Interactions between bacteria and fungi in polymicrobial biofilms influence the viability and global transcriptome of the interacting organisms that can affect the pathogenic potential of these biofilms

Abstract

Polymicrobial biofilms are often the cause of chronic infections and are resistant to treatment. The structure of biofilms increases their resistance to antibiotics thereby increasing the severity of infection. Rates of morbidity and mortality are high in immunocompromised patients who have been infected with polymicrobial biofilms. Thus, there is an urgent need to understand the nature of the interactions between microbial species in polymicrobial infections to better develop drug targets and treat infections in immunocompromised populations. In this thesis I set out to address the question ‘how do pathologically relevant bacteria and fungi interact?’. My aim was to investigate interactions between fungi and bacteria to identify if they could form polymicrobial biofilms. The mechanisms employed in antagonistic interactions between microbes have the potential to allow the identification of novel antimicrobial agents. Pathways identified as important in positive interactions between bacteria and fungi could be potential drug targets. I investigated the interactions between fungal and bacterial pathogens co-localised from lungs and gut, and their capacity to form a polymicrobial biofilm. Co-culture assays identified that Candida albicans was not viable post interaction with Escherichia coli, and Enterobacter aerogenes, while Staphylococcus aureus and Aspergillus fumigatus biomass were both stimulated post interaction by co-culture with one another. Confocal scanning laser microscopy (CLSM) analysis of polymicrobial biofilms, using fluorescent stains SYTO 9 (bacteria), Calcofluor white (fungus), and Concanavalin A Texas red conjugate (extracellular matrix ECM), revealed a close association between S. aureus and A. fumigatus cells as well as a statistically significant stimulation of hyphal development. CLSM analysis also allowed observation of a statistically significant inhibition of hyphal development in A. fumigatus cultured with P. aeruginosa. Transcriptome analysis was conducted on S. aureus and A. fumigatus polymicrobial culture using RNAseq. Upregulation of biofilm associated genes was observed in both species indicating biofilm formation was stimulated. Iron uptake stimulates the production of a biofilm in A. fumigatus, the observed upregulation of sidE and down regulation of ftrA suggested that siderophore-mediated iron uptake had been induced. Cell proliferation, metal binding genes, and the autolysin lytM, all of which are linked to increased virulence and pathogenesis were upregulated in S. aureus. The RNAseq profile of S. aureus in combination with A. fumigatus suggested that within a host, virulence of the bacteria would be enhanced.

Date of Thesis submission	2023
Original language	English
Awarding Institution	Western Sydney University
Supervisor	Oliver Morton (Supervisor), Thomas Jeffries (Supervisor) & Michael Phillips (Supervisor)