Real-time PCR for quantification of toxigenic Fusarium species in cereals

Fusarium species are potential mycotoxin producers in cereals. In near future EU is going to set the maximum limit values for Fusarium toxins in raw cereals and cereal products. Mycotoxin analyses are expensive and time-consuming hence a rapid and reliable quantification method for toxigenic Fusarium species is needed in order to be able to evaluate the mycotoxin risk in cereal-based industry. We have applied real-time PCR technique for the quantification of trichothecene-producing Fusarium species present in barley and malt samples. PCR results were compared to the amount of trichothecenes determined in the samples. Furthermore, highly toxigenic Fusarium graminearum was quantified in cereals by real-time PCR. DNA was extracted from ground kernels using FastDNA Spin Kit for Soil and analysed in a LightCycler„¥ system using the double-stranded DNA ¡Vbinding fluorescent dye SYBR Green I or the fluorigenic TaqMan probes. Both naturally and artificially contaminated grains were analysed. PCR results correlated with the trichothecene content of the grain samples. Non-specific products such as primer dimers were amplified in the SYBR Green I assay, which interfered the quantification. Non-specific amplicons were not detected with the TaqMan probes. In addition, this assay was regarded as sensitive and reproducible.