Generic viability qPCR for monitoring shelf life of microbial biological control agents coated on seeds based on the nucleic acid intercalating dyes EMA and PMAxx

• Published in: Biological control 2024-09, Vol.196, p.105569, Article 105569
• Main Authors: Köhl, Jürgen, Groenenboom-de Haas, Lia, Houwers, Ilse, Elena, Georgina, de Lange, Ezra, Butterbach, Patrick, van der Heijden, Liesbeth
• Format: Article
• Language: English
• Subjects: Antagonists; Biological control; EMA; PMAxx; Seed coating; Seed pathogens; Viability qPCR
• ISSN: 1049-9644; 1090-2112
• DOI: 10.1016/j.biocontrol.2024.105569

[Display omitted] •Biological control of pathogens feasible through microbial seed treatments.•Poor survival of microbial inocula on seeds is a main bottleneck for implementation.•Viability of microorganisms coated on seeds assessed by viability (v)-qPCR.•V-qPCR will support screening of antagonists with better survival on seeds.•V-qPCR will support development of seed technologies for microbial seed coating. Biological control of seedborne pathogens and soilborne seedling pathogens through antagonists applied on seeds is an alternative to chemical seed treatments. Information on the viability of inocula on treated seeds is essential for any development and use of beneficial fungi or bacteria on seeds. Generic fungal and bacterial qPCR assays were combined with the nucleic acid intercalating dyes ethidium monoazide (EMA) and propidium monoazide (PMAxx) for the quantification of viable cells of fungi and bacteria on seeds. The applied protocols for generic fungal viability qPCR (v-qPCR) in combination with EMA and PMAxx and for generic bacterial v-qPCR in combination with PMAxx allowed the viability quantification of fungal and bacterial isolates representing a broad range of species with the exception of fungal species with highly melanized conidia. A first application of v-qPCR to coated seeds of onion and spinach indicated a differential plant species effect on survival of a coated fungus and a yeast with a generally better survival on seeds of spinach compared to seeds of onions and a similar good survival of the bacterium L. enzymogenes 3.1T8 on both seed types. The v-qPCR protocols can be applied in screening assays aiming at the selection of new antagonists with higher survival potentials and the development of new seed processing technologies compatible with coated antagonists.