A species-specific PCR assay based on the calmodulin partial gene for identification of Fusarium verticillioides, F-proliferatum and F-subglutinans

Fusarium proliferatum, F. subglutinans and F. verticillioides are the most important Fusarium species occurring on maize world-wide, capable of producing a wide range of mycotoxins which are a potential health hazard for animals and humans. The ribosomal internal transcribed spacer and a portion of the calmodulin gene were sequenced and analysed in order to design species-specific primers useful for diagnosis. The primer pairs were based on a partial calmodulin gene sequence. Three pairs of primers (PRO1/2, SUB1/2 and VER1/2) produced PCR products of 585, 631 and 578 bp for F. proliferatum, F. subglutinans and F. verticillioides, respectively. Primer specificity was confirmed by analyzing DNA of 150 strains of these species, mostly isolated from maize in Europe and USA. The sensitivity of primers was 12.5 pg when the pure total genomic DNA of each species was analyzed. The developed PCR assay should provide a powerful tool for the detection of toxigenic fungi in maize kernels.