The Effect of Plasma Activated Water on Maize (Zea mays L.) under Arsenic Stress.

Plasma activated water (PAW) is a source of various chemical species useful for plant growth, development, and stress response. In the present study, PAW was generated by a transient spark discharge (TS) operated in ambient air and used on maize corns and seedlings in the 3 day paper rolls cultivation followed by 10 day hydroponics cultivation. For 3 day cultivation, two pre-treatments were established, "priming PAW" and "rolls PAW", with corns imbibed for 6 h in the PAW and then watered daily by fresh water and PAW, respectively. The roots and the shoot were then analyzed for guaiacol peroxidase (G-POX, POX) activity, root tissues for their lignification, and root cell walls for in situ POX activity. To evaluate the potential of PAW in the alleviation abiotic stress, ten randomly selected seedlings were hydroponically cultivated for the following 10 days in 0.5 Hoagland nutrient solutions with and without 150 µM As. The seedlings were then analyzed for POX and catalase (CAT) activities after As treatment, their leaves for photosynthetic pigments concentration, and leaves and roots for As concentration. The PAW improved the growth of the 3 day-old seedlings in terms of the root and the shoot length, while roots revealed accelerated endodermal development. After the following 10 day cultivation, roots from PAW pre-treatment were shorter and thinner but more branched than the control roots. The PAW also enhanced the POX activity immediately after the imbibition and in the 3 day old roots. After 10 day hydroponic cultivation, antioxidant response depended on the PAW pre-treatment. CAT activity was higher in As treatments compared to the corresponding PAW treatments, while POX activity was not obvious, and its elevated activity was found only in the priming PAW treatment. The PAW pre-treatment protected chlorophylls in the following treatments combined with As, while carotenoids increased in treatments despite PAW pre-treatment. Finally, the accumulation of As in the roots was not affected by PAW pre-treatment but increased in the leaves.

Physiological Responses of Young Pea and Barley Seedlings to Plasma-Activated Water.

This study demonstrates the indirect effects of non-thermal ambient air plasmas (NTP) on seed germination and plant growth. It investigates the effect of plasma-activated water (PAW) on 3-day-old seedlings of two important farm plants-barley and pea. Applying different types of PAW on pea seedlings exhibited stimulation of amylase activity and had no inhibition of seed germination, total protein concentration or protease activity. Moreover, PAW caused no or only moderate oxidative stress that was in most cases effectively alleviated by antioxidant enzymes and proved by in situ visualization of H2O2 and ˙O2-. In pea seedlings, we observed a faster turn-over from anaerobic to aerobic metabolism proved by inhibition of alcohol dehydrogenase (ADH) activity. Additionally, reactive oxygen/nitrogen species contained in PAW did not affect the DNA integrity. On the other hand, the high level of DNA damage in barley together with the reduced root and shoot length and amylase activity was attributed to the oxidative stress caused by PAW, which was exhibited by the enhanced activity of guaiacol peroxidase or ADH. Our results show the glow discharge PAW at 1 min activation time as the most promising for pea. However, determining the beneficial type of PAW for barley requires further investigation.

Cold Atmospheric Pressure Plasma Treatment of Maize Grains-Induction of Growth, Enzyme Activities and Heat Shock Proteins.

Zea mays L. is one of the most produced crops, and there are still parts of the world where maize is the basic staple food. To improve agriculture, mankind always looks for new, better methods of growing crops, especially in the current changing climatic conditions. Cold atmospheric pressure plasma (CAPP) has already showed its potential to enhance the culturing of crops, but it still needs more research for safe implementation into agriculture. In this work, it was shown that short CAPP treatment of maize grains had a positive effect on the vitality of grains and young seedlings, which may be connected to stimulation of antioxidant and lytic enzyme activities by short CAPP treatment. However, the prolonged treatment had a negative impact on the germination, growth, and production indexes. CAPP treatment caused the increased expression of genes for heat shock proteins HSP101 and HSP70 in the first two days after sowing. Using comet assay it was observed that shorter treatment times (30-120 s) did not cause DNA damage. Surface diagnostics of plasma-treated grains showed that plasma increases the hydrophilicity of the surface but does not damage the chemical bonds on the surface.

The Effects of Cold Atmospheric Pressure Plasma on Germination Parameters, Enzyme Activities and Induction of DNA Damage in Barley.

Climate change, environmental pollution and pathogen resistance to available chemical agents are part of the problems that the food industry has to face in order to ensure healthy food for people and livestock. One of the promising solutions to these problems is the use of cold atmospheric pressure plasma (CAPP). Plasma is suitable for efficient surface decontamination of seeds and food products, germination enhancement and obtaining higher yields in agricultural production. However, the plasma effects vary due to plasma source, treatment conditions and seed type. In our study, we tried to find the proper conditions for treatment of barley grains by diffuse coplanar surface barrier discharge, in which positive effects of CAPP, such as enhanced germination or decontamination effects, would be maximized and harmful effects, such as oxidation and genotoxic potential, minimized. Besides germination parameters, we evaluated DNA damage and activities of various germination and antioxidant enzymes in barley seedlings. Plasma exposure resulted in changes in germination parameters and enzyme activities. Longer exposures had also genotoxic effects. As such, our findings indicate that appropriate plasma exposure conditions need to be carefully optimized in order to preserve germination, oxidation balance and genome stability, should CAPP be used in agricultural practice.

Evaluation of the Impact of Cold Atmospheric Pressure Plasma on Soybean Seed Germination.

The present study aims to define the effects of Cold Atmospheric Pressure Plasma (CAPP) exposure on seed germination of an agriculturally important crop, soybean. Seed treatment with lower doses of CAPP generated in ambient air and oxygen significantly increased the activity of succinate dehydrogenase (Krebs cycle enzyme), proving the switching of the germinating seed metabolism from anoxygenic to oxygenic. In these treatments, a positive effect on seed germination was documented (the percentage of germination increased by almost 20% compared to the untreated control), while the seed and seedling vigour was also positively affected. On the other hand, higher exposure times of CAPP generated in a nitrogen atmosphere significantly inhibited succinate dehydrogenase activity, but stimulated lactate and alcohol dehydrogenase activities, suggesting anoxygenic metabolism. It was also found that plasma exposure caused a slight increment in the level of primary DNA damage in ambient air- and oxygen-CAPP treatments, and more significant DNA damage was found in nitrogen-CAPP treatments. Although a higher level of DNA damage was also detected in the negative control (untreated seeds), this might be associated with the age of seeds followed by their lower germination capacity (with the germination percentage reaching only about 60%).

Silicon triggers sorghum root enzyme activities and inhibits the root cell colonization by Alternaria alternata.

MAIN CONCLUSION: Silicon inhibits the growth of Alternaria alternata into sorghum root cells by maintaining their integrity through stimulating biochemical defense reactions rather than by silica-based physical barrier creation. Although the ameliorating effect of silicon (Si) on plant resistance against fungal pathogens has been proven, the mechanism of its action needs to be better understood on a cellular level. The present study explores the effect of Si application in sorghum roots infected with fungus Alternaria alternata under controlled in vitro conditions. Detailed anatomical and cytological observations by both fluorescent and electron microscopy revealed that Si supplementation results in the inhibition of fungal hyphae growth into the protoplast of root cells. An approach of environmental scanning electron microscopy equipped with energy-dispersive X-ray spectroscopy enabling spatial detection of Si even at low concentrations showed that there is no continual solid layer of silica in the root cell walls of the rhizodermis, mesodermis and exodermis physically blocking the fungal growth into the protoplasts. Additionally, biochemical evidence suggests that Si speeds up the onset of activities of phenylpropanoid pathway enzymes phenylalanine ammonia lyase, peroxidases and polyphenol oxidases involved in phenolic compounds production and deposition to plant cell walls. In conclusion, Si alleviates the negative impact of A. alternata infection by limiting hyphae penetration through sorghum root cell walls into protoplasts, thus maintaining their structural and functional integrity. This might occur by triggering plant biochemical defense responses rather than by creating compact Si layer deposits.

Silicification of Root Tissues.

Silicon (Si) is not considered an essential element, however, its tissue concentration can exceed that of many essential elements in several evolutionary distant plant species. Roots take up Si using Si transporters and then translocate it to aboveground organs. In some plant species, root tissues are also places where a high accumulation of Si can be found. Three basic modes of Si deposition in roots have been identified so far: (1) impregnation of endodermal cell walls (e.g., in cereals, such as Triticum (wheat)); (2) formation of Si-aggregates associated with endodermal cell walls (in the Andropogoneae family, which includes Sorghum and Saccharum (sugarcane)); (3) formation of Si aggregates in "stegmata" cells, which form a sheath around sclerenchyma fibers e.g., in some palm species (Phoenix (date palm)). In addition to these three major and most studied modes of Si deposition in roots, there are also less-known locations, such as deposits in xylem cells and intercellular deposits. In our research, the ontogenesis of individual root cells that accumulate Si is discussed. The documented and expected roles of Si deposition in the root is outlined mostly as a reaction of plants to abiotic and biotic stresses.

Tobacco plants (Nicotiana benthamiana) were influenced by silicon and were not infected by dodder (Cuscuta europaea).

The effect of silicon (Si) on tobacco (Nicotiana benthamiana) development and dodder (Cuscuta europaea) - tobacco interaction were studied. Three Si application approaches were tested: tobacco seed priming (2.5 mM Si and 5 mM Si; 2.5S, 5S), watering tobacco plants with Si solution (2.5 mM Si and 5 mM Si; 2.5W, 5W) and foliar application (1 mM Si and 2.5 mM Si; 1F, 2.5F). Dodder was not able to infect the host plant in almost all Si treatments. Only in the control and 2.5W treatments was dodder able to infect its host. A significant increase in all observed antioxidant enzymes activities (POX, CAT and SOD) occurred in the plants of 2.5W treatment after infection in comparison with the uninfected 2.5W treatment and control plants, which indicated the importance of antioxidant enzymes activities in the plant parasite - host interaction. Resistance of Si treated plants to dodder could have been due to the changes in the cell wall properties of the epidermis and cortex where activity of POX was confirmed histochemically. The growth and development of tobacco shoots were evaluated after four and eight weeks of cultivation in the individual Si treatments. The development of shoots was enhanced after eight weeks of cultivation in the 2.5S, 5S, 2.5W and 5W treatments in comparison with the control treatment. However, a negative effect of Si was observed in 1F and 2.5F treatments. In the majority of cases, the plants treated with Si had decreased chlorophyll content when compared to control, except for chl a in 5W plants after 8 weeks of cultivation. Contrary to this, carotenoids increased in all Si treated plants after eight weeks cultivation in comparison with the control. The secondary xylem formation in tobacco was enhanced after 4 and 8 weeks cultivation in shoots of plants receiving the 2.5S, 5S, 2.5W and 5W treartments. The cambium was the most active in producing secondary xylem in the 2.5S treatment. Protein profile and antioxidant enzymes activities (POX, CAT and SOD) were altered by Si treatment. After 8 weeks of cultivation, activities of POX were significantly decreased in 2.5S, 5S, 2.5W and 5W in comparison with control. Catalase was decreased in 2.5S, 5S and 5W in comparison with the control, however, 1F and 2.5F treatments had significantly increased CAT and SOD activities. The specific activity of POX was confirmed histochemically in Si treated plants in the cell walls of several stem tissues like the epidermis, cortex and pith. A small amount of H2O2 was detected in leaves in the control and Si treated plants. The amount of O2- decreased in all treatments with time. The highest Si concentration in the plants (almost 800 mg . kg-1 d. w.) was detected in the 2.5W, 5W treatments.

Ionome and expression level of Si transporter genes (Lsi1, Lsi2, and Lsi6) affected by Zn and Si interaction in maize.

Zinc (Zn) is an essential microelement involved in various plant physiological processes. However, in excess, Zn becomes toxic and represents serious problem for plants resulting in Zn toxicity symptoms and decreasing biomass production. The effect of high Zn and its combination with silicon (Si) on ionome and expression level of ZmLsi genes was investigated in maize (Zea mays, L; hybrid Novania). Plants were cultivated hydroponically in different treatments: control (C), Zn (800 µM ZnSO4 · 7H2O), Si5 (5 mM of sodium silicate solution), and Si5 + Zn (combination of Zn and Si treatments). Growth of plants cultivated for 10 days was significantly inhibited in the presence of high Zn concentration and also by Zn and Si interaction in plants. Based on principal component analysis (PCA) and mineral element concentration in tissues, root ionome was significantly altered in both Zn and Si5 + Zn treatments in comparison to control. Mineral elements Mn, Fe, Ca, P, Mg, Ni, Co, and K significantly decreased, and Se increased in Zn and Si5 + Zn treatments. Shoot ionome was less affected than root ionome. Concentration of shoot Cu, Mn, and P decreased, and Mo increased in Zn and Si5 + Zn treatments. The PCA also revealed that the responsibility for ionome changes is mainly due to Zn exposure and also, but less, by Si application to Zn stressed plants. Expression level of Lsi1 and Lsi2 genes for the Si influx and efflux transporters was downregulated in roots after Si supply and even more downregulated by Zinc alone and also by Zn and Si interaction. Expression level of shoot Lsi6 gene was differently regulated in the first and second leaf. These results indicate negative effect of high Zn alone and also in interaction with Si on Lsi gene expression level and together with ionomic data, it was shown that homeostatic network of mineral elements was disrupted and caused negative alterations in mineral nutrition of young maize plants.

Cuscuta europaea plastid apparatus in various developmental stages: localization of THF1 protein.

It was generally accepted that Cuscuta europaea is mostly adapted to a parasitic lifestyle with no detectable levels of chlorophylls. We found out relatively high level of chlorophylls (Chls a+b) in young developmental stages of dodder. Significant lowering of Chls (a+b) content and increase of carotenoid concentration was typical only for ontogenetically more developed stages. Lower content of photosynthesis-related proteins involved in Chls biosynthesis and in photosystem formation as well as low photochemical activity of PSII indicate that photosynthesis is not the main activity of C. europaea plastids. Previously, it has been shown in other species that the Thylakoid Formation Protein 1 (THF1) is involved in thylakoid membrane differentiation, plant-fungal and plant-bacterial interactions and in sugar signaling with its preferential localization to plastids. Our immunofluorescence localization studies and analyses of haustorial plasma membrane fractions revealed that in addition to plastids, the THF1 protein localizes also to the plasma membrane and plasmodesmata in developing C. europaea haustorium, most abundantly in the digitate cells of the endophyte primordium. These results are supported by western blot analysis, documenting the highest levels of the THF1 protein in "get together" tissues of dodder and tobacco. Based on the fact that photosynthesis is not a typical process in the C. europaea haustorium and on the extra-plastidial localization pattern of the THF1, our data support rather other functions of this protein in the complex relationship between C. europaea and its host.