Analysis of Iron Requirements and Siderophore Production
This chapter describes the methods for inducing, detecting, and purifying the Legionella pneumophila siderophore. The first protocol details the methods by which L. pneumophila is cultured to facilitate production of the siderophore, rhizoferrin. This chapter then describes how to purify siderophore from culture supernatants through sequential reversed-phase/weak-anion exchange chromatography and high-performance liquid chromatography. The next section describes assays which allow the detection of the iron-binding capability and the biological activity of the purified siderophore. Lastly, this chapter describes the growth of L. pneumophila in chemically defined liquid medium (CDM) containing various iron sources as a method to assess the iron requirements of L. pneumophila.
Key wordsLegionella pneumophila CDM BYE BCYE Iron Siderophore HPLC Rhizoferrin Bioassay CAS assay Iron salts
This work was supported by NIH grant AI034937 awarded to NPC.