Phylogenetic and functional characterization of azole-resistant Aspergillus fumigatus clinical isolates in individuals with cystic fibrosis

Aspergillus fumigatus is a globally distributed fungal pathogen responsible for pulmonary aspergillosis in immunocompromised individuals, including those with asthma, cystic fibrosis (CF), and immune deficiencies. Its capacity to induce diseases ranging from allergies to life-threatening invasive infections stems from rapid adaptation to host microenvironments. This adaptability, a hallmark of its genetic plasticity, drives the emergence of phenotypically diverse strains under selective pressures. Such genetic variation underpins metabolic and phenotypic heterogeneity, including resistance to first-line azole antifungals, compromising treatment efficacy and worsening patient outcomes. This PhD study aimed to characterize the interplay between genomic and transcriptomic regulation of A. fumigatus metabolism by integrating phenotypic microarray metabolomics with multi-omics analyses. Whole-genome sequencing profiling of 145 clinical isolates (azole-resistant [ARAf] and azole-sensitive) revealed two phylogenetically distinct clades: Clade A (n = 43) and Clade B (n = 102), with distinct azole-resistance polymorphisms. Clade-specific genetic divergence reflects independent azole-resistance evolution driven by environmental or clinical pressures, with no significant influence from host endotypes. Phenotypic variation was assessed in 42 isolates across 379 growth conditions using the BioLog OmniLog® platform. Significant differences in 28 metabolic parameters were observed between azole-susceptible and ARAf isolates from CF and non-CF hosts. Notably, CF isolates exhibited reduced mean growth rates under stress, indicative of a fitness cost linked to adaptation to the CF lung niche. To illustrate adaptive mechanisms, genome-wide association studies (GWAS) and transcriptomic (RNA-seq) analyses were performed under baseline conditions and CF-relevant stressors (histamine and NaCl). GWAS identified 140 significant SNPs mapping to 88 genes, while RNA-seq revealed over 8,000 differentially expressed genes. Gene Ontology (GO) analysis identified 10 genes linked to metabolism and pathogenicity. Integrative multi-omics studies revealed AFUA_2G05620, a histamine catabolism regulator, as a key mediator of phenotypic plasticity, governing critical adaptive traits for survival in dynamic host environments.