Moving from qPCR to Chip Digital PCR Assays for Tracking of some <i>Fusarium</i> Species Causing <i>Fusarium</i> Head Blight in Cereals

• Main Authors: Caterina Morcia, Giorgio Tumino, Giulia Gasparo, Caterina Ceresoli, Chiara Fattorini, Roberta Ghizzoni, Paola Carnevali, Valeria Terzi
• Format: Article
• Language: English
• Published: MDPI AG 2020-08-01
• Series: Microorganisms
• Subjects: molecular diagnostics; <i>Fusarium</i>; chip digital PCR; qPCR
• Online Access: https://www.mdpi.com/2076-2607/8/9/1307

<i>Fusarium</i> Head Blight (FHB) is one of the major diseases affecting small-grain cereals, worldwide spread and responsible for severe yield and quality losses annually. Diagnostic tools, able to track <i>Fusarium</i> species even in the early stages of infection, can contribute to mycotoxins’ risk control. Among DNA-based technologies for <i>Fusarium</i> detection, qPCR (single and multiplex assays) is currently the most applied method. However, pathogen diagnostics is now enforced by digital PCR (dPCR), a breakthrough technology that provides ultrasensitive and absolute nucleic acid quantification. In our work, a panel of chip digital PCR assays was developed to quantify <i>Fusarium graminearum</i>, <i>F.culmorum</i>, <i>F. sporotrichioides</i>, <i>F. poae</i> and <i>F. avenaceum</i>. The primers/probes combinations were evaluated on pure fungal samples with cdPCR technique, in comparison with the qPCR approach. Moreover, the cdPCR assays were applied to quantify <i>Fusarium</i> in durum wheat and oat samples, naturally contaminated or spiked with fungal DNA. For a better evaluation of infection level in plants, duplex assays were developed, able to co-amplify both plant and fungal DNA. To the best of our knowledge, this is the first study directed to the application of digital PCR to <i>Fusarium</i> diagnosis in plants.