XII INTERNATIONAL CONGRESS OF GENETICISTS AND BREEDERS OF THE REPUBLIC OF MOLDOVA September 17 – 18, 2025 Chisinau, Republic of Moldova SCIENTIFIC ASSOCIATION OF GENETICISTS AND BREEDERS OF THE REPUBLIC OF MOLDOVA INSTITUTE OF GENETICS, PHYSIOLOGY AND PLANT PROTECTION MOLDOVA STATE UNIVERSITY ACADEMY OF SCIENCES OF MOLDOVA ROMANIAN SOCIETY OF BIOENGINEERING AND BIOTECHNOLOGY MATERIALS PROCEEDINGS 1 Scientific Association of Geneticists and Breeders of the Republic of Moldova Institute of Genetics, Physiology and Plant Protection Moldova State University Academy of Sciences of Moldova Romanian Society of Bioengineering and Biotechnology XII INTERNATIONAL СONGRESS OF GENETICISTS AND BREEDERS OF THE REPUBLIC OF MOLDOVA September 17-18, 2025, Chisinau, Republic of Moldova MATERIALS PROCEEDINGS Chisinau, 2025 Editura USM 575+577.21(082)=135.1=111 https://doi.org/10.53040/cga12 I-58 Materials Proceedings of the XII International Сongress of Geneticists and Breeders of the Republic of Moldova, September 17-18, 2025, Chisinau, Republic of Moldova (approved by the Scientific Council of the Institute of Genetics, Physiology and Plant Protection, minute no 10, data 09.07.2025). Editorial Production: Dr. hab. Duca Maria, Moldova State University Dr. Clapco Steliana, Moldova State University Dr. Port Angela, Moldova State University Antonov Alexandr, Moldova State University Note! The Authors of papers submitted to the Congress of Geneticists and Breeders of the Republic of Moldova take full responsibility for their content/originality and for English language. The Congress is organized within the subprogramme 011101 Genetic and biotechnological approaches to the management of agroecosystems in the conditions of climate change, funded by the Ministry of Education and Research. ©Duca Maria et al. ©Moldova State University ©Editura USM DESCRIEREA CIP A CAMEREI NAŢIONALE A CĂRŢII DIN REPUBLICA MOLDOVA International Congress of Geneticists and Breeders of the Republic of Moldova (12 ; 2025 ; Chişinău). XII International Congress of Geneticists and Breeders of the Republic of Moldova : Materials Proceedings, September 17-18, 2025, Chisinau / scientific committee: Duca Maria (chair) [et al.]. – Chişinău : Editura USM, 2025. – 476 p. : fig. color, tab. Antetit.: Scientific Association of Geneticists and Breeders of the Republic of Moldova [et al.]. – Text : lb. rom., engl. – Referinţe bibliogr. la sfârşitul art. – Index de nume : p. 473-474. – 45 ex. ISBN 978-9975-62-897-6. 575+577.21(082)=135.1=111 I-58 3 SCIENTIFIC COMMITTEE CHAIR DUCA Maria, ORCID: 0000-0002-5855-5194, president of Scientific Association of Geneticists and Breeders of the Republic of Moldova (SAGBRM) Doctoral School of Natural Sciences, Moldova State University (MSU), Republic of Moldova. MEMBERS ANDRONIC Larisa, ORCID: 0000-0002-2761-9917, Institute of Genetics, Physiology and Plant Protection, MSU, vice-president of SAGBRM, Republic of Moldova CALALB Tatiana, ORCID: 0000-0002-8303-3670, Nicolae Testemitanu State University of Medicine and Pharmacy, Republic of Moldova CEPOI Liliana, ORCID: 0000-0002-7516-948X, Institute of Microbiology and Biotechnology, Technical University of Moldova, Republic of Moldova COMAROVA Galina, ORCID: 0009-0001-0063-4586, Technical University of Moldova, Republic of Moldova GROPPA Stanislav, ORCID: 0000-0002-2120-2408, vice-president of SAGBRM, Republic of Moldova JOITA-PACUREANU Maria, ORCID: 0009-0009-2797-6672, Center for Research for Agroforestry Biodiversity “Acad. David Davidescu”, Romania KAYA Yalcin, ORCID: 0000-0002-9297-8633, Plant Breeding Research Center, Trakya University, Turkey KOROL Abraham, ORCID: 0000-0002-3401-7068, University of Haifa, Israel LUPASCU Galina, ORCID: 0000-0003-3363-3595, Institute of Genetics, Physiology and Plant Protection, MSU, Republic of Moldova MASNER Oleg, ORCID: 0000-0002-6498-9095, National Institute of Applied Research in Agriculture and Veterinary Medicine, Republic of Moldova MEREUTA Ion, ORCID: 0009-0003-2151-9928, Institute of Physiology and Sanocreatology, MSU, Republic of Moldova MIHASAN Marius, ORCID:0000-0003-4439-4251, Al.I.Cuza University, Iasi, România PAUN Mihaela, ORCID: 0000-0002-3342-9140, National Institute for Research and Development for Biological Sciences, Bucharest, Romania ROSCA Ion, ORCID: 0000-0002-1304-8033, National Botanical Garden (Institute) “Alexandru Ciubotaru”, MSU, Republic of Moldova SIDOROFF MANUELA, ORCID: 0000-0001-8740-3803, Romanian Society of Bioengineering and Biotechnology, Bucharest, Romania SPIVACENCO Anatolie, National Center for Seed Research and Production, Republic of Moldova UNGUREANU Laurentia, ORCID: 0000-0003-4576-2810, Institute of Zoology, MSU, Republic of Moldova ZHAO Jun, ORCID: 0000-0002-3964-7755, Inner Mongolia Agricultural University, Huhhot, China 4 ORGANIZING COMMITTEE CHAIR DUCA Maria, President of the Scientific Association of Geneticists and Breeders of the Republic of Moldova, the Doctoral School of Natural Sciences, Center for Functional Genetics of the Moldova State University, Republic of Moldova MEMBERS COTENCO Eugenia, Institute of Genetics, Physiology and Plant Protection, MSU, Secretary of Scientific Association of Geneticists and Breeders ANTONOV Alexandr, Center of Functional Genetics, MSU BATIRU Grigorii, Faculty of Agricultural, Forestry and Environmental Sciences, Technical University of Moldova BILETCHI Lucia, Institute of Zoology, MSU CLAPCO Steliana, Center of Functional Genetics, MSU PORT Angela, Center of Functional Genetics, MSU SACARA Victoria, Institute of Mother and Child 5 SUMMARY I. GENERAL AND MOLECULAR GENETICS 1. DEAGHILEVA A., BELOUSOVA G., CUZNETOVA I., IGNATOVA Z. MULLER K. MOLECULAR DIAGNOSTICS OF SOME FUNGAL PATHOGENS IN GREENHOUSE GROWN TOMATOES 15 2. FRUNZE N. POOL AND DIVERSITY OF PROKARYOTES OF CARBONATE CHERNOZEM OF LONG-TERM FIELD EXPERIENCE OF MOLDOVA 21 3. GLADCAIA A., CHISNICEAN L. METHOD OF ACTIVATING ENTOMOPHAGES AND POLLINATORS HYMENOPTERA ORDER NATURAL POPULATIONS WITH THE AIM OF INCREASING THE AGROSYSTEM PRODUCTIVITY 28 4. GOLOVCOVA S. ASSESSING GENETIC DIVERSITY IN TRITICALE POPULATIONS USING iPBS MARKERS 34 5. IURCU-STRAISTARU E., BIVOL A., RUSU S., RUSU V. PARASITIC NEMATOFAUNA IN PEA CROPS (Pisum sativum L.) UNDER THE IMPACT OF THE UNSTABLE ENVIRONMENTAL CONDITIONS OF THE REPUBLIC OF MOLDOVA 40 6. SITNIC V., NISTREANU V., MUNTEANU V., LARION A., SOCHIRCA N. DETERMINATION OF THE PHYLOGENETIC LINEAGE OF THE SPECIES MICROTUS ARVALIS FROM THE REPUBLIC OF MOLDOVA BASED ON CYTB GENE SEQUENCING 48 7. UZUN T., GRĂJDIERU C. MOLECULAR-GENETIC IDENTIFICATION OF CERTAIN FRUIT TREE MYCOPATHOGENS TRANSMITTED BY APHID VECTORS 55 II. HUMAN AND MEDICAL GENETICS 1. BACALOV I., CRIVOI A., CHIRIŢA E., DRUŢA A. HOMEOSTASIS OF CARBOHYDRATE METABOLISM IN EXPERIMENTAL DIABETES UNDER THE INFLUENCE OF THE BIOPREPARATION SEV-3 62 2. COLIBAN I., UȘURELU N., SACARĂ V. FROM VALIDATION TO APPLICATION: IMPLEMENTING A COST- EFFECTIVE QPCR ASSAY FOR PRESYMPTOMATIC AND SYMPTOMATIC SMA DIAGNOSIS 70 3. DUCA M., ANTONOV A., SIDOROFF M. GENETIC MAPPING OF HUMAN DISEASES: A QUANTITATIVE AND FUNCTIONAL PERSPECTIVE ON GENE–PATHOLOGY RELATIONSHIPS 77 4. PODOLEANU D., MUNTEANU I., STARODUB E., CHIRMAN Gh., ENAKI N. 84 6 GENERATION OF PHOTONIC MOLECULAR COMPLEXES UNDER THE ACTION OF ULTRAVIOLET LIGHT IN THE DNA/RNA INACTIVATION PROCESS 5. SECU D., BLANITA D., USURELU N., SACARA V. CLINICAL, BIOCHEMICAL, AND GENETIC FINDINGS IN A GROUP OF PATIENTS EVALUATED FOR SUSPECTED MITOCHONDRIAL DISORDERS 95 6. TURUTA L., DUDNIC E. MANIFESTATIONS OF PHYSIOLOGICAL HOMEOSTASIS DYSFUNCTIONS 102 III. PLANT GENETICS AND BREEDING 1. ALİYEVA D., MUSAYEVA S. EVALUATION OF GENETIC DIVERSITY: A STUDY ON MORPHOLOGICAL AND PHYSIOLOGİCAL TRAITS OF LOCAL AND INTRODUCED DURUM WHEAT (TRITICUM DURUM DESF.) GENOTYPES 109 2. AVĂDĂNII L., GUȚU C., IACOBUȚA M. THE ROLE OF GENETIC RESOURCES IN THE IMPROVEMENT AND DEVELOPMENT OF NEW LEGUME CROP VARIETIES 117 3. BALAN V., BÎLICI I., ȘARBAN V., RUSSU S., BUZA C., TALPALARU D. NEW TRAINING SYSTEMS FOR HIGH-DENSITY CHERRY PLANTATIONS 121 4. BALMUȘ Z., COTELEA L., BUTNARAȘ V. EVALUATION OF INBRED LINES OF SALVIA SCLAREA L. 127 5. BATIRU G., COMAROVA G. USE OF BIOFORTIFICATION PRINCIPLES IN DEVELOPING A MAIZE GERMPLASM COLLECTION 133 6. BÎLICI I., BALAN V., ȘARBAN V., RUSSU S., BUZA C., TALPALARU D. INFLUENCE OF PLANTING DISTANCE AND CROWN SHAPE ON THE GROWTH, PRODUCTIVITY AND QUALITY OF CHERRY FRUITS, GRAFTED ON GISEALA 6 139 7. BOROZAN P., MUSTEAȚA S., SPÎNU V., SPÎNU A., DONICI R. RESULTS OF EARLY CORN BREEDING IN THE REPUBLIC OF MOLDOVA 148 8. BRINDZA J., HORČINOVÁ-SEDLÁČKOVÁ V. BREEDING THE DOMESTIC APPLE TREE (MALUS DOMESTICA BORKH.) AS A FAMILY CREATIVE ACTIVITY 155 9. BUTNARAS V., BALMUS Z., COTELEA L. PRODUCTIVITY INDICES IN LAVANDULA ANGUSTIFOLIA MILL. VARIETIES, SIXTH YEAR OF VEGETATION 163 10. CLIMENCO O. SCREENING OF CORN GENOTYPES FOR SALINITY RESISTANCE 170 11. COJOCARI D., BOUNEGRU S., BATIRU G. EVALUATION OF MID-PARENT AND BEST-PARENT HETEROSIS 176 7 FOR GRAIN YIELD IN SELECTED MAIZE INBRED LINES 12. CORLATEANU L., MIHAILA V., GANEA A., FOCSHA N. STORAGE POTENTIAL OF THE EGGPLANT (SOLANUM MELONGENA L.) COLLECTION ACCESSIONS FOR EX SITU CONSERVATION IN A PLANT GENE BANK 182 13. COTELEA L., BALMUȘ Z., BUTNARAȘ V. EVALUATION OF NEW HYBRIDS OF SALVIA SCLAREA L., WITH HIGH ESSENTIAL OIL CONTENT 188 14. CRISTEA N., LUPASCU G. RESEARCH ON THE INTERACTION PECULIARITIES OF COMMON WHEAT WITH FUSARIUM OXYSPORUM ISOLATES 195 15. CURSUNJI D. AGRONOMICAL CHARACTERISTICS THE BREEDING GERMPLASM OF CHICKPEA 204 16. CUTITARU D., BRINDZA J. INDUCTION OF ARTIFICIAL MUTATIONS IN LINUM USITATISSIMUM L. BY EXPOSING SEEDS TO DIFFERENT DOSES OF X-RAYS 211 17. DUCA M., CLAPCO S., MUTU A., MARTEA R., JOITA- PACUREANU M. COMPARATIVE DROUGHT RESISTANCE IN SUNFLOWER UNDER CONTROLLED CONDITIONS 217 18. ELISOVETCAIA D., IVANOVA R., POPOVSCHI E., FEDORENCO E., BRINDZA J. ADAPTATION OF BEECH PLANTS FROM DIFFERENT PROVENANCES TO NATURAL GROWTH CONDITIONS 224 19. EZERINA E., CHESNOKOV Y. INFLUENCE OF PHOTOPERIOD ON PRODUCTIVITY AND SPECTRAL INDICES OF LEAF BLADES OF GARDEN CRESS SAMPLES GROWN UNDER LIGHT CULTURE CONDITIONS 228 20. GORE A., LEATAMBORG S., ROTARI S. BREEDING OF COMON WHEAT (TRITICUM AESTIVUM) IN THE CENTRAL PART OF RM 232 21. IVANOVA R., LUTCAN E., VANICOVICI N., BOROVSKAIA A., ELISOVETCAIA D. IMPACT OF NON-OPTIMAL TEMPERATURE AND BIOREGULATOR ON ROOT AND SEEDLING VIGOUR OF CORN 239 22. LEATAMBORG S., ROTARI S., GORE A. STUDY OF VARIABILITY IN PRODUCTIVITY ELEMENTS OF WINTER TRITICALE COLLECTION 244 23. LUPASCU G., CRISTEA N., GAVZER S. INFLUENCE OF THE FUSARIUM OXYSPORUM FUNGUS ON THE TRANSGRESSIVE POTENTIAL OF GROWTH TRAITS OF COMMON WHEAT 250 8 24. MAKOVEI M. THE BREEDING OF HIGHLY RESISTANT TOMATO F1 HYBRIDS TO STRESS ABIOTIC FACTORS 258 25. MALII A. EVALUATION OF OSMOTIC STRESS TOLERANCE DURING THE GERMINATION STAGE IN SOYBEAN GENOTYPES 265 26. MARII L., ANDRONIC L., SAHANOVSCHIH M., URSACHI O. VARIATION IN TOMATO GENOTYPES RESPONSE TO X-RAY IRRADIATION AND ESTIMATION OF THE EFFECTIVE DOSES 271 27. MARINCIU C.-M., ȘERBAN G., MANDEA V., GALIT I., SAULESCU N. RANKS AND RANK STABILITY OF WHEAT CULTIVARS TESTED IN NATIONAL YIELD TRIALS IN ROMANIA 280 28. MIHNEA N., RUSU V., BREZEANU P., AMBĂRUS S. COMPARATIVE ANALYSIS OF RESISTANCE TO HIGH TEMPERATURE STRESS IN PROSPECTIVE TOMATO LINES 286 29. NIKOLIĆ V., ŽILIĆ S., SIMIĆ M., SARIĆ B., MILOVANOVIĆ D., BABIĆ V. VARIABILITY OF THE PHYSICOCHEMICAL PROPERTIES OF SELECTED MAIZE GENOTYPES AND PROSPECTS OF THEIR APPLICATION FOR VARIOUS PURPOSES 292 30. OGUDIN G., ARTEMYEVA A., SOLOV’EVA A. SOURCES OF ECONOMICALLY VALUABLE TRAITS FOR BRASSICA RAPA L. VEGETABLES CROPS BREEDING 297 31. PINTEA M. RESERCHES REGARDING ADAPTABILITY OF NEW WALNUT VARIETIES IN THE CONDITIONS OF REPUBLIC OF MOLDOVA 307 32. PORT A. TEMPORAL GENE EXPRESSION PATTERNS DURING MICROSPOROGENESIS IN SUNFLOWER 313 33. QURBANOVA Q., BABAYEVA S., RAHIMOV H., ABBASOV M. INITIAL GENETIC DIVERSITY ASSESSMENT OF ENDEMIC HYRCANIAN WILD FIG POPULATIONS IN AZERBAIJAN 321 34 RABKO S., PAPLAUSKAYA L., NOVIKOVA T., NOVIKOV A., CHESNOKOV Y. CLIMATYPE BREEDING FEATURES OF PINUS SYLVESTRIS L. CV. NEGORELSKAYA 329 35 ROMANCIUC G. THE POSSIBILITY OF INTEGRATING THE REPUBLIC OF MOLDOVA INTO THE MULTILATERAL SYSTEM OF THE INTERNATIONAL TREATY ON PLANT GENETIC RESOURCES FOR FOOD AND AGRICULTURE 334 36. RUDACOVA A., CHERDIVARA A., RUDACOV S. EFFECT OF DROUGHT ON THE TOTAL PROTEIN COMPOSITION IN 342 9 SUNFLOWER SEEDLINGS IV. BIOTECHNOLOGIES 1. ASLANOV R. INTEGRATION OF BIOINFORMATICS AND CHEMINFORMATICS FOR THE ANALYSIS AND CLASSIFICATION OF BIOACTIVE COMPOUNDS USED IN CARDIOVASCULAR DISEASES 350 2. BOBROVA O., GOLOSNA L., NAROZHNYI S., OSETSKY A., ZAMECNIK J. ANTIFUNGAL ACTIVITY OF LIPID FRACTIONS OF MEDICINAL PLANTS AGAINST FUSARIUM ISOLATES FROM INFECTED GARLIC CLOVES 355 3. CEPOI L. CHIRIAC T., RUDI L., ZINICOVSCAIA I. BIOTECHNOLOGICAL VALORIZATION OF ARTHROSPIRA PLATENSIS FOR HEAVY METAL AND RARE EARTH ELEMENT RECOVERY FROM MULTIMETALLIC EFFLUENTS 361 4. CIOBANU R. EVALUATION OF VARIABILITY OF QUANTITATIVE CHARACTERS IN SC4 TRITICLE SOMACLONES WITH VALUABLE AGRONOMIC PROPERTIES 367 5. CURIEV L. TESTING OF THE RIHTER WDG PREPARATION IN THE CONTROL OF APPLE SCAB (VENTURIA INAEQUALIS WINT) 374 6. FRON A., BATCO M. EVALUATION OF ZEUZERA PYRINA INFESTATION IN WALNUT ORCHARDS AND PROPOSAL OF AN ALTERNATIVE ECOLOGICAL CONTROL METHOD 379 7. HARCIUC O., SÜTYEMEZ M., CHISTOL M., MALII A., STINGACI A. OIL CONTENT IN SEEDS BY NMR RELAXATION METHOD 384 8. IORDOSOPOL E., BATCO M., ELISEEV S., FRON A., FILIMON V., IORDOSOPOL V., FRON A., MUNTEAN E. HIBERNATING ARTHROPODS’ COMPLEX IN THE PLUM TREE ORCHARD UNDER THE INFLUENCE OF GREEN MANURES AND NECTARIFEROUS PLANTS 390 9. IVANIŠOVÁ E., DLUGOŠ M., GRYGORIEVA O. EXPLORING THE PHYTOCHEMICAL PROFILE AND SENSORY PROPERTIES OF GREEN AND ROASTED COFFEA ARABICA L. BEANS FROM DIVERSE GEOGRAPHIC REGIONS 396 10. KULESHOVA T. INFLUENCE OF ROOT ENVIRONMENT HUMIDITY ON THE GENERATION OF POTENTIAL DIFFERENCE IN A PLANT BIOELECTROCHEMICAL SYSTEM 407 11. MOCANU L., GONTA M. CATALYTIC OXIDATION OF GALLIC: MECHANISMS, KINETICS, 412 10 AND BYPRODUCT ANALYSIS 12. PALADI I. STUDY OF THE EFFICIENCY OF SUNFLOWER BEETLE CONTROL METHODS IN THE CONTEXT OF INTENSIFICATION AND EXPANSION OF CULTIVATION AREAS 417 13. SAMOILOVA A. THE INFLUENCE OF UV PROTECTANTS ON THE BACTERIOPHAGE ABILITY TO SUPPRESS FIRE BLIGHT DEVELOPMENT IN THE PLANTS TISSUES 424 14. SCERBACOVA T., CURIEV L., STINGACI A., GORE A. TESTING OF A BIOPREPARATION BASED ON BACILLUS AMYLOLIQUEFACIENS FOR WINTER WHEAT PROTECTION AGAINST DISEASES 429 15. SECRIERU S. INFLUENCE OF THE PREPATATION ECOSTIM ON THE PARAMETERS OF PHOTOSYNTHETIC ACTIVITY AND PRODUCTIVITY OF WINTER BARLEY VARIETIES 435 16. SÎRBU T., MOLDOVAN C. AQUATIC BASINS - NATURAL SOURCE OF FUNGI WITH ANTIFUNGAL POTENTIAL 441 17. SÎROMEATNICOV I., COTENCO E., PALADI D., BREZEANU P., AMBĂRUS S. VARIABILITY AND EQUALITY OF BIOCHEMICAL QUALITATIVE CHARACTERISTICS VARIETIES AND LINES NEW TOMATO OBTAINED IN VITRO 447 18. SLANINA V., BALAN L. ANTIBACTERIAL ACTIVITY OF SOME BACILLUS STRAINS IN DEPENDENCE ON THE CULTURE MEDIUM 454 19. TODIRAS V., CORCIMARU S., SLANINA V., BALAN L., PRISACARI S. PHYTOSTIMULATORY POTENTIAL OF SOME BACTERIA FOR WHEAT AND MAIZE 461 20. VOLOȘCIUC L., ȘCERBACOVA T., STÎNGACI A., PÎNZARU B., SAMOILOVA A., HARCIUC O., ZAVTONI P., LUNGU A., CHISTOL M., CURIEV L. TRENDS IN COMBATING VINE PHYTOPATHOGENS WITH THE APPLICATION OF EPIPHYTIC MICROORGANISMS 465 11 THE HERITAGE OF A.E. KOVARSKY’S SCHOOL IN THE FIELD OF BREEDING AND GENETICS Yevgeny V. KOVARSKY In the life of an academician of the Academy of Sciences of the Republic of Moldova, a professor of the Chisinau Agricultural Institute named M. V. Frunze, a doctor of agricultural sciences Anatoly Efimovich Kovarsky (1904-1974) three periods of his activity are distinguished, related to the study of field crops in the Republic of Moldova, in the steppe part of Ukraine, in the mountainous part of Kyrgyzstan - in these three regions, which differ from each other by their climate, soils and gene pool of field crops that were of scientific interest to A. E. Kovarsky. The core of A.E. Kovarsky’s legacy is the School of A. E. Kovarsky, which includes theoretical and practical experience that Anatoly Yefimovich left to his students in the Republic of Moldova. It is well known that the students of A.E. Kovarsky grew up to become important scientists, academicians and corresponding members of the Academy of Sciences of the Republic of Moldova, headed scientific schools and institutes. To the heritage of A.E. Kovarsky belong scientific approaches of A.E. Kovarsky to the knowledge of Nature, based on the provisions of C. Darwin, practical recommendations of I.V. Michurin and N.I. Vavilova. Among them are the influence of growing conditions on plants, morphogenetic processes, self-pollination, the effect of inter-varietal and foreign mentor pollen, cross-pollination in self-pollinating plants, mentor lines, the method of distant-geographical hybridization, cytoplasmic male sterility (CMS), hybridization, intergeneric and interline hybridization, experimental mutagenesis, and recombinational variability. Born in 1904, in the countryside (S. Popovka, Sumy region) of Ukraine, in a large family of farmers engaged in sugar beet, Anatoly Yefimovich, like his older brother Mikhail, followed his father’s footsteps and became an agronomist, founding Kharkov Agricultural Institute. Since 1924, first in the state reserve of Ascania Nova, and then in Kharkov and Kherson agricultural institutes and in the Kherson labor colony. The fate of his older brother1, whom he loved very much, could not help but affect Anatoly Efimovich, but he continued to work, solving food problems. During this early period of his scientific and creative life, A.E. Kovarsky read extensively, both professional and literary works. He became deeply interested in the teachings of Charles Darwin and I.V. Michurin, published numerous independent studies, and authored books on peanuts and sesame, which he wrote in 1 Mikhail Kovarsky, was one of the founders of the Machine-Tractor Stations (MTS), was awarded for this by the Order of Lenin, worked as deputy head of the Tractor Center USSR, but was arrested by the Soviet authorities and shot in 1933 on a false report about counter-revolutionary rebel activities in Ukraine, rehabilitated in 1957. 12 his native Ukrainian language. These books were published in large print runs for distribution to collective and state farms (kolkhozes and sovkhozes). At a young age, while heading the phyto-breeding station at the Askania-Nova Nature Reserve, A.E. Kovarsky gained valuable experience in organizational and managerial work. He studied the geographic sowing strategies of the Vavilov Institute (VIR), introduced new field crops, and worked with cotton, sesame, peanuts, and legumes. He also acquired practical experience during the difficult years of famine and drought from 1926 to 1928. As a result of his expedition to the mountainous region of Crimea, he collected and described varieties of winter and spring wheat, as well as wild einkorn wheats. A.E. Kovarsky received his first official academic title while working as a staff member at the All-Union Plant Breeding Institute (VIR, named after N.I. Vavilov). Later, in 1939, he defended his doctoral dissertation on the topic "Introduction of Peanuts in the USSR." The second period of Anatoly Yefimovich Kovarsky’s scientific career took place during the evacuation of the Kherson Agricultural Institute to Southern Kazakhstan and Kyrgyzstan. Between 1941 and 1942, he served as Deputy Director and Head of Department at the Kaplanbek Republican Agricultural College (Kazakhstan). From 1942 to 1944, he was Head of the Department of Breeding and Seed Production, Head of the Experimental Field, and Chief Agronomist of the Training Farm at the Kyrgyz Agricultural Institute. In 1943, he was awarded an Honorary Diploma by the Presidium of the Supreme Soviet of the Kyrgyz SSR. During this period, A.E. Kovarsky carried out research on leguminous crops, in which his wife, agronomist D.A. Kovarskaya, also took part. The third and longest period of A.E. Kovarsky’s professional activity, lasting thirty years, took place in the Republic of Moldova. His main work was centered in Chișinău, where he was engaged in both scientific and teaching activities at the Agricultural Institute and the Academy of Sciences. He also managed the operations of the Experimental Breeding Base / Trial Station of the Agricultural Institute, located in the suburb of Costiujeni. In addition, he was active in public outreach, civic, and governmental work, which included founding the Society of Geneticists and Breeders of the Republic of Moldova, editing well-known journals and academic collections, participating in sessions of the Supreme Soviet, holding meetings with the leadership of the Republic of Moldova and the Ministry of Agriculture, and visiting collective farms throughout the territory of the Republic of Moldova. The practical heritage includes A.E. Kovarsky varieties and hybrids, as well as A.E. Kovarsky field crop collections, field crop farming experience and seed production. A. E. Kovarsky and S. L. Pynzar, along with their colleagues, collected and studied a large collection of samples of chickpea, bean, vetch, soybean, and cowpea. 13 For soybean and especially for cowpea and mung bean, they used the introduction of initial material from outside, from the center of its diversity (according to N. I. Vavilov, 1935) and distant-geographical and distant-genetic hybridization. By methods of distant-geographical, distant-genetic hybridization and experimental mutagenesis, they developed wheat varieties — Kishinevskaya 9, Kishinevskaya 24, Kishinevskaya 101, Kishinevskaya 102, Eritrospermum 103, Kishinevskaya Early, Kishinevskaya Large-Grained, new 56- and 42-chromosome triticale forms; soybean varieties — Kishinevskaya 1 and Kishinevskaya 5; bean varieties — Kishinevskaya Stambovaya and Delicates; chickpea varieties — Favorit and others. The collection of 709 local Moldovan maize samples, initiated by A. E. Kovarsky together with V. G. Uchkovsky in 1945 — the vast majority of which belong to the flint and semi-dent subspecies of maize — served as the source for obtaining male pollen sterility, known as the "Moldovan type of cytoplasmic male sterility" or "Moldovan CMS." Based on the Moldovan type of CMS, several maize hybrids zoned in the Republic of Moldova were developed and converted to a sterile cytoplasmic base, including VIR 42 MV, Kishinevsky 109 M, Kishinevsky 150 MV, Kishinevsky 161 MV, Kishinevsky 167 M, Doina, Vstrecha, Kishinevsky 183, Kishinevsky VL 115, and others. A.E. Kovarsky were collected new collections of samples of maize from different countries; attracted to the crossing with maize its closest relatives from triba maize; created a collection of spontaneous mutations, formed due to the action of natural factors of external environment; a collection of induced mutations resulting from the use of chemical and physical mutagenesis methods; a collection of so-called menthar lines resulting from the mutating effect of pollen decay products. The collection includes a large number of samples from Central Asia and Kazakhstan. The collection contains more than 1,000 samples from 45 foreign countries, which are mainly obtained through ERA. The maize collection is periodically (after 4-5 years) updated by pollinating inside a sample of 5-10 pollen plants with other 5-10 plants. This method aims to keep the samples in their original state. In order to preserve the heritage of A.E. Kovarsky School in the field of breeding and genetics, it is necessary to preserve, maintain and develop the collections of the gene pool created by A.E. Kovarsky, study of the works of A.E. Kovarsky and his students, continue research, Discussion of results at scientific conferences, preservation of the memory of Anatoly Efimovich Kovarsky. Session I. General and molecular genetics 14 Session I GENERAL AND MOLECULAR GENETICS Session I. General and molecular genetics 15 DOI: https://doi.org/10.53040/cga12.01 UDC: 632.4:635.64 MOLECULAR DIAGNOSTICS OF SOME FUNGAL PATHOGENS IN GREENHOUSE GROWN TOMATOES Angela DEAGHILEVA1, ORCID: 0000-0003-3659-2450 angela.deaghileva@sti.usm.md Galina BELOUSOVA1, ORCID: 0000-0002-9977-1248 galina.belousova@sti.usm.md Irina CUZNETOVA1, ORCID: 0000-0001-5810-6244 irina.cuznetova@sti.usm.md Zoia IGNATOVA1, ORCID: 0000-0003-1171-5928 zoia.ignatova@sti.usm.md Karel MULLER2, ORCID: 0000-0002-0817-8810 e-mail: muller@ueb.cas.cz 1Institute of Genetics, Physiology and Plant Protection, Moldova State University, Chisinau, Republic of Moldova 2Institute of Experimental Botany of the Czech Academy of Sciences Abstract. Tomato production in protected environments such as greenhouses has become a popular trend in agriculture. The main advantage of greenhouse crop production over open field systems is the ability to control the environment, limiting the impact of abiotic and biotic factors. However, under greenhouse conditions tomatoes are still susceptible to certain soil-borne fungal diseases. Early detection of fungal pathogens is paramount for controlling the diseases and their spread. The aim of the study was to monitor fungal pathogens affecting tomato plants grown in protected conditions. Molecular diagnostics of pathogens Alternaria spp., Fusarium spp., Penicillium spp. and Aspergillus spp. was carried out during the vegetative season in various organs of tomato plants planting in greenhouse using nested-PCR assay. A. alternata was the predominant pathogen among all tested tomato genotypes. Fusarium and Aspergillus species were not detected in greenhouse tomato plantings during vegetative season. Keywords: fungi greenhouse, pathogens PCR Tomato. INTRODUCTION Global temperature rise is a trigger for climate change and, as a result, a multitude of risks to organisms all over the world. Climate change has a major impact on the agricultural sector, decreasing crop productivity. High temperatures and extreme fluctuations in precipitation quantities distort plant growth cycle, facilitate the emergence of new pests and diseases, their propagation and widening the host range [1, 2]. Threats of epiphytoties caused by various pathogens are Session I. General and molecular genetics 16 currently increasing in agroecosystems in temperate regions [3]. The increasing impact of pathogens has led to the excessive use of chemicals, which have negative consequences for the environment, human and animal health [4, 5]. Plant breeding programs and long-term use of pesticides have provoked the emergence of strains with increased virulence [6]. Numerous studies on the impact of climate change on biological processes demonstrate the need for effective measures to reduce the severity of diseases, using more environmentally friendly methods of plant protection. Mitigation methods include phytosanitary control of seed and planting material, the use of resistant varieties and microclimate modification. In the last decade, tomato production in protected environments such as greenhouses has become a new trend in agriculture. The main advantage of greenhouse crop production over open field systems is the ability to control the environment, limiting the impact of abiotic and biotic factors [7]. Greenhouse tomatoes have better quality and are uniform in shape, size and color compared to those grown in the open field. However, under greenhouse conditions tomatoes are still susceptible to certain microbial diseases, as the closed environment can promote the development of infections. Soil-borne fungi are the main limiting factor for tomato production and cause significant economic losses [8]. Early detection of fungal pathogens is paramount for controlling the diseases and their spread. The aim of the study was to monitor fungal pathogens affecting tomato plants grown in protected conditions. Molecular diagnostics of pathogens Alternaria spp., Fusarium spp., Penicillium spp. and Aspergillus spp. was carried out during the vegetative season in various organs of tomato plants grown in greenhouse using nested-PCR assay. MATERIALS AND METHODS Monitoring of phytopathogenic fungi was carried out during the 2024 growing season. The following tomato genotypes were studied: the wild tomato species Solanum pimpinellifolium, Elvira and Rufina varieties (selection of IGFPP). Asymptomatic tomato plants at different phases of development were tested: seeds before sowing (2023 harvest), leaves (phase of 4-5 leaves, flowering phase), fruits, “new” seeds. Total DNA from plant material were isolated by a modified CTAB method [9]. At all stages of the analysis, DNA extraction from tomato plants was performed in triplicate to ensure reproducibility of the results. Molecular analysis by nested-PCR was performed using primers for the detection of Alternaria spp., Fusarium spp., Penicillium spp. and Aspergillus spp. Primer design was done based on species-specific sequences of each pathogen from NCBI nucleotide collection presented in GenBank database using Primer-BLAST designing tool: glyceraldehyde 3-phosphate dehydrogenase gene (GPD) of A. alternata and A. solani; translation elongation factor 1-alpha (tef1) gene of F. Session I. General and molecular genetics 17 verticillioides, F. oxysporum, F. solani, F. avenaceum; beta-tubulin (tub2) gene sequences of F. equiseti, F. sporotrichioides, Penicillium citrinum; oxygenase (fum6) gene sequences of F. proliferatum; o-methyltransferase A (aflP) gene of Aspergillus flavus. The PCR mixture, in a volume of 25 μl, contained: 66 mM tris-HCl (pH- 8,4), 16 mM (NH4)2SO4, 2,5 mM MgCl2, 0,1% Tween 20, 7 % glicerol, 100 μg ml- 1 Bovin Serum Albumin, 0,2 mM dNTPs of each, 1,25 U Taq DNA polymerase, 5 pM of each primer, and 5-10 ng DNA. The first round of the nested-PCR included 1 cycle: 95 0C – 4 min, 60 0C – 1 min, 72 0C – 1 min; followed by 29 cycles: 95 0C –1 min, 60 0C – 1 min, 72 0C – 1 min. The second-round conditions of the nested-PCR were: 35 cycles: 95 0C – 1 min, 60 0C – 1 min, 72 0C – 1 min. The amplification products were separated in 1,5 % agarose gel by electrophoresis (5-8 V/cm) and compared with molecular marker (100 bp DNA ladder, Thermo Fisher Scientific) in a migration buffer of Tris/borate/EDTA (pH 8.0) with ethidium bromide, viewed in the UV (302 nm) and photographed. RESULTS AND DISCUSSION Prior to molecular diagnostics of pathogens, total DNA extracted from plant material was tested for the presence of PCR inhibitors. DNA quality analysis was performed using primers designed based on plant 18S ribosomal RNA gene sequences [10]. The results of amplification selectively demonstrated in Figure 1. Figure 1. Electropherogram of PCR products obtained on total DNA extracted from tomato seeds using primers to the 18SrDNA gene. 1-3. Solanum pimpinellifolium, 4-6. Elvira, 7-9. Rufina. M - 100 bp DNA ladder. The arrow points at a 500 bp marker fragment. Specific fragment of expected size (315 bp) was revealed for all tomato DNA samples. The analysis indicated that DNA was free of PCR inhibitors and was suitable for downstream pathogen testing. Molecular analysis of pre-planting seeds using species-specific primers to Alternaria spp. revealed the presence of Alternaria alternata, but absence of Alternaria solani in all genotypes of tomato. Penicillium citrinum was detected only in the seeds of Solanum pimpinellifolium (Figure 2). The positive signals in the second round of nested-PCR for A. alternata Session I. General and molecular genetics 18 were marked with the presence of a 288 bp amplicon on electrophoresis (Figure 2A.) and 250 bp – for P. citrinum (Figure 2B.). It is known that infected seeds can serve as a source of primary infection, increasing the pathogenic load in plants during the growing season. A. B. Figure 2. Electropherogram of nested-PCR products obtained using primers for detection of A. alternata (A.) and P. citrinum (B.) and total DNA extracted from the seeds. 1-3. Elvira, 4-6. Rufina, 7-9. Solanum pimpinellifolium. M - 100 bp DNA ladder. The arrow points at a 500 bp marker fragment. Thus, in order to obtain healthy planting material, a phytosanitary assessment of their quality is necessary before storing and compliance with storage conditions. A. alternata was revealed in the tomato plants at the phase of 4-5 leaves in all studied tomato genotypes (Figure 3). Figure 3. Electropherogram of nested-PCR products obtained using primers for detection of A. alternata in total DNA extracted from the tomato plants at the of 4-5 leaves. 1-3. Elvira, 4-6. Rufina, 7-9. Solanum pimpinellifolium. M - 100 bp DNA ladder. The arrow points at a 500 bp marker fragment. Alternaria spp. was not found in leaves collected at the flowering phase. The single incidence of A. alternata infection was revealed in ′Rufina′ fruits, but absent in “new” seeds of all tomato genotypes. We have previously shown that fungi of the genus Alternaria infect tomato plants in open field during all phases of development [11]. Therefore, in greenhouse conditions it is possible to contain the development of this pathogen. During the vegetative season, Fusarium, Aspergillus and Penicillium species (F. verticillioides, F. oxysporum, F. solani, F. avenaceum, F. equiseti, F. sporotrichioides, F. proliferatum, A. flavus, P. citrinum) were not detected in greenhouse grown tomatoes. In previous studies we detected these fungal species Session I. General and molecular genetics 19 at different stages of tomato development in open field conditions [12]. It is known, that Alternaria can inhibit the growth and development of Fusarium [13]. However, the effectiveness of suppression can be influenced by many factors, including environmental conditions and the genotype of the host plant [14]. Thus, greenhouse growing conditions and the presence of Alternaria spp. probably limited the development of the named species of Fusarium, Aspergillus and Penicillium. As shown, A. alternata was the predominant pathogen in tomato plants during some phases of development. So, it can be used as a representative pathogen for presymptomatic assessment of phytosanitary status of tomato plantations to optimize fungal disease control under greenhouse conditions. The probability of tomato infection by soil-borne fungal pathogens exists at all stages of cultivation. To overcome this risk, it is necessary to use healthy seeds, genotypes with low susceptibility to pathogens and to optimize the planting conditions. Success in combating pathogens largely depends on the correct application of plant protection measures to obtain high-quality and healthy products. Of great importance is the detection of potential foci of infection before the appearance of external signs of disease in affected plants. Seasonal monitoring of pathogens using PCR-based diagnostic methods is necessary. Presymptomatic detection of tomato pathogens allows optimizing the use of chemical plant protection products and obtaining a more environmentally friendly and healthy product and seed material. CONCLUSIONS As a result of monitoring some fungal infections of tomatoes in greenhouse conditions, a decrease in the pathogenic load in plants was found, compared to open field systems. A. alternata was the predominant pathogen among all tested tomato genotypes. Fusarium and Aspergillus species were not detected in greenhouse tomato plantings during vegetative season. None of the pathogen species declared in the study were detected in the fresh seeds, which is favorable for their subsequent reproduction. Acknowledgments: This study was supported by the research projects 011101 “Genetic and biotechnological approaches of management of agroecosystems under climate change conditions”, funded by the Ministry of Education and Research; UNDP-IRH-00181 „Introduction and application of new genomic techniques in agricultural sector to face climate challenges” funded by UNDP and Czech Republic Development Cooperation. REFERENCES 1. LAHLALI, Rachid, TAOUSSI, Mohammed, LAASL, Salah-Eddine et al. Effects of climate change on plant pathogens and host-pathogen interactions. In: Crop and Environment. 2024, Session I. General and molecular genetics 20 vol. 3, pp. 159–170. https://doi.org/10.1016/j.crope.2024.05.003 2. FILHO, Walter, NAGY, Gustavo, GBAGUIDI, Gouvidé et al. The role of climatic changes in the emergence and re-emergence of infectious diseases: bibliometric analysis and literature-supported studies on zoonoses. In: One Health Outlook. 2025, vol. 7:12, pp. 1-12 https://doi.org/10.1186/s42522-024-00127-3, 3. IPPC SECRETARIAT. Scientific review of the impact of climate change on plant pests – A global challenge to prevent and mitigate plant pest risks in agriculture, forestry and ecosystems. 2021. Rome. FAO on behalf of the IPPC Secretariat. https://doi.org/10.4060/cb4769en 4. SINGH,  Brajesh, DELGADO-BAQUERIZO, Manuel, EGIDI,  Eleonora. Climate change impacts on plant pathogens, food security and paths forward. Nature Reviews Microbiology. 2023, vol. 21, pp. 640–656. https://doi.org/10.1038/s41579-023-00900-7 5. ASHWINI, Kumar, MAHANTA, Dibyajyoti, DANGE, Mohini et al. Global Challenges Facing Plant Pathology: A Review on Multidisciplinary Approaches to Meet the Food Security. In: Journal of Scientific Research and Reports. 2024, vol. 30 (6), pp. 884-892. https://doi.org/10.9734/jsrr/2024/v30i62106. 6. MILLER, Sally, FERREIRA, Jorge, LEJEUNE, Jeffrey. Antimicrobial Use and Resistance in Plant Agriculture: A One Health Perspective. In: Agriculture. 2022, vol. 12, 289, pp.1-27. https://doi.org/ 10.3390/agriculture12020289 7. ÁVALOS-SÁNCHEZ, Eugenio, MORENO-TERUEL, María, LÓPEZ-MARTÍNEZ, Alejandro et al. Effect of Greenhouse Film Cover on the Development of Fungal Diseases on Tomato (Solanum lycopersicum L.) and Pepper (Capsicum annuum L.) in a Mediterranean Protected Crop. In: Agronomy. 2023, vol. 13, 526, pp. 1-14. https:// doi.org/10.3390/agronomy13020526 8. ALLY, Nooreen, NEETOO, Hudaa, RANGHOO-SANMUKHIYA, Vijayanti, COUTINHO, Teresa. Greenhouse-grown tomatoes: microbial diseases and their control methods: a review. In: Int. J. Phytopathology. 2023, vol. 12 (01), pp. 99-127. 10.33687/phytopath.012.01.4273 9. ABOUL-MAATY, Nadia, ORABY, Hanaa. Extraction of high-quality genomic DNA from different plant orders applying a modified CTAB-based method. In: Bulletin of the National Research Centre. 2019, 43:1, pp. 1–10. https://doi.org/10.1186/s42269-019-0066-1 10. DIAGHILEVA, Angela, MITIN, Valentin, PASHA, Lilia, TUMANOVA, Lidia. Molecular examination of tomato plants whith TYLCV- like symptoms. In: Buletinul Academiei de Ştiinţe a Moldovei. Ştiinţele vieţii, 2016, nr. 3(330), pp. 104-108. ISSN 1857-064X. 11. GRĂJDIERU, Cristina, DIAGHILEVA, Angela, TUMANOVA, Lidia, MITIN, Valentin. Molecular analysis of some toxigenic fungi in Moldavian tomato varieties. In: Тенденции развития агрофизики: от актуальных проблем земледелия и растениеводства к технологиям будущего: Посвящено памяти академика Е.И. Ермакова, 2-4 octombrie 2019, St. Petersburg, Russia: ФГБНУ АФИ, pp. 302-307. ISBN 978-5-905200-40-3. 12. DIAGHILEVA, Angela. Identification of mycotoxin-producing species of fusarium in tomato ontogenesis. In: Genetica, fiziologia şi ameliorarea plantelor, 7-8 octombrie 2024, Chişinău, Republica Moldova, pp. 75-79. ISBN 978-9975-62-766-5. 13. SCHIRO, Gabriele, VERCH, Gernot, GRIMM, Volker, MÜLLER, Marina. Alternaria and Fusarium Fungi: Differences in Distribution and Spore Deposition in a Topographically Heterogeneous Wheat Field. In: J. Fungi. 2018, vol. 4, 63, pp. 1-24 10.3390/jof4020063 14. EBADI, Mostafa, EBADI, Ali. Genetic diversity and population structure of Alternaria alternata: An endophytic fungus isolated from various hosts. In: Fungal Biology. 2024, vol.128 (8), pp. 2305-2310. https://doi.org/10.1016/j.funbio.2024.11.005 Session I. General and molecular genetics 21 DOI: https://doi.org/10.53040/cga12.02 UDC: UDC: 631.445.4:631.46(478) POOL AND DIVERSITY OF PROKARYOTES OF CARBONATE CHERNOZEM OF LONG-TERM FIELD EXPERIENCE OF MOLDOVA Nina FRUNZE, ORCID: 0000-0001-7263-5863, ninafrunze@mail.ru Institute of Microbiology and Biotechnology, Technical University of Moldova, Chisinau, Republic of Moldova Abstract. For the first time in Moldova, changes in microbiological indicators obtained using polymerase chain reaction (PCR) in carbonate chernozem were studied depending on the type of land use under different anthropogenic loads. The work used soil samples from winter wheat crop rotation of long-term field experiments of different types of use: without fertilizers, mineral and organic background, as well as 75-year-old fallow land . A high diversity of genetic information with a spectrum consisting of 17 phyla was established. According to the International Committee on Prokaryotic Taxonomy (2021), they are new, identified and / or confirmed by metagenomic analysis. The prokaryotic pool varied from 0.01% to 46.53%: Actinomycetota (38.17-46.53%), Pseudomonadota (19.60-27.02%), Bacillota (7.32-22.40%), Bacteroidota (5.31-8.55%), Acidobacteriota (1.94-3.32%), Verrucomicrobiota (1.37-2.09%), Myxococcota (1.09-1.71%), Nitrospirota (0.15-0.65%), Planctomycetota (0.58-0.65%), Gemmatimonadota (0.36-0.58%), Patescibacteria (0.08-0.17%), Cyanobacteriota (0.05-0.08%), Chloroflexota (0.03-0.07%), Fibrobacterota (0.01-0.05%), Abditibacteriota (0.01-0.07%), Bdellovibrionota(0.01-0.011%), and Nitrososphaerota (7.32-22.4%). The first 16 are representatives of the Bacteria domain, and the Nitrososphaerota phylum belongs to the Archaea domain. Based on the results of multiple observations of the content of conserved DNA regions of different phylogenetic groups in the soil, it can be concluded that the prokaryotic community is susceptible to the type of chernozem use. As an alternative to soil conditions, bacteria act in the following sequence: soil without fertilizers → organic background → mineral background → fallow land, and archaea - vice versa. Keywords: prokaryotes, bacteria and archaea, 16S rAPH gene, metagenomic analysis. INTRODUCTION Prokaryotes, whose representatives include two domains – Bacteria and Archaea, are “the basic unit and universal basis of life” [6]. Approximately 5% of the total number of prokaryotes on Earth are concentrated in the soil [4], characterizing the soil environment as the largest reservoir of microbial diversity on Earth. The majority belongs to bacteria, while archaea account for 0.5 to 3.8% of the total number of all prokaryotes. However, the high level of genetic diversity of the soil microworld remained unknown for many years [1]. Only the advent of new methods of molecular ecology of microorganisms Session I. General and molecular genetics 22 revealed that at present, only about 0.1% of the total microbial diversity of the biosphere is taken into account by the cultivation of microorganisms [2]. With the introduction of methods of molecular ecology of microorganisms, science has gained the opportunity to take into account not only cultivated but also uncultivated forms of microorganisms [8, 12]. It is believed that the microbial community is a kind of "mirror" that reflects the features of the habitat, and its diversity represents the hidden potential of the soil microworld [1, 5] in connection with which the metagenomic characteristics of soils are in demand for assessing the sustainability of soil ecosystems under the influence of natural and anthropogenic factors, and interest in such studies is constantly increasing. The soils of Moldova have not been studied from this point of view. The aim of this study was to study the pool and diversity of prokaryotes of the carbonate chernozem of the Republic of Moldova using quantitative PCR and high-throughput sequencing of 16S rRNA gene libraries. MATERIALS AND METHODS The object of the study was microbial communities of carbonate chernozem: light loam with a humus content of 2.5-3.0%, mobile phosphate of 0.8-1.5 mg / 100 g, exchangeable potassium of 18-22 mg / 100 g and carbonates of 1.8-2.2% in the 0-20 cm layer. The studies were carried out in 2022 in a long-term stationary field experiment (founded in 1950) of the Ketrosy scientific and educational farm, district Anenii Noi . Soil samples were collected in the spring from variants with winter wheatof eight-field оf crop rotation : 1 - without fertilizers, 2 - with the application of mineral fertilizers, 3 - with the application of organic fertilizers and 4 - fallow land. Fertilizer application rates: total for crop rotation N675P480K480 (N90P60K60 for corn, N120P60K60 for winter wheat before corn and before sunflower, N45P60K60 for sunflower and peas); total for crop rotation 144 t/ha of semi-rotted manure (40 t for corn, 22 t for winter wheat, 20 t for sunflower). Metagenomic analysis was carried out using modern methods [3, 4] and equipment of the Collective Use Center “Genomic Technologies, Proteomics and Cell Biology” of the All-Russian Research Institute of Agricultural Microbiology, St. Petersburg, Russia. Taxonomic identification of OTUs was performed using the RDP (SILVA) database, the classification was refined using the online database https://www.ncbi.nlm.nih.gov/taxonomy) [9, 10, 11]. RESULTS AND DISCUSSION A study of long-term used carbonate chernozem soil revealed that its prokaryotic community consists of 17 phylums: Actinomycetota, Session I. General and molecular genetics 23 Pseudomonadota, Bacteroidota, Bacillota, Acidobacteriota, Verrucomicrobiota, Planctomycetota, Myxococcota, Nitrospirota, Gemmatimonadota, Cyanobacteriota, Patescibacteria, Chloroflexota, Fibrobacterota, Abditibacteriota, Bdellovibrionota, и Nitrososphaerota (Fig. 1). The first 16 are representatives of the Bacteria domain, and Nitrososphaerota belonged to the domain Archaea. According to the quantitative assessment of the content of conservative DNA regions of different phylogenetic groups in the soil, it can be concluded that the bacterial community is most responsive to the type of use of chernozems. As an alternative to soil conditions, bacteria act in the following sequence: soil without fertilizers → organic background → mineral background → fallow land, and archaea - vice versa. Figure 1. Taxonomic structure of prokaryotic communities of carbonate chernozem at the phylum level, %: A – without fertilizers. B – mineral background, C – organic background, D – fallow land The identified microbiome was similar in the composition of its components during the study, but the presence of phyla in the variants was different. All phyla were simultaneously present only in the soil of the crop rotation without fertilizers (control). In the fertilized variants, 15 phyla were identified, and in the fallow soil – only 13. The phyla also differed in their proportion of representation in the community. Among the identified prokaryotes, the highest abundance was recorded in microorganisms of the Bacteria domain (77.60–92.68%). The leaders among them were representatives of the Actinomycetota phylum (38.17–46.53%), reaching the highest value in soil without fertilizers and the lowest in soil fertilized with organic fertilizers for a long time and in the fallow land. Next, among the majority of components of the prokaryotic community, representatives of Pseudomonadota (19.60-27.02%) were widely encountered, which in general were almost 2 times inferior to actinobacteria in numbers, recording a similar development with actinobacteria in variants. The third in terms of representativeness was the archaeal phylum Nitrososphaerota (7.32-22.40%). 0% 20% 40% 60% 80% 100% A B C D Bacteria. Actinomycetota Bacteria. Pseudomonadota Archaea. Nitrososphaerota Bacteria. Bacillota Bacteria. Bacteroidota Bacteria. Acidobacteriota Session I. General and molecular genetics 24 despite the fact that it was represented almost twice less than the previous one, it recorded the highest ratios in the fallow and the lowest in the unfertilized soil, while the fertilized variants occupied an intermediate position. In addition, it should be emphasized that the contribution of archaea in the prokaryotic community doubled in fertilized variants (13.04-13.25%) in relation to unfertilized soil (7.32%), while at the same time noting a doubled representation in the fallow land soil (22.40%) in relation to fertilized variants (13.04-13.25%) and an almost 4-fold excess in relation to unfertilized crop rotation soil (7.32%). The phylum Bacillota was also characterized by high rates (6.08.-12.42%), distinguished by the highest values of abundance in the organic background soil and the lowest in unfertilized soil and fallow land. The Bacteroidota phylum had a taxa abundance of 5.31-8.55% with the highest indicators in the fertilized variants (6.90-8.55%).Further, attention is drawn to the phyla that did not differ in either large fluctuations or large values of abundance in the community: Acidobacteriota (1.94-3.32%) Verrucomicrobiota (1.37-2.09%) and Myxococcota (1.09-1.71%). They recorded the highest indicators in the soil of the organic background or in the fallow soil, and the lowest - in the unfertilized soil. Of these, the highest values were recorded by the phylum Acidobacteriota (3.32-1.94%) and the lowest - by the phylum Myxococcota, and Verrucomicrobiota (1.37-2.09%) occupied an intermediate position among them. The following 9 phyla had very small contributions to abundance prokaryotic community. Moreover, the first 5 phyla: Nitrospirota (0.15-0.65%), Planctomycetota (0.58-0.65%), Gemmatimonadota (0.36-0.58%) Patescibacteria (0.08-0.19%) Fibrobacterota (0.01-0.05%) were found comparatively more often and showed 0.03-0.65% contribution, in comparison with the rest. And the phyla Aditibacteriota and Bdellovibrionota with a contribution of only 0.01-0.07% were not identified in the soil of fertilized variants and in the fallow land soil. Ranking of phyla by individual “significance” abundence revealed that long-term agricultural use of carbonate chernozem provoked a strong restructuring of the prokaryotic community. The same composition of components hides a distinctive hierarchical role of taxa in the community (Fig. 2). Based on this, all phyla can be divided into 3 groups, which, being constant in composition, differ in their relative abundance throughout the study period. The first group with a dominant ecological role and an occurrence frequency of ≥5% had the highest representation among these 3 groups, individually differing significantly both in the variant and among them, accounting for about 91.56–93.63% of 16S rRNA genes from all sequences without significant differences among variants themselves.It consisted of 5 phyla, which in order of decreasing abundance were as follows: Actinomycetota (38.17–46.53%), Pseudomonadota (19.60–27.02%), Nitrososphaerota (7.32–22.40%), Bacillota (6.08–12.42%) and Bacteroidota (5.31– 8.55%). The second group, commonly found commonly encountered, was formed Session I. General and molecular genetics 25 from 3 phyla with a secondary ecological role and abundance ≤ 5% and averaged about 4.40–6.60%, recording an increase of 25% in the organic background soil and 50% in the fallow soil, compared to the rest. Moreover, the greatest contribution of Acidobacteriota (1.94–3.32%) and Verrucomicrobiota (1.37-2.09%) was recorded in the fallow soil, and Myxococcota (1.09-1.71%) in the organic background. The smallest contribution of this group of microorganisms was noted in the unfertilized soil of the crop rotation. Representatives of the second group of prokaryotes formed the following descending sequence: Acidobacteriota (1.94–3.32%) → Verrucomicrobiota (1.37–2.09%) → and → Myxococcota (1.09–1.71%) – in the soilorganic background. The third group included rarely seen representatives of prokaryotes with an abundance of ≤ 1%, having an insignificant ecological role in the community and amounted to only 1.84-1.97%, differing slightly in variants and at the same time constituting the smallest group of prokaryotes in terms of contribution and the largest group among them in terms of the number of phyla (9). Group ≥5%. 93,63 93,13 92,93 91,56 Group 1-5% 4,4 4,93 5,17 6,6 Group ≤ 1%. 1,97 1,94 1,9 1,84 A B C D 0 10 20 30 40 50 60 70 80 90 100 A b u n d an ce o f p ro ca ry o te s, % Figure 2. Redistribution of the relative participation of prokaryotes under winter wheat in carbonate chernozem, according to the degree of dominance (d.d.), %: A – unfertilized, B – mineral background, C – organic background, G – fallow land They were arranged in descending order of abundance as follows: Nitrospirota (0.15–0.65%) → Planctomycetota (0.58–0.65%) → Gemmatimonadota (0.36–0.58%) → Patescibacteria (0.08–0.19%) → Cyanobacteriota (0.05–0.08%) → Chloroflexota (0.03–0.07%) → Fibrobacterota (0.01–0.05%) → Abditibacteriota (0.01–0.011%) and → Bdellovibrionota (0.01– 0.07%). Consequently, under the influence of long-term agricultural use of carbonate chernozem, physicochemical and climatic factors from year to year, since the beginning of time, a significant restructuring and redistribution of the prokaryotic community occurs. Microorganisms, changing the degree of their dominance in the Session I. General and molecular genetics 26 community, losing or acquiring a new ecological status, survive and remain an important component of the microbiome of carbonate chernozem [7, 12]. It is these properties that characterize prokaryotes as structural elements of living matter in the soil, which interact with each other in the struggle for nutrients, thereby ensuring the role of the molecular alphabet of living matter. CONCLUSION 1. Carbonate chernozem is characterized by high genetic diversity of the most ancient representatives of the living world - prokaryotes, the spectrum of which consists of 17 phyla of both domains, similar to soils of other regions. But at the same time, all were present only in the unfertilized soil of the crop rotation. 2. The pool of prokaryotes ranged from 0.01% to 46.53% with the highest values in the unfertilized soil and the lowest in the organic background and fallow soil. The mineral background occupied an intermediate position among them. 3. According to the estimates of the International Committee on Prokaryotic Taxonomy (2021), the phyla are new, identified and / or confirmed by metagenomic analysis. Acknowledgments: The work was supported by the National Agency for Research and Development of the Ministry of Education and Science of the Republic of Moldova (project 20.80009.5107.08). The text of the article was prepared using the funds of the institute project No. 020101 of the Ministry of Education and Science of the Republic of Moldova “Innovative biotechnological solutions for agriculture, medicine and environmental protection”. REFERENCES 1. ANDRONOV, Е. Е., PETROVA, S.N., PINAEV, A.G., PERHINA, E.V. et. al. Analysis of the Structure of Microbial Community in Soils with Different degrees of Salinization Using T_RFLP and Real Time PCR Techniques // Eurasian Soil Science. 2012. Vol. 45, No. 2, pp. 147–156. https:// 10.1134/S1064229312020044. 2. ASLAM, Z. Too much bacteria still unculturable / Z. Aslam, M. Yasir, A. Khaliq, K. Matsui, Y.R. Chung // Crop & Environment. 2010. V. 1. P. 59–60. 3. BATES, S.T., BERG-LYONS, D., CAPAROSO, J.G. et al. Examining the global distribution of dominant archaeal populations in soil // ISME J. 2011. 5: 908–917. https://doi.org/10.1038/ismej. 2010.171. 4. CAPAROSO, J.G., KUCZINSKI, J., STOMBBAUGH, J. et al. Correspondence QIIME allows analysis of high- throughput community sequencing data Intensity normalization improves color calling in SOLiD sequencing // Nature Publishing Group. 2010. 7 (5): 335- 336. https://doi.org/10.1038/nmeth.f.303 5. CHERNOV, T.I., LEBEDEVA, M.P., TKHAKAKHOVA, A.K. et al. Profile analysis of microbiomes in soils of solonetz complex in the Caspian Lowland // Eurasian Soil Sci. 2017. 50:64–69. https://doi.org/10.1134/S1064229317010045. 6. DANIEL, R. The metagenomics of soil // Nature Reviews Microbiology. 2005. V. 3. P. 470– Session I. General and molecular genetics 27 478. 10.1038/nrmicro1160 7. IVANOVA, E.A., KUTOVAYA, O.V., TKHAKAKHOVA, A.K. et al. The structure of microbial community in aggregates of a typical chernozem aggregates under contrasting variants of its agricultural use // Eurasian Soil Sci. 2015. 48:1242–1256. https://doi.org/10.1134/S1064229315110083. 8. KIMEKLIS, A.K., DMITRAKOVA, Y.A., PERSHINA, E.V. et al. Analysis of microbiome of recultivated soils of the kingisepp area of phosphorite mining // Sel'skokhozyaistvennaya Biologiya. Agricultural Biology. 2020. 55:137-152. https://10.15389/agrobiology. 9. KUTOVAYA, O.V., LEBEDEVA, M.P., TKHAKAKHOVA, A.K. et al. Metagenomic Characterization of Biodiversity in the Extremely Arid Desert Soils of Kazakhstan // Eurasian Soil Science, 2015, Vol. 48, No. 5, pp. 493–500. https://10.1134/S106422931505004X 10. OREN, A., GARRITY, G.M. Valid publication of the names of forty-two phyla of prokaryotes // International Journal of Systematic and Evolutionary Microbiology. 2021. 71 (10): 5056. 10.1099/ijsem.0.005056 11. PARKS, D.H., CHUVOCHINA, M., WAITE, D.W. et al. A standardized bacterial taxonomy based on genome phylogeny substantially revises the tree of life // Nat Biotechnol. 2018. 36: 996-1004. 10.1038/nbt.4229 12. ZVEREV, A.O., KICHKO, A.A., PINAEV, A.G., Diversity Indices of Plant Communities and Their Rhizosphere Microbiomes: An Attempt to Find the Connection // Microorganisms. 2021, 9, 23-39. https://doi.org/10.3390/ microorganisms 9112339 1-11 Session I. General and molecular genetics 28 DOI: https://doi.org/10.53040/cga12.03 UDC: 595.79:631.527.82:631.147 METHOD OF ACTIVATING ENTOMOPHAGES AND POLLINATORS HYMENOPTERA ORDER NATURAL POPULATIONS WITH THE AIM OF INCREASING THE AGROSYSTEM PRODUCTIVITY Alla GLADCAIA, ORCID: 0000-0001-9182-4352 alla.gladcaia@sti.usm.md Lilia CHISNICEAN Institute of Genetics, Physiology and Plant Protection, Moldova State University, Chisinau, Republic of Moldova Abstract. Insects provide important ecosystem services of great economic value. We investigated a method for activating of entomophages and pollinators of Hymenoptera order natural populations ("field breeding"), consisting of a combination (synergy) of overseeding spice plants and placing Fabre hives nearby the fruit garden. The objects of the study were entomophages and pollinators of the Hymenoptera order. The experiment was conducted in three different biotopes: an apple orchard near a forest belt, a collection of spice plants near a pear orchard, and a field of Miscanthus sinensis. The maximum number of Megachilidae bee nests were found in artificial dark- colored shelters: made of cylindrical dark plastic (70,8%), and deltoid dark cardboard (20,8%). The bees preferred to build nests in the agrocenosis of the pear orchard nearby a plot of spice plants (91,7%). It has been established that the optimal strategy for the method of entomophage’s activating consists of combining a variety of different family’s spicy plants and ensuring continuous flowering (conveyor). Keywords: entomophages, field breeding, artificial shelters, Hymenoptera, aromatic plants. INTRODUCTION Insects provide important ecosystem services of great economic value. Proper management of agro-landscape resources can contribute to the biodiversity conservation. In this context, agri-environmental schemes (AES) were included in the EU agricultural policy in 1985 and have become the main instrument for biodiversity conservation in Europe [1]. We investigated a method for activating of entomophages and pollinators of Hymenoptera order natural populations ("field breeding"), consisting of a combination (synergy) of overseeding spice plants and placing Fabre hives nearby the fruit garden, which contributes to an increase in the beneficial insect’s survival and activity, and supports the formation of stable self- developing and self-regulating agrobiogeocenoses [2]. Since many predators are also anthophiles, their survival, productivity and performance are enhanced, when Session I. General and molecular genetics 29 flowering herbaceous plants are present within the agrocenosis. Research by scientists has shown, that strips of flowering plants near gardens and fields increase the biodiversity of pollinators and entomophages [3]. Flowering plants of the Apiaceae and Lamiaceae families significantly increase the number of beneficial insects in agroecosystems [4]. Wild bees (e.g. Osmia spp.) are more effective in pollinating fruit crops than honey bees. The placement of Fabre hives for Osmia increases the yield of fruit orchards several times [5]. Osmia bees prefer high-lipid pollen with a high content of sterols (fam. Asteraceae) [6]. Analysis of the nectar of essential oil plants showed that maltose and sucrose dominate in plants, pollinated by bees [4]. The key amino acid for the energy metabolism of solitary wasps (Sphecidae) and bees is proline, and the amino acids tryptophan and phenylalanine increase the survival and fertility of parasitoid wasps (Braconidae) [7, 8]. Terpenoids (linalool, geraniol) in the nectar of lavender and mint attract bees and repel unwanted insects [9]. To adapt the method to a specific garden, it was important to study local bee species and the optimal set of partner plants. The aim of our work was to study the method of entomophage’s natural populations activating, which involves the inclusion of a spice plant’s plot in the industrial garden array, in combination with the use of artificial shelters to attract of Hymenoptera entomophages and pollinators to the agrocenosis. MATERIALS AND METHODS The place and time of the research are laboratory and field conditions of the Institute of Genetics, Physiology and Plant Protection of the Republic of Moldova, 2024. The development of technological elements of artificial shelters for attracting economically important Hymenoptera order entomophages was carried out on the basis of certain modifications of Fabre hives. The objects of the research were entomophages and pollinators of the Hymenoptera order. In order to evaluate the most attractive filling materials, we used hollow tubes (reed and plastic) in each shelter. Four variants of the enclosure were considered: 1) white cardboard; 2) dark cardboard; 3) transparent plastic; 4) dark plastic. The experiment was conducted in three different biotopes: an apple orchard near a forest belt, a collection of spice plants near a pear orchard, and a field of Miscanthus sinensis (provides insects with natural tubular stems for nesting). The sowing of spicy plants, which are widely used in medicine, culinary and perfumery, contributed to the increase in the economic efficiency of the method. The diversity of species that make up the collection of spicy-aromatic plants differs not only in botanical origin, but also in the period of flowering and secretion of nectar and pollen. The attractiveness of artificial shelter options and agrocenoses was assessed by comparing the density of the tubes with Hymenoptera. The insects Session I. General and molecular genetics 30 were counted by sequentially inspecting of artificial shelter’s sections, determining the taxonomic affiliation of individuals and counting them. RESULTS AND DISCUSSION A dependence of the Hymenoptera nests number on the artificial shelter structure and the type of agrocenosis was established. The maximum number of solitary bees (Megachilidae) nests were found in cylindrical shelters made of dark plastic (70,8%), delta-shaped shelters made of dark cardboard (20,8%) and 8,4% in delta-shaped shelters made of white cardboard. Tubular reed stems with a diameter of 0,7–0,9 mm were populated, and bee nests were not found in plastic tubes. Thus, bees prefer dark-colored shelters and tubular reed stems (Figure 1). Figure 1. Population of artificial shelters by wild solitary bees depending on the type of body and material In the agrocenosis of the pear orchard near the spice plot nested 91,7 % of bees, in the agrocenosis of the apple orchard - 4,2 %, and 4,1 % - in the agrocenosis of the Miscanthus sinensis field. In our studies in 2024, we obtained imagoes and nests of solitary bees from two genera (Megachilidae): one of the largest genera of bees in the family, the genus Osmia Panzer, 1806 (mason bees) and the genus Stelis Panzer, 1806 (cuckoo bees) (Figure 2). Inside the cavity of the reed stems, Osmia bees divided the passage into brood chambers. Each chamber was supplied with enough pollen to feed one larva. The bee laid an egg on each pollen mass and sealed the chamber. The end of the tunnel was sealed with mud, plant resins, pieces of leaves or flower petals. Osmia bees pollinate an extremely wide range of flowering plants. Stelis bees lay eggs in the nests of other bees (Figure 3). Session I. General and molecular genetics 31 Figure 2. Population of artificial shelters and agrocenosis types by Megachilid family bees in various agrocenoses Figure 3. Nesting of solitary bees (Megachilidae). Bee nests in tubular reed stems with multi-colored plugs (1), open bee nests (2), imago of the genus Osmia bees (3), imago of Stelis (4). The hosts of Stelis are other Megachilinae. In most cases, the parasite, having discovered the host's nest, returns to it repeatedly to lay an egg in each of several host cells before they close. The ability of bees to populate anthropogenic cavities of suitable sizes and the wide polytrophism of species allows us to consider Megachilidae bees as very promising species for field breeding in artificial shelters in order to increase the economic efficiency of agroecosystems. In order to maintain and activate the population of Hymenoptera order entomophages and pollinators in the agrocenosis, a food base was provided from the continuous flowering of spice plants throughout the season. The flowering period in the collection of spice and aromatic plants in 2024 began on April 17 with the flowering of Bergénia crassifólia L. (Saxifragaceae) and Adonis vernalis L. (Ranunculaceae), which lasted for 18-20 days, respectively, and the visit (mainly Session I. General and molecular genetics 32 by bumblebees) was average. Then, from April 24 (flowering for 25 days), the flowering of Nepeta transcaucazica L., (Lamiaceae) and Veronica officinalis L. (Plantaginaceae) began. Their flowering is shorter 13-15 days, as well as Rumex acetosa L., Rheum officinale Baill. visits by insects, which are more numerous and varied. Starting from April 25 and for 25-30 days, species, such as Thymus vulgaris L., Thymus serpilium L., Thymus longicaulis L., Salvia officinalis L. (Lamiaceae) bloomed, and at the end of April, beginning of May – Rosa gallica L. and Sanguisorba officinalis L. (Rosaceae). From the beginning of May and for 20-25 days Thymus x citriodora Schreb., Asparagus officinalis L., Ruta graveolens L. (with inconspicuous flowers, but very much visited by insects), Veronica chamaedrys L., Matricaria chamomilla L., Plantago lanceolata L., Linum usitatisimum L., Centaurea cianus L., Allium shoenoprasum L. bloomed. In early June and for 25-30 days, most species of the Lamiaceae family bloom and secrete nectar, which are well visited by insects. Among them are Lavandula angustifolia Mill., Melissa officinalis L., Hyssopus oficinalis L., Dracocepalum moldavica L. The month of July is rich in flowering of the Apiaceae family species, which are visited by insects from the moment of the budding phase of plants (lasts from 5 to 13 days for different species). During this period, from July 2 to 5, insects "check" the umbrellas for nectar. The most frequent visits by insects are observed during the period of the flowering beginning and until the flowers in the umbrella have completely faded. This period begins on July 5 and lasts from 25 days for the annual species Anethum graveolens L. to 25 days and up to 30-35 days for the biennial species Passtinaca sativa L., Apium graveolens L., Foeniculum vulgare Mill., Ammi visnaga L., Anhtriscus cerifolium L. – equal to 25-35 days. In August- September, such species as Allium ramosum L., Satureja montana L., Inula helenium L. bloom. Thus, the site saw continuous flowering (conveyor) of 34 species of spice plants throughout the growing season (April-September). CONCLUSIONS 1. The degree of artificial shelters attractiveness for Hymenoptera order entomophages was established, depending on the design and materials from which they are made. The maximum number of bee’s nests of the Osmia and Stelis (Megachilidae) genera were found in artificial shelters of a dark color: from cylindrical dark plastic (70,8%), from deltoid dark cardboard (20,8%). Bees preferred to build nests in the agrocenosis of a pear garden near a plot of spice plants (91,7%); 2. It was established that the optimal strategy for the activating entomophages method consists in combining a variety of spice plants of different families and ensuring continuous flowering (conveyor). This composition of the Session I. General and molecular genetics 33 collection has a long flowering of species and provides sufficient food for insects living nearby, an eco-friendly environment, and is also important for creating favorable conditions for pollination and increasing seed productivity of spicy- aromatic and garden crops. Acknowledgements: The research was carried out within the framework of subprogram 011103 “Development of environmentally friendly means of reducing the impact of agricultural pests in the context of climate change”, funded by the Ministry of Education and Science. REFERENCES 1. GLADCAIA, A. А. Application of nest devices for the entomophages (Chrysopa, Chrysopidae, Neuroptera) accumulation in agrobiocenoses for biological protection of plants. In: Journal V. I. Ekosistemy. 2022 – Vol. 30, pp. 158–166. ISSN 2414-4738. 2. BEYER, N., Kulow J., Dauber J. The contrasting response of cavity-nesting bees, wasps and their natural enemies to biodiversity conservation measures. In: Insect Conservation and Diversity. 2023, pp. 1–15. ISSN: 1752-4598. 3. HAALAND, C., Naisbit, R. E., & Bersier, L. F. Sown wildflower strips for insect conservation: a review. In: Insect Conservation and Diversity, 2011. vol. 4, nr. 1, pp. 60-80. ISSN: 1752-4598. 4. FIEDLER, A. K., & Landis, D. A. Attractiveness of Michigan native plants to arthropod natural enemies and herbivores. In: Environmental Entomology. 2007, vol. 36, nr. 4, pp. 751- 765. ISSN: 0046-225X. 5. BOSCH, J., & Kemp, W. P. Developing and establishing bee species as crop pollinators: the example of Osmia spp. (Hymenoptera: Megachilidae). In: Fruit Trees. 2002, vol. 28, nr. 1, pp. 1-10. ISSN: 2151-8467. 6. RAGUSO, R. A. Wake up and smell the roses: The ecology and evolution of floral scent. In: Annual Review of Ecology, Evolution, and Systematics. 2008. nr. 39, pp. 549-569. ISSN: 1545-2069. 7. PETANIDOU, T., et al. What shapes amino acid and sugar composition in Mediterranean floral nectars? In: Oikos. 2006, vol. 115, nr. 1, pp. 155-169. ISSN:1600-0706. 8. CARTER, C., et al. Tryptophan and phenylalanine in nectar enhance parasitoid fecundity. In: Ecology Letters. 2006, vol. 9, nr. 12, pp. 1293-1301. ISSN:1461-0248. 9. WÄCKERS, F. L. A comparison of nectar- and honeydew sugars with respect to their utilization by the hymenopteran parasitoid Cotesia glomerata. In: Journal of Insect Physiology. 2001, vol. 47, nr. 9, pp. 1077-1084. ISSN: 1879-1611. Session I. General and molecular genetics 34 DOI: https://doi.org/10.53040/cga12.04 UDC: 633.11:575.224.2 ASSESSING GENETIC DIVERSITY IN TRITICALE POPULATIONS USING iPBS MARKERS Svetlana GOLOVCOVA, ORCID: 0009-0002-7541-1181 svetlana2004golovcova@gmail.com Moldova State University, Republic of Moldova Abstract. This article aimed to evaluate the efficacy of the iPBS (inter-Primer Binding Site) molecular marker system, leveraging mobile genetic elements, for genotyping diverse triticale accessions sourced from the collection of the Institute of Genetics, Physiology and Plant Protection. The objectives included the isolation and purification of genomic DNA from 7-day-old triticale seedlings of 'Ingen-54' and 'Ingen-93' genotypes, the empirical determination of optimal iPBS primer efficiency through a series of experimental amplifications, and the subsequent selection of highly informative primers for triticale genotyping. Standard polymerase chain reaction (PCR) amplifications were conducted utilizing specific iPBS primers (Pb13, Pb20, Pb21), followed by electrophoretic separation of amplification products on agarose gel. Amplification produced 33 clear, 100% polymorphic loci. POPGENE analysis revealed high informativeness (Nei’s gene diversity h=0.1290, Shannon index I=0.2403) despite low genetic distance (0.0074–0.0169) between the closely related varieties. Primers Pb20 and Pb21 generated the most polymorphic loci. These results confirm the high utility of iPBS markers for genetic diversity assessment and cultivar identification in triticale breeding, even for closely related genotypes. Keywords: genotyping, polymorphisms, retrotransposons molecular markers, triticale. INTRODUCTION Triticale (× Triticosecale Wittmack) is an artificial amphidiploid species derived from a cross between wheat (Triticum aestivum) and rye (Secale cereale). This hybrid was developed to combine the agronomically valuable traits of both progenitors: the high yield and grain quality of wheat with the cold tolerance, disease resistance, and environmental adaptability typical of rye.[1, 2] In terms of nutritional and feed value, triticale holds an intermediate position between wheat and rye. Triticale has high antioxidant activity due to its rich content of polyphenolic compounds and flavonoids, particularly phenolic acids such as 4- hydroxybenzoic acid, vanillic acid, caffeic acid, chlorogenic acid, p-coumaric acid, and rosmarinic acid [3]. Due to its combined traits, triticale isonsidered a promising cereal crop, especially in regions with harsh climatic conditions and marginal soils. The world production of triticale currently exceeds 20 million tons per year and is mainly concentrated in Europe, accounting for over 80% of the global output. In the countries of Central and Eastern Europe, including Romania and Ukraine, Session I. General and molecular genetics 35 triticale is actively introduced into crop rotations as an alternative to winter wheat on low-productivity soils.[4] Its adaptability, resistance to abiotic stress, and relatively low input requirements make it an important candidate for sustainable agriculture and food security in the context of climate change. In Moldova, national agricultural research institutions have prioritized the development of high- performing triticale varieties. [2, 5] Because of long-term efforts by the Institute of Genetics, Physiology and Plant Protection of the Republic of Moldova, was created a diverse collection of autochthonous triticale varieties, which has been developed to improve agronomic performance under Southeastern European conditions. Notable examples include Ingen 93 (1998), widely used as a reference standard in local research, as well as Ingen 33, Ingen 35, Ingen 40 (2015), Ingen 54 (2020), and Costel (2022) [2]. The existence of such a diverse gene pool within a relatively limited geographic area underscores the necessity for precise identification and differentiation of genetic variability among these cultivars. Genetic diversity analysis plays a fundamental role in crop breeding, conservation, and genotype selection. Various marker systems have been employed to evaluate genetic polymorphisms, including phenotypic, biochemical, cytogenetic, and molecular markers. Among these, molecular markers are considered the most reliable due to their high reproducibility, independence from environmental factors, and the potential for automation and scalability. Techniques such as RFLP, SSR, RAPD, AFLP, SNP, and iPBS have been used to assess genetic variation in cereal crops [6]. The iPBS (inter-Primer Binding Site) marker system, based on the conserved regions of long terminal repeat (LTR) retrotransposons, offers several advantages, including high sensitivity to polymorphisms, especially among closely related genotypes. This makes it a powerful tool for cultivar identification and genetic diversity studies in breeding programs [7]. Current work focuses on using iPBS markers for triticale genotyping as well as assessing molecular diversity within populations with different genetic background. MATERIALS AND METHODS Plant material. Two triticale genotypes – Ingen-54 and Ingen-93 were used in this study. The Laboratory of genetic resources of Institute of Genetics, Physiology and Plant Protection provided plant material. Seeds of 2022 yield were germinated on Petri dish and filter paper under 27°C during 7 days. Eight seedling of each genotype were selected for population analysis. Approximately 2 cm of apical meristem were cut off and used for DNA extraction. DNA extraction. DNA extraction was performed using the ISO 21571 method [8]. A sample of extracted DNA was subjected to a series of dilutions (1:4, 1:14). From the extracted samples, we randomly selected 16 genotypes, 8 genotypes Session I. General and molecular genetics 36 each of the Ingen-54 and Ingen-93 varieties, for PCR testing. Purified and diluted DNA was subsequently used for PCR tests. Primers. Several primers (Pb13, Pb20, and Pb21) designed to universal iPBS markers and described in (Kalendar R. et al., 2010) were used for genotyping and population analysis (Tab.1). Table 1. Primers used for qPCR analysis Primers Sequence Tm (°C) Pb13 CCATTGGGTCCA 45.7 Pb20 GGCTCAGATGCCA 50.5 Pb21 GCTCATCATGCCA 47.6 Amplification. PCR was performed in 25 μl of mix containing: Commercial buffer x10 (DreamTaq Buffer) – 2.5 μl, primer (forward + reverse) – 4.5 μl, dNTP mix – 0.5 μl, Taq polymerase – 0.3 μl, diluted DNA template – 10 μl, water – up to 25 μl volume. The amplification protocol was set up as follows: initial denaturation: 95°C, 3 minutes, followed by 45 cycles of denaturation at 95°C, 30 seconds, primer annealing at 55°C, 30 seconds and elongation at 72°C, 1 minute. Amplification concluded with final elongation: 72°C, 7 minutes. Product analysis. PCR results was visualized on a 1.5% agarose gel stained with ethidium bromide. Molecular weight of the amplicons was assessed using 100 bp DNA Ladder (Thermo Fisher Scientific). Data processing was carried out using the following software: GelAnalyzer, MS Excel, POPGENE. RESULTS AND DISCUSSIONS Total DNA quality test via agarose gel electrophoresis showed crisp and intense bands of high molecular weight without smears and low RNA content (Fig.1). This presumes high yield of integral DNA suitable for molecular genotyping. Figure 1. Electrophoregram of total triticale DNA The amplification profiles obtained using three iPBS primers (Pb13, Pb20, and Pb21) are shown in Figures 2–4. The banding patterns reveal the presence of polymorphic loci and demonstrate the capacity of the chosen primers to Session I. General and molecular genetics 37 discriminate between triticale genotypes. Figure 2. Electrophoregram of PCR products amplified with iPBS primer Pb13. 1-8 – Ingen-54; 9-16 – Ingen-93; M – 100 Plus Ladder (Thermo Fisher Scientific) Figure 3. Electrophoregram of PCR products amplified with iPBS primer Pb20. 1-8 – Ingen-54; 9-16 – Ingen-93; M – 100 Plus Ladder (Thermo Fisher Scientific) Figure 4. Electrophoregram of PCR products amplified with iPBS primer Pb21. 1-8 – Ingen-54; 9-16 – Ingen-93; M – 100 Plus Ladder (Thermo Fisher Scientific) Amplification with these primers produced a total of 33 clear and reproducible loci (attested in three amplifications), ranging in size from approximately 243 to 2912 bp. Session I. General and molecular genetics 38 According to POPGENE statistical analysis, all 33 loci were polymorphic (100%), indicating the high informativeness of the chosen primer set. Nei’s gene diversity (h) ranged from 0.0625 to 0.4188 with a mean of 0.1290, while Shannon’s Information Index (I) varied from 0.1426 to 0.6096 (mean = 0.2403). The highest polymorphism was observed at locus 257 (h = 0.4188), suggesting it as a key marker for genotypic discrimination, which was detected when amplifying with Pb20 primer. Nei’s genetic distance between the two triticale varieties was low (0.0074– 0.0169), with genetic identity ranging from 0.9832 to 0.9926, indicating a relatively close genetic relationship. Gst values (genetic differentiation) averaged 0.0642, and gene flow (Nm) was high (mean Nm = 7.29), reflecting limited genetic subdivision. Primers Pb20 and Pb21 generated the greatest number of polymorphic loci and showed higher values of gene diversity (h), making them more suitable for distinguishing closely related triticale genotypes. Consequently, these specific primers are highly recommended as potent tools for prospective genotyping and breeding programs. CONCLUSIONS iPBS markers proved to be effective for both discriminating between triticale cultivars with different genetic background, as well was evaluating population diversity for this species. It has potential application in triticale breeding programs and supplying national Gene bank information on triticale. PCR amplification using primers Pb13, Pb20, and Pb21 yielded a total of 33 clear and reproducible loci. All 33 loci demonstrated 100% polymorphism, confirming the high informativeness of the selected iPBS primer set. The highest level of polymorphism (h=0.4188) was recorded for locus 257. Nei's genetic distance between the two triticale varieties ('Ingen-54' and 'Ingen-95') is low, while genetic identity is high. Primers Pb20 and Pb21 generated the greatest number of polymorphic loci and exhibited higher gene diversity (h) values compared to Pb13, making them more suitable for distinguishing closely related triticale genotypes. REFERENCES 1. ГАЛИУЛЛИНА, С.А., ШУРЫГИНА, Ю.О., ГАЛИУЛЛИН, А.А. Тритикале – перспективная зерновая культура полифункционального назначения. В: АгроФорум. 2023, vol. 3, pp. 54-55 [online]. [accessed 16.03.2025]. Available: https://cyberleninka.ru/article/n/tritikale-perspektivnaya-zernovaya-kultura- polifunktsionalnogo-naznacheniya. 2. ВЕВЕРИЦЭ, Е., ЛЯТАМБОРГ, С. Селекция тритикале в Молдове. В: Світові рослинні ресурси: стан та перспективи розвитку. Киев, 2019, pp. 75-77. Session I. General and molecular genetics 39 3. CODINĂ, G.G. et al. Physicochemical Properties, Polyphenol and Mineral Composition of Different Triticale Varieties Cultivated in the Republic of Moldova. In: Molecules. 2025, vol. 30, nr. 1233. ISSN: 1420-3049 4. GHENDOV-MOSANU, A. et al. Effect of Brewers’ Spent Grain Addition to a Fermented Form on Dough Rheological Properties from Different Triticale Flour Cultivars. In: Foods. 2025, vol. 14, nr. 41. ISSN 2304-8158 5. GHENDOV-MOSANU, A. et al. Breadmaking Quality Parameters of Different Varieties of Triticale Cultivars. In: Foods. 2024, vol. 13, nr. 1671. ISSN 2304-8158 6. КАНУКОВА, К.Р и др. ДНК-маркеры в растениеводстве. В: Известия Кабардино- Балкарского научного центра РАН. 2019, vol. 6, pp. 220-232. ISSN: 1991-6639 7. KALENDAR, R. et al. iPBS: a universal method for DNA Fingerprinting and retrotransposon isolation. In: Theoretical and Applied Genetics. 2010, vol. 121, pp. 1419–1430. ISSN: 0040- 5752 8. ISO 21571:2005. Foodstuffs — Methods of analysis for the detection of genetically modified organisms and derived products — Nucleic acid extraction. International Organization for Standardization, 2015. 50 p. ISBN 2831886376 Session I. General and molecular genetics 40 DOI: https://doi.org/10.53040/cga12.05 UDC: 632.937:632.38:633.35(478) PARASITIC NEMATOFAUNA IN PEA CROPS (Pisum sativum L.) UNDER THE IMPACT OF THE UNSTABLE ENVIRONMENTAL CONDITIONS OF THE REPUBLIC OF MOLDOVA Elena IURCU-STRĂISTARU, ORCID: 0000-0003-3499 0084 iurcuelena@mail.ru Alexei BIVOL, ORCID: 0009-0003-5709-7173 Ștefan RUSU, ORCID: 0000-0002-3204-5436 Viorelia RUSU, ORCID: 0009-0006-2400-7030 Institute of Zoology of Moldova State University, Parasitology and Helmintology Laboratory, Chisinau, Republic of Moldova Abstract. The results of the present research estimated the efficiency of the phytosanitary helmintological control in peas grown in open field and elucidated the helmintological parasitic impact, establishing the range of the invasive nematofauna, the frequency and the abundance of the associations. As a result of the phytosanitary research and the analysis of the helmintological impact on pea plants,it was found that the parasitic nematode complexes consisted of 14 species included in 4 families: Aphelenchidae, Hoplolaimidae, Tylenchidae, Heteroderidae, of the order Tylenchida, class Nematoda, distributed according to the investigated areas and classified according to the spectrum of trophic specialization in 5 groups. A larger number of species was detected in the Center area 15 species from 8 genera. It was found that the values varied from 7 to 30%, the damage being caused mainly by invasive associations of parasite nematodes of the genera: Heterodera, Meloidogyne, Ditylenchus, Pratylenchus, Helicotylenchus, Aphelenchus. The results of the phytosanitary monitoring contributed to elucidating the degree of nematological damage and brought new evidence in favor of applying sustainable pest control measures suitable for fabaceae croops. Keywords: helmintological control, nematodes, parasitic impact, pea sectors, trophic specialization. INTRODUCTION Cultivating peas for grains, seed material and fodder solves three strategic problems: increasing seed production, increasing vegetable protein production and improving soil fertility, being 2-3 times richer in vegetable protein and having the advantage that it can be sown and harvested extra early [1, 2, 3, 4]. They are sown as a first priority under the environmental conditions of the Republic of Moldova, when the soil has sufficient moisture and allows sowing, usually in early March, but delayed sowing leads to significant production losses. Pea plantations for grain and fodder represent 8% of the arable land in the Republic of Moldova. They are Session I. General and molecular genetics 41 annually invaded by over 40 harmful species, which are also associated with invasive nematode complexes that trigger helminthiasis, which motivates specific annual monitoring to control the populations and their parasitic impact in the cultivation process of this crop [4, 5, 6]. Annually, according to the new institutional project 2024-2027, we conduct nematological research, including biodiversity estimates, morpho-ecological assessments and analyses of the structure of complexes of parasitic fauna, affecting various field crops, including peas [5, 6, 7, 8]. In the Republic of Moldova, helminthological research and monitoring of the biodiversity and structure of parasitic nematode complexes, detected in the agrocenoses of field crops cultivated in state-owned and private enterprises, were initiated and conducted by specialists in phytohelminthology and parasitology, renowned at the national level and recognized internationally, such as: Petru Nesterov, (1979-1980); Nicolai Ocopnnîi, (1982-1999); Melnic Maria et al., (1970- 2024); Poiras Larisa et al., (1996-2016) [4, 5, 6, 7, 9]. Currently, taking into account the specifics of areas with contrasting and unstable climates, we aim at investigating invasive nematode complexes from the families Heteroderidae Hoplolaimidae, Tylenchidae, affecting pea plants (Pisum sativum L), in the context of applying new modern cultivation technologies, comparatively by areas, production associations, plantations and purpose of production. Based on current events and the estimated purpose, the goal of our research has been to establish the structure and diversity of invasive helminth species from the families: Heteroderidae, Hoplolaimidae and Tylenchidae, associated with parasitic forms affecting pea crops (Pisum sativum L.), determining the parasitic impact by comparative analyses of frequency and abundance indices, in various production and experimental sectors, in the dynamics of phenological stages [5, 6, 7, 8]. MATERIALS AND METHODS The proposed program included specific, helminthological investigations in field crop plantations (legumes, winter cereals, technical crops), where samples of soil and plants affected by helminths were taken and surveys and periodic records were made comparatively across various investigated sectors, mainly in the Central area as compared with the Northern area. To establish the areas affected by helminthiasis, over 6 field trips, in 10 localities, 12 sectors, from 4 administrative districts in the Central area, were undertaken, where over 200 samples of soil and diseased plant organs were collected and analyzed (Fig. 1). Subsequently, indices of parasitic impact were determined, such as: species diversity, population density, frequency of attack (F%), intensity of attack (I%), estimation scales were used following phytosanitary control, field and laboratory records were made. Session I. General and molecular genetics 42 A B C Figure 1. A, B, C. Surveys with soil sampling and analysis of plants collected during the growing season (Criuleni district, March-June 2024-2025) A B C Figure 2. A, B, C. Logistics and helminthological extraction analyses and microscopic establishment of morpho-taxonomic indices of nematodes extracted from analyzed samples. The collected samples were subsequently analysed in the laboratory, according to the classical and new methods, with some modifications depending on the specifics of the nematode genera. Through specific techniques, nematodes were extracted from the soil and affected organs, applying the classic flotation-decantation method (Baermann funnel), with some methodological adjustments, with subsequent fixation in 4% formalin, at temperature of 60°C, for morpho-taxonomic studies, reflected in figure 2. The fixed material was analysed under a microscope, establishing the population size, trophic groups, as well as other parasitic indices in the investigated soil and plants. Methodological and logistical support was offered by the Parasitology and Helminthology Laboratory, with methodology according to; Парамонов А. А. 1970; Nesterov P.I. 1978-1988; Melnic M., Erhan D., Rusu Ș2014.; Poiras L. et al., 2016, Iurcu-Straistaru E. at al. 2018-2023. The taxonomic units were determined using nematological identification guidelines according to: Decker Н. 1972, Baldwin, J.G, Nadler, S.A., Adams, B.J. 2004; Perry, R.N., Moens, M.M. 2006 Decramer, W., & Hunt D. J. 2006; Siddiqi, M.R. 2010; Sasanelli, N. et al. 2018 [3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14]. Session I. General and molecular genetics 43 RESULTS AND DISCUSSIONS Biological control of pea plantations affected by helminths was verified in various private households and production associations in the Center of the Republic of Moldova and in the administrative districts of Criuleni, Anenii Noi, Nisporeni, located in the South-East and South-West, where winter cereals, corn, sunflower, fodder crops, etc. are also intensively cultivated. Peas are cultivated on large and significant areas, in the sectors of the Central area, but in the North the areas are limited, instead, soybean plantations for grains and fodder predominate. For this reason, our research was focused mainly on sectors of the Central area, making detailed surveys and analyses. Phytosanitary surveys were conducted monthly, in the period March-June, 2024-2025, which were characterized by relatively warm and humid weather conditions, which had an advantageous influence on the development of plants including but also of parasitic nematode complexes. The average frequency and intensity of helminthic attack were established, reaching values up to 25-30%. Abundant precipitation and higher temperatures have determined the severity of helminthiasis in association with various disastrous diseases such as: ascochytosis, fusariosis, which appeared early on the roots and stems during the vegetative phase. Simultaneously, on the preventively marked samples the presence of helminthic diseases was detected, the values of intensity and frequency of attack were analyzed comparatively, and the structure and predominance of certain genera and species with specific pathogenic invasive impact were highlighted. In all the investigated sectors, helminthic diseases and infestations were reported through local outbreaks, with retarded plants, in association with wilted, partially necrotic ones in variegated colors, presented in Figure 3. The periods with abundant rainfall and unstable daytime temperatures, in the months of April-June, caused an increase in the damage severity and in the numerical density of the pest populations, reaching average values in the Central area 50-250 individuals/100 gr./soil. It was found that, in all the investigated sectors, there were extensive nematode complexes that produce helminthiasis, in certain sectors affecting the plants severely. This is due to the adaptive resistance capacities in the soil. Meanwhile, nematodes form specialized associations in these crops, caused by non-compliance with maintenance techniques. Such conditions favored the increase in the degree of parasitic helminthological impact, affecting plants in the stages of leaf formation, until the formation of pods. Session I. General and molecular genetics 44 A B C Figure 3. A, B, C. Plants affected by mixed helminthic diseases associated with downy mildew and powdery mildew, on all plant organs, in the pod formation phase; A-productive sector May-June, Criuleni district, 2025 During this period, the reported helminth symptoms remain clearly accentuated and extensive, the visual symptoms including chlorosis, a reduced number of mature leaves, dwarf leaves, roots seriously affected by necrosis and rot caused by helminths. The species of pathogenic fungi in the soil estimated in ascending comparative values, by phenological stages, vary from 5% in the stage of 2-5 leaves up to 30% in the fruit development stage, presented in table 1. Table 1. The estimation of phytosanitary indices by comparative values of helminthic parasitic impact in various phenological stages of pea plants grown in the districts of the Central area, (March-June, 2025). Central area and pilot districts 26 March, stage of 2-5 developed leaves 25 April, stage of 10-15 developed leaves 27 May, flowering stage 26 June, fruit development – ripening stage D. (100 g soil) F.(%) I.(%) D. (100 g soil) F.(%) I.(%) D. (100g soil) F.(%) I.(%) D. (100 g soil) F.(%) I.(%) Nisporeni, Vărzărești village 5-17 7-10 5-7 17- 23 9-12 7-10 25-35 12-15 8-11 180 25-28 22-25 Criuleni district, Pașcani village 7-15 5-9 3-5 14- 18 8-9 6-8 20-28 10-13 9-12 250 27-30 24-27 Criuleni, Bălăbănești village 8-16 8-12 5-7 10- 15 7-10 6-9 28-33 13-18 7-10 220 25-28 22-25 Anenii Noi, Mereni village 6-14 5-8 2-5 13- 16 10-14 8-12 23-30 9-14 7-9 190 20-25 18-22 Legend: D. – density of nematode per 100 grams of soil; F (%) – frequency of attack; I (%) – intensity of attack. Session I. General and molecular genetics 45 A B C Figure 4. A, B, C. Pea plantations in th