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Real-time PCR for specific detection of three phytoplasmas from the apple proliferation group
Nataša Mehle1, Petra Nikolić, Kristina Gruden
1National Institute of Biology, Ljubljana, Slovenia. natasa.mehle@nib.si
Abstract:
In this chapter, we describe a real-time PCR detection system for fast, reliable, specific, and sensitive detection and discrimination of 'Candidatus Phytoplasma mali', 'Ca. P. prunorum', and 'Ca. P. pyri' from the 16SrX (apple proliferation-AP) group. These phytoplasmas are causal agents of fruit tree diseases within the Rosaceae family, namely apple proliferation, European stone fruit yellows, and pear decline. The assays use (hydrolysis) TaqMan(®) minor groove binder probes. The panel of assays comprises the same set of primers and specific probes for species-specific amplification, and an additional set of primers and probe for 18S rRNA as an endogenous quality control of DNA extraction. The assays described can be used in routine phytoplasma surveys and in certification programmes.
Insights
This study presents a real-time PCR system for rapid and accurate detection of three key phytoplasmas affecting fruit trees. The developed assay ensures reliable identification for disease management and certification programs.
Area of Science:
- Plant Pathology
- Molecular Biology
- Agricultural Science
Background:
- Phytoplasmas in the 16SrX group cause significant diseases in Rosaceae fruit trees, including apple proliferation, European stone fruit yellows, and pear decline.
- Accurate and rapid detection methods are crucial for managing these economically important plant diseases.
Purpose of the Study:
- To develop and validate a real-time PCR detection system for specific identification of 'Candidatus Phytoplasma mali', 'Ca. P. prunorum', and 'Ca. P. pyri'.
- To provide a reliable tool for routine phytoplasma surveys and certification programs.
Main Methods:
- Utilized real-time PCR with TaqMan® minor groove binder probes for species-specific amplification.
- Employed a common set of primers and probes for the target phytoplasmas.
- Incorporated an 18S rRNA assay as an endogenous control for DNA extraction quality.
Main Results:
- The developed real-time PCR assays demonstrated fast, reliable, specific, and sensitive detection and discrimination of the target phytoplasmas.
- The system effectively identified 'Candidatus Phytoplasma mali', 'Ca. P. prunorum', and 'Ca. P. pyri'.
Conclusions:
- The described real-time PCR system is a valuable tool for the accurate diagnosis of major phytoplasma diseases in fruit trees.
- This method supports routine phytoplasma surveillance and enhances the reliability of certification programs for fruit tree health.
