Month: November 2022

Our experimental results demonstrate that Cipro may interfere with the photosynthetic bio-energetic pathway, in addition to causing morphological deformities in higher plants. The range of EC50 values estimated from concentration-response models of the increase in inactive excited PSII Chl units and the decrease in root length are, respectively, 10.3-111.7 and 0.532-0.776 µM, both of which are higher than the respective LOEC values . Although these concentrations are higher than those typically found in surface waters , they are comparable with those found near industrial effluents . Therefore, the observed adverse in vivo response demonstrated here may be relevant to sensitive plants located near high-FQ-containing waters. Finally, the present study not only stresses the need, as pointed out previously , to consider sublethal molecular targets as potential meaningful phytotoxicity end points, but also takes a step toward the evaluation of FQ cumulative effects, a current challenge in the environmental risk assessment of pharmaceuticals , by providing correlations between FQ structures and specific molecular mechanisms of toxic action.Increasing population and consumption have raised concerns about the capability of agriculture in the provision of future food security.Recent estimates suggested that global agricultural production should increase by 70% to meet the food demands of a world populated with ca. 9.1 billion people in 2050. Food security is particularly concerning in developing countries, as production should double to provide sufficient food for their rapidly growing populations. Whether there are enough land and water resources to realize the production growth needed in the future has been the subject of several global-scale assessments. Te increase in crop production can be achieved through extensification and/or intensification. At the global scale, almost 90% of the gain in production is expected to be derived from improvement in the yield, but in developing countries,vertical plant rack land expansion would remain a significant contributor to the production growth.

Land suitability evaluations, yield gap analysis, and dynamic crop models have suggested that the sustainable intensification alone or in conjugation with land expansion could fulfil the society’s growing food needs in the future. Although the world as a whole is posited to produce enough food for the projected future population, this envisioned food security holds little promise for individual countries as there exist immense disparities between regions and countries in the availability of land and water resources, and the socio-economic development. Global Agro-Ecological Zone analysis suggests that there are vast acreages of suitable but unused land in the world that can potentially be exploited for crop production; however, these lands are distributed very unevenly across the globe with some regions, such as the Middle East and North Africa , deemed to have very little or no land for expansion. Likewise, globally available fresh water resources exceed current agricultural needs but due to their patchy distribution, an increasing number of countries, particularly in the MENA region, are experiencing severe water scarcity. Owing to these regional differences, location-specific analyses are necessary to examine if the available land and water resources in each country will surface the future food requirements of its nation, particularly if the country is still experiencing significant population growth.As a preeminent agricultural country in the MENA region, Iran has long been pursuing an ambitious plan to achieve food self- sufficiency. Iran’s self- sufficiency program for wheat started in 1990, but the low rate of production increase has never sustainably alleviated the need for grain imports. Currently, Iran’s agriculture supplies about 90% of the domestic food demands but at the cost of consuming 92% of the available freshwater. In rough terms, the net value of agricultural import is equal to 14% of Iran’s current oil export gross revenue. Located in a dry climatic zone, Iran is currently experiencing unprecedented water shortage problems which adversely, and in some cases irreversibly, affect the country’s economy, ecosystem functions, and lives of many people. Te mean annual precipitation is below 250 mm in about 70% of the country and only 3% of Iran, i.e. 4.7 million ha, receives above 500 mm yr−1 precipitation.

The geographical distribution of Iran’s croplands shows that the majority of Iran’s cropping activities take place in the west, northwest, and northern parts of the country where annual precipitation exceeds 250 mm . However, irrigated cropping is practiced in regions with precipitations as low as 200 mm year−1 , or even below 100 mm year−1 . To support agriculture, irrigated farming has been implemented unbridled, which has devastated the water scarcity problem. The increase in agricultural production has never been able to keep pace with raising demands propelled by a drastic population growth over the past few decades, leading to a negative net international trade of Iran in the agriculture sector with a declining trend in the near past . Although justified on geopolitical merits, Iran’s self-sufficiency agenda has remained an issue of controversy for both agro-ecological and economic reasons. Natural potentials and constraints for crop production need to be assessed to ensure both suitability and productivity of agricultural systems. However, the extents to which the land and water resources of Iran can meet the nation’s future food demand and simultaneously maintain environmental integrity is not well understood. With recent advancement in GIS technology and availability of geospatial soil and climate data, land suitability analysis now can be conducted to gain insight into the capability of land for agricultural activities at both regional and global scales. Land evaluation in Iran has been conducted only at local, small scales and based on the specific requirements of a few number of crops such wheat, rice and faba bean. However, there is no large scale, country-wide analysis quantifying the suitability of Iran’s land for agricultural use. Herein, we systematically evaluated the capacity of Iran’s land for agriculture based on the soil properties, topography, and climate conditions that are widely known for their relevance with agricultural suitability. Our main objectives were to: quantify and map the suitability of Iran’s land resources for cropping, and examine if further increase in production can be achieved through agriculture expansion and/or the redistribution of croplands without expansion. The analyses were carried out using a large number of geospatial datasets at very high spatial resolutions of 850m and 28m .

Our results will be useful for estimating Iran’s future food production capacity and hence have profound implications for the country’s food self-sufficiency program and international agricultural trade. Although the focus of this study is Iran, our approach is transferrable to other countries,what is a vertical farm especially to those in the MENA region that are facing similar challenges: providing domestic food to a rapidly growing population on a thirsty land.We classified Iran’s land into six suitability categories based on the soil, topography, and climate variables known to be important for practicing a profitable and sustainable agriculture. These suitability classes were unsuitable, very poor, poor, medium, good, and very good . This classification provides a relative measure for comparing potential crop yields across different lands. Four major land uses that were excluded from the suitability analysis comprised 19.3 million ha of Iran’s land , leaving 142.8 million ha available for agricultural capability evaluation .When land suitability was evaluated solely based on the soil and topographic constraints , 120 million ha of land was found to have a poor or lower suitability ranks . Lands with a medium suitability cover 17.2 million ha whilst high-quality lands do not exceed 5.8 million ha . Te spatial distribution of suitability classes shows that the vast majority of lands in the center, east and, southeast of Iran have a low potential for agriculture irrespective of water availability and other climate variables . As shown in Fig. 2, the potential agricultural productivity in these regions is mainly constrained by the low amount of organic carbon and high levels of sodium contents . Based on soil data, Iran’s soil is poor in organic matters with 67% of the land area estimated to have less than 1% OC. Saline soils, defined by FAO as soils with electrical conductivity >4 dS/m and pH<8.2, are common in 41 million ha of Iran. Although many plants are adversely affected by the saline soils , there are tolerant crops such as barley and sugar beet that can grow almost satisfactorily in soils with ECs as high as 20 dS/m, which was used as the upper limit of EC in this analysis .Although sodic soils are less abundant in Iran , soils that only have high ESP covers ~30 million ha . We used an ESP of 45% as the upper limit for cropping since tolerant crops such as sugar beet and olive can produce acceptable yield at such high ESP levels.As shown in Fig. 2, EC is not listed among the limiting factors, while we know soil salinity is a major issue for agriculture in Iran. This discrepancy can be explained by the higher prevalence of soils with ESP>45% compared to those with EC>20 dS/m, which can spatially mask saline soils. Tat is, the total area of soils with EC>20 dS/m was estimated to be about 6.4 million ha , while soils exceeding the ESP threshold of 45 constituted ~12 million ha i.e. almost double the size of saline soils. Iran’s high-quality lands for cropping are confined to a narrow strip along the Caspian Sea and few other provinces in the west and northwest .

In the latter provinces, the main agricultural limitations are caused by high altitude and steep slopes as these regions intersect with the two major mountain ranges in the north and west .Thus far, the land suitability analysis was based on soil and terrain conditions only, reflecting the potential agricultural productivity of Iran’s without including additional limitations imposed by the water availability and climatic variables. However, Iran is located in one of the driest areas of the world where water scarcity is recognized as the main constraint for agricultural production. Based on aridity index, our analysis showed that 98% of Iran could be classified as hyper-arid, arid, or semi-arid . August and January are the driest and wettest months in Iran, respectively, as shown in Fig. 3. Over half of the country experiences hyper-arid climate conditions for five successive months starting from June . This temporal pattern of aridity index has dire consequences for summer grown crops as the amount of available water and/or the rate of water uptake by the crop may not meet the atmospheric evaporative demand during these months, giving rise to very low yields or total crop failure. It must be noted that the high ratio of precipitation to potential evapotranspiration in humid regions could also result from low temperature rather than high precipitation. There is a high degree of overlap between regions that experience wetter conditions for most of the year and those identified as suitable for agriculture based on their soil and terrain conditions . This spatial overlap suggests that some of the land features relevant to cropping might be correlated with the climate parameters. In fact, soil organic carbon has been found to be positively correlated with precipitation in several studies. To incorporate climate variables into our land suitability analysis, we used monthly precipitation and PET as measures of both overall availability and temporal variability of water. We derived, from monthly precipitation and PET data, the length of the growing period across Iran . Growing period was defined as the number of consecutive months wherein precipitation exceeds half the PET. As shown in Fig. 3, areas where moisture conditions allow a prolonged growing period are predominately situated in the northern, northwestern, and western Iran with Gilan province exhibiting the longest growing period of 9 months. For over 50% of the lands in Iran, the length of the moist growing period is too short to support any cropping unless additional water is provided through irrigation . However, these areas, located in the central, eastern, and southeastern Iran, suffer from the shortage of surface and groundwater resources to support irrigated farming in a sustainable manner. Taking into account daily climate data and all types of locally available water resources can improve the accuracy of the length of growing period estimation.

Synergistic effects on plant growth under several conditions when PGPR and AMF are coinoculated are reported . In our study, however, the effects of combined inocculation with two PGPRs or with PGPRs and AMF never exceeded the sum of the effects of single inocculation with PGPRs and AMF. This indicates additive effects of PGPRs and AMF rather than clear synergistic effects. Roots of tomato after application of both bacteria strains were not only healthier but also showed a significantly higher colonization by AMF Glomus intraradices, indicating that AMF infection in the soils was suppressed directly by pathogens or indirectly as consequence of poor root development. Azcón and Linderman reported that unidentified PGPR have a strong stimulatory effect on the growth of AMF and increased mycelia growth of G. mosseae spores. P. fluorescens 92rk, alone or co-inoculated with P. fluorescent P190r, increased mycorrhizal colonization of tomato roots by G. mosseae BEG12 . Similarly to the results obtained by Marulanda-Aguirre et al. , where Bacillus megaterium inoculated with G. intraradices showed the highest percentage of mycorrhizal root length of Lactuca sativa plants compared to the single inoculation of G. intraradices. These results suggest that PGPR and AMF might be co-inoculated, at least in soils with a low AMF status,vertical hydroponic to optimize the formation and function of the mycorrhizal symbiosis. Both PGPR and AMF inoculation treatments directly and indirectly improved the nutrients acquisition and allocation to the shoots of tomato plants. The concentrations of P, Mn and Zn in tomato shoots were higher after inoculation with P. sp. ”Proradix®” and B. amyloliquefaciens FZB42 when compared to the untreated control. The ability of P. fluorescens and AMF to promote plant growth by improved nutrient acquisition and suppression of soil borne pathogens is well documented.

Both functions may promote plant growth but by different mechanisms. AMF facilitated mineral and water uptake, and increased the defense against soil borne pathogens . PGPRs induced the release of plant growth regulators . Siddique reported that Pseudomonas spp. can synthesize certain enzymes that can modulate plant hormone levels, might limit the available iron via siderophore production and can also kill pathogens by production of certain antibiotics. Our study confirmed that, B. amyloliquefaciens FZB42 can act as a PGPR, as described by De Freitas et al. , Kokalis-Burelle et al. , Kishore et al. and Marulanda-Aguirre et al. . Phae et al. reported that B. subtilis NB22 significantly reduce the occurrence of crown and root rot disease of tomato. Another aspect in the present study was to test if the mixtures of different bacteria species improve the control against FORL compared to one bacterium species alone. Our results did not confirm Pierson and Weller and Schisler et al. who proposed a strategy to increase the efficacy and the consistency of disease control by mixed application of antagonistic microorganisms with different modes of actions. Cordier et al. stated that dual or multiple inoculations of beneficial microorganisms can be neutral, positive or negative depending on the inoculants used. However, our study showed that combined application of two PGPRs improved tomato growth and suppressed FORL to the same extent as single application or further increases P shoot concentrations. Our results are in agreement with studies by Raupach et al. , Pierson and Weller and Duffy et al. , all of which demonstrated that certain mixtures of PGPR were significantly suppressive to cucumber pathogens and take-all disease. Different mechanisms of action for different PGPR strains may explain why combinations of P. sp. ”Proradix®” and Bacillus amyloliquefaciens FZB42 suppress disease similar to inoculation with single strains. Sung and Chung demonstrated that chitinase-producing Streptomyces spp. and B. cereus isolates used in combination with antibiotic-producing P. fluorescens and Burkholderia cepacia isolates had a synergistic effect on the suppression of rice sheath blight and Szczech and Dyśko who reported that among tested bacterial inoculations, only mixture of the bacteria B125 and PT42 tended to affect positively the growth of the plants and to reduce the density of Fusarium spp. in the rhizosphere of tomato plants.

These results indicate that the consistency of biocontrol agents in suppression of soil borne pathogens influenced by many factors, i.e. bacterial strains, soil borne pathogen species, species of plant, etc. Protein expression in plant systems has the potential to provide a safe, cost-effective, and scalable method to meet the increasing need for therapeutic protein production. Plant-based expression offers several advantages to the bio-pharmaceutical industry, including decreased cost of production, scalability, lack of susceptibility to mammalian pathogens, elimination of animal- or human-sourced raw materials, and the production of complex proteins with post-translational modifications such as N-glycosylation. For many therapeutic proteins, N-glycosylation is essential for protein folding, oligomerization, quality control, enzyme activity, ligand interactions, localization, and trafficking. Despite its potential, a possible barrier to the commercialization of plant-made glyco protein drugs is the difference between the N-glycan structures of human and plants. Of particular concern are plant-specific structures contained in complex type N-glycans, namely, α1,3 core fucose, β1,2 bisecting xylose, and the Lewis a epitope. Even though there is no definitive proof of adverse effects from plant-specific glycan structures, the presence of nonhuman glycans could potentially cause unwanted immunogenicity in humans, and the lack of sialic acid termination may lead to reduced blood circulatory half-life. A change in glycan structure could also potentially alter the protein’s structure or accessibility of its epitopes and, consequently, its function. Therefore, to ensure the efficacy of a plant-made bio-similar therapeutic, it is important that the N-glycans are compatible with both the protein’s function and the human immune system. Several strategies exist to modify a glyco protein’s N-glycan structures in planta, such as glycoengineering of the host cells using CRISPR/Cas9 genome editing to knock outβ-xylosyltransferase genes and α-fucosyl transferase genes and RNA interference technology to down regulate XylT and FucT genes, targeting of the protein to specific organelles, addition of compounds to alter the function of glycan-modifying enzymes, and in vitro glycan remodeling using chemoenzymatic reactions. In this work, we utilize kifunensine, a potent and highly specific inhibitor of α-mannosidase I in both plant and animal cells resulting in production of glycoproteins containing predominantly Man8GlcNAc2 and Man9GlcNAc2 structures, to a rice cell suspension culture grown in a bioreactor to inhibit α-mannosidase I activity. More than a few studies of kifunensine treatment in whole Nicotiana benthamiana plants successfully produced predominant Man9 structure glycoproteins; however, the study of kifunensine treatment in plant cell suspension cultures, including transgenic rice cell suspensions, is very limited.

In transgenic rice cell suspensions treated with 5 µM kifunensine cultivated in shake flasks, the productivity of a target glycoprotein, acid α-glucosidase , was significantly lower than the control, but the relative abundance of high-mannose structure GAA increased by 65% compared to the control. Here, we report the effects of kifunensine treatment on production and N-glycosylation of a glycoprotein conducted in a bioreactor. The culture media, method of cultivation, degree of glycosylation, and multi-merization of the product were different from the study by Choi et al.. Rice is generally recognized as safe by the FDA, and the rice alpha amylase 3D promoter, what is vertical farming a metabolically-regulated strong promoter, used in this study was well studied. In addition, semicontinuous bioreactor operations of transgenic rice cell suspensions proved the stability and robustness of transgenic rice cells under the RAmy3D promoter system for long-term recombinant protein production. In this study, we use a transgenic rice cell suspension culture to produce recombinant human butyrylcholinesterase , a bioscavenger hydrolase enzyme that can be used as a therapeutic and prophylactic treatment to counter organophosphorus nerve agents, as a model glycoprotein. Human BChE is a tetrameric glycoprotein with four identical 69-kDa monomers containing nine N-glycosylation sites per each monomer, with its activity, stability, and blood circulatory half-life highly dependent on the presence and structure of these glycans. The production of recombinant human BChE in transgenic rice cell suspension cultures is controlled by the RAmy3D promoter that is highly activated in the absence of sugar. Like other glycoproteins, N-glycosylation of a nascent rrBChE starts in the ER by co- or post-translational transfer of Glc3Man9GlcNAc2 from a dolichol lipid carrier onto Asn-X-Ser/Thr residues, where X is any amino acid except Pro. Since our rrBChE gene construct contains the RAmy3D signal peptide, a glycosylated rrBChE follows the secretory pathway for secretion, involving removal of glucose and mannose residues and addition of new sugar residues and N-acetylglucosamine , leading to complex-type N-glycans of rrBChE, as previously reported.Our goal in this study is to investigate the effects of kifunensine on N-glycosylation modification and the production of rrBChE in a transgenic rice cell culture bioreactor. By adding kifunensine to the medium during bioreactor cultivation at the end of the growth phase and throughout the induction phase, we demonstrate the production and N-glycosylation pattern of rrBChE in culture medium and within the cell aggregates .As shown in Figure 2a, the overall production level of active rrBChE increased by up to 80 µg/g FW, at least 1.5-fold higher than recently reported, using the same two-stage cultures. Table 2 also shows a significant improvement in the volumetric productivity and specific productivity . The increase in total active rrBChE, volumetric productivity, and specific productivity may be the result of increasing sugar-free medium concentration by 1.25 times and decreasing the bioreactor working volume during the media exchange by 1.25 times.Our hypothesis is that the rate of mannose trimming by ER and/or Golgi α-mannosidase I is faster than the rate of diffusion of kifunensine from cell aggregates to individual cells and from individual cells to Golgi and ER compartments.Although the concentration of rrBChE of 7.5 mg/L in the transgenic rice cell suspension culture in this study was significantly lower than the concentration of recombinant BChE of 1–5 g/L from the milk of transgenic goats, it is important to consider the long development time required between gene transfer and lactation for transgenic goats. There is limited quantitative information on production costs for rBChE from the milk of transgenic goats. However, since mammalian production systems can harbor and propagate human pathogens, regulatory requirements for the characterization of the transgenic founder, as well as feeding, housing, health monitoring, genetic stability assessment, and regulated disposal of ex-producer animals, could increase the production costs of human therapeutics in transgenic animals compared with plant-based systems. The expression vector design, cloning, transformation and selection of the callus, and media components were previously described. In brief, rice calli derived from Oryza sativa cv. Taipei 309 embryo/scutellum were co-cultured with Agrobacterium tumefaciens containing the binary vector with the RAmy3D promoter, codon-optimized human BChE gene to express in rice, the RAmy3D signal peptide, and the RAmy3D terminator. After eight rounds of screening starting with more than 300 transformation events, a stable transgenic rice cell line “9-2” was established, which was previously used in other studies as well as this study. The inoculum cultures were grown in 250 mL sugar-rich media in 1 L-shake flasks for 6 days in an Innova 4000 incubator/shaker at 140 rpm and 27 ◦C in the dark.Combined shake flasks described in Section 3.1 were inoculated at about 20% v/vin a 5 L stirred-tank bioreactor equipped with a pitched blade impeller and containing 2.5 L of sterile NB + S added through the head plate inoculation port inside a biosafety cabinet. Bioreactor conditions were controlled at 27 ◦C, 75 rpmagitation speed, 40% dissolved oxygen of air saturation , and 0.2 vvm of the overall mixed gas flow rate. The oxygen uptake rate was measured by the change in DO level in the culture when aeration is stopped but with continued agitation. The culture pH was monitored but not controlled. The cultures grown in the glass bioreactor were exposed to ambient light. Freshly prepared kifunensine solution was added to the bioreactor at 5 µM final concentration in the bioreactor medium at day 6 of cultivation, 24 h before the media exchange. At day 7 of cultivation, media exchange was performed to replace spent sugar-rich medium with sugar-free medium using the same method as previously described. Kifunensine solution was added at day 0 , day 2, and day 4 following induction, assuming 0 µM of kifunensine in the bulk medium prior to each kifunensine addition.

We identify key the data gaps that need to be filled in order to proceed with meaningful ecological risk assessments, whether they are more global/ regional in nature, or for site specific assessments. Finally, we attempt to draw conclusions from the literature about the relative sensitivity of different model organisms, as well as the importance of particle properties on fate, transport and effects.Nanoceria is used in electronic and optical devices, polishing agents for glass and of silicon wafers, exterior paints, metallurgy, and diesel fuel additives.Additionally, nanoceria is used in automotive catalytic converters.It is also used in catalysts in petroleum refining, in the fluid catalytic cracking process . Based on the amount of total global CeO2 annual production and global nanoceria production rates,roughly 15–25% of total CeO2 production is nano . Cerium oxide is used in these applications in both nano and non-nano form and quantitative estimates of cerium oxide use within specific applications do not distinguish between nanoceria and its bulk counterpart.There are few studies that quantify the release of engineered nanomaterials during use, and even less nanoceria specific studies. One of the few studies by Park et al., indicates that 6–100% of CeO2 will be released during the use phase of diesel fuel additives.This has not yet been validated by other researchers. In laboratory conditions, particles filters from diesel cars removed 99.9% of Ce present in fuel additives.However the manuscript does not specify whether the Ce additive was in the nanoscale. Considering the applications and the likelihood that the nanomaterials are released to the environment, the following assumptions were made. For example,vertical growers nanoceria in batteries is enclosed within a protective casing, which is likely to minimize release during use.

If the batteries are disposed of improperly, the most likely environmental compartment would be soil, with negligible release to air, water or wastewater treatment plants . Similar assumptions were made for metallurgical products, catalysts in FCC, polishing powders used in industry , and other applications. Experimental studies have been conducted to measure the release of various manufactured nanoparticles from surface paints on exterior facades. Kaegi et al. measured concentrations as high as 600 μg L−1 of nano-TiO2 in runoff from newly-painted building facades,and estimated that as much as 30% of nano-Ag is released from surface paints within a year of paint application.However, no data exist on nanoceria released from paints. Based on similar information, estimated nanoceria concentrations in treated WWTP effluent discharged to waterbodies are expected to be in the range of 0.003–1 μg L−1 . In biosolids, nanoceria concentrations are expected to be around 0.53–9.10 mg kg−1 .These estimated concentrations are expected to increase as nanoceria is used more widely, and there will likely be accumulation of CeO2 in soils and sediments, further increasing exposure concentrations in these media.The detection and characterization of nanoceria under conditions relevant to environmental, toxicological and biological systems remains a challenging, and frequently impossible, task. However, there is little or nothing that is ceria-specific, but applies to all nanomaterials. However, aspects of characterization are included here since it is fundamental to understanding of all nanomaterials, including nanoceria. Essential general aspects are listed below: i) In environmental systems, the specific and accurate detection and characterization of manufactured nanoceria remains essentially impossible,due to the gap between metrology and analysis and the complexity of the system . Total Ce detection is useful as it acts an upper limit of nano-ceria concentrations for risk assessment, but is not synonymous with manufactured nanoceria.

The discussion below applies primarily to spiked materials, mainly in the laboratory or mesocosm. ii) As with other nanomaterials, nanoceria should be fully characterized using suitable preparation methods and a multi-method metrological approach. In a multi-method approach, independent techniques operating on independent measuring principles provide cross-validation of measured properties. The source of the nanomaterial also needs to be fully reported, given the likely effects on properties. Fuller discussion is given elsewhere.iii) A number of properties require characterization which can be grouped as size, shape, morphology, aggregation/ agglomeration, surface charge and dissolution . These groups, or classes, contain several individual properties. For instance, for size, an average size should be reported, along with some measure of spread .iv) Given the changes that are well known to occur upon storage or changing media,it is essential to perform appropriate measurement over temporal and spatial scales which adequately capture the dynamics of the nanomaterial system. Although, none of the points above are ceria specific, nanoceria is capable of oxygen storage, which is size and shape dependent.Nanoceria is generally thought to have low solubility in water,although this is size and oxidation state dependent. Where dissolution and solubility are low, study is rendered simpler because dissolved ions should have little impact on toxicity. However, recent work has shown potential effects of even low level dissolution.Nano-ceria has two stable oxidation states IJCeIJIII) and Ce) under environmental conditions and cerium has the ability to transition readily between these two states.This redox activity gives nanoceria some of its key properties.However, oxidation state and morphology are usually poorly controlled or defined and spatially variable within an individual particle,giving rise to poorly reproducible data and uncertainties in understanding toxicity or exposure. These uncertainties, along with dynamic changes that occur in complex media, could explain the variable environmental and toxicological results that are seen in the literature for nanoceria.Table 1 shows a non-definitive selection of studies of nanoceria in a variety of different environmental, toxicological and standard complex media. These studies are examples of some of the most complete characterization in the literature, although there is still little consistency between studies and it is often not clear which nanomaterial properties require analysis because it is not well understood how each affects biological or environmental processes.

Lastly, because of logistical or other constraints, characterization is often not performed as fully as necessary to interpret such processes. Some of the most powerful techniques for the visualization of nanoparticles are transmission electron microscopy , atomic force microscopy and scanning electron microscopy . These techniques not only provide direct visual images but can be used to quantify other properties such aggregation, dispersion, sorption, size, structure and shape of the nanoparticles,although the sample preparation may alter considerably the sample. These techniques have been extensively applied to nanoceria, occasionally in complex media. Van Hoecke et al.and Rodea-Palomares et al. used TEM to visualize the interaction between the nanoceria and algal cells in order to test whether the nanoparticles are taken up or adsorbed by the algal cell wall. Zhang et al.used TEM to further investigate the needle like clusters on the epidermis and in the intercellular spaces of cucumber roots after treatment with nanoceria over 21 days. In some cases, TEM has been coupled with spectroscopy, for instance TEM coupled with EDS was used to determine the elemental composition of ceria clusters on both the root epidermis and in the intercellular regions of the cucumber plant.Merrifield et al.used AFM to image and quantify the size of PVP-coated nanoceria while compared them using TEM and DLS in toxicology exposure media. TEM confirmed that the larger particles are aggregates composed of smaller individual particles , but that nanoceria properties did not measurably change in the exposure media tested. In the same study, EELS was used to quantify the oxidation states showing that the smallest and the largest samples were composed of entirely CeIJIII),vertical grow with only small amounts of Ce present in the largest sample. Such spectroscopy is essential to microscopy imaging in complex media and is required to unambiguously identify the nanoparticles of interest in the presence of materials with similar sizes, shapes and electron densities/tip interactions. Microscopy, although a powerful single particle method, remains challenging when attempting to provide statistically meaningful measurements. Much data reported in the literature is pictorial and non-quantitative; careful design and timeconsuming analysis are required to be able to determine representative parameters with confidence. Nanoparticle tracking analysis is another widely used characterization technique which utilises microscopy to determine size distributions and number concentration of nanoparticles in liquid samples. NTA has been infrequently used for nanoceria, for instance to determine the mean size of nanoceria in green alga and crustaceans and to better understand the effect of natural organic matter on the particle-size distribution of nanoceria settling in model fresh water as a function of time.However, the methodology has some limitations in complex and realistic media.X-ray photoelectron spectroscopy has been used in only one relevant study, to our knowledge, in this case to understand the antioxidant capacity of nanoceria to DNA. The calculation of CeIJIII) : CeIJIV) ratios was performed,in an analogous manner to EELS, within a multi-method approach. Similarly, synchrotron-based X-ray spectroscopy has been used in several studies to assess Ce speciation. Studies using micro X-ray fluorescence coupled with X-ray absorption near edge structure in natural matrices have been conducted concluding that nanoceria can undergo biotransformations within a matrix, so the modifications, the mechanism and extent of these transformations should be fully addressed.Scanning transmission X-ray microscopy is an analytical microscopy which, with extended X-ray absorption fine structure spectroscopy, provided 2D quantitative maps of chemical species at concentrations which are environmentally relevant.

X-ray microscopy can in principle provide a spatial resolution down to ~30 nm while imaging the specimen in the aqueous state without the need for sample preparation.Synchrotron-based techniques provide direct structural information regarding the nanoparticles and their interaction with the environment.It is clear that X-ray spectroscopy, XPS and EELS are complementary methods for oxidation state analysis and combination may prove fruitful. Field flow fractionation has also been used on nanoceria to measure the size distribution of nanoceria in synthesized samples as well as to understand the aggregation behavior of other nanoparticles in the presence and absence of humic substances.ICP-MS can be used as a detector for FFF, but has not been for environmental or toxicological studies of nano-ceria, to our knowledge. Preliminary studies have shown the feasibility of ICP-MS for nanoceria analysis in single particle mode, although its further application in real systems has yet to be demonstrated. Infrared spectroscopy has also been used to study biotransformations in plants by comparing the molecular environment of the sample before and after exposure hence concluding that cerium speciation changes after incubation of nanoceria in different exposure media over 21 days. Ultraviolet-visible spectroscopy has been used to monitor the dynamic aggregation process of nanoceria in various waters with time along with DLS and TEM.Nanoparticles properties are altered by the water chemistry such as pH, ionic strength, nature of electrolytes or presence of NOM. One of the most important changes may be aggregation of nanoparticles: between the same nanoparticle, homoaggregation, or between nanoparticles and an environmental particle, heteroaggregation. The increase in size of the aggregates affects their transport, behavior, reactivity, uptake by organisms, and toxicity. In pure water, the stability of non-coated nanoparticles in solution depends on their surface charge. Nanoparticles brought into close contact via Brownian diffusion processes will repel each other if the charge is strong enough to overcome attractive forces. Nanoceria surface charge is dependent of the pH; nanoceria are positively charged at low pH, negatively charged at high pH and have an isoelectric point at approximately pH 8.The methods of synthesis and the cleanup of nanoceria have been shown to play a role in affecting the experimental point of zero net charge for nanoceria suspensions, which range from 6.5 to 8.1.Differences in the reported PZC may also come from differences in the method applied to determine the PZC, the order of titration process, and sorption of anions used in the titration.These authors measured the aggregation kinetic of nanoceria and compared to the theoretical prediction of Derjaguin–Landau–Verwey– Overbeek . At pH 11, the experimental critical coagulation concentration was higher for the monovalent , than the divalent salts, 80 mM and 16 mM respectively. They showed that DLVO theory could predict quite well the stability of nanoceria at this pH. However, this model fails to explain aggregation behavior as solution conditions become more environmentally relevant and non- DLVO forces may also play important roles between particles.In a water-saturated column packed with sand, water composition has also been shown to control the transport and deposition of nanoceria.

Reducing the “need” to fish by increasing available aquaculture derived food and materials does not necessarily equal reduced fishing pressure. Political/economic input will likely be needed to achieve the conservation goal of reduced fishing pressure as an outcome of implementing a FFP project.I would argue that new research paths in algae culture for bio-fuel production need to be followed in order to achieve results for application in the near-term. Maximum solar conversion efficiency are likely not necessary for commercial success with algae culture for bio-fuels— particularly in the case of food and fuel poly culture systems. A focus on simple easily applied culture techniques may yield a similar cost/benefit ratio per unit invested compared to employing the latest intensive technology to achieve maximum yield. Research and development efforts should be refocused toward small to medium scale ideas and projects with an emphasis on applied, relatively low-tech systems. The overwhelming majority of past and current work pertaining to algae culture for bio-fuel production is concentrated on microalgae. Microalgae holds great promises and this work should continue, but active work with macroalgae for bio-fuel production should be initiated and receive at minimum a similar level of effort. Screening of potential macroalgae candidate species for bio-fuel production, and developing technology transfer from current commercial algae farming operations for use in FFP systems are strongly recommended starting points. The concept of food and fuel poly culture is in its infancy, but with moderate investments in research to further develop the system’s components,greenhouse vertical farming the concept can play a role in providing food and energy to the world’s population while at the same time helping to conserve valuable natural resources— particularly those in at risk aquatic ecosystems.The rotation of the Earth acts to confer regular environmental changes in light availability and temperature and plants alter their physiology, biochemistry, and metabolism in response to these abiotic cues over the course of a day .

In addition, the inherent regularity of the transition between day and night also allows alterations in temperature and light to be predictive of subsequent abiotic stresses. For example, dusk is typically accompanied by a decrease in temperature and possible frost. It is therefore unsurprising that plants have developed an internal timing mechanism that allows prescient alterations in gene expression and biochemistry. Indeed, the circadian clock causes the regular oscillation of between 30% and 40% of genes in the model plant Arabidopsis thaliana , even when grown under constant light and temperature . These broad changes in gene expression precipitate a range of physiological responses including the regulation of hypocotyl growth , alterations to plant hormone production and sensitivity and time of flowering . In concert, such changes promote the fitness of plants grown in synchrony between endogenous and environmental cues . Circadian clocks are conceptually thought to be comprised of three parts: a central oscillator typically consisting of a negative feedback loop, input pathways to allow entrainment to local environmental conditions, and output pathways which act to modulate responses dependent on these endogenous cues. Although at its most basic level a circadian clock can consist of a single negative feedback loop with input and output pathways , evolution has typically led to the development of multiple interconnected molecular oscillators with varied levels of redundancy . The inclusion of partially redundant interlocking components likely allows greater flexibility in the modulation of clock inputs during evolution while also allowing greater accuracy of the timing mechanism itself . In addition to these core concepts, circadian clocks are recognised as having additional properties including temperature compensation and “gating”. Temperature compensation allows a circadian clock to oscillate at approximately the same frequency over a broad range of physiological temperatures while gating refers to the regulated sensitivity of the central oscillator to input stimuli. This latter mechanism enables the circadian clock to persist in plants grown in constant experimental conditions by reducing the responsiveness of core components to light during the subjective night .The other dominant zeitgeber of the Arabidopsis circadian clock is temperature, with steps as small as 4°C being capable of entraining the clock mechanism . The majority of large-scale mutant screens to identify Arabidopsis clock genes have used light as an entrainment signal and it is therefore unsurprising that comparatively little is known about temperature-sensitive entrainment of the clock.

It does appear, however, that the circadian regulation of individual genes may differ based upon the entrainment conditions used; CAB2 and TOC1 expression are similarly modulated by light and temperature whereas the phase of CAT3 expression is more sensitive to changes in temperature than light . Temperature inputs into the clock are at least in part incorporated via loops containing PRR7 and PRR9 as a prr7 prr9 double mutant is unresponsive to circadian phase changes induced by temperature and is arrhythmic if entrained to temperature steps . In contrast, it appears that TOC1 has a minor role in this mechanism as a toc1 mutant retains a wild-type entrainment response to temperature steps . Further characterization of this sensitivity is dependent upon identification of temperature sensors in Arabidopsis.Animal circadian clocks have long been recognized to contain a master clock which synchronizes multiple “slave” clocks in other tissues . This hierarchical arrangement of the clock permits individual tissues to utilize subsets of circadian genes . In comparison, plants are thought to measure time using cell-autonomous circadian oscillators , although it has remained unclear until recently whether each of these independent plant clocks sharecommon core components across different cell types. It now appears that certain loops of the plant clock act predominantly in certain tissues. PRR3 has been shown to be predominantly expressed in Arabidopsis vasculature while recent microarray analysis has indicated that only a subset of genes known to have a circadian expression pattern in aerial tissues oscillate in hydroponically grown roots . Such data suggest that circadian rhythmicity in roots is controlled by a simplified mechanism and is dramatic evidence that plant circadian rhythms need not be controlled by a uniform set of components. In support of this concept, experiments using RNAi to reduce PRR3 mRNA levels induce a greatly pronounced shortening of the circadian clock when measured using vasculature-specific luciferase reporter constructs in comparison to those with a broader range of expression . The use of modified clock circuitry in different plant tissues likely allows altered sensitivity to environmental stimuli and stresses and it will be interesting in the future to determine the functional role of tissue-specific circadian oscillations.Our understanding of the Arabidopsis circadian clock at a transcriptional level has progressed rapidly, aided by the high-throughput capabilities of luciferase reporter mutant screens and micro-array assays. While it is clear that a large percentage of the transcriptome is regulated by the circadian clock , our understanding of the molecular processes underlying these large-scale changes in transcription remains limited. Recent work has identified a correlation between histone acetylation and transcriptional activity at the TOC1 locus and similarly, a degree of histone methylation is associated with changes in transcriptional activity at CCA1 and LHY loci . Both of these observations suggest that epigenetic marks may regulate circadian gene expression and identification of proteins responsible for these epigenetic marks will allow a more thorough understanding of transcriptional regulation by the clock. While transcriptional regulation is clearly important for Arabidopsis clock function,vertical agriculture it is also increasingly clear that the rhythms generated by the transcriptional clock are modulated by a range of post translational modifications. TOC1 and PRR proteins are differentially phosphorylated and degraded over the course of a day while ZTL protein accumulates with a circadian rhythm despite being transcribed at a regular rate .

It is equally apparent that endogenous rhythms are regulated by changes in cytosolic composition, such as the concentration of free Ca2+ . Considering these examples, it is unlikely that we have either identified all components of the Arabidopsis clock or that we yet fully understand the subtleties of action and regulation of characterized transcripts and proteins. Ultimately, it will be important to transfer our understanding of the clock to real-world applications. Given the suggested role of the circadian clock in the regulation of plant responses to abiotic stresses , it is possible that altering expression of certain clock components may confer enhanced stress tolerance. Indeed, Arabidopsis prr5 prr7 prr9 triple mutants have recently been reported to have an enhanced cold, drought, and salt tolerance, caused by increased expression of stress-responsive genes . Such data are in agreement with work demonstrating that plants are differentially responsive to temperature over the course of a day and that this gating is controlled by the circadian clock . Further work to understand the process by which stress response pathways and the circadian clock interact will likely be a fruitful course of investigation.The biological oxidation of ammonia is carried out by two groups of chemotrophic bacteria , a process termed ‘nitrification’, results in transformation of NH4 + to NO3 – , making soil nitrogen susceptible to losses through multiple pathways – leaching and gaseous losses . Due to these nitrogen losses, a major portion of the soil nitrogen and applied fertilizer nitrogen is lost, in low nitrogen-use efficiency of agricultural systems . Also, nitrogen pollution is the single most environmental concern from agricultural systems, contaminating ground and surface water . Denitrification of nitrate-N is the major source of nitrous oxide emissions from agricultural systems and contributes significantly to global warming . Blocking the function of nitrifying bacteria or slowing the nitrifiers’ function can significantly reduce nitrogen losses associated with nitrification and extend the persistence of nitrogen as ammonium in the soil for uptake by plants, lead to improved nitrogen recovery and – use efficiency in agricultural systems . Recently, it was shown that some plant species have the ability to release nitrification inhibitors from roots that suppress nitrifiers’ function, a phenomenon termed ‘biological nitrification inhibition’ . Using a luminescent recombinant Nitrosomonas, an assay has been developed to detect and quantify this inhibitory effect in plant soil systems . With this newly developed methodology, it was shown that several tropical grasses and certain field crops including sorghum possessed the ability to release nitrification inhibitors from roots, termed BNI-capacity . We showed that sorghum roots release methyl 3-propionate , one of the active constituents with BNI activity of root exudates from sorghum . Here we present our findings on further characterization of BNI function in sorghum and reveal the identity of the two nitrification inhibitors released from its roots. In addition, we present evidence to show wide genetic variability in the release capacity of one of the major nitrification inhibitors from sorghum roots and also report preliminary field-based results for the existence of BNI-capacity in wild sorghum. Root architecture influences nutrient and water uptake, anchorage, and mechanical support, interactions with microbes, and responses to various abiotic stress factors . Since water and mineral supply are often limited in the soil, a plant with a more extensive root system exhibits higher performance with regard to the tolerance of drought and poor nutrient conditions . Several factors, including root angle, root growth rate, and root types, influence root architecture . Root growth requires the successive formation of new cells from stem cells in the root apical meristem , and the progeny of such stem cells divide rapidly and enter the elongation/differentiation zone . To maintain root meristem activity, the rates of cell division and differentiation have to be coordinated . In addition, the interaction between cytokinin and auxin determines the size of the RAM through the regulation of the genes involved in auxin signaling and/or transport to ensure an appropriate auxin gradient . The rice root system consists of one seminal root, numerous adventitious roots, and lateral roots that emerge from the other two types . Lateral roots are the major components involved in the absorption of nutrients and in interactions with the surrounding soil environment . Lateral root formation represents a complex developmental process modulated by several hormones, including auxin and ethylene.Well defined and closely coordinated cell division activities give rise to lateral root primordia.While lateral roots originate from pericycle cells adjacent to xylem poles in Arabidopsis,pericycle and endodermal cells located near phloem poles are the origins of lateral roots in rice and maize . Their development is initiated by the asymmetric division of the pericycle cells, and subsequent divisions result in the formation of dome-shaped, multilayered, lateral root primordia .

The resilience of these dynamics directly depends on the globalization of food through trade, including the amount of food traded, the number of links describing trade between countries, and the topological properties of the trade network. Using reconstructions of food production and trade based on FAO data , this analysis shows that in the past few decades the system has become increasingly vulnerable to instability as an effect of demographic growth, dietary shifts, and the increasing inter connectedness of the trade network. Indeed, some nodes are starting to show the first episodic signs of instability, particularly in water-poor and trade-dependent countries . This analysis of the long-term response to shocks is in agreement with the short-term propagation of perturbations in the trade network during a food crisis ; both approaches have shown how the fragility of the coupled food-demographic system has increased as an effect of the growing globalization of food through trade. If trade and interconnectedness have the effect of reducing the resilience of the system as a whole, it is paramount to investigate to what extent it would be possible to globalize without becoming more vulnerable. A possible solution of this problem is suggested by ecological systems, which often exhibit some degree of modularity.There is evidence that systems with a modular structure—that is, with groups of countries that interact more among themselves than with countries from other modules—are able to contain the spread of perturbations within the targeted module, whereas the other modules remain only marginally impacted . In other words, the modules act directly to buffer the propagation of shocks to other communities, thereby increasing the stability and resilience of the entire system . Interestingly,tower garden the virtual water trade network exhibits a growing degree of modularity with a ratio between internal and external fluxes that is approaching 70% . To date, however, the effect of modularity still needs to be investigated in the context of the resilience of trade and food security.

An alternative approach to understand the impact of virtual water trade on population growth is through carrying capacity plots , which distinguish different strategies and their success by showing the historical evolution of a region or population’s local food supply potential and net food imports relative to their local and post trade carrying capacity. Porkka and collaborators confirm that food import is the strategy nearly universally used to overcome local limits to growth. Nevertheless, they also highlight that these strategies are implemented to varying extent and with varying success . Therefore, whether dependency on imports is necessary and desirable, a clear policy priority at both local and global scale is needed and it ideally would attempt to keep the demand of food under control .There is wide agreement that humanity’s rate of resource use exceeds what can be sustainably generated and absorbed by Earth’s systems . Substantial uncertainty persists for an apparently basic question—by how much is food demand likely to grow in the coming decades—with estimates typically ranging from a required 60% to 110% increase by the year 2050 over circa year 2005 levels . More recently, Hunter et al. estimated that an increase in cereals production of 25–75% over 2014 levels could be sufficient to satisfy projected demand in 2050. The breadth of future GHG emissions trajectories—and the magnitude of their cascading consequences for agricultural productivity—leaves considerable unknowns regarding future food production under a changing climate . A radical transformation of the global food system is likely required in order to increase production while faced with the considerable uncertainties related to demand and climate impacts. New strategies for achieving FEW security worldwide may benefit from adoption of an integrated approach aimed at an improvement in the availability, access, and nutritional properties of food while enhancing the provision of affordable, clean, and reliable energy . Moreover, a secure FEW system will incorporate a sustainable use of natural resources, maintain environmental stream flows, and restore ecosystem services.

The FEW system will need to invert the ongoing trend of increasing vulnerability and enhance its resilience with respect to climate shocks, demographic growth, and consumption trends. The previous sections have highlighted the existence of several major challenges in this multi-objective strategy to food, water, and energy security. For instance, the sustainability of energy production can be improved by increasing the reliance on renewable energy sources, which would decrease the rate of fossil fuel depletion, reduce CO2 emissions, and consequently allow societies to improve their ability to meet climate change targets. Renewable energy sources based on bio-fuels, however, would claim huge volumes of water and large expanses of land, thereby inevitably competing with the food system . Moreover, bio-fuel production often entails direct and indirect land use change and associated GHG emissions, indicating that in the short term these energy sources might have a negative impact on the environment . Nonfuel-based renewable energy production may also require substantial amounts of water and therefore compete with food crops in water-stressed regions . The increasing demand for food and energy by the growing and increasingly affluent human population can hardly be met with the limited land and water resources of the planet unless we transform the FEW system. As noted in the previous sections, approaches focusing on ways to increase food and energy production instead of curbing the demand would ignore the existence of limits to growth imposed by the natural resources the planet can provide and are likely to achieve higher production rates at huge environmental costs , resilience losses , and increased food insecurity for the poor. To be sure, there are still margins for increased production through improvements in efficiency, technological innovation, and agroecologically efficient farming systems, but these measures are likely to be insufficient to meet long-term global food and energy security needs . There is the need for a complete rethinking of the FEW system to develop a comprehensive strategy for food, water, and energy security, based on enhancing the production and moderating the demand . Although a conclusive answer to the question of how to sustainably meet the food, energy and water needs of the rising and increasingly demanding human population is still missing, here we review a number of new and old approaches and ideas that could contribute to future food, water, and energy security.

Such approaches can be, in general, technological , cultural , or institutional .Large yield gaps, or the difference, between current and attainable yields still exist in many parts of the world, particularly in sub-Saharan Africa and offer the potential to increase global production of major crops by as much as 58% under currently available technologies and management practices . There is broad consensus that efforts to enhance crop yields on currently cultivated lands are crucial for avoiding additional agricultural expansion , the consequences of which would be profound and undesirable for natural systems and functioning . Agricultural intensification, however, is not free of environmental impacts in that it contributes to GHG emissions, freshwater and coastal water pollution, depletion of freshwater resources, and consequent loss of aquatic habitat . In light of the environmental impacts of conventional intensification, some scientists are advocating for an approach to food security that relies on a “sustainable intensification” of agriculture , which aims to close the yield gap while minimizing the environmental impacts . Moreover, as with historical yield trends, stacking flower pot tower harvest frequencies have generally increased through time, but many places with the potential to transition to double-cropping systems have yet to do so . On the one hand, yield and harvest gap closure offers great promise for increasing the food self-sufficiency of many developing nations because the areas with the largest potential for production increases are those places that currently rely heavily on food imports and have some of the highest rates of projected population growth to mid century . On the other hand, these remaining yield gaps raise questions about how best to promote the diffusion of high-yielding crop varieties and agricultural technologies, given that these agricultural advancements have yet to reach many places even 50 years after the start of the green revolution . Moreover, it is unclear whether additional inputs are adequately available in low-yield areas and, if so, how to avoid their unsustainable use . Thus, particularly with regard to non-mobile resources, such as land and water, it will likely be essential to ensure that increases in production occur in places where and when natural resources can support it . There are social, economic, and institutional factors that need to be accounted for while advocating for agricultural intensification versus alternative farming approaches . Most of the existing literature on this subject has recognized the pros of yield gap closure as an alternative to agricultural expansion at the expense of natural ecosystems , particularly in the tropics . “Land sparing” can, in many contexts, minimize habitat losses, land degradation, CO2 emissions, and declines in biodiversity . This approach, however, is not a panacea because its profound social impacts have often been overlooked by focusing on agrotechnological solutions without considering their effect on production systems, such as smallholder farmers . Intensification efforts require investments that are increasingly made by large-scale agribusiness corporations, particularly in the developing world. Such investments may affect the system of production and its inputs, for example, through contract farming or out grower schemes or land use, access, and tenure rights, as occurs for LSLAs , which are discussed in section 9.3. Agricultural intensification is most effective in countries where relatively large yield gaps still exist, such as sub-Saharan Africa, while ensuring that new fertilizer and water are used in the most efficient way possible . To boost crop yields, investment in modern agricultural technology is required, which many rural communities in lower-income countries cannot afford. If neither local land users nor domestic investors are able to improve crop yields, in years of increasing crop prices, foreign corporations or foreign-domestic joint ventures are not likely to miss the profit opportunities existing in under performing agricultural land .

Indeed, recent years have seen a wave of investments in agricultural land in the developing world, with import-dependent nations seeking to increase the pool of land and water resources under their control and targeted countries pursuing avenues to promote rural development and agricultural technology transfers . However, there is a growing body of scientific and anecdotal evidence showing that the development and food security goals of these land deals are often not achieved and, instead, often bring substantial social and environmental consequences . Such land deals ultimately may result in the displacement of subsistence or small-holder farmers by large-scale commercial agriculture, as well as the development of new agricultural land at the expense of savannas, forests, or other ecosystems . Because most of the cultivated land worldwide is managed by small-scale farmers , this ongoing shift in systems of production may strongly reshape the agrarian landscape around the world with important impacts on rural livelihoods because it increases the dependence on a volatile food market. Thus, agricultural intensification may be the result of important transformations in land tenure, farming systems, and livelihoods. Developing countries may enhance crop yields by introducing modern agricultural technology while promoting greater efficiency in food production through a transition in their agricultural sector toward commercial-scale farming. Commercial agriculture lends itself better to capital inputs from investors and could result from LSLAs or other forms of investment, such as contract farming , or mixed out grower schemes . However, there could be negative impacts on rural communities and their livelihoods because LSLAs may turn farmers into employees and increase their vulnerability to food price volatility The transition to large-scale farming, however, might be unnecessary: small-scale farms, which account for most of the global calorie and nutrient production , can be very productive. There is evidence in the economic literature about the inverse relationship between farm size and productivity, meaning that smallholder farmers, when provided with adequate inputs, may often achieve yields that outperform commercial, large-scale agriculture. Identifying mechanisms that support yield enhancements, technology transfers, and secure land tenure to these critical stakeholders is a key component of advancing global food security, promoting poverty alleviation, and enhancing food system resilience. Overall, intensification typically requires the introduction of modern green revolution technology in areas of the developing world in which relatively large yield gaps exist.

Water consumption for food production, including crops and livestock, accounts for about 86% of the total societal water consumption, though, locally, household and industrial uses can be predominant, particularly in major urban areas. Thus, securing water resources for agriculture, while reconciling the competing water needs of growing cities and surrounding rural areas, is a major challenge of our time. Climate change is expected to further enhance local water scarcity, especially in the subtropics. In fact, while climate warming is slightly increasing global precipitation, the global patterns of rainfall distribution are expected to become more uneven with an intensification of aridity in the dry subtropics, and an increase in precipitation in the wet tropics and the midlatitude temperate zone . The temporal variability of precipitation will likely increase, thereby enhancing the probability of drought and flood occurrences .Despite recent developments in desalinization technology , 2016, most human activities related to food and energy production rely on the consumptive use of freshwater. Desalinization remains limited to specific uses that require relatively small amounts of water and to societies that can sustain the associated costs . The freshwater available for human activities is stored in continental land masses either in soils or in surface-water bodies and groundwater aquifers. Often referred to as “green water,” soil moisture is retained in the ground by capillary forces and can be extracted only when it is subjected to a suction that overcomes the action of capillarity. Plants exert such suction through root uptake. Although most of terrestrial vegetation in natural ecosystems relies on green water , soil moisture remains for most part unavailable to direct human use because it is difficult to extract. In contrast, water stored in surface water bodies and aquifers, referred to as “blue water,” hydroponic net pots is more mobile and contributes to surface-water and groundwater runoff. Thus, green water leaves land masses in the water vapor phase as evapotranspiration , whereas blue water flows to the ocean in the liquid phase as runoff .

Since antiquity, human societies have engineered systems to withdraw blue water from rivers, lakes, and aquifers and have transported it through channels and pipes to meet the needs of a variety of human activities. Today, the main consumptive use of blue water is for irrigation , which strongly increases green water flows at the expense of blue water flows. Irrigation is a major human disruption of the water cycle ; indeed, many rivers are so strongly depleted that they no longer reach the ocean , while lakes in basins with internal drainage are drying out . Irrigation can modify the local climate, possibly by increasing evapotranspiration and effectively cooling the near-surface atmosphere . Irrigation may also moderately enhance precipitation downwind of irrigated areas and induce mesoscale circulations driven by the contrast between irrigated areas and the surrounding dry lands . It has been estimated that globally, irrigation uses a water volume that is roughly 2.56 × 1012 m3 /year , which accounts for about 2% of the precipitation . Although water is a renewable resource that is conserved in the Earth system, freshwater stocks can be depleted when their use exceeds the rates of natural replenishment. A typical example is groundwater that is often used for agriculture andis being depleted in many regions of the world , including the North American Southwest, Northern Africa, the Arabian Peninsula, and India . In some cases, groundwater use is depleting water stocks that accumulated in epochs with a wetter climate. In these aquifers “over pumping” leads to a permanent extraction of water resources, a phenomenon that is known as “groundwater mining” to better stress its unsustainability and the irreversible loss of resources that will not be available to future generations. However, even when the depletion of water resources is reversible, its environmental impacts may not be. Excessive water withdrawals from rivers and streams destroy the aquatic habitat and lead to extinction of riparian species. Interestingly, freshwater ecosystems are particularly vulnerable because the extinction rate of freshwater aquatic species is much greater than that of terrestrial organisms . Thus, sustainable use of water resources should prevent not only their permanent depletion but also the irreversible damage of downstream ecosystems.

A rich body of literature has discussed criteria to define minimum flow requirements and minimum flow variability required to conserve the aquatic habitat . A reevaluation of those efforts within the context of water sustainability has led to the formulation of the concepts of “planetary boundaries” and “safe operating space” that define a cap for sustainable water use . Such a cap is typically expressed as a fraction of the natural river flow, ranging from 20% to 60% , though recent studies have suggested referring to season-dependent fractions . Although globally, the current use of water for irrigation is smaller than the planetary boundary for blue water and accounts for only 5.4% of the global blue water flows , in many regions of the world those boundaries are locally exceeded, thereby causing habitat loss . Overall, irrigation is critical to sustaining the present rates of agricultural production. Although only 20% of the global agricultural land is irrigated , it sustains about 40% of the global crop production owing to the typically much higher yields in irrigated systems . Collectively, irrigated and rainfed agriculture accounts for about 10% of global precipitation over land, with green water flows from agroecosystems contributing to roughly 16% of the global evapotranspiration from terrestrial ecosystems . These figures give us a sense of the proportion of the water cycle that has been appropriated by agriculture. Moreover, other economic activities, such as mining, manufacturing, and energy production further increase the human demand for freshwater.Since the onset of civilization, agriculture has claimed land from natural ecosystems, such as forests, savannas, and grasslands. By converting these landscapes into agricultural land, humankind has profoundly altered the water and bio-geochemical cycles . Decades of research on deforestation have highlighted the profound hydroclimatic impacts of land use and land cover change . Compared to forests, rainfed farmland sustains lower evapotranspiration rates because of the smaller leaf area index, surface roughness, and root depth, and the greater albedo . The infiltration rates are also smaller because agricultural soils are often more compacted, typically from leaving the land fallow for part of the year and cultivated with heavy machinery. Smaller evapotranspiration and infiltration rates are expected to lead to higher runoff . However, in areas where agriculture is irrigated, water withdrawals for crop production deplete surface-water bodies and aquifers . Land use change also has an impact on the regional climate.

Land use change alters the surface energy balance and land-atmosphere interaction; these changes modify near-surface temperature, boundary layer stability, and the triggering of convection and convective precipitation . Some of these effects can alter the rainfall regime within the same region in which land cover change occurs, though it has been suggested that the impact also can be on adjacent ecosystems . Moreover, land cover change may modify the rate of emission of biological aerosols, thereby affecting cloud microphysics and cloud processes . The reduced evapotranspiration has the effect of reducing precipitation recycling, which is the fraction of regional precipitation contributed by atmospheric moisture from regional evapotransporation , a phenomenon that is relevant to policies and therefore is receiving the attention of social scientists , despite the great uncertainties with which it can be evaluated . Overall, forest or woodland conversion to cropland over large regions is expected to reduce precipitation and increase diurnal temperatures , though these effects depend on the size of the cleared area . The direct and indirect impacts of human activities on freshwater resources may strongly affect their availability to meet the competing needs of food or energy production and the environment,blueberry grow pot raising questions on the type of institutional arrangements that could improve water governance.Water is by its own nature fluid, renewable, and difficult to quantify , and its biophysical characteristics, such as the fact that it is a key input into biological processes and that is relatively plentiful and widely distributed , make the political economy of this resource very different from other similarly important strategic natural resources . From early human history, water use has led to complex dynamics of competition and cooperation . In a world with increasing societal pressure over scarce water resources and aggravating hydro climatic change, water governance is fundamental in the policy and development dimensions of water management. Even though access to safe water and sanitation is recognized as one of the UN-SDGs , about 4 billion people face water scarcity at least 1 month per year . Water availability may be affected by water quality, particularly in the case of drinking water, as the cost of treatment may become prohibitive in some locations, creating physical water scarcity of costly water resources. The reliance on water markets historically has been, and still is, strongly influenced by neoliberal governance approaches based on privatization, liberalization, and extension of property rights. The core principle behind these approaches is that water markets provide the correct economic incentives to promote the reallocation of water to higher valued uses and improve efficiency. These approaches treat water as a commodity and thus require the recognition of property rights that define the use, management, and trade of water resources . Easter et al. describe a strong legal system as the main institutional condition necessary for water markets to function properly. The creation of water markets in the Western United States and in Chile have been used as exemplary policy and governance models that could be exported and promoted in developing countries .

Since the 1980s, the World Bank has been the main promoter of water markets in developing countries, while also supporting the development of lucrative transnational opportunities in the water sector for private investors . In the context of the FEW nexus debate, however, a water market economy may lead to water resources previously used to produce food being transferred to other uses, such as industrial production or household needs in urban areas. In fact, the economic yield of food commodities may typically be orders of magnitude lower than that of the energy and water utility sectors . Food as a basic human need means that market approaches to water governance also can be evaluated in the context of their impacts on food security, particularly for the poor. For example, water markets could be structured with special consideration for certain industries, including agriculture, to avoid losing water allocations for production of food. A counterargument in favor of water markets stresses their positive environmental outcomes such as when water is partly acquired to reestablish environmental flows and improve aquatic habitat, or if the market sets a cap on the amount of water that can be withdrawn for human uses . The contemporary neoliberal trends of water commodification, that is, the multidimensional process through which goods that traditionally are not priced enter the world of money and markets , could be in stark contrast with the principle that access to water is a fundamental human right . Ostrom describes water resources as an iconic example of common-pool resources, which often have been successfully governed through diverse community and communal-property institutional arrangements. The multiple characterizations of freshwater by different cultures and societies make it difficult for freshwater to be reduced to a monetized commodity. Water can be perceived as a sacred commodity, a human right , a political good, an ecosystem medium, and a security asset . Moreover, the water sector has intrinsic characteristics that can be associated with structural market failures, with large externalities, and interconnectedness that make the level of individual and collective interdependence particularly critical . From a social and environmental justice perspective, the idea that water is not treated as a “common good” but as a commodity has generated criticism around the perpetuation of inequality and violation of fundamental human rights . Different narratives and political perceptions about the value, the meaning, and the function of water in society make a clear and uniform definition of “good water governance” difficult. As described by Meinzen-Dick , rather than considering single solutions for water governance, it may be more productive to have multiple institutions work together in an adaptive learning process.

However, if this is indeed the case, there must be a mechanism and a third component in the system which causes the inhibition of the diffusion, and which, by extension, exerts a pressure that opposes the surface tension and the external pressure, along with the internal pressure. The third component is suspected to be the hydroxide ion, which is always present in aqueous solutions, and which is detected around collapsing microbubbles, and has already been applied to wastewater treatment. This ion tends to aggregate around the nanobubble surface, and is suspected to be present in the form of a cloud of ions around the bubble, attracted to the surface by an as yet unconfirmed force, but widely thought to be physical bonds of the nature of van der Waal’s force, and plays a part in the inhibition of gas diffusion out of the bubble. The exact mechanism, distribution of the ions, the extent to which they inhibit the diffusion, and other concerns regarding their roles in the mechanism of stabilization is not yet determined, but several theories have been proposed as to their role, and more also exist which do not take into account their role, or do not require them to play any role in the process at all. The role of the ions is suspected to be due to the repulsion of the ions toward each other, which in some way opposes the external pressure and the surface tension, but this is yet to be confirmed. Thus, while there are several approaches to the question, as of recent efforts it still remains unresolved. Several theoretical approaches have been proposed, many of which are highly specific to the circumstances for which the study was conducted, and none thus far have proposed an overarching theory as to the formation and evolution of bulk nanobubbles. As far back as 1997 Ljunggren and co-workers proposed theoretical explanations for colloid-sized gas bubbles based on diffusion of the gas into the liquid,fodder system for sale which could now be considered nanobubbles. Seddon et. al. also contributed to the emerging idea around the same time, but there have been few contributions to understanding their stable presence since then.

Explanations for specific cases of phenomena such as surface nanobubbles, nanobubbles generated electrochemically, and so forth have been offered so far. Early on, the Young Laplace equation was used to describe nanobubble stability, but the internal pressures required are far higher than would be possible at ambient temperature for the amount of gas that is contained within the bubble. Attard and co-workers analysed the thermodynamic stability of bulk nanobubbles, but it was found that the radius of nanobubbles could not be accurately predicted from thermodynamic considerations, nor was an expression offered for the rate of decrease in nanobubble size. Brenner and Lohse presented a model for predicting the radius of surface bubbles based on the dynamic equilibrium between diffusion into and out of nanobubbles situated at a surface. Further work in specific cases by Weijs and Lohse suggested the use of increased length scales to counter the problem of high internal pressures due to the relatively high surface tension of a bubble in that size range. Sverdrup and colleagues offered explanations as to the rates of decrease in size based on diffusion in all directions possible through the gas-water interface at the nanobubble surface. In their models they consider the possibility of diffusion both into and out of the nanobubble, with a sufficiently high mass transfer coefficient. Their models consist of a combination of Henry’s Law and Taylor series expansion. The equations are plotted, taking time as a function of radius and show coherence with previous models given by Ljunggren. However, no comparisons with experimental data are provided. The Young-Laplace equation seems inadequate to completely describe the phenomenon as it requires extremely high internal pressures of the gas to balance the surface tension that causes the nanobubble to shrink, as summarized by Attard and coworkers. However, the interface through which the diffusion occurs has thus far been considered to have constant properties of being composed only of water molecules and gas molecules.

Yasui and colleagues also detail several theories that attempt to explain bulk nanobubble stability, based on the armoured bubble model, a particle crevice theory, a skin theory, the dynamic equilibrium model and electrostatic repulsion. Among these theories, it appears that electrostatic repulsion has the most experimental support. Studies of interfaces between water and practically all surfaces such as glass are negatively charged, assumed to be due to the accumulation of hydroxide ions physisorbed to the monolayer as reported by Zangi and Engberts. Thus, it is reasonable to suppose that the water-gas interface is also negatively charged due to similar congregation of hydroxide ions at the bubble surface. Furthermore, studies conducted by Takahashi and others have shown that nanobubbles are indeed negatively charged, with oxygen nanobubbles having a zeta potential about -35 mV. Thus, it is evident that hydroxide ions physisorbed onto the surface of the nanobubble play a role in the interactions between the molecules present there. Jin et. al. proposed a model for bulk nanobubble stability involving the electrostatic repulsion, terming the pressure due to the electrostatic force as Maxwell pressure. One rationale involving the surface charge density of a bulk nanobubble has been proposed by Ahmed and colleagues that involves electrostatic repulsion balancing the surface tension. In the following, we consider a theory of electrostatic repulsion and what it requires of the conditions imposed for nanobubbles to have the long-term stability that has been observed experimentally. Several applications have been discovered, such as for wastewater treatment, fish farming, shrimp breeding, and hydroponics. These are further substantiated by Agarwal and coworkers, for such specific issues as the disinfection of infected surfaces, the degradation of organic compounds, and the disinfection of the water itself. The effects of increased yield of fish due to higher dissolved oxygen content are summarised by Endo et al..

The usage of hydrogen nanobubbles in gasoline to improve the calorific yield is also reported by Oha et al.. Other projected uses include the use of nanobubbles as contrast agents for the ultrasound imaging of tumours, as reported by Cai and co-workers, as well as reduction and removal of deposits of calcium oxalate, which is similar to the composition of kidney stones in rat kidneys, as presented by Hirose et al. Another application of the nanobubble’s ability to permit salts to crystallize is the design of self-cleaning membranes for desalination of water,fodder growing system which use nanobubbles as electrically conductive spacers and pass current through them to force the salts to crystallize on the nanobubble surface, which will permit easy removal of the accumulated salts. This was demonstrated and presented by Abida et al. The pressure balance of the nanobubble is considered to be given by the Young-Laplace equation, which, as explained above, equates the internal pressure, external pressure and the surface tension. The first of the four forces that we consider in the Young-Laplace equation is internal pressure. It is proportional to the surface area of the nanobubble, and is assigned a positive sign since it acts to increase surface area. The second is the external pressure, given by the hydrostatic pressure acting on the surface of the bubble, which also decreases the surface area and is negative. The third is the surface tension, which acts along the surface area at the molecular level. The surface tension acts to decrease the surface area, hence the radius and size, and can also be assigned the negative sign. However, a fourth force which is thought to be integral to the stability is the electrostatic repulsion between hydroxide ions adsorbed to the surface of the nanobubble, or, possibly in the cloud surrounding the surface. This repulsion seeks to reduce the contact between the ions on the surface of the bubbles, which also acts to increase the distance between the ions, thus increasing the surface area, and therefore results in a positive pressure. The nature of the interaction between ions can be characterized by the expression for Coulombic repulsion. Since one hydroxide ion is of the order of 1 nanometre in diameter, and most nanobubbles are two orders of magnitude greater in size, we can ignore the curvature of the distance between them and take it to be linear. The repulsion should, in theory, affect all neighbouring hydroxide ions, but is assumed to be insignificant beyond the nearest neighbours. We also assume the spatial arrangement of these ions over the surface to be close-packed in nature, since the repulsion is equal in all directions, and they would ideally assume a close-packed formation. This arrangement of ions is shown schematically below, in Fig. 1a, and as shown in Fig 1b it is assumed, due to close-packing, that they assume the formation of a rhomboidal unit cell, of side and diagonal length denoted by x, which will be referred to subsequently as the inter-ionic distance. That the nanobubble shrinks due to outward diffusion of the gas contained within is, of course, undisputed, but the precise methods and the rate of diffusion are highly debated. Previous theoretical studies have always assumed a model with a higher mass transfer coefficient, or longer time scales for the process to account for the reduced rate and the high lifetime of the nanobubble.

However, it is reasonable to suggest that the change in the rate of diffusion can be attributed to two things: the velocity due to the Brownian motion of the nanobubble, and the inhibition of the diffusion due to the adsorbed hydroxide ions on the surface. In this chapter, the possible effects of Brownian motion are examined for the effect on the rate of diffusion that they may possess. Earlier studies have shown that nanobubbles can be formed by supersaturation, where the solubility limit of the gas, when surpassed will permit the gas to precipitate and form bulk nanobubbles as reported by Matsuki and co-workers . The shrinkage of nanobubbles has so far been thought to be governed by Fick’s Laws, since it is a case of how fast the gas can dissolve into the surrounding fluid. Thus, according to the first law, it must be directly proportional to the outward gas flux, but the constant is still the diffusion constant D0 for the diffusion of the gas into water. However, this only holds true where the surface area of the nanobubble remains constant. It is, however, possible, that the outward diffusion is a case of Fick’s second law, since the surface area that is available to the gas to diffuse outward also changes according to size, and that this surface area determines the rate of shrinkage and thus the lifetime of the bulk nanobubble. It is then reasonable to suppose that the cause of the change of surface area available for diffusion is the change in the surface area occupied by hydroxide ions combined with the decreasing radius of the bulk nanobubble. The rationale for the assumption that the hydroxide ions adhere to and are released the nanobubble surface is based on two observations, as mentioned before. Firstly, the observation that all interfaces formed by water are negatively charged, and we consider nanobubbles to be a special case of a gas-water interface which may be charged in the same way. Secondly, the zeta potentials measured for nanobubbles are all negative, indicating that a negative ion present in pure water is responsible for the negative charge, which by elimination is the hydroxide ion. Further observations also indicate higher negativepotentials for more electronegative gases, such as oxygen and nitrogen, than for other reported gases such as argon and xenon as reported by Ushikubo et. al.. That nano- and microbubbles release hydroxide ions as they shrink is a well-known phenomenon. The stabilization and the shrinkage can be considered to be related to the same phenomenon; thus, the ideal case can be taken to be a nanobubble that is newly formed with no hydroxide ions at the surface at the instant of its formation of an interface. Here, the hydroxide ions present in the water immediately surrounding the bubble, in the hydrodynamic layer, adhere almost instantaneously, the time taken for the adsorption to occur being too small in comparison to the overall timescale to be important.

Plants are the primary producers on earth, assimilating carbon dioxide by daytime photosynthesis for the bio-genesis of all essential structures.This carbon assimilate is partitioned primarily into sugars and starch in the autotrophic ‘sources’ with a portion of the sugars allocated to the heterotrophic ‘sinks’ to support growth of the latter.In the absence of photo assimilation, the starch stored in the source is degraded to replenish cellular sugars in order to avoid carbon starvation.Therefore, carbon assimilation and utilization is carefully balanced for optimal plant development.Adverse environmental conditions can disrupt the normal starch and sugars levels with repercussions for the ability of the plant to sustain growth.Drought is associated with reduced starch or sugar levels in source Thissues.Salinity stress can induce higher starch accumulation in the source or sink of some species, but trigger starch reduction in others.Similarly, chilling stress is associated with accelerated source-starch accumulation or degradation.These observed increases in starch or sugars may be adaptive responses for stress-survival, or may be ‘injury’ responses resulting from the under-utilization of carbon because of growth cessation, regardless, documenting these changes is necessary for a deeper understanding of plant stress response.Feeding plants with 14CO2 is useful for tracking carbon movement, and can inform on changes in carbon allocation due to stress.Salinity increased flux from source to developing fruits in tomato and to the roots in transgenic rice seedlings.Water-stress elicited a similar distribution pattern in Arabidopsis, with higher 14C allocated to the roots,in beans, where 14C flux to the pods increased, dutch buckets and in rice, where it stimulated 14C mobilization from the stem and allocation to the grain.

Additional 14C-allocation studies under varied stress conditions could help to clarify whether or not higher source-sink flux is a universal stress response.Te observed changes in local and distant carbon fuxes in plant Thissues during stress result from multiple activities – epigenetic, transcriptional, post-transcriptional and post translational changes, occurring across different spatial and temporal scales, which must be integrated to deliver a cohesive response to stress.Te trehalose-6-phosphate/Sucrose non-Fermented Related Kinase 1signaling cascade may function in this way.It is critical for plant survival under low carbon and energy conditions, in part through changes in starch metabolism.Te T6P/SnRK1 can also modulate source-sink interactions; therefore, key elements of this regulatory network could potentially be activated for a ‘rewiring’ of whole plant carbohydrate use under stress.Because of the many issues with respect to plant carbon use under stress that remain unresolved, our aim in this work was to investigate changes in carbon partitioning and allocation in response to short-term drought, salinity, and cold stresses.14CO2-labeling of a single source leaf was used to map whole-plant and intra-Thissue changes in carbon use, as it can provide partitioning and allocation data in the same system.Single-leaf labeling permits more accurate tracking of 14C-movement than can be obtained by exposing the entire rosette to the label.By comparing plants exposed to different stresses it may be possible to identify convergent and divergent adaptive responses associated with each unfavorable condition.Starch content was also assayed in the source leaf and the roots of the stressed plants and the data were compared to 14C-starch fuxes to identify how starch metabolism may be regulated to alter sugar distribution.Finally, the transcriptional activity of key genes in the T6P/ SnRK1 pathway was assessed to identify genes associated with changes in carbohydrate levels under abiotic stress.By integrating these data, we present one of the first comprehensive pictures of how Arabidopsis changes carbon flux under short-term environmental stress.This information could be combined with that generated from the wealth of -omics data to broaden our understanding of plant stress response.

Our first aim was to investigate how plant source and sink Thissues use carbon over the diurnal cycle under normal conditions.One hour before the middle of the day , a single mature, but still developing source leaf was fed with 14CO2 for 5 min.Te labeled source leaf, unlabeled sink leaves, and the roots were harvested separately at MD, at the end of the day , and at the end of the night.MD, ED and EN correspond to 6h, 12h and 24h after dawn.Te percentage of 14C distributed among the source and the sinks was determined.Within each Thissue, the incorporation of 14C into the main metabolites pools: sugars, amino acids, organic acids, starch, protein, and ‘remaining insoluble compounds’ , was established.First, we calculated the percentage of 14C distributed from the source to the sinks.During the day, ~60% of the 14C was retained in the source leaf, but by EN, the percentage of total 14C was evenly distributed among all Thissues.Nighttime export of 14C from the source, and its subsequent allocation into the sinks, accounted for the re-distribution.Second, we examined the 14C partitioning between the source and sinks to create a full picture of how allocation and subsequent partitioning were altered.Partitioning in the roots was more dynamic than in the sink leaves, and this difference was amplified most at ED.In the roots, there was increased incorporation of 14C into metabolites used for growth — i.e.sugars, amino acids, and RICs — and less into those used for storage —i.e.protein and starch — compared to the source.Te pattern of 14C-partitioning in source leaf vs.roots therefore reflected the prioritization of biological processes in each Thissue type.Te other change of note occurred at EN, when both sinks incorporated less 14C into organic acids but more into starch compared to the source.This may indicate that the sinks had greater sufficiency with respect to carbon with a relatively reduced need for organic acids as sources of energy compared to the source.Finally, we examined changes in 14C-partitioning over the diurnal cycle.Data at ED and EN were compared to that generated at MD to fully assess how the day-night cycle affected carbon partitioning in different Thissues.Te metabolic pools in the source leaf were variable, while those in the sinks were relatively stable.Relative to MD, there was less 14C in the sugar and starch fraction, but an almost 2-fold greater flux into organic acids at EN in the source.

Organic acids may serve as the primary substrate for respiration after reductions in the sugar pool.In the roots, at EN, the 14C percentage in sugars decreased, but increased in starch.This indicates that the starch in the roots was accumulated constantly during the diurnal cycle, with more accretion during the night than the day.In contrast, in the sink leaves, the carbon flow into sugars and starch were stable at EN, but there was a 4-fold increase in the 14C partitioned into the RICs, suggestive of nighttime growth processes.Carbon allocation was negatively affected by osmotic stress, and the inhibition grew in severity as the stress progressed.By EN, mild and severemannitol stress increased the percentage of 14C in the source, and decreased it in the roots.This could reflect reduced carbon export due to enhanced source activities, inhibited carbon export from the source, reduced sink strength, or a combination thereof under osmotic stress.Carbon partitioning within the source was also modulated to a greater extent than in the sinks.At MD, both mild and severe osmotic stress reduced the 14C-partitioned into starch but increased 14C-partitioning into organic acids in the source, presumably for respiratory use.Six hours later, only severe osmotic stress had this effect leading to greater 14C flux into osmoprotectants — sugars, organic acids, and amino acids — at the expense of the storage compounds.Te 14C-fux into these osmoprotectants also increased in both sinks at the expense of the RICs, with the latter decreasing drastically in the roots.Te most obvious change was the percentage of 14C allocated from source leaf into roots, which decreased significantly by EN under both mild and severe NaCl stress.Te 14C-use in source leaf was more responsive to salinity compared to the sinks.Severe salinity stress decreased 14C-partitioning into starch but increased partitioning into sugars, amino acids,grow bucket and organic acids during the day in the source.At MD, more 14C was partitioned into sugars in the sink leaves, but 6h later at ED the 14C in sugars was stable, with reduced fux into starch and proteins.This indicates that 12h after the stress treatment, carbon was diverted from storage and preferentially partitioned into sugars for osmoprotection.In the roots, less 14C was partitioned into the RICs at ED and EN compared to the control, which suggest a shift away from investing 14C into resources normally used for root growth under salinity.This may have led to increased 14C accumulation into sugars at the end of night because they were under-metabolized.Interestingly, proteins were the only metabolite affected by both mild and severe salinity stress in both source and sink leaves, while it was unchanged in the roots.Carbon fux into this pool decreased compared to the control at ED in both source and sink leaves.Further, unlike sink leaves, the source had increased 14C label in protein at MD.Te changes in 14C partitioning and allocation in response to different levels of salinity stress are summarized as follows:the source leaf partitioned less 14C into storage compounds but more 14C into osmoprotectantsin response to severe salinity stress;sink This issues showed a differential response to salinity stress: similar to the source leaf, the sink leaves showed reduced 14C in storage compounds, however, roots This issue had reduced 14C in structural compounds; and the amount of 14C imported into roots This issue was inhibited by salinity; this might be due to reduced sink activity, inhibited phloem transport, or a combination there of.

The percentage of 14C in root This issues was significantly reduced by cold stress at the end of night, showing similarity to This issues under osmotic and salinity stress.Carbon allocation was not affected by low temperature during the day , but carbon partitioning was highly regulated in the source leaf , especially at the end of day.Te most notable difference was that the 14C-fux into starch and RICs decreased relative to the control plants.Te decrease in starch was high at MD but lessened during the diurnal cycle, while the opposite was true for the RICs, where inhibition intensified over the day.In the source, there were also higher fluxes into sugars, amino acids, and organic acids from MD to ED.Cold also triggered increased 14C into the protein pool at MD, and decreased it at ED.At EN, the 14C in RICs strongly decreased, with a corresponding strong increase in sugars.Cold stress therefore stimulated more 14C partitioning into sugars over the diurnal cycle in the source leaf.Te sinks were less affected by cold than the source.In sink leaves, there was increased carbon fow into sugars during the day and decreased carbon into starch at night, with no difference in RICs.In contrast, the roots had increased 14C in the sugar pool at night, and reduced partitioning into the RICs.This change of 14C partitioning suggests reprioritization of reserves with a greater fux towards sugars for osmoprotection at the expense of other pathways.Te 14CO2 labeling experiment showed that starch is the most dynamic metabolite pool that changed under all types of abiotic stresses used in this study.14C-fux into starch was down-regulated by abiotic stress, and the regulation depended on the time of day and This issue type examined.Under control conditions, 14C-partitioning into starch was stable during the day but decreased at night in the source leaf.However, this pattern was disrupted under salinity and cold stress due to reduced carbon flow into starch.In contrast to the source leaf, 14C in starch in sink leaves did not change during the day even under stress.In roots, the percentage of 14C into starch normally increased by EN, and interestingly, this partitioning was maintained under osmotic stress, but not under salinity and cold stress.Source and sink This issues therefore partitioned carbon into starch differently in response to abiotic stress.Source leaf showed reduced 14C partitioning into starch at MD under both mild and severe salinity and osmotic stress, and at ED under severe stress only.Sink This issues had very little changes of 14C in starch under stress.Since the plants were labeled 5 h after the stress treatment, the 14C flux into starch cannot provide a whole picture of starch metabolism changes during the entire stress period.It only informs on percentage change in partitioning and allocation.Therefore, absolute starch content measurements were made in source leaf and roots , to determine changes in accumulation over the time-course.

Salinity stress is a major abiotic stress that affects plant growth, resulting in a loss of crop yield, especially rice, which is one of the most salt-sensitive plants in comparison to other cereals.Salt stress affects plants via both osmotic and ionic effects.Osmotic effects result in a reduction of water absorption ability such that the effects are similar to drought stress.Ionic stress causes Na+ toxicity, which disrupts photosynthesis, protein synthesis, and enzyme activity.Numerous reports have shown negative effects of salt stress on rice growth and productivity based on the total chlorophyll content, protein concentration, photosynthetic CO2 fixation, stomatal conductance, transpiration, shoot dry weight, tiller number per plant, spikelets per panicle, and grain yield.Ca2+ is a crucial second messenger consisting of a transient elevation of cytosolic [Ca2+].The Ca2+ signals are transduced and decoded via Ca2+ binding protein, and then the information is relayed to downstream responses.The signals are mainly transduced through kinases mediating the phosphorylation cascade, resulting in downstream response regulation, including changes in gene expression through the regulation of transcription factors.Calcium signaling is used to respond to environmental stimuli, as well as to coordinate growth and development in plants.In the plant calcium signal transduction process, calcium sensors, including calmodulin ,hydroponic nft channel calcineurin B-likeprotein and Ca2+-dependent protein kinase , play important roles in the transduction of various stimuli.CaM is a protein that contains characteristic EF-hand motifs that bind Ca2+ ions with high affinity and specificity.CaM binding to Ca2+ leads to the exposure of hydrophobic regions on the molecule surface and subsequent interactions with target proteins or nucleic acids.

Rice carries 5 CaM-encoding genes: OsCam1–1, OsCam1–2, OsCam1–3, OsCam2 and OsCam3.The expression of OsCam1–1 increases to a great extent in response to NaCl, mannitol and wounding treatment.Several lines of evidence have revealed that calcium sensors are involved with an enhanced abiotic tolerance capacity in plants.Evidence has shown that the constitutive expression of bovine calmodulin in tobacco results in a shortened germination time of transgenic tobacco seeds under salt stress.Arabidopsis over expressing GmCaM4exhibit increased expression of AtMYB2-regulated genes, including proline-synthesizing enzymes, suggesting that this feature confers salt tolerance to the transgenic Arabidopsis by enabling the accumulation of proline.Our previous report have shown that transgenic rice over-expressing OsCam1–1 grow better under salt stress than wild type.Wu H.and colleagues have found that the biphasic Ca2+ signal and enhancement of OsCam1–1 expression in rice cause heat stress-mediated expression of downstream heat shock-related genes, and OsCam1–1 over expression Arabidopsis are more tolerant to heat stress than its wild type.In another report, AtCam3 knockout mutant Arabidopsis showed a clear reduction of thermotolerance after heat treatment at 45 °C, and when AtCam3 was over expressed in mutant and wild type Arabidopsis, the thermotolerant ability was rescued and increased, respectively.Moreover, co-expression of some heat shock protein genes with AtCaM3 suggested that AtCam3 plays a key role in the Ca2+-CaM heat shock transduction pathway.The versatile functions of CaM are interesting, especially the role in the regulation of gene expression.CaM proteins directly modulate transcription factors , and some of these TFs have been verified to play roles in stress signaling pathways; however, the Ca2+ and Ca2+/CaM-regulating TF mechanisms remain incompletely understood and require further investigation.The aim of the present study was to investigate the downstream effects of OsCam1–1 over expression on gene expression regulation in rice under salt stress using a transcriptomic approach and to identify the interacting proteins to elucidate the role of OsCam1–1 in the salt stress response mechanism.

CaM is a multifunctional protein that regulates the activities of numerous target proteins.Genome-wide analysis techniques such as transcriptome profiling are particularly suitable for identifying the downstream components that are potentially regulated by CaM.In our previous report, rice over expressing OsCam1– 1 showed a significantly higher relative growth rate than wild type when grown under salt stress.Here, transcriptome profiling of the 3-week-old rice leaves of transgenic rice over-expressing OsCam1–1and its wild type under normal condition and salt stress conditions for 4 h was conducted.More than 185 million reads from eight libraries from single-end RNA-Seq by Illumina Hi-Seq 2000 were obtained, with a total read of each library between 22 and 25 million reads.The reads were processed by POPE, which provided a total clean read per library of more than 99% of the total reads.At least 93% of the clean reads were mapped to the rice genome reference, Michigan State University rice annotation project’s MSU7 and less than 11% of the clean reads were multiple alignment reads.To compare the transcriptome profiles of the rice, differential gene expression analysis of the transcriptome data using DESeq was carried out, which provided the number of differentially expressed genes summarized in Table 2.Analysis of the wild type identified 12,184 DEGs between the transcriptome profile under normal and salt stress conditions , in which 5842 and 6342 genes were up-regulated and down-regulated, respectively.For transgenic rice over-expressing OsCam1–1, comparisons between normal and salt stress conditions revealed a total of 13,259 DEGs with 6434 and 6825 up-regulated and down-regulated genes, respectively.Furthermore, the transcriptome profiles of the transgenic rice were compared with those of the wild type.Under normal conditions , 2022 DEGs were identified, with 892 and 1130 DEGs expressed at higher or lower levels in the transgenic rice, respectively.Under salt stress, comparisons of transgenic rice with wild type rice revealed 1677 DEGs, with 957 and 720 DEGs expressed at higher or lower levels in the transgenic rice, respectively.The scatter plots showed quantitative overview of the four transcriptome profile comparisons.

OsCam1–1 was found to be highly expressed in transgenic rice under both normal and stress condition, with an average RPKM of 1758.67 and 1644.62, while the average RPKM of wild type under normal and stress conditions was 91.94 and 97.84, respectively.The expression of OsCam1–1 in the wild type was not induced at 4 h after salt stress , in good agreement with a previous study.According to a gene expression study conducted by Chinpongpanich et al., the transcript level of OsCam1–1 determined by qRT-PCR was highly induced at 1 h after 150 mM NaCl treatment and then sharply decreased after 1 h.This result validated the over expression of OsCam1–1 in transgenic rice with an approximately 18-fold change in RPKM compared with wild type.Based on a differential transcriptome analysis, the gene expression levels of those 2022 and 1677 DEGs were thus likely affected by OsCam1–1 over expression.In our previous report, OsCam1–1-overexpressing lines showed a significantly higher relative growth rate than wild type when grown under salt stress.Based on the genes identified herein, among which several were involved in central energy pathways, sucrose and starch levels were determined in the three independent lines under normal and salt stress conditions at day 3 and 5 after treatment.Salt stress led to a significant reduction of the starch level and slightly decreased sucrose levels in both wild type and transgenic rice lines.Noticeably, at day 3, the transgenic lines could maintain the sucrose and starch levels better than the wild type under salt stress conditions.At day 5, the trends observed for sucrose and starch levels in transgenic rice under salt stress conditions were similar to those in wild type.In addition, the photosynthesis rate , stomatal conductance , intercellular carbon dioxide and transpiration rate were examined in the transgenic rice over-expressing OsCam1–1.Under salt stress, Pn, gs and E decreased at both day 3 and day 5, while Ci decreased slightly at day 3 of treatment.Interestingly, transgenic rice had slightly lower Pn values than wild type rice at both day 3 and day 5 and tended to have lower gs and E values at day 5 of salt stress treatment.In contrast, the Ci measurements did not reveal significant difference between the transgenic and wild type.For FV′/FM′, which reflects the maximum efficiency of photosystem II, no change was observed under the given salt stress conditions,nft growing system and the transgenic rice did not exhibit difference either under normal or salt stress conditions compared with the wild type.In a previous study, the northern blot results showed the highest expression levels of OsCam1–1 in transgenic rice line L1 among the three transgenic rice lines.Under both normal and salt stress conditions, the sucrose and starch content correlated with the expression level of OsCam1–1 in those transgenic rice lines.CaM does not possess functional domains other than EF hand motifs, so it functions by binding to and altering the activities of various interacting proteins.To understand how CaM1 mediates Ca2+-signal responses, its specific interacting proteins were identified using a cDNA expression library with 35S-labeled rOsCaM1 protein as the probe.The purity of the prepared 35S-labeled rOsCaM1 protein was examined by SDS-PAGE.To test its specificity, PVDF membrane spotted with various amounts of CaMKII peptide, calcineurin, and BSA was incubated with the probe.The autoradiograph showed that the probe only interacted with CaMKII peptide and calcineurin but not BSA, and the intensity of the signal on the X-ray film was dose-dependent.The results indicated that the 35S-labeled rOsCaM1 protein could specifically bind to well-known target proteins in the presence of Ca2+.After screening the cDNA library, the purified clones from the tertiary screening were titered before performing single-clone excision.As a result, 10 distinct positive cDNA clones were obtained.All unique pBlue script SK plasmids obtained from the single-clone excision were sequenced to determine the cloned cDNA insert sequences.The resulting sequences were BLAST searched against the Rice Genome Annotation Project and the Rice Annotation Project databases.The functions of 8 OsCaM1 targets were identified , which were diverse and potentially involved in various cellular processes, including metabolism, transcription, movement of organelles and vesicles, membrane transport, and signal transduction.Four known CaM-binding proteins previously identified in other plants were obtained from this screening, which included a cyclic nucleotide-gated ion channel , a glutamate decarboxylase, a CaM-binding transcription activator, and a kinesin motor domain-containing protein.The six identified putative novel CaM1-binding proteins comprised a transferase family protein , a response regulator receiver domain-containing protein , a lipin , a myosin heavy chain-containing protein , and two proteins with unknown function, LOC_Os08g34060 and LOC_Os02g13060.Interaction of eight putative target proteins with OsCaM1 was confirmed by protein blot analysis.

The mitogen-activated protein kinase cascade is a highly conserved central regulator of diverse cellular processes.CaM plays role in the MAPK/MPK cascade by binding to mitogen-activated protein kinase and/or mitogen-activated protein kinase phosphatase.A rice MAPK, BWMK1 encoded by an HT salt-responsive DEG, could phosphorylate the OsEREBP1 transcription factor for binding to the GCC box element , which is a basic component of several pathogenesis-related gene promoters.Inositol 1,3,4-trisphosphate 5/6-kinaseencoded by two HT salt-responsive DEGs, phosphorylates inositol 1,3,4-trisphosphate to form inositol 1,3,4,5-tetrakisphosphate and inositol 1,3,4,6, tetrakisphosphate, which are ultimately converted to inositol hexaphosphate and play roles in plant growth and development.In rice, the T-DNA mutant of an IPTK gene showed reduced osmolyte accumulation and growth under drought conditions, and some genes involved in osmotic adjustment and reactive oxygen species scavenging were down-regulated.In addition, over expression of DSM3resulted in a decrease in inositol trisphosphate , and the phenotypes were similar to the mutant under salt and drought stress conditions.These findings suggested that DSM3 might play a role in fine-tune balancing the inositol phosphate level when plants are exposed to stress or during development.Diacylglycerol kinaseen coded by an HT salt-responsive DEG, catalyzes the conversion of diacylglycerolto phosphatidic acid, and PA plays a role in the stress signaling pathway, including the MAPK/MPK cascade.A report has shown that the expression of OsBIDK1 encoding rice DGK is induced by benzothiadiazole and fungal infection.Moreover,transgenic tobacco constitutively expressing OsBIDK1 was more tolerant to plant pathogenic virus and fungi.These findings suggest that several genes in the signaling process might be enhanced by OsCam1–1 under salt stress.Interestingly, a universal stress protein gene was identified as an HT salt-responsive DEG.Evidence has shown that the expression of tomato USPis induced by drought, salt, oxidative stress and ABA, and over expression of spUSP improves tomato drought tolerance via interactions with annexin, leading to the accumulation of ABA.In addition, a xylanase inhibitor protein gene , was highly expressed and induced by salt stress and OsCam1–1 over expression.A previous report has shown that OsXIP can be induced by methyl jasmonate and wounding, so it was suggested that OsXIP may play a role in pathogen defense.As many OsCam1–1 and/or salt stress affecting DEGs involve both biotic and abiotic stresses, OsCam1–1 may be a component that mediates the crosstalk of biotic stress and abiotic stress responses.

Developing detailed spatial below-ground information about both plant and microbial location was considered to be both a priority and a major hurdle by many attendees.Current approaches include infrared, X-ray, chemical sensing, and acoustic imaging although all have limitations.The EcoPOD provides an opportunity to collaborate with teams who are developing these methods, such as those at the Danforth Center and the ARPA-E TERRA program, to test existing methods over a range of soil types and conditions such as temperature and soil saturation, as well as develop new methods.The EcoPOD user has access to the full depth of the soil column.This gives an opportunity to place sensors through the column.Many sensors exist, but may benefit from miniaturization, as space is still relatively limited in the EcoPOD quadrants.This is a good opportunity to develop collaborations with both academia and industry partners for the engineering expertise needed to miniaturize sensors.The group noted that above ground phenotyping and sensing is relatively advanced, primarily due to the physical ease of access.As a result, there are many more field derived datasets to compare to experiments performed in the EcoPOD.This will be useful for bench marking.In particular, multiple participants raised concerns about whether the artificial lighting was sufficient.Previous Ecotron efforts have often been criticized for the poor quality lighting but recent advances in LED lighting may resolve much of this and the EcoFAB team has recently found that they can simulate field lighting conditions using new PHYTOFY RL LED lights which are tunable in 6 different spectra from UVA to far red that could also be used in EcoPODs.Collecting sensor and imaging data on aerial growth of plants under a set of standard conditions in a defined field soil should be a priority.

There has been much recent interest in predictive phenotyping, where data on young plants is predictive of mature traits such as yield and grain quality.This has been developed in field systems,ebb and flow trays including via Advanced Research Projects Agency–Energy funded projects such as Transportation Energy Resources from Renewable Agriculture , but the EcoPOD would allow further interrogation of these models, in particular the response to individual climatic parameters e.g., drought vs.temperature.The addition of EcoPOD capabilities to bridge the gap between small-scale EcoPOD and large scale field capabilities will enable scientists to address critical DOE missions in energy and the environment.For example these capabilities will enable elucidation of molecular mechanisms by which microbial communities and abiotic constraints control key geochemical processes such as soil carbon cycling.They will also enable rapid development and translation of beneficial microbial communities from benchtop to field applications to support efforts in sustainable bio-energy and bio-products.Importantly, the containment and control afforded by EcoPODs and EcoFABs will enable pioneering studies in secure bio-systems design to provide key insights into the persistence, fate, and control of engineered microorganisms within soil micro-environments.Finally, there are opportunities to collaborate with other fabricated ecosystem projects at various scales across the globe.There was a lot of enthusiasm from the participants about using the EcoPODs for experiments that are challenging to do in the field due to regulatory or safety concerns.Examples include understanding the persistence and fate of engineered microbes within contained and controlled environments to identify risks and effective containment strategies.It was also noted that it’s an opportunity to monitor the effects of potentially environmentally hazardous materials such as plastic microfibers or carbon nanotubes.There was also interest from National Labs with secure facilities who could adapt technologies such as the EcoBOT, a robot that enables automated EcoFAB experiments, and EcoPOD, once they are derisked and developed further, for work which requires secure facilities that are not available at Berkeley Lab.

Due to the breadth of scientific background among our workshop participants, many great experimental ideas were discussed including topics such as carbon sequestration experiments, for which the EcoPOD’s semi-closed system can facilitate mass balance calculations easier than in field experiments.Furthermore, deep soil processes were discussed as most data from soil experiments does not exceed the top 10 cm.Additionally, soil atmosphere gas exchange was mentioned, which we will keep in mind as we are aiming to develop future prototypes that can provide gas-tight conditions.Similarly, climate change simulations that include warming or elevated CO2 concentrations are a future experimental goal that will require prototype updates that are achievable.All participants were excited about the possibility of separating individual environmental parameters to observe their effect on the plant-microbe-soil atmosphere ecosystem.This includes being able to better predict the phenotype of engineered bio-energy crops under different environmental conditions, reducing the need for expensive and complex multi-site field trials.The EcoFAB team has made access to the units purchased by Berkeley Lab a priority.For example, detailed protocols on how to fabricate them have been published , and TEAMS, a project funded by DOE, is dedicated towards the dissemination of EcoFAB supplies and protocols including model micro-biomes to foster interlaboratory science and experimental standards.Through these efforts, laboratories across the globe are now using Berkeley Lab-designed EcoFABs as well as developing their own iterations, with a new EcoFAB ring-trial study with B.distachyon and a synthetic microbial community planned for early 2021.As discussed above, EcoFABs and EcoPODs are complementary technologies with different strengths and weaknesses.One way Berkeley Lab is leveraging this fact is through the use of higher throughput EcoFABs experiments to assess important parameters that can later be implemented within EcoPODs.

The EcoBOT being developed at Berkeley Lab through the Trial Ecosystems for the Advancement of Microbiome Science project will support remote, high throughput EcoFAB experiments to improve to improve turnaround, standardization, and reproducibility for EcoFAB experiments.ESM analyses are typically performed to evaluate and optimize space mission payloads to minimize launch costs as a function of mass, volume, power, cooling, and crew time needs.NASA’s exploration medical system trade study tools, which includes a systems engineering model and a medical risk analysis model, have the potential to serve as a foundation for this analysis.There are many obstacles ahead before making pharmaceutical foundries in space a reality.What has not been thoroughly discussed in this review is the downstream processing of a molecular medical foundry, which will depend on the purity needed for the pharmaceutical formulation, delivery method, production host, etc.Downstream processing, the purification of the target molecule from the production host, is a resource-intensive aspect of bio-pharmaceutical production across all platforms.There is a lack of downstream processing technology that translates well from Earth-based constraints to those of space, as they often require a high quantity of consumables, raw materials, equipment, and cleaning.This bottleneck will need to be addressed for pharmaceutical foundries in space to succeed.One approach is to conduct research on novel drug delivery modalities to reduce the need for downstream processing, and another is to diminish the resource demands of the processing itself.A growing emphasis on distributed and just-in-time pharmaceutical production for healthcare on Earth is already driving solutions to these downstream challenges.The other major hurdle is in regulatory compliance.Production and administration of pharmaceuticals in space will require extensive quality control; manufacturing a small molecule might have 50 critical tests, while manufacturing a biologic may have over 250 tests.Here, the advent of personalized medicine on Earth will illuminate a path forward.The shift from mass produced to individualized patient-specific medicine hinges on re-structuring the path to regulatory approval and quality control.While there are many challenges ahead that need to be addressed to pave the way for Earth-independent life support, the rewards of this pursuit will include great insights into supporting life on Earth and beyond.Understanding this value, we aim to highlight the critical importance of developing Earth-independent systems in the future of human exploration.We illustrate that molecular pharming provides a diverse production tool set that could be used to establish a robust molecular medical foundry subsisting on a small fraction of food crop needs.In addition to advocating for molecular pharming as a synergistic asset of space life support systems,4×8 flood tray we focus on the need for multi-faceted utilization of resources in limited environments such as space and extraterrestrial bodies.Sunflower is an oil seed crop of great importance worldwide, due to the excellent quality of the oil extracted from its seeds that are consumed in various ways.Cultivation of the sunflower is becoming increasingly significant globally.In 2008–2009, the world sunflower seed production was about 33 million tones, around 8.5% of the world’s total oil seed production , the leading producers of which are the EU, Russia, Ukraine, Argentina, USA,China, India, and Turkey.Foliar fertilization is an increasingly popular practice with particular importance for the production of high value crops such as sunflowers with many examples of positive responses to foliar application of micro-nutrients, including zinc , iron ,boron , manganese , and molybdenum , on the seed yield and seed quality of sunflower.Foliar fertilization has particular value in overcoming nutrient deficit resulting from stress conditions, such as salinity and drought, which often compromise root growth and decrease root absorption capacity.Foliar application of Zn resulted in a greater improvement in Zn densities in rice and wheat grain when compared with soil applied Zn.

Foliar fertilization is theoretically more immediate and target oriented than soil fertilization since nutrients can be directly delivered to plant.Thissues during critical stages of plant growth.Optimizing the efficacy of the foliar applied nutrients is therefore of great importance from an economic, agronomic and environmental point of view.Our understanding of the factors that determine the ultimate efficacy of foliar applications remains poor and the response of plants to foliar Zn applications is highly variable.Many reports indicate that foliar application of Zn may significantly increase the concentrations of Zn in the applied leaves but may have little effect on foliar concentrations in non-sprayed Thissues or Thissue that develop subsequent to the foliar application.The factors that control the uptake and subsequent translocation of foliar applied nutrients out of the leaf, and the effect of spray formulation on these processes are poorly understood.While various approaches have been used to determine the efficacy of foliar applied nutrients using stable and radioactive isotopic labeling , it remains challenging to determine the pathways of mobilization from leaf to shoot and to monitor the influence of foliar formulation on phloem loading and micro-nutrient transport.To address the inherently low efficiency of many foliar Zn formulations, a wide range of commercial products have been developed and marketed.Recently, there has been much interest in the incorporation of organic moleculesor bio-stimulants into foliar fertilizers with the rationalization that these additives will enhance the uptake, or subsequent mobility of the applied nutrient.The term ‘bio-stimulant’ is used to describe a substance or material, with the exception of nutrients and pesticides, which when applied to plants has the capacity to beneficially modify plant growth.Currently there is very little scientific evidence that bio-stimulants can specifically enhance the uptake and utilization of foliar applied fertilizer materials.The technique of X-ray fluorescencehas been widely used in the research of elemental distribution in plant Thissues, and has proved to be a promising tool to study in vivo localization of metals in plants due to its high-resolution and sensitivity.XRF analyses can be performed to visualize cellular and subcellular distribution of elements in plants without significant pretreatment of the samples.We have previously applied this technique to characterize the location and to monitor changes in concentration and distribution of Zn during plant development or following foliar applications.In this current study, we will utilize μ- XRF to obtain high spatial quantification of elemental distribution and transport following the application of various Zn formulations with the aim of:increasing our understanding of the processes that govern the localization and transport of foliar applied nutrients with emphasis on Zn, and to determine if the formulation of the foliar applied Zn, with addition of macro-nutrients or bio-stimulant, alters the mobility of the element following its absorption by the leaf of sunflower.Because of the complex nature of the commercial products used, it was not possible to prepare a control spray treatment that contained equivalent amounts of all nutrient elements present in Kick-Off or CleanStart.At the rate used here, GroZyme contains negligible concentrations of all essential plant elements.To avoid the possibility that the effect of the foliar spray was a consequence of alleviation of secondary nutrient deficiency, all plants were grown with continuous and abundant soil nutrient.Leaf analysis was conducted and all nutrients were found to be present at adequate levels and plants showed no sign of nutrient deficiency.