Widely dispersed clonal expansion of multi-fungicide-resistant Aspergillus fumigatus limits genomic epidemiology prospects

Invasive aspergillosis, caused by Aspergillus fumigatus, represents a critical public health concern, particularly due to increasing resistance to triazole antifungals linked to TR₃₄/TR₄₆ cyp51A haplotypes. In our genomic epidemiology study of 157 A. fumigatus isolates from Dutch environmental hotspots and two clinical centers, we identified near-identical genomes in several environmental and patient isolates, indicating a probable link. However, the geographic and temporal data alone are not sufficient to explain direct transmission pathways. Furthermore, a comparison with more than 1,200 globally sourced genomes revealed the extensive dissemination of certain clonal groups across multiple distant regions, raising significant challenges for the utility of genomic epidemiology. The discovery of high genetic diversity and the widespread distribution of some clonal groups challenges current understanding, suggesting that in most cases, tracing the precise source of individual infections will remain extremely difficult, even with increased sampling. In addition, we uncovered that the multi-triazole-resistant TR₃₄/TR₄₆ cyp51A haplotypes are associated with resistance to non-triazole fungicides such as benzimidazole, succinate dehydrogenase inhibitor, and quinone outside inhibitor classes, strongly suggesting an exposure history to multiple agricultural fungicides in these environmental hotspots. This resistance beyond the azole class suggests that strategies targeting only triazoles may be insufficient. Our findings challenge current paradigms and carry significant implications for One Health research and global public health strategies, underscoring the urgency of multidisciplinary approaches to tracking and monitoring fungal resistance.