A multiplex direct PCR method for the rapid and accurate discrimination of three species of spider mites (Acari: Tetranychidae) in fruit orchards in Beijing

Can Lu ¹, Shao-Dong Hao ¹, Pa-Zi Ha ²

⁰Key Laboratory of Urban Agriculture, Ministry of Agriculture, College of Bioscience and Resource Environment, Beijing University of Agriculture, Beijing, 102206, North China, China.

Experimental & Applied Acarology +

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March 15, 2024

View abstract on PubMed

Summary

A new direct multiplex PCR method allows rapid, simultaneous identification of three key spider mite pests in Beijing orchards. This technique bypasses DNA extraction, significantly reducing time and cost for effective pest management.

Area Of Science

Agricultural Entomology
Molecular Diagnostics
Pest Management

Background

Spider mites (Tetranychidae) are significant agricultural pests, with species like Tetranychus urticae, Amphitetranychus viennensis, and Tetranychus truncatus posing threats to fruit production in Beijing.
Accurate and rapid identification of these spider mite species is crucial for effective pest control and quarantine measures in orchards.
Traditional identification methods are challenging due to limited morphological characteristics, and existing molecular techniques like PCR require time-consuming DNA extraction.

Purpose Of The Study

To develop a direct multiplex PCR method for the simultaneous identification of three common spider mite species: A. viennensis, T. truncatus, and T. urticae.
To provide a rapid and accurate diagnostic tool for differentiating these pest species in Beijing's orchards.
To offer technical support for phytosanitary measures and pest management strategies.

Main Methods

Designed three pairs of specific primers targeting the mitochondrial cytochrome c oxidase subunit I (COI) gene for T. urticae and T. truncatus, and the 18S gene for A. viennensis.
Optimized primer concentrations to establish a direct multiplex PCR system.
Validated the method for its ability to discriminate between the three species without prior DNA extraction and purification.

Main Results

Successfully established a direct multiplex PCR method for simultaneous identification of A. viennensis, T. truncatus, and T. urticae.
Achieved high sensitivity, with detection limits of 0.047 ng for T. truncatus and T. urticae DNA, and 0.0002 ng for A. viennensis DNA.
Eliminated the need for DNA extraction and sequencing, significantly reducing identification time and costs.

Conclusions

The developed direct multiplex PCR method provides a rapid, accurate, and cost-effective solution for identifying key spider mite pests in orchards.
This technique facilitates efficient pest monitoring and quarantine, supporting integrated pest management strategies.
The method holds potential for integration into field-based laboratory systems for real-time pest surveillance.

Keywords:

Amphitetranychus viennensis Zacher Tetranychus truncatus Ehara Tetranychus urticae Koch Direct multiplex PCR Species identification