F-025. Deletion of the Aspergillus fumigatus Protein Kinase A (PKA) Regulatory Subunit, pkaR, Decreases Virulence in a Murine Model of Chronic Granulomatous Disorder

J. R. Fortwendel, R. S. Gibbons, H. L. Allen, G. S. Deepe, S. L. Newman, D. S. Askew, J. C. Rhodes;
Univ. of Cincinnati, Cincinnati, OH.

Background: Chronic Granulomatous Disorder (CGD) patients produce phagocytes that are deficient in their ability to undergo the oxidative burst process, leading to an increased risk of infection with microorganisms. Aspergillus fumigatus is an opportunistic fungal pathogen that, despite improvements in anti-fungal therapy, is a major cause of morbidity and mortality in individuals with CGD. Deletion of pkaR, the cAMP-dependent PKA regulatory subunit, has demonstrated an important role for PKA activity in regulating morphology, virulence, and oxidative-stress response in A. fumigatus. In a pilot study involving a CGD mouse model, ΔpkaR conidia were found to be reduced in virulence, implicating the PKA pathway as important to the development of disease in the CGD patient. We hypothesized that PKA pathway activity contributes to the non-oxidative stress response in A. fumigatus and therefore decreases virulence in the CGD mouse model. Methods: Animal survival studies and peritoneal polymorphonuclear leukocyte (PMN) extractions employed C57/BL6 and Cybb mice as the wild type (wt) and murine CGD model, respectively. Fungal growth (CMFDA) and viability (XTT/menadione) assays were preformed as previously published. Results: CGD mice displayed a significantly higher survival rate when infected with the ΔpkaR strain (58% survival) when compared to wt (0% survival). Interestingly, dissemination was kept to a minimum in CGD mice, with occurrence of extra-lung organ involvement occurring in no mice infected with ΔpkaR and only 8% infected with wt and complemented strains. Lesion size and inflammation caused by infection with the ΔpkaR strain were greatly decreased as compared to infection with wt. In addition, there was a trend towards increased susceptibility of ΔpkaR hyphae to PMN extracted from CGD mice. When examined further, ΔpkaR hyphae displayed a small, but consistent, increase in susceptibility to lactoferrin, a non-oxidative component released by PMN. Conclusions: Our results suggest that deregulation of PKA pathway activity decreases virulence in CGD mice and increases susceptibility of A. fumigatus hyphae to lactoferrin.