Label-free detection of a bacterial pathogen using an immobilized siderophore, deferoxamine

Abstract

Pathogenic bacteria obtain the iron necessary for survival by releasing an iron chelator, termed a siderophore, and retrieving the iron-siderophore complex via a cell surface siderophore receptor. We have exploited the high affinity of Yersinia enterocolitica for its siderophore, deferoxamine, to develop a rapid method for capture and identification of Yersinia. In this methodology, a deferoxamine-bovine serum albumin conjugate is printed onto a gold-plated chip in a parallel line pattern. After flowing a suspension of Yersinia across the siderophore-derivatized chip, any Yersinia that binds to the chip is detected by dark-field microscopy analysis of the scattered light, followed by Fourier transform analysis of the scattering pattern. Since peak intensities are found to correlate with pathogen concentration, pathogen titers as low as 10³ cfu/ml can be readily detected. Moreover, immobilized deferoxamine can distinguish Y. enterocolitica, which binds ferrioxamine (deferoxamine-Fe), from Staphylococcus aureus, Mycobacterium smegmatis and Pseudomonas aeruginosa, which don't. Because human pathogens cannot easily mutate their iron retrieval systems without loss of viability, we suggest that few if any mutant Yersinia will emerge that can avoid detection. Together with previous results demonstrating selective capture of Pseudomonas aeruginosa by its immobilized siderophore (pyoverdin), these data suggest that pathogen-specific siderophores may constitute effective and immutable capture ligands for rapid detection and identification of their cognate pathogens.