Depending on the outcome measure, detectable effects may take weeks or months for the intervention. Only a limited number of studies exist assessing the impact of nut or berry intake on the incidence or severity of diseases or metabolic dysfunction, which require durations of months or years. Moving Forward: Precision Nutrition, Multi-omics, and Biomonitoring Precision nutrition evaluates an individual’s unique biological characteristics such as genotype and phenotype, including DNA expression, influences of the gut microbiome, and metabolic response to specific foods or dietary patterns, as well as dietary habits and external factors influencing outcomes such as social determinants of health, to determine the most effective dietary strategies to improve health and prevent disease. Understanding the sources of interindividual variability that contribute to metabolic heterogeneity and applying mathematical modeling and computational algorithms will be essential to refining dietary recommendations. Several recent publications comprehensively review research gaps and study design considerations in the field of precision nutrition and specifically concerning phenolic-rich plant foods. Precision nutrition will lead to important discoveries pertaining to interindividual responsiveness to the intake of nuts and berries. Ultimately, this information can be applied via targeted recommendations to individuals and groups for achievable and sustainable dietary intake of nuts and berries to promote optimal health. The incorporation of bio-monitoring technologies into study designs may also be used for precision nutrition. Current and emerging mobile devices can provide continuous data collection in free-living populations with minimal participant burden. The study of nuts and berries would be enhanced with the use of devices that can capture real-time physiological outputs at home that reflect normal living conditions.

Further collaborative efforts in the fields of bioengineering and artificial intelligence hold promise for advancing the understanding of benefits from nuts or berries. An emerging personal bio-monitoring technology is the Precision Health Toilet,square pot which collects and evaluates human urine and stool, which are then analyzed using artificial intelligence to determine flow rate and volume of urine, as well as fecal analysis via the Bristol Stool Scale. A second type of toilet seat, the Heart Seat, has recently been approved by the US Food and Drug Administration for home use to monitor heart rate and oxygen saturation, with future plans to add sensors that monitor systolic and diastolic blood pressure. Assessment of metabolites in the excreta seems like a feasible goal for future development, which may be useful, for example in the detection of urinary and fecal metabolites that can reflect the metabolism of ellagic acid to urolithins and of -epicatechin to γ-valerolactone. A third example is an ingestible capsule containing a biological photosensor that can detect gut inflammation. Bioluminescence can be monitored from bacteria that have been engineered to illuminate when they come into contact with a molecule for which they have been coded, such as urolithins from berries or lipid-sensitive metabolites from nuts. Finally, another type of ingestible capsule has recently been detailed that collects samples from multiple regions of the human intestinal tract during normal digestion. This device has been used to explore the role of the gut microbiome in physiology and disease, with novel findings that intestinal and stool metabolomes differ dramatically. The ability of nut or berry intake to alter such metabolomes, and their association with changes in physiological function and health outcomes, would be an interesting area for future research. Although these technologies are still in their infancy, they have promise to further precision nutrition research efforts on nuts and berries. Research addressing the issue of “responders” compared with “nonresponders” is important in understanding the metabolic discrepancies in many studies on nuts and berries. For example, platelet aggregation phenotypes can vary significantly by individual responsiveness to oxylipins, bio-active lipid mediators derived from polyunsaturated fatty acids present in nuts as well as in extra virgin olive oil.

Variations in circulating metabolites and microvascular function following the intake of freeze-dried strawberry powder have been reported. Those individuals producing increased nitrate and nitrite levels showed favorable changes in function whereas those showing no change in nitrate or nitrite levels did not. Another example is illustrated by a recent letter in response to a systematic review of almond intake and inflammatory biomarkers. The letter notes that while the review included amounts of almonds ranging from 10 to 113 g/d, favorable responses only occurred at intake of <60 g/d. Further, the authors note that although the review reports beneficial effects of almond intake on reduction in C-reactive protein and interleukin-6, subgroup analyses showed that the effects on these 2 outcomes were not significant among those with obesity or who were rated as unhealthy prior to the intervention. Characterizing participants according to precision nutrition, including the use of genetic phenotyping to identify target genes that may result in “responders” and “nonresponders” prior to enrollment may be helpful for clinical trials but does not reflect responses in a free-living population. Furthermore, in addition to physiological variations, sociobehavioral differences among individuals that may modulate responses to berries and nuts must also considered. Nonetheless, innovative precision nutrition models that can identify inter individual differences would be useful in defining mechanisms of action and potentially who would benefit the most from regular nut or berry consumption. Plasma and serum concentrations are useful to identify the bio-availability and bioefficacy of key nutrients and phytochemicals found in nuts and berries. Some compounds, such as small molecular weight polyphenols, are first absorbed in their native state in the small intestine. Other polyphenols can be biotransformed via the host microbiota to a second set of compounds that are subsequently absorbed and confer additional bio-activity beyond that obtained from the parent molecules. Monitoring both host and microbial metabolites in the blood and urine, and those that may accumulate in tissues of interest such as the liver and gastrointestinal epithelium, among other tissues, would be useful in understanding the dynamics of nut and berry bio-activity and specific association with site of actions.

Broader application of orthogonal approaches that combine untargeted with targeted metabolomic platforms and combined with the use of advanced informatics will support new understanding about the absorption, distribution, metabolism, and excretion of compounds found in nuts and berries. For example, the UC Davis West Coast Metabolomics Center conducts both targeted and untargeted assays that assess plasma microbial metabolites using a bio-genic amine panel that identifies and quantifies acylcarnitines, trimethylamine N-oxide, cholines, betaines, nucleotides and nucleosides, methylated and acetylated amines, di- and oligo-peptides, and a number of microbially modified food-derived metabolites. Some inter individual differences in response to nut or berry intake have been attributed to the composition of the gut microbiome. For example, ellagitannins are polyphenolic compounds present in strawberries, raspberries, and walnuts that are metabolized by gut bacteria into an array of urolithins . The production of urolithins relies on the capacity of specific microbes, Gordonibacter pamelaeae and Gordonibacterurolithinfaciens. Urolithins may decrease symptoms of chronic metabolic diseases, including inflammation and dyslipidemia. Following a single intake of red raspberries, individuals with prediabetes and insulin resistance had lower concentrations of circulating urolithins compared to levels found in those who were metabolically healthy, a result related to gut microbiome composition. In the same population, consuming red raspberries for 4 wk improved hepatic insulin resistance and total and LDL cholesterol in the prediabetes group, and the effects were related to decreased R. gnavus and increased E. eligins. Overall, including a practical amount of red raspberry in the diet regularly is a low-calorie dietary strategy that improves gut microbiota composition and function in individuals with prediabetes and insulin resistance resulting in improvements in metabolic health. With a sustained emphasis on the role of gut microbiota in nutrition research, advances in our understanding of food-gut dynamics will provide new insights about the role of nuts and berries in human health and performance. Although research on a specific nut or berry provides insight into bio-activity and potential mechanisms of action,square plastic planter such focus also creates the potential for fragmentation because the search for overall dietary patterns is not addressed. The composition of fruits and nuts differ at the molecular level, and a broader view assessing similarities in chemistry and health benefits is critical for translational research as well as for messaging purposes. For example, blueberries, strawberries, pomegranate, walnuts, and grapes all have reported benefits for cardiovascular health, driven largely by the presence of similar polyphenols, which are present at varying quantities in each of these foods. Although health professionals and consumers often hear messaging on a single berry or nut, the potential benefits of increasing consumption of the broader category may be obscured or lost. This challenges the ability to maintain consistent messaging and align better with translatable dietary guidance. Future interventions that combine nuts and berries with one or more other foods within a food matrix at dietary achievable doses and in more diverse populations are warranted. To date, multi-omics technologies have provided valuable insights into exposure-disease relationships.

Coupled with artificial intelligence, predictive modeling and continuous, personalized monitoring, these data-intensive outcomes can provide further insights about the health benefits associated with regular intake of nuts or berries. Use of highly personalized data collection devices will require secure data repositories. One of the challenges of similar foods being studied in differing formats and by various research groups is the utility of the data as a combined set. Differences in test materials and experimental designs make integration of data difficult. The proper curation of combined data, whether physiologic, metabolomic, or genomic, is critical to ensure that combined datasets provide synergy, statistical power, and enhanced usefulness.The cardiometabolic benefits from regular consumption of nuts or berries are widely reported and include improved vascular function, reduction of cardiovascular disease risk factors, improved insulin sensitivity, and reduced risk of type 2 diabetes mellitus. Antioxidant and anti-inflammatory capacity and activity have also been noted. Metabolic outcomes may be context-specific and related to the physiologic state of the individual and host microbiome composition, among other factors. Examples include findings of ellagitannin and ellagic acid rich foods resulting in differential responses in healthy individuals compared to those with prediabetes, who are dependent on gut microbial-derived metabolite profiles. Many factors contribute to inter individual variability in response to diet that can extend to context-specific aspects influencing the magnitude of health benefits and reinforces the importance for further research aimed at advancing discoveries in precision nutrition. Additional health outcomes related to nut or berry intake are outlined below.Adding nuts or berries to the daily diet may be advantageous for weight management for several physiological reasons. One is that these foods produce feelings of satiety, helping to reduce the desire to consume calorie-rich snacks that are low in vitamins, minerals, and fibers, ultimately improving body composition over time. A second possibility is due to urolithins, secondary metabolites produced from ellagitannins in nuts and berries. Urolithins increase the activation of the adenosine monophosphate-activated protein kinase pathway, resulting in anti-obesogenic properties in vitro and in animal models. AMPK increases fatty acid oxidation and decreases triglyceride accumulation. Phosphorylation of AMPK may also decrease cholesterol synthesis and lipogenesis by down regulating 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and sterol regulatory-element binding protein expression. In clinical studies exploring the relationship between food and body composition, the incorporation of nuts and berries into the diet was associated with weight loss or maintenance.Regular consumption of nuts or berries has been reported to support brain health and cognitive function, motor control, mood, and executive function at physiologically relevant intakes. Middle-aged and older adults experienced improvements in balance, gait, and memory, and children experienced higher executive function and positive affect after acute and regular intake of both strawberries and blueberries. These beneficial effects may be the result of direct effects on brain signaling or indirect effects through oxidant defense and anti-inflammatory properties of polyphenols and other bio-active compounds in nuts and berry foods. The gut-brain axis is an emerging area of research. Most studies are preclinical in nature using animal models but are suggestive of a significant role of gut microbial-derived ellagitannin metabolites on brain health and neuroprotection.Regular intake of almonds, a good source of fatty acids and polyphenols, has been associated with a significant decrease in facial hyperpigmentation and wrinkle severity.

Numerous studies on blueberry treatment of murine cell lines use TLR4-activated models, through the addition of the bacterial cell wall component LPS at doses varying from 0.01 to 10 μg/mL . This produces a strong inflammatory response through, but not limited to, the expression of IL-6, TNF-α, or NO . The capacity of blueberry components to reduce proinflammatory marker gene expression and secretion inLPS-induced murine cell models has been shown in several studies, although the anti-inflammatory effect seems to be carried out by an array of compounds rather than an individual blueberry phenolic fraction. Diverse blueberry products, including blueberry pomace, whole blueberry extracts, polyphenol-rich extracts, or specific phenolic fractions lowered the production of cytokines by the cells in a dose-dependent manner. Further details on the treatments and doses used in the studies are reported in Table 1. For instance, blueberry phytochemicals reduced the gene expression and secretion of proinflammatory cytokines induced by LPS, particularly IL-6 , IL-1β , and TNF-αin RAW 264.7 cells and BMDMs compared with the LPS-induced control. However, it is not clear which fraction or specific phenolic compound exerts a more potent effect, especially since their activities seem to be cytokine specific. For example, Esposito et al. reported a better ability of a proanthocyanidin fraction to reduce IL-1β compared with the polyphenol extract, anthocyanin, or phenolic acid fractions. However, IL-6 gene expression was not suppressed by that same proanthocyandin fraction, and monocyte chemoattractant protein -1 expression , a chemokine that regulates monocyte infiltration ,drainage planter pot was inhibited by the blueberry anthocyanins but not by the other phenolic fractions. Although the large variation between the blueberry treatment doses used across different reports hinders direct comparisons between the studies, most have shown a dose-dependent regulation of cytokines .

Cheng et al. , however, reported 40–60% inhibition of IL-1β gene expression with ≤200 μg/mL blueberry extract, but no effect when the treatment dose was increased to 400 μg/mL . This observation was tentatively explained by the increased phagocytic activity of the macrophages in the presence of a high polyphenol concentration. iNOS is an enzyme responsible for the synthesis of NO, a mediator secreted by neutrophils and macrophages to induce vasodilation, mediate the immune response, or regulate apoptosis . After stimulation of RAW 264.7 cells by LPS, blueberry extracts inhibited iNOS gene expression and NO production , with extracts from anthocyanin-rich cultivars and blueberry proanthocyanidin fractions being particularly effective . COX-2 also plays a central role in the induction of inflammation. It is involved in the formation of prostaglandins, including prostaglandin E2 responsible for the induction of pain . Blueberry extracts have consistently reduced COX-2 gene expression in RAW 264.7 cells , although Mueller et al. and Grace et al. did not report a significant reduction of iNOS and COX-2 gene expression by whole blueberry or polyphenol-rich extracts. Although most studies have focused on blueberry polyphenols, Gu et al. reported the anti-inflammatory effect of the volatile extracts of several berries compared with their phenolic counterpart. Volatile and phenolic blueberry extracts showedsimilar inhibitory effects on inflammatory cytokines, NO, and COX-2 production, even though the volatile fraction was tested at a lower concentration than the phenolic fraction. This provides initial evidence that the volatile fraction of blueberries also contains molecules with anti-inflammation properties, but this remains to be validated. The anti-inflammatory activities of blueberry phytochemicals have also been demonstrated in cell lines derived from humans , including the U-937 and THP-1 monocytelike cells, that can be differentiated into macrophages after stimulation with phorbol-12-myristate-13-acetate , and human primary peripheral blood mononuclear cells . In the context of blueberry studies, a variety of compounds have been used to induce inflammation, including Fusobacterium nucleatum bacteria , LPS , or cytokines .

Blueberry extracts exerted an inhibitory effect toward cytokine secretion and matrix metalloproteinase -8 and 9 production , in cells triggered by bacteria or LPS. Blueberry extracts decreased TNF-α gene expression induced by LPS in THP-1 monocytes and U937 macrophages , but on the contrary, increased its expression in THP- 1 differentiated macrophages . The regulatory effect reported in most studies was associated with a decrease in NF-κB translocation in THP-1 cells . In PBMCs and THP-1 cells alternatively induced with either IFN-γ or TNF- α, the effects of a blueberry treatment were less robust . Cytokine secretion and adhesion molecule gene expression were inhibited by a blueberry extract in IFN- γ -induced PBMCs, but the same blueberry extract further increased the proinflammatory marker secretion when the cells were induced with TNF-α . These observations were tentatively explained by looking at the pathways activated by the different cytokines : the signal transducers and activators of transcription pathway activation was inhibited by coincubation of IFN-γ with the berry treatment, but NF-κB was enhanced by the addition of TNF-α combined with a blueberry extract . A follow-up study demonstrated that the IFN-γ receptor 2, responsible for transducing the signal conveyed by the proinflammatory cytokines, was inhibited by the blueberry anthocyanins . These observations on cell models induced with non-LPS ligands suggest that the immunomodulatory effects of blueberry compounds are context and pathway specific. In summary, blueberry phenolic and polyphenolic extracts have been shown to dampen inflammation in RAW 264.7, U-937, BMDMs, and PBMCs challenged with inflammation inducers, through the reduction of proinflammatory cytokine gene expression and secretion, and inhibition of NF-κB translocation to the nucleus. No specific fraction emerges as being more potent, suggesting a general effect of multiple phytomolecules rather than a single compound. More studies are warranted to better define molecular targets of blueberry-derived molecules and to assess the involvement of TLR-dependent and -independent pathways.ROS and free radicals are natural by-products of enzymatic reactions produced during metabolism .

When controlled, ROS production is used for signaling in metabolic processes . Environmental factors, lifestyle, and pathologies contribute to an unbalanced state, where ROS production overwhelms the defense capacity of the cells and induces oxidative stress . This state leads to protein and nucleic oxidation, and lipid peroxidation, which can impair enzymatic processes, induce breakage of DNA strands, and may lead to cell death . Endogenous antioxidant defense mechanisms exist in the body to limit the production and deleterious effects of ROS. Superoxide dismutase , found in the membrane or cytosolic fractions of cells, converts superoxide radicals to H2O2 and O2 . Glutathione peroxidase , via the oxidation of glutathione S-transferase, reduces lipid peroxide and converts H2O2 to H2O . Enzymes, including DNA glycosylases, repair damaged DNA . Oxidation and inflammation are intricately related, as cytokines and chemokines secreted by inflammatory cells can trigger ROS production. In turn, ROS activate proinflammatory pathways, including NF- κB, and sustain the cycle of oxidative and inflammatory stress . These conditions favor the development of chronic pathologies such as cancer , cardiovascular , inflammatory , and neurodegenerative diseases . The effect of blueberry phytochemicals on oxidative stress has been evaluated using several cell models, including neurons , fibroblasts , hepatocytes , enterocytes , and epithelial cells . Despite the diversity in models used, studies overlap in terms of the endpoints measured, which focus on evaluation of the modulation in ROS production and lipid peroxidation, increases in antioxidant enzyme activities, and protection of DNA against oxidative damage . The induction of oxidative stress in a variety of cells was attenuated by treatment with blueberry extracts, principally through the decreased formation of ROS , but also increased scavenging activity , and/or reduction of lipid peroxidation . This effect, however,plant pot with drainage was only partly explained by the regulatory effect of the blueberry compounds on antioxidant enzymes, which were upregulated in neuronal cells treated with blueberry juice and Caco-2 cells incubated with a polyphenol-rich blueberry extract . Glutathione concentrations, however, remained unchanged in Caco-2 cells treated with an anthocyanin fraction . In addition, a blueberry pomace extract tested on Chinese hamster ovary epithelial cells and a human colon cancer cell line failed to stimulate the transcription of detoxification enzymes such as heme oxygenase and NADPH quinone oxidoreductase- 1 , both involved in the antioxidant/oxidant balance . In contrast with those observations made on blueberry parent compounds, phloroglucinol aldehyde increased thetranscription activity of nuclear factor erythroid 2-related factor 2 , which when induced by oxidative stress stimulates the transcription of HO-1 and NQO-1 .A consequence of oxidation is DNA damage and potentially increases in cell death. Blueberry extracts demonstrated a protection against DNA damage induced by hydroxide peroxide and tert-butylhydroperoxide , although in Caco-2 cells, only the blueberry phenolcarbonic acid fraction reduced DNA damage compared with whole blueberry extract, anthocyanin, and polymeric fractions . Protective effects in terms of DNA damage and cell death were attributed to blueberry anthocyanin fractions in liver cells and pulmonary epithelial cells exposed to light or ionizing radiation, through modulation of apoptosis and cell cycle regulatory gene expressions . However, no improvement of cell cycle perturbation induced by 2,2-azobisdihydrochloride was reported in intestinal epithelial cells IPEC-1 treated with blueberry anthocyanins . To summarize, blueberry extracts and phenolic fractions demonstrated protective effects against oxidative stress, which were mainly explained through reduction of ROS production and protection against DNA damage induced during oxidation. Additional studies comparing cell models and/or blueberry fractions using similar experimental parameters are necessary to demonstrate the effects of the treatment on antioxidant enzymes, and fully understand the antioxidant contribution of blueberry phytochemicals versus their metabolic by-products.

There is also a need to evaluate the potential effects of blueberry volatiles in this regard.Atherosclerosis, a chronic inflammatory disease of the arterial wall , is characterized by the buildup of plaques in arteries and is the most frequent underlying condition for the development of cardiovascular diseases . Human umbilical vein endothelial cells and human microvascular vein endothelial cells provide a model to study normal as well as oxidation and inflammation-related dysfunctions. Studies on the effect of blueberry compounds in endothelial cell models are detailed in Table 3. Risk factors including smoking, aging, hypercholesterolemia, and hyperglycemia promote the retention of lipids, particularly LDL prone to oxidation in the vascular wall, causing the activation of inflammatory processes . The treatment of endothelial cells exposed to oxidative stress triggers with blueberry anthocyanins demonstrated protective effects toward ROS secretion and lipid peroxidation . A similar observation was reported with cells treated with blueberry exosome-like nanoparticles ,an extracellular messenger vesicle presents in plants that contains proteins, lipids, mRNA, and microRNA . In addition to reducing ROS production, blueberry ELN also regulated gene expression involved in endothelial activation and leukocyte recruitment [MAPK1 and intercellular adhesion molecule ] and inflammation . The antioxidant effect of blueberry extract on endothelial cells is likely due to the activity of several phytochemicals, but the extent to which other compounds contribute to the effect remains unclear, particularly due to the low number of studies focusing on nonphenolic compounds. The secretion of chemokines and adhesion molecules by the endothelium is primarily regulated by TNF-α and Creactive protein , and leads to monocyte recruitment . After infiltration, monocytes differentiate into macrophages and phagocytose LDL . Macrophages that accumulate lipids eventually turn into foam cells, becoming surrounded by smooth-muscle cells and a collagen matrix, ultimately resulting in plaque formation . Blueberry anthocyanins reduced the adhesion of THP-1 monocytes to HUVEC endothelial cells with a better efficacy than the phenolic acid fraction derived from the same extract . The action of blueberry anthocyanins was further investigated and the individual compounds malvidin and cyanidin 3-glucoside, protocatechuic, and gallic acid reduced THP-1 adhesion . Blueberry extracts also decreased platelet- and endothelial-derived microvesicles through the inhibition of P2X7 transcription and Akt phosphorylation, both contributing to the release of extracellular vesicles associated with monocyte interaction with endothelial cells . Su et al. observed that blueberry extracts led to down regulation of noncoding miR-21, miR-146a, and miR125b, miRs typically increased in macrophages involved in plaque formation . In cardiovascular disease, endothelial cell migration and angiogenesis are reduced, leading to structural and functional alterations of the endothelium . Akt is a major signaling pathway in angiogenesis, regulating cell survival, cell cycle, and migration . Treatment with blueberry polyphenols increased angiogenesis in endothelial cells through the upregulation of the Akt pathway . However, abnormal angiogenesis promoted by vascular endothelial growth factor was counterbalanced by blueberry extract treatment through the inhibition of ERK and Akt phosphorylation . In addition, blueberry polyphenol extract did not modulate polyphospholipase C expression and phosphorylation, involved in angiogenesis, in HUVEC cells following induction with VEGF or LPS .

As the US/ Brazilian admixture weight is much less than the European admixture weight, this was likely due to a migration event from the Americas into Irish populations. The other out-of-US admixture event, from the Western United States to South Korea , was seen when m ¼ 6, 8, and 10. F3 statistics all have significantly negative values, and the F4 statistics and are significantly positive, supporting a Western US/South Korea admixture. However, using nine migration edges Treemix reported the reverse direction; as F3 and F4 statistics cannot easily infer directionality, more heavily sampling of the Asian populations or alternate methods may be needed to determine whether flow is occurring in both directions.To determine if invasive populations have experienced loss in genetic diversity, we used the software ANGSD to estimate average pairwise nucleotide diversity in 20 kb increments across the 20 largest contigs of genome for each population. Invasive populations can sometimes exhibit reduced levels of diversity early on in their history due to a founder effect , whereas ancestral populations tend to have the greatest amount of diversity as they have had many generations to accumulate mutations. A Welch one-way test found a significant difference in mean pairwise nucleotide diversity between clusters. We then used pairwise Games-Howell tests and found each cluster to be significantly different , except for the Eastern United States, Brazil, and Italy when compared with each other. As Asia is the ancestral home of D. suzukii, it is no surprise that South Korean wild populations exhibit the highest diversity levels . Similarly, the laboratory populations from Japan and Hawaii have half as much pairwise diversity as the wild South Korean population,drainage pot consistent with a small laboratory population size. The invasive populations display an intermediate level between these extremes. To assess whether invasive populations may have experienced a bottleneck or population shrinkage, we also estimated Tajima’s D in the same genomic intervals.

Extremely positive values can indicate a loss of rare alleles, which can occur during a population shrinkage, whereas extremely negative values can indicate a recent bottleneck followed by rapid expansion . A Welch one-way test again indicated significant differences in mean Tajima’s D between clusters , and pairwise Games-Howell tests found all clusters to betatistically different except for Western US against Brazilian flies. Strains from Hawaii and Japan both had high genome-wide levels of Tajima’s D, indicative of a loss of rare alleles that can occur during a population shrinkage . The remaining populations had neutral values of D, except for Ireland’s relatively high value. Based on this, we conclude there are no strong signals for a recent bottleneck, although the high genome-wide D value for Ireland suggests a recent population shrinkage. As our Irish samples were collected in 2016 only 1 year after its discovery in Ireland, we could be observing the founder’s effect in action .Based on population allele frequencies, we have shown that D. suzukii exhibit population structure based on region and invasion history. In the New World populations, we find that Eastern and Western US samples appear to be distinct populations. While this could be the result of continuous population variation from East through Central to the West coast, it is more likely the case that the two populations experience little gene flow due to strong geographical barriers such as the Sierra Nevada or Rocky Mountain ranges, and the fact that key target fruit crops such as cherries,raspberries, blueberries, and strawberries are primarily grown in states that we sampled . Any genetic exchange between these regions would likely be the result of human activity, such as could be the case with samples collected from Alma Research Farm, Bacon County, Georgia clustering with the Western US populations. As other nearby collections failed to share this signal, the Alma research population could represent a recent and isolated migration event.

Otherwise, we see little evidence of migration events or admixture between the Eastern and Western United States, which is somewhat surprising as the country’s supply of fresh blueberries, cherries, and caneberries are concentrated in a few states and shipped across the country . However, recent changes to cultural management such as more frequent harvesting and post-harvest chilling may be responsible for disrupting the D. suzukii lifecycle and limiting cross-country transport . While we were able to detect population structure between eastern and western locations in the United States, we were surprised to discover a lack of structure on a finer scale, either based on latitude or simple geographical distance, given the large number of loci analyzed. In a similar study using 3484 SNPs in 246 Hawaiian D. suzukii samples, researchers were able to identify three distinct populations roughly seperated by islands . The fact that D. suzukii has been present in Hawaii since 1980, in addition to the isolation by island, are likely the strongest factors in providing enough genetic drift to create such differentiation. As the continental US D. suzukii have only been present since 2008, it may be too early for finer structure to have developed. Alternatively, continual dispersion and transportation of D. suzukii around the United States may be hindering the development of more local structure. Several studies have reported a low probability of D. suzukii surviving when exposed to freezing temperatures, based on cold survival assays of wild-caught specimens , suggesting that flies collected in cold-winter regions such as Washington, Michigan, Maine, and New York could be annual migrants to the area from nearby warmer locations. The lack of north-south population structure supports the hypothesis that flies are regularly re-migrating into colder climates after the harsh winters have ended. Alternatively, flies could be tolerating winters by surviving inside human structures , or by having evolved resistance to freezing temperatures . Studies using D. suzukii collected from different locations have reported different levels of rapid cold-hardening response, suggesting that there could be regional selection present .

If populations in northern regions undergo strong seasonal fluctuations in allele frequencies, such as has been demonstrated in wild D. melanogaster populations collected in Pennsylvania , by only sampling sites in the summer we may be missing signals of population differentiation between the north and south. Likely, some combination of these factors is responsible for the success of D. suzukii in these regions, and further studies will be needed to identify the causes. North-south clines in specific traits such as diapause and circadian rhythms have been previously identified in drosophilids and could be at play here as well . Further analyses using methods such as those recently used to detect SNPs correlated with invasive success could be applied to this dataset to find signals of selection. Fst values between populations from the United States, Brazil, and South Korea were low and agree with previously published Fst estimates based on Pool-Seq data; Olazcuaga et al. 2020 observed that Fst between US, European, Asian, and Brazilian populations varied between 8.86 and 9.02%. However, we were surprised to see that our Italian and Irish samples had much higher values of Fst compared with the other populations, and even to each other. This discrepancy could be due to the small sample sizes we had from Europe; in this scenario, pooling larger number of samples can improve power to estimate Fst, and we instead rely on comparing the relative Fst values between populations for our analysis. High Fst values between our Japanese and Hawaiian populations were expected, however, as these have likely experienced strong drift during their time in captivity. In general, we find that our treemix and migration results largely coincide with the proposed invasion pathway inferred from microsatellites , as well as a recent preprint that re-analyzed invasion pathways with pooled sequencing data . We see that European and US/Brazilian populations form two distinct clades,large pot with drainage emphasizing these regions were invaded by two independent migrations from Asia. Hawaii is the first population to diverge in the Americas, followed by the Western United States, then the Eastern United States and Brazil. Additionally, in the Western United States, we detected a strong signal of admixture from Hawaii, which could be due to multiple or ongoing migration events. We also detected signals of admixture from the Eastern United States/Brazil to Ireland, which matches the predicted initial invasion pathway and suggests multiple migration events. Unique to our analysis, we recover support for admixture of Western US samples in Asia, suggesting that migrations could be ongoing in both directions. Invasive species transport is strongly associated with international trade of live plants and plant products , and indeed agricultural export data supports the possibility of this migration as Japan receives almost 15% of all US blueberry exports, and Oregon recently became the first state to begin shipping blueberries to South Korea in 2012 . It should be noted that while Treemix infers direction of migration, the model can occasionally infer the incorrect direction, particularly when populations are closely related without an available out group .

In conjunction with evidence of this widespread ongoing migration, we observed nucleotide diversity levels of all invasive populations to be only moderately below that of the wild South Korean population, a trend also observed in Fraimout et al. . Typically, recent invasion events are characterized by reduced diversity relative to the ancestral populations due to founder or bottleneck effects . However, successive invasion events can provide relief from any initial bottlenecks by providing increased genetic diversity. This has been observed to occur in multiple animal studies and could lead to increased ability to adapt and evolve to new climates. Correspondingly, in our analysis, we did not find populations with broadly low values of Tajima’s D, suggesting little bottleneck effect. As measures to reduce impacts of invasive species are often hindered by repeated migrations , it will be important to enforce that fruits being exported and imported internationally are free of live D. suzukii as required by the US Department of Agriculture, even though this species is already internationally established.We anticipate that the genomic data provided here will prove useful in many fields of biology beyond the scope of this study. Knowledge of genetic variation and alternate alleles present within a species can be informative for the design of probes and micro RNAs , such as for the purpose of creating gene drives to control invasive species. Gene drive mechanisms to eliminate D. suzukii have been experimentally tested on multiple lines to ensure the miRNAs are broadly effective , but a large dataset of wild population sequencing allow researchers to more confidently select target sites that are nonvariable and thus susceptible to Cas9 targeting . Drury et al., demonstrated that minor natural polymorphisms in target sites reduce gene drive effectiveness in flour beetles, and tools have been developed to help researchers design gRNAs accounting for population variation . Similarly, with the recent development of a CRISPR-Cas9 editing and RNAi knockdown protocols for D. suzukii , prior knowledge of allelic variation will allow researchers to design targeting oligonucleotides more precisely to avoid loci with variability. Most recently, our dataset has been used to study sensory receptor evolution in D. suzukii, giving insights into its evolution toward becoming a major agricultural pest . Other future uses of this trove of genomic data could involve insecticide resistance studies or the development of diagnostic assays for rapid detection in the field.Since domestication efforts began in the early 1900s , higbush blueberry has rapidly become a high-value fruit crop worldwide. Higbush blueberry, compared to hundreds of closely related blueberry species in the Ericaceae, is widely cultivated due to its adaptation to temperate climates, excellent fruit quality, yield, and composition of phytonutrients. As a result for the demand for fresh blueberries as a ”super fruit”, high bush blueberry production has increased 600% during the past three decades and steadily grown to a multi-billion dollar industry. In addition to its short domestication history, high bush blueberry is unique in being one of only three major commercially valuable fruit crops, accompanied by cranberry and the garden strawberry, with wild progenitor species native to North America. Blueberries have a single epidermal layer that expresses a rich profile of anthocyanins during ripening that, in combination with epicuticular wax, generates its characteristic ”powdery blue” color.

From the perspective of crop pollination services provision, this effect is important since the pollination service can decrease not only due to lack of neighboring natural areas but also nearby flowering crop area. The vast nutritional benefits of a diet containing a wide variety of plants have long been known. However, benefits distinct from simple nutrition, such as phytochemicals have recently become clear. Diets rich in a plethora of phytochemicals can promote a healthy and diverse gut microbiota, reduce intestinal and systemic inflammation, and decrease the risk of colorectal cancer and type 2 diabetes mellitus. Some of these benefits can be observed around the world. Many parts of India have historically low colon cancer incidence rates. The Indian subcontinent has been continuously settled for millennia. Ancient cities in the Indus valley have been dated to the third and fourth millennia BC and some sites are even older. Archaeological evidence of grain cultivation, including several varieties of barley and wheat, has been found in excavations dated to the sixth millennium BC. Wheat is still a staple crop in northern India, and many other grains, including barley, were commonly cultivated and eaten until the 1950s, when wheat and white rice became dominant. Although the country has many diverse cultures, some customs remain common and conventional throughout the nation. One such tradition is the form of main meals where a large round platter, the thali, holds rice or bread and several smaller bowls, or katori, which hold a separate condiment or curry to be eaten with the rice or bread at the diner’s preference. Typical dishes include, but are not limited to, dal , yogurt ,vertical indoor hydroponic system and assorted spices and vegetables. The development of agriculture early in its history has allowed India to develop rich traditions around food.

These traditions have been deeply influenced by Ayurveda, the ancient Indian system of medicine. In Ayurvedic practice, food is a source of nourishment and medicine, used to both prevent and treat illness. Maintaining a proper balance of Ayurvedic elements through diet is considered an effective way to live a healthy lifestyle. According to the Ayurvedic principles, each meal should contain a balance of the six major flavors. This calls for the many small portions of a thali meal which also easily incorporate variety. A variety of flavors in a meal often indicates the presence of many classes of bio-active compounds . Although these substances may not be macronutrients, vitamins, or minerals, they still impact human health. Polyphenols are perhaps the largest class of bio-active compounds, containing sub-classes such as flavonoids, isoflavones, stilbenes, lignans, and tannins. As a flavonoid subgroup, anthocyanins are included in this class. Anthocyanins are of interest in the food industry as nontoxic and water-soluble pigments, as most are colored red, purple, or blue, and many display antioxidant and anti-inflammatory activity. A class of phytochemicals called polyphenols is also found in virtually all plant foods, though their quantity may be reduced by preparation methods. Rich sources of anthocyanins include deeply colored fruits and vegetables, such as blueberries, eggplants, and certain carrot and potato cultivars. Given that many phytochemicals exert anti-inflammatory activity via promoting gut bacterial diversity, there is a growing interest in a food-based approach to countering the growing epidemic of inflammation-promoted chronic diseases such as colon cancer.We have learned that no discussion of diet is complete without consideration of the intestinal microbiota. Trillions of bacteria, distributed throughout the gastrointestinal tract from mouth to anus, facilitate digestion and intestinal homeostasis. Structural factors greatly impact the overall makeup of each community.

For example, low pH prohibits many pathogenic bacteria from colonizing the stomach and the upper small intestine. The depths of the large intestine, on the other hand, is an ideal habitat for many anaerobes. The gut microbiota is a dynamic community, composed of living organisms that can alter in response to diet, disease, and other environmental pressures. Changes in the intestinal microbiota were first correlated with illness in 1681 when Anton van Leeuwenhoek recorded that the microbial composition of his diarrhea differed from normal fecal samples. Since then, the intestinal microbiome has been closely studied to show how it can be implicated in a variety of conditions ranging from obesity to colon cancer. A great deal of investigation into microbiota has been accomplished in the last decade. Many of these observed changes result in an overall loss of bacterial diversity in the microbiota, indicating that species diversity is associated with health. However, the opposite may be true for cause-consequence relations, but not enough research has been brought to light. High-throughput technologies have driven advances in identifying the trillions of microbes and the metabolic functions that live in the colon. This led to a critical insight that gut plays as dynamic of a role in metabolism as the liver. The proximity of these microbes to the intestinal mucosa and gut lymphoid tissue explains the critical role they play in health and disease. Indeed, dysbiosis plays a significant role in the development of inflammatory bowel disease, obesity, and colon cancer. Emerging evidence suggests that diet can directly influence the content and composition of gut microbiota. Thus, understanding the complex interactions between diet, gut microbiota, and the host are crucial in prevention and treatment of chronic diseases that plague our society. Studies in murine models have shown rapid changes in the gut bacteria of mice being switched quickly from a standard diet to a high-calorie diet back to a standard diet.

In humans, surveys show that diets high in fiber correlate with higher microbial diversity and reduced populations of Enterobacteriaceae, including Escherichia and Shigella species. Marked differences are also seen during consumption of animal- vs. plant-based diets. While nutrients in the diet will affect intestinal microbes, other substances present in food may also have an effect. For example, most anthocyanins are not absorbed into the bloodstream in the small intestine, and so they stay in the gastrointestinal tract until they reach the colon. There, they can affect the colonic microbiota in multiple ways. Firstly, anthocyanins have antioxidant activity that can reduce inflammation-induced oxidative stress on the gut bacteria. Secondly, anthocyanins are a potential carbon source, which bacteria can metabolize, resulting in increased growth of certain microbes. Lastly, bacterial metabolism of anthocyanins produces a variety of metabolite byproducts, some of which have antimicrobial effects on enteric pathogen species including Escherichia coli.Chronic intestinal inflammation is a hallmark of certain bowel disorders, such as ulcerative colitis and Crohn’s disease, which are two major forms of inflammatory bowel diseases , and IBD is also considered a risk factor for colorectal cancer. In the latter, inflammation is generally low-grade but persists over a long period of time. Diet composition can promote or suppress chronic inflammation. Low-fiber high-calorie diets, which are typical in Western countries, may directly promote inflammation, or as already discussed, indirectly promote this through dysbiosis. Indeed, some dietary patterns associated with chronic inflammation are also linked to the reduction of total microbial diversity and imbalances in intestinal microbial groups. Furthermore, some bacteria, including E. coli, can flourish during low-grade inflammation, where thinning of the intestinal mucus layer occurs and allows for more direct interaction between the host’s cells and the intestinal bacteria. This condition can cause a feedback loop in which contact between bacteria and epithelial cells leads to dysregulation of mucosal immune response. This contact can lead to a bacterial biofilm, formed when bacteria attach themselves to the surfaces of the aqueous environment in the gut and begin to secrete substances that allow them to affix onto the epithelium. The interaction between bacteria and epithelial cells elevates inflammation,vertical farming tower for sale leading to increased thinning of the mucus and direct host-bacteria interaction. The thali approach, however, combats this cycle in two different ways: by suppressing bacterial growth with anti-microbial phytochemicals , and by reducing the opportunity for inflammation to occur. One molecular pathway involved in such a cycle involves interleukin 6 . This cytokine is normally expressed during acute inflammatory responses, and among other effects, upregulates the transcription factor STAT3. In the nucleus, STAT3 promotes cell proliferation and differentiation as well as upregulating anti-apoptosis genes. When IL6 is chronically elevated, it can lead to an apoptosis-resistant, constantly expanding T-cell population in the intestinal mucosa.

These cells can further contribute to chronic inflammation. Just as a certain diet may promote chronic inflammation, a change in diet can help to restore health. Various bio-active compounds, including anthocyanins, have demonstrated antioxidant activity, reducing local amounts of reactive oxygen species. Low levels of reactive oxygen species can lower the expression of some inflammatory genes, including IL6, and relieve the stresses on both the intestinal microbiota and epithelial cells caused by chronic inflammation. In a study of pigs, we found that supplementing a high-calorie diet with purple potatoes that contains anthocyanins led to a six-fold reduction in levels of interleukin-6 compared to high-fat diet control. Colorectal cancer killed nearly 774,000 people worldwide in 2015, and nearly an estimated 50,630 deaths in 2018 in America making it the third leading cause of cancer-related deaths in the United States in women and second in men. Virtually all cases of CRC are considered to result from an interplay of exogenous and endogenous factors with respect to the variable contribution from each factor. Some non-modifiable risk factors include old age and family history of CRC. Other risk factors, however, are associated with lifestyle or behaviors and thus can be changed. These modifiable risk factors include smoking, obesity, low physical activity, deficiency of dietary fiber, deficiency of vitamin D, deficiency of folate, high intake of red and processed meat, and alcohol consumption. Some of these risk factors, however, are closely related. For example, inadequate fiber intake and excessive fat intake are dietary risk factors which tend to lead to a lack of exercise which ultimately may contribute to obesity, particularly in combination. In the US, 40 percent of adults are obese, and so the risk factors discussed are common mainly due to the modern Western lifestyle. Therefore, it is no surprise that nearly half of the CRC cases arise in the developed nations. The Western diet in its current form contains more risk factors than the calorie and fat content. Foods that contain heterocyclic amines , polycyclic aromatic hydrocarbons , and emulsifiers can also contribute to carcinogenesis. HCA and PAH are produced in meats when they are fried or grilled over an open flame. These substances have been proved to damage the DNA of colonocytes and potentially promote risk of colon cancer. Emulsifiers are used in foods like ice cream to ensure an even distribution of fat molecules. Recent evidence suggests, however, that emulsifiers promote intestinal inflammation, creating an environment that favors colon carcinogenesis in mice. Some of these risk factors, however, are closely related. For example, inadequate fiber intake and excessive fat intake are dietary risk factors. These tend to lead to a lack of exercise, which ultimately contributes to obesity. In the US, 40 percent of adults are obese, and so the risk factors discussed are common mainly due to the modern Western lifestyle. Therefore, it is no surprise that nearly half of CRC cases arise in developed nations. However, colon cancer has a long development period . This gives ample time for lifestyle changes to take place, including diet-based intervention. Chronic inflammation, a condition that is promoted by dietary risk factors also contributes to the development of cancer, even in humans. Patients with inflammatory bowel disease have a significantly increased risk of developing CRC, while long-term aspirin treatment is associated with a significantly decreased risk of CRC. For example, the IL6/STAT pathway discussed earlier is also implicated in cancer formation. Over expression of IL6 leads to excess STAT3 transcription, causing unwanted cell proliferation not only in T cells but also in the intestinal epithelium. Another inflammatory cytokine of note is TNF α. While the intestinal bacteria can promote inflammation, they may also affect the likelihood of CRC more directly. Once the intestinal mucus layer is thinned, and direct bacterial-epithelial cell interactions occur, certain bacterial strains promote tumor development. E. coli strains bearing the pks island are of particular interest. This genetic locus codes for the secondary metabolite colibactin, along with the enzymes necessary for its production.

Comparing estimates from columns and suggests that the inclusion of time invariant parcel characteristics, such as parcel size and distance to the coast, as well as time-varying lagged crop choice and lagged recycled water deliveries is important.Our empirical approach is deliberate in conditioning on an array of factors that are likely correlated with both crop choice and salinity. The inclusion of the annual time trend and water prices in column provides an opportunity to test if our estimates of WTP are robust to their inclusion. The stability of our estimates between columns and lends credibility to our identifying assumption that, conditional on observables, salinity is uncorrelated with unobservables that may impact crop choice. A look at the marginal effects reveals how crop shares in the region change due to salinity. Using the estimates from column of Table 3.3, we hold the control variables constant at their average levels and show the predicted share of parcels in each crop type under a constant basin-wide TDS level of 2000 mg/L in Table 2.3. This is depicted relative to the average share of crops by type in the sample period. We see that given this increase in TDS of almost 1,360 mg/L, fallow ground would increase from 10.6% of parcels to 12.3% of parcels, holding all other variables at their averages. Likewise, vegetables and strawberries would increase slightly by 1.3 and 1.7 percentage points, respectively, and caneberries would decline by 4.5 percentage points. An increase in vegetable crops relative to fallowed land due to a change in salinity, holding all else constant, is intuitive due to their insensitivity to salt relative to the other regional crops, as shown in Figure 3.4. Interestingly,round plastic pots shares of strawberries also increase and the greatest declines are observed in caneberries, despite strawberries being relatively more salt sensitive. This may reflect the relative value per-unit of these crops or the fact that strawberries are often times rotated with vegetable crops such as broccoli, lettuce, and cauliflower for pest and soil management.

We next use these estimates to deduce growers’ willingness to pay for a reduction in groundwater salinity. We focus on crops whose estimates in Table 3.3 were significant at the 95% confidence level or greater, namely, vegetables, strawberries, and caneberries. The willingness to pay estimates for a reduction in groundwater salinity of 10 mg/L are reported in Table 4. While these willingness-to-pay estimates are wide-ranging, the high dollar values indicate that growers highly value water with low salinity levels. These WTP estimates are on the same order of magnitude as the expected revenue losses from a drop from 100% to 90% yield capacity as shown in Figure 3.4. To put these magnitudes into perspective, row cropland in Monterey and Santa Cruz Counties in 2021 ranged in value from $28,500 to $75,000 per acre, representing some of the most expensive cropland in the state. Contrast this to the range of values observed in 2021 for well-dependent cropland in Fresno county which ranged from $10,000 to $16,000 per acre . To assess the robustness of our results, we test the sensitivity of our results to two modeling choices. First, we take a closer look at our measure of salinity, which can be measured in different ways. We chose to focus on TDS in our main estimation, since it is widely used in the agronomic literature and is a measure of general salinity. This allows us to look at results that may be applicable across multiple types of salinity problems . One concern may be that growers actually respond to an alternative salinity metric when faced specifically with seawater intrusion. Chloride is a measure of salinity that specifically captures seawater intrusion, rather than salinity from other sources, such as soil, rock, or other natural materials. Table 2.5 reports marginal effects based on the estimation of our preferred specification from the last column of Table 3.3, except we replace the salinity variable with chloride measurements. On average, chloride levels are 15% of TDS values, so an increase in chlorides to 300 mg/L is similar to a shift in TDS to 2,000 mg/L. As shown, results are largely the same across these two highly correlated measures of salinity.

Larger marginal effects are estimated for vegetable row crops and caneberries when using chlorides, which may be due to the fact that the spatial distribution of chloride concentrations is different than that of TDS. Finally, we take into account the possibility that other water resources may be available to a subset of growers in the Pajaro Valley. Farmers located near the coast experience some of the highest salinity levels relative to the rest of the basin, and simultaneously impose the greatest externality on others when they pump groundwater. PV Water recognizes this, and in collaboration with the growers in the basin and the City of Watsonville, set up the Delivered Water Zone and a distribution system to deliver recycled water and other alternative supplies to the growers most impacted by seawater intrusion. The distribution system serves roughly 15%, or 5,000 of the 30,000 acres farmed in the Valley. The total quantity delivered has slowly increased from 667 AF in 2005 to 4,203 AF by 2016. This is a small fraction of irrigation water used within the DWZ, but it is likely still relevant for farm-level decisions. It is plausible that growers inside the DWZ have different projections of their future access to high quality water and are less responsive to groundwater salinity, compared to growers without access to delivered recycled water. In addition, anecdotal evidence from conversations with growers in the Valley suggests that growers inside the DWZ who had been unable to plant strawberries before they started receiving deliveries are now able to plant the salt-sensitive crop once again. While we conditioned explicitly on recycled water deliveries to farmers inside the DWZ in our initial estimation, we perform a robustness check by removing all parcels within the DWZ, to focus solely on the effects of salinity on crop choice when there are no other water sources available. The fraction of parcels planted in various crop categories differs for this subset so marginal effects in this case need to be compared to a different baseline. Marginal effects from the estimation of salinity impacts on this sub-sample are shown in Table 2.6.

Results are very similar in magnitude and significance to those reported in Table 2.3, suggesting that the existence of the DWZ is not biasing our estimation of the relationship between salinity and crop choice. Finally, we ask the question of what would happen to crop choices and to consumer welfare under a scenario in which the quality of groundwater deteriorates significantly across the basin. We simulate and compute the utility-maximizing crop choices for each parcel in each year under a high TDS scenario and plot the distribution. To do this, we keep the estimated marginal utilities for the attributes of crops, parcels, and climate variables the same as in the panel mixed logit model displayed in column of Table 3.3. For each parcel, we estimate the probability of choosing each crop type and use these to predict each parcel’s baseline crop choice. Then,hydroponic bucket we recalculate the probabilities of each crop being grown for each parcel after altering the vector of TDS values to reflect a higher salinity scenario. Ideally, we would simulate the change in TDS predicted by a climate change model of sea-level rise. This exercise is challenged by the fact that model predictions of sea-level rise, while necessary, are insufficient for deploying our model of crop choice. To use climate model output in our simulation, we would need to map sea levels to the salinity concentration in the groundwater wells on each parcel throughout the Pajaro Valley. Doing this would require the use of a hydrologic model of the groundwater basin, which is beyond the scope of this paper. Further, seawater intrusion is highly influenced by demand for groundwater resources, so the decomposition of changes in salinity to either sea-level rise or to groundwater overdraft is challenging. Climate change is predicted to cause higher temperatures and more variable precipitation, which may lead to increasing demand for groundwater, in addition to a rise in sea levels . Instead, we opt to model a realistic increase in TDS by looking at how much groundwater salinity has increased in the basin over our sample period. The average spring TDS during our sample period is 644 mg/L. From 1990-2020, four years had an average TDS greater than double this value, with an average increase of 10% annually. For a relatively straightforward simulation, we increase the current-period TDS by 100%, to see how crop choice would evolve with this plausible shift in salinity. All other variables, including weather, remain stable for this analysis, which allows us to focus on how salinity specifically impacts cropchoice distributions. The estimated change in the distribution of crop choices is plotted in Figure 2.8. The graph plots the difference between the original model estimates of crop choice and the simulated estimates of crop choice under an increase in TDS conditions of 100%. Caneberries experience the largest shift in parcels planted under the 100% increase, which coincides with their sensitivity to salinity, as well as their relatively lower profitability when compared to strawberries, another salt-sensitive crop. Vegetables experience the largest increase in the probability of being planted, as they are the least salt-sensitive. Fraction of land left idled also increases substantially under the high TDS scenario.Seawater intrusion, which occurs when saline water from the ocean enters a freshwater aquifer, can manifest from two primary drivers: groundwater extraction and sea-level rise. Pumping groundwater faster than the natural rate of recharge can move seawater to freshwater zones, and sea-level rise alters where saltwater sits relative to freshwater in the aquifer .

Salinity is a major concern for coastal agricultural production that is dependent on groundwater for its water supply , but it can also have significant impacts on inland irrigated agriculture as salts accumulate in the soil over time. Increased salinity levels in agricultural water lead to declines in agricultural productivity, and farmers are left with few mitigation strategies. In this paper, we empirically evaluate the likelihood that farmers switch crops in response to changing groundwater salinity, with an application to the Pajaro Valley, a coastal region in California. Unique spatial panel data on groundwater quality and land use spanning 11 years lends itself to a panel mixed logit model of crop choice. This revealed preference approach allows us to estimate the marginal willingness-to-pay for improvements in irrigation water salinity. We find evidence that growers are more likely to shift away from crops that are saltsensitive, such as strawberries and caneberries, relative to fallow ground, when facing an increase in groundwater salinity. The marginal WTP to pay for a 10 mg/L reduction in TDS varies by crop, and ranges from $1,613 for strawberry growers to $16,369 for caneberry growers. Our simulation of a 100% increase in TDS across the basin speaks to potential land use changes in the basin if salinity trends continue. We estimate that a change in TDS of this magnitude, which could realistically occur in a future defined by sea-level rise and continued groundwater overdraft, would result in a welfare reduction of $140 million. While our WTP and marginal damages estimates are restricted to an agricultural region defined by the jurisdiction of a single water management agency, most groundwater management decisions and investments are made at this scale. The paucity of robust, geospatial groundwater quality data that can be paired with accurate planting information precludes us from deriving estimates in other regions. While salt sensitivity is crop specific, which will drive differences in the WTP across regions, this methodology is generalizable and can be applied elsewhere to determine the benefits of reducing groundwater salinity. Salinity is becoming an increasingly common issue across the United States and the globe as sea levels rise and groundwater aquifers become more stressed under climate change. Estimating marginal damages from changing salinity can provide new context for the cost of climate change and the cost of groundwater overdraft, both of which are broadly important for groundwater management.The stability of water resources for agricultural production has always been an important topic, but the scale and urgency of the issue has dramatically increased in recent decades.

The principle is that the features of samples belonging to the same category are close to each other, while the features of samples belonging to different categories are far from each other. The earliest representative work is Siamese Network, which is trained with positive or negative sample pairs . Vinyals et al. proposed the Matching Networks, and they borrowed the concept “seq2seq+attention” to train an end-to-end nearest neighbor classifier. Snell et al. proposed Prototypical Network, which learns to match the proto center of class in semantic space through few samples. Sung et al. proposed Relation Network, which concatenates the feature vectors of the support samples and the query samples to discover the relationship of classes. Li et al. proposed CoveMNet based on the covariance presentation and covariance metric of the consistency of distribution. The network extracts the second order statistic information of each category by an embedding local covariance to measure the consistency of the query samples with the novel classes. Chen et al. proposed Meta-Baseline method, which achieves good performance on some FSL benchmarks. The accuracy achieves at 83.74% with 5-way, 5-shot task of Tiered-ImageNet, and 90.95% with 1-way, 5-shot task of Mini-ImageNet. Recently, FSL has started to be used in research on plant disease identification. Argüeso et al. used Siamese Network on the dataset Plant Village . Jadon proposed SSM-Net that uses the Siamese framework and combines two features from a Conv and a VGG16. Zhong et al. proposed a novel generative model for zero-shot and few-shot recognition of citrus aurantium L. diseases by using conditional adversarial auto-encoders. Afifi et al. compared Triplet network, Baseline, Baseline++,strawberry gutter system and DAML on PV and coffee leaf datasets. The results show that the Baseline has the best performance. Li and Chao proposed a semi-supervised FSL method and tested it with PV.

Nuthalapati and Tunga introduced transformer into plant disease recognition. Chen et al. used meta-learning on Miniplant-disease dataset and PV. Li and Yang used Matching Network and tested cross-domain performance by mixing pest data. These methods have been tried from various perspective and have made important progresses. Nevertheless, FSL still has two common challenging issues: limited features extracted from few samples are less representative for a class ; the generalization requirements are very high and various. In this work, we tackle the two issues by using multi-scale feature fusion and improving training strategies. CNN is widely used in image-based deep learning methods. In a CNN architecture, the local features with more details and small perceptive fields are extracted from low-level layers, while the global features with rich semantic information and large perceptive fields are extracted from high-level layers . MSFF is the technology using multi-scale features which are extracted from different layers of CNN . In object detection and semantic segmentation, many excellent networks are proposed by using MSFF, such as Feature Pyramid Network , Unet , Fully Convolutional Network etc. MSFF is also used in image restoration, image dehazing and image super resolution etc. . These methods fuse features by using dense connection, feature concatenation or weighted element-wise summation . In common, the mentioned methods have encoder-decoder framework. The multi-scale features extracted from encoder are reused in decoder to enhance feature representation. However, in conventional classification task, MSFF is seldom used because the network does not have decoder. Generally, only the top semantic features are fed into classifier, but other scale features are abandoned. But in fact, the high-level features and the low-level features are not subordination relationship. The local features including rich fine-grained features can be an effective compensation to formulate a richer feature representation of sample . In the data-limitation condition, it requires to extract as many features as possible from a limited amount of data.

Therefore, in this work, we propose to leverage the MSFF to enhance feature representation. Multi-scale features can be fused in different ways. In our work, we use cascaded multi-scale feature fusion . The channels of feature maps increase after feature fusion. But it does not mean that all channels are the same significance. The contribution of each channel is different. Some channels should be emphasized and some should be suppressed. Attention can help to focus on the meaningful channels. Attention mechanism plays important role in human perception to selectively focus on salient parts in order to capture visual structure better . It has been leaded into some areas of machine learning such as computer vision, natural language processing etc. and has significance to improve performance . It not only tells where to focus, but also improves the representation of interests. Recently, some light-weight attention modules have been proposed. Wang et al. proposed Residual Attention Network that uses encoder-decoder style attention module. Hu et al. introduced a compact module to exploit the inter-channel relationship, which was named as Squeeze-and excitation module. Woo et al. proposed Convolutional Block Attention Module that includes channel attention and spatial attention. These light-weight attention modules can be easily embedded into deep learning networks as plug-ins. In this work, we use the CA to weight the accumulated channels obtained from CMSFF. As the definition of FSL, it is asked to generalize to novel categories or novel domains. Generalizing to new categories within the same domain of training is defined as intra-domain classification, while generalizing to novel domain is defined as cross-domain classification. Long-tail distribution of data is common in plant disease datasets. To identify the part of categories with few samples, the model can be trained with the part of diseases that have more samples.

This generalization happens in the same domain. Cross-domain happens when a set of categories with few shots is required to be identified but does not belong to any dataset. Cross-domain adaption happens between different datasets, which is more difficult than intra-domain adaption. However, researchers found that it is frequently encountered situation and inescapable for boosting FSL to practical application. Guo et al. established a new broader study of cross-domain few-shot learning benchmark and pointed out that all meta-learning methods underperform in relation to simple fine-tuning methods, which indicates that the difficulty of the cross-domain issue. Adler et al. proposed a method of representation fusion by an ensemble of Hebbian learners acting on different layers of a deep neural network, which is from feature representation perspective. Li W.-H. et al. proposed a task-specific adapters for cross-domain problem from the perspective of network architecture. Qi et al. proposed a meta-based adversarial training framework for this problem, which is also from the perspective of network architecture. As we know, there is no research that has been done from a training strategy perspective. These efforts are the kind of general explorations of using general benchmarks and rarely discuss specific domains. In fact, different domain has its own characteristics and resources to utilize when crossing domains. Hence, in this work, we propose a set of training strategies to match various cases of generalization using the available data resources. The contributions of this work are summarized as: we propose a Meta-Baseline based FSL approach merging with CMSFF and CA for plant disease recognition; we propose a group of training strategies to meet different generalization requirements; through extensive comparative experiments and ablation experiments,hydroponic fodder system we validate the superiority of our method and analyze various factors of FSL. Comparing with the existing related works under the same data conditions, our method has achieved at the best accuracy.In this research, three public datasets are used in our experiments. Mini-ImageNet is a subset of the ImageNet, which includes 100 classes and 600 images per class. We select 64 classes in our experiments. The second is PV released in 2015 by Pennsylvania State University. It is the most frequently used and comprehensive dataset in academic research up to now in plant disease recognition. Totally, it includes 50,403 images which crosses over 14 crop species and covers 38 classes, as shown in Table 1. Because the number of samples in PV is unbalanced, we use the data after augmentation and select 1,000 images per class to keep balance. The third is the dataset of apple foliar disease , which was published in FGVC8 Plant Pathology 2021 Competition. All images of AFD were taken in wild with complicated backgrounds, as shown in Figure 1A. Like classical classification structure, our framework contains two components: an encoder and a classifier, which is illustrated in Figure 2A.

The encoder noted as fθ is a CNN-based network merging with CMSFF and CA. It is trained in two stages: base-training and meta-learning. In base-training, the network contains fθ and base-training classifier, which is trained with image-wise data. The goal in this stage is to learn the general features as prior knowledge. Some large-scale general datasets with more classes and diverse data, such as ImageNet, Mini-ImageNet etc. are good choices for learning prior knowledge. The classifier can be linear classifier, fully connected layer, SVM, or other classifiers. The cross entropy loss is calculated to update the parameters of fθ during back propagation. After base-training is completed, the classifier is removed and the trained model is delivered to the meta learning stage. In meta-learning, fθ is initialized by the trained model from base-training. Meta-learning is a concept of learning to learn. So, the purpose is not to learn the knowledge of the training classes, but to learn how to differentiate between classes. Aiming at the objective, the classifier in meta-learning is replaced by a distance measurement module. The classification result is decided by the distances from the support samples to the query sample. Meta-learning is a task-driven paradigm where training data is formulated as N-way, K-shot tasks. Based on a simple machine learning principle: test and training conditions must match , the data of Cnovel is also formatted into tasks in test. After embedding, the 2D color image has been a high dimensional vector in semantic space. The distance of query sample to the class centroid is calculated by a distance metric. Distance metric uses distance function which provides a relationship metric between each element in the dataset. In many machine learning algorithms, distance metric is used to know the input data pattern in order to make any data-based decision. The most common used measures to calculate the distance between two vectors are cosine similarity, dot product and Euclidean distance. Cosine similarity is a measure of similarity between two non-zero vectors of an inner product space. It is measured by the cosine of the angle between two vectors and determines whether two vectors are pointing in roughly the same direction. It is the same as the inner product after normalization . In Euclidean geometry, the dot product of the Cartesian coordinates of two vectors is widely used. It is often called as inner product or projection product of Euclidean space. The length of projection represents the distance of two vectors. In mathematics, the Euclidean distance between two high-dimensional vectors is the square root of the sum of the squares of the distances in each dimension. Basically, the structure of MSFF includes two categories: parallel multi-scale feature fusion and cascaded multi-scale feature fusion . The two fusion methods are illustrated in Figure 2B. The PMSFF concatenates the features from different layers of CNN simultaneously. The different resolutions of feature maps are uniformed before concatenation. Comparatively, the CMSFF fuses the different resolution feature maps step by step. Taking Resnet12 as backbone network, four convolutional blocks are linked. A group of feature maps of double times of channels and half resolution is generated after each block forwarding. In the backward fusion, small size feature maps are two times up-sampled and concatenated with the feature maps of previous block. After a series of up-sampling and concatenation, all channels are fused together to be the fused full-scale feature, noted as F. The CMSFF is used in this work. The domain of training is noted as source domain , and the domain of test is noted as target domain . Data from different domains can be used in the three stages: base-training, meta-learning, and test. It is special that there are two training stages of our method, and the datasets used in the two stages could be different. We just consider the domain of meta-learning stage as SD. When SD is the same as TD, it is intra-domain adaption, otherwise, it is cross-domain adaption. In order to mimic different adaption situations, we design different data configurations.

If, in contrast, gibbons would react competitively as other great ape species did in previous social dilemmas, we would expect them to try to maximize their food rewards by hesitating to act and by taking advantage of their passive role—placing themselves in front of the release mechanism. In that sense, we would expect passive partners to benefit from their position and obtain more rewards than the actors. Furthermore, we would also expect gibbons to be more likely to act in direct food test trials given that they could easily obtain at least one direct reward from their actions.Therefore, actors were still able to obtain the majority of the five released rewards in both conditions. Furthermore, actors obtained the extra rewards in 87% of the direct food test trials in which they acted. In line with this finding, Fig. 2 shows the percentage of trials in which individuals release food against the number of rewards that each individual obtained in indirect and direct food test trials. Interestingly, in only one dyad the more active individual obtained less benefits than the passive partner in both conditions. We also found significant effects of our control variables age and length of dyad suggesting that younger individuals obtain more food and that dyads who have been longer together distributed the five rewards more equally. Next, we evaluated whether passive gibbons acted strategically by placing themselves in front of the ramp at the moment the actor released the five food rewards and whether they would take advantage of their position in direct food test trials compared to indirect food test trials given that the actor gibbon would spend some time retrieving the food baited inside the handle.We also explored whether gibbons would take an advantageous position after the food had been released.

For that we measured whether the passive individual was in front of the ramp by the time the actor arrived at the location where the food had been released. After releasing the food,hydroponic nft the actor moved to the food location in 237 of those 287 trials . Passive individuals were in front of the ramp by the time the actor arrived in 76 occasions . We found no differences between conditions . We found that actors did not approach the released food in 50 trials in which they released it. A majority of those trials were direct food test trials . This makes sense since actors spent some time obtaining the food reward placed inside the handle. Nevertheless, in a great majority of these trials the actors ended up obtaining the food reward from the handle before the passive individual finished her last reward. Interestingly though, this behavior predominantly occurred in three of the six dyads. Furthermore, we only found one case in which the actor showed an intention to approach the partner by getting closer to him.Te results of the study suggest that dyads of gibbons managed to solve a competitive food task where one dyad member had the opportunity to pull and activate a release mechanism containing five potential food rewards for both partners. When gibbons had the opportunity to obtain an extra reward from their actions , they almost always acted, avoiding mutual defection. In contrast, in an indirect food test condition where gibbons could not obtain the extra rewards from their pulling efforts, their likelihood to pull and release potential food rewards dropped significantly in comparison to the direct food test condition. In our opinion, two primary reasons could explain this pattern of results. First, it is possible that gibbons were more motivated to pull when they could directly benefit from the extra reward inside the handle. Similarly, gibbons would have significantly pulled more often in the indirect food test condition compared to the no food control because they could still obtain some rewards. Clearly, gibbons showed that they did not just act for the sake of pulling the rope. Most likely, their actions were motivated by the prospect of obtaining rewards, especially when those were easier to access. While this reason is very plausible, it does not necessarily account for why gibbons did not pull for 90 s in almost 40% of the indirect food test trials.

This is especially surprising if we consider that actors benefitted more than passive partners in this condition. In other words, gibbons did not seem to interpret the situation as beneficial for actors. If that were the case, we would have expected gibbons to pull more often in indirect food test trials. Thus, the second possible explanation for our results is that gibbons interpreted the situation as a conflict of interest and hesitated to pull to avoid losing rewards in favor of the passive partner. This interpretation would be in line with previous findings in chimpanzees, bonobos and common marmosets. Thus, given the two non-mutually exclusive explanations, it remains unclear whether gibbons defected in indirect food test trials due to a reduced motivation to act because they could not access the extra reward attached to the handle or because they wanted to avoid the possibility of losing rewards to their partner. So far, we have mainly discussed gibbons’ decisions whether to pull or not. The next question concerns whether gibbons strategized when they took a passive role. In other words, did they try to maximize their own rewards when their partners pulled? The main source of potential conflict between participants lied on the possibility that passive individuals could position themselves in front of the release mechanism during direct and indirect food test trials. In direct food test trials, this could be particularly beneficial for passive partners given that actors could lose some valuable seconds trying to obtain the extra food reward attached to the handle. However, we found that actors actually obtained most of the rewards in both conditions, with a special advantage over passive partners during indirect food test trials. In fact, in only one dyad of gibbons the passive individual obtained more rewards than the actor in both test conditions. Furthermore, passive individuals rarely took advantage of the situation and they did not distinguish between conditions. That is, during direct food test trials passive partners did not benefit from the time that the actors spent trying to obtain the reward located inside the handle. In that sense, gibbons did not interpret the situation as a social dilemma in which they could benefit more than their partner.

Yet, gibbons also did not solve this task cooperatively. For that to be the case, dyad members would have needed to benefit more or less equally on both conditions and they would have not hesitated to manipulate the handle in indirect food trials. It is also very unlikely that gibbons’ decisions were driven by proactive prosocial motivations such as releasing food rewards to favor their dyad members given that actors benefited the most from their own actions. One possible explanation to understand why actors obtained more rewards than passive partners is that both individuals approached the handle during direct food test trials to obtain the reward, although only one individual pulled the handle. This occurred in 27.6% of the direct food test trials in which an individual pulled the handle. This possibility could partially explain why passive gibbons rarely position themselves in front of the release mechanism during direct food test trials, but it cannot explain why in indirect food test trials they did not take advantage of their passive role. In fact, in indirect food test trials passive partners approached the handle in 10% of trials. However, there are numerous reasons to suggest that a dominance component cannot fully explain our pattern of results. First,hydroponic channel only three of 12 individuals released rewards in less than 20% of the direct and indirect food test trials. Te result suggests that although they did not solve the task cooperatively, in half of the dyads both individuals exchanged roles relatively often. Furthermore, our results are also in line with literature suggesting that there is no clear dominance of one sex over the other and that gibbons engage in food sharing in both captive and natural settings. Importantly, conflict avoidance does not seem to be relevant here either. Cofeeding events occurred three times as often than displacements events, suggesting that individuals were usually tolerant with each other, as it has been observed in other populations. Relatedly, gibbons never hoarded the blueberries. They ate them as soon as they got them. Furthermore, these tolerance may be mediated by the length of time that dyads had spent together. In our study, the length of dyad predicted how equally gibbons divided the five food rewards. A third alternative explanation is that once a gibbon decided to manipulate the handle, the other one totally disengaged from the task, as if acknowledging a sort of property claim. This could explain why passive partners did not take an advantageous position in front of the release mechanism and why, as a consequence, actors did not face a social dilemma in many trials. After all, actors benefit more than passive partners despite the costs to pull the rope, understood as the possibility to lose rewards due to the distance they had to cover and the time lost to return to the location where the food was released. Nevertheless, this is unlikely because passive individuals still obtained a significant fraction of the food rewards .

All these previous arguments cannot fully explain why passive partners rarely took advantage of their position, especially given how successful this strategy was: passive partners obtained almost 75% of rewards when they positioned themselves in front of the release mechanism by the time the actor manipulated the handle. We thus propose that passive gibbons did not always take advantage of their partner actions because of the combined processes of motivation from the side of the actor and a general high level of social tolerance towards inequities. Tat is, individuals that were more motivated to obtain food and more attentive were also more likely to take the actors’ role in our task. Trough participation, they could become more aware of the situation as a whole and react faster to obtain the rewards despite the potential costs of pulling the rope. This gave them an advantage over their passive partners, whom at the same time tolerated actors obtaining the majority of rewards . Importantly, tolerance towards reward inequities also came from the perspective of the actors. Indeed we found that in fly trials individuals from five of the six dyads seemed to adopt a prosocial attitude towards their passive partners by letting them access all the released rewards. Future studies should improve different aspects of our setup to continue exploring gibbons’ decision-making strategies when individuals’ interests collide. Te main weakness of our design is that we were not able to separate dyads of gibbons before the experimental sessions. In that sense we could not train them with the different task contingencies as it is usually the case in this type of settings but see. It is thus possible that some individuals were more skillful than others, and that might have affected our results. Nevertheless, all individuals approached the apparatus and obtained rewards and only one never pulled from the handle during the course of the study. Moreover, despite the fact that gibbons distinguished between conditions with food and the no food control, their latencies to act did not differ between the indirect food test trials and the no food control trials. This could be interpreted as strategic behavior in the indirect food test trials if the gibbons were waiting for their partners to act. It is possible that with longer trials we would have found a significant difference between gibbons’ latencies in indirect food test trials and no food control trials. In that sense, future changes in the trial time or the food rewards can better assess whether gibbons strategize to solve conflicts of interest. Relatedly, it is possible that gibbons were more motivated to act during direct food test trials compared to indirect food test trials because there were more rewards in play . It has been found that at least great apes are able to distinguish between these two amounts when they are presented at the same time.

Using standardized observations of floral visitation and seed set measurements of yellow starthistle, we test the hypotheses that increasing urbanization decreases 1) rates of bee visitation, 2) viable seed set, and 3) the efficiency of pollination . In addition to contributing to a better understanding of how change in landscape use, particularly urbanization, affects pollination-plant interactions, the study illustrates the importance of use of neighboring lands for pollination services.Our study system was located around Brentwood, in east Contra Costa County, California, where natural, agricultural, and urban areas intersect with each other within a 20620 km region . A county water district , regional park district , and California state park all fall within the region, leaving large areas of land protected from development. This protected land consists mainly of grasslands and oak woodlands, some portions of which are managed for grazing. East Contra Costa County has had a farming community presence since the late 19th century. The agricultural areas of Brentwood, Knightsen, and Byron mostly consist of orchards , corn, alfalfa, and tomatoes. A housing boom in the 1990s led to massive residential growth in the area. The city of Brentwood has grown from less than 2500 people in the 1970s to over 50,000 today , and nearby Antioch has now over 100,000 residents .We selected 12 sites dominated by yellow star thistle in a stratified design to span the different land use types . Yellow starthistle is a common weedy plant that forms homogenous flowering patches in grassy areas throughout this region. Many different bee taxa in a range of functional groups and size classes have been observed to visit yellow starthistle, in part because it flowers late in the season relative to other floral resources. Despite being considered a serious introduced weed, yellow starthistle is unusual as an invasive species in that it depends on animal pollinator visits in order to set seed. Within each site we selected a 50 m650 m plot such that each plot was at least 2 km away from all others,hydroponic gutter a distance larger than the maximum assumed typical bee foraging ranges.

Although certain bee species have been recorded foraging as far as 1400 m, most bees in this type of habitat have nesting and foraging habitat within a few hundred meters of each other. Within each plot we estimated number of flowering yellow starthistle blooms by randomly placing 10, 1 m61 m quadrats and counting the number of flowering blooms in each. We also measured the spatial area of yellow starthistle patches within each 50 m650 m plot to obtain an estimate of total flowering blooms within each plot. We categorized total blooms/plot on a log scale: ,103 , 103 –104 , and .104 . Using NOAA’s 2006 Pacific Coast Land Cover dataset , a 500 m buffer was created around each plot, and the number of pixels classified as agricultural, urban, natural, water, or bare land was extracted. These categories were obtained by lumping finer categories in NOAA’s classification scheme using the following definitions: Urban–‘‘High Intensity Developed’’, ‘‘Medium Intensity Developed’’, ‘‘Low Intensity Developed’’, and ‘‘Developed Open Space’’; Agricultural–‘‘Cultivated’’, ‘‘Pasture/ Hay’’; Natural– ‘‘Grassland’’, Deciduous Forest’’, ‘‘Mixed Forest’’, ‘‘Scrub/Shrub’’. Each plot was classified as a proportion of each of the 3 different land use categories, as well as for the category that was dominant. By this latter measure, of our 12 sites, 4 of each were classified as ‘‘urban’’, ‘‘agricultural’’, and ‘‘natural’’.We observed visits by all bee species to yellow starthistle at all sites 3 times for a 30 min period for a total of 90 min of total observation time per site within the same 2 wk period in August 2011. AM was defined as being between the hours of 9:30–11:30, Mid-Day as between 11:30–13:30, and PM as between 13:30–15:30. Also recorded at each observation period were approximate number of blooms, and wind and temperature simultaneously . Bees were not netted for later identification as we did not want to interfere with visitation to starthistle during this study. Instead, we used a modified protocol of citizen scientist observation surveys with 15 expected bee morphotypes that correspond to 30 possible genera known to occur in the region .

The observer slowly walked through the yellow starthistle patch, and upon reaching patch edge, returned on a path at least 3 m away from the previous, and recorded the morphotype classification of all bee visitors within 1.5 m on either side of the transect.Yellow starthistle has composite flowers, which are aggregations of anywhere from 20–80 florets. At each site, 12 yellow star thistle buds were randomly selected from different plants and covered with a mesh bag. Yellow starthistle blooming cycles have been described in detail in other publications. We selected buds at stage BU-4, when buds had no yellow petals exposed, but had well-developed straw-colored spines. When in full flowering, 10 bags were opened for a 4 hour period from 10 am to 2 pm, while 2 were kept closed as controls to verify that self-pollination was not occurring. At the opening and re-closing of the bags, the number of florets that had their stigmas extended were counted. Later, when flowers were fully mature , seed heads were collected, and later dissected in the lab. Viable and non-viable seeds in yellow starthistle seed heads are easily distinguishable based on color and shape. Because yellow starthistle requires pollination to produce viable seeds , non-viable seeds represent pollen limitation occurring during the 4-hour period that the flowers were exposed to pollinators. All seeds were counted to compare ratios of viable to non-viable seeds. Any seed predation was noted, and when possible, the seed predator was identified.All analyses were done in R 2.15.1 . Because each site had an AM, Mid-Day, and PM observation event, there were a total of 36 observation events, each with unique wind and temperature recordings, and visit observations of the 15 bee morphotypes. From these, we calculated the total number of bee visitors, total number of bee morphotypes, Shannon diversity of morphotypes, and morphotype evenness. Shannon diversity and evenness were calculated using the R package vega. The spatial auto-correlation of all bee visitor response variables was assessed by Mantel tests in R package ade4, using the average values for each time of day at each site. Spatial auto-correlation was not detected . To test for the effect of land use type on each of the response variables we used a generalized linear mixed model using the R package lme4. We designated land use type, bloom category of flowering patch, observation time period, wind, and temperature as fixed effects and site as a random effect.

Natural land use and AM observation time period were the model baselines for the categorical variables of land use type and observation time. Shannon diversity and evenness were fit with Gaussian distributions while all other variables were fit with Poisson distributions. In comparing the ratios of viable seeds to total seeds vs. the ratio of viable seeds to counted stigmas, we found that there was a strong correlation between these metrics. To look at the effect of land use type on seed-set, we therefore decided to utilize the ratio of viable seeds to total seeds in each seed head that did not experience seed predation, because of error in counting the number of stigmas . We then used a generalized linear mixed model fit with a Binomial distribution, with land use type as a fixed effect and site as a random effect. Finally,u planting gutter we tested for an effect of floral visitor observations on yellow starthistle seed set at each site. We averaged the number of visits from each morphotype across temporal observation events at the same site. Morphotypes that averaged at least one visit per 30 minute observation window were included as fixed effects in alinear mixed model fit with a binomial distribution, with site as a random effect and the ratio of viable to total seeds as the response variable. We also modeled the effects of total bee visitation, morphotype richness, and morphotype diversity on seed set ratios.Our results show that rates of bee visitation and seed set vary among urban, agricultural, and natural landscapes, demonstrating the importance of land use in the dynamics of plant-pollinator interactions. We suggest that these effects are at least in part explained by floral availability, a vital bee resource, which can be highly variable among different land use types. For example, in August there are few plants in flower besides yellow starthistle in the natural areas of Contra Costa County, California, whereas in urban and agricultural areas there are many exotic plants and supplementary inputs available . From pantrapping of bee specimens in the region , we know that total bee abundance is highest in the spring in natural areas. However, towards the end of the summer when yellow starthistle is in flower, there is little difference in collected bee abundance between human-altered landscapes and natural areas, and human-altered areas may even exhibit overall higher bee abundance. Our results of bee visitation to yellow starthistle support this pattern. Agricultural areas have large populations of managed honey bee colonies, so one would predict visitation to yellow starthistle by honey bees to be positively associated with surrounding agricultural land use. By contrast for native bees , the highest rates of visitation to yellow starthistle were in sites with more surrounding urban land use. Urban gardens have many exotic plants, often selected for aesthetic purposes, many of which are in flower later in the season than most California native plants. In addition, many of the plants in urban areas both directly and indirectly receive supplementary resources, particularly water, that further extend their flowering time. Even though agricultural areas also have supplementary resources, the main crop in flower in East Contra Costa County later in the season is maize, which is wind pollinated. There may be multiple impacts of exotic plants in urban areas. By filling the phenological flowering gap noted above, they may help attract even larger populations of bees into the urban landscape. In addition, bees in urban sites may be behaviorally more likely to visit non-native plants due to the increased encounters they have with novel plants. In agricultural and natural landscapes, a positive correlation between pollinator visitation and seed set is typical. Surprisingly in our system, in human-altered landscapes, higher total observed bee visitation did not result in higher proportions of seed set, as would be expected. In fact, urban areas, despite receiving the highest rates of native bee visitation, exhibited the lowest rates of seed set. Conversely, natural areas, which received the lowest amount of total bee visitation, had the highest rates of seed set. We suggest 2 possible explanations for this discrepancy between pollinator visitation and rates of seed set: 1) pollinator efficiency; and/or 2) the composition of the local flowering community. Depending on the plant, certain pollinator species are much more effective than others. For example, Osmia, Habropoda, and Apis, have been found to produce varying amounts of seed set as a result of a single visit to blueberry, but these results vary slightly depending on the blueberry variety. In the case of yellow starthistle, it is likely that the most frequent visitors are perhaps not the most efficient. When we directly compared average seed set at each site against visitation rates, we found a significant positive association with the medium hairy leg bees. The medium hairy leg bee morphotype includes those species which fall in both the Tribes Emphorini and Eucerini. Emphorini are known to largely be oligolectic , meaning they specialize on certain plant groups, which theory suggests would make them more efficient pollinators than generalists. The medium hairy leg bee morphotype was not significantly associated with any of the land use typesIt was also the only group that was observed most frequently during morning sampling, perhaps reflecting a difference in when yellow starthistle is most receptive to pollination. Despite the overwhelming abundance of honey bees in agriculture areas, we did not observe higher seed set in those regions, consistent with the observation that honey bees can be poorer pollinators than other species.It is also important to note that this study used a morphotype classification, and there may be multiple species that fit within the same morphotype that provide varying degrees of pollination services.

Going forward, as researchers across various scientific domains and sectors come closer to a unified definition of resilience and perhaps agree to the use of a standard checklist for designing and reporting on resilience studies, there is greater opportunity to harmonize the science and develop more empirical evidence of resilience outcomes.Optimizing performance also includes building resilience in order to enhance the ability to perform tasks and ensuring resilience in order to prevent illness, injury, and disease. Within the US Department of Defense, researchers are able to study different models of physical and psychological stress and the application of different nutritional interventions with Service Members throughout their careers. Various models of stress are introduced, including initial military training , advanced military training courses , service academies , and extreme environments , along with examples of various interventions and outcome measures collected to date. The importance of nutrition on readiness and resilience was identified in military populations more than a decade ago and continues to be of interest. Two specific examples are provided to further explore nutrition interventions aimed at optimizing performance in the Department of Defense. The first, a completed double-blind, randomized, placebo-controlled trial, used a calcium and vitamin D fortified food product to optimize bone health during initial military training of Marine Corps recruits. Using a supplement or food intervention for calcium and vitamin D, participants received 2000-mg calcium and 1000-IU vitamin D per day. The primary outcomes of the study showed that bone markers and vitamin D status improve, but the supplementation did not affect skeletal parameters. A second,bato bucket forth coming study aims to evaluate the effectiveness of adding spices and herbs to increase vegetable intake among junior-enlisted Service Members.

Their current research is focused on the stressors and challenges to enhance resilience. Using a cycle of basic science/discovery that advances to clinical trials with various review steps helps move the field of nutrition science forward in a “total force fitness” approach. Total force fitness was introduced as a framework to help Service Members, their families, and military units reach and sustain optimal, holistic health, and performance in a way that aligns with their mission, culture, and identity. Other examples of frameworks focused on a holistic approach to research include Whole Person Health proposed by the National Center for Complementary and Integrative Health, Whole Health developed by the Department of Veterans Affairs, and a recent consensus study report by the National Academies entitled Achieving Whole Health. A focus on improving resilience as a model outcome highlights the opportunities and complexities of conducting optimal health and nutrition research in this space.As the number and proportion of older adults in the population increase, the prevalence of age-related deficits in mobility and cognition also increases. Such deficits may be because of normal aging or to pathologic processes. For instance, cognitive impairments like declines in memory and speed of processing may result from normal brain aging or neurodegenerative diseases like dementia. When considering hallmarks of optimal nutrition and health, improving resilience from cognitive decline has strong promise and impact. Although the etiology of age-related mobility and cognitive changes is multifactorial, it is well established that vulnerability to oxidative stress and inflammation increases as we age. Strategies that target oxidative stress and inflammation may improve resilience to processes that lead to cognitive decline. For example, a healthy diet may help combat both oxidative stress and inflammation in the body, but a diet rich in bio-active polyphenolics from fruit, vegetables, walnuts, and coffee may be especially important in improving resilience and health outcomes. Polyphenols have antioxidant and anti-inflammatory activities, so consuming them could slow or prevent age-related changes.

As previously shown, foods high in polyphenols, e.g., dark-colored berry fruits, prevent age-related neuronal and behavioral deficits in animal models of aging. In particular, studies from animal models of aging have found that polyphenolic compounds from walnuts and berries hold promise in slowing—and perhaps even reversing—age-related motor and cognitive declines. These polyphenolics possess antioxidant and anti-inflammatory properties and may also influence the brain directly through various mechanisms, including altered cell signaling and increased neurogenesis, arborization of dendrites, and autophagy in the brain. In recent randomized, double-blind, placebo-controlled pilot studies in healthy older adults , blueberry or strawberry supplementation was able to improve some aspects of cognitive performance, but not gait or balance. In a randomized, double-blind, placebo-controlled trial in 44 healthy older adults , supplementation with freeze-dried blueberry powder for 3 months improved 1 measure of executive function and 1 measure of learning and memory. In a similarly designed trial, supplementation of freeze-dried strawberry powder in 39 healthy older adults for 3 months improved 2 measures of learning and memory compared with placebo but had no effect on executive function. Both trials found that berry powder supplementation did not affect mobility, including measures of balance and gait, likely because the study subjects had no mobility detriments at baseline. Berry supplementation did not decrease serum levels of inflammatory biomarkers compared with placebo, but when serum from berry-supplemented subjects was applied directly to cultured microglia cells, there was a reduction in LPS-induced inflammatory markers relative to placebo-treated subjects. Interestingly, the serum was protective when taken during fasting as well as post prandially. Although these studies are preliminary, they add to the evidence that berry supplementation may help protect against age-related cognitive declines. In addition to single nutrients, healthy dietary patterns have been shown to slow the rate of cognitive decline. In particular, the Mediterranean-DASH diet intervention for neurodegenerative delay diet, which highlights increased intake of plant-based foods, such as berries and green leafy vegetables, is associated with lower risk of cognitive impairment in older adults.

Further investigations examined mechanisms and other factors involved in the beneficial effects of berry fruits. For example, changes in circulating levels of specific phenolic compounds were correlated with changes in cognition. Furthermore, cognitive performance and inflammation were related, as serum collected from berry-supplemented animals reduced LPS-induced inflammatory-stress-mediated signals in stressed highly aggressively proliferating immortalized microglia in vitro relative to serum from placebo-fed controls, and nitrite levels following supplementation were positively correlated with cognitive performance. Therefore, the inclusion of additional servings of polyphenolic-rich foods, such as nuts and berries, in the diet may be one strategy to forestall age-related neuronal deficits, perhaps via decreases in inflammation and suppression of microglial activation, to help increase cognitive resilience and preserve cognitive function. Other non-nutritive natural compounds derived from plants should also be considered as bio-active compounds that contribute to optimal health and improving resilience. The BENFRA Botanical Dietary Supplements Research Center at Oregon Health & Science University studies Botanicals Enhancing Neurological and Functional Resilience in Aging. Two botanicals of interest are Centella asiatica and Withania somnifera . The Center has considerable experience with Centella asiatica, which is used in Ayurvedic medicine to improve memory. It is a popular dietary supplement for “brain health” and shows potential to be developed as a FDA approved “botanical drug” for the treatment of Alzheimer’s disease. The rational use of botanicals, whether as dietary supplements or botanical drugs, requires their evaluation through optimized clinical trials. These trials must be based on sound preclinical studies providing evidence for functional effects, mechanisms of action, and active compounds. The use of preclinical models is critical to inform the optimal design and implementation of future nutrient or botanical clinical intervention trials in healthy older adults and in patients with neurologic diseases, such as Alzheimer’s disease. However, due to the limitations of preclinical models in representing human health, disease, and responses, evaluation of the efficacy of an intervention through clinical trials in humans is essential. Research needs to focus on product authentication, identification of the biologically active compounds and their mechanisms of action, and detection of relevant biomarkers that translate to humans. Preclinical studies also need to address efficacy and safety of the botanical to advance translational research in cognitive resilience. Preclinical studies at Oregon Health & Science University have confirmed the cognitive effects of Centella asiatica in aged mice and that the antioxidant response gene Nrf2 is a molecular target of this herb. Triterpenes and caffeoylquinic acids have been identified as active compounds in C. asiatica and may account for its neuroprotection.

A phase I clinical trial examining the pharmacokinetics of Centella asiatica compounds in older adults with mild cognitive impairment was recently published. A recently initiated clinical trial will examine safety of C. asiatica and also characterize the biologic signatures of its cognitive effects in a population of cognitively impaired older adults. In the case of Ashwagandha, work at the BENFRA Center has focused on water and hydroethanolic extracts of the root,dutch bucket hydroponic as these preparations are commonly used in dietary supplements and in previously reported scientific studies. In one study, the effects of aqueous and hydroethanolic extracts of Ashwagandha root were compared in Drosophila melanogaster models of sleep, cognition, locomotion , and stress-induced depression. Treatment with the hydroethanolic extract improved age-related sleep fragmentation in male flies. Surprisingly, Ashwagandha root aqueous extract showed stronger effects than the hydroethanolic extract in Drosophila melanogaster models of cognition and locomotion and a model of stress-induced depression. Treatment with the aqueous extract of W. somnifera improved age-related locomotor declines in females at lower doses than the hydroethanolic extract. The aqueous extract also provided some resilience against stress-induced depression both when given prophylactically and continuously in a Drosophila model of depression. By contrast, the hydroethanolic extract was only effective when given continuously. The withanolides, commonly regarded as Ashwagandha’s active compounds, are present in greater amounts in the hydroethanolic than aqueous extracts. Together, this suggests that different Ashwangandha compounds may modulate the botanical’s effects on cognition, mood, and sleep and that compounds other than the well-known with anolides may be involved in some of its biologic effects. Studies are underway to explore these unknown active compounds in resilience to age-related cognitive decline and stress. Knowledge of the bioactive compounds associated with each potential clinical use of Ashwangandha will be important in optimizing products for clinical trials of Ashwangandha for those conditions. In summary, as we pivot to emphasize the promotion of optimal health, we need alternative indices of health besides disease outcomes. An individual’s ability to be resilient, including the ability to respond to stressors and to thrive and retain functionality while maintaining a high quality of life, should be considered. Moreover, both essential nutrients as well as other nonessential bioactive compounds should be considered as key factors that promote optimal health.As we define optimal health, it is clear that the potential solutions will vary depending on many individual and environmental factors. Nutritional interventions in healthy adults are known to produce a variety of responses, and work is underway to identify and characterize the different phenotypes that result in unique metabolic needs, with the goal to design personalized dietary approaches to maximize individual health. Although reasonable skepticism regarding the consumer readiness for precision and personalized nutrition exists, efforts to better illuminate the goals and challenges to this emerging technology are being openly discussed in the research community. For instance, in August of 2021, the National Academy of Science, Engineering, and Medicine held a public workshop titled “Challenges and Opportunities for Precision and Personalized Nutrition”. At this workshop, participants raised important perspectives on current opportunities and information gaps in our understanding and approaches to variability in nutritional responses, the shift in the personalized nutrition industry, and numerous studies demonstrating the potential utility for research in this area to aid in our understanding of variable nutritional responses as well as how in certain circumstances they may be beneficial in tooling both dietary guidance for glucose control and therapeutic interventions for weight loss. Addressing these knowledge gaps and refining our expectations and applications for individualized nutrition will be critical to realizing the full potential of this knowledge as another tool in our arsenal to improve human health. In 2020, the NIH also launched a new strategic plan for nutrition that emphasizes Precision Nutrition. In 2022, the NIH invested $170M in a new Precision Health program that leverages the All of Us research program and will allow unprecedented opportunities to combine metabolism, microbiome, diet assessment methods, and data sciences together to provide new insights in precision nutrition. Instead of a one-size-fits-all, we can envision a time where a person’s unique characteristics will be effectively used in a proactive approach to health promotion and disease prevention, and importantly, allowing personalized strategies based on these characteristics.

The family Betaflexiviridae comprises twelve different genera , including Vitivirus, Trichovirus and Foveavirus. After in-silico and in-vitro analyses of trichoviruses and foveaviruses, we did not find evidence for cross-reaction by the universal assay. A single test for all known grapevine vitiviruses can be a useful tool for improving efficiency and reducing costs of large-scale surveys. Potentially, this generic assay may detect novel Vitivirus species in grapevine and other hosts given its unbiased nature. Similar assays have been developed for carlaviruses, nepoviruses and different members of the family Betaflexiviridae. Grapevine is clonally propagated, consequently, to prevent the spread of vitiviruses, it is critical to use virus tested material. The assay developed here will be made available to diagnostic labs and will facilitate the production of certified virus-tested propagation material and the effective control of vitiviruses.As people age, their risk for cognitive decline increases. An estimated 40% of people 65 years and older have age-associated memory impairment characterized by self perception of memory loss and a standardized memory test score demonstrating lower objective memory performance compared with young adults. While genetic factors play an important role in age-related memory decline, such nongenetic lifestyle factors as diet and exercise contribute as well. In particular, a growing body of evidence indicates that accumulation of oxidative damage to macromolecules increases progressively during the aging process and contributes to neurodegeneration in Alzheimer’s disease. Epidemiological studies have suggested a link between antioxidant consumption and cognitive protection,blueberry packing boxes and studies using a rodent model of Alzheimer’s disease suggest that tau phosphorylation occurs as a compensatory response to oxidative stress. Several clinical trials of antioxidant use in subjects with normal aging or Alzheimer’s disease suggest memory benefits, while others yielded negative results.

While many fruits are rich in various antioxidants, including ascorbic acid, carotenoids, and phenolics, commonly consumed fruits show large differences in antioxidant capacity, as determined by the ferric reducing/antioxidant power assay. In particular, there is special interest in the pomegranate polyphenols, which have been studied extensively in animal models of Alzheimer’s disease. The emerging data on pomegranate fruits and their inherent polyphenols suggest positive benefits ranging from neuroprotective effects to staving off effects of senescent neurodegeneration in animal models of Alzheimer’s disease. Previously, we found that pomegranate juice has the highest antioxidant capacity among fruit juices using five different in vitro antioxidant assays. In comparison, apple juice, which is low in antioxidant capacity, was less effective than pomegranate juice in affecting antioxidant capacity based on FRAP assays in 26 elderly subjects consuming pomegranate juice or apple juice over a 4-week period. Over expression of antioxidant enzymes or supplementation of some antioxidants appears effective in extending the life span in several animal models, and a few studies have specifically found memory-enhancing effects of polyphenols in mice . However, evidence for memory enhancement from polyphenols in human-controlled trials is limited. One preliminary study found beneficial effects of grape juice on memory in five older adults with ageassociated memory impairment compared to seven placebo controls; to our knowledge this small study is the only placebo-controlled human trial of polyphenols that assessed memory change. Therefore, more well-controlled studies are needed to determine potential beneficial effects of antioxidants that occur naturally in various food sources and to elucidate the possible underlying mechanisms. Moreover, studies that examine possible brain mechanisms of polyphenol treatment in humans are limited. One potential method for identifying mechanisms of recovery in drug trials is functional magnetic resonance imaging .

While structural MRI studies measure long-term changes in metrics, such as cortical thickness and volume , fMRI measures blood flow changes in response to memory challenge, which has proven to be effective in determining brain differences among subjects with Alzheimer’s disease, mild cognitive impairment, or non-demented subjects at risk for Alzheimer’s disease. Further, fMRI has proven to be sensitive in measuring changes in neural activation patterns among older subjects in response to memory-enhancing drugs, such as donepezil and memantine, with results showing increased activation or restoration of resting state brain networks following treatment. One investigation used fMRI to examine the effects of antioxidants in young healthy controls and demonstrated increased fMRI activity to cognitive challenge with flavonoid administration; however, no studies to date have examined fMRI effects of antioxidant therapy in older adults with or without memory complaints. To address this knowledge gap, we explored potential neuroprotective effects derived from polyphenols in pomegranate juice in older volunteers with mild memory complaints in a placebo-controlled, randomized, and double blind trial and measured three aspects: metabolites of pomegranate juice using blood biomarkers; effects of pomegranate juice on memory performance; and evidence of functional MRI changes during memory activation. We hypothesized that pomegranate juice compared to placebo would increase metabolites, improve memory, and increase task-related brain activation in the left and right hemispheres for verbal and nonverbal memory tasks, respectively. Together these metrics provide preliminary evidence of bio-availability, clinical efficacy, and brain mechanisms of pomegranate juice in older adults with memory complaints.Thirty-two volunteers were initially enrolled into the study out of 39 who were screened; of these, 28 completed the clinical trial . Subjects were recruited via advertisements through local newspapers, flyers, posters, lectures, and word-of-mouth. We specifically recruited nondemented, older right-handed subjects with self-reported age-related memory complaints. The subjects did not carry a diagnosis of mild cognitive impairment and were screened using the mini-mental state examination . Subjects randomized to placebo versus control groups did not differ in mean age, percentage of women, mean MMSE scores, or baseline memory performance .

Potential subjects were excluded who had documented neurological, psychiatric, or major medical conditions, including heart disease, diabetes, and gastrointestinal disease, that might affect cognitive function or interfere with study procedures. We also excluded subjects who routinely ate more than three servings per day of fruits and vegetables or were taking vitamin supplements, any medication, or dietary supplement that interferes with the absorption of polyphenols. Potential subjects participating in regular vigorous exercises other than ordinary daily walks or who were unwilling to maintain a sedentary lifestyle during the 28-day study were excluded, as were those for whom MRI was contraindicated . Seven potential subjects were screened out based on one or more of these exclusion criteria. Subjects were instructed that they would be removed from the study if they failed to follow the prescribed low polyphenol diet during the one-week run-in period based on self-report in-person interview with a study dietitian. The study was approved by the UCLA Institutional Review Board; all subjects gave informed consent to participate in all procedures and were reconsented for followup evaluations. Funding for this study was provided by the UCLA Center for Human Nutrition from a “various donors” account. The investigators were blind to the contributors to this fund, and there was no communication or input from donors or commercial entities at any stage of the study.Following phone screening, volunteers participated in an initial clinical evaluation,package of blueberries which included an explanation of the study procedures, consent, medical screening, MMSE, and routine CBC and chemistry panel. Prior to the intervention phase, each subject met with a registered dietitian and was instructed on a low polyphenol diet, which required that they restrict their intake of several fruits and vegetables, onions, tea, chocolate, and dried beans, for one week prior to the baseline visit and for the duration for the study. At baseline, subjects underwent neuroimaging and neurocognitive testing . We also acquired blood samples to perform the antioxidant assay and to verify compliance with the protocol. Subjects were then randomized to either the pomegranate or placebo groups; investigators and subjects were blind to group membership. After these procedures, subjects were given a one-week supply of 8-ounce containers of pomegranate or placebo juice with instructions to consume 8 ounces of juice daily. The pomegranate drink was the commercial Pom Wonderful product. The flavor-matched drink contained sugar, citric acid, and food color to match the calorie, taste, and color of the juice. Each week, subjects returned their empty juice containers to ensure compliance, picked up their juice supply for the following week, and met briefly with the dietitian to ensure compliance with the low polyphenol diet. At the end of the study , subjects again underwent neuroimaging, cognitive assessment, and blood draws. A placebo drink using sugars and natural colorings and flavoring to imitate the taste and appearance of pomegranate juice was used. The research personnel were blinded as to the assignment of subjects to placebo or pomegranate juice.Each subject underwent fMRI and brief memory testing immediately prior to beginning the trial and following 28 days of pomegranate juice or placebo. Subjects received a memory testing battery and underwent functional MRI scanning on the same day. Memory tests administered were the Buschke-Fuld selective reminding task, from which we calculated two performance measures, total items recalled and consistent long term retrieval; an experimental unrelated word pair associates learning task, based on the Wechsler memory scale paired associates learning in which we used identical procedures to that used in the fMRI scan but with different stimuli. We also obtained behavioral data from the MRI tasks administered in the scanner .

Of the 28 subjects completing the study, 26 had behavioral data on both occasions . Subjects were administered one of two alternate forms of each task, with the order of forms counterbalanced across subjects, to avoid practice effects. To assess changes in verbal memory, we compared performance on total number of items recalled on the Buschke-Fuld selective reminding task and on the consistent long term retrieval score . The TR score emphasizes immediate recall, while CLTR depends upon memory consolidation processes. Because memory performance in this age range is highly variable, we used time 2 minus time 1 difference scores as our dependent variables. At baseline, there were no differences between groups on either TR performance or CLTR . The spatial navigation task involved learning and recalling routes on a virtual map, while subjects play the role of a “taxi driver.” During the encoding phase, subjects watched and rewatched two sets of video clips. In each video, subjects first saw the name of the imaginary town, such as “Johnsberg,” then for 15 second blocks they saw a first-person view of “driving” to a virtual town laid out in 5×5 block grids, to a storefront destination with unique characteristics for specific storefronts, stopping for 3 seconds before proceeding to a new store within the same town starting from a different location. After each city, subjects performed a 30-second distractor task where they pressed a left or right button in response to a probe. This was repeated while subjects learned 6 cities in total. The fMRI data were then collected during memory retrieval in three conditions. Subjects were asked to remember one of three characteristics about each storefront for the separate tasks. Two conditions focused on the physical features of the environment . In condition 1, participants were asked to identify an appearance change of the store , such as signs, colors, and doors. In condition 2, they were asked to identify the spatial location change of the store within the town, relative to other buildings , respectively. Condition 3 required subjects to identify the store for a specific item that might be purchase there, requiring the participant to encode the association between an item and a spatial location. For this source task, subjects had to recall whether the store was in the first or second town . Only the latter condition tapped into the associative memory processes. The order of the three conditions was counterbalanced across subjects.All functional data were analyzed using FSL version 4.1.4 . Preprocessing included motion correction to the mean image, spatial smoothing , and high-pass temporal filtering .Twenty three subjects were able to complete the visual memory fMRI task without significant head motion, while for the verbal memory task, eight pomegranate juice and nine placebo subjects were able to complete both scanning sessions with high quality images. Each functional scan was registered to its corresponding coplanar high-resolution image with rigid body transformations and to the MNI152 standard brain using nonlinear transformation . Additionally, a high-pass temporal filtering of 100 sec was applied to the functional images. Individual preprocessing and statistical analysis of each subject’s functional scan was run using FSL FMRI Expert Analysis Tool ; FEAT was then used to analyze data at the group level.