Clinical isolates of Mycobacterium avium complex reveal an M. intracellulare -associated IL-17/neutrophilic pulmonary immune program in a murine disease model

ABSTRACT Mycobacterium avium complex (MAC) is the leading cause of nontuberculous mycobacterial pulmonary disease (NTM-PD) and mainly comprises M. avium (MAV) and M. intracellulare (MI). Although MAV- and MI-associated pulmonary disease is currently treated using the same guideline-based regimens, whether MAC species and individual isolates elicit distinct pulmonary host responses remains unclear. Here, we established a panel of MAC clinical isolates exhibiting persistent pulmonary infection in mice and performed transcriptomic analyses to evaluate pulmonary immune responses. Although MAC infections induced broadly shared inflammatory responses, MI-infected lungs exhibited a robust IL-17/neutrophilic inflammatory signature, whereas MAV infection showed relative enrichment of IFN-γ-/cytotoxicity-associated responses, indicating differences in the balance of their immune gene expression programs. The MI-associated IL-17/neutrophilic program remained evident at the isolate level and after adjustment for pulmonary colony-forming unit (CFU) values. RT-qPCR and flow cytometric analyses using the representative isolate pair FKJ-1 (MI) and FKJ-8 (MAV) further supported the differential induction of these immune signatures in infected lungs. Single-cell RNA sequencing of lungs infected with FKJ-1 and FKJ-8 demonstrated that infection with the representative MI isolate was associated with increased Il17a -expressing CD4 + T cells and γδ T cells, neutrophilic inflammation, and the expansion of inflammatory macrophages. Together, these findings suggest that, within a selected panel of persistent MAC clinical isolates, MI infection is associated with a robust IL-17/neutrophilic pulmonary immune program. This study provides a preclinical framework for dissecting species- and isolate-associated host-pathogen interactions in MAC-PD. IMPORTANCE Mycobacterium avium complex pulmonary disease (MAC-PD) is an increasing clinical challenge with heterogeneous disease courses and outcomes. Although current guideline-based treatment does not differ between M. avium (MAV)- and M. intracellulare (MI)-associated PD, examining whether these species elicit distinct host responses may help explain heterogeneity in disease progression and clinical outcomes and support future patient stratification. We used a panel of MAC clinical isolates that persist or proliferate in mouse lungs to compare host responses induced by MAV and MI. Although both species induced broadly shared inflammation, MI infection was associated with a robust IL-17/neutrophilic program, whereas MAV infection showed relative enrichment of IFN-γ-/cytotoxicity-associated responses. The MI-associated program remained evident at the isolate level and after adjustment for pulmonary colony-forming unit (CFU) values. These findings suggest that MAC species and isolates may shape pulmonary immune landscapes, providing a preclinical framework for understanding MAC-PD heterogeneity.