocytosis of Mav in a dose-dependent manner at non-cytotoxic concentrations. In the mousemouse model, prior Mav infection significantly delayed Af clearance and permitted hyphal morphologies histologically, whereas PBS controls predominantly showed cleared conidia. Conclusions Soluble factors (s) from NTM enhance Af biofilm and antifungal tolerance, while Mav infection suppresses macrophage antifungal recognition through Dectin-1 downregulation, reducing conidial phagocytosis. Conversely, Af-derived gliotoxins impair the phagocytosis of Mav by macrophages. These reciprocal interactions provide a mechanistic rationale for the high burden and poor outcomes of NTM-PD complicated with CPA and highlight potential therapeutic targets (biofilm-directed strategies, host-directed restoration of Dectin-1 signaling, and toxin neutralization). This abstract is funded by: None
Takazono et al. (Fri,) studied this question.