Biology subjects

Adams, E. M.

Publications and source records attributed to Adams, E. M..

3 recordsLinked to original sources

bioRxiv2023

Aspergillus fumigatus hypoxia adaptation is critical for the establishment of fungal keratitis.

Purpose: The poor visual outcomes associated with fungal keratitis (FK) underscore a need to identify fungal pathways that can serve as novel antifungal targets. In this report, we investigated whether hypoxia develops in the FK cornea and, by extension, if fungal hypoxia adaptation is essential for virulence in this setting. Methods: C57BL/6j mice were inoculated with Aspergillus fumigatus and Fusarium solani var petroliphilum via topical overlay or instrastromal injection. At various time points post-inoculation (p.i.), animals we were injected with pimonidazole for the detection of tissue hypoxia through immunofluorescence imaging. The A. fumigatus srbA gene was deleted through Cas9-mediated homologous recombination and its virulence was assessed in the topical infection model using slit-lamp microscopy and optical coherence tomography (OCT). Results: Topical inoculation with A. fumigatus resulted in diffuse pimonidazole staining across the epithelial and endothelial layers within 6 h. Stromal hypoxia was evident by 48 h p.i., which corresponded to leukocytic infiltration. Instrastromal inoculation with either A. fumigatus or F. solani similarly led to diffuse staining patterns across all corneal cell layers. The A. fumigatus srbA deletion mutant was unable to grow at oxygen levels below 3% in vitro, and corneas inoculated with the mutant failed to develop signs of corneal opacification, inflammation or fungal burden. Conclusions: These results suggest that fungal antigen rapidly drives the development of corneal hypoxia, thus rendering fungal SrbA or related pathways essential for the establishment of infection. Such pathways may therefore serve as targets for novel antifungal intervention.

pathology↗

bioRxiv2023

Basal UPR activity in Aspergillus fumigatus regulates adaptation to nutrient stress and is critical for the establishment of corneal infection

The Aspergillus fumigatus unfolded protein response (UPR) is a two-component relay consisting of the ER-bound IreA protein, which splices and activates the mRNA of the transcription factor HacA. Spliced hacA accumulates under conditions of acute ER stress in vitro, and UPR null mutants are hypovirulent in a murine model of invasive pulmonary infection. In this report, we demonstrate that a hacA deletion mutant is completely unable to establish infection in a model of fungal keratitis, a corneal infection and an important cause of ocular morbidity and unilateral blindness worldwide. Contrary to our initial prediction, however, we demonstrate that hacA splicing is not increased above baseline conditions in the cornea, nor is the expression of genes classically associated with UPR activation, such as protein chaperones. We employed transcriptomics on wild-type and{Delta} hacA strains in gelatin media, as a proxy for the corneal environment, and found that hacA supports the expression of numerous primary and secondary metabolic processes that likely promote adaptation to nutrient limitation. Taken together, our results support a model in which the cornea, similar to growth on protein in vitro, is a source of sub-acute ER stress for A. fumigatus, but one nevertheless that requires the UPR pathway for proper adaptation. The data also suggest that this pathway could be a target for novel antifungals that improve visual outcomes for fungal keratitis patients. AUTHOR SUMMARYFungal keratitis has emerged as a leading cause of ocular morbidity and unilateral blindness worldwide. Relative to other infectious contexts, however, little is known about the fungal genes or pathways that regulate invasive growth and virulence in the corneal environment. In this report, we demonstrate that genetic disruption of the Aspergillus fumigatus unfolded protein response (UPR) abolishes the ability of the mold to establish infection in a mouse model of FK. Despite this critical role for virulence, however, we did not detect a concerted activation of the pathway beyond levels observed on standard medium, suggesting that the host environment is not an acute source of endoplasmic reticulum stress. Transcriptomic profiling of the wild-type and UPR-deficient strains under host-relevant nutrient conditions revealed a critical role for the pathway in regulating primary and secondary metabolism, cell wall biology, and mitochondrial function, all of which likely modulate fungal growth within and interactions with the host. These results expand our understanding of UPR regulation and function in this important mold pathogen and suggest the pathway could serve as a target for novel antifungals that improve visual outcomes in the setting of fungal keratitis.

microbiology↗

bioRxiv2022

Genomic and phenotypic trait variation of the opportunistic human pathogen Aspergillus flavus and its non-pathogenic close relatives

Fungal diseases affect millions of humans annually, yet fungal pathogens remain understudied. The mold Aspergillus flavus is a causative agent of both aspergillosis and fungal keratitis infections, but species closely related to A. flavus are not considered clinically relevant. To study the evolution of A. flavus pathogenicity, we examined genomic and phenotypic traits of two strains of A. flavus and three closely related non- pathogenic species: Aspergillus arachidicola (two strains), Aspergillus parasiticus (two strains), and Aspergillus nomiae (one strain). We identified over 3,000 orthologous proteins unique to A. flavus, including seven biosynthetic gene clusters present in A. flavus strains and absent in the three non-pathogenic species. We chose to characterize secondary metabolite production for all seven strains under two clinically relevant conditions, temperature and salt concentration. Temperature impacted metabolite production in all species. Conversely, we found a lack of impact of salinity on secondary metabolite production. Strains of the same species produced different metabolites. Growth under stress conditions revealed additional heterogeneity within species. Using the invertebrate model of fungal disease Galleria mellonella, we found virulence of strains of the same species varied widely, and A. flavus strains were not more virulent than strains of the non-pathogenic species. In a murine model of fungal keratitis, we observed significantly lower disease severity and corneal thickness for A. arachidicola compared to other species at 48 hrs, but not at 72 hrs. Our work identifies key phenotypic, chemical, and genomic similarities and differences between the opportunistic human pathogen A. flavus and its non-pathogenic relatives.

evolutionary biology↗