Easy detection of siderophore production in diluted growth media using an improved CAS reagent

Siderophores are low molecular weight organic compounds produced by various microorganisms, especially pathogenic bacteria including rhizobacteria, and have a high affinity for iron. Although most microorganisms are thought to secrete siderophores under iron-depleted conditions, it is unclear how many microorganisms produce siderophores in the natural environment. Also, the chrome azurol sulfonate (CAS) assay, which is widely used for the detection of siderophores, needs to be improved for wider applicability. We developed a simple, high-throughput CAS assay in a 96-well microplate with a concentrated CAS reagent and commonly used diluted growth media in the absence of artificial iron depletion. The improved microplate CAS shuttle assay revealed that it could easily detect siderophores released from Pseudomonas (P.) fluorescence, P. putida, Burlkholderia stabilis, and Ottowia oryzae, as models of siderophore-producing bacteria. This CAS shuttle assay employed along with diluted growth media is a promising tool to screen new siderophore-producing bacteria. •Diluted growth media without iron-depleting treatment induce siderophore production of microbe.•Developed a simple, high-throughput CAS assay in a 96-well microplate with a CAS reagent and used diluted growth media•Confirmed that diluted growth media in the absence of artificial iron depletion induce siderophore production of microbe•We demonstrated that the diluted media can be applied to measure low concentrations of fluorescent siderophores.•Fluorescent siderophores were observed in new specie by our assay.