Horticulture

Often, PGPR must be repeatedly added to a soil to maintain high population densities. One of the greatest research gaps limiting this field is inconsistent results when cultures are taken from lab to field. To better manage the ecosystem function of PGPR it is critical to study the survival and activity of both inoculated and native soil microorganisms in non-sterile soil.Carrier materials can influence inoculum success by providing protective habitats and also by affecting soil aggregate formation, which provide protection from predation . Soil inoculants are commonly prepared in formulations of powders, granules, and liquids . Previous research has evaluated the use of carrier materials to improve survival and distribution of inocula, much of which was conducted for soil inoculation with rhizobia . Peat moss is commonly used a microbial carrier and its benefits increase markedly if sterilized prior to inoculation . Vermiculite, lignite, and sodium alginate encapsulation have all been studied as alternative carriers to peat. Also,hydroponic gutter charcoal maintains high populations of inoculum suitable for use after 280 days of storage .

While this information has existed for years, very little work has been continued to examine the suitability of charcoal as a carrier for microorganisms.There is a variety of well-studied carrier materials, all of which have limitations that restrict their widespread use. For example, peat has had success in the rhizobia industry, but it is a highly variable material and the extraction of peat from bogs is unsustainable . The process of obtaining vermiculite also requires mining, which is associated with negative environmental impacts. As a result of limited sources, both peat and vermiculite are sometimes unavailable in regions where they are not naturally present . Alginate encapsulation of inoculum preparations offers a promising alternative to liquid inoculum, clay, and peat formulations. However, production of alginate beads is more expensive than the alternatives . Although the technology behind bioencapsulation has been reported since the 1980’s, the c rrent devices that prod ce the beads are still predominantly limited to lab-scale .Although countless studies have been performed on plant responses to soil inoculants, far less research has focused on inoculum preparation. A literature survey by Xavier et al.found that less than 0.5% of publications that cover rhizobia research discuss inoculum formulation. Many factors will differentially influence the survival of inoculum when applied to soils with diverse physical and chemical characteristics. For example, several abiotic soil parameters affected the survival of Azospirillum brasilense in bulk soil, including soil texture, water holding capacity, soil nitrogen, and organic matter . Accordingly, for PGPR to be considered a successful means to address major agricultural challenges, several things must be considered.

Carrier materials must be affordable, sustainable, and widely available. Also, the formulation of the inoculum must ensure that high PGPR populations can be stored and distributed into soils, and that once applied to soil, the PGPR can thrive, colonize roots, and commence plant-beneficial activities.Although biochar is a relatively new term to the scientific community, it is rooted in an ancient tradition of native Amazonians. Their method for disposing organic wastes involved heating it in deep earth pits, under low levels of oxygen. This left behind carbon rich “Terra Preta”, or black earth, which is still stable hundreds of years later and has proven to be an excellent soil amendment . Terra Preta soils contain high concentrations of nitrogen, phosphorus, potassium, calcium, and stable organic matter . These highly fertile pockets of soil are a stark contrast to the acidic indigenous soils, which are low in nutrients and organic matter and considered to be incapable of supporting agriculture . Interestingly, similar soil patches can be found throughout the world, and their high fertility is consistently associated with an abundance of black carbon . The use of charcoal also has a long history in agriculture and has been used to promote agronomic productivity for centuries . This tradition can be modernized by way of pyrolysis machinery specially engineered to heat solid or liquid biomass at designated temperatures in zero to low levels of oxygen. The resulting product has been termed biochar and biochar materials are receiving a lot of attention from scientists, engineers, farmers, and entrepreneurs .Given the current annual increase in atmospheric carbon dioxide of 4.1 tons year-1 , reduction of anthropogenic greenhouse gas emissions is critical .

Products of pyrolysis are carbonaceous and recalcitrant and can be incorporated into soils thereby serving as a stable carbon sink and climate change mitigation strategy . Biochars have been estimated to have mean residence times in soils of temperate climates of about 2000 years whereas fresh organic matter may be degraded in less than a decade . Pyrolyzing waste materials typically sequesters 50% of the source carbon as compared to traditional slash-and-burn techniques, which sequester only 3%, and natural decomposition, which retains 10– 15% . Biochar production has been credited as a tool that could offset 12% of anthropogenic carbon dioxide carbon equivalents annually if implemented on a global scale . Therefore, biochar production from all bio-wastes offers a sustainable mechanism for land and waste management while providing a carbon negative system.The process of pyrolysis generates porous, charred particles that structurally resemble the parent material but have carboxylated aromatic cores with slight negative charge . During pyrolysis a lack of oxygen available to the system results in a residual material rich in carbon. As pyrolysis temperatures are increased the resulting biochars become increasingly aromatic as oxygen-bound functional groups escape. NMR- based diagrams of slow and fast pyrolysis chars prepared with pyrolysis temperatures of 500°C, and gasification produced char generated at 750°C, display typical aromatic clusters that comprise biochar materials . Chen et al.used elemental analysis and FTIR to examine the sorption behavior of eight pine needle biochars produced at pyrolysis temperatures ranging from 100°C to 700°C. Pyrolysis temperatures up to 300°C mark the initial removal of OH, aliphatic C-O, and ester C=O groups from outer surfaces of such structures. At 400 ° C there is complete destruction of aliphatic alkyl and ester C=O groups that shield the aromatic core. At temperatures above 500 °C, there is further removal of aromatic COand phenolic –OH groups. The removal of these outer groups and exposure of the aromatic core is a key determinant of the sorption behavior and cation exchange capacity of the biochar . In general,u planting gutter for lignocellulosic materials, carbonization increases as combustion temperatures rise to 500°C and the materials approach full carbonization as temperatures reach 1000°C .The exposure of aromatic structures lends these biochars the property of having many fine pores. The abundance of nano to meso-sized pores contributes to the large surface areas associated with higher-temperature chars as well as to their enhanced ability to adsorb non-polar compounds . Biochar materials often retain the cellular structure of the feedstock, which can provide and intricate network of pores on the order of tens of micrometers in diameter . Hence, factors such as feedstock and pyrolysis conditions, the highest treatment temperature achieved during pyrolysis in particular, will affect the porosity, specific surface area, cation exchange capacity , and adsorptivity of the resulting biochar. This allows the opportunity to derive biochar materials with optimized properties for specific uses, such as agronomy, soil remediation, water filtration, or soil inoculum delivery. Meta-analyses of publications containing biochar field trials and greenhouse studies reveal that biochar application resulted in average increased above ground biomass ranging from a conservative 10% to 30% and a recent meta-analysis, which incorporated publications up to April, 2013 demonstrated an overall mean increase in crop productivity of approximately 11% with biochar application . From each of these meta-analyses it becomes apparent that biochar will have different effects on plant biomass and yield that is influenced by soil and biochar characteristics, application rate, crop variety, and time post amendment.

These reviews concur that biochar had the greatest positive influence on crop productivity when incorporated into acidic, clay, or sandy soils with low water holding capacity or low organic matter . The economic cost associated with biochar production and application has become a major limiting factor to its wide-spread use . Estimates by Brown et al.projected that current biochar production will only be profitable if bio-oil is simultaneously generated and carbon offsets have values in the range of 20 USD or more per metric ton. Shackley et al.provided a total assessment of the costs and benefits associated with biochar deployment and determined that biochar production costs are best reduced when feedstocks are waste feedstocks that would otherwise have a gate fee or landfill charge associated with their disposal. The greatest economic benefits associated with biochar use were related to energy production, as agronomic cost benefits can be highly inconsistent . Hence, adjustment to biochar products to ensure greater agronomic benefits could transition biochar use into a profitable sector. It is important to gain a fundamental understanding of how biochar application will affect soil biota before its use can be recommended on a broad scale. When applied to soils, biochar has many effects on soil physical and chemical properties that, in turn,affect the properties of the soil as a habitat for microbial growth. To date several exploratory studies have assessed the response of bacteria, fungi, and enzymes to biochar incorporation in a soil. In Figure 1.5, scanning electron micrographs depict clear images of fungal hyphae extending into biochar pores and bacterial cells located on char surfaces . Furthermore, microscopic, chromatographic, and spectroscopic studies have shown root hairs entering water-filled macropores or bonding to biochar surfaces . At this interface the biochar particles can adsorb organic compounds released from growing roots. Thus, biochar pores may serve as an ideal microenvironment for biological activity. However, contrary to these findings Quilliam et al.report minimal colonization of biochar by native soil microorganisms 3 years post 2– 4% amendment with a wood-derived biochar. In examining the influence of biochar on native soil bacteria many investigations have focused on diversity profiles. In a pioneering study, Kim et al.compared Terra Preta soils to adjacent pristine soils and found the Terra Preta to contain 25% greater bacterial species richness and hundreds of novel bacteria taxa . Also, Kolton et al.found that changes in bacterial community structures were observed after soils were amended with a fresh, citrus wood-derived biochar. Pyrosequencing of 16S rRNA genetic markers revealed a decrease in the total numbers of proteobacteria when biochar was added to a soil and an increase in bacteroidetes, and particularly flavobacteria . These are noted chitin degraders that secrete antifungal compounds. Nielsen et al.utilized an ultra high-throughput sequencing platform to obtain 16S rRNA gene sequences of bacteria with low abundance, as low as 0.01% of the total population. Their results agreed with previous findings, that biochar applications resulted in shifts in abundances of various taxonomic groups and also indicated that taxa correlation patterns are altered with biochar application . A phospholipid fatty acid analysis by Steinbeiss et al.revealed that once incorporated into soils, biochar prepared from a protein rich feedstock selected for fungi while biochar derived from a cellulose-based feedstock selected for bacteria. In agreement with this finding, another recent study demonstrated a positive correlation between the C:N ratio of biochar-amended soils and soil total PL A’s and bacterial PL A’s, in particular . However, Jindo et al.report a negative correlation between C:N ratio and bacterial biomass in biochar-compost mixtures. Additionally, biochar products, particularly those prepared at low pyrolysis temperatures, commonly contain a large number of adsorbed volatile organic compounds that may affect microbial growth and plant responses to biochar . Soil enzyme activities are differentially affected by biochar application . Most notably enzymes with enhanced activities in alkaline conditions showed higher activity post biochar amendment . This is reasonable, as pH’s of many biochars tend to be alkaline, a property that is dependent on pyrolysis temperature. Furthermore, Harter et al.used molecular techniques to assay abundance and expression of bacterial genes involved in nitrogen cycling as affected by biochar application. They found that N2O reductase transcript numbers were increased when soils were amended with 10% biochar . This altered microbial activity is especially important when considering greenhouse gas emissions and reduced N2O emissions from soil, a common phenomenon reported with biochar application .

The F3 peptide is a 31-amino acid fragment of the high mobility group protein HMG2N. F3 preferentially binds to nucleolin, a shuttle protein that is highly expressed on the plasma membranes of tumor cells.Nucleolin- targeted delivery of drug-loaded nanoparticles using F3 peptide or nucleolin-specific aptamers has been previously demonstrated.For example, F3-functionalized PEG-PLA nanoparticles led to deeper tumor penetration into 3D glioma spheroids and prolonged the survival of mouse bearing intracranial C6 glioma when loaded with paclitaxel.In this work, we conjugated F3 to TMVlys following an established protocol .In brief, TMVlys was mixed with a maleimidePEG4-NHS bifunctional linker using 10 equivalents linker per TMVlys coat protein in 10 mM KP buffer for 2 hrs. The mixture was then purified by ultra centrifugation at 112,000 g for 1 h on a 40% sucrose cushion. The F3 peptide synthesized with a C-terminal Cys was then added to the mixture at 0.5 equivalents peptide per TMVlys coat protein and reacted for 2 hrs. The final TMVlys-F3 was purified with 10,000 MW cut-off Slide-A-Lyzer MINI dialysis units. A higher excess of the F3 peptides led to aggregation and a lower excess did not show sufficient conjugation yields.

Cy5-labelled TMVlys-F3 was also prepared for cell uptake studies. Using the aforementioned protocol, SDS-NuPAGE gel electrophoresis confirmed covalent attachment of F3 peptides,bato bucket as indicated by higher molecular-weight bands The TMV coat protein has a molecular weight of ~17.5 kDa; a slightly higher molecular weight indicates the addition of F3 peptide . ImageJ software was used to quantify the degree of F3 conjugation, and data indicate that over 20% of the TMVlys coat proteins was modified with F3 peptide. The A260:A280 ratios of Cy5-TMVlys-F3 and Zn-Por:TMVlys-F3 were 1.21 and 1.29, respectively, which are indicative of intact TMV preparations . Furthermore, transmission electron microscopy indicated that TMVlys maintained its structural integrity post chemical modifications .To assess the targeting efficacy, we compared the cellular uptake of Cy5-labeled TMVlys and TMVlys-F3 particles using HeLa cells. HeLa cells were chosen because this cervical cancer cell line is known to express high levels of nucleolin.For cell targeting studies, HeLa cells were incubated with 150,000, 300,000, or 750,000 particles per cell at 37°C and 5% CO2 for 3 h in DMEM supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin. Flow cytometry was performed as described above.

Flow cytometry showed an over 50% increase in cell uptake of conjugated Cy5-TMVlys-F3 particles . Compared to cells incubated with native TMV particles, the mean fluorescence intensity increased by 40-fold in the presence of nucleolin-targeted Cy5-TMVlys-F3 particles . Because flow cytometry does not provide insights into the fates of the nanoparticles, we also used confocal microscopy to study where the particles localized within the cells. HeLa cells were incubated with 6,000,000 particles per cell for 6 h in culture medium. Then cell membranes were stained with Alexa Fluor 555- conjugated wheat germ agglutinin and mounted with Fluoroshield™ with DAPI. Slides were imaged using a Leica TCS SPE confocal laser scanning microscope and the data was analyzed using ImageJ software. The confocal images were in agreement with flow cytometry, showing high cell interactions of the F3-targeted TMV formulation. The Cy5-TMVlys-F3 partiles appear bundled up at the surface of HeLa cells, where nucleolin is over expressed . Although others have shown intracellular trafficking of F3-functionalized polymeric nanoparticles,cellular uptake of Cy5-TMVlys-F3 was not observed in our study. This possibly indicates that the TMV’s high aspect ratio shape may not be suitable to be shuttled by nucleolin. Nevertheless, the accumulation of Cy5-TMVlys-F3 particles on the cell membrane may be advantageous for PDT as cell membrane targeting may prevent trapping of zinc porphyrin in endolysosomes and therefore enhance its cytotoxic efficacy. We moved on to explore the therapeutic efficacy of the F3-targeted Zn-Por drug delivery system. The enhanced cytotoxic efficacy of Zn-Por3+ loaded in TMVlys-F3 was reflected in the decreased survival of HeLa cells after white light treatment . TMVlys particles were loaded with 2,000:1 molar excess of Zn-Por3+ as described above prior to the conjugation of F3 peptides. From the MTT cell proliferation assay, the IC50 values for Zn-Por3+ in HeLa cells were equal to 0.034, 0.38 and 0.19 μM for Zn-Por3+ -TMVlys-F3, Zn-Por3+ -TMVlys and free Zn-Por3+ respectively. No cell killing was observed with drug-free TMVlys-F3 particles at maximum concentration.

Without light treatment, data indicated that Zn-Por3+ -TMVlys-F3 is non-toxic to HeLa cells . Compared to the data reported above, the efficacy of free Zn-Por3+ drug with white light treatment was 6-fold higher in B16F10 cells than in HeLa cells. This may be attributed to the biochemical differences between a mouse cell line and a human cell line. The drug activity decreased by half after loading into TMVlys-F3. This level of activity decrease after loading was not as significant as our previous data with TMV and TMGMV particles in B16F10 cells, yet the trend of decreased activity after loading into VNP did agree. Meanwhile, the targeted Zn-Por3+ -TMVlys-F3 particles showed a 5-fold increase in cell killing efficacy compared to the free drug. The increase PDT activity of Zn-Por3+ -TMVlys-F3 vs. Zn-Por3+ -TMVlys may be explained as follows: i) a significantly larger amount of particles targets cancer cells when using TMVlys-F3 vs. its native counterpart; and ii) TMVlys-F3 targets the cell membrane, light activation may lead to cell toxicity through cell membrane disruption, and iii) it is also possible that the PS cargo is released at the cell surface, and since the Zn-Por molecule is cell permeable and positive charged, cell uptake maybe favored – in contrast TMV without the F3 ligand accumulates in the endolysosomal compartment. In recent years, plant virus-based nanoparticles have been investigated for vaccine and immunotherapy applications to combat infectious diseases, cancers, and autoimmune diseases.Plant viruses are noninfectious to mammals and therefore are safter than their mammalian counterparts that are often used in oncolytic therapies.They can be manufactured in a cost effective manner and in large scales as viral nanoparticles as well as non-replicative viruslike particles devoid of their genomic payload.Cowpea mosaic virus VNPs and VLPs thereof have been proposed as in situ vaccine for cancer immunotherapy. The native form of CPMV consists of a bipartite ssRNA virus forming a 31 nm icosahedron with pseudo T = 3 symmetry. CPMV is made of 60 identical copies of a large and small coat proteins encapsidating RNA-1 and RNA-2 in separate but identical CPMV particles. When applied as in situ vaccine, the CPMV or eCPMV formulation is administered directly into a tumor to reprogram the tumor micro-environment and launch systemic anti-tumor immunity. While both CPMV and eCPMV demonstrated potent antitumor response in mouse models and canine patients,the RNA containing CPMV formulations demonstrated higher efficacy than eCPMV through the activation of additional cytokines and immune cells, which ultimately led to an extended survival rate of tumor-bearing mice.The proteinaceous nanoparticle presents danger signals that activate the immune system through pattern recognition receptors ,dutch bucket hydroponic and the presence of the RNA provides an additional danger signal.RNA activates TLR7/8, and induces type I IFN secretion, which increases the potency of the CPMV- based vaccines.This phenomenon was also reported using papaya mosaic virus as an in situ vaccine.While CPMV is not infectious to mammals, it remains infectious to legumes including Vigna unguiculataplants. From a translational point of view, it is thus important to develop RNA laden but non-infectious CPMV that is safe from to the environment and plants.

CPMV is stable in a variety of environmental conditions, such as temperature , pH , and in the presence of harsh chemicals, such as dimethyl sulfoxide.CPMV particles are not sensitive to certain standard methods of virus inactivation, including peptidase or hypochlorite treatment,but showed good response to ultra-violet light.Here we investigated UV treatment vs. chemical treatment of CPMV to render it non-infectious while maintaining its potent efficacy as a cancer immunotherapy. We compared β-propiolactone or formalin treatment with the previously reported UV inactivation method. These chemical treatment modalities are commonly used to produce non-virulent vaccines such as polio, hepatitis A, enterovirus, and influenza viruses vaccines.Of particular interests, these methods do not remove the RNA from the VNP, but rather create RNA damage preventing its transcription and translation, and therefore viral replication. UV light promotes RNA-protein crosslinking and dimerization of adjacent urcacils.βPL promotes the alkylation or acylation of cytidine, deoxyadenosine, and deoxyguanosine. βPL treatment also leads to a large extend of protein modifications; for example poliovirus proteins are more extensively modified by βPL than nucleic acid during viral inactivation.Specifically, βPL acylates and alkylates to a great extent cysteine, histidine, and methionine, and to a lesser extent with aspartic acid, glutamic acid, lysine, serine, threonine, and tyrosine.Lastly, formalin causes protein-protein and RNA-protein covalent crosslinking.To gain further insights into whether and to what degree RNA-protein crosslinking occurred, intact and denatured CPMV particles pre- and post-inactivation were analyzed by native gel electrophoresis and denatured samples were analyzed by SDS-PAGE. All gels were imaged after staining for proteins and nucleic acid under white light and UV light, respectively. In the native agarose GE, two distinctive protein bands were observed due to the presence of two electrophoretic forms of CPMV particles ; the fast form of CPMV is the result of a 24 amino acid loss at the C-terminus of the small coat protein due to proteolysis in plant cells.In all samples, nucleic acids and proteins traveled through the agarose gel together, confirming the entrapment of the RNA within the CPMV particles. CPMV treated with doses of 5, 7.5, 10 and 15 J cm-2 of UV showed signs of particle aggregation, as indicated by the presence of a smear instead of two distinct bands. Compared to CPMV, βPL-CPMV and Form-CPMV traveled farther through the gel proportionally to the concentration of βPL and formalin employed to inactivate CPMV. These results could reflect a change in overall particle charge due to the action of βPL and formalin on proteins and RNA. Samples were denatured by SDS-PAGE to separate the L and S coat protein subunits, visualized as single bands at 42 kDa and 24 kDa, respectively . The band intensity of UV-CPMV coat proteins decreased with increasing dose of UVradiation, indicative of the presence of particle aggregates. βPL-CPMV showed no sign of protein breakage or aggregation regardless of the dose of βPL used during treatment. In contrast, the higher the dose of formalin, the more inter-CP cross linking was observed at doses as low as 0.5 mM . GelRed staining was added to SDS-PAGE gels to assess the integrity of the RNA content of particles. RNA from UV-CPMV and Form-CPMV particles did not travel through the gel, most likely due to intra-RNA and RNA-coat protein cross linking; the samples remained stuck in the wells. RNA from βPL-CPMV matched the profile of that released from CPMV; however, at high βPL concentrations RNA breakage was observed – this is also consistent with UV/visible spectroscopy and SEC data as described above. Therefore, data indicate that at doses higher than 1 J cm-2 UV light or 1 mM βPL or formalin RNA and protein modifications and crosslinking occur; at high concentration of UV light, CPMV particle aggregation is observed; and at high βPL concentrations, RNA breakage is indicated. Agarose gel electrophoresis confirmed that leaves infected with CPMV inactivated through UV treatment at doses of 7.5 J cm- 2 or higher were indeed effectively inactivated and CPMV infection was not detectable by RTPCR . Similarly, formulations treated with 50 mM βPL-CPMV and 1 mM FormCPMV were confirmed to be fully inactivated. It is worth mentioning that leaves inoculated with 10 mM βPL-CPMV showed no visual symptoms of infection, yet these leaves tested positive by RT-PCR – the latter is a more sensitive assay. Based on the inactivation studies and infection assays in plants, for all subsequent experiments, we used the inactivated CPMV formulation obtained from 7.5 J cm-2 UV, 50 mM βPL, and 1 mM formalin. At these doses UV-CPMV was structurally intact but to some degree aggregated, and there was evidence of RNA-RNA and RNA-protein crosslinking. βPL-CPMV remained structurally sound and monodisperse but its RNA was severely damaged. FormCPMV also retained its size and monodispersity even though protein-protein, protein-RNA, and RNA-RNA cross linking occurred.

Bioavailability of CVTMGMV and free CV in C. elegans was investigated in liquid culture. C. elegans nematode motility was classified as either totally immobilized, impaired motility, or completely mobilized nematodes. To illustrate the data that was collected, a series of snap shots of C. elegans incubated with no treatment, 10 μM CV, and 10 μM CVTMGMV was taken every second for 60 seconds. The corresponding videos can be found in the supporting information. Figure 3.5 A-C illustrates the nematodes observed after 3 h of incubation. Five nematodes were selected in each treatment regime and pseudo-colored to illustrate their motility. Untreated C. elegans showed no impaired motility . For example, the nematode colored in pink moves across the frame within the 40 sec interval, while other nematodes disappear from or appear in the frame during that time interval. Although the motility of these nematodes is evident, most nematodes do not travel far but rather move within a restricted area,blueberry packaging such as the nematode colored in yellow. C. elegans treated with 10 μM of CV or 10 μM of CVTMGMV behaved differently and showed severe motility impairment .

All pseudo-colored nematodes in Figure 3.5 B+C were paralyzed or dead and did not move. However, this is not true for all nematodes, as a population of nematodes showed little to no motility impairment when treated with CV or CVTMGMV. From the imaging data there were no apparent differences between the two treatment groups, free drug vs. CVTMGMV .To quantitatively analyze the motility effects of CV on C. elegans, nematodes were treated with various concentrations of free CV, CVTMGMV, or TMGMV for 24 h at 22°C. At specific time points nematodes were observed under a white light microscope and the percent of affected nematodes and was quantified as a function of time. The effective concentration , defined as the concentration of CV at which half of the maximum immobilization of C. elegans was reached, was determined for free CV and CVTMGMV . Sixty percent of nematodes treated with 100 μM CV were paralyzed/dead within 1 h and no further improvements were observed within 24 h . When treated with 10 μM or 1 μM of CV, only ~30% or ~15% of nematodes were paralyzed/dead within the first hour, respectively. In those cases, maximum efficacy was observed after 6 h of incubation, when ~50% and ~25% of nematodes were affected. In both treatment regimes, a decrease in efficacy was observed after 6 h of incubation – this phenomenon may be explained because remaining unaffected population of nematodes continued to progress through their life cycle; consequently eggs were laid and nematodes hatched, which led to an overall increase in population and a decrease in percent of nematodes affected by the treatment. Furthermore, it is possible that at low doses of CV, nematodes are able to recover and slowly become mobile again.

At doses of CV lower than 1 μM, there was no significant effect on nematode motility compared to the untreated population. The EC50, defined as the concentration of CV at which half of the maximum immobilization of C. elegans was reached, was quantified at various time points and was determined to be 3.7 μM. CVTMGMV showed a similar trend to free CV , and, as expected, TMGMV alone did not show any nematicide properties When treated with 100 μM of CVTMGMV, ~40% of nematodes were paralyzed/dead within the first hour, and maximum efficacy was reached in the first 3 h. Therefore the efficacy of 100 μM of CV and CVTMGMV is identical after 3 h of incubation. However, when the concentration of CVTMGMV was dropped to 10 μM, the maximum efficacy was ~30% and was reached after ~8 h of incubation. Interestingly, CV release from CVTMGMV in nematode media conditions revealed a half-life of 8 h , thus supporting the idea that CV was released from TMGMV and made available to treat the nematode infestation. All studied concentrations of CVTMGMV lower than 10 μM led to no significant treatment of the nematode infestation compared to the untreated population. The calculated EC50 of CVTMGMV is 13.8 μM, which is approximately 4 times greater than the EC50 of free CV. While reduced efficacy was observed in the petri dish experiments, I envision that CVTMGMV will outperform free CV in the field based on the enhanced drug delivery aspect . Next, I set out to understand the bio-distribution of CV in the nematodes. I prepared fluorescently labeled TMGMV and analyzed whether TMGMV would interact with or be ingested by C. elegans.

Briefly, diazonium coupling and click chemistry was used to conjugate a Cy5 dye to TYR side chains on TMGMV, as structural studies indicated that TYR2 is solvent-exposed . I conjugated ~160 dyes per full length TMGMV, or about 7.5% of CPs were modified with Cy5 . We have previously demonstrated that a minimum conjugation of Cy5 to ~8% of TMV coat proteins is sufficient to yield maximum fluorescence intensity,237 thus the prepared samples were thought to be sufficient for imaging experiments. Fluorescent TYR-Cy5TMGMV was incubated with C. elegans nematodes for 3 h at 22°C and subsequently analyzed by fluorescent microscopy . Results indicate that nematodes ingest the proteinaceous TMGMV carrier and that while TMGMV distributes throughout the entire nematode body, the majority of TMGMV accumulates in the gastrointestinal tract. A soil mobility test was designed to establish the leaching of CVTMGMV and free CV in soil. Briefly, top soil was packed in a plastic column up to a height of 4 cm and saturated with deionized water. CVTMGMV or free CV was applied atop the soil columns, followed by DI water. Fractions were collected from the soil column, purified, and analyzed by UV/visible spectroscopy for the presence of TMGMV and CV. The λ260 and λ280 wavelengths were monitored to quantify the amount of TMGMV that leached through the soil. A background 260:280 absorbance was observed in a CV soil leaching column, which most likely corresponds to the absorbance of organic matter present in top soil . CVTMGMV showed enhanced mobility over free CV in the soil column,blueberry packaging box eluting from the column at high concentrations in the 5th to 15th elution fractions . In stark contrast, the efflux of CV from the soil column was delayed until the 25th to 50th elution fractions at a concentration 3.6 times lower than CVTMGMV . CV is hydrophobic and has a strong binding affinity to soil particles rendering the drug mostly immobile in soil, which explains the delayed efflux and lower concentrations eluted. Taken together, the data demonstrates the potential of TMGMV as a drug carrier to enable penetration of CV or other nematicides through soil to reach nematodes feeding on the roots of plants. In this study, I have demonstrated the potential of tobacco mild green mosaic virus as a carrier for anthelmintic drugs, such as crystal violet , to treat plants infected with parasitic nematodes. After careful analysis of the TMGMV structure, I identified solvent exposed TYR2 on the exterior surface enabling chemical modification. I also identified solvent exposed carboxylates, GLU145 and ASP66 on the exterior surface and GLU95 and GLU106 on the interior surface, and established the chemical addressability of these residues. I also showed the potential for electrostatic encapsulation of positively charged guest molecules in TMGMV. Further studies are needed to identify which of the identified GLU and ASP residues are chemically reactive. Electrostatic drug loading using crystal violet was achieved, yielding TMGMV carriers loaded with ~1500 CV per CVTMGMV nanocarrier.

Treatment efficacy, while lower compared to free drug, was demonstrated using liquid C. elegans nematode cultures . Diffusion experiments revealed significantly increased soil mobility of CVTMGMV vs. free CV; the latter was unable to sufficiently diffuse and disperse through soil. Overall CVTMGMV demonstrates efficacy and superior soil motility, and as such makes a promising platform technology as a drug carrier targeting agricultural application. Pesticides are needed to protect our crops and thus maximize crop yields.However, the efficacy of chemical pesticides is limited by their instability and strong binding to organic matter in soil, which can render them inactive or prevent their accumulation at the root level, where many pests reside.Large doses are applied to compensate, resulting in the accumulation of pesticide residues in soil, water and agricultural products.Long-term exposure to these chemicals is a risk to human health and threatens the biodiversity of an already fragile ecosystem.Advances in nanotechnology have led to the development of more effective drug delivery and medical imaging methods , and the same innovations are now being applied to smart agrochemical delivery systems, known as nanopesticides.These involve the use of nanomaterials for the adsorption, encapsulation or conjugation of pesticides, improving the biodegradability, stability, permeability and dispersion of the active pesticide ingredient. Nanopesticides have a much greater surface area than conventional pesticides, increasing their potential for interaction with target pests at lower doses. The encapsulation of pesticides within nanoparticles also prevents premature degradation and the risk of direct human exposure to the active ingredient. There is also evidence that nanopesticides and conventional pesticides differ in their environmental behaviour, so it is necessary to understand the fate of nanopesticides in detail in order to ensure they comply with regulatory guidelines and legislation.Most of the nanopesticides investigated thus far are based on synthetic or natural polymers, metallic compounds or liposomes, which tend to persist in the environment.As a biodegradable alternative, nanopesticides can be developed from plant viruses.One example, already EPA-approved, is the application of Tobacco mild green mosaic virus as the herbicide Solvinix, which is produced by BioProdex for deployment against invasive tropical soda apple weed in the state of Florida.The safety profile and possible risks of TMGMV have been reported.TMGMV cannot self-disseminate and is not transmitted by vectors such as insects, seeds, or pollen. Mechanical transmission through insects or contact between plants is thus the only route of transmission. Only plants of the Solanaceae are susceptible to TMGMV infections. Therefore, TMGMV offers a good safety profile for crops that are not part of the Solanaceae. Nonetheless, TMGMV as well as other plant virus-based systems could be inactivated through ultraviolet radiation for safe use on any crop.To investigate the potential of plant virus nanoparticles and virus-like particles as nanopesticides in more detail, I compared the behaviour of three viruses and two synthetic particle formulations in soil column experiments and computational models, as a way to gauge their ability to deliver pesticides to the rhizosphere and thus prevent infestation by root pests . I tested two VNPs, based on the rod-like TMGMV and the icosahedral Cowpea mosaic virus , and a virus-like particle based on Physalis mosaic virus . These were compared to mesoporous silica nanoparticles and a poly formulation , which have already been developed as synthetic nanopesticides.TMGMV was oβtained from Bioprodex, DegraFluorex Fluorescent PLGA nanoparticles were purchased from Phosphorex, and MSNPs functionalized with propylcarboxylic acids were obtained from Sigma-Aldrich. I resuspended 3 mg ml-1 of PLGA and 1 mg ml-1 of MSNP in distilled water and sonicated them using a Branson 2800 device for 10 min to obtain homogeneous solutions. CPMV was propagated in Burpee black-eyed pea plants and purified as previously described.PhMV VLPs were prepared in ClearColi BL21 cells as previously described.USDA Permits were obtained for any work with plant viruses. TMGMV comprises 2,130 identical coat proteins arranged helically around a single-stranded RNA genome, forming a hollow rigid rod measuring 300 × 18 nm with a 4-nm internal channel.The external surface features two solvent-exposed tyrosine side chains , which can be functionalized using diazonium coupling reactions. I used sulfo-Cy5-azide to modify these Tyr residues as previously described.10 Briefly, I mixed 25 μl 0.68 M 3-ethynylaniline with 75 µl 3 M sodium nitrite in 400 μl 0.3 M p-toluenesulfonic acid monohydrate for 1 h on ice. I then added 15 equivalents of the resulting diazonium salt to 2 mg ml-1 TMGMV in 10 mM borate buffer for 30 min on ice. The particles were centrifuged at 112,000 g for 1 h on a 30% sucrose cushion to separate the TMGMV-alkyne particles from the excess DS. TMGMV-alkyne was resuspended in 10 mM KP buffer overnight before adding sulfo-Cy5-azide via a Cu-catalysed alkyne-azide cyclo-addition reaction. I added two equivalents of Cy5 per coat protein to 2 mg ml-1 TMGMV-alkyne in the presence of 2 mM aminoguanidine, 2 mM L-ascorbic acid sodium salt and 1 mM copper sulfate in 10 mM KP buffer on ice for 30 min.

The reduced bio-degradation in the SB microcosms may have resulted from the ~40% higher carbon content in the SB microcosms, which would be expected to increase the soil-water distribution coefficient by a comparable amount. Reduced TCS concentration in soil pore water would be expected to slow bio-transformation, potentially in a nonlinear fashion. Another possible contributor to the slower degradation of TCS in SB is the greater availability of alternative, likely more easily degradable, carbon sources in SB than soil microcosms, reducing the use of TCS as a substrate. Selective bio-degradation of one carbon source, and inhibition of the degradation of other chemicals also present, has been observed for mixtures of chemicals in aquifers . To assess which of these mechanisms was controlling, measured Freundlich isotherm parameters for TCS adsorption on bio-solid amended Yolo soil were used to calculate equilibrium pore water concentrations in the soil and SB microcosms over the course of the experiment. Using estimated pore water concentrations of moistened soil and SB samples,pe grow bag instead of total soil concentrations to perform half-life calculations, resulted in modest increases in the rate constants and decreases in half-lives of soil samples and did not narrow the significant gap between half lives in soil and SB .

This suggests that the primary reason for the slower degradation of TCS in bio-solid amended soils is the increase in more labile forms of carbon because organic material is highly porous and has a lower particle density. Previous research shows that TCS biodegrades within weeks to months in aerobic soils , although Chenxi et al., found no TCS degradation in bio-solids stored under aerobic or anaerobic conditions, Kinney et al., observed a 40% decrease in TCS concentrations over a 4-month period following an agricultural bio-solids application. Because the slopes of the lines in Fig. 1 are not significantly different as a function of spiking level , the slopes were averaged for each treatment type, yielding apparent first order rate constants of 0.093±4% d−1 for soil samples and 0.024±41% d−1 for SB samples where the percent error represents the relative percent difference between the 10 mg/kg and 50 mg/kg degradation curves. These apparent rate constants translate to half-life estimates of 7.5 d in soils and 29 d in bio-solid amended soil. The estimated half-life of TCS in soil is within the range of previously reported half-lives of from 2.5 to 58 d in soil . The half-life determined here in bio-solid amended soils is lower than the one available literature value of 107.4 d . The microbial biomass decreased in the TCS spiked samples after 7 or 30 days of incubation in comparison with the unspiked controls, for both soil and SB, and the decline was statistically significant at 50 mg/kg .

Although exposure to TCS caused declines in biomass in both soil and SB microcosms, the total microbial biomass was two times higher in SB than soil probably due to the increased availability of nutrients and/or possibly due to addition of bio-solid associated microorganisms in the latter . The total number of PLFAs ranged from 42–47 in soil and 48–59 in SB . No significant change in numbers of PLFAs was evident with increasing dosage of TCS for any incubation time suggesting that TCS addition did not adversely affect microbial diversity. Microbes respond to various stresses by modifying cell membranes, for example by transforming the cis double bond of 16:1ω7c to cy17:0, which is more stable and not easily metabolized by the bacteria, reducing the impact of environmental stressors . Consequently, the ratio of cy17 to its precursor has been employed as an indicator of microbial stress that has been associated with slow growth of microorganisms . Increases in this stress biomarker were observed in both soil and SB samples as TCS concentrations increased , suggesting that TCS has a negative effect on the growth of soil microorganisms. The overall level of cy17 to its precursor is lower in SB than soil samples, suggesting that nutrients contributed by the bio-solids reduce stress on the microbial community. Our study agreed with a previous study showed that carbon added to soil led to a reduction in the cy17 fatty acid TCS additions, however, increased the stress marker compared with that detected in the corresponding samples with no added TCS.

A broader implication of this result is that presence of bio-solids may mitigate the toxic effects of chemicals in soil, or chemicals added in combination with bio-solids, on soil microbial communities. Groupings of microbial communities, based on CCA analysis of their composition as estimated by PLFA, were distinguished primarily by whether they were in soil or SB treatments and secondarily by time since spiking . To isolate the effects of bio-solids and TCS amendments on microbial community composition, the data was analyzed using pCCA considering TCS and bio-solid amendment as environmental variables, and incubation time as a covariable . This confirmed the results of the CCA indicating that the strongest determinant of microbial community composition was addition of bio-solids to soil. TCS concentration, on the second axis, described only 3.6% of the variation, showing TCS effects were overshadowed by the effects of bio-solid amendment. Bio-solid amendments caused an approximately two-fold increase in PLFA biomarkers for Gram-positive bacteria, actinomycetes and eukaryotes in SB compared to soil samples . Even larger increases were observed in biomarkers for fungi and Gram-negative bacteria, which were up to three times higher in SB than soil. Again, these changes were likely due to increased nutrient availability in the bio-solid amended samples and/or the biomass added along with the bio-solids, growing bags consistent with previous studies that found that the fatty acid 18:2 ω6, 9c and monounsaturates were increased by addition of these materials . The effect of TCS on microbial community composition was greater in soil than SB. Spiking with 10 or 50 mg/kg TCS decreased the abundance of Gram positive and Gram negative bacteria as well as fungi, with reductions ranging from 14 to 27% by day 30. Additionally, actinomycetes, which are Gram positive bacteria, were reduced in the 50 mg/kg TCS samples after 30 days of incubation . Eukaryotes were negatively affected after 7 and 30 days of incubation at both concentrations of TCS in soil but not SB samples. Biomass results for all microbial groups were consistent in suggesting that the presence of bio-solids mitigated the potential toxicity of TCS. It is important to note that the spiking levels used here are similar to levels found in the upper half of U.S. bio-solids, but would be unlikely to be achieved in bio-solid amended soils even after continued long term application. Therefore, the effects observed at the 10 or 50 mg/kg spiking levels should be viewed as a conservative upper bound on potential effects expected in the field. In addition, since all of the results in this study are based on an observation period of 30 d, the extent to which the observed effects persist is not known. Future studies should, in particular, investigate longer term changes in community structure in response to addition of bio-solids both with and without specific contaminants.

Over the past 30 years, nanoparticle engineering has led to the development of novel delivery systems for active ingredients with medical, veterinary, and agricultural applications. The increasing cost of research and development combined with the growing number of competitive manufacturing entities, short patent cycles, and the tightening regulatory guidelines for active ingredients, have made it difficult to bring new formulations from the bench to the market.Furthermore, the efficacy of many drugs is limited by their low solubility and/or stability, as well as off-target effects following systemic delivery. For example, cancer therapy is often unsuccessful due to the toxicity of cancer drugs towards healthy cells and/or the development of resistant cells over expressing efflux transporters and multi-drug-resistance proteins.The resulting low bio-availability of the active ingredient in the tumor requires the administration of larger doses to ensure the drug concentration stays within the therapeutic window, which in turn increases off target toxicity. Nanocarriers can address this challenge by delivering active ingredients via the enhanced permeability and retention effect, a well-established phenomenon based on the combination of leaky vasculature and poor lymphatic drainage at the tumor site.The EPR effect only increases the tumor homing of nanoparticles by two-fold compared to normal tissue,so nanoparticles can also be functionalized with targeting ligands, aptamers, antibodies, or antibody fragments to promote their binding to receptors overexpressed on tumor cells or in the surrounding extracellular matrix.The entrapment of active ingredients in nanocarriers also reduces the clearance rate via renal elimination and phagocytosis, which increases the active ingredient circulation time and therefore its therapeutic longevity. The medical and veterinary applications of nanocarriers are analogous, but only experimental veterinary applications have been reported.Most research in veterinary drug delivery has focused on diseases in animals that can be translated to humans. However, the importance of animal welfare per se is increasingly important to consumers, and nanocarriers that improve the efficacy and safety of active ingredients are demanded in the context of companion animals such as cats, dogs and horses, as well as farm animals such cattle, sheep, swine and poultry.Pet owners consider companion animals as an extension of the family and are willing to pay their bills, including the high cost of cancer treatment, with the cost of veterinary care in the USA therefore rising from $7 billion in 2001 to $19 billion in 2019.This increase most likely reflects a combination of inflation, high drug costs, better treatment options , and an increased willingness to care for pets. In contrast, the food industry works with low profit margins and would only treat animals suffering from temporary and low-risk diseases, such as infections.Veterinary nanocarriers must therefore combine low costs with the release of active ingredients for sustained periods to minimize the frequency of animal handling and improve therapeutic efficacy. For example, animals are often subject to bacterial infections, and a nanomedicine approach could achieve the targeted delivery of drugs to pathogens, killing them on demand. This avoids the unnecessary use of antibiotics, which can encourage the emergence of resistant strains. The controlled delivery of agrochemicals and nutrients to plants is conceptually similar to drug delivery in humans and animals. However, agricultural delivery takes place in an open field, with variable weather and geographic features and no specific transport pathway to the target, in contrast to the closed and regulated nature of the bloodstream. Nanocarriers can be administered via the foliage, where they are taken up passively through stomata and any wounds, or can be transported through the soil and taken up via the roots.Among the agrochemicals that can be delivered using nanoparticles, pesticides are particularly suitable candidates because they are effective at very low doses but are difficult to apply in such small amounts due to their non-uniform distribution in the field.To compensate, the active ingredient can be diluted within a mixture of liquid or solid diluents. However, the active ingredient is often unstable, sparingly soluble, and binds with high affinity to soil particles, thus reducing its efficacy against target pests and increasing the amount required to achieve an effective dose.In an analogous manner to the off-target effects caused by systemic drugs, the persistence of large quantities of pesticides in the environment is toxic to other species, and contaminates the soil and groundwater leading to health problems in domestic animals and humans, including cancer and infertility.In one strategy, the active ingredient is enveloped in organic or inorganic coatings for protection against photolysis or bio-degradation, allowing the controlled release of the ingredient.But even microencapsulation is limited by the poor chemical and thermal stability of the capsules, and degradation promotes the acidification of soil, which can impair its fertility. As discussed in more detail below, these drawbacks can be addressed by a new generation of nanocarriers based on polymers, lipids and other materials. The definition of a nanomaterial is not yet harmonized, but the International Organization for Standardization defines nanoparticles as objects with dimensions of 1–100 nm, because the physicochemical properties of the material at this scale differ from the bulk material. Unfortunately, this ISO definition excludes most nanomaterials that are relevant in the medical, veterinary, and agricultural sectors.

In this chapter I have presented an attempt to estimate the effect of rural out-migration on rural wages. I find a strong positive and robust effect of rural out-migration on rural wages in Brazil during the period 1991-2000. Using a cohort analysis my results suggest that rural out-migration flows between 1991 and 2000 have increased rural wages in Brazil by 6.5%. One concern is that rural out-migration may be accompanied with changes in the workforce composition in rural areas since those who migrate do not constitute a random sample of the initial rural population. I find that changes in the workforce composition do account for some of the wage increase due to larger out-migration rates. Controlling for observable measures of workforce composition such as educational attainment and gender composition, I find that the wage effect of rural out-migration flows between 1991 and 2000 drops from 6.5% to about 3%. Due to data limitation, the analysis in this chapter did not measure the short-run effect of rural out-migration on wages by taking into account dynamic adjustments in physical capital. Future research in this areas could she light on which rural population groups lose or gain in the short-run as rural out-migration increases.Because of their dependence on rain-fed agriculture a large proportion of households in developing countries are particularly vulnerable to rainfall shocks.

Moreover the usual mechanisms for smoothing income or consumption may be missing or limited in such economies. In addition,flower bucket since shocks such as weather and pests are likely to affect the income of all households, they cannot be insured locally through non-market mechanisms. Households and individuals inability to transfer resources across time and states of the nature may lead them to adopt coping strategies that are detrimental to asset and human capital accumulation. For instance, a negative income shock may lead households to draw on monetary or liquid savings to smooth income and consumption. Households who experience a negative shock can also smooth income by increasing labor supply or reducing spending in some food or investment groups. Increasing labor supply may entail putting children to work, while the need to reduce spending can result in less health and schooling investment. Jensen finds that among households who experienced a rainfall shock in Cote d’Ivoire, enrollment rates and child growth drop considerably relative to one year before the shock. Jacoby and Skoufias find that household income fluctuations in India lead to year-to-year variations in school attendance. Beegle et al. find evidence of increased child labor following crop losses using longitudinal household data from Tanzania. Examples include Funkhouser for the debt crisis in Costa Rica, Thomas et al. who examines household response to the financial crisis in Indonesia and Rucci for the Argentine peso crisis.

However considerably less attention has been devoted to the medium to long-run consequences of income shocks on children’s schooling in developing countries. In the presence of state dependence, an income shock may have permanent effect on a children schooling. De Janvry et al. find strong evidence of state dependence in child school enrollment using a panel of households in rural Mexico. I contribute to this literature in this paper by examining to which extent short-run drops in enrollment rates affect medium-run enrollment decisions and years of education using a period of drought in Southern Africa. This paper is related to Meng and Qian’s analysis of the long term link between famine and educational attainment in China; but it also differs in several ways. The two papers have different notion of “long term”. While Meng and Qian analyze the impact of the 1959-1961 Famine in China over 30 years after, my medium to long-run analysis is carried out within the 10 years following the droughts. This allows me to observe individuals before they have completed their schooling which, in turn, permits an analysis of their school enrollment decisions adding to our understanding of their educational attainment. Following this approach, I show that individuals exposed to the droughts tend to stay in school at older ages, and that this leads to partial catch-up in educational attainment, especially in regions where the intensity of the shock was not too high. The policy implication of this finding is that simple means testing could help target policies aimed an dampening the negative effect of agricultural shocks. In 1991/1992 and 1994/1995, the southern region of Africa experienced two major droughts.

The 1992 drought was qualified as one of the most devastating droughts in the region and followed a 60-year low rains and over two million cattle lost. Figure 3.1 shows the logarithm of the ratio between rainfall and its mean between 1940 and 1995. The graph confirms that during this 26 year period rainfall was lowest in 1992 and 1995. In 1991/1992 rainfall was about 65.8 per cent of average rainfall in the sample period, while in 1994/1995 rainfall was about 66.5 of average rainfall. These low rains have had a large negative impact on food production. Figure 3.2 shows an index of food production in Zambia between 1960 and 2005 and its deviation from a quartic time trend. In 1991/1992 food production was 14 per cent below the trend. While 1992/1993 and 1993/1994 were relatively good years, in 1994/1995 food production dropped by 8 per cent relative to the estimated production. 2 . In this paper I use data from Zambia to explore the short-run and medium-run consequences of rainfall shocks on children’s schooling. I use three cross-sections of the Zambian Demography and Health Survey, one collected during the first months of the 1991/1992 rainy season and two other surveys in 1996 and 2001/2002, to investigate the impact of the droughts on school enrollment and years of schooling of children exposed to the drought. A major drawback of the ZDHS is the absence of measures of income and consumption,square flower bucket or information on children’s time allocation or work inside or outside of the household. Thus I focus on a reduced form analysis where I compare the schooling of school aged children affected by the drought with children of the same age-group interviewed before the drought. I control for trends in schooling using enrollment and years of schooling of older youth and adults. The estimates using this strategy are biased if other aggregate shocks correlated with schooling took place during the same period. To address this issue, I employ a triple difference strategy to confirm that the effects on schooling can be attributed to the drought. I use rainfall data from actual rainfall gauges and exploit variation across provinces in the intensity of rainfall deficit during the peak of the drought. My triple difference approach consists in comparing differences in schooling across highly and moderately affected provinces between school-aged children affected by the drought and children of the same age-group interviewed before the drought. I find that exposure to the drought reduced enrollment rates by 10 percentage points and years of schooling by 8 percentage points in the short-run. I also find some evidence of partial catch-up in the medium-run in provinces moderately affected by the drought which suggests that children exposed to the drought remained in school at older ages. However, in the provinces most affected by the drought, I find no evidence of such accumulation of delayed education.

Within such provinces, young children who were in school during the drought were up to 7 percentage points less likely to be enrolled in school five to six years after the drought. Given the existing literature, the medium-run consequences on delayed entry in the job market, forgone earnings, lower wages might be large. These findings have important policy implications. They suggest that technologies to reduce the impact of rainfall shocks and safety nets may have large benefits in reducing delays and increasing the rate of human capital accumulation. Moreover education policies should target regions and individuals exposed to agricultural or income shocks in order to limit drops in enrollment rates and facilitate the return of students who temporarily left school. The remainder of this chapter is organized as follows. In section 3.2 I develop a simple dynamic model of income shock and investment in school to motivate the empirical analysis. Section 3.3 describes the timing of the data collection and the rainfall season in Zambia and some basic summary statistics. Section 3.4 explains the empirical strategy, and section 3.5 presents the main results and some robustness checks. Section 3.6 concludes. The onset of the rainy season in Zambia is normally during October or November. Rains are usually recorded up to March or April of the following year. Since I am using the 1992 ZDHS to control for an age-group’s pre-drought schooling, my estimates of the impact of the 1991/1992 and 1994/1995 droughts will be biased if, for instance, households started reducing investment in schooling during the first months of the raining season. My estimates will also be biased if data collection occurred too early relative to the raining season for households to be affected and to adjust their schooling decisions. As a consequence it is important to understand the timing of the ZDHS data collections relative to the onset and offset of the raining season in Zambia; the diagram below provides a basis for this. Data collection of the ZDHS-1992 was carried out between January and May 1992 meaning that the issue that households may have started reducing investment in schooling during the 1991/1992 raining season. In the analysis below I present a robustness check to judge the severity of the concern. The robustness of the results are tested by restricting the ZDHS-1992 sample to households surveyed during the 1991/1992 raining season, period during which the impact of the drought on harvests was not yet felt. On the other hand, the ZDHS-1996 data was collected between July and December 1996, so 3 months after the end of the poor 1994/1995 raining season suggesting that I should be able to capture the impact of the two droughts on schooling. Primary and secondary education in Zambia is divided in three levels : Primary education , Junior Secondary , and Upper Secondary. My analysis focus on school enrollment, and years of schooling for individuals 6 to 40 years old living in rural areas. I first present some graphical analysis of educational attaintment before and after the drought to motivate the empirical strategy. Figures 3.3 to 3.6 report school enrollment rates and average years of schooling by age for the ZDHS in 1996 and 1992. The general patterns reported here are similar using non-parametric regression of enrollment rate and years of schooling on age. Figures 3.3 to 3.4 show enrollment rates in the full sample and for males and females separately. Enrollment status is only recoded for residing household members between 6 to 24 years old. Figure 3.3 shows that for individuals from 6 to 13 years old, current enrollment in school is higher in 1992 relative to 1996. For individuals ages 15 to 19 years there is a less clear ranking of enrollment rate between individuals interviewed in 1992 and 1996. For individuals 20 to 24 years old, the difference in enrollment rates in 1992 relative to 1996 is much smaller. Turning to years of schooling, figures 3.5 to 3.6 show years of schooling in the full sample, and for males and females separately. For individuals 6 to 13 years old, years of schooling is larger for individuals interviewed in 1992 relative to those interviewed in 1996. There is no difference in education between 1992 and 1996 for individuals 14 to 30 years old. For older cohorts, individuals interviewed in 1996 have slightly higher schooling consistent with the overall increase in education over time. Taken together these graphics suggest a strong negative impact of the drought on individuals that were 6 to 13 years old in 1996. This cohort of children is a judicious group of interest to analyze the schooling consequences of the drought for two main reasons. First, the threshold of 13 years old is important as it represents the final year of primary education for children who started school at age 6 and had a normal progression through school. Moreover about 91 per cent of the sample has completed 7 years of schooling or less. Second, exposure to drought for children ages 2 to 5 years old is likely to affect children’s educational attainment.

From 2000 and 2015, the organic production of spring mix lettuce increased 153% in Monterey County. Short-term lettuce data in the County reflect this trend. Since the passage of the 2012 Ag Waiver , organic head lettuce production has increased 155%, from 112 acres to 174 acres, organic romaine lettuce has increased by roughly the same percentage, from 2,750 to 4,096 acres, yet organic leaf lettuce production in the county has increased decreased slightly from 1,088 acres to 1,066 acres. An upward trend in organic production is also true for the two other crops in Monterey County for which there are longitudinal data—organic cauliflower has more than quadrupled, from 180 acres to 780 acres between 2000 and 2013, and organic strawberries production has grown exponentially, from 48 acres in 2000 to 2,082 acres in 2015. Despite the appeal of using fewer pesticides for human health, the environment and higher profit margins, a possible drawback associated with less pesticide use and/or organic production is increased pest damage resulting in crop loss. The amount of crop loss depends greatly on the pest pressure in a particular area and crop type and stage. Even within the same cropping system, pests can have varying levels of destruction.

For example,procona valencia cabbage maggots can cause yellowing, retarded growth or even plant death on brassicas , but in some propitious situations , the same brassica plants could survive cabbage maggot infestations unimpeded . The severity of pest damage can also differ within a particular region, as it does with cabbage maggot in the Salinas Valley . For example, the acceptance of pest pressure on farms could also greatly vary. The two survey responses that referenced pest damage, both from San Luis Obispo County and both who previously used diazinon demonstrate the varying degrees of frustration growers have with accepting crop loss. One respondent who previously used diazinon to control beetle populations shared his sentiments of surrendering to the pests, “We are accepting cucumber beetle damage on annual crops.” The second survey response offered a more exasperated reaction to increasing pest pressure, alluding to the fact that he wished there was alternative pesticide to use, “There is no substitute. The ants are thriving.” The two responses could be representative of varying degrees of pest pressure on two different targeted pests , different value systems, or different financial circumstances allowing one farm to accept pest pressure more readily than another. Using fewer pesticides, however, does not automatically mean a farm will experience more pest damage and lower yields . For example, a study comparing organic and conventional apple production in the Central Coast showed not only increased profits from transitioning to organic production, but also increased yields .

Additionally, a recent study in Nature found that organic farming methods promote a stronger pest control among natural enemies as well as yield larger plants than management practices typical under conventional farming systems .The costs of chlorpyrifos and diazinon also could have played a small part in some farmers abandoning their use. Clearly, data on chlorpyrifos and diazinon pricing varies substantially based on the size and cropping system of the agricultural operation and the volume discounts that large farms might receive. UC Extension, however, has estimated operating costs in their detailed Cost and Return studies, including specific material and labor costs related to insecticide use for a variety of California crops. In 2009, Smith and his colleagues at UC Extension published a Cost and Return study for leaf lettuce producers in the Central Coast region. This report estimated roughly 1 lb/acre of diazinon use on lettuce at a price of $10.45/acre. Compared to the costs of other insecticides, such as Radiant SC , or other herbicides, such as Kerb 50W , diazinon was a minor cost, and only 4% of overall insecticide expenditures . Additional costs associated with diazinon include cultural costs . In the report, these costs were estimated as an aggregate totaling $128/acre for several pest control agents, including diazinon. Weighing these costs against estimated net returns per acre is complicated by the range of farm productivity and prices received for lettuce; for example, net returns for a head lettuce farm producing 400 12-3 count cartons per acre at an average market price of $11/acre was estimated around $-2,407/acre, whereas a production of 1000 12-3 count cartons per acre at the same price was estimated to yield $111/acre in net gains.

None of the recent Cost and Return studies on broccoli in the Central Coast include estimations on chlorpyrifos use in their calculations. However, a UCE study on a related crop, cauliflower, estimates about 7.00 lb/acre of chlorpyrifos is needed for root maggot control, the target pest for both cauliflower and broccoli. The cost of the chemical was valued at $2.80/acre for a total of $19.60/acre . Compared to total operating costs for cauliflower and broccoli , expenditures on chlorpyrifos for this specific pest were relatively small . As with lettuce, net returns on broccoli vary substantially by productivity and price. A farm producing 545 boxes/acre receiving an average price of $6.80/box had an estimated net loss of $569/acre whereas a farm producing 785 boxes/acre receiving the same price was also in the red with a net loss of $112/acre. These data suggest that cost was not a persuasive factor in growers’ decision to cutback on chlorpyrifos and diazinon use. The lack of survey responses highlighting cost to be a major impetus in decision-making corroborate with these data; only one survey respondent cited the cost of diazinon and chlorpyrifos as playing a part in his decision-making to stop using them.In the Central Coast region, broccoli has historically been one of the top three crops with the heaviest use of chlorpyrifos, and lettuce is the chief crop with the highest diazinon use. The region’s year-round mild climate offers the ideal growing conditions for these cool season crops; however, the cool, wet weather is also favorable to cabbage maggots ,flower bucket the predominant target pest of chlorpyrifos on broccoli. Monterey is the leading broccoli-producing county in the state, with 40 percent of the acreage and production . The Salinas Valley, located in Monterey County is the “salad bowl of the world,” producing 80% of the salad greens consumed in the U.S.

A closer look at chlorpyrifos and diazinon application on broccoli and lettuce in Monterey compared to other regions and the state as a whole sheds light on why crop type may be a pivotal factor in allowing Central Coast growers to give up chlorpyrifos, while growers in other regions haveheld on to it for survival, and why crop type did not have as powerful an effect on diazinon’s demise.Chlorpyrifos has been one of a handful of insecticides that broccoli farmers rotate into their pest management plan to slow potential pest resistance . According to pest advisors and the UC ANR IPM program, growers tend to use chlorpyrifos on broccoli prophylactically, targeting cabbage maggots at the larval stage before the pest hatches and before it can cause damage to crops. Encouragingly, despite cutting back on chlorpyrifos, the Monterey County broccoli industry appears to be thriving. The number of acres in production and the total amount of broccoli produced continue to climb . At the same time, the market price for the crop is also on the rise, making the value per ton the highest it has been in recent years. In just over a decade, from 2000 to 2013, the value of Monterey County’s broccoli crop rose from $280.4 million to $426.9 million . These data suggest that regional broccoli growers on average are not only surviving, but thriving without the use of chlorpyrifos. As with broccoli, Monterey is the leading producer of lettuce in the state with 57% of production. Monterey and the second-lettuce producing county in the state, the Imperial County, together account for 70% of all lettuce produced in California, or roughly half of all lettuce produced in the U.S. Unlike chlorpyrifos, diazinon is used on lettuce for a variety of insect controls; and for each pest, there are a handful of readily available chemical alternatives. Consequently, the world-renowned “salad bowl” was unscathed by diazinon cutbacks. Both the production in lettuce acreage and crop totals have steadily increased in Monterey County as well as the price/value of the crop . In 2014, the most recent data available, Monterey’s lettuce crops were the highest they had ever been, valued at $1.2 billion. Comparing these same data with the second highest broccoli and lettuce-producing county in the state, the Imperial County, demonstrates that the Central Coast’s unique cool season cropping systems may be at the core of why agricultural production can thrive without chlorpyrifos, while growers in other regions are not willing or able to give it up so readily. The Imperial Valley is located in southeastern Southern California in the Colorado River Basin Region. With high summer temperatures, the Valley is well known for its number one agronomic crop, alfalfa, grossing $220 million in 2014. The region also has a reputation for its midwinter vegetable crops, including head lettuce, leaf lettuce, cauliflower, broccoli and cabbage. Alfalfagrowers in the Imperial Valley have become ever more reliant on chlorpyrifos due to the increased pest pressures from blue alfalfa aphids.

Chlorpyrifos is preferred by alfalfa growers for the suppression of these aphids over alternative insecticides, and is a fundamental tool in most growers’ IPM programs . Consequently, the region has not experienced the same downward trend in chlorpyrifos use that Monterey and the Central Coast have. Though broccoli farmers in the Imperial County were able to curtail chlorpyrifos application, alfalfa farmers in the region use a much larger share of the chemical, trumping any decline benefited by chlorpyrifos cutbacks on broccoli . Diazinon use in the Imperial Valley has declined on broccoli as well as on all crops.Fresno County, the third largest broccoli and lettuce-producing region in the state, has a parallel story to the Imperial County. Fresno County is located in the Central Valley and is characterized by its hot mediterranean climate. Taking advantage of Fresno’s ideal growing conditions as well as the lucrative almond market, farmers have been steadily converting land to almond production. In 2014, almonds were farmed on 170,711 acres up from 82,700 acres just a decade earlier. In 2013 and 2014 almonds grown in Fresno surpased the billion dollar mark, outdoing grapes for the number crop value in the county. As of 2013, almonds had the highest economic value of any California nut crop and were the highest export value of any American specialty crop .The county is the third largest producer of broccoli and lettuce in the state, and produces a variety of other vegetables including tomatoes, onions and melons. In 2013, chlorpyrifos had the greatest percentage increase in use among insecticides and most of this increase was dedicated to almond production . Almond growers became increasingly dependent on the chlorpyrifos due to budding populations of two crop pests: leaffoted bugs and navel orangeworms . Chlorpyrifos use on broccoli, on the other hand, steadily decreased as it did in Monterey and the Imperial Valley, and was trumped by use on almonds and other crops .Over the past decade, diazinon use in Fresno has declined overall as well as specifically on lettuce crops, mirroring Monterey, Imperial Valley and statewide trends. Comparing chlorpyrifos use between Monterey County, the Imperial County and Fresno County underscores the importance of crop type and pest pressure on growers decision to apply the chemical or not. This comparison leads to several interesting policy questions: Would the Imperial Valley or Fresno County have stopped using chlorpyrifos on alfalfa or almonds if it were held to Central Coast Tier 3 requirements? Or would growers have complied with Tier 3 requirements to continue producing their most profitable crops? Or, lastly, would growers have given up the alfalfa and almonds altogether to escape individual monitoring mandates? The widespread decline in chlorpyrifos use on broccoli in three different regions under three different regulatory programs gives considerable credence to other macro-conditions, besides the 2012 Ag Waiver, as causes for the pesticide’s demise. However, before jumping to this conclusion, other factors, such as differing pest pressure, must be considered.

The bio-assessment studies show a clear relationship between increased water pollution and increased agricultural and urban land use . Nationwide, agricultural non-point pollution is the chief impediment to achieving national water quality objectives . The EPA lists the chief components of these non-point source agricultural pollutants as nitrogen and phosphorus from fertilizers, pesticides, animal sources, soil erosion, and salts from irrigated fields. The National Rivers and Streams Assessment, conducted by the U.S EPA in 2004 and again in 2008/9, uses separate monitoring data from the 303 listings. Over the course of five years, between 2004 and 2009, the Assessment found seven percent fewer stream miles were in good biological condition. Similar to the Central Coast, throughout the U.S. changes to water quality in streams were variable over time and space. Overall, the report found that U.S. streams and rivers are “under significant stress and more than half exhibit poor biological condition” . Despite the diverse datasets,30 litre plant pots bulk frequency and consistency of monitoring data are still not sufficient to verify the effectiveness of the Agricultural Waiver .

The following two sections will assess the value of the Ag Waivers requirements, particularly the monitoring provisions.A closer look at the requirements themselves highlights why compliance may not lead to improved water quality. The Agricultural Waiver, in theory, uses an approach that gradually increases compliance requirements, called an “iterative approach,” meaning dischargers implement increasingly improved management practices until the region has achieved clean water. This approach recognizes that progress towards achieving water standards can take time. Logically, the 2012 Waiver should be significantly more rigorous than its predecessor. While Tier 3 farms might have more stringent requirements, a handful of significant provisions for Tier 1 and 2, which make up 99% of all growers, have been so watered-down and in some cases eliminated that the 2012 Ag Waiver has been regarded as “only marginally stronger than the 2004 Ag Waiver” . Several examples illustrate this point. First, in its modifications to the 2012 Agricultural Waiver, the State Board eliminated the only enforceable provision that would control nitrogen pollution—the nitrogen balance ratio target2 . Instead, grower snow only need to report the total N applied. Even with the 100% compliance rate of this mandate, the total N reporting provides substantially less information about which farms have nitrogen surpluses and might be contributing to pollution. Second, and arguably most importantly, the Ag Waiver does not have any quantifiable mechanisms to determine if management practices implemented by Tier 1 and 2 farms reduce pollution . Third, choosing which management practices to implement is largely up to the discretion of agricultural operators.

The Ag Waiver does not define what management practices should be implemented or verify if those practices are actually improving water . Though management practices are a means to reduce pollution discharges and achieve water quality, the California’s Non-point Source Policy establishes that “management practices may not be substituted for actual compliance with water quality standards” . One new requirement that can aid the Regional Board in estimating improved water quality is the mandate to report all water quality management practices and outcomes. The online form requires growers to check all nutrient, irrigation, pesticide and sediment management practices that are being implemented and the number of acres on which the practices are applied. While this new tool will provide baseline data for the Regional Board to better understand how growers say they are managing their land and crops, there are no means to verify if those management practices are effective. Growers have the opportunity to report if they have seen a positive outcome from their implemented management practices, yet outcomes are measured by the grower’s perception of change rather than a numeric or quantifiable water quality data. For example, in the 2014 annual compliance form, the most commonly used method to confirm sediment reduction was by walking the perimeter of the property to verify erosion controls were in place and that sediment did not leave the ranch/farm during irrigation events and/or storm events; the least commonly used method to confirm sediment reduction was to measure turbidity in stormwater runoff.

The Agricultural Waiver has significant monitoring limitations. In the 2012 Ag Waiver, the Regional Board acknowledged that a critical limitation of the 2004 Ag Waiver was “the lack of discharge monitoring and reporting… and the lack of public transparency regarding on-farm discharges” . The 2015 Superior Court Judge ruling reiterated this point: “The 2004 Waiver has not been successful because it lacks adequate standards and feedback mechanisms to assess the effectiveness of implemented management practices in reducing pollution and preventing further degradation of water quality.” Despite adding a handful of modest monitoring requirements to contend with these limitations, the updated 2012 Ag Waiver suffers from the same shortcomings as its predecessor. The biggest deficiency in the monitoring program is that data collected are neither comprehensive enough to verify the effectiveness of the management practices nor to identify individual operations that cause impairments . This issue points to the most controversial AgWaiver topic: public disclosure and transparency of information. The most effective means of identifying a polluter is to conduct individual discharge monitoring at the edge of a discharger’s field where pollutants enter the water. Because of its controversial nature, and the difficulty to collect data from thousands of individual farms, the 2012 Ag Waiver compromised by mandating that only the highest risk polluters—Tier 3 farms—need to report individual surface discharge monitoring. The biggest fear among growers is that of being identified as a point source polluter, and subsequently regulated under WDRs or NPDES permits, rather than a Waiver. As one Regional Board staff member put it, growers “don’t want to deal with a government agency managing their land and water, and they don’t want to be called part of the problem.” With individual discharge monitoring requirements as the driving force,wholesale plant containers growers did anything they could to get out of Tier 3. Farm operations split their ranches into sub-parcels, stopped using certain pesticides, or stopped farming altogether. To depict the drastic exodus out of Tier 3, in 2010, over 10% of farms were categorized in Tier 3, yet as of September 2015, only 1% of all farms in the Region are regulated under that Tier. As a result of the shift to lower tiers, monitoring and regulatory provisions, and the overall Ag Waiver itself, have been severely hindered, since most growers are not held to sufficiently strict mandates. A goal of requiring individual surface water monitoring of Tier 3 farms was to evaluate effects of waste discharge on water quality and beneficial uses; it remains to be seen whether data from such a small subset of growers will adequately achieve this objective. In contrast, the 99% of other growers must report surface receiving water monitoring, either cooperatively or individually. Surface receiving monitoring is conduced on the main stem of a river, rather than near a grower’s fields. For growers, this is a much more attractive scenario: data are reported as an aggregate and pollutants detected from surface receiving water data can rarely be traced back to its source. Additionally, the cost is generally less than the fees associated with the individual surface water discharge Sampling and Analysis Plan and Quality Assurance Project Plan .

Monitoring challenges are exacerbated by the diffuse nature of non-point source pollution. Because agricultural runoff does not enter a stream at a well-defined point, and often occurs episodically , continuous or targeted monitoring are needed to evaluate the rapidly changing and dynamic local environmental conditions. Growers and the cooperative monitoring program are not required to collect data at the same time or even during the same rain event, making it difficult to compare results and establish trends. A nonprofit, the Coastal Watershed Council , has attempted to address this problem by testing several water parameters in watersheds throughout the region during the first rain event, or “First Flush”, in their annual Snapshot Day. By collecting water quality data during the first rainfall, the CWC attempts to capture the most concentrated pollutants washing off the landscape in significant levels at the same time from year to year. The CWC’s Snapshot Day found nutrients and turbidity from agriculture and urban areas to be a major source of regional water contamination. However, the CWC program is volunteer-based and has a limited capacity to carry out high quality comprehensive monitoring.A related complaint by growers is that they will be substantially harmed by the cost of compliance. A 2012 Ag Alert article reported that the regulatory requirements in the 2012 Order amounted to more than $230 million in lost revenue and an estimated 2,500 to 3,300 in lost agricultural jobs . The Growers-Shippers Association of Central California added that the adopted regulations are “over-board and intrusive on grower operations” . Some growers claim that the compliance costs are unwarranted because farm management practices similar to the ones being mandated are already in effect. A representative from the Santa Cruz Farm Bureau voiced the agricultural community’s frustrations, “in general, there has been a lot of concern about the regulations being applied. In particular, the Regional Board did not take into consideration what was already being done on the farm. [The Agricultural Waiver] adds a financial and time burden on growers.” Several growers and agricultural organizations, including seven county Farm Bureaus, put into writing the perceived economic burden in their appeal to the 2012 Ag Order. In their request, agricultural petitioners claimed the cost of compliance would amount to 1.3-2.5%, 0.13-0.3%, and 0.8-1.5 % of gross crop revenues per acre for leaf lettuce, strawberry, and head lettuce, respectively. Some asserted that the methods employed in the agricultural group’s cost analysis were “not credible” and the numbers were “inflated,” and “self-serving” . In its argument against the agricultural industry’s estimated compliance costs, the Regional Board claimed that because the potential costs vary widely from farm to farm it is impossible to estimate the range over all farms. Another example of cost discrepancies was in the estimated monitoring expenditures for the two pesticides regulated in the Ag Waiver, diazinon and chlorpyrifos: the Regional Board estimated the total cost to monitor these two pesticides would be $250 per farm, whereas the agricultural petitioners estimated $7,000 to $11,000 per farm. It is nearly impossible to put a dollar value on the public health and ecological benefits gained from the two Agricultural Waivers, but it is worth mentioning some potential benefits from the Ag Waiver. In their rebuttal to the request for a “stay,” the Regional Board listed several environmental benefits that would result from the 2012 Agricultural Waiver including improved drinking water, overall public health, decreased pollutant loadings in surface and groundwater, reduced threat to sensitive aquatic habitats, and more stabilization of stream banks in riparian areas. Whether these improved societal and environmental conditions outweigh the estimated 0.8- 2.5% of gross crop revenues it would cost to comply will largely depend on who is asked.Issues of equity are at the heart of public policy controversies , and can be used to measure policy effectiveness two ways: fairness or redistribution . Factors that may play a role in measuring equity through the lens of environmental justice include the level of participation among stakeholders and/or distributive outcomes of pollution . These types of concerns harken back to the founder of policy studies, Harold Lasswell , who encouraged policy scholars to ask: “Who benefits? Who gets what, when, and how?” Answers to such questions attempt to uncover the inevitable unequal allocation of resources that result the dynamic relationship of power and bargaining inherent in the making of any set of rules and regulations . In the case of the Central Coast Ag Waiver, three main distributional consequences of compliance have been highlighted as unfair. The first two are contestations among growers themselves. First, Tier 3 growers contend that the three tiered system is imbalanced because it distributes a substantially higher burden on a small number of farms. This assertion represents a classic policy paradox: “equal treatment may require unequal treatment; and the same distribution may be seen as equal or unequal, depending on one’s point of view” .

Exclusion discourse placed Chinese as more efficient than whites, whose “manhood” is wasted in repetitive tasks such as mining and service work. Lye shows that the Chinese were for this reason seen as more “modern,” more suited to a future of proletarian work. The Anglo-Saxon’s supposedly large, violent, inefficient body had been well adapted to hunting and Indian-killing, an era that had ended. Thus the anxieties around Chinese Exclusion were partially a result also of the closing of the frontier. The Octopus represents the Chinese as docile and feminized, but for that very reason as more refined. Unlike anti-Black racism, where the African is animalized in contrast to the European, in the American West it is Anglo-Saxons who are closer to animals because they are more alive, virile, and so on. They are “red-blooded Americans.” When the Anglo-Saxon farmhands and the Chinese cooks appear together, it is when the former are eating, and Norris’s description emphasizes the scale of the operation,best grow pots the frenzy of impersonal activity: “The half hundred men of the gang threw themselves upon the supper the Chinese cooks had set out in the shed of the eating-house…

The table was taken as if by assault; the clatter of iron knives upon the tin plates was as the reverberation of hail upon a metal roof” . The way the Chinese “set out” the food appears refined in comparison to the naturalist cacophony of “the gang’s” eating, as unstoppable as the weather. The cooks are set apart from the farmhands, but are an integral component of the large-scale ranch. Like the new machinery, the Chinese cooks allow for the rationalization and division of labor of the new agriculture, as opposed to the archaic animal nature of the farmhands: “It was a veritable barbecue, a crude and primitive feasting, barbaric, homeric. But in all this scene Vanamee saw nothing repulsive… this feeding of the People, this gorging of the human animal, eager for its meat.” The feasting is both “barbaric,” those outside of civilized discourse, and “homeric,” the origin of Western civilization—this is a tension internal to the Anglo-Saxon race, internal to “the People” and their holy destiny of Indian-killing, and is best captured by the phrase “the human animal.” The otherness of the Chinese, who are not hungry for meat, is of a different order, on par with the introduction of machines into the landscape.

The labor of “the gang” is what is being replaced by mechanized agriculture, whereas the Chinese domestic servants’ positions are secure, as they work in the house feeding the ranch’s owners as well. Norris signals the defeat of the white workers when, for all the talk of Chinese famine, it will be a white farmer who actually starves to death in the novel. Mrs. Hooven, the widow of a German immigrant farmer killed at the irrigation ditch, is dispossessed of the family’s land by the railroad and travels to San Francisco with her two daughters. This account of proletarianization is formally the most elaborate section of the book, with the events presented out of temporal sequence. The teenage daughter, Minna, becomes separated from her mother and young sister, and after witnessing the “horrors” of Chinatown she is the first to face the shock of starvation: “The idea of her starving, of her mother and Hilda starving, was out of all reason. Of course, it would not come to that, of course not. It was not thus that starvation came” . After long descriptions of her physical hunger, she is able to survive by becoming a prostitute. The narrative then juxtaposes the mother’s plight and a dinner party held by San Francisco elites including Cedarquist and one of the railroad barons, as well as Presley due to a series of convenient accidents. The dinner party takes only a few hours while Mrs Hooven’s story unfolds over many days, but the narrative cuts back and forth between the rising dramatic tension of the two plotlines. After twenty pages of intercutting, the section climaxes with “‘My best compliments for a delightful dinner’… ‘she has been dead some time—exhaustion from starvation,” .

The Hoovens lose each other because they are country folk unaccustomed to the scale of the city, and because, as German immigrants, they do not speak perfect English. In the racial schema employed by Gompers and Norris, they are not Anglo-Saxons, and so are technically not part of the labor competition. In the new world of U.S.-Chinese relations, their Europeanness is a comical archaicism that cannot survive: they must either die, as the parents do, or assimilate into degrading wage labor, as Minna does.This is part of Lye’s general focus on how U.S. East Asia policy is related to representations of Asian Americans. But the boycott as a social movement shows that the political consequences of American China policy operate in both North America and East Asia. Any contradictions in The Octopus’s view of political economy are familiar from neoliberal globalization at the turn of the twenty-first century, which again advocated for restrictions on racialized immigration but not on goods. We can see this already in the nineteenth-century Exclusion discourse itself, which Cedarquist’s speech does not in fact contradict. Consider this 1886 memorial to Congress, after Chinese had allegedly struck so as not to work alongside white workers: “To begin with, they have a hive of 450,000,000 Chinese to draw from,plant in pots ideas with only one ocean to cross, and behind them an impulsive force of hunger unknown to any European people” . The Chinese are not rational but propelled by impulse, not individuals but a group mind. This naturalist conception of the world, where hunger is an external force, driving them out of the hive from behind, is of a piece with both Turner’s view of forces and what we will encounter in The Octopus. The insect comparison is typical, as shown in an address by Morris M. Estee before the State Agricultural Society at Sacramento, in which the Chinese are so hungry as to be counterproductive in agricultural labor because they eat more than they harvest. He recommends barring from “our orchards, vineyards, hopfields and grain fields […] the thieving, irresponsible Chinaman, who like the locusts of Egypt, are eating out our substance” . It is apparently when working in the fields that such a voracious hunger comes to the fore, unlike the machinic and docile mode when engaged in modern service work. The broader point, however, is that Chinese hunger also underlies U.S. imperial reach in Asia.

As we saw above, Norris describes a shift from war and empire to trade and markets. Lye tends to focus on the rupture between these two, so that empire is now a euphemism or exciting metaphor for what is actually the cold economic logic of the market. I am instead following Eperjesi in emphasizing the continuities, seeing economic power as a new form of empire.For Western understandings of China have long taken famine to be a crucial component of its political system. In The Spirit of the Laws –the most influential text of the Enlightenment on comparative political systems–Montesquieu argued that while the Chinese had the most advanced methods of intensive farming, this could never keep up with their large population. This imbalance somehow derives from a contradiction between low fertility of the land and high fertility of the Chinese female body: “The climate of China is surprizingly favourable to the propagation of the human species. The women are the most prolific in the whole world. …[However,] China, like all other countries that live chiefly upon rice, is subject to frequent famines” . Writing in the mid-eighteenth century, the author does not even explain, let alone support, these fertility dynamics. By comparison, Norris’s version at the turn of the twentieth century is relatively sophisticated, with its pseudo-scientific theory of the decline of Chinese rice’s nutritive value. I raise this long history of the idea of Chinese hunger here to argue that, far from a projection of immigration discourse, it is the historically-primary way of thinking about China in terms of food. Thus the authoritarian rulers are compelled to rule well and provide enough food for the people lest they be overwhelmed by the hungry population. Montesquieu here applies his famous theory of checks and balances to the Chinese system, but these checks are not a separation of powers among institutions but the environmental power of fertility and hunger that checks the emperor. It is a domestic economy rather than a political economy. We see these same themes repeated in the U.S. in the late nineteenth century, that China is a land of overpopulation and famine, and that individual Chinese are satisfied with less. It underlies Cedarquist’s vision of Empire marching westward: whoever can feed the Chinese is their rightful imperial ruler, and so the mandate falls to American industry. However, whereas Europeans already viewed the Chinese in terms of food in the eighteenth century—unmatched agricultural innovations coexisting with famine and limited dietary needs—this was in the context of an inquiry into social questions, above all religious, political, and economic organization. By the time we reach the mid- to late-nineteenth century, and specifically in the labor market of the western U.S., these food characteristics were reconceptualized in terms of biological race. One aspect of this is the dehumanization following from extreme hunger, which animalizes them, as in the above quotations where the Chinese emerge from a “hive,” feed like “locusts,” and are biologically different from the “beef-and-bread man.”

These are the characterizations that the Chinese boycott proponents are fighting and they show what is at stake in refusing consumption. Where the Americans use food distribution as a weapon to conquer China, the Chinese politicize food consumption. Where the Exclusion supporters politicize the theory of evolution through biological races, the boycott supporters apply the theory of evolution to the history of political forms to show the need for a strong nation.American Studies scholars have often noted the contradictory attitudes of love and hate that Americans have felt toward China, which in this period can be seen in the contradiction between the twin discourses of the dream of the China market and the nightmare of Chinese invasion, the yellow Peril . While psychoanalytic and other theoretical frameworks can illuminate the dynamics between these two tendencies, we can understand the more direct connections between them by foregrounding the Chinese experience of the U.S. In the U.S. this is primarily a class difference, between the workers who oppose competition, that is, who want a monopoly on the labor market, and the merchants and industrialists who want to increase exports to China. This is not, however, always understood as a direct class conflict, as for example Gompers argues that the boycott threat will never become a reality, and so Chinese workers can be excluded without hurting American capital in Asia. Frank Norris, on the other hand, is more dubious about the China trade’s effect on American workers, as we have seen when Mrs. Hooven starves in San Francisco. The larger point is that U.S. texts from this period generally do not see a causal link between the China market and Chinese immigration; they are two contemporary phenomena which must both be managed. The Chinese proponents of a boycott of American goods, however, articulate a direct connection between the two: cutting off foreign trade is an appropriate response to the mistreatment of Chinese nationals in the U.S. While this is partly a practical matter—the only way ordinary Chinese can affect the us, in however small a way—it also reflects the historical and economic links between Western imports into China and Chinese immigration to the Americas, both of which are the result of Chinese decline and Western military interventions. While many American Studies scholars tend to treat Chinese Exclusion as an internal development in the unfolding national history of race and labor, we should see that this is also a pivotal moment in the history of China. The campaign in support of coolie labor is the first Chinese social movement to be articulated in nationalist terms. On the other hand, while the boycott is often described by historians of China as the first Chinese social movement to define itself in national terms, this relationship with the U.S. at the origin is generally seen as somewhat incidental and not central to the development as Chinese nationalism as a whole. In this chapter I have been arguing that we should recenter this relationship in considering the early twentieth-century development of both countries.

Our results suggest that the yield effects of crop rotational histories in cotton are relatively modest in magnitude: the posterior means for effects of any specific crop were mostly under 15%. However, given the tight profit margins of commercial agriculture, a 15% change in yield could translate into a far greater percentage change in profit, and could therefore be of substantial economic significance to a grower. As we seek to feed a growing worldwide population while doing minimal harm to the environment, crop management practices that increase yield while reducing the need for costly and damaging pesticides and fertilizers are of great value. Crop rotation is one such method, and we are optimistic that ecoinformatics approaches may be helpful in elucidating the details of how to optimally implement crop rotation.On February 5th, 2014, over thirty U.S. food production organizations joined together to form the Coalition for Safe Affordable Food with the aim of lobbying Congress to affirm the safety of genetically modified foods. The organizations of the coalition—the Grocery Manufacturers Association, the National Council of Farm Cooperatives,large plastic garden pots the National Corn Growers Association, and the American Soybean Association, among others—were on the defensive first of all against ballot initiatives seeking compulsory labeling of genetically modified foods over the last two years in over half of the fifty states.

Secondly, China’s recent ban on certain genetically modified food imports had led to the return of 600,000 tons of American corn in 2013, and perhaps explaining why the front page of the Coalition’s website features a photograph of a smiling East Asian child against a pastoral background, happily biting into a large corn cob . In China, by comparison, the issue had become explicitly nationalist, with GM crops depicted as a threat not only to public health but to national food security and with struggling farmers protesting the “traitors” in the agricultural ministry who had continued some imports. In a video made for army officers but later leaked online, the voice-over makes this clear: “America is mobilizing its strategic resources to promote GM food vigorously. This is a means of controlling the world by controlling the world’s food production” . This dissertation will show that these contemporary conflicts between and within the United States and China over food security have a long history that has defined uneven political and sociocultural relationships between these two countries, and moreover that literature has played an important role in imagining and defining those relationships. Frank Norris’s 1901 novel The Octopus famously calls for a rapid increase in food exports to China to sustain the industrialization of agriculture in the American West. A character in the novel declares that for American farmers and industrialists to survive in the coming century, “we must march with the course of empire, I mean we must look to China,” . Suggesting his own views hued closely to this fictional voice, Norris elsewhere linked the close of the frontier to American marines landing in Beijing to help put down the Boxer Rebellion .

In addition to celebrating American power directly through the military, and indirectly through control of the food supply, Norris also contributed to so-called “yellow peril” discourse with stereotypical portrayals such as the Chinese kidnapper in Moran of the Lady Letty . While scholars have explored the connections and tensions in American literature of this period between the fantasy of the China market as an engine for American prosperity and anti-immigrant racism, what remains unexplored is how Chinese writers understood and engaged with these same issues. In 1905, to protest the treatment of Chinese workers in the U.S., activists in southern China organized a boycott of American imports that lasted over a year . In the propaganda literature and other works of cultural production popularizing the boycott, the act of giving up American food products appears as particularly poignant. One popular song, for example, politicizes Norris’s celebrated commodity: “American wheat flower is made with Chinese blood” . In the novella Extraordinary Speeches of the Boycott , meanwhile, the protagonist decides that the only way for China to be truly independent of the U.S. is to develop its own food sovereignty, and at the close of the narrative he moves to the countryside to promote large-scale agriculture. In reading these American and Chinese texts comparatively for the first time, this project shows the multifaceted roles of both imaginative literature and other rhetorical forms, and especially literatures of food, on both sides of U.S.-China political struggles.

Furthermore the project demonstrates how these early conflicts informed the later Cold War competition between the two countries, when each attempted to export its own versions of rural modernization to the rest of Asia, from the Korean War era all the way up to the 2014 military video. More broadly, by reading American and Chinese literature about agriculture together, we begin to appreciate how writers and their reading audiences of this period imagined the modernization of food production in their respective national contexts as entwined with transnational exchanges and tensions. In addition to the prominent conflicts over U.S. and Chinese food power during the past century, transnational collaborations between Americans and Chinese are perhaps less well known. Most importantly, from the 1890’s to the 1937 Japanese invasion of China, a steady stream of American agriculturalists travelled to China to research Chinese agricultural practices and recommend improvements. While certain interests, such as International Harvester, hoped to promote U.S. farming technologies, this project will focus on the larger group of researchers with institutional funding—above all the Rockefeller Foundation—trying to solve what they saw as a global food shortage1 . These visitors were graduates of the new U.S. agricultural colleges, and an increasing number of Chinese also travelled to the U.S. to become trained agronomists. The most influential of these consultants was Lossing Buck, husband of novelist Pearl S. Buck. Buck travelled with Lossing during his fieldwork, and edited his first book, Chinese Farm Economy,raspberry plant pot in the year before she wrote The Good Earth. I argue that this novel’s extraordinary binational success—it was widely read and debated in China—can be best understood through the longer history of academic collaboration between Chinese and American agricultural experts attempting to define the nature of Chinese rural society through reference to the U.S. This is furthermore why there is actually significant overlap between her representations of the Chinese countryside and those of the Communist writers ostensibly at the other end of the political spectrum. World War II largely put an end to U.S.-Chinese collaboration in the countryside, and when the Communists came to power in 1949 they renewed the terms of food conflict from the turn of the century. When the U.S. ambassador was recalled to Washington in August 1949, Mao Zedong characterized him as a fleeing colonial governor, and in a widely-read series of essays Mao criticized the American practice of distributing famine relief flour, saying that it was bait intended to catch the Chinese people and devour them. The image is reminiscent of the ending of The Octopus, where the figure of the American industrialist prepares a shipment of famine relief wheat as the first step to expand U.S. food export channels to China and India.

With the opening of the Cold War in Asia during the 1950s, the U.S. and China each launched propaganda campaigns to promote how their competing visions of rural modernization could help the rest of Asia. The earlier work of the agronomists in China, such as Lossing Buck, served as the foundation for the Cold War U.S. program of non-redistibutive, technical improvements aimed at international development, a series of projects in India and other countries that were later collectively dubbed the green revolution. The Chinese Communist Party, meanwhile, went on to implement nearly all of the agronomists’ earlier suggestions, but combined them with land redistribution. As evident in contemporaneous literature and film, both nations championed their own programs as promising the utopian end to rural hunger, in contrast to the failure of the other. In this way the Cold War period shows a synthesis of the earlier two periods, as the old political rivalry renewed but this time through technical discourses of production and development. Just as the two major national powers advancing the Cold War in Asia developed their agricultural and rural modernization programs out of their respective work in the Chinese countryside before the war, so did a third approach that we can now recognize as the prototype of the modern non-governmental organization . Another transnational project of the late 1920s and early 1930s was the Mass Education Movement , headed by James C. Yen, who was born and raised in Sichuan and who attended Yale and Princeton. Through his network of contacts with the YMCA, Yen secured Rockefeller funding for the MEM, giving talks throughout the U.S. and publishing multiple essays and pamphlets for an American audience. In contrast to the technical focus of the agronomists, Yen sought to reform village life as a whole, beginning with literacy and only then moving on to rural economy, health, and government. Literature again played a central but slightly different role in this project, which focused on literacy for the rural population rather than literary representations of those rural communities. American academics also participated in the MEM project, but here they were anthropologists seeking to understand and affirm local traditions, rather than scientists seeking to maximize crop yields. Most notably, Yen strove to remain independent of both the Nationalists and Communists, and after the Communists ultimately came to power, he neither stayed in China nor settled in Taiwan, but instead moved to the Philippines and founded the Institute for Rural Reconstruction. This non-governmental organization has continued to promote grass-roots, community-centered programs throughout rural Asia, later expanding to Latin America and Africa. When in the 1970’s and 1980’s a new generation of international development workers began to question the top-down technical programs that the U.S. promoted throughout the Third World, James C. Yen’s work fifty years earlier in China was rediscovered . In short, this dissertation argues that American and Chinese writers imagined the other through tropes of agriculture, food, and hunger during the early- and mid-twentieth century. Writers producing fiction, poetry, and other forms of literature about agriculture and rural life mediated the material connections between the two countries, including their food trade and agricultural modernization projects. In pursuing these literary and cultural comparisons, the dissertation proposes dialectical relationships along two axes. It begins with the dialectic between imaginative literature and political-economy, specifically the political-economy of agricultural development. Literary and cultural studies as fields have shown that literature mediates readers’ understanding of and engagements with historical processes, which cannot be apprehended in their totality. The relationship between literature and political-economy is dialectical because the former does not simply mirror the latter, but also provides ways of understanding that influence future actions. Many scholars have examined American literature’s dialectical relationship with agricultural development and larger environmental transformations.2 For example, William Conlogue has shown that during the twentieth century U.S. literary authors crystalized for their reading publics various competing directions for future agricultural development. In Chinese literature, understanding and reimagining the countryside and the rural people has been explicitly tied to national transformation since the early twentieth century, and during the Maoist period aesthetic treatments of agricultural production were tightly regulated.In particular, this dissertation builds on Allison Carruth’s work showing that many twentieth-century U.S. literary writers responded to and mapped the growth of what she calls “American food power” abroad. I argue that how Americans and Chinese imagined their competing food interests had long-term consequences for the emergent global food system. As my literary and cultural analyses will show, the most common trope through which the relationship between the U.S. and Chinese countryside—and in turn the shifting political economic relationships between the two nations—was imagined was that of famine, for which I propose two main reasons. First, famine displays most completely the perceived failures of the current rural system, and so the need for modernization. As Marx noted, during the smooth functioning of the capitalist economy, people in daily life experience production, distribution, and consumption as only abstractly connected, but during periods of crisis their direct underlying unity becomes become painfully evident. In this way the representation of rural crisis as famine actually gives the most complete picture of the total food system as analyzed by food studies.

The Southern California Comprehensive Water Reclamation and Reuse Study , for example, provides a comprehensive assessment of existing reuse and reuse potential in Southern California.The following is a review of key moments in history that changed water policy, from passage of water rights legislation to the voter approval of the State Water Project . Although there are other significant events, however, they are not deemed as relevant and significant to this research and hence are not included. As shown in Figure 2-5, the first Colorado River delivery was made to the farmlands of the Imperial Valley was in 1901 and the major compact agreement was signed in 1922, designating specific allocation to the upper and lower Colorado basins for all seven states . The Central Valley Project and State Water Project were authorized for construction in 1933 and 1951 respectively. In 1973, the first State Water Project delivery was made to Southern California and the Federal Safe Drinking Water Act was passed in 1974 setting the first ever standard for drinking water throughout the country.

In 1998-2003,plastic seedling pots the Colorado Quantification Settlement Agreement was reached between California and other Colorado River Basin states and the Federal Government. As a result of this agreement, California received its allocation of 4.4 MAF which still serves as the state’s allotted water share even until today. Many state agencies are involved with California water management as shown in Table 2-3. While overlapping responsibilities might occur in terms of broad objectives, generally, there is not duplication of functions. Most agencies focus on a specific subset of water management, for example, the State Water Resources Control Board and the Department of Water Resources are the two leading water management agencies. They have mandated water supply objectives, however, their roles differ greatly. DWR focus on water delivery, water supply planning, and infrastructure development, while SWRCB is more of a regulatory body, managing water rights and water quality permitting . These roles are complementary and often require the two agencies to work in concert to address water management at the state level. The management of California water systems consists of three main components: water supply, water quality, and flood control.

Most agencies involved in one or more of these components also have responsibilities for scientific activities and monitoring and administering financial assistance for local water infrastructure. For example, several financial assistance programs attempt to jointly address water quality and water supply needs at the local levels, thereby providing more comprehensive local water supply reliability. Other state agencies not listed may be involved with water management as part of their greater mission . At the federal level, most agencies have distinct roles, for example, the United States Environmental Protection Agency focuses on water quality, while the United States Bureau of Reclamation focuses on water supply. However, these roles can overlap and potentially duplicate state efforts, for example, both state and federal entities estimate the state’s water supply resources, although the state has a more comprehensive role through the efforts of DWR. At the local and tribal levels, most entities play multiple roles including both water supply and water quality ones. Local entities can be both regulated and regulatory entities, receiving permits from state agencies for water quality while in return regulating their constituents to meet those permitting requirements as shown on Table 2-4. In some respects, these roles may duplicate those of state and federal efforts. For example, federal, state and local water agencies may each be independently investigating the development of new water supply sources to potentially serve the same region of the state. Water regions defined by DWR and SWRCB are similar but not identical.

SWRCB works in conjunction with nine semiautonomous regional boards while the DWR divides the state into ten hydrological regions governed from Sacramento headquarters. Although, some activities of DWR and SWRCB require coordination among regions and between the two state agencies, their differences in regional definitions can pose a challenge for implementing programs, planning or accounting for California water resources. Figure 2-6 shows the difference in regional boundaries between DWR and SWRCB.The Federal government holds the most water rights in the state with over 112 MAF of water rights mainly for delivery through the federal Central Valley Project. Second to this area are the water rights held by Imperial Irrigation District serving mainly farms in the Colorado River region. Water rights exceed actual total water volume availability on almost all river systems of the state. This is partly because water may be reused as it runs off farms or may be returned to the river after use for a non-consumptive purpose such as energy production. In some cases, water rights are oversubscribed and exceed actual water availability .As described above, California’s water supplies originate from many sources including local surface water projects, groundwater, inter-regional surface water deliveries such as State Water Project, Colorado River imports, treated wastewater, and natural stream flow. In 2005, DWR estimated that California used about 40.2 million acre-feet of water. Of this amount, roughly 78% was used by the agricultural sector, while the remaining 22% was used by urban users. Table 2-5 shows historical estimates of urban water use published by DWR for the years 1972 to 2005.

Urban water appears to have grown along with the state’s population . Water declined during the drought in the 1990’s, but appears to have rebounded. Per capita urban water use does not appear to have changed significantly over time . From 2000 to 2005, per capita water use averaged 229 gallons per capita per day. The data appear to show that statewide average per capita urban water use has changed little over time over the period from 1972 to 2005 . Compare to the year 2000, a typical water year, total water supply of 43.1 MAF was used to supply agricultural and urban sectors. Figure 2-11 shows the proportions of water use by each sector and the corresponding non-stream supply for California for this year. Almost 80% of non-environmental water supply is used by the agricultural sector,container size for raspberries and more than half of the urban use is by households. One third of all supply originated from local sources and another third is from groundwater and reuse. The remaining third came from the big water management projects – the State Water Project, the Central Valley Project and the Colorado River. Figure 2-12 shows a graphic that illustrates the significant movement of water across the state. The figure shows that about a quarter of the water that flows into the Bay-Delta region from the Sacramento and San Joaquin Rivers and other tributaries is diverted from the Bay-Delta region for distribution throughout the southern portion of the state. Much of the supply exported from the Bay-Delta is delivered to agricultural regions in the southern Central Valley, supplementing their surface water and groundwater supplies . The remaining Bay-Delta exports are delivered via the California Aqueduct to urban regions along the Central Coast and to Southern California. Southern California also imports substantial water supply from the Colorado river to supplement its local resources. Water and energy are inseparable. The two resources are inextricably entwined. Energy is needed to pump, treat, transport, heat, cool and recycle water. Likewise, the force of falling water turns the turbines that generate hydroelectric electricity, and most thermal power plants are dependent on water for cooling . In California, concurrent demands for energy and water usage have continued to rise. As a result, the need to plan and implement efficient technologies and using alternative sources are critical to the success of California’s future.

Water usage for energy generation in California varies greatly, depending on the primary energy source, conversion technologies, and cooling technologies used. Figure 2-13 illustrate the state’s typical water use cycle. Water is first extracted from a source. It is then transported to water treatment facilities and distributed to end users. What happens during end use depends on whether water is for agricultural or urban use. Wastewater from urban uses is collected, treated and discharged back to the environment, where it might become a source for someone else. Energy is required in all stages of the water use cycle. It is difficult to measure the amount of water-related energy that is actually consumed. Figure 2-14 indicate the total water-related energy consumption used in California at approximately 19% of all electricity and 32% of all natural gas generated in the state in the year 2005 . Due to significant variations in energy used to convey water supplies form one place to another, the average energy intensity of water use cycle in Southern California is much greater than in Northern California. This is due to the fact that Southern California imports about 50% of its water from the Colorado River and from the State Water Project . Each of these supply sources is more energy intensive than any single source of water supply used in Northern California . Table 2-6 illustrates the combined energy intensity of the water use cycle for urban communities in Northern and Southern California. By and large, California’s reservoirs and water delivery systems were designed and operated based on historical hydrology. However, with climate change, this mode of traditional operation may no longer be valid . When it comes to climate impacts in the next 20- 50 years, “utility planners will have to grapple with many of them prospectively rather than as phenomena that are already observable” . According to DWR, temperatures across California has risen one degree Fahrenheit on average. This increase in temperature has decreased the spring snow pack in the Sierra Nevada Mountains by about 10 percent which is equivalent to a reduction of 1.5 million acre-feet of water. Seasonal snowpack in the Sierra Nevada has always been the largest surface water storage for the state. Sea level along the California’s coast has also risen by about 7 inches . Climatology experts advise that the warming of air temperatures may cause our normal precipitation to shift from snow to rain. This would lead to serious reduction in the amount of snowpack, an important natural reservoir for storing water in the winter and supply California with water in the spring snowmelt. Figure 2-16 provides an estimate of the average reduction in snowmelt water content based on rising temperatures. California water planners further suggest that more changes are expected to come as we head towards the year 2050 and beyond. Some of those changes may include another rise in mean temperature by 1.5 degrees Fahrenheit which may decrease the Sierra Nevada snowpack further to 25 percent, a near storage volume of about 3.8 million acre-feet. Figure 2-17 provides an estimate of the snowmelt reduction based on projected future temperatures. Figure 2-18 shows the increased in flood events as recorded by DWR from 1910 to 2000. Climate change is already having a profound effect on California’s water systems as evidenced by the changes in snowpack storage, river flows, and sea levels. Scientific studies show these changes will increase stress on the water systems in the future. Because some levels of the climate change is inevitable, California water systems must be adaptable to change . The impacts of these changes will gradually increase during this century and beyond. California needs to plan for water system modifications that adapt to the impacts of climate change. Figure 2-19 shows a possible global sea level rise of 4 to 6 inches by mid-century. Higher sea levels will increase the salinity in the Delta, disrupting the freshwater supply and quality that many Californians traditionally rely on. While California’s urban water use has grown steadily with population, some areas of the state have succeeded in lowering per capita water use. For example, in 2011, urban water use in Los Angeles was 123 gpcd, among the lowest in the state . A Study by DeOreo et al. 2011 found that more than half of California water use for single-family residences was for outdoor uses. DWR 2011 also reported that residences account for about two-thirds of urban water use. In 2005, residential use accounted for an estimated 66% of total urban water use in California. Water planners believe that this number can be curtailed with improvements in water-use efficiency and conservation by means of rebates and incentives. Water agencies and utilities are also encouraged to implement conservation and efficiency programs, termed best management practices, outlined in the of California Urban Water Conservation Council’s Memorandum of Understanding .