The RNA interference (RNAi) pathway has evolved numerous functionalities in eukaryotes, with many on display in Kingdom Fungi. RNAi can regulate gene expression, facilitate drug resistance, or even be altogether lost to improve virulence potential in some fungal pathogens. In the WHO fungal priority pathogen, Aspergillus fumigatus, the RNAi system is known to be intact and functional. To extend our limited understanding of A. fumigatus RNAi, we performed a multi-condition mRNA-seq analysis comparing expression of several RNAi double knockout mutants with the wild-type strain in conidia and mycelium grown for 24 or 48 hours. The analysis linked the A. fumigatus dicer-like enzymes and RNA-dependent RNA polymerases to regulation of conidial ribosome biogenesis. Cumulatively, A. fumigatus RNAi appears to play an active role in defense against double-stranded RNA species alongside a previously unappreciated housekeeping function in regulation of conidial ribosomal biogenesis genes. Overall design: Comparative gene expression profiling analysis of poly(A) RNA-seq data for double knockouts (?dclA/B, ?ppdA/B, ?rrpA/B) in the RNAi pathway of Aspergillus fumigatus conidia, 24-h-mycelium, and 48-h-mycelium.