Detection of Fusarium culmorum in wheat by a surface plasmon resonance-based DNA sensor

A surface plasmon resonance (SPR) sensor based on DNA hybridization has been developed for the detection of Fusarium culmorum, a fungal pathogen of cereals. A 0.57 kbp DNA fragment of F culmorum was amplified by specific primers and a 25-mer oligomicleotide probe was selected within the sequence of the PCR amplicon. After biotmilation, the probe was immobilized on a streptavidin sensor chip and tested for biospecific interaction with PCR products of E culmorum. The effect of denaturating agents (formamide and urea) and ionic strength (NaCI) on hybridization efficiency of double-stranded PCR products with the immobilized probe and the specificity of the probe were investigated. The SPR biosensor was successfully used for the detection of E culmorum in culture material of different strains and in naturally infected wheat samples. Tested on fungal cultures, it showed a good selectivity for F culmorum against other species of either Fusaritini or other fungal genera. A background signal was observed in wheat samples strictly depending on the DNA amount of the testing matrix. Testing 30ng of durum wheat DNA the detection limit was 0.06pg of F culmorum DNA. The developed PCR-SPR assay allowed to detect F culmorum with sensitivity and specificity higher than gel-electrophoresis analysis. (c) 2006 Elsevier B.V. All rights reserved.