Investigating the influence of arachidonic acid on Candida albicans and its interaction with Pseudomonas aeruginosa
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Date
2020-01
Authors
Fourie, Ruan
Journal Title
Journal ISSN
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Publisher
University of the Free State
Abstract
Infection causes the release of arachidonic acid (AA) by the host that not only alters pathogen clearance by the immune system, but can also affect virulence and the antimicrobial susceptibility of pathogens. Furthermore, it may alter the interaction between co-infecting agents. Candida albicans exhibits multiple phenotypes and morphological plasticity, that are crucial to its virulence. Furthermore, it is known to form associations with commensal and co-infective bacteria in polymicrobial biofilms that can affect patient outcomes. The interkingdom interaction between C. albicans and the ubiquitous bacterium, Pseudomonas aeruginosa, can be regarded as a model for polymicrobial infection with detrimental cause to the host. The role that AA plays during this interaction is unknown. Arachidonic acid is not endogenously produced by C. albicans and P. aeruginosa, however, exogenously added AA has been reported to increase the susceptibility of C. albicans to antifungal agents, with reduction in virulence associated characteristics. The mechanism of this is unknown. To address this lack of knowledge, transcriptomic profiles of single species and polymicrobial biofilms, in the absence or presence of a sub-inhibitory concentration of AA, were compared. Focusing on C. albicans, genes of interest were identified, and homozygous deletion mutants were constructed. The roles of these genes were then evaluated during in vitro characterisation of biofilms as well as virulence in a Caenorhabditis elegans infection model. Regulatory mechanisms of C. albicans phenotypes and morphologies were evaluated in the presence of P. aeruginosa, along with their contribution to virulence during polymicrobial infection. Deletion of a component of the Set3/Hos2-histone deacetylase complex, involved in morphogenesis via chromatin remodelling, showed limited effect on the interaction between fungus and bacterium in vitro, but abrogated virulence, even in the presence of P. aeruginosa. Furthermore, deletion of WOR1, the principle regulator of phenotypic switching, did not modify population dynamics or virulence of C. albicans with P. aeruginosa. Genes associated with membrane organisation and antifungal resistance were shown to be induced by AA in C. albicans single species biofilms. Three genes, CDR1, IPT1 and RTA3, were chosen due to their importance in antifungal susceptibility. In addition to AA, co-incubation with P. aeruginosa induced their expression. However, only deletion of IPT1 was able to affect survival of C. albicans in the presence of P. aeruginosa. Furthermore, deletion of IPT1 and CDR1, but not RTA3, altered the virulence of C. albicans. Deletion of CDR1, a multi-drug transporter, increased oxidative stress, possibly due to lipid peroxidation, in the presence of AA. This may indicate that Cdr1p may be responsible for the efflux of AA. Although AA increased CDR1 mRNA and protein levels in a dose dependent manner, its function was diminished. This may be due to a combination of factors, including mislocalisation and subsequent loss of function of Cdr1p, reduced mitochondrial function, as well as competitive inhibition of AA with xenobiotic compounds. As Cdr1p provides antifungal resistance to C. albicans by efflux of antifungal agents, this provides a possible mechanism whereby AA increases antifungal susceptibility in C. albicans. This study addressed the influence of AA, a polyunsaturated fatty acid (PUFA), present in ample amounts during infection, on C. albicans. The influence of this fatty acid is frequently ignored during in vitro studies. Furthermore, the results obtained in this study prompts investigation into other PUFAs and their effects on pathogenic yeast, and their associations with pathobionts.
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Keywords
Thesis (Ph.D. (Microbial, Biochemical and Food Biotechnology))--University of the Free State, 2020, Arachidonic acid, Biofilms, Candida albicans, CRISPR-Cas9, Polymicrobial, Pseudomonas aeruginosa, Transcriptomics, Microbial - Biochemical - Food Biotechnology