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Molecular genetics of Aspergillus pathogenicity
D W Holden1, C M Tang, J M Smith
1Department of Infectious Diseases and Bacteriology, Royal Postgraduate Medical School, Hammersmith Hospital, London, UK.
Abstract:
Aspergillus fumigatus is the most frequent cause of Invasive Pulmonary Aspergillosis (IPA), a life-threatening disease of immunosuppressed patients. In addition to a number of general physiological attributes of this fungus, it has been suggested that extracellular elastase and toxins might facilitate its growth in lung tissue. We have investigated the roles of two extracellular proteins, an alkaline protease with elastase activity (AFAlp), and the ribotoxin restrictocin in murine models of IPA. Gene disruption was used to create stable null mutant strains of the fungus lacking one or other protein, and their virulence and histopathological features were compared with an isogenic parental strain in steroid-treated and neutropenic mice. We have been unable to demonstrate any significant differences between the three strains, which shows that, considered independently, these proteins are not important virulence determinants. We are also interested in identifying fungal-specific gene products involved in general metabolism and which are required for growth in the lung, because these could represent new targets for antifungal drugs. For this work a model of murine IPA involving Aspergillus nidulans was established, to take advantage of the many well characterised mutations affecting metabolic pathways. Pathogenicity tests with strains carrying one of two auxotrophic mutations, lysA2 and pabaA1, have shown while lysine biosynthesis is not essential for the fungus to cause pulmonary disease, biosynthesis of p-aminobenzoic acid is essential. We are now in the process of cloning the A. fumigatus pabaA homologue to determine its function and whether this gene is required for growth of the clinically important species in the lung.
Insights
Extracellular elastase and restrictocin from Aspergillus fumigatus do not significantly impact invasive pulmonary aspergillosis (IPA) virulence. However, p-aminobenzoic acid biosynthesis is essential for fungal growth in the lung, indicating a potential drug target.
Area of Science:
- Mycology
- Infectious Diseases
- Medical Microbiology
Background:
- Aspergillus fumigatus is a primary cause of Invasive Pulmonary Aspergillosis (IPA), a severe lung infection in immunocompromised individuals.
- Extracellular virulence factors like elastase and toxins are hypothesized to aid fungal growth in lung tissue.
Purpose of the Study:
- To investigate the role of extracellular alkaline protease with elastase activity (AFAlp) and restrictocin in the virulence of Aspergillus fumigatus during IPA.
- To identify essential fungal metabolic genes required for growth in the lung as potential antifungal drug targets.
Main Methods:
- Generation of Aspergillus fumigatus null mutant strains lacking AFAlp or restrictocin using gene disruption.
- Comparison of virulence and histopathology of mutant strains versus the parental strain in murine models of IPA (steroid-treated and neutropenic mice).
- Establishment of a murine IPA model using Aspergillus nidulans with auxotrophic mutations (lysA2, pabaA1) to study metabolic pathway requirements.
Main Results:
- No significant differences in virulence or histopathology were observed between wild-type and mutant strains lacking AFAlp or restrictocin, indicating these proteins are not critical independent virulence factors.
- Lysine biosynthesis (lysA2) was not essential for pulmonary disease development in the Aspergillus nidulans model.
- p-aminobenzoic acid (PABA) biosynthesis (pabaA1) was found to be essential for fungal growth and pathogenicity in the murine lung.
Conclusions:
- Independently, extracellular elastase (AFAlp) and restrictocin are not major determinants of Aspergillus fumigatus virulence in IPA.
- p-aminobenzoic acid biosynthesis is a critical metabolic pathway for fungal survival and growth within the lung environment.
- The pabaA gene in Aspergillus fumigatus represents a promising target for the development of novel antifungal therapies.