Month: October 2023

Posted on

Flag leaves were harvested from plants four days after panicle emergence

To use the SPQV, all QTL with a sufficiently high logarithm of the odds score should be assessed. The function provides the higher and lower confidence limits, as well as the combined CI for each mathematically related set of QTL. If any QTL within a set exceeds the combined CI for that set, then the whole set can be deemed successful. QTL with observed gene counts that exceed the upper confidence limits should of course be considered to have attained significance. If at least one QTL in an experiment is determined to be significant by the SPQV, the mapping experiment on a whole can be deemed a success. The QTL that contain lower numbers of genes might be identifying previously unknown genes, and therefore their non-significance does not detract from the significance of other QTL. Because of the potential for the identification of new phenotype associated genes in QTL mapping, QTL containing significantly low numbers of identified genes might also be of interest, as they may have been placed on ’empty’ regions due to previously unidentified genes that have a large impact on the phenotype. The QTL containing significantly low numbers of genes should only be considered interesting when at least one other QTL in the same mapping experiment has been proven to be significant so as to avoid the suggestion that all QTL in a faulty mapping study were significant. The intake, transportation, and storage of elements within a plant is vital to its appropriate growth and development. Plants must maintain balance between excessive uptake, which can cause toxicity and necrosis through the production of free radicals or the exclusion of other nutrients, and sufficient uptake of essential inorganic nutrients . Tight control of ion homeostasis allows plants to respond appropriately to environmental conditions such as temperature, soil pH, and water availability . This regulatory control must be responsive to both the concentration of ions within the soil and to those concentrations within the plant. Understanding the genetic control of elemental uptake and transportation will contribute to efforts to improve crops,vertical farming tower for sale and to the ultimate sustenance of the growing global population.

Examining the concentration of elements within a plant sample allows for exploration of the genetic and physiological processes involved in adaptation to a particular environment. To this end, we and others have developed a pipeline that is designed to cheaply and efficiently measure the concentrations of 20 different elements via inductively coupled mass spectrometry . This process is known as ionomics, which is defined as the quantitative study of the mineral nutrient and trace elemental content of an organism; that is to say, its ionome . In cereal crops such as Setaria italica, the flag leaf emerges just before the panicle, and therefore marks the specific developmental time point at which the plant has taken up the majority of its total mass. This tissue is therefore ideal for ionomic inquiry. Additionally, nutrient loading of the grain is accomplished through the remobilization of organic and inorganic materials from the leaves . The flag leaf specifically is instrumental in the loading of photo assimilates and other micro-nutrients , and is therefore commonly used as representative tissue to assess the composition of a grass . The species Setaria viridis, or green foxtail millet, is a member of the Panicoideae that utilizes C4 photosynthesis . It is therefore a good model system for several related, economically important crops, including sorghum and maize. The compact stature, short life span, and sequenced genome of S. viridis have also contributed to its status as an emerging model organism . In addition, S. viridis is the wild ancestor of the crop species Setaria italica ; these two species have a semi-permeable boundary between them, as their primary difference is phenotypic and they are still readily crossed . Foxtail millet is a member of the small millet species, a group of ancient grains that are relatively nutritionally dense when compared to rice and wheat and which are often cultivated as subsistence crops . Because of the combination of the high nutritional value of the S. italica grain and its resistance to abiotic and biotic stressors, breeding elite cultivars of S. italica for increased nutritional content is an attractive prospect. Understanding the contribution that different regions of the S. italica genome make to total nutrient content is an important first step for breeding purposes.

Structured populations are useful tools for dissecting the relationship between elemental accumulation and the genetic content of a species. Recombinant inbred lines have various advantages when it comes to quantitative trait loci analyses. Repetitive selfing allows for the break up of large linkage blocks, which in turn allows for finer mapping . Additionally, once established, RILs may be continuously maintained in a fixed homozygous state. This makes it possible to assay the same combinations of alleles in multiple different environments . The resulting phenotypic and genotypic data can then be compared through various statistical means in order to identify QTL. Here, we use elemental profiling of a RIL population resulting from a wide cross between S. italica and S. viridis grown in multiple environmental conditions to identify QTL associated with the ionic content of leaf material. Overall, we identified 251 QTL, 171 of which were associated with a single element and 80 of which were associated with a principal components analysis of the ionome. The use of traits defined by the differences between treatments in an experiment allowed for the quantification of the influence of the environment on Setaria’s ionome. Experiments were conducted in the summers of 2013 and 2014. Experiments assaying the effect of density of planting on ionic content were conducted in 2013 and 2014. A single drought experiment was conducted in 2014. A total of 189 F7 RILs resulting from a wide cross between the B100 cultivar of S. italica and the A10 line of S. viridis, together with their parent lines, were used as the study material . In every experiment, lines were planted in triplicate in a block design in the field in Creve Coeur, MO. Treatments in the density experiment consisted of either five centimeter spacing between neighboring plants, or twenty centimeter spacing between neighbors. Plants in the drought experiment were either well watered until the time of sample collection, or were subjected to drought stress from eight weeks post planting. The data used for this work included measurements for 20 different elements in flag leaf tissue collected from a recombinant inbred line population resulting from the cross of the B100 cultivar of Setaria italica and the A10 line of Setaria viridis. A drought experiment was conducted in Creve Coeur, Missouri in 2013 ; both a drought and a density experiment were conducted in the same location 2014 . 179 of the RILs were planted in at least two of the three experiments, while 116 were grown in all three.

The leaf samples from all experiments were treated in an identical fashion; samples were dried and stored in temperature and humidity controlled rooms before ionomic analysis. Each sample was profiled for the quantity of 20 elements using ICP–MS . The resultant measurements were normalized to the sample weight and technical sources of variation using a linear model. Experiment level analytical outliers were removed as in Davies and Gather 1993. Pursuant to this, the measured values for each element were transformed to normality using the Box Cox family of transformations, and Studentized deleted residuals were used to identify and eliminate further outliers within the measurements for each element. After outlier removal,hydroponic vertical farm phenotypes were derived by averaging the values for each line within an experiment and treatment. Both environment and genotype impacted the variation present in these data. Repeatability was generally lower than within experiment heritability , indicating that there was less variation in genotypic replicates within an individual experiment than across experiments. The broad sense heritability of 9 elements in the DN13 experiment, 14 elements in the DN14 experiment and 16 elements in the DR14 experiment exceeded 0.4. Certain elements including selenium, sulfur, and boron showed low repeatability; this is likely due to the fact that these elements tend towards analytical artifacts, as they accumulate to levels that are near the limits of detection of the methods described in this paper. The heritability of individual elements varied by up to 0.533 between different experiments. The function stepwiseqtl from the R package was used in order to identify a multiple QTL model for each of the elemental phenotypes. This function moves iteratively through the genome to test for significant allelic effects of each marker on the phenotype in question. When a significant locus has been identified, this is added to the model. A combination of forward and backward regression ultimately produces a genome wide QTL model for each trait. Each element was considered individually, as well as in combination with the others as a contributor to a principal components analysis that was run for each experiment . Five different metrics were used as the phenotype for each element in each experiment. These phenotypes include the ‘raw’ values for each treatment and the differential values for that trait .

The significance of a QTL was computed using the 95th percentile threshold resulting from 1000 iterations of the scanone function as a penalty for adding the QTL to the model. When all experiments are considered, a total of 251 QTL were identified . As expected from the heritability measurements, the majority of these were identified in the 2014 drought experiment . The 2013 and 2014 density experiments allowed for the identification of 75 and 71 QTL, respectively. Approximately a third of the QTL were identified within treatments; the remainder were identified using either the difference, relative difference, or ratio of the phenotypic values measured within the different treatments in a single experiment . Of the 251 QTL, 80 were identified for the mapping based on the principal components analysis; 39 of these resulted from the drought experiment, 21 from the 2013 density experiment, and the remaining 20 from the 2014 density experiment. Of the 251 QTL, 55 were located on chromosome 2 . The locations of the QTL were assessed for overlap with the locations of known ionic genes . Forty five of the QTL contained at least one gene within their 95% confidence intervals. Of the QTL that coincided with genes, 35 were identified using PC as the mapped trait. The QTL were assessed for significance using the Scanning Probabilistic QTL Validator. The QTL were divided into mathematically related sets based on experiment and the phenotypic metric that was used when they were mapped. The results of this assessment are reported in Table 3-3; each phenotypic metric that was used for mapping was associated with at least one set of QTL that identified a significant number of genes, indicating that the data curation was done effectively. There were several regions in which QTL were remarkably concentrated. Fifteen QTL were identified on chromosome 2 between 89.4 and 95.9 cM ; these QTL were discovered in both the 2013 density experiment and the 2014 drought experiment. The traits associated with these QTL included As, Al, Co, Cu, Mo, P, Rb, and Sr, as well as PC2 for the 2013 density experiment. In the context of the ionome, principal components analysis allows for the identification of regions of the genome that would not otherwise be found. While some of the PC QTL identified regions that overlapped with those identified by ion specific QTL, the majority of them, including many QTL identified for PCs which explained a large amount of the variance present in the data, did not. It is possible that for a single ion, the signal associated with the PC QTL regions is not sufficient for their identification, while the additive signal that is inherent in a principal component suffices. Moreover, many of the first few principal components overlay regions associated with water use efficiency , with concentrated regions of QTL identified on chromosomes 2, 5, 7, and 9 at positions 94, 111.9, 99.9, and 123.7, respectively.

Posted on

An additional experiment was carried out in an airtight container without aeration

Briefly, 400 mg sub-samples of the dried, ground plant matter were freeze-dried for 12 h, weighed, and extracted in polypropylene tubes by sequential sonication and centrifugation with 20 mL methyl tert-butyl ether and then again with 20 mL acetonitrile. The combined extracts were evaporated under nitrogen to less than 1 mL, and mixed with 5 mL methanol and 20 mL water. A 6 mL aliquot of the extract was taken for analysis by LSC to determine the fraction of activity as extractable residue. Selected 150 mg sub-samples of the solventextracted plant matter were combusted on the Biological Oxidizer as described above to determine the fraction of 14C present as non-extractable residue. When nutrient solution and jars were exchanged, the volume of remaining nutrient solution in each jar was gravimetrically determined. A 9 mL aliquot of the solution was mixed with 13 mL Ultima Gold scintillation cocktail and the 14C was quantified by LSC. Water loss from evaporation during each 3 d period was found to be negligible in the no-plant control containers. It is likely that microbial activity in the nutrient solution may have resulted in transformation of the spiked 14C-compounds and that plants may have accumulated both parent PPCP/EDCs and transformation products. To discern the contribution of transformation products to plant accumulation, the used nutrient solution from day 21 was preserved with 2 g sodium azide and 100 mg ascorbic acid, extracted, and fractionated using high performance liquid chromatography . Solutions from 14C-BPA, DCL, or NPX treatments were first filtered through a What man #4 filter paper and then passed through a HLB solid phase extraction cartridge . Before use, the cartridges were sequentially conditioned with 5 mL each of MTBE, methanol , and water. The filtered solution was drawn through the conditioned HLB cartridges under vacuum and followed by 50 mL deionized water. A sub-sample of the filtrate that passed through the cartridge was collected for analysis by LSC to quantify 14C that was not retained by the cartridge.

The cartridges were dried with nitrogen gas,hydroponic bucket and then sequentially eluted with 5 mL of MeOH:MTBE and 5 mL MeOH. The collected eluent was dried under nitrogen to 100 μL. The concentrated eluent was transferred to an HPLC vial equipped with a 250 μL insert. The condensing vial was rinsed with 130 μL of methanol, and the rinsate and 20 μL of non-labeled parent standard were added to the HPLC vial. Preliminary experiments showed that the recovery of this extraction procedure from the initial solution to HPLC analysis was 81.5 ± 7.1% for BPA, 85.8 ± 2.5% for DCL, and 74.0 ± 1.9% for NPX. Nutrient solutions from the 14C-NP treatment were extracted by a simple liquid-liquid extraction method. Each nutrient solution sample was shaken with 50 mL hexane for 30 min, and then the upper layer of the sample was transferred to a centrifuge tube and centrifuged at 3500 rpm for 30 min to reduce emulsification. The hexane phase was transferred to a 15 mL glass tube, concentrated under nitrogen to 300 μL, and transferred to an HPLC vial. The condensing vial was rinsed with 180 μL of methanol, and the rinsate and 20 μL of non-labeled NP standard were added to the HPLC vial. The recovery of this extraction method from the initial solution to HPLC analysis for NP was determined to be 66.8 ± 12.0%. Young plants of lettuce and collards were grown for 21 d in nutrient solution containing one of the four 14C-labeled PPCP/EDCs. No significant differences in plant mass were observed between treatments at the end of the experiment. During the experiment, three plants died . Figure 2 shows the mean mass balance for the systems at the end of the experiment, depicting the fractions of the spiked 14C present in plant tissues, in the used nutrient solution, and as unaccounted activity. The unaccounted activity reflected the 14C that was not found in the nutrient solution at the time of solution renewal or in the plant tissues after harvest and may include losses via unidentified processes, such as volatilization, microbial mineralization in the nutrient solution , or stomatal release. Activity in each fraction varied across compounds and to a lesser degree across plant species, suggesting specificity to uptake. Figure 3 shows the cumulative 14C dissipation from the nutrient solution as calculated from the difference in activity in the solution at the beginning and end of each 3 d interval of solution renewal, representing 14C loss from plant uptake and other processes. Dissipation followed the decreasing order of BPA > NP > DCL > NPX for all treatments and occurred at a similar rate throughout the 21 d cultivation. The presence of plants significantly increased the dissipation of PPCP/EDCs from the nutrient solution, except for NP.

For example, the initial concentration of 14C-DCL in the nutrient solution was 105.3 ± 0.3 dpm/ mL, but it decreased to only 32.8 ± 1.9 dpm/mL after 3 d in the presence of lettuce, while 91.2 ± 3.2 dpm/mL remained in the no-plant control. Lettuce and collards treatments had different levels of chemical dissipation in the nutrient solution. For example, the overall dissipation of BPA in the lettuce treatment was 69.1 ± 8.7%, as compared to 88.4 ± 5.3% in the collards treatment . Different compounds also dissipated at different rates. For instance, in the presence of collards, the cumulative loss was 88.4 ± 5% for BPA, 55.6 ± 11.8% for DCL, and 45.5 ± 4.3% for NPX.The dissipation of NP in the solutions with plants was found to be similar to that in the noplant control, especially for the lettuce treatment . The loss of NP from the noplant control was likely associated with volatilization, as continuous aeration was used to maintain the oxygen level in the nutrient solution throughout the experiment. The Henry’s Law constant for NP is 1.09 ×10−4 atm m3 mol−1 , suggesting a tendency for volatilization. The loss of NP in the solution was found to be insignificant, as all of the spiked 14C was found in the solution , and a solvent rinse of the system showed little sorption of 14C-NP on the container wall . Doucette et al. found that in a hydroponic set up, about 13% of the spiked NP was lost to volatilization in the absence of plants. The increased volatilization losses in the current study were likely due to specific aeration and temperature conditions used. Despite volatilization losses, significant amounts of 14C were detected in plant tissues, suggesting that both collards and lettuce accumulated NP . Noureddin et al. studied the uptake of 5 mg/L BPA from hydroponic solution by water convolvulus and found that approximately 75% of the spiked BPA was removed after 3 d. This removal was comparable to that observed for BPA with lettuce in this study, but was smaller than that with collards . Calderón-Preciado et al. evaluated hydroponic uptake of triclosan, hydrocinnamic acid, tonalide, ibuprofen, naproxen, and clofibric acid by lettuce and spath and showed that the removal of NPX from solution was about 70% for lettuce and 10% for spath after 3 d. In comparison, Matamoros et al. observed less than 10% removal of NPX after 3 d of hydroponic growth with wetland plants , while 46% removal of NPX was measured in the collards treatment in the present study.

Matamoros et al. also showed that DCL did not dissipate appreciably in treatments with wetland plants, which was in contrast to the high removal of DCL by leafy vegetables observed in this study . It is likely that the smaller plant mass and the use of non-aerated nutrient solution in the earlier study contributed to the limited plant uptake. The range of variation suggests that plant species,stackable planters along with other factors such as plant mass and environmental conditions, affect the actual accumulation of PPCP/ EDCs into plant tissues. Plant tissues were collected after 21 d of cultivation, rinsed with deionized water, and separated into roots, stems, new leaves, and original leaves for analysis of both extractable and non-extractable 14C. Table 1 shows concentrations of 14C in plant tissues, expressed as parent-equivalents. In agreement with the dissipation trends in solution, plant accumulation followed the decreasing order of BPA > NP > DCL > NPX. Concentrations based on dry plant mass ranged from 0.22 ± 0.03 to 12.12 ± 1.91 ng/g in leaves and stems. Statistical analysis showed that the accumulation in leaves and stems was not significantly different between lettuce and collards, or among the different compounds. In contrast, roots accumulated significantly more 14C than all the other plant tissues, with concentrations that ranged from 71.08 ± 12.12 to 926.89 ±212.89 ng/g. Accumulation of 14C in plant tissues exhibited several apparent trends. In whole collards plants, significantly greater accumulation was found for the neutral compounds BPA and NP than the anionic compounds DCL and NPX , suggesting that the charge state of PPCP/EDCs may greatly influence plant uptake . Similar effects have been frequently observed for anionic herbicides, and are attributed to exclusion of negatively charged molecules by cell membranes . Between lettuce and collards, lettuce significantly accumulated less PPCP/EDC when all test compounds were pooled , although the interaction effect for individual compounds was not significant . Accumulation of BPA or NP in plant roots was significantly higher for collards than lettuce , while portion of DCL accumulated into lettuce and collards roots was not significantly different. Analysis of tissue extracted with solvent showed that essentially all of the 14C was non-extractable; only the root samples from NP-collards treatment contained a detectable fraction of 14C in extracts .

Combustion of extracted plant tissues confirmed that almost all 14C remained as non-extractable residue, one possible endpoint for xenobiotics taken up by plants . Only a few studies have examined the plant uptake of some of the same PPCP/EDCs considered in this study. Wu et al. grew iceberg lettuce and spinach for 21 d in hydroponic solution initially spiked with a suite of 19 PPCPs, including DCL and NPX, each at 500 ng/L and found no detectable residues of DCL or NPX, except for NPX in spinach at 0.04 ng/g. Calderón-Preciado et al. analyzed apple tree leaves and alfalfa from fields irrigated with water containing BPA, DCL, and NPX. DCL was detected at 0.354 ng/g in apple leaves and 0.198 ng/g in alfalfa; NPX was detected at 0.043 ng/g and 0.04 ng/g, respectively. The low concentrations found in these studies generally agree with the findings of this study, but there is some variation in the tendency for specific compounds to accumulate. This variation may be partly attributed to the different analytical approaches. In other studies, uptake of PPCP/EDCs by plants was evaluated using non-labeled compounds, and accumulation was measured by targeted chromatographic analysis for the extractable parent compound. The use of 14C-labeled compounds in the current study should have provided “worst-case” estimates of human exposure, as the concentrations included non-extractable residue and likely also included transformation products. Transformation products may be an important component of potential risk since the metabolites of some PPCP/EDCs have higher biological activity than their parents and studies have shown that a large portion of PPCP/EDCs that are taken up by plants may be transformed in vivo . A translocation factor , which was the total 14C in stems, new leaves, and original leaves divided by the 14C in roots, was calculated . The derived TFs were consistently very small, demonstrating the poor translocation of these PPCP/EDCs from roots to upper tissues after uptake. The TF values followed the decreasing order of NPX > DCL > NP > BPA, the opposite observed for plant accumulation. Lettuce displayed lower TFs than collards for the same PPCP/EDCs. For example, the mean TF for BPA was only 0.010 ± 0.003 for lettuce, but was 0.051 ± 0.008 for collards. The much greater accumulation of PPCP/EDCs in roots, as compared to leaves, has been observed in previous studies. For instance, Herklotz et al. found that the levels in leaves were 0.00952 – 0.00503 of those in roots for cabbage grown in nutrient solution spiked with carbamazepine, salbutamol, sulfamethoxazole, and trimethoprim. Doucette et al. reported that the accumulation of NP in leaves was 0.0233 – 0.0167 of that in the roots of crested wheatgrass grown in solution.

Posted on

A two compartment model for SA dynamics on the human skin was developed and fitted to data

Some parameters are identifiable to a reasonable degree through model fitting, but there is a large degree of uncertainty in the viral transport efficiencies and the AD kinetic parameters. While this could be a consequence of fitting a limited number of data points with several parameters, the viral load at harvest and risk estimates were well constrained. This large variation in parameters and ‘usefully tight quantitative predictions’ is termed the sloppiness of parameter sensitivities, and has been observed in physics and systems biology. Well-designed experiments may simultaneously reduce uncertainty in the parameters as well as predictions and therefore increasing confidence in predictions. One possible experiment to reduce parameter uncertainty is recording the transpiration and growth rate to fit eq. independently to get at and bt . An interesting outcome of my analysis is the strong association of risk with plant growth conditions. The health risks from consuming lettuce irrigated with recycled wastewater are highest in hydroponic grown lettuce, followed by soil grown lettuce under Sc2 and the least in soil grown lettuce under Sc1 . This difference in risk estimates stems to a large degree from the difference in AD kinetic constants . Increasing katt,s will decrease risk as more viruses will get attached to the growth medium, while increasing kdet,s will have the opposite effect , as more detached viruses are available for uptake by the plant. The combined effect of the AD parameters depends on their magnitudes and is portrayed in Fig. A.4. This result indicates that a better understanding of the virus interaction with the growth environment can lead to an improved understanding of risk. More importantly,livestock fodder system this outcome indicates that soil plays a vital role in the removal of viruses from irrigation water through the adsorption of viral particles. An investigation focused on understanding the influence of soil composition on viral attachment will help refine the transport model.

The risk predicted by this dynamic transport model is higher than the EPA annual infection risk as well as the WHO annual disease burden benchmark. The reasons for this outcome are many-fold. First, there is a significant variability in the reported internalization of viruses in plants. In research of data for modeling NoV transport in plant, I filtered the existing data using the following criteria: 1) human NoV used as the seed agent, 2) presence of quantitative viral results in the growth medium and different locations of the plant. Based on these criteria, the data from represent the best available data on viral internalization and transport in lettuce. However, it is also important to note that a similar study by did not observe human NoV internalization in lettuce. This discrepancy could be due to the specific subspecies of the plant and growth conditions used in the studies. Besides, minor changes such as damages in roots or decrease in humidity of the growing environment can promote pathogen internalization. Alternatively, tracking viral transport through the growth medium and the plant is challenging, which may yield false results due to reaction inhibitions in genome amplification and inferior detection limit. The risk outcome of this study is conservative because it assumes an individual consumes the wastewater irrigated lettuce daily for an entire year. This assumption and the corresponding higher risk estimates are only applicable to a small portion of consumers, while most consumers in the U.S. are likely to have a more diverse diet. While the model outcomes presented here represent the best attempt given the available data, it is also possible that the internalization observed by is an extreme case and and typically internalization occurs to a lesser extent.As previously discussed by others , risk estimates by different NoV dose-response models differed by orders of magnitude. This study primarily aims to introduce a viral transport model without advocating any one dose-response model. The future refinement of pathogen dose-response models will reduce variability in risk estimates.

The risk of consuming lettuce grown in soil as predicted by is higher than my predictions, although I used the results of in both studies. This is a consequence of considering the greater adsorption capability of soil, which is not reflected when assuming a simple input:output ratio. Using different inoculating concentrations of NoV, body weight and consumption rate distributions also contributed to the difference in the outcomes but to a lesser extent. In addition to a transport model predicting the NoV load in lettuce, I explored strategies to reduce the risk of NoV gastroenteritis by increasing holding time of the produce after harvesting or using larger hydroponic culture volumes. Although neither strategy could significantly alleviate the risks, the process highlights two strengths of modeling: 1)It provides analytical support for arguments that would otherwise be less convincing; 2) It predicts outcomes of experiments without the physical resources required to perform them. For instance, the model can be used to explore alternate irrigation schedules to reduce the NoV internalization risk. Modeling also helps encapsulate our understanding of the system and generate hypotheses. For example, simple first-order decay did not produce the trend observed in the water, which suggests that additional mechanisms are at play. I postulated the attachment of virus particles on the walls of the hydroponic system as one possible mechanism and examined the fit of the model. Although viral attachment to glass or other materials has been observed before, here it stands as a hypothesis that can be tested. In addition to generating and testing hypotheses, some of my model assumptions raise broader questions for future research. For example, I assumed that viruses are transported at the transpiration rate from the growth medium to the roots. However, not much is known regarding the role of roots in the internalization of viruses. Investigating the defense mechanisms of plants’ roots to passive viral transport, i.e., through rhizosphere microbiome interactions, may shed light on the broad understanding of plant and microbe interactions. The question of extending this model to other pathogen and plant systems draws attention to the dearth of data in enabling such efforts. While modeling another virus may not require changes to the model, understanding transport in other plants can be challenging.

Data required includes models for growth rate and transpiration, plant growth characteristics including density, water content, as well as internalization studies to determine transport efficiencies. However, from the perspective of risk management, lettuce may be used as the worst-case scenario estimate of risk in water reuse owing to its high consumption with minimal pathogen inactivation by cooking. This worst-case scenario can be used to set water quality standards for irrigation water for the production of fresh produce eaten raw. The models can also be extended to include pathogen transport to the plant tissue from manure/biosolids that are used as organic fertilizer. By assuming that SA transitions from an un-adapted state to an adapted state, the model is grounded in first principles. The stochastic aspect of dose-response emerges naturally from a stochastic simulation of the growth kinetics. In addition, the model predicts carrier outcomes without additional data. Armitage et al. interpret results from several studies to posit that pathogens, including bacteria,hydroponic nft gully show an initial exponential increase in all individuals. We argue that this is not inconsistent with the initial decrease assumption for three reasons. Firstly, the exponential increase is observed in organs like the liver or spleen, and not the whole body or site of inoculation . This does not refute the possibility of an initial decrease at the inoculation site or the whole body. Secondly, the posited decrease is transient, and samples may not have been collected during this window. Thirdly, the magnitude of decrease is low at higher inocula and consequently less detectable. Further, compared to the initial decrease observed when all bacteria are in the S1 state, one would expect 1) no initial decrease if seeding with bacteria all in the S2 state, and 2) a smaller initial decrease if seeding with a mixture of bacteria in the S1 and S2 state. These trends have been observed when pathogens from in-vivo cultures were used for infecting the host . We note that the transition from S1 to S2 is perhaps not instantaneous, and the pathogen population may constitute a continuum of states between S1-S2. When loads were measured in the whole body, a transient decrease was observed in some cases . Clumping of bacteria was offered as a possible explanation, but this does not rule out an actual reduction in viable counts observed in other systems . Armitage et al. also note that non-responders show a subsequent decrease after the initial exponential increase. These were substantiated by measurements from survivors who were killed at later time points. This decrease is probably due to the activation of the adaptive immune response inside the host, which could be incorporated in a within-host variant of the 2C model. Using the concept of IED to evluate reponse, I am able to explain the data with a single IED. It has been observed that the toxic dose of a chemical can vary between individual subjects or with the season.

A similar stochasticity may be expected in IED between individuals which can be attributed to differences in covariates such as body weight, sex, immune history and biological noise. However, assuming this was not necessary to produce an acceptable fit. The model was fit to data by following a two step optimization procedure. Direct multi-objective optimization was not pursued since the objective functions were very different from each other. The deterministic ODE model was easy to evaluate and a global optimization algorithm was employed to guard against local minima while fitting the growth data. Fitting the dose-response data was computationally challenging for 3 reasons: a non-smooth objective function, stochastic simulations have to be repeated many times, and the number of stochastic entities being modeled is not small. Hence, a simple brute-force optimization was adopted. The RH model exhibits a sharp initial decline in SA density and predicts values lower than the observed minimum for each initial load . The 2C model only goes as low as the lowest load observed on the skin. Experiments similar to that of with greater time resolution are necessary to ascertain the time of true minimal SA density. The 2C model stochastic model does not perform as well as the RH model . However, the 2C model fit to dose-response data improves along the Pareto front . It is possible that exploring solutions with a higher growth objective may yield a solution that fits as well as, if not better than, the RH fit to the dose-response data. Moreover, the proposed approach offers advantages over the existing approach in that 1) it is fully mechanistic, and hence is more applicable in other scenarios , and 2) in addition to response outcomes, the proposed approach also accounts for carrier outcomes. Perhaps the most interesting outcome of this study is the incorporation of quorum sensing in dose-response modeling. The rejection of the absence of cooperativity in SA pathogenesis and the adequate fit of cooperativity make a strong case for the cooperativity in action hypothesis. Experimental support for this hypothesis include the well studied Agr system of quorum sensing . In the words of Le. et al, the Agr system “generally enhances pathogenesis by increasing expression of aggressive virulence determinants such as toxins and degradative enzymes”.This system is activated when bacteria reach a certain density, which results in a disease response such as a murine abscess. However, the 2C model posits that quorum sensing enhances bacterial growth rate, for which I propose two possible explanations. The direct explanation is the existence of an as yet undiscovered signaling mechanism responsible for density dependent growth enhancement. A second explanation relates to the events initiating response in a host, which is the interaction of the toxins/enzymes produced by SA with the host tissue. The 2C model captures these dynamics at a higher level of abstraction, with the mathematical variable i representing the amount QS signals and toxins. We can interpret b2 as the rate of enhanced production of these factors.

Posted on

Credible and prediction intervals in the shoot at harvest were similar for both models

However, Southern California, a region that suffers from a similar degree of water shortage, currently uses less than ~3% of municipal wastewater in agriculture, while discharging ~1.5 million acre-feet effluent per year into the Pacific Ocean . Secondary municipal wastewater effluent for ocean discharge is often sufficient to support both the nutrient and water needs for food production. Water reuse in agriculture can bring municipal water reclamation effluent to nearby farms within the city limit, thus promoting local agriculture and also reducing the rate of farmland loss to urban development. While the use of reclaimed water in agriculture offers a multitude of societal and agronomical benefits, broader adoption faces great challenges. One of the important challenges is ensuring the safety of food products in light of a plethora of human pathogens that may be present in recycled wastewater. Past studies have identified risks associated with irrigating food with recycled wastewater through the retention of the irrigation water on edible plant surfaces during overhead irrigation . With the emphasis on water conservation and reduction of evapotranspiration, subsurface drip irrigation is gaining popularity . Since there is lesser contact between water and the plant surface, the chance of surface contamination of pathogens is reduced. However, this new practice presents risk of uptake of microbial pathogens into plants. Such internalized pathogens are of greater concerns as washing, even with disinfectants, may not affect pathogens sheltered in the vasculature. Although pathogen transport through root uptake and subsequent internalization into the plant has been a growing research area, results vary due to differences in experimental design, systems tested, and pathogens and crops examined . Among the array of pathogens causing food borne illness that may be carried by treated wastewater, viruses are of the greatest concern but least studied. According to the CDC, 60% of U.S. food borne outbreaks associated with eating leafy greens were caused by noroviruses ,stacking pots while Salmonella and E. coli only accounted for 10% of the outbreaks . Estimates of global food borne illness prevalence associated with NoV surpass all other pathogens considered.

Viruses are also of concern because they persist in secondary wastewater effluents in high concentrations . They do not settle well in sedimentation basins and are also more resistant to degradation than bacteria . Therefore, in the absence of solid scientific understanding of the risks involved, the public are likely less receptive to adopting treated wastewater for agricultural irrigation. NoV internalization in hydroponic systems has been quantified by DiCaprio et al. . Internalization in crops grown in soil is considered lesser but nevertheless occurs. However, the only risk assessment that considered the possibility of NoV internalization in plants assumed a simple ratio of viruses in the feed water over viruses in produce at harvest to account for internalization. The time dependence of viral loads in lettuce was not explored and such an approach did not permit insights into the key factors influencing viral uptake in plants. In this study, we introduce a viral transport model to predict the viral load in crisp head lettuce at harvest given the viral load in the feed water. It is parameterized for both hydroponic and soil systems. We demonstrate its utility by performing a quantitative microbial risk assessment . Strategies to reduce risk enabled by such a model are explored, and a sensitivity analysis highlights possible factors affecting risk.The plant transpiration rate was adopted as the viral transport rate ) based on: 1) previous reports of passive bacterial transport in plants , 2) the significantly smaller size of viruses compared to bacteria, and 3) the lack of known specific interactions between human viruses and plant hosts . Accordingly, viral transport rate in hydroponically grown lettuce was determined from the previously reported transpiration model , in which the transpiration rate is proportional to the lettuce growth rate and is influenced by cultivar specific factors . These cultivar specific factors used in our model were predicted using the hydroponic crisp head lettuce growth experiment carried out by DiCaprio et al. described in Section 2.3 . Since the transpiration rate in soil grown lettuce is significantly higher than that in the hydroponic system, viral transport rate in soil grown lettuce was obtained directly from the graphs published by Gallardo et al. using WebPlotDigitizer . The shoot growth rate for soil grown lettuce was determined using Eq. 9 . In the absence of a published root growth model for lettuce in soil, a fixed root volume of 100 cm3 was used. In the viral transport model, viral transfer efficiency was used to account for the potential “barrier” between each compartment .

The existence of such a “barrier” is evident from field experiments where some microbial pathogens were internalized in the root but not in the shoot of plants . In addition, viral transfer efficiencies also account for differing observations in pathogen internalization due to the type of pathogen or lettuce. For example, DiCaprio et al. reported the internalization of NoV into lettuce, while Urbanucci et al. did not detect any NoV in another type of lettuce grown in feed water seeded with viruses. The values of ηgr and ηrs were determined by fitting the model to experimental data reported by DiCaprio et al. and is detailed in Section 2.3. The viral removal in the growth medium includes both die-off and AD, while only natural die-off was considered in the lettuce root and shoot. AD kinetic constants as well as the growth medium viral decay constant in the hydroponic case were obtained by fitting the model to the data from DiCaprio et al. . Viral AD in soil has been investigated in both lab scale soil columns and field studies . In our model, viral AD constants in soil were obtained from the experiments of Schijven et al. , who investigated MS2 phage kinetics in sandy soil in field experiments. As the MS2 phage was transported with the water in soil, the AD rates changed with the distance from the source of viruses. To capture the range of AD rates, two scenarios of viral behavior in soils were investigated. Scenario 1 used the AD rates estimated at the site closest to the viral source , while scenario 2 used data from the farthest site . In contrast to lab scale soil column studies, field studies provided more realistic viral removal rates . Using surrogate MS2 phage for NoV provided conservative risk estimates since MS2 attached to a lesser extent than NoV in several soil types . The viral decay rate in the soil determined by Roberts et al. was adopted because the experimental temperature and soil type are more relevant to lettuce growing conditions compared to the other decay study . Decay rates in the root and shoot were used from the hydroponic system predictions.The transport model was fitted to log10 viral concentration data from DiCaprio et al. , extracted from graphs therein using WebPlotDigitizer . In these experiments, NoV of a known concentration was spiked in the feed water of hydroponic lettuce and was monitored in the feed water, the root and shoot over time.

While fitting the model, an initial feed volume of 800 mL was adopted and parameters producing final volumes of b200 mL were rejected. To fit the model while accounting for uncertainty in the data, a Bayesian approach was used to maximize the likelihood of the data given the parameters. A posterior distribution of the parameters was obtained by the differential evolution Markov chain  algorithm,strawberry gutter system which can be parallelized and can handle multi-modality of the posteriors distribution without fine tuning the jumping distribution. Computation was carried out on MATLAB R2016a and its ParCompTool running on the High Performance Computing facility at UC Irvine.Table 3 lists the parameters estimated by model fitting and their search bounds. Fitting data from DiCaprio et al. without including viral AD to the tank walls was attempted but the results were not used in the risk estimates due to the poor fit of model to the data. The rationale behind the model fitting procedure and diagnostics are discussed in Supplementary section S1H.A summary of the model fitting exercise for viral transport in hydroponic grown lettuce is presented in Fig. 2. Under the assumption of first order viral decay, NoV loads in water at two time points did not fall in the credible region of model predictions, indicating that mere first order decay was unsuitable to capture the observed viral concentration data. The addition of the AD factor into the model addressed this inadequacy and importantly supported the curvature observed in the experimental data. This result indicates the AD of viruses to hydroponic tank wall is an important factor to include in predicting viral concentration in all three compartments .The adequacy of model fit was also revealed by the credible intervals of the predicted parameters for the model with AD . Four of the predicted parameters: at, bt, kdec, s and kp, were restricted to a smaller subset of the search bounds, indicating that they were identifiable. In contrast, the viral transfer efficiency η and the kinetic parameters spanned the entirety of their search space and were poorly identifiable. However, this does not suggest that each parameter can independently take any value in its range because the joint distributions of the parameters indicate how fixing one parameter influences the likelihood of another parameter . Hence, despite the large range of an individual parameter, the coordination between the parameters constrained the model predictions to produce reliable outcomes . Therefore, the performance of the model with AD was considered adequate for estimating parameters used for risk prediction.Risk estimates for lettuce grown in the hydroponic tank or soil are presented in Fig. 4. Across these systems, the FP model predicted the highest risk while the 1F1 model predicted the lowest risk. For a given risk model, higher risk was predicted in the hydroponic system than in the soil. This is a consequence of the very low detachment rates in soil compared to the attachment rates. Comparison of results from Sc1 and Sc2 of soil grown lettuce indicated lower risks and disease burdens under Sc1 . Comparing with the safety guidelines, the lowest risk predicted in the hydroponic system is higher than the U.S. EPA defined acceptable annual drinking water risk of 10−4 for each risk model. The annual burdens are also above the 10−6 benchmark recommended by the WHO . In the case of soil grown lettuce, neither Sc1 nor Sc2 met the U.S. EPA safety benchmark. Two risk models predicted borderline disease burden according to the WHO benchmark, for soil grown lettuce in Sc1, but under Sc2 the risk still did not meet the safety guideline. Neither increasing holding time of the lettuce to two days after harvesting nor using bigger tanks significantly altered the predicted risk . In comparison, the risk estimates of Sales-Ortells et al. are higher than range of soil grown lettuce outcomes presented here for 2 of 3 models. The SCSA sensitivity indices are presented in Fig. 5. For hydroponically grown lettuce, the top 3 factors influencing daily risk are amount of lettuce consumed, time since last irrigation and the term involving consumption and ρshoot. Also, the risk estimates are robust to the fitted parameters despite low identifiability of some model parameters . For soil grown lettuce, kp appears to be the major influential parameter, followed by the input viral concentration in irrigation water and the lettuce harvest time. Scorr is near zero, suggesting lesser influence of correlation in the input parameters.In this study, we modeled the internalization and transport of NoV from irrigation water to the lettuce using ordinary differential equations to capture the dynamic processes of viral transport in lettuce. This first attempt is aimed at underscoring the importance of the effect of time in determining the final risk outcome. The modeling approach from this study may be customized for other scenarios for the management of water reuse practices and for developing new guidelines for food safety. Moreover, this study identifies critical gaps in the current knowledge of pathogen transport in plants and calls for further lab and field studies to better understand risk of water reuse.

Posted on

Carbon captured in soil already exists and is merely captured rather than permanently destroyed

This proposed bill did not pass, but CARB is in the process of constructing a webpage that will provide information on proposed offset programs, and the agency affirmatively notes that it will be considering additional offset programs as a part of future rule making activities.Thus, an agricultural soil carbon sequestration offset program may not be far from being proposed and considered as a possible offset program under California’s cap and trade program.Ensuring that emission reductions are quantifiable, permanent, and additional are important considerations for any type of offset program but are particularly difficult in relation to agricultural soil carbon sequestration offset programs. Quantifiability and permanency are especially central concerns about offset programs, and it has been suggested that agricultural soil carbon sequestration plays such a minimal role in major carbon markets because soil carbon is considered difficult to measure, verify, and track.Deciding how to allocate offset credits can be challenging in any carbon sequestration program because it is difficult to accurately quantify how much carbon has really been sequestered.Soil carbon sequestration depends on a complicated living system that is constantly changing and not easy to directly quantify. A variety of factors determine how much carbon a unit of soil can sequester, including seasonal variations, weather, precipitation, plant species present, and the variation in soil type and quality.This problem does not arise in offset programs that decrease emissions from point sources, such as smokestacks or manure lagoons,rolling benches for greenhouse where measurement is more concentrated and accurate methods of measurement are established and verifiable. For example, a manure lagoon equipped with a BCS can use a meter to determine methane emissions from the entire lagoon. Adding to the complication of quantifiability, some studies dispute whether conservation tillage practices actually sequester carbon at all.

The goal of permanency is problematic in agricultural soil carbon sequestration programs because the carbon reduction is easily reversible.When carbon is sequestered in soil, it can be re-released into the atmosphere from a disturbance such as increased intensity of tilling, wind or water erosion, or a natural disaster such as an earthquake, fire, or disease outbreak.One agricultural soil carbon sequestration offset program incorporates a 60% discount into its program to account for the uncertainty of permanency.Compare this to destroying a unit of methane with a BCS or reducing a unit of carbon emissions from a smokestack by installing new technology. When that methane or carbon unit is destroyed or never created, that reduction is not reversible because it never existed.Simple disturbances can cause the loss of some or all of the carbon that was stored in the soil and essentially negate any climate change benefit.Additionality is also at issue with agricultural soil carbon sequestration offset programs because cropland conservation practices such as no till and conservation tillage are already widely used in at least some parts of the United States due to incentives programs set up by the USDA starting in the 1960s and 1970s.The USDA study on cropland conservation practices in the Missouri River Basin indicates that within the 95 million acres of cropland studied between 2003 and 2006, 46% of the cropland met no-till criteria and 97% of the cropland “had evidence of some kind of reduced tillage on at least one crop in rotation.”Considering that cropland conservation practices seem to be common in at least some parts of the country, it may be difficult to tell if any given offset project under an agricultural soil carbon sequestration offset program would have occurred anyway in a business-as-usual scenario for purposes of determining additionality.The checkpoints for offset programs—that the offset credits generated are quantifiable, real, permanent, and additional—do not explicitly include an analysis of the trade offs or incidental effects of an offset program. However, in agricultural soil carbon sequestration offset programs, the considerations of incidental effects caused by the offset program should be a critical checkpoint to consider.

Some of the conservation practices that most effectively sequester carbon in agricultural soil can present trade offs that bring new problems for farmers that must be fixed through alternative means. Primarily, tilling decreases weed growth, so farmers who infrequently or never till typically use more herbicide to keep weeds out of their field.Farmers using the other sequestration practices encouraged under soil carbon sequestration programs besides no-till and conservation tillage are also reported as using much larger quantities of herbicide. For example, the Kenya Agricultural Carbon Project does not address the use of herbicides, and the World Bank reported that herbicides are heavily used on farms that are involved with the project.An environmental and social monitor for a soil carbon sequestration program reported that “the herbicides are applied . . . without due regard to environmental consequences.”At least one assessment reported contrary findings, that less herbicide was used when conservation practices were employed.However, this assessment utilized many types of conservation practices including improved pesticide management practices, which could explain the decrease in herbicide use in this study. Increased herbicide use can be detrimental for reasons including environmental harm, pollutant emissions, and human health. Herbicides can migrate into the surrounding environment through soil, air, and waterways.The resulting chemical residues can negatively affect the natural surroundings, as any chemical might.The effects would depend largely on the toxicity of the chemicals used in the herbicide, the quantity used and leached, and the sensitivity of the surrounding environment. Additionally, harmful air pollutants, including greenhouse gases, are released when using herbicides.Herbicides release a large amount of nitrous oxide, a powerful pollutant with an estimated 298 times the global warming potential of carbon dioxide.This can be seen as similar to the problem of copollutants. Co-pollutants are pollutants that are released simultaneously and from the same source as the greenhouse gas or pollutant at issue. Increasing emissions of the pollutant at issue will often increase co-pollutant emissions, which can be more localized and harmful in smaller quantities. Similarly, increasing herbicide use will increase nitrous oxide emissions that would not have otherwise occurred if not for increased herbicide use. Thus, even if an agricultural soil carbon sequestration offset program is measured to be carbon neutral, it may unintentionally provide an avenue for increased nitrous oxide emissions and harm to the environment. Another incidental effect of increased herbicide use is that more chemicals will be put onto our food products and affect human uses of soil, water, and air.Herbicides have been linked to serious diseases, such as non-Hodgkin’s lymphoma, soft-tissue sarcoma, and Parkinson’s disease.Due to these health risks, some countries have started to mandate that farmers reduce the amount of herbicide used on their crops due to the harmful human health effects of herbicides.

These circumstances have led some to sharply oppose the increased use of herbicide. If CARB considers including an agricultural soil carbon sequestration offset program in AB 32’s repertoire of offset programs,cannabis grow systems the issues of quantifiability, permanency, additionality, and incidental effects of the offset projects should be addressed. Implementing a new agricultural soil carbon sequestration offset protocol under AB 32 without considering and compensating for these issues would jeopardize the purpose of AB 32’s cap and trade program and likely inflate AB 32’s carbon market with credits that do not actually represent the additional sequestration of one ton of carbon dioxide or its equivalent.These issues may be most completely and accurately addressed by using an ecosystem approach to design the offset program and to approve and implement the resulting offset projects in a case-by-case manner. An ecosystem management approach acknowledges the inter connectivity of the parts within an ecosystem and sees the environment as a single functioning landscape.This approach recognizes that considering only a single species, pollutant, or practice can be detrimental when it successfully decreases one harm but incidentally increases another harm that may be just as, if not more, harmful to the ecosystem. Accordingly, any increase in herbicide use, and subsequent nitrous oxide emissions, or any other potentially harmful externality would be accounted for in the offset program. Under an ecosystem approach, offset programs would not favor projects or regulations that induce harms of a larger or more detrimental magnitude than the harm which is to be prevented by the program. The need for an ecosystem approach, and the regulating agency’s response to this need, is illustrated by the Endangered Species Act.Certain species are listed and protected under the Endangered Species Act and a federal budget is allocated to preserving those listed species. However, many believe that the environment and society would be better served by protecting and managing ecosystems on a larger scale as opposed to individual species.Recognizing this and similar needs in different areas under their jurisdiction, the U.S. Fish and Wildlife Service published An Ecosystem Approach to Fish and Wildlife Conservation, which includes guidelines the FWS strives to use in order to incorporate the ecosystem approach into their conservation work. The Pacific Islands Forum Fisheries Agency , a group formed to help its Pacific Island members to manage, control, and develop the fisheries within the Exclusive Economic Zones, encourages its countries to utilize an ecosystem approach to manage their fisheries.The FFA’s Ecosystem Approach to Fisheries Management consists of four steps.The first step is to determine the scope of the assessment by clearly identifying what is to be managed.The second step is to identify all the issues to be assessed within five key areas and to agree on the values that are to be achieved for each issue.The third step is to determine which issues should be directly managed.The last step is to determine acceptable performance levels, what management arrangements will achieve these levels, and the review process for assessing performance.During the creation of the offset program, an ecosystem approach could be utilized to determine whether the program should be created at all. If no agricultural soil carbon sequestration offset project could ever in theory have a net benefit when considering all the greenhouse gas sources and sinks and other externalities created by an individual project, then the analysis under an ecosystem approach may indicate that the offset program should not be approved. If the analysis of the offset program under an ecosystem approach indicates that only certain types of projects could result in a net benefit to the environment, the program could be limited to those particular types of projects. The ecosystem approach could also be used to assist in determining on a case-by-case basis whether an agricultural soil carbon sequestration offset project should be approved under the offset program. Because the variables associated with each agricultural soil carbon sequestration project will be different for each project and have the potential to vary greatly, a case-bycase ecosystem approach for the approval process for each project would help decision makers to properly determine whether the offset project is quantifiable, permanent, real, and additional. Which externalities should be included in an ecosystem approach analysis of agricultural soil carbon sequestration projects would be a basis for much disagreement, and would depend on scientific and policy analysis beyond the scope of this Comment. However, at a minimum, the effects of increased herbicide use on the surrounding ecosystem and the increase of nitrous oxide emissions should be included in the analysis, as those are some of the more egregious oversights in certain existing agricultural soil carbon sequestration programs, as discussed in Part V.B. In addition to legitimizing a future offset program and resulting offset projects in general, applying case-by-case and ecosystem approaches have the potential to resolve specific issues regarding quantifiability, additionality, and incidental effects identified in Part V.At least two main methods of quantifying agricultural soil carbon sequestration for the purposes of allocating credits for agricultural soil carbon sequestration offset projects could be envisioned. One is a simpler standards-based approach and the other follows a case-by-case process. Although a case-by-case approach may be impracticable in practice, this example illustrates why a case-by-case approach would be more appropriate and crucial for an agricultural soil carbon sequestration offset program. The standards-based option is to give an offset credit per a certain acreage of land covered by an offset project.That particular acreage of land would, on average, sequester one metric ton of carbon dioxide or carbon dioxide equivalent regardless of individual features of the land.

Posted on

The main reason was a reduction in direct and indirect nitrous oxide emissions

Phase I also considered agricultural adaptation strategies that addressed regional issues such as hydrology, growers’ attitudes toward climate change, and urbanization versus preservation of farmland. These topics are explored in more quantitative ways here.Since 1960, total crop acreage in Yolo County has been declining. Vegetable and orchard crop areas have increased, while field crop acreage has declined . There has been an increase in higher‐revenue‐per‐acre crops, especially a shift out of barley, and a shift into more processing tomatoes, wine grapes, and walnuts. Many factors affect changes in acreage, including changes in market conditions , input supplies, and climate. Among factors affecting acreage decisions, we investigated whether changes in climate have affected acreage allocations across crops. If responses to climate changes in the past continue to hold in the future, we can use hisorical information to learn more about how crop acreages are likely to change in response to the forecasted Yolo County climate changes from 2010 to 2050. We developed econometric models that relate acreages of each major crop to relative prices and key climate variables . The models are applied to the data including 60 years of acreage for major crops and 100 years of local climate history. Our climate history indicates that during the past century, the increase in annual temperature appears to be mainly due to warmer winters rather than to warmer summers . There was a decrease of about 150 winter chill hours in the last 100 years. Using historical reationships between climate and acreage allows investigation of how forecasted climate changes in Yolo County may affect Yolo acreage patterns. Acreage projections use climate projections for the B1 and A2 scenarios from 2010 to 2050 with GCM data from GFDL‐BCCA. Acreage projections hold constant relevant drivers of crop acreage, except for local climate variables. Among field crops,planting gutter warmer winter temperatures were projected to cause wheat acreage to decline and alfalfa acreage to rise . Thus, future decisions to increase alfalfa acreage present an interesting implication for water use: wheat uses little irrigation; whereas, alfalfa is one of the more intense water users.

By 2050, tomato acreage is projected to increase compared to the current level . This is also related to the increase in growing degree days in the winter months. A warmer climate in late winter/early spring has allowed early planting and provided favorable conditions for establishment. The forecasted climate changes have only moderate impacts on projected tree and vine crop acreage, in part because the climate changes that have occurred have not yet affected key variables enough to induce a significant change in the acreage of perennials when market conditions have been favorable. Almond acreage is projected to increase slightly with warmer temperatures in 2035–2050 . Almonds have a relatively low winter chill hour requirement. Walnut acreage, however, would decline slightly ; it has a higher winter chill requirement. This is consistent with the finding that surveyed orchard growers express concern about a decrease in winter chill hours . These projections rely on using historical relationships between acreage change and climate variables change. They are based actual past responses of acreage to climate. However, no attempt is made to forecast the relative prices, technical changes, new markets, or other factors that will also affect acreage. Water supply vulnerabilities for agriculture and other sectors can be mediated through traditional infrastructure improvements or alternative water policies . Local stewardship that is implemented by water managers and agricultural users tends to be more economical and have less environmental impact than developing new supplies. One tool that has helped water resource managers integrate climate change projections into their decision making process is the Water Evaluation and Planning system . WEAP, a modeling platform that enables integrated assessment of a watershed’s climate, hydrology, land use, infrastructure, and water management priorities , is used here for the Yolo County Flood Control and Water Conservation District service area. It covers 41 percent of the county’s irrigated area and is located in the western and central portion of the county .Recognizing the key role that land‐use planning will play in achieving the goals of AB 32, legislators passed Senate Bill 375 in 2008, requiring sustainable land‐use plans that are aligned with AB 32 .

Local governments must address GHG mitigation in the environmental impact report that accompanies any update to their general plan or carry out a specific “climate action plan” . Emissions of GHG from agriculture are often missing from existing inventory tools geared to local planners. The local government of Yolo County was among the first in California to pass a climate action plan . This project contributed to this climate action plan, and developed a set of guidelines to estimate GHG emissions from agriculture within a local inventory framework . The Tier 1 methods used here have been adapted for local activity data largely from three main sources: the California Air Resources Board Technical Support Document for the 1990–2004 California GHG Emissions Inventory ; the U.S. Environmental Protection Agency Emissions Inventory Improvement Program Guidelines ; and the 2006 IPCC Guidelines for National GHG Inventories . In Yolo County, total agricultural emissions declined by 10.4 percent between 1990 and 2008 .Lower fertilizer use was driven by two important land use trends: a 6 percent reduction in the county’s irrigated cropland; and a general shift away from crops that have high N rates coupled with an expansion in alfalfa and grape area, which require less fertilizer . In both years, emissions of CO2, N2O, and methane from diesel‐powered mobile farm equipment were responsible for 20.0 to 23.0 percent of total agricultural emissions in Yolo County between 1990 and 2008 . Fuel consumption per unit area for several important crops offset the small decline in irrigated cropland. Using the Tier 1 method prescribed by ARB, emissions of CH4 from rice cultivation were estimated to increase from 25.9 to 31.2 kilotons carbon dioxide equivalent between 1990 and 2008, entirely due to an expansion in the area under rice cultivation. Studies also suggest that cultivation practices that combine straw incorporation and winter flooding tend to generate more CH4 emissions than burning rice straw . Thus, estimates generated using the DeNitrification‐DeComposition model showed a larger increase in emissions over the study period because the Tier 3 method accounted for changes in residue and water management made in compliance with the state air quality regulations that have phased out rice straw burning, and the increase in cultivated area . 

Many agricultural practices to mitigate GHG emissions offer agricultural co‐benefits. For example, economic factors are prompting local farmers to shift more of their land to crops that happen to require less N fertilizer and diesel fuel, and to adopt practices that reduce these inputs. Growers cite rising cost and market volatility of inputs, rather than mitigation per se, as a more immediate motivation to use fertilizer and fuel more efficiently. In 1990, emissions sources associated with urban areas accounted for approximately 86 percent of the total GHG emissions countywide,gutter berries while unincorporated areas supporting agriculture were responsible for 14 percent . If calculated on an area‐wide basis the county’s urban areas emitted approximately 152.0 tons CO2e per hectare per year . By contrast, this inventory results indicates that in 1990 Yolo County’s irrigated cropland averaged 2.16 t CO2e ha‐1 yr‐1 and that livestock in rangelands emitted only 0.70 t CO2e ha‐1 yr‐1 . This 70‐fold difference in the annual rate of emissions between urbanized land and irrigated cropland suggests that land‐use policies that protect existing farmland from urban development are likely to help stabilize and or reduce future GHG emissions, particularly if they are coupled with “smart growth” policies that prioritize urban infill over expansion .Many factors affect farmers’ perceptions and response to climate change; for example, characteristics of the individual farmer and their farm; social networks and involvement in programs run by local institutions, agricultural organizations, and extension services; and views on government programs and environmental policies. The goal of this sub-project is to: examine Yolo County farmers’ perceptions of climate change and its risks to agriculture; and develop a better understanding of how such factors might influence farmers’ adoption of proposed adaptation and mitigation practices. We conducted semi‐structured interviews with eleven farmers and two agricultural extension workers in the fall of 2010. The sampling strategy recruited respondents from a cross section of farm sizes, local cropping systems, and market orientations. Interviewers followed a set of open‐ended questions to minimize prompting and interviewer bias, and were used to develop a quantitative survey which was mailed to farmers in Yolo County during February and March of 2011. The survey sample was drawn from a list of 572 individuals who have submitted conventional or certified organic pesticide use permits to the Yolo County Agriculture Commissioner’s office. The final response rate was 34.0 percent. Results of the survey indicated that 54.4 percent of farmers agreed to some extent with the statement “the global climate is changing” . A minority indicated that local summer temperatures had decreased over time, while only 5.6 percent observed an increase. While contrary to statewide mean temperatures, this corresponds with local climate records which show little change in maximum summer temperature over the last century . A majority of farmers indicated that rainfall, drought, and flooding had not changed over the course of their career, but a sizable minority reported water availability had decreased and <1 percent said it had increased. In 1976, the newly constructed Indian Valley Reservoir began supplementing the District’s surface water supplies to local growers.

However, a recent drought in 2009 and 2010 reduced water releases in those years to less than 40 percent of the average for the preceding decade . The memory of this recent a drought may therefore occupy a central place in farmers’ perception of water related trends. Respondents with greater concern for drought and less reliable water were more likely to pump groundwater, drill new wells, and adopt drip irrigation . A farmer’s views on climate change affected the inclination to implement voluntary mitigation practices. More specifically, farmers who disagreed with the statement “The global climate is changing” were less likely to adopt mitigation practices than those who agreed with the statement. Likewise, skepticism that human activities are an important cause of climate change meant less inclination to adopt mitigation practices. Farmers who had frequent contact with local agricultural organizations were more likely to implement mitigation strategies Farmers are often more concerned about the future impact of government regulations than they are about the direct impacts of climate change. This ranking of concern is not surprising given the gradual nature of climate change. However, it does underscore the importance of understanding how farmers view environmental regulations and the information needed to influence their likelihood to adopt mitigation and adaptation practices. Strategies to expand the reach of local agricultural organizations and government conservation programs by improving farmer participation in their activities are thus seen as an important way to strengthen adaptation and mitigation efforts.UPlan relies on a number of demographic inputs . Attractors are given a positive value . Discouragements are given negative values . A system of weights is used to rank the attractive or discouraging property of each variable. We modified UPlan to allow development within existing urban areas, on the assumption that a significant urban redevelopment is likely within the 2010–2050 time frame. The A2 scenario loses two times more acreage of high quality soils to urbanization compared to B1 . One of the most striking findings is just how little land is required to house future populations at higher densities. The B1 and AB 32+ scenarios require 44 percent and 7 percent of the urbanized land of the A2 scenario, respectively. Even holding population increase constant at B1 levels, these scenarios use 63 percent and 38 percent of the land of the A2 scenario, most or all of it within existing urban areas, and also greatly reduce GHG emissions from transportation. These results suggest that the most important climate change mitigation policy that Yolo County could adopt would be to restrict urban development to infill locations within existing cities, and to keep existing farmland in agriculture.

Posted on

Many ecological processes governing agricultural pest abundance occur over a large spatial scale

Additionally, as the amount of land in cropland increases, opportunities for invasion or refuge from pesticide applications may be reduced, thus leading to a negative effect of landscape simplification on pesticide use. Three recent reviews of empirical, landscape-scale ecological studies evaluating the effect of landscape complexity on insect pests reported similarly equivocal results, with some studies finding reduced pest pressure, pest abundance, or pest diversity, whereas others find no relationship or the opposite relationships . The variability in the literature may reflect the inadequacy of current study designs to disentangle the net effect of landscape simplification on pesticide use. Confounding variables, such as crop type, or endogenously determined variables, such as farm size or income, could give misleading results if not properly controlled for. Alternatively, studies that are small scale or over short time periods may miss important underlying drivers of pest abundance. Pests disperse large distances, both naturally and aided by the movement of people and goods. Agricultural pests are thus likely governed in large part by meta population processes . Within an agricultural landscape pests may go locally extinct from crop patches because of pesticide use or because of stochasticity influencing small populations, only to be recolonized from a persistent meta population existing in the surrounding agricultural matrix or from a new invasion into the system. Natural enemies too may require resources outside of individual crop fields for alternative prey and shelter for overwintering or from disturbances, such as pesticide application or harvest . Furthermore,dutch buckets the periodic disturbance of crop fields may disrupt predator–prey dynamics by reducing natural enemies directly or by temporarily reducing pest populations to the level below which predators can be supported.

As a result of pest and natural enemy dispersal and immigration, the effect of local processes on regional abundances may be small, despite large effects on within-field abundances. Thus, small-scale studies that fail to account for the landscape-scale dynamics of agricultural pests and their natural enemies could result in spurious associations of what promotes or limits pest abundance. For these reasons, landscape-scale studies provide the best insight into the effect of habitat simplification on pests . Beyond meta population dynamics and trophic interactions, invasion and spread of insect pests and natural enemies are partly stochastic processes influenced by yearly environmental conditions and by the timing of insect pest and natural enemy arrival . Thus, temporal scale may be equally as important as spatial scale to disentangle the effects of landscape simplification on pest abundance. For example, a heat wave at the right time of the growing season may result in widespread pest mortality and high crop yields, whereas a heat wave at a different time of the season may stress crops, making them more susceptible to pest outbreak but having little effect on the pests themselves. This variability over time could appear like ambivalent results of landscape simplification when it is instead the result of the interaction between insect pests and weather. If we are to mitigate the effects of pesticide use on both human health and ecological systems, it is necessary to understand the underlying abiotic or biotic factors resulting in differences in pesticide use. Here I take advantage of longitudinal data from the US Department of Agriculture Census of Agriculture to revisit whether landscape simplification is a consistent driver of insecticide use. I perform cross-sectional analyses for five USDA census years in seven Midwestern US states at the county level. I follow this with a panel data analysis using a fixed-effects model, which identifies the effect of landscape simplification on insecticide use using year-to-year variation within counties.

I specifically focus on insecticides in these states to compare this multiyear analysis with a recent single-year study by Meehan et al. . I check the robustness of these results by comparing data from the USDA Census of Agriculture to the National Agricultural Statistics Service Cropland Data Layer , and check different selection criteria for included counties. I compare these results to that of Meehan et al. , who used the same data sources and model specifications for 2007 only, and find that incorporating multiple years of data as I do here provides insights impossible to glean from a single data year.Annual expenditure on insecticides is over 4 billion dollars in the United States , which equates to the use of almost 100 million pounds of active ingredients . Given the many health and environmental consequences related to insecticide exposure, it is critical to understand what farm, landscape, or environmental characteristics drive the insect pests that motivate insecticide use. It has long been thought that landscape simplification is one of these characteristics. Reviews of empirical evidence for this theory have been largely inconclusive , although a recent statistical analysis of the Midwestern United States in 2007 found a strong, positive relationship between landscape simplification and insecticide use . Here I analyzed data from five USDA Census of Agriculture years using cross-sectional and fixed-effects models. The cross sectional results show that landscape simplification does not consistently drive higher insecticide use. Although the coefficient on proportion of county in cropland, my metric for landscape simplification, is positive and significant in the 2007 analyses, that relationship is absent or reversed in prior census years. Furthermore, adjacent census years, such as 2002–2007 and 1992– 1997, show large changes in the magnitude and changes in significance of the landscape-simplification coefficient.It is evident that the drivers of insecticide use may not be easily or reliably identified using single time-period studies. Using a fixed-effects model to remove unobserved characteristics, I find a non-significant relationship between landscape simplification and proportion of county in cropland. Counter intuitively, these results suggest that as cropland increases, the proportion of cropland sprayed with insecticides is unaffected.

The existence of a null relationship between landscape simplification and insecticide use is not unlike the results of Hutchison et al. , who reported large reductions in the European corn borer in non-Bacillus thuringiensis corn as a positive externality from B. thuringiensis corn plantings. Although pesticides may have negative effects on public health, biodiversity, and ecosystem services,grow bucket the application of pesticides by a nearby farm may reduce pest incidence on surrounding farms because of pesticide drift or pest suppression . Additionally, as the amount of land in cropland increases, opportunities for invasion from natural or untreated areas may be reduced. As a result of landscape simplification, natural lands have been isolated to farm boundaries, fallow lands, or wood lots . Numerous ecological studies have found that these fragmented natural or less intensively managed areas can act as a source for natural enemies and pest species that recolonize species poor crop fields . If the cost of pest invasion is greater than the benefits of natural enemy pest suppression stemming from non-crop land, these habitats can have a net negative impact on the farmer in terms of pest control. The above mechanisms may explain why a null relationship is observed in the fixed-effects model; however, they do not account for the importance of year. What could explain the wild variation in the landscape simplification coefficient in the cross sectional analyses and why year fixed effects are so important? There are a number of drivers that could be behind the year-to year variability, and deciphering which mechanism is at play is critical because different policy measures are needed to address different types of drivers. For example, a stochastic driver such as weather could be the culprit. Insect development is strongly influenced by weather conditions, such as temperature and precipitation, and thus yearly differences in these or other environmental conditions could have an important effect on insecticide demand and the relationship between landscape simplification and insecticide use. Preliminary analysis indicates that the effect of weather on this relationship is complex. [Preliminary analysis using growing season precipitation and degree days based on the National Climatic Data Center Global Historical Climatology Network Daily file does not explain the variation in the cross-sectional relationship between landscape simplification and insecticide use.] This finding may be because the timing of pest arrival relative to the growing season may determine the likelihood of pest outbreaks and the benefits of applying insecticides . Furthermore, temperature and precipitation affect the survival and development of different pests differently, and thus which pests and enemies are present may determine the effect of weather on the relationship between landscape simplification and insecticide use. Refined data on pest outbreaks or type and timing of insecticide use are currently not available for the study area examined. However, the development of such data or further empirical study focusing on abiotic conditions would greatly increase our understanding of the link between weather events and insect outbreaks, and thus increase our ability to forecast variation in insecticide use both now and under future climate change. It is also conceivable that the change in the relationship between landscape simplification and insecticide use between 2007 and all previous years reflects a systematic and predictable trend in insecticide use. For example, in 1996 there was a major change in the regulation of pesticides in the form of the Food Quality Protection Act .

FQPA prompted the reevaluation of all registered pesticides, and promoted the use of more selective, less persistent “reduced-risk” pesticides via a fast-track registration process . FQPA could affect the relationship between landscape simplification and insecticide use because insecticides that are effective against a multitude of insect pests and persist in the environment for longer periods of time may have provided higher positive externalities to surrounding crop fields, thus necessitating less insecticide use in landscapes dominated by agricultural fields. The implementation of FQPA and the resulting use restrictions took 10 y, and phasing out of certain chemicals is still in progress . Because changes in available insecticides were occurring between 1996 and 2007, it is difficult to statistically evaluate the effect of FQPA on the results reported here. Future Census’ of Agriculture or more refined insecticide data that include information on the active ingredient in use could elucidate how policy changes are interacting with the relationship between landscape simplification and insecticide use. Agriculture has vast impacts on the Earth’s environment and these impacts are only expected to grow as demand increases in the coming decades . The challenge, as Balmford et al. discuss, is how to meet the increasing demand with the least effect on native biodiversity and the ecosystem services intact ecosystems provide. There are various advantages and disadvantages to whether increased demand should be met by increased intensity of farming on current agricultural land or by increased land conversion to agriculture to be farmed with more biologically harmonious farming methods . In the Midwestern United States, it appears that land-sparing at the county level does not lead to consistent increases in the proportion of cropland treated with insecticides. However, without understanding what is behind the year-to-year variation in the relationship between landscape simplification and insecticide use, it is impossible to predict how land sharing or land-sparing as a policy initiative would affect insecticide use in the future. As suggested by this study and recent empirical reviews , the presence and direction of the relationship between landscape simplification and insecticide use can be positive, negative, or null. If this variation is driven by variation in yearly weather, whether simplified landscapes cause more or less insecticide use could flip flop unpredictably. If the variation is driven by extreme weather or weather characteristics that will be altered with climate change, perhaps there will be some directionality. If the relationship between landscape simplification and insecticide use is an indirect consequence of management policies, perhaps 2007 is a glimpse of the future. The data available are currently inadequate to decipher the underlying mechanisms. However, given the different policy implications of a stochastic driver, such as weather, versus a predictable driver, such as regulatory change, developing the necessary data sources to tease apart the underlying causes is imperative. Perhaps most importantly, this study emphasizes the need for longer-term research agendas, especially when investigating a politically, economically, and ecologically important question, such as insecticide or pesticide use.

Posted on

Terpenoid synthesis showed clear segregation into the mevalonate and the MEP/DOXP pathways

Airborne dust was ever-present while we sampled, and despite extensive containment efforts, some exposure of the apparatus to dust occurred . However, the rest of the DOM metagenome is remarkably distinct from the other samples, suggesting that, with the exception of the known contaminants, it is still representative of the deep groundwater. We eliminated all suspected contaminant genomes from further analysis.We assembled genomic bins using the following pipeline: IDBA_UD initial assembly, REAPR breaking of misassemblies, VizBin binning of contigs into draft genomes using 5-mer frequency and coverage information for visual aid, additional manual bin cleanup based on contig coverage distribution , taxonomic assignment of bins based on RAPSEARCH to the UniProt UniRef100 database. Selected bins were assembled further by PRICE targeted assembly and REAPR correction of misassemblies. Genome quality was assessed throughout using CheckM. Further details are in the methods. Overall we assembled and refined 79 unique genomic bins from the three groundwater samples . An additional 51 bins were made from the LAG sample and were used to determine probable contaminants in other samples, but no refinement of these bins was attempted, as we were interested in the properties of the groundwater communities. Contamination was only detected for the DOM sample as previously discussed. No evidence of overlap with the surface water was seen in either MW5 or MW6. The most abundant taxa in the partial genomic bins were from the CPR Parcubacteria , followed by the DPANN Woesearchaeota , and the CPR Microgenomates . While the OD1 and OP11 lineages have previously been found in association with anammox communities. The high relative abundance and diversity of CPR and DPANN genomes at over 50% of the community is notably higher than previous reports . The next highest abundance of our genomic bins were from the Planctomycete Brocadiaceae family . We made genomic bins of many of the other key members of the wastewater anammox community, including Omnitrophica , Nitrospirae , Chloroflexi , Chlorobi , Bacteroidales , Acidobacteria , and γ-Proteobacteria . Additionally, we assembled partial genomes from the Nitrospinae/Tectomicrobia group , Spirochaete , a Planctomycete from the OM190 family, an archaeon from the TACK radiation, a Firmicute that was dominant in the DOM well, a Bacteroidetes , an α-Proteobacterium, and a β- Proteobacterium. Finally, we assembled numerous phage bins.

Only one is included here ,hydroponic gutter but evidence of bacteriophage was abundant.While many potentially interesting and novel genomes were isolated from this community, we focus here on the Brocadiaceae Planctomycetes, which oxidize ammonium under anaerobicconditions in a specialized organelle called the anammoxosome that protects the cells from the toxic hydrazine intermediate products of the biochemical reaction. Anammox genomes are highly prevalent in the MW5 and MW6 samples and also occur in the DOM sample . We were able to make several near-complete assemblies of these genomes , however, the genome size of 2 Mb for many of these genomes was well below the 4 Mb seen in other members of this family. The coverage of some of these small genomes is rather high , making it seem plausible that the true genome size is reduced. In an attempt to resolve this discrepancy, we rebinned the MW5 anammox genomes using less stringent criteria, and found increased but still incomplete coverage of the Brocadiaceae reference genomes . These composite genomes were multi-strain chimeras, as indicated by conserved single copy gene occurrences increasing above one . The fact that merging multiple strains of the same species did not give complete coverage of the single copy genes is in agreement with the hypothesis that the true genome size is small but further sampling would be needed to confirm the hypothesis. In any event, we have not been able to resolve the discrepancy in genome size in the present study. The phylogenetic placement is apparent by homology with the Brocadia sinica and Jettenia caeni reference genomes . The separation of genomic bins shown by pentanucleotide clustering suggests multiple Brocadia-like genomes coexist in MW5, MW6 and DOM. A 16S phylogeny supports this observation . We refer to these genomic bins herein as MW5-59_1, MW5-59_2, MW6-02, MW6-03, MW6-13, DOM-02, DOM-03, and DOM-40 . We also assembled an 8.3 Mb Planctomycete genome with similarity to the Planctomycetaceae family within the Planctomycetes . The larger genome size indicates the genome is not of the Brocadiaceae, which have genome sizes around 4 Mb. Whole genome sequence comparison of MW6-09 to the available reference Planctomycetes showed highest similarity to Singulisphaera acidiphilus , however, the similarity even to Singulisphaera was not especially high, indicating that this genome is truly diverged from the reference genomes. Examining the 16S alignment suggests the genome could be from the OM190 group of Planctomycetaceae , a group with no sequenced genomes . We caution, however, that while we could link the EMIRGE-assembled OM190 16S gene with theMW6-09 genome using targeted assembly , multiple 16S fragments could be linked to the genome, thus our placement MW6-09 as an OM190 Planctomycetaceae should be revisited when new, related genomes are discovered.

To determine whether the anammox strains were unique to their respective bins or overlapping, we used VizBin to perform additional kmer distribution-based clustering of all the Planctomycete contigs together. Six distinct clusters are apparent , with MW5-59_1, MW5- 59_2, and MW6-13 overlapping. We next refined the genomic bins by combining the anammox genomes from MW5, MW6, and DOM and repicking chimeric bins. We then performed a single round of assembly using PRICE in order to merge the contigs. Overall, little improvement in bins was made. However, inter-strain contamination was reduced, and the DOM-02 bin was substantially improved by adding 20% more contigs from MW6 that coclustered . The 16S phylogengy indicates the bins come from three distinct lineages . The abundant MW5 and MW6 bins come from a new lineage that is intermediate between Jettenia and Kuenenia. The abundant DOM bins and one of the MW6 bins come from two different lineages within the Brocadiaceae W4 group. As there are no sequenced members of these lineages to use as reference, we aligned our bins to the closest available reference draft genomes of Brocadia, Jettenia, Kuenenia and Scalindua species . Reflecting the 16S phylogeny, the best homology was to Jettenia for the abundant MW5 and MW6 bins, and lower homology was seen for the DOM bins and MW6-02. As previous reports have noted low diversity of anammox genomes within a given sample , we find it noteworthy that as many as three distinct anammox genomes coexist within a single groundwater well. To confirm that all of these genomes were true anammox metabolizers, we checked for hydrazine conversion genes by BLASTX. Confirming that they are indeed anammox organisms, all Brocadiaceae genomes showed good coverage of the hydrazine database, and MW6-09, which is phylogenetically places as a nonanammox Planctomycete, did not have BLASTX hits.We analyzed the biochemical potential of the genomic bins in two ways focusing on pathways and modules rather than on individual proteins . This analysis was based on taxonomic placement rather than on the well of origin. First, we mapped the contigs from each draft assembly to the database of KEGG orthologs and used KEGG Mapper to visualize the results. Second, we used antiSMASH to detect potential secondary metabolite bio-synthetic gene clusters. The results reveal variation between genomic bins as well as pathways for potential community interactions linking nitrogen and sulfur metabolic pathways in the groundwater.To determine what functional genes are present in the water microbiota, we first aligned all of our contigs in each of the dairy water samples to the KEGG prokaryote database and evaluated trends at the whole metagenome level. Overall, we see enrichment of phosphotransferase systems, two-component systems, ABC transporters, and terpenoid production. The PTS systems are particularly high in the nutrient poor DOM sample,hydroponic nft channel consistent with the idea that there is a selective pressure driving acquisition of nutrients in nutrient-poor environments. However, no clear signature of different modes of nitrogen metabolism is indicated when examining the aggregated data for each sample. Thus, we examined the individual genomic bins to get a broad understanding of their biochemical potential. We focused on the KEGG pathways for nitrogen metabolism, sulfur metabolism, flagellar assembly, chemotaxis, ABC transporters, two-component systems, terpenoid synthesis, ATPase family, secretion systems, cofactor F420 , and B12 production since these pathways showed the most variability across the genomic bins. We include nucleotide synthesis as a positive control, since all of the complete bins have good coverage of the nucleotide metabolism pathways.

Because many of the genomic bins were partially incomplete, we aggregated KEGG maps from related species in order to get a more coherent picture of the pathway representation as a function of phylogeny . Overall, we see sparse coverage of nitrogen metabolism by the CPR, and DPANN genomes, while Methylomirabilis, Omnitropica, Nitrospira, Brocadia, and Nitrospinae had high coverage. The Bacteroidales also had sparse coverage of nitrogen metabolism, and the Chlorobi had intermediate coverage of the pathway, indicating that not all genomes in the community are directly involved in nitrogen metabolism. The same pattern was true of sulfur metabolism, with the exception that OP11 has the module for assimilatory sulfate reduction, which is consistent with the work of Canfield showing that sulfur and nitrogen redox pathways are coupled in the oxygen minimum zone of the oceans. Methane metabolism, indicated by presence of the coenzyme F420, was present in one DPANN bin, one OD1 bin, Methylomirabilis, and Nitrospinae. Intermediate coverage of this module was seen for Chlorobi, supporting the observations of Speth et al that species have diverse and overlapping niches within the anammox community and Shen et al that methane oxidation co-occurs with anammox. For oxidative phosphorylation, distinct ATPases were seen between the phyla. OP11, OD1, Methylomirabilis, Omnitrophica, Chlorobi, and Nitrospinae have the F-type ATPase, while DPANN, has the A-type ATPase, and Nitrospira, Brocadiaceae, and Bacteroidales have both F-type and A-type ATPases.In terms of acquiring nutrients from the environment, the CPR genomes were deficient in ABC transporters besides phosphate. The DPANN have slightly more, but the rest of the genomes each have significant coverage of  15 ABC transporters each. Coverage of two-component systems was consistent across all genomes for phosphate, while either nitrogen or nitrate was present for all except OP11. Twitching motility was indicated for OP11 as well as for OD1, and Chlorobi. High coverage of the chemotaxis pathway was seen only in the Nitrospira, Brocadiaceae, and Nitrospinae, with moderate coverage seen in the DPANN, OP11, OD1, Chlorobi, and Bacteroidales. Methylomirabilis and Omnitrophica appear to lack pathways for both chemotaxis and flagellar assembly, whereas Nitrospira, Brocadiaceae, and Nitrospinae have the complete pathways, and Chlorobi and Bacteroidales have most of the chemotaxis pathway but Chlorobi has the complete flagellar pathway and Bacteroidales lack it entirely. In terms of biosynthetic capabilities, Omnitrophica, Nitrospira, the Brocadiaceae, and Nitrospinae have complete vitamin B12 pathways, while Chlorobi and Bacteroidales have the latter half of the pathway as does one OP11 genome, while the other OP11 genomes as well as the OD1 genomes lack B12 production entirely. Methylomirabilis has sparse coverage of the pathway, and the DPANN genomes have genes for converting B12 to the active form. These results indicate that B12 sharing is likely an active part of the anammox community metabolism.Methylomirabilis, Omnitrophica, Nitrospira, Brocadiaceae, Bacteroidales, and Nitrospinae all have only the non-mevalonate pathway, whereas Chlorobi has both pathways, and DPANN use either one pathway or the other but not both. Within the CPR, OP11 has the mevalonate pathway, and OD1 has neither pathway. Wide variation was seen in the secretion systems, with the DPANN, OP11, and OD1 using only the SecSRP system, Bacteroidales using the SecSRP and Tat systems, Methylomirabilis and Omnitrophica using the SecSRP, Tat, and Type II systems, and Nitrospira using the SecSRP, Tat, Type I and II systems. The Brocadiaceae and Chlorobi use the SecSRP, Tat, Type II, IV and VI systems. And finally, Nitrospinae uses the the SecSRP, Tat, Type I, II, IV and VI systems. Comparison of KEGG pathway coverages with the available reference genomes showed similar results, supporting the hypothesis of niche specialization in a shared community metabolism.

Posted on

The tax was finally suspended indefinitely, pending a new proposal from the government

As with past reforms, the Commission’s proposal to place a cap on direct income payments was completely rejected due to widespread opposition from the member states. The final result of the Commission’s proposal income payment cap was a form of degressivity, under which payments over a certain threshold may be reduced. Degressivity was left to the member states as an option, with a very small level made mandatory . Moreover, member states could choose to garnish their total national envelope, as opposed to applying degressivity directly to their largest earnings, allowing them to reduce or entirely eliminate the loss to large farmers. Finally, greening measures were watered down, the end result being that most farmers were exempted from compliance, and overall standards were lowered. Essentially, anything that was contentious or in some way resisted by a member state was handled one of three main ways: the standards for the policy were lowered; the qualifications for an exemption were widened; or discretion for what standards to set and how to enforce the policy was given to the individual member state. In the end, nearly 50% of arable land covered by the CAP and just under 90% of arable goods farmers were exempted from the new greening rules and standards. The remaining more minor elements of the initial reform proposal were also adjusted in the final agreement. Those member states whose direct income payments were 90% or more below the EU average were afforded more flexibility in transferring funds between Pillar 2 and Pillar 1 . The proposal to more rigorously define who counts as a farmer and thus what land can be considered agricultural was significantly weakened. The Commission’s effort to restrict a base payment to individuals engaged in farming as their primary source of income was also completely defeated,hydroponic grow systems meaning anyone with agricultural land could receive this payment, even if they were a non-farmer. A.2.3 which summarizes the changes for these small proposals can be found in the Appendix.

The MFF agreement, which fixed the budget and also included some CAP provisions shaped the course of the CAP negotiations that operated concurrently and ultimately subsequent to the battle over the budget. This effect was perhaps most profound in the area of greening. Greening measures to support the environment and battle climate change were, at least at one point, considered an essential part of justifying the increasingly criticized direct payment scheme. In the initial stages of the CAP reform, when the process began in 2010 before the MFF talks were underway, it was expected that farmers would be required to provide a direct, clear, “public good” in the form of following particular environmental practices in exchange for the receipt of direct payments. However, once the MFF was agreed, the greening push was fatally weakened. With the budget fixed, the threat of imposing cuts if greening standards were not raised was no longer credible. The greening case was hurt more broadly by the fact that there was general uncertainty of the extent to which these measures would deliver real, significant environmental benefits. In addition, many worried about the added financial costs for farmers and more broadly the bureaucratic cost of administering these programs. In the end, the final agreement fell well short of initial proposals. A perhaps overly pessimistic view was expressed by multiple DGVI officials who lamented that rather than making the CAP fairer, greener, and simpler, reform left the CAP still unfair, barely greener, and far more complex . This characterization understates the reform. In fact, there were meaningful changes to reduce systemic inequalities. That said, the reform did make the CAP far more complex and also did little to improve greening. The previous four chapters have explored the major rounds of CAP reform, illustrating my claims about the conditions that affect policy reform, the nature of farmer power, and the links between welfare state reform tactics and agricultural policy reform, including the key strategy of pairing compensation with reform. These chapters have demonstrated that reforming agricultural policy is an exercise in navigating farmer power.

Theories of welfare state retrenchment help to explain how and why the process of agricultural policy unfolds in the manner that it does. Most often, reformers rely on the kinds of strategies described by Paul Pierson in his analysis of welfare retrenchment: compensating farmers, making smaller adjustments to correct existing policies , and introducing changes that may open the door to more far-reaching shifts down the road . When CAP reform is initiated at a time of disruptive politics, like trade negotiations or enlargement, it is much more likely to succeed. The 1992 MacSharry and 2003 Fischler reforms occurred at times of disruptive politics and the end result was major reform to the operation of the CAP payment system. Of course, as my argument requires, these reforms 1) were paired with generous compensation and 2) largely consisted of recalibration, in which the modes of payment were changed, but the amounts remained the same. Under politics as usual, reform proposals are almost always defeated. The 1999 and 2013 CAP reforms occurred in “normal” times- no trade negotiations were occurring and enlargement was not looming. In both cases, despite proposals for extensive change, the CAP remained virtually unchanged. Any minor reforms that did occur were either accompanied by extensive compensation or qualified by massive exemptions. The previous four chapters have demonstrated how theories of welfare state retrenchment, combined with an awareness of the broader context of the reforms, can help explain the process and outcome of the CAP reform. The question remains, however, if the analytical efficacy of welfare state retrenchment theories stems from circumstances particular to the CAP and the EU or if this approach can be applied more broadly. In order to assess the analytic usefulness of my approach, I test it under a variety of different conditions and circumstances in three mini-cases: austerity-driven domestic policy in Europe, domestic agricultural policy reform in Japan, and agricultural trade policy negotiations in the GATT Uruguay Round.

My claims about the politics of agricultural policy reform and farmers’ influence over policy making are not restricted to the CAP and the specific circumstances of EU politics. They can also explain agricultural reform at the domestic level. To make this case, I explore domestic politics and policy making in the wake of the 2008 financial crises. The mini case proceeds in two parts. I begin with a close look at France, focusing on domestic policy reforms, which are not specifically agricultural in nature. An “eco tax” 41, proposed in France in 2013, that was to affect truck transport of goods, including agricultural products, was abandoned due to pressure in large part from farmers. Pension-related cuts, however, went ahead despite mass protests. The second part of the mini case looks at domestic reform in Europe more broadly, and confirms that the French case is not an outlier. Indeed, I find that at the domestic level, European governments largely did not cut national discretionary spending on agriculture, while they did impose significant cuts on pensions. In the wake of the Great Recession and sovereign debt crisis, austerity programs were adopted across the European Union. Spending was cut, programs of support for those suffering financial hardship were canceled or suspended,hydroponic growing and new taxes were created to help generate revenue for cash-strapped governments. One social group that would have appeared to be particularly vulnerable to austerity-imposed policies is the agricultural community. Farmers are a tiny population that is continuing to decline. Any spending cuts or new tax burdens would therefore apply to only a small portion of the population, in theory allowing politicians and policymakers to minimize the negative backlash faced by aggrieved constituents. Moreover, farmers already receive a disproportionate share of financial support, given their share of the population, making their programs and policies low-hanging fruit for retrenchment-minded officials. Yet, since the 2008 financial crisis, farmers have felt little if any of the budgetary pressure that austerity has brought to bear on communities across Europe. As is observed in EU policy making, farmers at the domestic level successfully resist the imposition of new costs that are not paired with compensation. The parallels between the ways that farmers defend their policies and thwart unwanted policy changes at the domestic and EU levels can be made clear by looking at a case in which a national government attempted to impose new costs on their agricultural community without offering compensation. In 2013, Socialist French President François Hollande attempted to implement the so called “eco tax” first put forward by his conservative predecessor, Nicolas Sarkozy. The eco tax was intended to promote greener commercial transportation by imposing a tax on heavy vehicles. Under the plan, any vehicle over 3.5 tons would be taxed a flat rate of .13€ per kilometer traveled on 15,000 kilometers of roads included in the scheme. The government expected the tax to generate over €1 billion in revenue annually. The eco tax was slated to come into effect beginning 1 January 2014. The government’s proposal was immediately met with criticism from the main French farmers’ organization, the FNSEA. The organization described the tax as an “usine à gaz”, a situation where pipes are going everywhere and the system is overly complex. Through thus turn-of-phrase, the FNSEA meant to convey that the eco tax was a complicated procedure with little actual value or payoff. The FNSEA argued that the tax would place a significant burden on the agricultural community, particularly farmers in Brittany, who had suffered significantly from the financial crisis, and demanded that it be suspended immediately.

Other critics raised concerns that Breton farmers might be driven out of business as a result of higher transportation costs. In addition to the concerns about its effects on Breton farmers, the FNSEA warned that French goods would pass through the tax gates more often than trucks carrying foreign goods, putting French farmers at a disadvantage compared to farmers’ goods arriving from abroad. Xavier Beulin, the leader of the FNSEA, promised immediate action against the proposal, directing members to target the “portiques” that were intended to scan the trucks as they passed underneath. Beulin called on farmers from other parts of France, even from those areas without the tax scanners, to join the protests. The call for action was successful, as a wave of angry protests erupted in Brittany and across France. In Brittany, the heart of the demonstrations, protesters gathered in main town squares, many wearing red caps, or bonnets rouges in a reference to a 17th-century protest against a stamp tax proposed by Louis XIV. Some protestors threw stones, iron bars, and potted chrysanthemums at riot police, while others destroyed the electronic scanners intended to collect the fee from passing trucks. The protesters included not just farmers, but also the broader public, who were rallying to oppose taxes, with some also supporting the farmers specifically. In addition to the violent actions in Brittany, farmers elsewhere blocked roads with their tractors, including around Paris. Despite the disruptions these protests caused to the daily life of the average French citizen, the farmers did not face any negative public backlash, a further indication of the deep support and connections between farmers and urban France. Indeed, public polling concerning the image of farmers revealed that the public has a strong, positive image of farmers. According to a 2014 survey, shortly after the mass protests by farmers, just 26% of respondents were willing to describe farmers as selfish and only 16% of respondents agreed that farmers were violent. A resounding 80% agreed with the statement that farmers were trustworthy42 . After Prime Minister Jean-Marc Ayrault met with local officials from Brittany, the government proposed to “suspend” the tax until January. This concession, though it was expected to cost the government €800 million in revenue, was seen as insufficient, and tens of thousands of protesters continued to gather in the epicenter of resistance to the proposal, the town square of Quimper in Brittany.France’s eco tax then, like to efforts to change CAP income support systems or greening policies, demonstrates that it is nearly impossible to impose new costs on farmers, without some degree of compensation or widespread exemptions. For example, new CAP greening standards that are costly for farmers to adhere to are typically coupled with subsidies for compliance.

Posted on

Fischler truly viewed his task in these reforms as saving the CAP from collapse

Germany also took action, replacing its SPD farm minister, Karl-Heinz Funke, with Green Party member Renate Künast, a Fischler ally and vocal advocate of CAP reform. Künast advocated strict standards for animal welfare, tough environmental regulation, and greater oversight of and limitations on industrial farming. Prior to the MTR, commitments to meaningful environmental measures were tepid at best. Environmental policies were optional, and implementation was left to the discretion of member states, who mostly ignored them due to farmer resistance. The series of food scares increased pressure on the CAP from consumers, environmentalists, and animal welfare advocates. Public opinion of the CAP in these matters was quite negative. While 90% of respondents in a 2001 Eurobarometer poll expressed a belief that the CAP should “ensure that agricultural products were healthy and safe”, only 36% thought that “food bought could be safely eaten” and just 34% felt that “food bought is of good quality” . As a result, Fischler saw an opening to push for meaningful, mandatory reforms that would increase food safety and security, including the adoption of environmental and animal welfare regulations. There was also a growing recognition among the public that farmers were significant polluters . According to Eurobarometer surveys in 2001 and 2002, just under 90% of respondents stated that the CAP should be used to “promote respect for the environment” while only 41% of respondents across the EU 15 felt that the current version of the CAP actually “promoted respect for the environment” . The environmental goal was second only to the objective of “ensuring the agricultural products are healthy” . The CAP bore the brunt of the blame for agricultural pollution,hydroponic container system given that it allowed for the industrialization of agriculture and by extension tacitly promoted the use of environmentally damaging farming practices, designed to extract the highest possible yields. A final problem confronting the CAP concerned the distribution of benefits. The CAP directed most of its support to only a small number of farmers. For example, in France, 40% of all aid went to fewer than 10% of French farmers, overwhelmingly the large cereal cultivators .

This problem, known variously as the 80/20 problem or the “Queen of England Problem” had plagued the CAP for a number of years. Reformers were under pressure to, if not correct this imbalance in support distribution entirely, at least attenuate it. The unequal allocation of CAP benefits, and reports in the press about disparities in payments received by large and small farmers, led Fischler and his associates to be concerned that public opinion would turn against the CAP. Declining public support was a real worry because EU officials, particularly other Commissioners, already questioned why such a large sum of money was being spent on an increasingly small faction of the population. The challenges and opportunities posed by enlargement, WTO negotiations, and poorly operating CAP programs offered Fischler and his associates an opportunity to propose far reaching reforms of the CAP under the auspices of the MTR. Such reforms were seen by Fischler and his team as necessary to “reduce the ammunition of those demanding large budget cuts and [to] create a new support base for the CAP” . If the CAP remained unreformed, unsustainable spending and environmental destruction would make it an easy target for other commissioners and member states who preferred a much slimmer CAP budget. Their case would be helped by increasingly negative attitudes from the general public. As Pirzio-Biroli, Fischler’s deputy, noted, “[we] concluded that, if we wanted to preserve the CAP, we needed to change it” . Fischler’s agenda was not just about cutting spending . Rather, he sought to reform the CAP to save the CAP- most notably by making it financially sustainable, viable under enlargement, compatible with WTO rules, and responsive to public concerns about food safety and the environment. Some critics saw Fischler as a bean counter, looking to make cuts wherever he could to get CAP spending under control. While it is true that Fischler was attempting to radically restructure CAP spending, these critics misread the motives behind his actions. Fischler was himself a former agricultural minister. He was seeking to reform the CAP and to make cuts not to please his colleagues in Brussels, but rather to make sure the CAP continued to be viable.

Fischler benefited from a high degree of personal credibility within the Commission, having been a minister of agriculture and the leader of Austria’s accession negotiations. He was considered to be both an expert on agricultural policy and a reform-minded official, who was willing to pursue the tough changes necessary for preserving the overall health of the CAP, no matter the criticism he might face from farmers or their member state representatives. As evidence of the high regard in which Fischler was held, Commission President Romano Prodi completely devolved agenda-setting competency to Fischler and reportedly “had no firm ideas one way or the other about agriculture and issued no directive to Fischler about how or whether to reform” . Although the MTR was intended only to be a health check and not a revision of existing CAP policies, Fischler believed that the CAP needed much more than a status report on the functioning of existing programs. A program that could neither work within the EU budget nor meet basic environmental and health standards ran the risk of being cut drastically by European technocrats, if not entirely eliminated. In addition, without reform, the CAP would stand to be an anchor inhibiting the EU’s ability to negotiate in the Doha Round of WTO negotiations. Fischler thus constructed his reform objectives around making those changes necessary to ensure the CAP’s long-term survival. If he did not reform the income payment system, new member states would explode the budget beyond sustainability. If the CAP remained an obstacle for European services and manufacturing in trade negotiations, these external actors would force deep and unpleasant changes to CAP programs such that European agriculture no longer tied their hands. In other words, failure to reform the CAP could spell the end of the CAP. Fischler’s central goals were to decouple CAP payments from production and to expand the environmental scope and standards of CAP policy . The first goal, decoupling, would help reduce the vast disparity in CAP income payments, whereby 80% of CAP income payments went to only 20% of eligible farmers. It would also put CAP spending into conformity with WTO rules. The second goal, improving environmental standards, would address growing public dissatisfaction with the effects of CAP policy on food safety and the environment. Decoupling would go a long way towards ensuring that the CAP remained financially viable in an enlarged Europe,planter pots drainage while mandatory environmental standards and regulations would make certain that newly-added agricultural land was protected as well as guarantee that food produced in Eastern Europe would meet the higher quality and safety standards already in place in the West. While there had recently been high-profile incidents of food-borne illnesses, overall safety and quality standards and rules in the West were much higher than those in the East. Without these changes, the CAP would be overwhelmed by the financial strain of supporting the new member states, would be challenged by Europe’s trading partners, and would lose public support due to its social and environmental consequences. Fischler had learned several lessons from the Agenda 2000 negotiations. During Agenda 2000, the early publication of reform proposals allowed vested interest groups the opportunity to mobilize and undermine initiatives before they could even be launched .

Farmers in particular, once tipped off about the contents, had moved to lobby their governments to oppose reforms before any discussion or formal presentation and explanation of the proposals could occur. Fischler, therefore, resolved to develop the MTR behind closed doors, like MacSharry had done with his 1992 reform. By working first in secret, Fischler would be able to propose a reform that called for dramatic and far-reaching change—more than was expected under the MTR. A more open development process would leave the agricultural directorate susceptible to interference from the member states. Given that a number of member states wanted the MTR to be nothing more than a review, Fischler opted to keep his proposals under wraps until the time was right. Secrecy also gave Fischler and the Commission a research advantage: they could collect and gather evidence so that when the reform proposal was presented, Fischler would be able to provide data to support his proposals . The member states would then be forced to play from behind in order to mount specific, evidence based criticisms of the Commission’s reform package. Because the content of proposals would be a surprise to the member states, they would be unable to make specific, detailed claims about the effects of reform, and any general reactions, like claiming the reforms would hurt farmer incomes, could quickly be refuted by the Commission, armed with data and evidence. Another key and more personal lesson that Fischler had learned was to be wary of French President Jacques Chirac. When speaking about his adversaries in CAP negotiations, Fischler stated, “my biggest opponent was Mr. Chirac” . The two had tangled multiple times in the past. Fischler described Chirac as an “intelligent and crafty politician who knew a great deal about agriculture and could manipulate political rules to his advantage” . Fischler recounted that at the Berlin Summit, where Agenda 2000 would be formally adopted by the European Council , Chirac “used a trick” to reopen and revise the agreement . First, Chirac exploited his personal relationship with German Chancellor Gerhard Schröder who held the rotating presidency at the time. Because the CAP was part of the new Multi-annual Financial Framework , the summit chair could open any component of the MFF for debate and reform. Second, Chirac used a complicated rule in the operation and calculation of the CAP budget to compel other member states to abandon a dairy reform that Chirac did not like29. Fischler “was furious because the reform was already agreed to, but Chirac went back and undid the work” . Fischler learned the hard lesson that even when the agreement was concluded in the Council of Ministers, he still needed to make sure the holder of the rotating presidency did not reopen the CAP portion of the MFF at the European Council summit meeting. Fischler’s rocky relationship with Chirac continued after the Berlin Summit. Following the Agenda 2000 reform, Chirac unsuccessfully appealed to Commission President Prodi to not renew Fischler’s post as agriculture commissioner for a second term. When asked specifically about the MTR, Fischler again identified Chirac as one of his biggest adversaries . During the MTR negotiations, Fischler, however, benefitted from the lessons he had learned from the Agenda 2000 reform process and was better positioned to manage and respond to Chirac.Fischler’s initial plan was sent to the Commission on 10 July 2002 after months of study and work conducted largely in secret by Fischler and a small group of associates, including his Deputy Director, Corrado Pirzio-Biroli. The small group included only the top officials at DGVI. These officials relied on studies and analyses by experts within the DGVI administration, who conducted preparatory analysis and calculated potential effects of the reforms but were not fully informed of overarching agenda. Otherwise, Fischler and his associates preferred to keep the civil servants in the dark. Because CAP reform would force DGVI civil servants to change how they worked and adopt new, often complex and cumbersome systems, Fischler thought it unwise to reveal the extent of the plan to them. Fischler was also concerned that the civil servants might leak aspects of the program to their permanent representations, allowing the member states to begin to mount a defense before he could announce his reform package . Retrenchment-minded welfare state reformers are also known to work in secret. Keeping potential reform proposals out of the public debate and masking the costs of new policies and reforms are strategies commonly used by welfare state reformers to avoid resistance from those who stand to lose.