Countrywide screening supports model-based predictions of the distribution of Batrachochytrium dendrobatidis in Ukraine.

The chytrid Batrachochytrium dendrobatidis (Bd) is a widespread fungus causing amphibian declines across the globe. Although data on Bd occurrence in Eastern Europe are scarce, a recent species distribution model (SDM) for Bd reported that western and north-western parts of Ukraine are highly suitable to the pathogen. We verified the SDM-predicted range of Bd in Ukraine by sampling amphibians across the country and screening for Bd using qPCR. A total of 446 amphibian samples (tissue and skin swabs) from 11 species were collected from 36 localities. We obtained qPCR-positive results for 33 samples including waterfrogs (Pelophylax esculentus complex) and fire- and yellow-bellied toads (Bombina spp.) from 8 localities. We found that Bd-positive localities had significantly higher predicted Bd habitat suitability than sites that were pathogen-free. Amplification and sequencing of the internal transcribed spacer (ITS) region of samples with the highest Bd load revealed matches with ITS haplotypes of the globally distributed BdGPL strain, and a single case of the BdASIA-2/BdBRAZIL haplotype. We found that Bd was non-randomly distributed across Ukraine, with infections present in the western and north-central forested peripheries of the country with a relatively cool, moist climate. On the other hand, our results suggest that Bd is absent or present in low abundance in the more continental central, southern and eastern regions of Ukraine, corroborating the model-predicted distribution of chytrid fungus. These areas could potentially serve as climatic refugia for Bd-susceptible amphibian hosts.

Structure, Physicochemical Properties and Biological Activity of Lipopolysaccharide from the Rhizospheric Bacterium Ochrobactrum quorumnocens T1Kr02, Containing d-Fucose Residues.

Lipopolysaccharides (LPSs) are major components of the outer membranes of Gram-negative bacteria. In this work, the structure of the O-polysaccharide of Ochrobactrum quorumnocens T1Kr02 was identified by nuclear magnetic resonance (NMR), and the physical-chemical properties and biological activity of LPS were also investigated. The NMR analysis showed that the O-polysaccharide has the following structure: →2)-ß-d-Fucf-(1→3)-ß-d-Fucp-(1→. The structure of the periplasmic glucan coextracted with LPS was established by NMR spectroscopy and chemical methods: →2)-ß-d-Glcp-(1→. Non-stoichiometric modifications were identified in both polysaccharides: 50% of d-fucofuranose residues at position 3 were O-acetylated, and 15% of d-Glcp residues at position 6 were linked with succinate. This is the first report of a polysaccharide containing both d-fucopyranose and d-fucofuranose residues. The fatty acid analysis of the LPS showed the prevalence of 3-hydroxytetradecanoic, hexadecenoic, octadecenoic, lactobacillic, and 27-hydroxyoctacosanoic acids. The dynamic light scattering demonstrated that LPS (in an aqueous solution) formed supramolecular particles with a size of 72.2 nm and a zeta-potential of -21.5 mV. The LPS solution (10 mkg/mL) promoted the growth of potato microplants under in vitro conditions. Thus, LPS of O. quorumnocens T1Kr02 can be recommended as a promoter for plants and as a source of biotechnological production of d-fucose.

N-Stearoylethanolamine Inhibits Integrin-Mediated Activation, Aggregation, and Adhesion of Human Platelets.

N-stearoylethanolamine (NSE), a lipid mediator that belongs to the N-acylethanolamine (NAE) family, has anti-inflammatory, antioxidant, and membranoprotective actions. In contrast to other NAEs, NSE does not interact with cannabinoid receptors. The exact mechanism of its action remains unclear. The aim of this study is to evaluate the action of NSE on activation, aggregation, and adhesion of platelets that were chosen as a model of cellular response. Aggregation of platelets was measured to analyze the action of NSE (10-6-10-10 M) on platelet reactivity. Changes in granularity and shape of resting platelets and platelets stimulated with ADP in the presence of NSE were monitored by flow cytometry, and platelet deganulation was monitored by spectrofluorimetry. In vivo studies were performed using obese insulin-resistant rats. Binding of fibrinogen to the GPIIb/IIIa receptor was estimated using indirect ELISA and a scanning electron microscopy (SEM). It was found that NSE inhibits the activation and aggregation of human platelets. Our results suggest that NSE may decrease the activation and subsequent aggregation of platelets induced by ristocetin, epinephrine, and low doses of ADP. NSE also reduced the binding of fibrinogen to GPIIb/IIIa on activated platelets. These effects could be explained by the inhibition of platelet activation mediated by integrin receptors: the GPIb-IX-V complex for ristocetin-induced activation and GPIIb/IIIa when epinephrine and low doses of ADP were applied. The anti-platelet effect of NSE complements its anti-inflammatory effect and allows us to prioritize studies of NSE as a potent anti-thrombotic agent. SIGNIFICANCE STATEMENT: N-stearoylethanolamine (NSE) was shown to possess inhibitory action on platelet activation, adhesion, and aggregation. The mechanism of inhibition possibly involves integrin receptors. This finding complements the known anti-inflammatory effects of NSE.

Lipopolysaccharide and flagellin of Azospirillum brasilense Sp7 influence callus morphogenesis and plant regeneration in wheat.

In vitro somatic callus culturing is used widely in plant biotechnology, but its effectiveness depends largely on the donor plant genotype. Bacteria or components of their cells are rarely used to activate morphogenesis. In this work, inoculation of explants from immature wheat (Triticum aestivum L.) embryos with a suspension of living cells of the bacterium Azospirillum brasilense Sp7 resulted in callus death after 7 days of growth, in contrast to explant treatment with a suspension of heat-killed whole cells of Sp7. The experiments used two wheat lines, LRht-B1a and LRht-B1c, which differ in morphogenic activity. Growing calluses with the lipopolysaccharide of A. brasilense Sp7 increased the yield of regenerated plants 2- to 3.5-fold in both lines. This increase was through the activation of regenerant formation from morphogenic calluses. We have demonstrated for the first time the effects of bacterial flagellin on plant tissue culture. The polar-flagellum flagellin of A. brasilense Sp7 leveled the genotypic differences in the morphogenic ability of callus tissue. Specifically, it increased the yield of morphogenic calluses in the weakly morphogenic line LRht-B1a to the yield value in the highly morphogenic line LRht-B1c but lowered the yield of regenerants in the highly morphogenic line LRht-B1c to the yield value in the weakly morphogenic line LRht-B1a. Thus, bacterial lipopolysaccharides and flagellins can be used to regulate the formation of morphogenic calluses and regenerants in plant tissue culturing in vitro.

AGE-RELATED MORPHO-FUNCTIONAL CHANGES IN RATS' PANCREAS UNDER HIGH-FAT DIET-INDUCED INSULIN RESISTANCE AND ITS PHARMACOLOGICAL TREATMENT.

OBJECTIVE: The aim: To determine the set of structural and functional changes in pancreatic islets (PI) of obesity-induced insulin resistant (IR) rats of different age (young and old) fed with prolonged (6 month) high-fat diet (HFD) (58% of fat) and further treatment with N-Stearoylethanolamine (NSE), a bioactive N-Acylethanolamine. PATIENTS AND METHODS: Materials and methods: Alimentary obesity-induced IR model in rats of two age groups was used to investigate the influence of age and NSE treatment on pancreas morphology (using histological, histochemical and immunohistochemical techniques) and on several biochemical parameters associated with DM onset. RESULTS: Results: The NSE administration normalized pancreas morphology which was more affected in the old IR group; the signs of inflammation, edema, fibrosis and steatosis were somehow diminished and PI area became significantly increased. The amount of the A-F-positive insulocytes increased and TUNEL-positive - decreased. Compensatory hyperplasia in the affected pancreas of both age was an important indicator of NSE stimulating effect. CONCLUSION: Conclusions: Protective effects of NSE on morpho-functional state of pancreas in HFD-induced IR rats of both age are associated not only with its anti-inflammatory, anti-oxidant and anti-dyslipidemic properties but also with activation of PI hyperplasia and ß-cells compensatory mechanisms.

Ochrobactrum cytisi IPA7.2 promotes growth of potato microplants and is resistant to abiotic stress.

Bacteria in natural associations with agricultural crops are promising for use in the improvement of clonal micropropagation of plants. We clarified the taxonomic position of Ochrobactrum cytisi strain IPA7.2 and investigated its tolerance for salinity, high temperature, and glyphosate pollution. We also tested the strain's potential to promote the growth of potato (Solanum tuberosum L.) microplants. Using the IPA7.2 draft genome (no. NZ_MOEC00000000), we searched for housekeeping genes and also for the target genes encoding glyphosate tolerance and plant-growth-promoting ability. A multilocus sequence analysis of the gap, rpoB, dnaK, trpE, aroC, and recA housekeeping genes led us to identify isolate IPA7.2 as O. cytisi. The strain tolerated temperatures up to 50 °C and NaCl concentrations up to 3-4%, and it produced 8 µg ml-1 of indole-3-acetic acid. It also tolerated 6 mM glyphosate owing to the presence of type II 5-enolpyruvylshikimate-3-phosphate synthase. Finally, it was able to colonize the roots and tissues of potato microplants, an ability preserved by several generations after subculturing. We identified the development phase of potato microplants that was optimal for inoculation with O. cytisi IPA7.2. Inoculation of in vitro-grown 15-day-old microplants increased the mitotic index of root meristem cells (by 50%), the length of shoots (by 34%), the number of leaves (by 7%), and the number of roots (by 16%). Under ex vitro conditions, the inoculated plants had a greater leaf area (by 77%) and greater shoot and root dry weight (by 84 and 61%, respectively) than did the control plants. We recommend O. cytisi IPA 7.2 for use in the growing of potato microplants to improve the production of elite seed material.

Functioning of plant-bacterial associations under osmotic stress in vitro.

The search for effective plant-growth-promoting strains of rhizospheric bacteria that would ensure the resistance of plant-microbial associations to environmental stressors is essential for the design of environmentally friendly agrobiotechnologies. We investigated the interaction of potato (cv. Nevsky) microplants with the plant-growth-promoting bacteria Azospirillum brasilense Sp245 and Ochrobactrum cytisi IPA7.2 under osmotic stress in vitro. The bacteria improved the physiological and biochemical variables of the microplants, significantly increasing shoot length and root number (1.3-fold, on average). Inoculation also led a more effective recovery of the plants after stress. During repair, inoculation contributed to a decreased leaf content of malonic dialdehyde. With A. brasilense Sp245, the decrease was 1.75-fold; with O. cytisi IPA7.2, it was 1.4-fold. During repair, the shoot length, node number, and root number of the inoculated plants were greater than the control values by an average of 1.3-fold with A. brasilense Sp245 and by an average of 1.6-fold with O. cytisi IPA7.2. O. cytisi IPA7.2, previously isolated from the potato rhizosphere, protected the physiological and biochemical processes in the plants under stress and repair better than did A. brasilense Sp245. Specifically, root weight increased fivefold during repair, as compared to the noninoculated plants, while chlorophyll a content remained at the level found in the nonstressed controls. The results indicate that these bacteria can be used as components of biofertilizers. A. brasilense Sp245 has favorable prospects for use in temperate latitudes, whereas O. cytisi IPA7.2 can be successfully used in saline and drought-stressed environments.

Effect of bacterial lipopolysaccharides on morphogenetic activity in wheat somatic calluses.

We evaluated the effect of lipopolysaccharides from the plant-growth-promoting associative bacterium Azospirillum brasilense Sp245 and from the enteric bacterium Escherichia coli K12 on the morphogenic potential of in vitro-growing somatic calluses of soft spring wheat (Triticum aestivum L. cv. Saratovskaya 29). A genetic model was used that included two near-isogenic lines of T. aestivum L. cv. Saratovskaya 29 with different embryogenic capacities; one of these lines carries the Rht-B1 dwarfing gene, whereas the other lacks it. When added to the nutrient medium, the lipopolysaccharide of A. brasilense Sp245 promoted the formation of calluses with meristematic centers and stimulated the regeneration ability of the cultured tissues in both lines. By contrast, the lipopolysaccharide of the enteric bacterium E. coli K12 barely affected the morphogenetic activity of callus cells and the yield of morphogenic calluses and regenerated plants. These findings indicate that the lipopolysaccharide of the plant-growth-promoting associative bacterium A. brasilense Sp245 specifically enhances the morphogenetic activity of wheat somatic tissues, which increases the efficacy of culturing of genotypes with a relatively low morphogenic potential. The results of the study may contribute to the improvement of the efficacy of plant cell selection and gene engineering and to a better understanding of the mechanisms responsible for plant recognition of lipopolysaccharides of associative bacteria.

N-Stearoylethanolamine Exerts Cardioprotective Effects in Old Rats.

BACKGROUND: Aging is associated with the slowing down of metabolic processes, diminished physiological processes, changes in hormonal activity and increasing exposure to oxidative stress factors and chronic inflammation. The endocannabinoid system (ECS) is a major signaling network that plays a pro-homeostatic role in the central and peripheral organs of the human body. A class of minor lipids, N-acylethanolamines (NAEs), which do not activate cannabinoid receptors, except for anandamide, but can potentiate the action of endocannabinoids and have a wide spectrum of biological activity and significant adaptogenic potential, belongs to ECS. The results of different studies over the past decades have established the protective effect of NAE on many pathological conditions. OBJECTIVE: This study aimed to investigate the cardioprotective effects of C18:0 NAE- N-stearoylethanolamine (NSE) in aged rats. In this study, we focused on investigating the effects of C18:0 NAE- N-stearoylethanolamine (NSE) on the intensity of oxidative/ nitrosative stress, antioxidant potential, lipoprotein profile and inflammation markers of blood plasma, phospholipid composition and age-related morphological changes of old rat heart tissues. METHODS: The study was conducted on Sprague Dawley male laboratory rats. The three groups of rats were involved in the study design. The first group consisted of young rats aged 4 months (n=10). The second (n=10) and third (n=10) groups included old rats aged of 18 months. Rats from the third group were administered a per os aqueous suspension of NSE at a dose of 50 mg/kg of body weight daily for 10 days. All groups of rats were kept on a standard vivarium diet. The blood plasma, serum, and heart of rats were used for biochemical and histological analysis. RESULTS: The cardioprotective effect of N-stearoylethanolamine in old rats was established, which was expressed in the normalization of the antioxidant system condition and the level of proinflammatory cytokines, positive modulation of blood plasma and lipoprotein profile, normalization of heart tissue lipid composition, and significant reduction in age-related myocardium morphological changes. CONCLUSION: The revealed effects of N-stearoylethanolamine can become the basis for developing a new drug for use in complex therapy to improve the quality of life of older people.

Rhizobacteria Increase the Adaptation Potential of Potato Microclones under Aeroponic Conditions.

Adaptation ex vitro is strongly stressful for microplants. Plant-growth-promoting rhizobacteria (PGPR) help to increase the adaptation potential of microplants transplanted from test tubes into the natural environment. We investigated the mechanisms of antioxidant protection of PGPR-inoculated potato microclones adapting to ex vitro growth in an aeroponic system. Potato (Solanum tuberosum L. cv. Nevsky) microplants were inoculated in vitro with the bacteria Azospirillum baldaniorum Sp245 and Ochrobactrum cytisi IPA7.2. On days 1 and 7 of plant growth ex vitro, catalase and peroxidase activities in the leaves of inoculated plants were 1.5-fold higher than they were in non-inoculated plants. The activity of ascorbate peroxidase was reduced in both in vitro and ex vitro treatments, and this reduction was accompanied by a decrease in the leaf content of hydrogen peroxide and malondialdehyde. As a result, inoculation contributed to the regulation of the plant pro/antioxidant system, lowering the oxidative stress and leading to better plant survival ex vitro. This was evidenced by the higher values of measured morphological and physiological variables of the inoculated plants, as compared with the values in the control treatment. Thus, we have shown some PGPR-mediated mechanisms of potato plant protection from adverse environmental factors under aeroponic conditions.

Rhizobacteria Inoculation Effects on Phytohormone Status of Potato Microclones Cultivated In Vitro under Osmotic Stress.

Water deficits inhibit plant growth and decrease crop productivity. Remedies are needed to counter this increasingly urgent problem in practical farming. One possible approach is to utilize rhizobacteria known to increase plant resistance to abiotic and other stresses. We therefore studied the effects of inoculating the culture medium of potato microplants grown in vitro with Azospirillum brasilense Sp245 or Ochrobactrum cytisi IPA7.2. Growth and hormone content of the plants were evaluated under stress-free conditions and under a water deficit imposed with polyethylene glycol (PEG 6000). Inoculation with either bacterium promoted the growth in terms of leaf mass accumulation. The effects were associated with increased concentrations of auxin and cytokinin hormones in the leaves and stems and with suppression of an increase in the leaf abscisic acid that PEG treatment otherwise promoted in the potato microplants. O. cytisi IPA7.2 had a greater growth-stimulating effect than A. brasilense Sp245 on stressed plants, while A. brasilense Sp245 was more effective in unstressed plants. The effects were likely to be the result of changes to the plant's hormonal balance brought about by the bacteria.

Structure, gene cluster of the O antigen and biological activity of the lipopolysaccharide from the rhizospheric bacterium Ochrobactrum cytisi IPA7.2.

Lipopolysaccharide (LPS) of Ochrobactrum cytisi IPA7.2, a bacterium isolated from the roots of Solanum tuberosum L., was extracted from dry bacterial cells and chemically characterized. The O-specific polysaccharide was obtained by mild acid hydrolysis of the LPS and studied by sugar analysis and 1H and 13C NMR spectroscopy, including 1H,1H COSY, 1H,1H TOCSY, 1H,1H ROESY, 1H,13C HSQC, and 1H,13C HMBC experiments. The polysaccharide was linear and consisted of trisaccharide repeating units of the following structure: A putative O-antigen gene cluster of O. cytisi IPA7.2 was identified and found to be consistent with the O-specific polysaccharide structure. The LPS of Ochrobactrum cytisi IPA7.2 promoted the growth of potato microplants in vitro.

Study of total antioxidant capacity of red raspberry (Rubus idaeous L. ) shoots. / Estudio de la capacidad antioxidante total de brotes de frambuesa roja (Rubus idaeous L. ).

Background: Today, cardiovascular, oncological, and neurodegenerative diseases are the main causes of death in the world, according to official World Health Organization (WHO) statistics. Antioxidants are used to treat and prevent these diseases. In order to develop optimal technology for obtaining drugs based on plant extracts with antioxidant action, it is necessary to determine the total antioxidant capacity of raspberry shoots. Objectives: The study aimed to determine the total antioxidant capacity of red raspberry shoots, study the content of biologically active substances (BAS), and the antioxidant activity of red raspberry shoot extracts obtained during subsequent exhaustive extraction. Methods: The number of phenolic compounds, catechins, flavonoids, and hydroxycinnamic acids was determined by a spectrophotometric analysis method, whereas organic acids were determined by the alkalimetric method in red raspberry shoot extracts; the antioxidant activity of obtained extracts was evaluated by potentiometric method. Results: The total antioxidant capacity of red raspberry shoots was 164.12 mmol-equiv./m dry weight, the sum of the total content of phenolic compounds was 24.40 mg gallic acid (GA)/mL, catechins ­ 21.36 mg epigallocatechin-3-O-gallate (EGCG)/mL, flavonoids ­ 0.77 mg rutin (R)/mL, hydroxycinnamic acids derivatives ­ 2.56 mg chlorogenic acid (ChA)/mL and organic acids ­ 1.88 mg citric acid (CA)/mL in red raspberry shoot extracts obtained during subsequent exhaustive extraction. The analysis showed that there is a very high positive correlation between antioxidant activity and total phenolic compounds, catechin, flavonoid, hydroxycinnamic acids derivatives, and organic acids content in red raspberry shoot extracts. Conclusions: Total red raspberry shoots' antioxidant capacity has been determined. The study results can be used to develop optimal technology for obtaining drugs based on the extract of red raspberry shoots, which has an antioxidant effect

Contexto: Hoy en día, las enfermedades cardiovasculares, oncológicas y neurodegenerativas son las principales causas de muerte en el mundo según estadísticas oficiales de la Organización Mundial de la Salud OMS. Los antioxidantes se utilizan para tratar y prevenir estas enfermedades. Para desarrollar una tecnología óptima para la obtención de fármacos a base de extractos de plantas con acción antioxidante, es necesario determinar la capacidad antioxidante total de los brotes de frambuesa.Objetivos: El estudio tuvo como objetivo determinar la capacidad antioxidante total de los brotes de frambuesa roja, estudiar el contenido de sustancias biológicamente activas (SBA) y la actividad antioxidante de los extractos de brotes de frambuesa roja obtenidos mediante extracción exhaustiva. Métodos: La cantidad de compuestos fenólicos, catequinas, flavonoides y ácidos hidroxicinámicos se determinó por método de análisis espectrofotométrico, mientras que los ácidos orgánicos por método alcalimétrico en extractos de brotes de frambuesa roja; La actividad antioxidante de los extractos obtenidos se evaluó por método potenciométrico. Resultados: La capacidad antioxidante total de los brotes de frambuesa roja fue de 164.12 mmol-equiv./m de peso seco, la suma del contenido total de compuestos fenólicos fue de 24.40 mg gálico ácido (GA)/mL, catequinas ­ 21.36 mg epigalocatequina-3-O-galato (EGCG)/mL, flavonoides ­ 0.77 mg rutina (R)/mL, derivados de ácidos hidroxicinámicos ­ 2.56 mg clorogénico ácido (ChA)/mL y ácidos orgánicos ­ 1.88 mg cítrico ácido (CA)/mL en extractos de brotes de frambuesa roja obtenidos durante extracción exhaustiva. La correlación analizada mostró que existe una correlación positiva entre la actividad antioxidante y el contenido de compuestos fenólicos totales, catequinas, flavonoides, derivados de ácidos hidroxicinámicos y ácidos orgánicos en extractos de brotes de frambuesa roja. Conclusiones: Gracias a nuestros resultados se ha determinado la capacidad antioxidante total de los brotes de frambuesa roja. Los resultados del estudio se pueden utilizar para desarrollar una tecnología óptima para la obtención de fármacos basados en el extracto de brotes de frambuesa roja, que tiene un efecto antioxidante