High concentrations of Na+ in the cytoplasm disrupt the ionic balance and the uptake of essential mineral nutrients, such as K+, which in turn causes adverse effects on many metabolic pathways. To cope with salt stress, plants have evolved various tolerance mechanisms including two transport processes at the single cell level. Either exporting Na+ out of the cell, or compartmentalizing excessive Na+ into the vacuole. These two transport mechanisms act in a coordinated manner to maintain a low Na+ concentration in the cytoplasm. However, it remains unknown if they are regulated by the same or different signaling pathways. The SOS pathway is generally viewed as a signaling mechanism for the activation of the Na+ efflux through SOS1, a NHX-type Na+/H+ exchanger in the plasma membrane. The loss of function of SOS genes thus results in hypersensitivity to NaCl, coupled with the Na+ over-accumulation in the cytoplasm. On the other hand, some Na+/H+ exchangers are localized in the tonoplast and may be involved in transporting Na+ from the cytoplasm to the vacuole. However, the exact role of different NHX isoforms responsible for salt tolerance remains unclear. Interestingly, the two distinct but inter-connected salt transport processes appear to be both regulated by calcium signaling, in which calcineurin B-like proteins are thought to be the primary calcium sensors during salt stress adaptation. Among them, CBL4 and CBL10 display distinct tissue expression patterns and subcellular localizations. The spatial specificity of these two calcium sensors may contribute to their functional diversification in salt stress adaptation. In order to understand how they work synergistically in the regulation of salt tolerance,plastic pots 30 liters we genetically analyzed the salt-sensitive phenotype of the cbl4 cbl10 double mutant in comparison with the single mutants.

The cbl4 cbl10 double mutant was dramatically more sensitive to salt as compared to the cbl10 and cbl4 single mutants, suggesting that CBL4 and CBL10 either functionally overlap or each directs an independent salt-tolerance pathway. If the two CBLs are functionally overlapping, they should regulate the same transport processes and then the double mutant should not only show more severe phenotype but also show more severe deviation in the Na+ and K+ contents as compared to the wild-type plants. However, that was not the case: cbl4 and cbl10 displayed generally opposite Na+ and K+ profiles. Although the cbl4 cbl10 double mutant plants showed Na+ over-accumulation compared to the wild type, but significantly lower Na+ content than the cbl4 single mutant . This suggests that CBL10 should not be involved in the CBL4-regulated Na+ extrusion process , although these two calcium sensors interact with a common downstream kinase CIPK24 . Instead, CBL10 should regulate a distinct Na+-transport process in response to high salt, probably the Na+ sequestration into the vacuole, as suggested by its tonoplast localization and the lower Na+ content in the cbl10 mutants. This is consistent with the general theme that the Na+ efflux or Na+ sequestration into the vacuole both contribute to salt tolerance and disrupting either may result in elevation of the Na level in the cytoplasm and thus leading to salt sensitivity. Certainly disrupting both transport processes would lead to more severe salt sensitivity, which match the more sensitive phenotype of cbl4 cbl10. Previous studies suggested that CIPK24 serves as the common downstream target of CBL4 and CBL10 by forming CBL4-CIPK24 or CBL10-CIPK24 complex at the plasma or vacuolar membrane separately. Although our findings in this study supported this hypothesis, they also suggested that other CIPKs, in addition to CIPK24, should be also involved in the CBL10-mediated pathway based on the genetic evidence that double mutants of cbl4 cbl10 and cipk24 cbl10 displayed a significant enhancement in Na+ sensitivity as compared to cipk24 .

Indeed, screened by the yeast two-hybrid assay, we found that CBL10 did interact with other CIPKs in addition to CIPK24 . Various combinations of CBL10 with different CIPKs may target different target proteins and exhibit diverse functions. To examine whether SOS1 is a downstream component of CBL10 in the pathway, we also compared the salt sensitivity between sos1 cbl10 and sos1. In our test conditions, the salt sensitivity of cbl4 cbl10 and sos1 cbl10 was comparable to sos1 , suggesting that SOS1 may serve as aconverging point for the two CBL pathways. However, the double mutants cbl4 cbl10 and sos1 cbl10 accumulated much lower Na+ content than the single mutants of cbl4 and sos1, respectively, under salt conditions , which implies that CBL10 and SOS1 functions in two different transport processes in regulating Na+ homeostasis. For instance, in the sos single mutants in which the Na+ efflux is blocked, the CBL10 pathway functions to transport Na+ into the vacuole leading to the over-accumulation of Na+ in plant tissues. When the vacuole sequestration is defective in the cbl10-associated double mutants, the Na+ uptake is inhibited as a feedback of lacking storage space, leading to less accumulation and thus lower Na+ content in these double mutants as compared to the sos single mutants . Despite overall lower Na+ content in plant tissues, the double mutants showed similar salt sensitivity as sos1 because the majority of Na+ in these double mutants is in the cytoplasm effectively causing toxicity. Our results thus provide an example where a two-tier evaluation system must be implemented for dissecting salt tolerance mechanism in plants: First by whole-plant phenotyping and further by the analysis of Na+/K+ homeostasis . Concerning the target transporters for CBL10, all evidence so far supports the hypothesis that the CBL10-CIPK pathway may regulate Na-transporters in the tonoplast. Sequestration of Na+ into the vacuole is presumably fulfilled by an array of Na + transporters that include the vacuole-localized NHX-type Na+ /H+ transporters.

However, recent genetic evidence indicates that vacuole-localized antiporters NHX1-4 have Na+-transport activities but may not contribute much to the vacuolar Na+ compartmentation, because the quadruple knockout mutant nhx1/2/3/4 is not more sensitive to NaCl than the wild type. Furthermore, vacuoles isolated from the quadruple mutant still retain the Na+ uptake that is independent to the pH gradient, implicating the presence of NHX-independent Na+ transporters in Arabidopsis vacuoles. We speculate that some of these unknown transporters may serve as CBL10-CIPK targets. On the other hand, endosomal compartments emerge as critical players that may be directly involved in controlling Na+ homeostasis. A possible but yet to be proved model is that the Na+ sequestration into the plant vacuole may actually be achieved, at least in part,round plastic pots through endosomal Na+ scavenging processes and subsequent fusion to the vacuole. NHX5 and NHX6 are localized to endosomal compartments and associated with protein trafficking from the Golgi/Trans-Golgi Network to vacuoles. Supporting this hypothesis is the finding that disruption of two endosomal NHXs in the nhx5 nhx6 double mutant showed increased sensitivity to salinity . Considering the fact that a proportion of the CBL10 protein was also localized to the dynamic endosomal compartments, NHX5/6 could also act as the candidate targets of the CBL10-CIPK complexes. In a recent work, translocon of the outer membrane of the chloroplasts 34 was identified as a novel interaction partner protein of CBL10 at the outer membrane of chloroplasts, clearly indicating that CBL10 can relay Ca2+ signals in more diverse ways than currently known.Identification of target transporter directly regulated by the CBL10-CIPK module is an important and challenging task for future research, which would also unravel the pathway through which Na+ is deposited into the plant vacuole. Treated wastewater, commonly called reclaimed or recycled water, is a valuable water source in arid and semi-arid areas where fresh water sources are becoming increasingly scarce due to urbanization and climate change . Reclaimed water may have many beneficial applications, including agriculture irrigation and landscape irrigation. In the state of California, these irrigation uses account for 37% and 18%, respectively, of the 650,000 acre-feet per year of water reuse . State policy calls to increase the use of reclaimed water to more than 2.5 million acre-feet per year by 2030 . Accompanying increased reuse, the presence and environmental risks of unregulated organic contaminants in reclaimed water are drawing attention . Pharmaceutical and personal care products and endocrine disrupting compounds are typically anthropogenic chemicals with known biological effects that may interfere with normal metabolism and behaviors of organisms .

Many PPCP/EDCs are routinely found in reclaimed water , as well as in surface water impacted by wastewater treatment plant effluent and in groundwater . When reclaimed water is used for irrigation, the associated PPCP/ EDCs may interact with the soil matrix and may contaminate groundwater and food crops . Accumulation of PPCP/EDCs into food crops that are consumed fresh, such as many leafy vegetables, is relevant due to the likelihood of unintentional human exposure. If research demonstrates that accumulation of PPCP/EDCs by crops is unlikely to result in human health risks, this will provide scientific basis to promote use of reclaimed water, as well as enhance positive public perception of water reuse. Many factors influence the uptake of organic compounds into plants, such as by affecting diffusion through cell membranes. Briggs et al. suggested that chemical hydrophobicity is an important factor affecting uptake by diffusion and that chemicals with a log Kow of 1 – 3.5 have the greatest plant uptake potential because lipid and aqueous solubility are balanced . In addition to hydrophobicity, molecular ionization has also been shown to influence plant accumulation, such as of herbicides . Charged molecules may have a reduced potential for plant uptake, since ionization may reduce their ability to permeate cell membranes . However, the role of ionization is poorly understood and exceptions have been noted . To date only a handful of studies have considered plant uptake of PPCP/EDCs . While these studies have clearly shown the ability for plants to take up PPCP/EDCs, the state of knowledge is limited to a few compounds or plant types. Due to the analytical challenges of detecting chemicals at trace levels in plant matrices, most studies also relied on the use of artificially high concentrations, with a few exceptions . In this study, we comparatively determined the accumulation of four commonly occurring PPCP/EDCs, i.e., bisphenol A , diclofenac , naproxen , or nonylphenol , at relevant environmental levels into two leafy vegetables, lettuce and collards, and examined the composition and distribution of accumulated residues. These compounds have been frequently detected in reclaimed water and surface water , and have different ionization states at neutral pH. To achieve realistically low concentrations while affording quantitative measurement, 14C-labeled compounds were used. Results were used to infer effects of plant type and compound characteristics on plant accumulation and estimate probable human intakes. Following 21 d of hydroponic cultivation, plants were sacrificed for analysis of 14C accumulation and distribution. Each whole plant was rinsed with DI water, and then separated into roots, stems, new leaves, and original leaves.Individual plant samples were placed in pre-weighed metal screen pouches, weighed to determine wet weight, and dried at 50 °C for 60 h. After drying, each plant sample was weighed to measure the dry weight, and then chopped and mixed in a stainless steel coffee grinder. The grinder was rinsed between samples with DI water and methanol to prevent cross contamination. Multiple 150 mg sub-samples of each plant sample were analyzed until standard deviation of the sub-samples was below 20%, due to notable variation in plant tissue activity. Sub-samples were combusted on an OX-500 Biological Oxidizer at 900 °C for 4 min, and the evolved 14CO2 was trapped in 15 mL of Harvey Carbon-14 cocktail . The 14C was measured on a Beckman LS 5000TD Liquid Scintillation Counter . Recovery was 91-96% for spiked standards,which was used to correct for the actual activity. The activity and weight of the sub-samples were used to determine the total radioactivity accumulated in different tissues of each plant. Analysis of 14C by combustion provided information on total residue in plant tissues. To better understand the nature of the residue, plant samples were solvent extracted using a method modified from Wu et al. . The fractions of 14C in solvent-extractable and nonextractable forms were separately determined.

However if the timing and controls on hot moments are unknown or sporadic, less frequent sampling may significantly underestimate N2O emissions . Our results suggest that roughly 8,000 randomized individual chamber flux measurements would be needed to accurately estimate annual N2O budgets from these agricultural peat lands with a 95% confidence interval and 10% margin of error, assuming the drivers of hot moments were not well understood. Approximately 500 individual measurements would yield a 50% margin of error. Given the more sporadic nature of CH4 hot moments, our results suggest that it is even more difficult to accurately estimate CH4 fluxes with periodic sampling in these ecosystems. Analyses found that at least 17,000 and 2,500 individual flux measurements would be needed to estimate annual CH4 budgets within a 10% and 50% margin of error, respectively. The agricultural maize peat land soil studied here was a much larger source of soil GHG emissions than other maize agroecosystems. While agricultural peat soils are highly productive, average annual GHG emissions were 3.6-33.3 times greater on an area-scaled basis and 3-15.6 times greater on yield-scaled basis relative to other agricultural maize emissions estimates. We conducted an upscaling exercise as a first approximation of the potential impacts of maize peat land fluxes on regional GHG budgets. Our estimates suggested that maize agriculture on similar peat soils in the region could emit an average of 1.86 Tg CO2e y-1 .This value is significantly higher than previous estimates for the region and highlights the importance of including high frequency N2O measurements to capture hot moments in N2O fluxes,plastic pots 30 liters the disproportionate impact N2O emissions have on agricultural peat land GHG budgets, and that these agricultural peat lands are significant N2O sources.

We also found that irrigation timing and duration, not fertilization, was the predominant driver of N2O and CH4 emissions and a significant source of the total GHG budget. Determining management strategies that reduce soil N2O and CH4 emissions, particularly changes in flood irrigation timing and duration, could have a disproportionate impact on reducing total agricultural peat land GHG emissions .Although legends of humans using coffee in Ethiopia date back as early as 875 A.D., the earliest verifiable evidence of human coffee consumption occurs in Yemen in the 15th century. At this time, it was illegal to bring unroasted coffee out of Arabia, and strict measures were taken to ensure that viable coffee seeds did not leave the country. The birth of coffee production in India is attributed to the Indian Muslim saint Baba Budan, who, on his return from a pilgrimage to Mecca, allegedly smuggled seven coffee beans out of Arabia by hiding them in his beard. In 1670 he planted these seeds in Karnataka, and cultivation soon spread throughout the state and into neighboring regions. The first large-scale plantations arose with British colonization and spread rapidly throughout South India, fueled by increasing demand for export to northern latitudes. The proliferation of coffeehouses in Western Europe during this era proved to have substantial social consequences. Also known as “Penny Universities” since the price of entry and a cup of coffee was commonly one penny, coffeehouses in 17th century Britain came to play an important role in social and political discourse. In a society with such a rigid socioeconomic class structure, coffeehouses were unique because they were one of the only places frequented by customers of all classes.Thus they became popular establishments for discourse and debate, open to all classes and unfettered by the structure of academic universities. Intellectuals found in “the hot black liquor a curious stimulus quite unlike that produced by fermented juice of grape.”English coffeehouses “provided public space at a time when political action and debate had begun to spill beyond the institutions that had traditionally contained them,” and because of this, are widely accepted as playing a significant role in birthing the age of Enlightenment in Europe.While coffee was bringing the Enlightenment to Western Europe, the commodity was having opposite effects in the regions where it was being produced.

In India, the age of British plantations was rife with suffering and oppression, as slavery and forced labor were common practice. Historical research reveals that “during Europe’s industrial revolution and rise of bourgeois society, slavery, coffee production, and plantations were inextricably linked.”Historical records indicate that in the 1830s, the East India Company held over 247,000 slaves in Wayanad the Malabar coast alone.Even after slavery was officially abolished in 1861, so-called “agricultural slavery” and indentured labor on plantations continued. 8 According to historical accounts, indentured laborers were treated almost identically as they were during the height of slavery. To this day, an estimated 18.3 million people in India and 46 million people worldwide live in conditions of modern defacto slavery, such as bonded labor, human trafficking, and forced marriage. The global coffee market has always been volatile. Plagued by unpredictable harvests, susceptibility to weather events, and massive disease outbreaks, regional coffee production has risen and fallen dramatically over the centuries. For example, in the late 19th century in Sri Lanka an outbreak of the fungal pathogen known as “coffee rust” caused 90 percent of area under coffee cultivation on the island to be abandoned. 11 This past century has been no different for India. As the Great Depression affected coffee exports around the world in the 1930s, the Coffee Board of India was established to protect farmers and promote consumption of coffee. The Coffee Board of India, run by the federal government’s Ministry of Commerce and Industry, pooled farmers’ coffee for export at a set price. This provided price stability for farmers but also eliminated incentives to improve quality. From 1991 – 1996 a series of economic reforms relegated the coffee market in India entirely to the private sector. Immediately thereafter, the price of coffee fell from its 1997 levels of around $2.50 per pound to a staggering 45 cents per pound in 2002, the lowest it has been in over fifty years.India was not alone in this plight. While certainly not the only cause of financial insecurity among farmers, the spread of neoliberalism and free trade in the global commodity market has historically been associated with large increases in price volatility and overall downward trends in price, which has had deleterious effects for small-scale producers who depend on these markets for their livelihoods.

Especially in the 1980s and 1990s, growth and consolidation among multinational commodity traders led to a relative loss of market power among producing nations, while foreign pressure from international donors forced many of those nations to privatize their commodity export authorities against their own best interests.This has led to income instability and poverty for many coffee farmers around the world. The coffee farmers of Kerala are facing many of the same challenges that currently plague coffee farmers all over the world. In recent years the global price of coffee has fell drastically from $2.88-per-pound in 2011 to 93 cents-per-pound as of May 2019.While maintaining its downward trend over the past decade, the price continues to fluctuate wildly, making it impossible for farmers to budget their yearly expenses. It is not unheard of for the price to even dip below an individual farmer’s production costs,round plastic pots leaving powerless farmers forced to sell their harvest at a loss, or let it spoil in the fields and get nothing at all. How is it possible that coffee farmers are selling their harvest for less than what it cost them to produce it? While this seems paradoxical to the very basis of economics, it is a common situation facing farmers of many different cash crops, where prices are determined by what are called “buyer-driven supply chains.” While many factors go into the creation of buyer-driven supply chains, some of the few largest factors are discussed below. All this to say, farmers do not have the capacity to determine the price they get for their own products. Prices are driven by market conditions, speculation, futures contracts, and corporate interests who control the majority of world-market shares. With the growth of powerful commodities traders and the liberalization of international markets, prices for coffee and incomes for farmers have reached historic lows. This has led to an increasingly tenuous existence for those who already struggle to get by. Historically, coffee cultivation consisted of only one plant species, Coffea arabica. Today, Coffea arabica still makes up most of the world’s coffee production , but cultivation of another species, Coffea canephora, also known as robusta coffee, is growing due to its higher levels of hardiness and productivity.In addition, a very small amount of a third species Coffea liberica is grown. Although modern coffee production is currently limited to the scope of these three species, a large diversity of sub-varieties and hybrids are grown throughout the world, each with their own unique flavors and characteristics. Coffea arabica is widely lauded as having the best cup quality, and consistently fetches a higher price on the global commodity market. It also tends to grow better in slightly shaded conditions, making it conducive to traditional inter cropping methods.

In India, Coffea arabica is usually grown under the shade of other cultivated trees, such as jackfruit and areca nut, or under the shade of native forest trees, which are used to support vines of black pepper.In the under story below the coffee plants ginger, clove and turmeric are grown. In addition to sustaining families of farmers for generations, a recent study has shown that these multi-species farms support much higher levels of animal biodiversity than conventional monocultures, and that they sequester soil carbon at the same rate as surrounding rain forests. However, the rise of C. canephora as a cash crop has changed things in Kerala. Due to its higher yields and tolerance to pests such as coffee rust C. canephora plantations have replaced multi-species C. arabica farms over huge swaths of India in recent decades. Today, nearly 80% of coffee grown in Wayanad and surrounding regions is C. canephora. Since this robusta species prefers full-sun conditions, this shift away from C. arabica is associated with the removal of shade trees and a proliferation of full-sun monoculture coffee plantations. This has had substantial consequences for biodiversity, erosion, watershed management, and other ecosystem services.This has the potential to negatively impact the small amount of C. arabica that remains in Kerala. Studies indicate that deforestation can lead to a hotter and drier local climate.Coffea arabica is a finicky plant, thriving in a narrow temperature range between 18˚ – 21˚ Celcius.It follows that this pattern of tree removal could lead to conditions in Kerala becoming less ideal for Coffea arabica. This would suggest the potential for a feedback loop, in which robusta production and the associated deforestation lead even more farmers to convert to robusta in order to cope with changing environmental conditions. If climate change is occurring in Kerala, it would not only be threatening cultivated coffee, but also a multitude of wild species. At least six species of wild coffee are known to occur in India.According to a recent study there are now 124 known species of wild coffee, each with their own under-studied and potentially useful characteristics, such as drought or pest resistance, unique flavor profiles, or naturally decaffeinated beans.Of these, an estimated 60% are threatened with extinction due mostly to climate change and habitat loss.The following analysis examines the local climate of Wayanad in recent decades to determine if any changes are occurring. Farmers interviewed during a field visit to Kerala assert that local conditions have become hotter and drier, especially during specific times of the year that are important to the life cycle of the coffee plant. The farmers of Wayanad have suggested an increasingly unpredictable monsoon season, a failure of the “blossom rains” in early spring, and a decrease in November showers. The following study was conducted to corroborate the personal experience of these farmers, and, in the event that trends are found, to determine if causal factors point to global-scale or local forcings. The district of Wayanad in the State of Kerala, India is a mountainous tropical region with altitudes ranging from 700 to 2100m above sea level, daily temperature minimums from 14˚ – 20˚ C, and daily temperature maximums from 25 – 32˚ C.