Quantification of the fungal pathogen Didymella segeticola in Camellia sinensis using a DNA-based qRT-PCR assay

The fungal pathogen Didymella segeticola causes leaf spot and leaf blight on tea plant (Camellia sinensis), leading to production losses and affecting tea quality and flavor. Accurate detection and quantification of D. segeticola growth in tea plant leaves are crucial for diagnosing disease severity...

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Bibliographic Details
Published in:Plant methods 2024-10, Vol.20 (1), p.157-9, Article 157
Main Authors: Zhang, You, Tu, Yiyi, Chen, Yijia, Fang, Jialu, Chen, Fan'anni, Liu, Lian, Zhang, Xiaoman, Wang, Yuchun, Lv, Wuyun
Format: Article
Language:English
Subjects:
Biochemical assays
C2H2-ZNF
Didymella segeticola
Disease progression
Diseases and pests
Distribution
DNA-based qRT-PCR
Environmental aspects
Fungi, Phytopathogenic
Genetic aspects
Genetic screening
Methods
Polymerase chain reaction
Quantification
Tea (Plant)
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Summary:The fungal pathogen Didymella segeticola causes leaf spot and leaf blight on tea plant (Camellia sinensis), leading to production losses and affecting tea quality and flavor. Accurate detection and quantification of D. segeticola growth in tea plant leaves are crucial for diagnosing disease severity or evaluating host resistance. In this study, we monitored disease progression and D. segeticola development in tea plant leaves inoculated with a GFP-expressing strain. By contrast, a DNA-based qRT-PCR analysis was employed for a more convenient and maneuverable detection of D. segeticola growth in tea leaves. This method was based on the comparison of D. segeticola-specific DNA encoding a Cys2His2-zinc-finger protein (NCBI accession number: OR987684) in relation to tea plant Cs18S rDNA1. Unlike ITS and TUB2 sequences, this specific DNA was only amplified in D. segeticola isolates, not in other tea plant pathogens. This assay is also applicable for detecting D. segeticola during interactions with various tea cultivars. Among the five cultivars tested, 'Zhongcha102' (ZC102) and 'Fuding-dabaicha' (FDDB) were more susceptible to D. segeticola compared with 'Longjing43' (LJ43), 'Zhongcha108' (ZC108), and 'Zhongcha302' (ZC302). Different D. segeticola isolates also exhibited varying levels of aggressiveness towards LJ43. In conclusion, the DNA-based qRT-PCR analysis is highly sensitive, convenient, and effective method for quantifying D. segeticola growth in tea plant. This technique can be used to diagnose the severity of tea leaf spot and blight or to evaluate tea plant resistance to this pathogen.
ISSN:1746-4811
1746-4811
DOI:10.1186/s13007-024-01284-2