High‐affinity iron uptake is required for optimal Epichloë festucae colonization of Lolium perenne and seed transmission

<p dir="ltr"><i>Epichloë festucae</i> uses a siderophore-mediated system to acquire iron, which is important to maintain endophyte–grass symbioses. Here we investigate the roles of the alternative iron acquisition system, reductive iron assimilation (RIA), via disruption of the <i>fetC</i> gene, which encodes a multicopper ferroxidase, either alone (i.e., Δ<i>fetC</i>) or in combination with disruption of the gene <i>sidA</i>, which encodes a siderophore biosynthesis enzyme (i.e., Δ<i>fetC</i>/Δ<i>sidA</i>). The phenotypic characteristics of these mutants were compared to Δ<i>sidA</i> and wild-type (WT) strains during growth under axenic culture conditions (in culture) and in symbiosis with the host grass, perennial ryegrass (in planta). Under iron deficiency, the colony growth rate of Δ<i>fetC</i> was slightly slower than that of WT, while the growth of Δ<i>sidA</i> and Δ<i>fetC</i>/Δ<i>sidA</i> mutants was severely suppressed. Siderophore analyses indicated that Δ<i>fetC</i> mutants hyperaccumulate ferriepichloënin A (FEA) at low iron concentrations and ferricrocin and FEA at higher iron concentrations. When compared to WT, all mutant strains displayed hyperbranching hyphal structures and a reduced ratio of <i>Epichloë</i> DNA to total DNA in planta. Furthermore, host colonization and vertical transmission through infection of the host seed were significantly reduced in the Δ<i>fetC</i>/Δ<i>sidA</i> mutants, confirming that high-affinity iron uptake is a critical process for <i>Epichloë</i> transmission. Thus, RIA and siderophore iron uptake are complementary systems required for the maintenance of iron metabolism, fungal growth, and symbiosis between <i>E. festucae</i> and perennial ryegrass.</p>