Spatial and Temporal Assessment of Brassica napus L. Maintaining Genetic Diversity and Gene Flow Potential: An Empirical Evaluation

Unpredicted persistence of all forms of B. napus present in the agro-ecosystem is the most common consequence of preservation and self-recruitment of seeds originating from soil seed bank. In nature, spontaneous intra- and inter-specific hybridization of B. napus is possible with sexually compatible species from the Brassicaceae family. The aim of this chapter is (a) to identify the distribution pattern and population dynamics of volunteers and feral populations along statistical regions in Slovenia; (b) to assess the global diversity of naturally appearing B. napus plants; (c) to evaluate the genetic differentiation between volunteers and feral populations; (d) to obtain the spatial and temporal distribution of spontaneous pollination potential and estimation of gene flow conservation; (e) to find the empirically assigned out-crossing rate of B. napus under a fragmented landscape structure, during 4-year monitoring; and (f) to observe that ecologically, evolutionary, and agronomically oriented studies could be conducted at the DNA level using short sequence repeat (SSR) markers. In total, we collected 261 samples of volunteer and feral populations. Our results showed that alleles from both volunteer and feral populations were distributed in three genetic clusters with relatively similar levels of diversity. Naturally occurring out-crossing rate is 13.71%. The global Mantel correlation coefficient of genetic and spatial relatedness between genotypes is 0.044

Uporaba DNA markerjev za genetsko diferenciacijo navadnega (Avena sativa L.) in golega ovsa (Avena nuda L.)

The usefulness of genetic identification of varieties for seed quality analyses becomes important, when we suspect the presence of another variety, species or even genus, based on morphological seed traits in the purity analysis and germination test, With genetic analyses, the doubt about the authenticity of the naked oats (Avena nuda L.) variety \u27Kamil\u27 was successfully solved. There were atypical seeds with chaff among the samples of this variety, so it was not possible to confirm with certainty, whether these were seeds of the same species/variety or impurities of common oats (Avena sativa L.). Based on the seed phenotype of the \u27Kamil\u27 variety (two independent samples 284 and 285), four sub-samples were prepared (284AN, 284AS, 285AN, 285AS); AN label was the sub-sample with naked seeds and AS label was the sub-sample with chaffed seeds. In addition, another variety (\u27Noni\u27) and four accessions of common oats (ACC378, ACC379, ACC380, ACC381) from the Slovene Plant Gene Bank were included as common oat standards. A total of 36 plant samples of Avena sp. were included in the genetic differentiation. Using a set of six highly informative SSR markers and the results of diversity parameters at individual loci and paired genetic comparisons, we were able to confirm that the analysed sub-samples belonged to the same variety, i.e. \u27Kamil\u27. In addition, despite the morphological similarity, the chaffed seeds of naked oats variety ‘Kamil’ were sufficiently (genetically) different from the analysed variety and/or accessions of common oats.Uporabnost genetske identifikacije sort se kaže pri analizah kakovosti semena, ko se na osnovi morfoloških znakov semena pri analizi čistote in/ali klic v postopku kalivosti pojavi sum na drugo sorto, vrsto ali celo rod. Z aplikativno uporabo genetskih analiz smo uspešno rešili dvom o pristnosti sorte \u27Kamil\u27 vrste golega ovsa (Avena nuda L.). Med semenom omenjene sorte je bilo namreč prisotno tudi seme s plevo, a se ni dalo z gotovostjo potrditi ali gre za seme iste vrste s plevo ali za primes semen navadnega ovsa (Avena sativa L.). Na podlagi fenotipa semena sorte \u27Kamil\u27 (dva neodvisna vzorca 284 in 285) so bili pripravljeni štirje pod-vzorci (284AN, 284AS, 285AN, 285AS), kjer AN pomeni golce, AS pa plevence. Kot standardi navadnega ovsa so bili v genetsko identifikacijo vključene še sorta navadnega ovsa \u27Noni\u27 in štiri akcesije navadnega ovsa (ACC378, ACC379, ACC380, ACC381) iz Slovenske rastlinske genske banke. Skupno je bilo v genotipizacijo vključenih 36 vzorcev rastlin rodu Avena sp. Z uporabo seta šestih visoko informativnih SSR markerjev in rezultatov analize parametrov raznolikosti na posameznih lokusih ter parnih genetskih primerjav smo lahko potrdili, da so analizirani pod-vzorci pripadali isti vrsti in sorti, t.j. golemu ovsu \u27Kamil\u27. Poleg tega so bili kljub morfološki podobnosti plevenci sorte Kamil‘ genetsko dovolj različni od analizirane sorte in/ali akcesij navadnega ovsa

Drought Stress Response in Agricultural Plants: A Case Study of Common Bean (<em>Phaseolus vulgaris</em> L.)

Drought is one of the major threats to common bean (Phaseolus vulgaris L.), affecting its growth and productivity and, thus, contributing to considerable losses in yield in many regions worldwide. The development of varieties tolerant of drought stress has, therefore, become one of the primary goals in many common bean breeding programs. Plants have developed various mechanisms of their adaptation, to a greater or lesser extent, to drought. These are expressed, on the molecular level, by changes of gene expression and of protein content, together with responses at physiological and morphological levels. The response of common bean to drought is still not sufficiently well characterized due to its genetic complexity and its diverse, often ambiguous, phenotypic effects. Understanding these mechanisms is thus of fundamental importance for developing varieties that are better adapted to such stress conditions. In this chapter, we present research that provides an insight into the morpho-physiological adaptation and its underlying molecular changes in common bean plants exposed to drought. We include our contribution to establishing the basis for breeding of common bean with greater tolerance to this abiotic stress that uses molecular markers and identification of quantitative trait loci (QTLs)

The Influence of Different Substrates on the Growth, Yield and Quality of Slovenian Sweetpotato Cultivars under Greenhouse Conditions

A greenhouse experiment was conducted to evaluate the genetic relatedness between three Slovenian sweetpotato cultivars; and to assess the effects of different growing substrates on selected agronomic and nutritional traits. Tubers of three cultivars (‘Lučka’, ‘Janja’ and ‘Martina’) with different skin/flesh color were produced in planters under glasshouse conditions in five different growing substrates (perlite, peat, expanded clay, vermiculite and garden soil) from prior raised seedlings. Genetic analysis was performed using a set of eight SSR markers. According to Nei’s genetic distance and pairwise population Fst analysis, the most related cultivars are ‘Janja’ and ‘Martina’. The following agronomic traits were evaluated: vine length, thickness of vine-base, number of branches, weight of above ground part, number of leaves plant−1, number of tubers plant−1 and tubers weight plant−1. Among nutritional traits, total phenolic content (TPC), antioxidant potential (AOP) and ascorbic acid content (AA) were determined. Significant interactions of growing substrates (factor A) × cultivar (factor B) were observed for thickness of vine-base, weight of above ground part, AOP, TPC and AA. Overall results show different response of cultivars in different growing substrate. Growing substrate provide a discriminant classification of the sweetpotato cultivars according to their agronomic and nutritional traits

Validation of a One-Step Reverse Transcription-Droplet Digital PCR (RT-ddPCR) Approach to Detect and Quantify SARS-CoV-2 RNA in Nasopharyngeal Swabs

Background. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has rapidly spread worldwide from the beginning of 2020. Quantitative reverse transcription-PCR (RT-qPCR) is, to this day, the preferred methodology for viral RNA detection, even if not without problems. To overcome some of the limitations still existing for the detection and quantification of nucleic acids in various applications, the use of one-step reverse transcription-droplet digital PCR (RT-ddPCR) has been established. The purpose of this study was, then, to evaluate the efficacy of ddPCR for the detection of SARS-CoV-2 RNA in nasopharyngeal swabs, optimizing the detection of low-viral load-burdened samples. Methods. The RT-ddPCR workflow was validated for sensitivity, specificity, linearity, reproducibility, and precision using samples from 90 COVID-19-infected patients referred to the Department of Laboratory Medicine of the University Hospital of Udine (Italy). Results. The present study shows that RT-ddPCR allows the detection of as low as 10.3 copies of a SARS-COV-2 E-gene per sample with a higher level of accuracy and precision, especially at low concentration. Conclusion. During the postpeak phase of the SARS-CoV-2 pandemic, it is essential to rely on a highly robust molecular biology method to identify infected subjects, whether they have symptoms or not, in order to prepare appropriate containment measures

Genetic diversity of Lathyrus sativus L. collection and characteristics of seeds grown in Slovenia and Serbia

Grass pea (Lathyrus sativus L.) is a less common and nearly forgotten crop in many countries, including Slovenia and Serbia. In the present study, genetic analysis and seed characteristics of the collection of 22 grass pea accessions from several Southeast European plant gene banks were examined. The collection was produced in an open-field experiment at two countries/locations, i.e. Slovenia/Ljubljana – Jablje and Serbia/Novi Sad – Rimski Šančevi, during the 2019 growing season. For analysis of genetic diversity, specific high polymorphic SSR markers were applied. Initial screening at 12 species-specific SSR loci using binary data revealed a great genetic diversity among the grass pea accessions within a collection and three genetic groups were formed. Differences in seed size were measured using descriptors for seed length, width and thickness, and 100-seed weight. Furthermore, several seed colour characters using IBPGR descriptors were visually assessed. The average seed length, width and thickness of the seeds in the collection were 9.0 mm, 8.3 mm and 5.4 mm, respectively, while 100-seed weight ranged considerably from 11.7 g to 38.9 g. The highest differences between growing location were seen for 100-seed weight where the coefficient of variability reached 23.5%. The examined grass pea accessions had less monochromatic (one colour) and more colourful seeds, with two or three colours

Morpho-Agronomic Characterisation of Runner Bean (Phaseolus coccineus L.) from South-Eastern Europe

In South-Eastern Europe, the majority of runner-bean (Phaseolus coccineus L.) production is based on local populations grown mainly in home gardens. The local runner-bean plants are well adapted to their specific growing conditions and microclimate agro-environments, and show great morpho-agronomic diversity. Here, 142 runner-bean accessions from the five South-Eastern European countries of Slovenia, Bosnia and Herzegovina, Serbia, North Macedonia and Romania were sown and cultivated in their respective countries and characterised using 28 quantitative and qualitative morpho-agronomic descriptors for Phaseolus spp. based on inflorescences, leaves, plants, pods and seeds. For each of these morpho-agronomic descriptors, the accessions can be classified into two or three specific groups. The highest correlations were observed within the fluorescence, seed and pod traits. The highest variability, at 76.39%, was between the different countries, representing different geographic origins, while the variability within the countries was 23.61%. Cluster analysis based on these collected morpho-agronomic data also classified the accessions into three groups according to genetic origins. The data obtained serve as useful genetic information for plant breeders for the breeding of new bean varieties for further studies of the morpho-agronomic traits of the runner bean

Examination of Genetic Diversity of Common Bean from the Western Balkans

In this study, genetic diversity of 119 accessions of common bean (Phaseolus vulgaris) from five former Yugoslav republics constituting the western Balkans was assessed by 13 microsatellite markers. This set of markers has proven before to efficiently distinguish between bean genotypes and assign them to either the Andean or the Mesoamerican gene pool of origin. In this study, 118 alleles were detected or 9.1 per locus on average. Four groups (i.e., Slovene, Croatian, Bosnian, and Serbian) showed similarly high levels of genetic diversity as estimated by the number of different alleles, number of effective alleles, Shannon's information index, and expected heterozygosity. Mildly narrower genetic diversity was identified within a group of Macedonian accessions; however, this germplasm yielded the highest number of private alleles. All five germplasms share a great portion of genetic diversity as indicated by the analysis of molecular variance (AMOVA). On the basis of the scored number of migrants, we concluded that the most intensive gene flow in the region exists in Bosnia and Herzegovina. Cluster analysis based on collected molecular data classified the accessions into two large clusters that corresponded to two gene pools of origin (i.e., Andean and Mesoamerican). We found that Andean genotypes are more prevalent than Mesoamerican in all studied countries, except Macedonia, where the two gene pools are represented evenly. This could indicate that common bean was introduced into the western Balkans mainly from the Mediterranean Basin. Bayesian cluster analysis revealed that in the area studied an additional variation exists which is related to the Andean gene pool. Different scenarios of the origin of this variation are discussed in the article

Trypsin inhibitor activity in grass pea seeds (Lathyrus sativus L.)

Grass pea (Lathyrus sativus) is a valuable grain legume known for its high protein content and rich essential amino acid profile. Its exceptional characteristics such as drought tolerance, high adaptability to extreme conditions, disease resistance, and low cultivation inputs, make it particularly suitable for cultivation by resource-poor farmers. However, the potential use of grass pea is limited due to the presence of anti-nutritional factors, including protease inhibitors, especially trypsin inhibitor. This study aimed to develop a rapid and reliable method for measuring TI activity in seeds of grass pea and to investigate the influence of genotype and environment on trypsin inhibitor (TI) activity in seeds of grass pea. A set of 25 grass pea accessions from seven European countries was cultivated in Slovenia at the Agricultural Institute of Slovenia, and a set of 12 grass pea accessions from the Serbian gene bank was grown in Serbia at the Institute of Field and Vegetable Crops. The TI activity varied considerably among the grass pea accessions studied, with values ranging from 26.7 to 90.3 TUI/mg. To further evaluate the effects of environmental conditions on TI activity, eight grass pea accessions originating from Serbia were grown in both Slovenia and Serbia. The TI activity of the accessions grown in Slovenia ranged from 26.7 to 81.0 TUI/mg, while the activity of the accessions grown in Serbia ranged from 40.3 to 57.0 TUI/mg. The correlation of TI activity between grass pea accessions grown in Slovenia and those grown in Serbia was 0.39, with genotype diversity being the largest contributor (55.9%). This study provides a valuable insight into the variability of TI activity in grass pea and shows the possible influence of environmental conditions on this trait. However, since the data are only from a one-year field trial, further studies are needed to fully evaluate the influence of different environmental factors on TI activity

Evaluation of Lathyrus sativus L. seed for protein content and trypsin inhibitor activity

Grass pea (Lathyrus sativus L.) is a legume with high protein content and good quantities of essential amino acids. It has high tolerance to drought, high adaptability to extreme conditions, disease resistance and low input requirement for cultivation, which makes it very useful for cultivation by resource-poor farmers. Potential usage of grass pea is limited due to the presence of anti-nutritional factors including protease inhibitors, especially trypsin inhibitors. They have a major impact on nutritional value, by inhibiting pancreatic serine proteases (thus impairing protein digestion), but are also likely to protect legume seed against attack by predators. Although once extensively grown throughout Balkan Peninsula, grass pea was nearly forgotten. In recent years there was an effort to reintroduce and increase the interest of researchers and farmers in this crop. The aim of this study was to establish whether genotype and environment have significant influences on trypsin inhibitor activity (TIA) in grass pea and to determine whether TIA is correlated with seed protein content