Aspergillus fumigatus In-Host HOG Pathway Mutation for Cystic Fibrosis Lung Microenvironment Persistence

The prevalence of Aspergillus fumigatus colonization in individuals with cystic fibrosis (CF) and subsequent fungal persistence in the lung is increasingly recognized. However, there is no consensus for clinical management of A. fumigatus in CF individuals, due largely to uncertainty surrounding A. fumigatus CF pathogenesis and virulence mechanisms. To address this gap in knowledge, a longitudinal series of A. fumigatus isolates from an individual with CF were collected over 4.5 years. Isolate genotypes were defined with whole-genome sequencing that revealed both transitory and persistent A. fumigatus in the lung. Persistent lineage isolates grew most readily in a low-oxygen culture environment, and conidia were more sensitive to oxidative stressinducing conditions than those from nonpersistent isolates. Closely related persistent isolates harbored a unique allele of the high-osmolarity glycerol (HOG) pathway mitogenactivated protein kinase kinase, Pbs2 (pbs2(C2)). Data suggest this novel pbs2(C2) allele arose in vivo and is necessary for the fungal response to osmotic stress in a low-oxygen environment through hyperactivation of the HOG (SakA) signaling pathway. Hyperactivation of the HOG pathway through pbs2(C2) comes at the cost of decreased conidial stress resistance in the presence of atmospheric oxygen levels. These novel findings shed light on pathoadaptive mechanisms of A. fumigatus in CF, lay the foundation for identifying persistent A. fumigatus isolates that may require antifungal therapy, and highlight considerations for successful culture of persistent Aspergillus CF isolates. IMPORTANCE Aspergillus fumigatus infection causes a spectrum of clinical manifestations. For individuals with cystic fibrosis (CF), allergic bronchopulmonary aspergillosis (ABPA) is an established complication, but there is a growing appreciation for A. fumigatus airway persistence in CF disease progression. There currently is little consensus for clinical management of A. fumigatus long-term culture positivity in CF. A better understanding of A. fumigatus pathogenesis mechanisms in CF is expected to yield insights into when antifungal therapies are warranted. Here, a 4.5-year longitudinal collection of A. fumigatus isolates from a patient with CF identified a persistent lineage that harbors a unique allele of the Pbs2 mitogen-activated protein kinase kinase (MAPKK) necessary for unique CF-relevant stress phenotypes. Importantly for A. fumigatus CF patient diagnostics, this allele provides increased fitness under CF lung-like conditions at a cost of reduced in vitro growth under standard laboratory conditions. These data illustrate a molecular mechanism for A. fumigatus CF lung persistence with implications for diagnostics and antifungal therapy.