Vidmantas Stanys

Expression and Anthocyanin Biosynthesis-Modulating Potential of Sweet Cherry (Prunus avium L.) MYB10 and bHLH Genes

Anthocyanins are essential contributors to fruit coloration, an important quality feature and a breed determining trait of a sweet cherry fruit. It is well established that the biosynthesis of anthocyanins is regulated by an interplay of specific transcription factors belonging to MYB and bHLH families accompanied by a WD40 protein. In this study, we isolated and analyzed PaWD40, PabHLH3, PabHLH33, and several closely related MYB10 gene variants from different cultivars of sweet cherry, analyzed their expression in fruits with different anthocyanin levels at several developmental stages, and determined their capabilities to modulate anthocyanin synthesis in leaves of two Nicotiana species. Our results indicate that transcription level of variant PaMYB10.1-1 correlates with fruit coloration, but anthocyanin synthesis in Nicotiana was induced by another variant, PaMYB10.1-3, which is moderately expressed in fruits. The analysis of two fruit-expressed bHLH genes revealed that PabHLH3 enhances MYB-induced anthocyanin synthesis, whereas PabHLH33 has strong inhibitory properties.

Characterization of apple NADPH oxidase genes and their expression associated with oxidative stress in shoot culture in vitro

Genes encoding Rboh (respiratory burst oxidase homolog) have been described in a variety of plant species. Transcriptional regulation of rboh genes has been shown during plant response to stress or phytohormone treatment. In vitro conditions often induce plant stress that leads to slow plant growth, early senescence, or even recalcitrance to in vitro growth. Production of reactive oxygen species (ROS) is an important manifestation of plant environmental stress. Expression of rboh orthologs in plant tissues grown under in vitro conditions and their role in response to abiotic stress is not fully understood. Therefore the aim of this study was to identify rboh homologues in apple (Mdrboh) and to characterize their expression during the senescence of apple shoot culture in vitro. Similarity searches using Arabidopsis Rboh (AtRboh) protein sequences revealed nine homologous rboh genes in the apple genome. Phylogenetic analysis using conservative N-terminal half region sequences of the apple, Arabidopsis, pear, peach and wild strawberry Rboh proteins revealed four groups of related sequences that were linked to AtRboh D, E, F and H. A homologue with unique sequence was annotated as Rboh K in plants of the Rosaceae family. Expression of MdrbohD1–3 and F was detected in apple leaves, in vitro cultures of shoots and cell suspension, and MdrbohE2 and H1–2 varied among the tissues. The MdrbohD1–2 and F genes were differentially expressed during transfer, growth and senescence stages of apple in vitro shoot culture having varying levels of oxidative stress damage, suggesting transcriptional regulation of the Mdrboh genes in the apple shoot culture, and a potential for distinctive functions of the three Rboh D orthologs.

Cold acclimation efficiency of different Prunus and Fragaria species and cultivars in vitro@@@Įvairių Prunus ir Fragaria rūšių bei veislių augalų užsigrūdinimo in vitro efektyvumas

Winter hardiness is one of the main traits affecting survival, productivity and cultivar choice for important horticultural plants in a temperate climatic zone. Given the complexity of the trait and the often fluctuating winter weather, extensive field testing over many years would be required to gain reliable results. Testing for cold tolerance under controlled conditions to grade genotypes has been used in several plant breeding programs to identify superior genotypes of different species. The aim of the present study was to investigate cold tolerance of microshoots of different genotypes of the genera Prunus and Fragaria after 7, 14, 28 and 56 days of cold acclimation, by measuring ion leakage and evaluating the critical temperature (CT50) after freezing in vitro. Genotypes of Rosacea family demonstrated distinct patterns of CT50 change during cold acclimation. Cold acclimation for 56 days decreased the CT50 value by 1.3–2.0°C for Prunus microshots and by 0.8–2.1°C for Fragaria microshoots in comparison to nonacclimated plants. The results of cold acclimation and freezing treatments of Rosacea family plants in vitro show that for maximal cold hardiness, acclimation for 56 days or longer is required. Sour cherry (Prunus cerasus) cv. ‘Orkolija’, garden strawberry (Fragaria × ananassa) cv. ‘Melody’, Virginia strawberry (F. virginiana) and musk strawberry (F. moschata) were the most winterhardy after 56 days’ cold acclimation. The data obtained in the study can be used to improve breeding and cryopreservation technologies of Rosacea plants, and lay the foundation for identification of genes responsible for efficient cold acclimation and low temperature tolerance.

Computational analysis of thaumatin-II allergenicity and prediction of antigenic elements of thaumatin-like family proteins@@@Taumatino II alergeniškumo įvertinimas ir antigeniškumo elementų nustatymas bioinformatikos metodais

Thaumatin-II is an intense sweet tasting protein isolated from fruits of Thaumatococcus daniel lii. Thaumatin is a member of the pathogenesis-related protein family referred to as thaumatinlike proteins (TLPs). Expression of thaumatin-II in plants was associated with an enhanced resistance against pathogens. In addition, thaumatin was used to provide the sweet taste quality to plant products. Thus, TLP proteins have a potential application for improving plant stress resistance and taste qualities using the recombinant DNA technology. However, several members of the TLP family were characterized as food allergens. The allergenic properties of thaumatinII have not been characterized. In this study, a putative allergenicity of thaumatin-II was established using computational analysis of sequence similarity with known human allergens. Antigenicity analysis and multiple sequence alignment of related TLP sequences identified six putative allergenic epitopes. The residues Thr12, Leu74, Gln133 and Thr161 of thaumatin-II or equivalent residues of other TLPs were proposed as a target for mutagenesis aimed to develop protein isoforms with reduced allergenicity.

Endophytic Bacillus and Pseudomonas spp. Modulate Apple Shoot Growth, Cellular Redox Balance, and Protein Expression Under in Vitro Conditions

Interactions between host plants and endophytic microorganisms play an important role in plant responses to pathogens and environmental stresses and have potential applications for plant stress management under in vitro conditions. We assessed the effect of endophytic bacteria on the growth and proliferation of domestic apple cv. Gala shoots in vitro. Further, a model apple cell suspension system was used to examine molecular events and protein expression patterns at an early stage of plant–endophyte interaction. Among the seven strains used in the study, Bacillus spp. strains Da_1, Da_4, and Da_5 and the Pseudomonas fluorescens strain Ga_1 promoted shoot growth and auxiliary shoot proliferation. In contrast, Bacillus sp. strain Oa_4, P. fluorescens strain Ga_3 and P. orientalis strain G_12 inhibited shoot development. In the cell suspension, the effects of the association between endophytic bacteria and plant cells were specific to each strain. Modulation of the cellular redox balance was monitored in the apple cells using a 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA) probe, and strain-specific effects were observed that correlated with the in vitro shoot development results. Proteomic analysis revealed differences in protein expressions in apple cells co-cultivated with different Bacillus spp. strains that had contrasting effects on cellular redox balance and shoot development. The Bacillus sp. strain Da_4, which enhanced shoot development and oxidation of H2DCFDA, induced differential expression of proteins that are mainly involved in the defense response and regulation of oxidative stress. Meanwhile, treatment with Bacillus sp. strain Oa_4 led to strong upregulation of PLAT1, HSC70-1 and several other proteins involved in protein metabolism and cell development. Taken together, the results suggest that different cell signaling and response events at the early stage of the plant–endophyte interaction may be important for strain-dependent regulation of cellular redox balance and development of shoot phenotype.

Antimicrobial activity of berries extracts of four Ribes species, their phenolic content and anthocyanin composition

Abstract Phenolic compounds are widespread in berries and determine their antimicrobial activity. The aim of our study was to establish the amounts of phenolic compounds and the anthocyanin composition in berries of four Ribes species, and to evaluate the effect of berry extracts on the growth of common Gram-positive and Gram-negative bacteria, and also yeasts isolated from food processing plants. The phenolic content and anthocyanin composition were estimated spectrometrically and by HPLC, respectively. The highest amount of phenolic compounds, and also anthocyanins, was found in extracts of R. aureum ‘Corona’. The anthocyanin content was the lowest in berries of R. aureum Au Gs-5, with equal amounts of delphinidins and cyanidins. Delphinidins were predominant (68.6%) in berries of R. nigrum ‘Ben Tirran’, while cyanidins dominated in R. uva-crispa. The berry extracts of R. aureum Au Gs-5 and R. uva-crispa ‘Lûðiai’ had the largest growth-suppressing effect on yeasts and most of the bacteria tested. All of the berry extracts suppressed the growth of pathogenic and conditionally pathogenic bacteria. The industrially important Lactococcus lactis was the most resistant to the Ribes berry extracts. There was no correlation between the amount of anthocyanins in the extracts and their antimicrobial properties. Extracts with a lower anthocyanin–to-phenolics ratio more effectively inhibited the growth of bacteria.

Role of Endophytic Bacteria in Stress Tolerance of Agricultural Plants: Diversity of Microorganisms and Molecular Mechanisms

Bacterial endophytes are a group of endosymbiotic microorganisms widespread among plants. An association of plants with endophytic bacteria includes a vast diversity of bacterial taxa and host plants. In this review we present an overview of taxonomic composition of the bacterial endophytes identified in common agricultural crops with special emphasis on the most recent results obtained using metagenomic analysis. Endophytic microbiome constitutes a part of larger soil microbial community and is susceptible to direct or indirect effect of agricultural practices: soil tillage, irrigation, use of pesticides and fertilizers has a major effect on function and structure of soil and endophytic microbial populations. Therefore, the use of agricultural practices that maintain natural diversity of plant endophytic bacteria becomes important element of sustainable agriculture that ensures plant productivity and quality of agricultural production. On the other hand, the endophytic microbiome itself have been shown to have multiple effects on their host plant, including modulation of phytohormone signaling, metabolic activity, and plant defense response pathways. It has been demonstrated that these effects could be helpful for plant adaptation to abiotic or biotic stresses. Therefore, application of endophytic bacteria to improve crop performance under cold, drought, salinity, and heavy metal contamination stress conditions or to enhance disease resistance presents an important potential for sustainable agricultural production.

Agronomical Characters of Introduced New Blackcurrant Cultivars

The blackcurrant cultivars ‘Abanos’, ‘Ronix’, ‘Deea’, ‘Geo’ (Romania), ‘Almo’ (Estonia), ‘Narve Viking’, ‘Varde Viking’ (Norway), ‘Mikael’ (Finland), and standard cultivar ‘Ben Tirran’ (Scotland) were tested at the Institute of Horticulture, Lithuanian Research Centre for Agriculture and Forestry in 2009-2012. Two-year-old bushes were planted in an orchard in 2009. Bush parameters (vigour and width, m), resistance to anthracnose (Pseudopeziza ribis) and leaf spot (Septoria ribis), berry weight (weight of 100 fruits), yield (kg/bush) and biochemical composition were determined. The study showed that ‘Geo’ was most resistant to fungal disease. Fruits of ‘Abanos’ and ‘Almo’ were the largest. Yield of cultivars ‘Ronix’, ‘Ben Tirran’ and ‘Deea’ were the highest. ‘Abanos’ and ‘Geo’ had the highest content of soluble solids, ‘Ben Tirran’ of titratable acid, ‘Varde Viking’ of anthocyanins and phenols, and ‘Ronix’ and ‘Narve Viking’of ascorbic acid. Upeņu šķirņu pētījumi veikti Lietuvas Lauksaimniecības un Meža pētījumu centra Dārzkopības institūtā laikā no 2009 līdz 2012. gadam. Pētījumos iekļautas šķirnes ‘Abanos’, ‘Ronix’, ‘Deea\ ‘Geo’ (Rumānija), ‘Almo’ (Igaunija), ‘Narve Viking’, ‘Varde Viking’ (Norvēģija), ‘Mikael’ (Somija). Stādījums ierīkots 2009. gadā, izmantojot divgadīgus stādus. Pētīti krūmu parametri (augstums, platums, m) izturība pret ogulāju iedegām (Pseudopeziza ribis) un ogulāju sīkplankumainību (Septoria ribis), svērta ogu masa (100 ogu masa, g ), raža (kg/krūma) un analizēts bioķīmiskais saturs. Pētījumu rezultāti rāda, ka izturīgākā pret sēņu slimībām bija šķirne ‘Geo’. Lielākās ogas bija šķirnēm ‘Abanos’ un ‘Almo’. Augstākā ražība bija šķirnēm ‘Ronix’, ‘Ben Tirran’ un ‘Deea’, bet šķirnēm ‘Abanos’ and ‘Geo’ ogās bija augstākais šķīstošās sausnas saturs. Šķirnes ‘Ben Tirran’ ogas saturēja visvairāk titrējamās skābes , ‘Varde Viking’ antocianīnu un fenolu , ‘Ronix’ un ‘Narve Viking’ - askorbīnskābi.