Author:
S Liu1#, Y Abu Yousef2,3#+, DC Sheppard2, 3, S Zhang1, F Le Mauff2,3*
Author address:
1Jiangsu Key Laboratory for Microbes and Functional Genomics, Nanjing Normal University, Nanjing, China
2Microbiology and Immunology, McGill University, Montreal, Canada
3Infectious Diseases and Immunity in Global Heath Program, Research Institute of the McGill University Health Center, Montreal, Canada
+Current address: McMaster University Medical School, Hamilton, Ontario, Canada
#Contributed equally to the work
Full conference title:
10th Advances Against Aspergillosis and Mucormycosis
Date: 2 February 2022
Abstract:
Purpose:
Adherence to cells is a key step in fungal pathogenesis. In Aspergillus fumigatus, hyphal adherence to host cells is mediated by the exopolysaccharide galactosaminogalactan (GAG). While several studies have identified genes whose product is required for GAG biosynthesis, little is known about the genetic regulation of GAG production and adherence.
Methods:
A library of 400 A. fumigatus transcription factor knock-outs was screened for their capacity to form adherent biofilms using the crystal violet assay. Transcription factor mutants with impaired biofilm-forming capacity were re-constructed to confirm the role of each candidate gene in the regulation of adherence. Mutants were then tested for potential growth defects by visual observation and XTT metabolic activity. GAG synthesis was quantified by ELISA and immunofluorescence microscopy. Cell wall composition was assessed by gas chromatography/mass spectrometry.
Results:
Out of 400 transcription factor knock-outs, 9 strains exhibited a reduction of > 50% in biofilm formation as compared with the parent strain Ku80. After reconstruction of the 9 deletions, the simultaneous study of biofilm adherence and growth allowed the classification of these mutant strains into 4 categories: 1 mutant had no growth defect and exhibited impaired formation of adherent biofilms throughout the growth period, 4 mutants had no growth defect, and reduced biofilm formation that could be restored with longer incubation, 3 mutants exhibited both a growth defect and a defect in biofilm formation that persisted despite prolonged incubation. Finally, 1 mutant displayed a severe germination defect and was excluded from further study. Interestingly, all strains except one produced both cell wall-associated or secreted GAG. Further studies of the cell wall polysaccharides in these mutants suggested a wider dysregulation of cell wall biosynthesis.
Conclusion:
This study highlights the role of several novel transcription factors in the regulation of A. fumigatus adherence and cell wall synthesis. Further, the inability of several of these strains to form adherent biofilms despite the production of GAG may provide insights into other GAG-interacting or independent factors required for fungal adhesion and biofilm formation. The
identification of these new adherence actors and their precise role may identify new therapeutic targets to prevent the development of A. fumigatus biofilms.
Abstract Number: 51
Conference Year: 2022
Link to conference website: https://aaam2022.org/
URL Conference abstract: