P366 Role of the Sit1 siderophore transporter in iron uptake, germination, and virulence of Mucor lusitanicus

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PMMucormycosis is a life-threatening systemic infection caused by certain members of the filamentous fungal order Mucorales (e.g., Rhizopus delemar, Lichtheimia corymbifera, and Mucor circinelloides). Mucormycosis is associated with a high mortality rate, which can be nearly 100% depending on the underlying condition of the patient. One of the main risk factors of mucormycosis has been the iron-chelating treatment. Iron is an essential nutrient for both the host and the pathogen. Therefore, organisms developed various strategies to acquire iron from the environment. Three strategies are available to microorganisms to uptake iron from the environment: acidification of the environment, reduction of ferric iron to the more soluble ferrous form, and secretion of soluble iron-chelating molecules. All fungi produce hydroxamate-type siderophore, except Mucorales fungi, which secrete a polycarboxylate siderophore, rhizoferrin. However, they also can utilize hydroxamate siderophores as xenosiderophores. In the Mucor genome database, one putative sit1 gene encoding a ferrioxamine B transporter, which belongs to the ARN/SIT subfamily of the major facilitator superfamily (MFS), coding was found. In Saccharomyces cerevisiae, Candia albicans, and Aspergillus fumigatus, siderophore-iron chelates are taken up through Arn3/Sit1 transporters. The relative transcript level of sit1 gene was measured after iron-starvation, and different siderophore treatments using quantitative real-time PCR. We have started to create and characterize (growth and germination ability, in vivo virulence assay, etc.) a sit1 knock-out mutant constructed using a CRISPR-Cas9 system. Sit1 gene showed significantly increased transcript levels using deferoxamine-iron, deferasirox-iron, and enterobactin-iron complexes. The colony diameter of the mutant strain was measured on Blood and Cas agar with (CasFe) or without iron (Cas-Fe). On CasFe agar, the growth ability of the mutant was significantly decreased compared to the control strain. In Galleria mellonella model, disruption of the sit1 gene resulted in decreased virulence. Relative transcript level of the putative rhizoferrin synthase gene (rfs) was measured in the sit1 knock-out mutant, where the gene showed an increased transcript level. Our result suggested that Sit1 is required for germination and virulence of Mucor lusitanicus and it may influence rhizoferrin production. This study was supported by the NKFI project K131796 and the grant ITM NKFIA TKP-2021-EGA-28. GN is grateful for the support of the Premium Postdoctoral Fellowship Program of the Hungarian Academy of Sciences (460 050).