Quantitative Real-Time PCR for <i>Fusarium graminearum</i> sensu stricto and <i>F. asiaticum</i> Infection in Wheat Kernels

To develop effective measures for controlling Fusarium head blight and mycotoxin contamination in wheat, accurate quantification of Fusarium infection is essential. We developed a species-specific real-time PCR assay as a reliable quantification method for Fusarium graminearum sensu stricto (s. str.) and F. asiaticum infection in wheat. Two types of assays were evaluated: SYBR Green and TaqMan assays. Total DNA was extracted from wheat kernels harvested from a field where F. graminearum s. str. and F. asiaticum infection in wheat was artificially promoted, and EF1α gene copy numbers were quantified using both assays. Strong correlations were observed between the copy numbers obtained from the two assays (r = 0.97 for bulk samples), as well as between the EF1α gene copy number (SYBR Green assay) and ergosterol content (r = 0.88). These results indicate that real-time PCR provides a reliable method for quantifying F. graminearum s. str. and F. asiaticum infection. In particular, the primer and probe sequences prepared in this study were specific to F. graminearum s. str. and F. asiaticum, making the TaqMan assay suitable for quantifying infections by these species.