Trifolium Medium Research Articles

Inoculation with Arbuscular Mycorrhizal Fungi Supports the Uptake of Macronutrients and Promotes the Growth of Festuca ovina L. and Trifolium medium L., a Candidate Species for Green Urban Infrastructure.

Green roofs and walls play an important role in promoting biodiversity, reducing the urban heat island effect and providing ecosystem services in urban areas. However, the conditions on green walls/roofs (low nutrient and organic matter content, drought, high temperatures) are often unfavorable for plant growth. Arbuscular mycorrhizal fungi (AMF) can improve the growth and development of plants under stress conditions as they can increase nutrient and water uptake. In a 6-month pot experiment, we investigated the effect of AMF inoculation on the growth and NPK uptake of Festuca ovina L. and Trifolium medium L., which are used for green roofs and walls. Two variants of mycorrhizal inoculation were used in the experiment: a commercial mycorrhizal inoculant AM Symbivit (Symbiom Ltd., Lanskroun, Czech Republic) and a mycorrhizal inoculant collected from calcareous grassland in the Silesia region (Poland). Funneliformis mosseae was the most abundant species in the roots of F. ovina and T. medium with IM inoculum. In the CM variant, a dominance of F. mosseae was observed in the roots of F. ovina. In contrast, Archaeosporaceae sp. node 317 dominated in the roots of T. medium. Both inoculations had a positive effect on the increase in dry weight of the shoots of T. medium, but only the commercial inoculum had a positive effect on the growth of F. ovina. Both inoculations improved the P uptake by the roots and the P and K uptake into the shoots of T. medium. In addition, both inoculations improved the K uptake by the roots of F. ovina and the N, P and K uptake into the shoots. In conclusion, both AMF communities included in the inoculations had a positive effect on plant growth and nutrient uptake, but the effect depends on the plant and the mycorrhizal fungus species.

Ecological and floristic features of modern plant groupings on the territory of the Kipchak archaeological complex

The article presents the results of the study of the peculiarities of the existing vegetation cover on the territory of the Kipchak archaeological complex (Republic of Bashkortostan). The data of ecological and botanical description are presented, which contribute to the interpretation of the natural state of vegetation groupings on the territory where archaeological surveys are conducted. The investigated burial ground and settlement are part of the objects of the Kipchak archaeological complex and are located near the village of Kipchakovo. The article includes a list of vascular plants that were discovered and described in this area during the survey in 2022, indicating their relevance to the trial areas of the Kipchak archaeological complex. According to the survey data the most widespread species of vascular plants were identified, they include: Vicia tenuifolia Roth, Fragaria vesca L., Trifolium alpestre L., Galium verum L., Achillea millefolium L., Achillea millefolium L., Dactylis glomerata L., mountain clover Trifolium montanum L., Trifolium medium L., Galium boreale L., Elytrigia repens (L.) Nevski. To date, the study area is assessed as slightly disturbed.

The Effect of Combined Drought and Temperature Stress on the Physiological Status of Calcareous Grassland Species as Potential Candidates for Urban Green Infrastructure.

Nature-based solutions are promising for climate adaptation and environmental management in urban areas, but urban conditions are stressful for vegetation. In particular, the interaction of drought and high temperatures may be detrimental. Guiding plant selection for urban greening with native species requires a far better knowledge of plant adaptations and stress acclimation. We tested the physiological responses of four candidate calcareous grassland species for green roofs and walls to the combined effects of drought and high temperatures under controlled conditions. The tested species proved relatively resistant to stress despite different strategies to protect the photosynthetic apparatus, maintain water balance, and repair damages. Based on the physiological responses, we rank the species in descending order of resistance to the stress factors tested: Trifolium medium > Festuca ovina > Carex flacca > Potentilla reptans, but all four can serve as potential candidates for green walls and roofs. Physiological stress screening of plant species for use on green roofs and walls supplements the habitat template approach to provide a stronger and wider base for prioritizations.

Investigating the Origin and Evolution of Polyploid Trifolium medium L. Karyotype by Comparative Cytogenomic Methods.

Trifolium medium L. is a wild polyploid relative of the agriculturally important red clover that possesses traits promising for breeding purposes. To date, T. medium also remains the only clover species with which agriculturally important red clover has successfully been hybridized. Even though allopolyploid origin has previously been suggested, little has in fact been known about the T. medium karyotype and its origin. We researched T. medium and related karyotypes using comparative cytogenomic methods, such as fluorescent in situ hybridization (FISH) and RepeatExplorer cluster analysis. The results indicate an exceptional karyotype diversity regarding numbers and mutual positions of 5S and 26S rDNA loci and centromeric repeats in populations of T. medium ecotypes and varieties. The observed variability among T. medium ecotypes and varieties suggests current karyotype instability that can be attributed to ever-ongoing battle between satellite DNA together with genomic changes and rearrangements enhanced by post-hybridization events. Comparative cytogenomic analyses of a T. medium hexaploid variety and diploid relatives revealed stable karyotypes with a possible case of chromosomal rearrangement. Moreover, the results provided evidence of T. medium having autopolyploid origin.

A de novo assembled high-quality chromosome-scale Trifolium pratense genome and fine-scale phylogenetic analysis

BackgroundRed clover (Trifolium pratense L.) is a diploid perennial temperate legume with 14 chromosomes (2n = 14) native to Europe and West Asia, with high nutritional and economic value. It is a very important forage grass and is widely grown in marine climates, such as the United States and Sweden. Genetic research and molecular breeding are limited by the lack of high-quality reference genomes. In this study, we used Illumina, PacBio HiFi, and Hi-C to obtain a high-quality chromosome-scale red clover genome and used genome annotation results to analyze evolutionary relationships among related species.ResultsThe red clover genome obtained by PacBio HiFi assembly sequencing was 423 M. The assembly quality was the highest among legume genome assemblies published to date. The contig N50 was 13 Mb, scaffold N50 was 55 Mb, and BUSCO completeness was 97.9%, accounting for 92.8% of the predicted genome. Genome annotation revealed 44,588 gene models with high confidence and 52.81% repetitive elements in red clover genome. Based on a comparison of genome annotation results, red clover was closely related to Trifolium medium and distantly related to Glycine max, Vigna radiata, Medicago truncatula, and Cicer arietinum among legumes. Analyses of gene family expansions and contractions and forward gene selection revealed gene families and genes related to environmental stress resistance and energy metabolism.ConclusionsWe report a high-quality de novo genome assembly for the red clover at the chromosome level, with a substantial improvement in assembly quality over those of previously published red clover genomes. These annotated gene models can provide an important resource for molecular genetic breeding and legume evolution studies. Furthermore, we analyzed the evolutionary relationships among red clover and closely related species, providing a basis for evolutionary studies of clover leaf and legumes, genomics analyses of forage grass, the improvement of agronomic traits.

In Vitro Evaluation of the Antioxidant Activity and Chemopreventive Potential in Human Breast Cancer Cell Lines of the Standardized Extract Obtained from the Aerial Parts of Zigzag Clover (Trifolium medium L.).

The aboveground parts of Trifolium medium L. (zigzag clover), a little-known representative of the family Fabaceae, collected during flowering in a wild stand (Sławin-Szerokie district, Lublin, Poland), were used in this study. Our previous investigations confirmed the higher content of phytoestrogenic isoflavones (especially biochanin A and formononetin derivatives) in T. medium compared to the closely related medicinal plant T. pratense (red clover) and the involvement of these compounds in anti-osteoporotic effects in ovariectomized female rats. The current study focused on evaluating other antibiodegenerative (antioxidant, chemopreventive, and cytostatic) effects for the lyophilisate (TML) obtained from wild zigzag clover. For this purpose, efficient ultrasound-assisted extraction (UAE) was employed, followed by vacuum drying and phytochemical standardization using a newly developed reversed-phase high-performance liquid chromatography (RP-LC) coupled with a PDA detection. Malonylglycosides of biochanin A and formononetin were the predominant compounds and were found to contribute more than 54% to the total isoflavone content determined in the standardized extract of zigzag clover. The antioxidant potential of TML was examined in vitro using the Folin–Ciocalteu and cupric ion-reducing (CUPRAC) methods in addition to the free radical (DPPH• and ABTS•+) scavenging assays. The cytotoxic effects of TML, formononetin, and ononin were evaluated on MCF-7 (estrogen-dependent) and MDA-MB-231 (estrogen-independent) human breast cancer cell lines using the MTT assay. The important role of malonyl isoflavone derivatives has been indicated both in chemoprevention and potential cytotoxic effects of TML against certain types of breast cancer.

CONTENT OF POTASSIUM AND CALCIUM IN PLANTS OF THE MIDDLE MOUNTAIN DAGESTAN

The results of a study on the content of potassium and calcium in plants of the Middle Mountain province of Dagestan are presented. The content of macroelements was determined on «Savant AA Σ» atomic absorption spectrometer. The K content in plants varied from 0.16 to 3.21%, Ca - from 0.02 to 3.47%. The content of mobile forms of elements on the northern slope is slightly higher than on the southern (1.2 times). The coefficient of biogeochemical mobility K ranges from 5 to 100.3, Bx Ca – from 1.2 to 204.1. On the southern slope, K and Ca are most actively absorbed from the herb-cereal phytocenoses Plantago media, Plantago lanceolata, Lotus corniculatus, Trifolium medium, Inula britannica, Achillea millefolium, Psephellus daghestanicus, Galium verum, Astragalus onobrychioides, Medicago glutinosa. On the northern slope, K, Ca Astrantia biebersteinii, Inula germanica, Trifolium campestre, Vicia truncatula, Vicia cracca, Betonica macrantha, Origanum vulgare, and Betonica orientalis accumulate most actively. The least accumulation of K and Ca is observed in plants of the Poaceae family. The difference in the content of K and Ca in plants depends on the content of mobile forms of elements in the soil, biological characteristics of plants, and environmental factors.

Legumes in natural post-fire successions of forest meadows and pastures in Northern Bulgaria

We have witnessed a drastic increase in global fires in recent years that are strongly linked to climate change. On the other hand, the prescribed fires are applied in agricultural practice; however, their effectiveness is controversial. It is important to study the effects of both types of fires – wild and prescribed to predict their impact on biota. In this study, four different locations in Central Norther Bulgaria that recovered after the fire were studied concerning the biodiversity and abundance of the first colonizer species. Two generations from the locations were studied and demonstrated that the legume plants are dominant. The abundant species and their morphological and physiological characteristics are described. The annual species like Bituminaria bituminosa, Vicia striata, and Medicago minima propagated with seeds and perennial rhizome Lathyrus pratensis and Trifolium medium show the best potential to resprout after the fire events. Presented results will help responsible authorities of our county to take adequate actions for burned areas recovery. The directions for future studies are highlighted. URL: https://www.upjs.sk/pracoviska/botanicka-zahrada/odborne-aktivity/contents-abstracts/

In-depth phytochemical and biological studies on potential AChE inhibitors in red and zigzag clover dry extracts using reversed–phase liquid chromatography (RP-LC) coupled with photodiode array (PDA) and electron spray ionization-quadrupole/time of flight-mass spectrometric (ESI-QToF/MS-MS) detection and thin-layer chromatography-bioautography

In the study, Trifolium medium L. and T. pratense L. were used to obtain lyophilisates which were named as TML and TPL, respectively. The former clover taxon represents a little explored species, while the latter one is often found in dietary supplements and functional foods due to the content of isoflavones that alleviate various menopausal symptoms. Detailed phytochemical profiles of both lyophilisates were examined and compared using coupled chromatographic (RP-LC) and spectroscopic PDA/ESI-QToF/MS-MS methods. A total of 54 and 55 compounds were identified in TML and TPL, respectively. Close chemotaxonomic similarities were confirmed for both clover taxa examined, especially in terms of the most abundant isoflavones and hydroxycinnamates. For the first time, neurotropic polyphenols (AChE inhibitors) were identified in clover extracts using TLC-bioautography. In this group, caffeoylmalic acid exhibited significant AChE inhibitory effect confirming the potential of TPL and TML to support physiological functions of the endocrine and nervous systems.

Sowing Mixtures of Native Plant Species: Are There Any Differences between Hydroseeding and Regular Seeding?

Hydroseeding is a convenient, low-cost way to plant seeds. Traditionally, fast-growing commercial species that are cheap to obtain are preferred in hydroseeding, while native species have limited use. Nowadays, the use of native species is often desired in revegetation projects. However, there is a paucity of information about hydroseeding native species in Northern areas of Europe. Therefore, we aimed to determine whether hydroseeding has any effects on native plant cover formation, species richness and abundance, the development of plant morphological features, or aboveground biomass. A total of 40 native plant species in Lithuania were sowed using hydroseeding and regular seeding. The experimental plots were assessed for two years. The results show a relatively small and short positive effect of hydroseeding on plant cover formation. No significant differences were found in species richness between the sowing treatments. However, a comparison of species composition revealed significant differences between the sowing treatments that were more associated with species abundance than species diversity. Hydroseeding was favoured by legume species, such as Onobrychis viciifolia, Ononis arvensis, Lotus corniculatus, and Trifolium medium, while Festuca rubra favoured the regular seeding treatment. Overall, our findings emphasize that legume species that display more competitive growth traits should be included in the seed mixture in lower proportions when hydroseeding is applied.

Resources of birch trees and their productivity for honey collection in conditions of the Leningrad region

The Leningrad Oblast has a rich food base for bees on forest land. Still, currently, there is no regulatory basis for assessing the honey resources of birch forest land at the regional level. Therefore, forest areas with woody, shrubby or herbaceous plants as part of the woody, shrubby or herbaceous layer are used as a forage base for bees. The birch forests of the Leningrad region are the second largest in terms of the area covered. The research aims to carry out an inventory of honey resources and determine honey productivity for the primary honey plants under the birch canopy and forest edges. We used a proven survey methodology for counting plants of the understorey at the experimental sites. The study presented 71 species from 31 families of melliferous plants of blueberry, wormwood and herb-meadow birch forests. Significant melliferous plants under the birch canopy are Aegopodium podagraria L. (herb-twine and sagebrush birch, 57 kilograms/hectare); Vaccinium myrtillus L. (birch bilberry and acidic birch, 27 kilograms/hectare); Veronica chamaedrys L. (acidic birch, 24 kilograms/hectare). Rubus idaeus L. (107 kg/ha) has maximum honey production in the forest margins. The honey yield of Frangula Alnus Mill. and Chamaenerion angustifolium (L.) Scop is 50-60 kg/ha; the honey yield of Trifolium medium L. is 32 kg/ha. The evaluation of the birch woodlands showed that the studied components of the forest phytocenosis are characterized by a significant number of honey-bearing plant species that have a wide range of honey productivity.

К синтаксономии сегетальной растительности Среднего Урала

The decrease in the crop area and changes in their ratio have led to changes in weed vegetation in the Middle Urals, where its cenoflora was studied relatively well (Tretyakova, 2006; Tretyakova, Kondratkov, 2018; Kondratkov, Tretyakova, 2018; 2019 a, b), while community diversity so far has not yet been assessed. The first results of the ecological and floristic classification of basic crop communities (wheat, barley, oats, corn, potatoes, peas, rapeseed etc.) in six botanical and geographical regions of the taiga and forest-steppe zones (Kulikov et al., 2013) within the Sverdlovsk region, based on the analysis of 160 relevés performed by A. S. Tretyakova, P. V. Kondratkov and N. Yu. Grudanov in 2019, are presented. Communities are classified within the order Aperetalia spicae-venti J. Tx. et Tx. in Malato-Beliz et al. 1960 of the class Papaveretea rhoeadis S. Brullo et al. 2001 nom. conserv. propos. and assigned to the alliance Scleranthion annui (Kruseman et Vlieger 1939 nom. conserv. propos.) Sissingh in Westhoff et al. 1946), which represents the most mesophytic types of weed vegetation in the Urals. Three associations allocated in alliance (two for the first time): Solano nigri–Erodietum cicutarii ass. nov. hoc loco (Table 2, syntaxon 7; Table 4); nomenclature type (holotypus hoc loco) — Table 4, rel. 11 (number in database — 25): Sverdlovsk Region, Sysertskiy district, Kunarskoe village environs, 23.07.2019, authors — A. S. Tretyakova, P. V. Kondratkov);Lamio amplexicaulis–Stellarietum mediae ass. nov. hoc loco (Table 2, syntaxon 8; table 5; nomenclature type (holotypus hoc loco) — Table 5, rel. 10 (number in database — 27): Sverdlovsk Region, Sysertskiy district, Shaidurovo village environs, 22.07.2019, authors — A. S. Tretyakova, P. V. Kondratkov); ass. Euphorbio helioscopiae–Fumarietum officinalis Khasanova et al. 2018. The last one includes two variants. Two unranked communities are also described. Distribution areas of syntaxa cover the southern taiga and pre-forest-steppe pine-birch forests subzones of the taiga zone and the northern forest-steppe subzone of the forest-steppe zone. Habitats are confined mainly to gray forest soils, in rare cases — leached chernozems. The mostly wide distributed is the ass. Euphorbio helioscopiae-Fumarietum officinalis, also common to the south of the study area, in the Republic of Bashkortostan. Аss. Lamio amplexicaulis–Stellarietum mediae is identified in five botanical and geographical districts within the Sverdlovsk region. The confluence with crops and the accompanying system of agriculturе is different. The species composition of the established syntaxa was compared with the associations of the alliance Scleranthion annui described earlier in the Republic of Bashkortostan. The species richness of the latter is generally higher. Differences in species composition of diagnostic groups were revealed. There is decreasing of constancy of Tripleurospermum inodorum, Centaurea cyanus, Polygonum aviculare, Raphanus raphanistrum in the cenoflora of the first one, and both diagnostic species of the more xerophytic alliance Caucalidion lappulae von Rochow 1951 (Galeopsis bifida, Persicaria lapathifolia, Silene noctiflora, Galeopsis ladanum, Melilotus officinalis),apophytes of meadow pasture and forest edges (Medicago lupulina, Trifolium hybridum, Trifolium medium, Achillea millefolium,etc.), xeromesiphytic species (Avena fatua, Amaranthus retroflexus, Setaria viridis, Euphorbia virgata). The latter is probably associated with significant reduction in the field area of some crops (sunflower, flax, etc.). The differences in species composition and ecology of the compared syntaxa are illustrated by the results of ordination analysis. Weed communities of the Southern Urals are localized on the left side of the DCA-ordination diagram, these of the Middle Urals — on the right one. There are changes from the most xerophytic communities of the ass. Linario vulgari-Lactucetum serriolae to the most mesophytic ones of associations Lamio amplexicaulis–Stellarietum mediae and Euphorbio helioscopiae–Fumarietum officialalinae along the first axis. The second axis reveals the change in species composition under the influence of the agronomic factor — culture and the corresponding agricultural system and communities of winter cereals (associations Consolido regali–Centaureetum cyanae and Galeopsetum bifidae) are replaced by communities of spring cereals and row crops (corn, potatoes, peas, rapeseed), which are assigned to the associations Linario vulgari–Lactucetum serriolae and Solanо nigri–Erodietum cicutarii.

Communities of broadleaved woodlands of the lower part of the forest belt of the Belaya river basin (North-Western Caucasus)

The study of phytocoenotic diversity of oak and oak-hornbeam forests of the lower part of the forest belt in the Belaya river basin (Western Caucasus) was performed using the Braun-Blanquet approach. Five floristically different groups of releves were identified based on quantitative classification (Ward's method, Jaccard Distance Measure). An association rank was established for each group. According to the results of comparative syntaxonomic analysis, three associations of moderately masic oak and oak-hornbeam forests - Geranio sanguinei-Quercetum petraeae ass. nova hoc loco (diagnostic species: Pinus sylvestris, Calamagrostis arundinacea, Rhododendron luteum, Geranium sanguineum, Vaccinium arctostaphylos), Inulo salicinae-Quercetum petraeae ass. nova hoc loco (diagnostic species: Brachypodium pinnatum, Inula salicina, Dorycnium graecum, Laser trilobum, Trifolium medium, Viola alba, Stachys officinalis, Pyrethrum poteriifolium, Phleum phleoides, Teucrium chamaedrys, Genista patula) and Acero tatarici-Carpinetum betuli ass. nova hoc loco (diagnostic species: Phalacroloma annuum, Thalictrum minus, Torilis japonica, Prunus divaricata, Physalis alkekengi, Lamium maculatum, Acer tataricum) were included in the alliance Crataego-Carpinion Passarge 1981, order Lathyro-Carpinetalia Passarge 1981, class Carpino-Fagetea Jakucs et Passarge 1968. The correct syntaxonomic position of two associations of mesic hornbeam and oak-hornbeam forests - Dryopterido filicis-maris-Carpinetum betuli ass. nova hoc loco (diagnostic species: Dryopteris filix-mas, Circaea lutetiana, Aegopodium podagraria, Paris incompleta, Abies nordmanniana, Impatiens noli-tangere, Festuca gigantean, Symphytum grandiflorum, Dipsacus pilosus, Aconitum orientale, Asperula caucasica, Moehringia trinervia, Polygonatum orientale) and Vincetoxico scandentis-Carpinetum betuli ass. nova hoc loco (diagnostic species: Viola hirta, Vincetoxicum scandens, Ligustrum vulgare, Lathyrus vernus, Polygonatum multiflorum, Rubus caucasicus, Euphorbia squamosa) has not been determined, however their relations to beech and beech-hornbeam forests of the order Rhododendro-Fagetalia orientalis Passarge 1981 were noted. The described associations of oak-hornbeam forests correspond to the regional ecological series of communities along the moisture gradient.

Pedigree analysis of pre-breeding efforts in Trifolium spp. germplasm in New Zealand

BackgroundPrebreeding in plants is the activity designed to identify useful characteristics from wild germplasm and its integration in breeding programs. Prebreeding aims to introduce new variation into the populations of a species of interest. Pedigree analysis is a valuable tool for evaluation of variation in genebanks where pedigree maps are used to visualize and describe population structure and variation within these populations. Margot Forde Germplasm Centre (MFGC) is New Zealand’s national forage genebank and holds a collection of ~ 75 species of the genus Trifolium, of which only a dozen have been taken through prebreeding programs. The main objective of this study was to construct pedigree maps and analyse patterns of relatedness for seven minor Trifolium species accessions contained at the MFGC. These species are Trifolium ambiguum, Trifolium arvense, Trifolium dubium, Trifolium hybridum, Trifolium medium, Trifolium subterraneum and the Trifolium repens x Trifolium occidentale interspecific hybrids. We present a history of Trifolium spp. prebreeding in New Zealand and inform breeders of possible alternative forage species to use.ResultsPedigree data from accessions introduced between 1950 and 2016 were used and filtered based on breeding activity. Kinship levels among Trifolium spp. remained below 8% and no inbreeding was found. Influential ancestors that contributed largely to populations structure were identified. The Australian cultivar ‘Monaro’ had a strong influence over the whole population of accessions in T. ambiguum. T. subterraneum and T. repens x T. occidentale had the largest number of generations (3). T. ambiguum and T. medium had the highest cumulative kinship across the decades.ConclusionsWe conclude that there are high levels of diversity in the seven Trifolium spp. studied. However, collection and prebreeding efforts must be strengthened to maximize utilization and bring useful genetic variation.

Синтаксономия растительности карьеров на месте сведенных сосняков лишайниковых и зеленомошных (среднетаежная подзона европейского северо-востока России)

Синтаксономия растительности карьеров на месте сведенных сосняков лишайниковых и зеленомошных (среднетаежная подзона европейского северо-востока России)

Development and use of synanthropic phytocenoses with complex invasion in Kaluga region

Currently, less than 33% of arable land is used in Meshchovskoye Opolye. Optimizing technology elements of resource-saving development of synanthropic fallow phytocenoses with complex invasion is relevant. The article is devoted to analysis of development of self-organizing phytocenoses with varying invasion degrees and their use on lands temporarily withdrawn from active agricultural use. Field experiments were carried out on postagrogenic gray forest loamy soils in Kaluga Research Institute of Agriculture in 2006-2018. Influence of mineral fertilizers as an optimization element of technology of fallow development was studied using transects and permanent survey sites on the area of 12.0 ha. The reasons for change in productivity and its determining elements in hayfields in autogenous - allogenic phytocenoses were analysed. We established that economic value of plant communities was determined by potential of constituent species and variability of their productivity in years with various environmental conditions. 12 years later phytocenoses become homogeneous and consist of 10-12 main plant species, determining green mass productivity by 75%. Compared to native species invasive plant species have 1.4-2.0 fold higher productivity which accounts for 60% and more productivity of phytocenoses. Transforming role of Erigeron canadensis L., Lupinus polyphyllus Lindl. and Solidago gigantea Ait. on their expansion into aboriginal herb (share in mowed mass - 40% or more) communities was shown; their high adaptive potential for ecological-soil conditions of Meshchovskoye Opolye (center of Nonchernozem Zone of Russia) was established. Application of mineral fertilizers (P40K90) in the secondary Trifolium medium phytocenoses (more than 40%) contributed to 2.4-fold increase in its productivity (from 1.38 to 3.29 kg/m2) and increased productivity stability by 60%, reducing low-value species in crop structure by 1.6 times. The total area of cenopopulations increased by 10.4-21.5%, yields - from 2.31 to 4.41 kg/m2.

Red Clover (Trifolium pratense) and Zigzag Clover (T. medium) - A Picture of Genomic Similarities and Differences.

The genus clover (Trifolium sp.) is one of the most economically important genera in the Fabaceae family. More than 10 species are grown as manure plants or forage legumes. Red clover’s (T. pratense) genome size is one of the smallest in the Trifolium genus, while many clovers with potential breeding value have much larger genomes. Zigzag clover (T. medium) is closely related to the sequenced red clover; however, its genome is approximately 7.5x larger. Currently, almost nothing is known about the architecture of this large genome and differences between these two clover species. We sequenced the T. medium genome (2n = 8x = 64) with ∼23× coverage and managed to partially assemble 492.7 Mbp of its genomic sequence. A thorough comparison between red clover and zigzag clover sequencing reads resulted in the successful validation of 7 T. pratense- and 45 T. medium-specific repetitive elements. The newly discovered repeats led to the set-up of the first partial T. medium karyotype. Newly discovered red clover and zigzag clover tandem repeats were summarized. The structure of centromere-specific satellite repeat resembling that of T. repens was inferred in T. pratense. Two repeats, TrM300 and TrM378, showed a specific localization into centromeres of a half of all zigzag clover chromosomes; TrM300 on eight chromosomes and TrM378 on 24 chromosomes. A comparison with the red clover draft sequence was also used to mine more than 105,000 simple sequence repeats (SSRs) and 1,170,000 single nucleotide variants (SNVs). The presented data obtained from the sequencing of zigzag clover represent the first glimpse on the genomic sequence of this species. Centromeric repeats indicated its allopolyploid origin and naturally occurring homogenization of the centromeric repeat motif was somehow prevented. Using various repeats, highly uniform 64 chromosomes were separated into eight types of chromosomes. Zigzag clover genome underwent substantial chromosome rearrangements and cannot be counted as a true octoploid. The resulting data, especially the large number of predicted SSRs and SNVs, may have great potential for further research of the legume family and for rapid advancements in clover breeding.

Secondary Metabolites of Micromycetes in Plants of the Family Fabaceae, Genus Trifolium

The component composition and content of mycotoxins in perennial grasses of five species of the genus Trifolium, selected from natural biocenoses in different periods of vegetation, were studied. In the terrestrial parts of plants, the permanent presence of alternariol, cyclopiazonic acid, and emodin was shown. It was noted that differences in contamination are most clearly expressed in meadow clover Trifolium pratense L. and white clover Trifolium repens L., while alsike clover Trifolium hybridum L. occupies an intermediate position. It was found that zigzag clover Trifolium medium L. and mountain clover Trifolium montanum L., morphologically close to meadow clover, practically did not differ in the composition of toxic metabolites. The ways of formation of metabolic equilibrium in plants with participation of microscopic fungi are discussed.

Interspecific hybrids of Trifolium medium L. × 4x T. pratense L. obtained through embryo culture.

Interspecific hybrids of Trifolium medium L. × 4x T. pratense L. obtained through embryo culture.

Impact of interspecific hybridization of T. pratense × T. medium and backcrossing on genetic variability of progeny

Red clover (Trifolium pratense L.) is a high-quality fodder crop which has been hybridized successfully with its wild relative zigzag clover (T. medium L.). The aim of this study was to evaluate the genetic impact of interspecific hybridization and subsequent repeated backcrossing on the variability within hybrid progeny genomes. Nuclear DNA content of 800 and 753 hybrid plants from F7/F8 and F8/F9 generations, respectively, was measured by flow cytometry. Resulting values were converted to estimated chromosome counts, which were successfully validated on a sample of 28 plants by counting mitotic chromosomes. The two generations showed a similar distribution of various chromosome counts ranging from 22 to 47 chromosomes. In total, 24.0% and 34.3% of plants from the two generations had different numbers of chromosomes from their parental plants. Variability within the hybrid population was assessed by fluorescent in situ hybridization using rDNA probes. Individual plants had a pattern of 5S and 45S rDNA loci rather more similar to that of T. pratense than of T. medium. Numbers of chromosomes with clusters of 5S rDNA ranged from 6 to 14 while those of 45S rDNA varied between 4 and 13. Individual arrangements were almost unique, and some plants possessed also novel formations which were not present in any of the parental species, such as a cluster of 5S rDNA surrounded by 45S rDNA clusters or a 45S rDNA cluster surrounded by 5S rDNA clusters. This suggests complex rearrangements connected with post-hybridization stabilization of hybrid genomes.