The quarantine pathogenic fungus Phyllosticta citricarpa is the causative agent of citrus black spot disease. Due to the risk of its spread to the European region, air and rain sampling is taking place in endangered orchards. The samples were analysed as part of this master's thesis, in which the presence of the fungus is monitored using an optimized DNA extraction protocol and qPCR tests. In addition to the control tests – 18S (Applied Biosystems), Yeast (Hierro et al., 2006), and FungiQuant (Liu et al., 2012) – the tests PC (van Gent-Pelzer et al., 2007) and PC-2 (Schirmacher et al., 2019) are used for specific fungal identification. Tests based on the ITS sequence are used to distinguish P. citricarpa from P. capitalensis and P. citriasiana, but not from P. paracitricarpa. Therefore, our goal was to establish and validate a new qPCR assay TEF1 that would allow this. With the set experiments, we demonstrated its strict specificity for P. citricarpa, its high sensitivity and robustness. We demonstrated the linearity of the test over a wide range and proved its repeatability by repeated experiments. It was evaluated as suitable for further use in diagnostics. We have analysed real air samples and found that the target pathogen is not present in them, which means that it has not yet spread to the European region.
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