Melon Fly-LAMP: Bringing Molecular Pest Detection from Lab to Land

By Suman Barman and Shashank P. R.

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Fruit flies (Diptera: Tephritidae) are among the most destructive insect pests affecting fruits and vegetables across the world. With more than 5,000 species recorded globally and nearly 200 of them causing serious economic losses, fruit flies pose a constant threat to agriculture, food security, and international trade. These insects cause significant damage as they eat and grow inside infested fruits, often crossing borders unnoticed. The rapid growth of global trade, human travel, and climate change has further increased the spread of invasive fruit fly species into new regions. A major challenge in managing fruit flies is accurate species identification. Many economically important species look extremely similar, particularly during the egg, larval, and pupal stages, which are the most commonly intercepted forms at quarantine and inspection points. Traditional identification relies mainly on adult morphological features, meaning that immature samples must be reared in the laboratory until they become adults. This process is slow, labour-intensive, and risky. A single misidentification can either result in unnecessary trade restrictions or, even worse, allow a dangerous invasive species to establish itself in a new area. To improve accuracy, molecular techniques such as DNA barcoding, PCR, and real-time PCR have been introduced into fruit fly diagnostics. These methods are highly reliable, but they require expensive equipment, uninterrupted electricity, clean laboratory conditions, and trained personnel. Because of these requirements, their use is limited in ports, markets, farms, and border inspection stations where fast decisions are needed.

Figure 1. Schematic representation of on-site LAMP-based detection of the melon fly Zeugodacus cucurbitae. Different pest life stages encountered from infested fruits and fields are subjected to rapid DNA extraction, followed by isothermal LAMP amplification at 60–65 °C for about 40 minutes. The results are visualized by colour change, where yellow indicates a positive reaction (pest DNA detected) and pink indicates a negative reaction (no pest DNA). (Illustration improved using AI tools)

This gap between laboratory science and field reality has been bridged by Loop-mediated Isothermal Amplification (LAMP). LAMP is a DNA-based technique that works at a constant temperature, usually between 60 to 65 °C. Unlike PCR, it does not require a thermal cycler. A simple heating block or even a water bath is sufficient. The method uses 2-3 pairs of primers that recognize multiple regions of the target gene, giving it very high specificity. Within 30 to 60 minutes, LAMP can amplify large amounts of DNA if the target species is present. The results are often visual, using colour-changing dyes, allowing positive samples to be identified by the naked eye. This makes LAMP ideal for field use, even by personnel with limited laboratory training. LAMP was first applied to fruit fly detection in 2009 for the Mediterranean fruit fly, Ceratitis capitata. Since then, it has been developed for many important pest species, including Bactrocera tryoni, Z. scutellatus, Dacus ciliatus, and B. minax. These successes proved LAMP a powerful tool for quarantine and pest surveillance (Dermauw et al., 2022).

Among economically important fruit fly pests, the melon fly Zeugodacus cucurbitae is one of the most destructive species in India and many parts of Asia. It infests a wide range of cucurbit crops, including cucumber, melon, pumpkin, bitter gourd and ridge gourd. The larvae develop inside the fruits, rendering them unmarketable and causing severe economic losses. Under unprotected conditions, Z. cucurbitae can result in up to 100% yield loss by puncturing the fruit surface with its stout ovipositor and depositing four to ten eggs per oviposition event. Owing to its high invasiveness and economic importance, it is listed as a quarantine pest by major plant protection organizations, including the Asia and Pacific Plant Protection Commission (APPPC), the European and Mediterranean Plant Protection Organization (EPPO), the Comité de Sanidad Vegetal del Cono Sur (COSAV), the Caribbean Plant Protection Commission (CPPC) and the Organismo Internacional Regional de Sanidad Agropecuaria (OIRSA). Given its high damage potential and quarantine status, early and accurate detection is critical for effective management.

To meet this need, a melon fly-LAMP assay was developed specifically to detect Z. cucurbitae by ICAR-Indian Agricultural Research Institute team. This test targets the COI (cytochrome c oxidase subunit I) gene, a well-established marker used for insect species identification. The assay runs at 60 °C and provides results within 40 minutes, making it highly suitable for rapid field diagnosis. The test showed excellent specificity. When evaluated against five closely related fruit fly species, Z. tau, Bactrocera dorsalis, B. divenderi, B. zonata, and B. correcta, only Z. cucurbitae produced a positive result. This confirms that the assay can clearly distinguish the target melon fly from similar species. It also demonstrated very high sensitivity, detecting extremely small amounts of DNA, even from eggs, larvae, and damaged tissues. Importantly, the assay works on all life stages, including eggs, larvae, pupae, and adults. Another major advantage is its simplicity: insects can be crushed in double-distilled water, and the extract can be used directly in the reaction without complex DNA extraction. The assay was validated using melon fly samples from different parts of India and performed consistently across all locations, proving its reliability across geographical populations (Barman et al., 2025).

Figure 2. Workflow for species-specific LAMP assay development for the melon fly, Zeugodacus cucurbitae. The process includes selection of the target pest, identification of a species-specific mitochondrial COI gene region, design of LAMP primers, optimization of the amplification reaction, integration of a simple crude-DNA template preparation method, and final validation of the assay for accurate detection of the melon fly.

In summary, Melon fly-LAMP offers a fast, accurate, sensitive, and field-friendly method for detecting the melon fly, Z. cucurbitae. By bringing molecular diagnostics out of the laboratory and into the field, it supports better quarantine decisions, early pest detection, reduced pesticide misuse, and safer agricultural trade.

For more details, refer:

Barman, S., Shashank, P. R., Diksha, D., Sharma, S. K., Gupta, N., Kumar, A., Dhillon, M.K. & Ray, S. (2025). A Simple Template Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid On-Site Detection of Melon fly, Zeugodacus cucurbitae. Journal of Agriculture and Food Research, 25(2026), 102581.

Dermauw, W., Van Moerkercke, Y., Ebrahimi, N., Casteels, H., Bonte, J., & Witters, J. (2022). A loop-mediated isothermal amplification (LAMP) assay for rapid identification of Ceratitis capitata and related species. Current Research in Insect Science, 2, 100029.

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About the authors:

Suman Barman is a Ph.D. Student at the Division of Entomology, ICAR- Indian Agricultural Research Institute, New Delhi. He worked on LAMP-based on-site detection of Melon fly, Zeugodacus cucurbitae.

Email: sumanbarman21302@gmail.com 

Shashank P. R. is a Senior Scientist at the Division of Entomology, ICAR- Indian Agricultural Research Institute, New Delhi. His field of specialization is insect taxonomy, molecular diagnosis and invasive pests. He is the Founding Managing Editor of Indian Entomologist.

Email: spathour@gmail.com

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