Dr. Le Thi Mai Linh and her colleagues are analyzing nematode samples

Plant parasitic nematodes (PPN) are one of the most serious pests in plants. They can cause damage to all different parts of the plant such as: stem, leaves, flowers, fruit or roots. In particular, nematodes that damage roots have the ability to reduce crop yield and quality. Research on nematodes is essential for species identification, as a basis for effective management measures to prevent outbreaks of nematode pests, especially in developing countries. develop like Vietnam. Supplementing missing DNA data for nematode groups that are harmful to coffee, pepper, citrus and some intercropped crops focused on research in Vietnam.

From the above reality, scientists from the Institute of Ecology and Biological Resources proposed and carried out the task: "Developing DNA barcoding, multiplex PCR, Real-time PCR techniques in identifying dangerous parasitic nematodes on coffee, pepper and citrus trees in Vietnam", code: VAST04.08/22-23, Dr. Le Thi Mai Linh is the project head. The project aims to determine the diversity and phylogenetic relationships of plant parasitic nematodes in Vietnam; Develop and optimize DNA barcoding and Multiplex PCR methods to help quickly identify plant-parasitic nematodes; Designing a process using the Realtime-PCR method to quickly classify dangerous nematode groups in Vietnam.

After the research process, the research team added and analyzed morphological data on groups of harmful plant parasitic nematodes, including root-knot nematodes, moving endoparasitic nematodes (Pratylenchus coffeae), and gill nematodes. virus transmission (Xiphinema bevicolle) and some ectoparasitic varieties. At the same time, scientists have successfully designed a pair of 18S rDNA primers to serve the identification of dangerous nematode species belonging to the group that transmits viruses. The team discovered that Root-knot nematodes (except the tropical nematode Meloidogyne incognita group (MIG)) can use nuclear genes (ITS, 28S) and mitochondrial genes (COI), the MIG group is necessary to use using NAD5 gene; For the group of migratory endoparasitic nematodes, the D2D3 28S rDNA gene can be used, and for nematodes transmitting X. bevicolle virus, the 18S rDNA gene can be used in phylogenetic and identification analyzes.

Dr. Linh and colleagues have perfected and optimized the use of Multiplex PCR primer pairs for rapid identification of root-knot nematode species complexes in Vietnam. The team has successfully designed and built a Real-time PCR reaction process for the species Meloidogyne incognita, which has the most variable morphology in Vietnam. At the same time, the research team completed a set of 300 specimens for morphological assessment of important plant-parasitic nematode species collected at the investigation sites with detailed and complete information of 07 species belonging to 05 genera. Scientists have also published 40 DNA sequences (18S, 28S and COI gene regions) of several species on the international Gen Bank.

Microscopic image of the nematode Hemicaloosia was recorded for the first time in Vietnam

The Multiplex-PCR method amplifies DNA fragments with multiple primer pairs in the same PCR reaction (multiple-primer PCR) can help quickly classify many species in the same reaction and is applied to many groups of serious pests. Therefore, the Multiplex PCR method in rapid classification of three species of root-knot nematodes Meloidogyne incognita, Meloidogyne arenaria, Meloidogyne javanica of the research team can be applied in quarantine and plant protection. The research results of the project are a new direction, contributing to the accurate assessment of important groups of plant-parasitic nematodes on key crops with economic value such as coffee, pepper, Citrus trees or some intercropped trees in Vietnam. Multiplex PCR as well as Realtime-PCR methods will provide maximum support for plant nematode quarantine by simplifying inspection steps while still ensuring speed and accuracy. Furthermore, this method can be widely applied in practice because the technical requirements are not too high, and technicians with basic skills in PCR practice can perform it.

The research results were published in 03 articles in the SCIE category, 01 article in the VAST02 category and were graded A by the Acceptance Council of the Vietnam Academy of Science and Technology. With the above successes, scientists from the Institute of Ecology and Biological Resources wish to continue in-depth research on the Realtime-PCR method in quickly classifying dangerous nematode groups in Vietnam to improve the results of the project and soon put it into practice.

Translated by Quoc Khanh
Link to Vietnamese version