DEVELOPMENT OF NEW TEST SYSTEMS FOR MOLECULAR GENETIC IDENTIFICATION DITYLENCHUS DESTRUCTOR THORNE, 1945

The potato rot nematode (Ditylenchus destructor) has been recorded from several countries and causes considerable yield loss of potato. During 2018-2020, the potato rot nematode, D.destructor, Thorne, 1945 were extracted from potato tubers collected from six regions of Iran and the Russian Federation. These populations were described and compared for the first time based on their morphological, morphometric, and molecular characteristics. Furthermore, two new primer pairs were developed to exactly identify the nematode. The morphological and morphometric features were generally in agreement with those published for D. destructor. The rRNA gene of the nematodes was amplified and sequenced using two primer sets: dsn.1 R/F and dsn.2 R/F. New sequences were deposited in the GenBank under accession numbers MN122076, MN307126, MN307128, MN493767, MN658597, MN658599, MN658637, MN658638. The identification was further supported by PCR with designed species-specific primers for this species

Primary Screening of Microorganisms against <i>Meloidogyne hapla</i> (Chitwood, 1949) under the Conditions of Laboratory and Vegetative Tests on Tomato

Highly adapted obligate endoparasites of the root system, root-knot nematodes (Meloidogyne spp.), cause great damage to agricultural crops. Our research is aimed at the assessment of nematicidal activity and effectiveness of antagonist fungal and bacterial strains against the most common type of root-knot nematode in the south of Russia. By means of molecular genetic identification, it was found that in the south of Russia, the species Meloidogyne hapla Chitwood, 1949 and Meloidogyne incognita (Kofoid and White, 1919) Chitwood, 1949 cause galls on the roots of open-ground and greenhouse tomato. Screening of microbial agents against second-stage juvenile (J2) M. hapla was carried out in the laboratory. At the end of the experiment, two liquid fungal cultures of Paecilomyces lilacinus BK-6 and Metarhizium anisopliae BK-2 were isolated, the nematicidal activity of which reached 100.0 and 70.2%, and exceeded the values of the biological standard (Nemotafagin-Mikopro) by 38.4% and 8.8%. The highest biological efficacy was noted in the liquid cultures of P. lilacinus BK-6, M. anisopliae BK-2, and Arthrobotrys conoides BK-8 when introduced into the soil before planting tomato. The number of formed galls on the roots was lower in comparison with the control by 81.0%, 75.5%, and 74.4%

Molecular diagnostics of Ditylenchus destructor based on the ITS-rDNA from Iran and Russia federation

Several nematode species related to potato have been documented. Among these, the potato rot nematode, Ditylenchus destructor (Thorne) may cause important losses in potato yields. Proper identification of nematode species and isolates is important. To verify species identification, the populations of D. destructor from Iran and the Russian Federation were tested for differences using PCR with species-specific primers for the rapid and reliable result. New sequences deposited in the Genbank under accession numbers MN307126, MN307128 and MN493767 from Iran populations, and MN658597, MN658599, MN658637 and MN658638 from Russian populations. Phylogenetic studies of the ITS-rDNA region of D. destructor put it in a distinct clade. © 2020, Gaurav Society of Agricultural Research Information Centre. All rights reserved

Developing PCR primers for identification of potato rot nematode (Ditylenchus destructor) in the Russian Federation and Iran

The potato rot nematode (Ditylenchus destructor) species is an influential pathogenic potato nematode. Many countries impose quarantine restrictions on the presence of D. destructor in imported plant and soil materials. Therefore, the present investigation was conducted from 2018 to 2020 at the All-Russian Plant Quarantine Center of Russia Federation with samples collected from the Severtsov Institute of Ecology and Evolution, RAS, Russia and Iran. In the current study, we developed PCR with species-specific primers for the rapid and reliable separation of D. destructor using gel electrophoresis and melting curve analysis. A species-specific primer set was designed based on the nucleotide sequence of the gene. The PCR protocol was verified using samples of D. destructor and the closely related species, D. dipsaci and D. gigas. The species-specific primer set was able to detect D. destructor from samples containing mixtures of Ditylenchus species. The PCR species-specific protocol should allow for more rapid identification of Ditylenchus species recovered from plant materials than previously possible. © 2020, Gaurav Society of Agricultural Research Information Centre. All rights reserved

Preserved Microarrays for Simultaneous Detection and Identification of Six Fungal Potato Pathogens with the Use of Real-Time PCR in Matrix Format

Fungal diseases of plants are of great economic importance causing 70&#8315;80% of crop losses associated with microbial plant pathogens. Advanced on-site disease diagnostics is very important to maximize crop productivity. In this study, diagnostic systems have been developed for simultaneous detection and identification of six fungal pathogens using 48-well microarrays (micromatrices) for qPCR. All oligonucleotide sets were tested for their specificity using 59 strains of target and non-target species. Detection limit of the developed test systems varied from 0.6 to 43.5 pg of DNA depending on target species with reproducibility within 0.3&#8722;0.7% (standard deviation). Diagnostic efficiency of test systems with stabilized and freeze-dried PCR master-mixes did not significantly differ from that of freshly prepared microarrays, though detection limit increased. Validation of test systems on 30 field samples of potato plants showed perfect correspondence with the results of morphological identification of pathogens. Due to the simplicity of the analysis and the automated data interpretation, the developed microarrays have good potential for on-site use by technician-level personnel, as well as for high-throughput monitoring of fungal potato pathogens