Introduction
Greenhouses are vital platforms for agricultural production, contributing significantly to food security. High yield potential and efficient land utilization are key advantages of greenhouse cultivation of tomatoes and cucumbers in Herat and Farah provinces of Afghanistan. The cultivation of greenhouse cucumbers and tomatoes is a relatively new practice in Afghanistan, particularly in the Herat region, resulting in limited knowledge about the pathogens affecting these crops. Detailed information on plant-parasitic nematodes impacting cucumbers, tomatoes, and various other crops in Afghanistan, particularly in the Herat region, remains limited. Each year, significant crop losses are caused by root-knot nematodes (Meloidogyne spp.). These challenges prompted the present study to identify root-knot nematode species associated with cucumber and tomato crops in these regions.
Materials and Methods
From 2019 to 2021, 40 soil and root samples were collected from greenhouses and fields in Herat and Farah provinces. After collection, soil samples were labeled and transported to the Plant Disease Research Laboratory, Faculty of Agriculture, Ferdowsi University of Mashhad, for analysis. Nine soil samples infested with root-knot nematodes were reared on tomato (Solanum lycopersicum L. cv. Early Urbana) in the Plant Disease Research Greenhouse. After 60–65 days, nematode-infected plant roots were transferred to the Plant Disease Laboratory for species identification. Species characterization was performed using morphological and morphometric data of adult females and second-stage juveniles. Two populations (HJI1 from Harat and HJI2 from Farah Provinces) selected for more morphometric, morphological and molecular studies and comparisons. Molecular studies were conducted only for HJI1 & HJI2 populations and based on D2-D3 of 28S rRNA region.
Results and Discussion
Morphometric and morphological features of M. incognita distinguished this nematode from other species. For more detailed studies HJI1 & HJI2 populations were selected from Harat and Frah provinces, respectively. Both populations were similar to each other, morphologically. Females: Pear-shaped, white, with prominent neck, body length 700 to 900 μm, fine stylet 15 to 17 μm long, large and oval mid-esophageal bulb with a distinct valve. The perineal pattern oval to rounded, the dorsal arch high, rounded and sometimes rectangular, flatten at the tip or trapezoidal, the lines of lateral fields indistinct and sometimes with fork-like striae. The vulval slit 19 to 24 μm, its distance from the anus was 17 to 23 μm, and the phasmids visible. Males: not founded. Second-stage juveniles: Long, worm-shaped 330-450 μm, lateral field with four incisures. Stylet 10.5 to 14 μm, the conical part of the stylet is almost equal to the cylindrical part, and knobs prominent and round. The dorsal esophageal gland 2 to 3.5 μm below the stylet knobs. Tail conical, 33 to 56 μm long, hyaline region 10.5 to 14 μm. However, there are some overlaps in the morphometric and morphological data of Meloidogyne spp., so sequencing of the D2-D3 region of 28S rRNA was used for more precise identification. The results grouped the studied species with tropical root-knot nematodes. A phylogenetic tree was constructed based on 31 sequences from the 28S ribosomal DNA region, including two sequences coded HJI1 and HJI2 from the present study, two sequences from Pratylenchus penetrans and Radopholus similis as outgroups, and 27 other sequences from the NCBI gene bank. In this tree, the sequences from the two populations HJI1 and HJI2 were placed in a monophyletic multilineage clade (clade A) that also includes sequences from other tropical root-knot nematode species, including M. incognita, M. javanica, M. arenaria, M. luci, M. morocciensis, M. konaensis, M. paranaensis, and M. phaseoli. Therefore, M. incognita was distinguished from other tropical root-knot nematode species by its trapezoidal, rounded, high and occasionally flattened dorsal arch, closely spaced striae that exhibit a very wavy to zigzag pattern, and the indistinct lateral field and sometimes marked by breaks in stria. This is the first study of morphometric, morphological and molecular characteristics of M. incognita species in Harat and Frah Provinces greenhouses of Afghanistan.
Conclusion
Preventing damage to greenhouse cucumber and tomato cultivation and improving crop yields requires an understanding of factors that reduce the quality and quantity of these crops. Root-knot nematodes are destructive plant-parasitic pathogens that reduce agricultural productivity, and threaten food security. Notably, M. incognita was isolated and identified from Herat and Farah provinces and two selected populations were similar. The high similarity between the two populations and the neighborhood of Herat and Farah provinces may indicate that the spread of M. incognita infections has been happen through plant exchanges without regard to their health. Therefore, it is necessary to observe plant quarantine in infected areas and greenhouses to prevent the spread of this pathogen. The obtained data and characteristics of this study can be used to facilitate future studies and more easily identifying M. incognita species.