Results from the pseudo mixed model indicated the interaction effect was more important than the treatment effect. Thus, “alcohol hotness” will not be included in any further discussion of significant attributes for BA wines. The significant difference in malic acid content in the wines among treatments appears to have had little impact on sensory evaluation given that there was no significant difference in the perception of sourness in the wines. From the PCA generated from BA descriptive analysis results , the control and sort wines appear to be correlated more closely with “alcohol” . Wines made from these treatments were higher in ethanol content, which may explain this trend. However, the small number of significant attributes indicate that BA wines made by different treatments were very similar in sensory properties.Analysis of wine color revealed that there were perceivable differences among treatments for all three varieties . For BA the reject treatments were rated lighter in color compared to the control and sort treatments, whereas a similar trend was observed in the CS treatments. This was expected because berries with less color were removed by the optical sorter and included in the reject fermentations. This agrees with results from Table 6; the rejected treatments were significantly lower in anthocyanin content for BA and CS, which can explain the difference in color perception. For GN wines, the control treatment was perceived to be slightly darker than the sort and reject treatments. Although fermentations were prepared to have similar solid-to-juice ratios in the must among treatments, grow bag for tomato it is possible that variations between replicates may have resulted in the control treatments being slightly more concentrated, which could provide an explanation for this result.

It can be concluded that optical sorting was generally successful in removing berries with less color; however, this did not lead to a large difference in the final color of the wines between the sort and control treatments.Multiple Factor Analysis was performed for each variety using all sensory attributes and only volatile compounds that differed significantly among treatments . This was done to observe the association, if any, of the significant volatile compounds and sensory attributes. For GN wines, the only significant attribute was “SO2”. From Figure 7, isobutanol, which can impart a solvent like aroma in wine, is grouped closely with “SO2”. It is possible that wines with a higher isobutanol concentration were perceived to be higher in “SO2” aroma. For BA wines, there does not appear to be a trend among sensory attributes and volatile compounds . For CS wines, “apple” is grouped closely with ethyl esters , which provides evidence that this may have caused the increased perception of this attribute in the control and sort treatments .Overall, optical sorting had minimal impact on the sensory properties of the three varieties tested. It is possible that the chemical differences noted earlier were too small to result in consistent differences by descriptive analysis. Even though the wines made from reject material contained significantly higher concentrations of higher alcohols, it did not result in a difference in sensory perception. Higher alcohols have a relatively high sensory threshold . It is possible that the concentration of these compounds in the reject wines was below the sensory threshold.The purpose of this study was to determine what effects, if any, optical berry sorting had on wine made from different red grape varieties, and to investigate the potential to use optical sorters to sort for different ripeness levels using color as a main criterion.

Given the observed differences in Brix and final ethanol content, optical sorting seemed to be successful in removing underripe berries for CS and possibly for BA; however, this did not result in a significant difference in the final ethanol content between the sort and control treatments. The removal of underripe berries was also evident by the difference in color among treatments. For BA, the rejected treatments were significantly lighter in color; however, the color of the sort and control treatments was very similar, whereas a similar trend was observed in the CS treatments. Wines made from GN generally did not follow these trends; possibly because sorting parameters were too aggressive for this cultivar, resulting in a high percent rejection of optimal berries. This may have minimized potential differences between reject wine with the other treatments. Another possibility is that color differences in the GN fruit did not correspond to differences in sugar content. From these results, it may be concluded that, when using color as a criterion, optical sorting based on ripeness level was successful but may be dependent on variety and fruit variability. Additionally, the impact on the resulting wine is likely dependent on the initial variability in grape ripeness. The optical sorter was successful in removing MOG. This result was reflected in the phenolic analyses; reject treatments were generally higher in total phenolics and tannin, most likely due to the greater proportion of MOG included in the must. The decrease in anthocyanins is likely due to the higher percentage of green, underripe berries in the reject treatment musts. A study that made wine with the addition of MOG found that this addition significantly increased the phenolic and tannin content in the resulting wines. Despite the differences observed in the phenolic composition of the reject wines, the control and sort treatments were very similar for all three varieties. This is in contrast with some previous studies that have found wine made from optical sorted fruit had significantly different levels of phenolics.

One study found that optical sorting led to wines with higher levels of total phenolics. It should be mentioned that the researchers here did whole cluster pressing for their control wines , whereas the sorted wines were destemmed. It is possible that higher levels of phenolics were extracted due to the damage caused by the destemming process on the seeds and skins. Another study found that wine made from optically sorted grapes that were machine harvested generally had lower levels of phenolics; levels that were similar to the same wines made from a handpick treatment. Given that the rejects were, in general, significantly higher in total phenolics and tannin than the control and sort treatments, it can be suggested that optical sorting has the potential to decrease the phenolic content in wine; however, there was not enough MOG to show a large impact in the current study. Optical sorting likely has a greater impact on mechanically harvest fruit due to generally higher levels of MOG observed from this harvest method. Some differences were found among treatments in the aroma profiles of the wines. Few compounds differed significantly between sort and control treatment and, in general, the reject treatments had greater concentrations of higher alcohols and control and sort treatments had greater concentrations of ethyl esters. The higher ethanol content of the sort and control treatments as well as their lower pH can lead to a higher production of esters. In general, reject treatments contained significantly more suspended solids then the control and sort treatments for all varieties studied. Research has shown that high levels of suspended solids during fermentation can lead to greater production of higher alcohols. Descriptive analysis indicated only one significantly different attribute among GN treatments and only two significantly different attributes among BA treatments. BA control and sort wines were associated with the “alcohol” descriptor which correlated with the higher ethanol levels in these treatments compared to the reject treatment. Similarly, there were only three significant attributes among the CS treatments. “Alcohol hotness” related to ethanol content as previously described. The control and sort treatments were also rated significantly higher in “apple” and “sweet” aromas compared to the reject treatment. Some studies have shown that higher levels of ethanol can increase the perception of sweetness in a wine. However, as King et al. noted, grow bag for blueberry plants there is disagreement in this regard, as other studies have shown that ethanol content can either decrease or have no effect on the perception of sweetness. Thus, this may not be a sufficient explanation as to why the control and sort wines were rated significantly higher in sweetness. Perhaps the higher concentration of total phenolics and tannin in reject wines could explain the difference given that phenolics in wine contribute to bitterness and astringency. From the PCA in Figure 6, it can be noted that “bitter” and “drying” are more associated with reject wines. Although these attributes are not significantly different among the treatments there appears to be a trend which could impact the perception of sweetness.

One study found that increasing bitterness in coffee decreased the perception of sweetness. It is possible that reject wines were rated lower in “sweet” due to the higher concentration of phenolic compounds thus decreasing the perception of sweetness. The higher perception of sweetness in the control and sort wines may also be attributed to the higher intensity of the “apple” aroma, which the judges could have associated with a sweet taste. One study found that retronasal aromaperception of fruity compounds increased with an increasing level of sweetness in a model wine solution. The authors also noted several other studies which found that aroma compounds can enhance the perception of sweetness in different foods and beverages. Another study found that samples described as “fruity” were also often associated with a “sweet” aroma. This provides further evidence that the judges in the current study may have associated these attributes together. The overall sensory differences were minimal, and the wines were determined to be similar. The results from this study largely agree with results from previous studies investigating the effects of optical sorters. It is possible that there was not enough variation in the starting material of the current study for optical sorting to have a large impact. Optical sorters may be used to greater effect during vintages with inconsistent ripening, issues with raisining, or large amounts of berry damage, possibly caused by either birds and/or fungal infections. Future research should investigate the impact of optical sorters in these scenarios.Grapevine has indeterminate growth habits compared to other perennial fruit crops. Latent growth of the dormant grapevine bud may be induced by favorable conditions with little to no dormancy period required . Therefore, semi-tropical regions may raise two crops a year, and in fact, it is not uncommon for the latent bud to produce some fruit when correlative inhibition is removed in temperate regions. Furthermore, the grape berry does not have the same fruit abscission mechanism as apple or peach revealed under carbon starvation. It is therefore possible for grapevine canopy size and crop level manipulations leading to a wider range of source or sink limiting conditions within a growing season. The crop level of a perennial crop is initially determined by organogenesis at the basal buds. The number and size of the flower primordia is associated with number of clusters and berries per cluster through the formation flowers and fruit set . However, fruit set is largely variable among years, weather, location, and cultivars . Poor fruit set may be a limitation to crop yield, although weather is often considered to be the leading cause. However, the mechanism of poor fruit set is not fully understood. Carbon supply or mineral nutrition are related to the amount of fruit set , which is an acclimation mechanism to unfavorable conditions. Ultimately, yield of grapevine is affected by berry size, and within the berry, pulp enlargement is the largest contributor to yield gain rather than skin or seed biomass . Conversely, vegetative growth is far less influenced by latent bud formation, as competition amongst growing buds tends to buffer the impact of growing shoot tips on its length and total leaf area . This is likely due to the great limiting effect of nitrogen among other nutrients or hydraulic pressure . The ratio between leaf area and fruit mass is closely related to the amount of carbohydrates accumulated in the must . Thus, an excessive crop level or less than ideal canopy size may result in over cropping and may lead to delayed ripening . Conversely, in under cropping, where there is excessive vigor or reduced crop level, this is not necessarily deleterious for speed of ripening . However, it may be a wasteful management of resources if there is not a trade-off with farm-gate prices.

While ant activity only significantly increased after string placement on connected coffee plants, we also observed lesser increases in ant activity on control coffee plants and nest trees . This unexpected result could mean that strings, a novel element in the environment, acted as a form of habitat modification or disturbance, which increased overall ant activity in the local area. However, if our manipulation was the cause, we would have expected the ants to attack the jute strings , a behavior that we did not observe during the experiment. Experiments in tropical forests have shown that the long-term removal of lianas can influence ant richness on trees , and therefore may possibly also affect overall ant abundance and activity when promoted. It is also possible that other factors could potentially explain this result in control plants, such as changes in local abiotic factors that we did not measure systematically in our experiment. Future research which expands on the temporal scope of this study may be useful in assessing the long-term effects of artificial connectivity in this system. Ant activity after string placement was negatively affected by distance to the nesting tree . This result is consistent with previous studies suggesting that within 5 m A. sericeasur dominance in the leaf litter decreases with distance to the nesting tree . However, in our study, grow bag the effect of distance after string placement was significant only on control plants, but not on connected plants. This suggests that connections could buffer the negative effects that larger distances from the nesting tree pose to ant activity and potentially increase antprovided biological control services in these plants. Connected coffee plants also had significantly higher CBB removal than control plants .

Overall, greater ant activity on coffee plants was associated with higher CBB removal rates , suggesting that ant activity directly influenced CBB removal rates. However, while this effect was significant on control coffee plants, it was only marginally significant on connected plants. While we believe that these results support the hypothesis that connectivity enhances ant foraging and bio-control services on coffee, the use of dead CBB in this experiment as a proxy to measure bio-control may explain the only marginally significant effect of ant activity on CBB removal in connected plants. It is possible that dead prey exhibit more variable recruitment responses from ants than live prey. Despite this, it is likely that strings facilitated ant movement to coffee plants by providing a smooth, linear substrate and indirectly increased CBB removal . In other systems, the leaf-cutting ant Atta cephalotes uses fallen branches to rapidly move between areas and thereby quickly discover new food resources . Similarly, these resources allow scouts to return quickly to the colony, minimizing the time taken for information transfer and recruitment of other foraging workers . The role of trunk trails and fallen branches has received extensive attention in the leaf-cutting ant system; however, fewer studies have looked at the influence of connectivity resources on foraging behavior of predatory arboreal ants. Surprisingly, CBB removal did not follow the same trend as ant activity with distance to the nesting tree. While control plants tended to have lower CBB removal rates than connected plants as distance to the tree increased, we did not find a significant effect of distance on CBB removal in either control or connected plant groups. Collectively, these results suggest that connections in the arboreal stratum have the potential to increase ant activity and therefore enhance plant protection from CBB attack, particularly in connected plants.

Further studies should assess the effect of distance on CBB removal using plants located at distances larger than 3.5 m from the tree. It is important to note that enhanced ant activity on coffee plants could lead to increases in the density of ant-tended hemipterans, such as the green coffee scale, which if severe enough could reduce the productivity of coffee plants. However, the green coffee scale is not a major pest in the region of study, in contrast to the economically significant coffee berry borer . Furthermore, a recent study evaluating the benefits associated with the indirect Azteca–Coffea mutualism found that the protective benefit ants provide to coffee plants is positively associated with high densities of the scale . This suggests that the enhanced CBB control by ants outweighs the costs associated with scale damage. However, these interactions may be context-dependent and still need to be fully evaluated in the field to provide a holistic understanding of the impact of connectivity on scale density and coffee yield. Other ant species could also benefit from the addition of connections between coffee plants and shade trees, such as C. basalis and P. simplex, which were observed using these connections during our study. The ant P. simplex has been previously reported as an important CBB bio-control agent, acting in conjunction with other species of ants to effectively suppress CBB at various life stages . Therefore, this technique could support Azteca ants as well as other ant species that play an important role in suppressing CBB populations. Our results support the general hypothesis that connectivity, one measure of habitat complexity, can sustain important ecological processes in natural and managed ecosystems. In aquatic systems, more complex habitats with macrophytes allow for greater food capture and maintain higher levels of diversity . In terrestrial systems, higher complexity can influence trophic dynamics . In coffee agroecosystems, ants are highly sensitive to habitat change and management intensification, generally expressed as the reduction of shade, elimination of epiphytes, and use of chemical inputs . Such intensification can have a negative effect not only on vegetation connectivity and ant foraging, but may also cascade to affect ecosystem services, such as biological control.

Our study supports the idea that promoting complexity at a local scale, in this case providing structural resources for ants in agroecosystems, can significantly enhance connectivity within the arboreal strata, and potentially improve biological control of coffee pests. This idea has already been successfully implemented in other agricultural systems, placing “ant bridges” made of bamboo strips or strings connecting neighboring trees in , and could be incorporated as a management strategy in coffee systems. Future research should evaluate the practical feasibility of adding connections between vegetation strata to enhance bio-control. For example, studies in timber plantations have estimated that the presence of ants increases timber production by 40%, and that ants can be maintained at lower costs by providing intra-colony host tree connections using rope, poles or lianas . It is important that future studies in coffee also consider the costs of other CBB control methods, such as the application of the pesticide endosulfan, which can lead to the development of resistance, can negatively impact natural enemies, and can have harmful impacts on human health . Further investigation into promoting ant bio-control with artificial connections in coffee should: assess economic trade-offs, management applicability, and farmers’ perceptions of this method in large and small coffee plantations, compare the cost between string placement and other management approaches , grow bag gardening and assess coffee yields on connected and not connected plants to provide management recommendations. More broadly, incorporating conservation bio-control strategies in combination with vegetation connectivity is consistent with criteria identified as key for the sustainability of biological control, such as increasing local habitat quality and enhancing species’ dispersal ability . Generally, the maintenance of shade trees and natural vegetation in agroforestry systems may increase vegetation complexity and natural connectivity between plants to promote ant foraging and subsequent biological pest control.The HPA axis maintains a diurnal rhythm marked by a daily peak after waking, a subsequent decline over the course of the day, and a nadir shortly after onset of continuous sleep . The diurnal pattern of HPA activity plays important roles in a variety of metabolic, immunological, and psychological processes that support our day-to-day functioning . In studies of children, the preferred assessment method of HPA axis activity is the collection of saliva and the measurement of cortisol . Cortisol is the “end-product” hormone released into the bloodstream from the adrenal glands—the final step in a biological cascade initiated by the hypothalamus and perpetuated by the pituitary gland. In addition to supporting the orchestration of several other processes , moderate cortisol levels are thought to support effective neural transmission and optimal learning and high-order cognition . In times of acute physiological or psychological stress, the HPA axis mounts a particularly pronounced response, culminating in high levels of cortisol that reach glucocorticoid receptors throughout the body and brain. Working with the ANS, these acute HPA stress responses coordinate the physiological and psychological resources needed to overcome the stressor . Yet, given negative feedback processes, high cortisol levels also play important regulatory roles in down-regulating HPA axis activation, allowing it and other systems to return to baseline . Collectively, these complex within- and cross-system dynamics support an organism’s ability to both respond to and recover from the effects of environmental stressors . HPA axis reactivity and regulation are evident very early in life. Newborn infants can mount an HPA axis response to environmental stimuli , and normative circadian rhythms tend to stabilize as infants begin to forego their afternoon naps .

However, the span from infancy through early childhood is also a time of meaningful developmental change. Indeed, a growing theoretical and empirical literature indicates that children’s early experiences play a critical role in the organization of their emerging adrenocortical systems .Low-income ecologies present a confluence of distal and proximal risk factors thought to influence children’s developing physiological stress systems and undermine optimal cognitive and social development . For example, children growing up in low-income contexts are more likely to face distal stressors, such as inhospitable and dangerous neighborhoods and inadequate access to services and social capital . Such distal risks are known to have trickle-down effects that undermine parents’ abilities to effectively read, interpret, and respond to their children’s needs . In turn, a convergent literature comprising experimental work with animals as well as observational studies of young children indicates that sensitive and responsive caregiving can support adaptive HPA axis functioning . This is evident with respect to children’s acute stress responses. For example, young children with secure attachment relationships and more sensitive caregivers tend to show better regulated HPA axis responses when faced with acute psychological stressors . Changes in the quality of children’s caregiving environments have also been linked with their baseline, or resting levels of HPA axis activity. For example, at the more extreme end, children who are moved from very high-risk households into foster care have been found to evince comparatively lower resting cortisol levels than their peers who remain in high-risk homes . For instance, in prior work with the same sample as used in the present study, our group showed that higher levels of maternal sensitivity in infancy are predictive lower levels of resting cortisol, after adjusting for income and a number of potential confounds . Beyond psychosocial risks, children growing in the context of economic adversity are more apt be exposed to households that are more densely populated, noisy, disorganized, and unpredictable—aspects typically discussed under the umbrella term chaos . A growing literature suggests that chaotic environments may alter children’s ANS and HPA axis functioning in early and middle childhood. Recent work by researchers in our laboratory suggests similar effects with respect to infants and toddlers , with within-child increases in chaos predictive of contemporaneous increases in resting salivary cortisol in later infancy and toddlerhood.Notably, young children growing up in low-income contexts spend substantial amounts of time in settings outside of their homes—such as non-parental child care. Indeed, in the United States approximately 43% of children in poverty attend regular non-parental care by 9 months of age . A well-developed literature indicates that young children’s early child care experiences also play a meaningful role in their HPA axis functioning. Meta-analytic findings indicate that—compared to their normal diurnal patterns experienced at home—children tend to show cortisol increases across the day on days when they attend child care . Some work suggests these patterns are particularly strong in toddlerhood and the beginning of the early childhood years and for children who attend lower quality child care . There is also some, albeit limited, evidence of long-term effects; for example, Roisman and colleagues found that spending greater proportions of time in center-based care in infancy and early childhood was predictive of children’s subsequent cortisol awakening response in adolescence.

Consistent with studies affirming the influence of vegetation connectivity on predatory arthropod movement and predation range, our results illustrate how vegetation connectivity facilitates A. sericeasur foraging mobility and pest removal. In coffee systems, higher degrees of vegetation connectivity are associated with shade trees, as well as more heterogeneous habitat complexity and variability in plant structure. In other studies, ants generally increase predation services in shaded systems as compared to monocultures and, in coffee plants, more effectively remove CBB in shaded coffee systems as compared to sun monoculture systems. Interestingly, most studies find the opposite effect of structural complexity on parasitoid behavior, with higher degrees of plant structural complexity leading to decreased parasitoid foraging efficiency. This negative relationship between parasitism and habitat complexity transfers to coffee systems, where the parasitic phorid flies exert a greater inhibiting effect on Azteca ants in simple, low-shade farms than in complex, high-shade farms. Together with the aforementioned study, our combined results illustrate how habitat complexity at the landscape scale and vegetation connectivity at the plot scale dually facilitate A. sericeasur-mediated pest removal: by facilitating ant mobility and by reducing the efficiency of the parasitoid that interferes with their pest removal ability. In order for A. sericeasur to provide ant-mediated pest removal services, coffee agroforests must include enough shade trees to provide sufficient habitats for ant nests. Planting coffee plants close enough to shade trees to allow for direct connectivity and leaving some vegetation connections between coffee plants and shade trees rather than chopping them or relying on herbicides can facilitate ant-provided ecosystem services by providing foraging paths through naturally occurring structural connectivity.

By enhancing the A. sericeasur effectiveness in controlling CBB populations, vegetation connectivity can potentially reduce chemical pesticide use. Our results offer management insight into one piece of a complex ecological puzzle. Because A. sericeasur tend C. viridis, square black flower bucket they could indirectly reduce coffee plant growth by contributing to high-scale densities and an associated damaging sooty mold. However, high densities of C. viridis also beneficially attract Lecanicillium lecanii, which attacks coffee leaf rust, a devastating coffee fungal disease. Moreover, the CBB is regarded as a far more damaging coffee pest than C. viridis. Furthermore, facilitating the mobility of A. sericeasur as a single ant species is not necessarily the most effective pest management approach, as higher ant diversity can improve pest control through the cooperation of complementary predatory species. Enhanced A. sericeasur activity on coffee plants could alter the behavior of other ant species, which could have positive or negative effects on overall pest control services due to spatial complementarity or potential negative interactions between predators. However, studies find that increasing connectivity generally increases species richness, and so, vegetation connections that increase A. sericeasur mobility likely facilitate the mobility of other predatory ants in coffee systems, even by providing alternative paths to avoid aggressive altercations with A. sericeasur. Although A. sericeasur occupies only 3–5% of the shade trees at our research site, other ants known to contribute to CBB regulation would likely also use vegetation pathways, facilitating additional pest control. Future research should examine how vegetation connectivity impacts the abundance and diversity of other ant species on coffee plants and the associated spatial complementarity between specific predators of the CBB. Future studies could also investigate how phorid attacks on Azteca vary on different foraging pathways to better understand the mechanisms behind their preference for vegetation pathways.

Connectivity affects arboreal ant distribution, behavior, and interactions with other organisms in agroecosystems, profoundly impacting ant community diversity and ant provided ecosystem services. Our results demonstrate how vegetation connectivity increases A. sericeasur activity, recruitment to resources, and CBB removal, and that naturally occurring vegetation connectivity, in the form of branches and natural substrates, accounts for this enhancement. As climate change increases coffee’s susceptibility to CBB damage, agroecological and economically feasible forms of pest control are increasingly necessary for coffee-producing communities. Farm management conducive to forest conservation, habitat and structural complexity, and the associated higher degrees of vegetation connectivity will facilitate ant-provided pest control services in coffee agroecosystems.Wild birds provide many ecosystem services that are economically, ecologically, and culturally important to humans . On a global scale, insectivorous birds consume an estimated 400–500 million tons of insects annually and have the capacity to decrease arthropod populations and increase crop yields of both temperate and tropical farms . While these beneficial effects are not always observed , attention has focused on promoting avian diversity and abundance on farms to leverage these benefits . The fact that birds consume agricultural pests does not ensure that they can control them, in the sense of substantially reducing densities of rapidly-growing pests. Here, we evaluate the capacity of birds to suppress agricultural pests, specifically the coffee berry borer, aninvasive pest found in almost every coffee-producing region worldwide. The coffee berry borer is one of the most economically significant pests of coffee worldwide , causing an estimated annual global loss of US $500 million . These small beetles damage coffee crops when a female bores into a coffee cherry and excavates chambers for larvae to grow, consuming the coffee bean. Control of CBB can be accomplished by spraying fungal bioinsecticide Beauvaria bassinia, increasing harvest frequency or continually removing, by hand, over-ripe and fallen cherries, which serve as reservoirs for infestations .

The last, and most laborious, control method appears to be the most economically effective In addition to human-mediated control, natural predators such as ants, parasitoid wasps, and nematodes are being explored as potential bio-control agents . Birds have also been identified as a significant biological control agent of CBB . Field experiments in Central America have shown that CBB infestation dramatically decreases when birds are present . For example, Karp et al. reported that bird predation suppresses CBB infestation by 50% and saves farmers US $75– 310/ha per year; another estimate values bird predation at US $584/ha . Suppression is done by both resident foliage-gleaning insectivores, such as rufous-capped warblers , and Neotropical migrants like the yellow warbler . Similar to other agriculture systems, avian abundance is higher on farms with heterogenous landscapes in close proximity to native habitat , suggesting low-intensity shade coffee farms are better not only for supporting biodiversity, but also in providing pestmediating ecosystem services . Several lines of evidence support the notion that birds depredate CBB in coffee plantations, and that their effects are biologically significant. Firstly, we know that a variety of bird species consume CBB from assays of avian fecal and regurgitant samples , though the detection rate is quite low . Low detection rates might be due to low consumption rates; detectability of DNA in feces depends on number of CBB eaten, and time since feeding, as well as fecal mass . Secondly, bird and bat exclosure experiments are associated with greater CBB infestation within enclosures . At the same time, it is not clear how birds can effectively suppress CBB at most sites, and throughout the season. Exclosure experiments that report avian suppression appear to be at sites with relatively low CBB infestations , whereas coffee-producing regions with more recent introduction of CBB have infestations of up to 500,000 CBB in a season . We also do not know whether suppression is effective throughout the reproductive cycle of the CBB, or just when abundances are relatively low. Finally, CBB field traps often capture large numbers of CBB, even in the presence of birds . Consequently, while there is clear evidence that birds consume CBB, the degree to which CBB populations can be suppressed is less clear, particularly because of the species’ population growth potential . Here, square black flower bucket wholesale we use a CBB population growth model to assess the capacity of birds at naturally occurring densities to reduce CBB populations, as a function of a starting infestation size. We created an age-based population growth model for CBB using data from a life-stage transition matrix published by Mariño et al. . We converted their matrix into a female-only, daily time-step, deterministic Leslie matrix; we could not estimate population growth directly from the original matrix because it did not use a common time step . We incorporated a skewed adult sex ratio to mimic real populations , and added a life-stage for dispersing females, the stage at which CBB are vulnerable to predation by birds. Since the entire CBB lifecycle occurs within the coffee cherry, CBB are vulnerable to predation by birds for a short time window when adult females disperse between plants and burrow into a new cherry .

Birds do not eat coffee cherries, with the exception of the Jacu , which is found in southeastern South America. Consequently, we assumed that only adult CBB females are vulnerable to bird predation. With our Leslie matrix, we projected population growth for a closed population during a single CBB breeding season. We projected growth at three levels of initial starting populations of CBB , calculated from published estimates of CBB densities from alcohol lure traps in coffee farms from Colombia, Hawaii and Costa Rica. We then determined the degree to which dispersing female survival rate would have to be decreased to result in a 50% depression in the adult population size at the end of the coffee season at all three infestation levels. Finally, we assessed the plausibility of this degree of CBB suppression by birds as a function of avian energy requirements, reported avian densities on coffee farms, prey composition of avian diets, estimated caloric value of CBB, and the starting population size of CBB females.Coffee phenology is directly related to rainfall patterns that differ among coffee producing regions, leading to distinct seasons, and timing of harvest. Our model assumes environmental conditions of Costa Rica, and thus describe the coffee phenology of this region. In regions of Costa Rica with marked seasonality, coffee flowering is triggered during the dry to wet season transition by the onset of acute precipitation . Areas with relatively consistent rain patterns have more continuous flowering events and a longer harvest season In the Central Valley of Costa Rica, flowering typically begins in March, with three flowering events spread over a month . Flowers are short-lived, lasting only a few days before fruit begin to develop. Maturation of coffee cherries is slow, with immature green cherries taking up to 240 days to develop into red, ripe fruit that is ready for harvest in mid-October through January . After harvest, coffee plants are left to recuperate until flowering is initiated again the following year by the next onset of rain.Following the coffee flowering period and initiation of cherry growth, adult female CBB emerge and disperse via flight in search of new cherries to colonize . Timing of emergence appears to be driven primarily by relative humidity and temperature, with dispersal peaks occurring around the end of the coffee harvest, from December through March . Females begin ovipositing in chambers carved out of the coffee endosperm roughly 120–150 days after coffee flowering, when the dry content of the seed is 20% or higher . It is this dispersal period, and subsequent drilling into the coffee cherry, when CBB are vulnerable to predation by birds, as the remainder of the CBB life cycle occurs within the coffee cherry. There are five main CBB developmental stages: egg, larva, pupa, juvenile, and adult. Females can oviposit daily for up to 40 days, averaging 1–2 eggs per day . After a week, eggs hatch and larva take 17 days to develop into pupa. Following pupation , juveniles emerge and reach sexual maturity after about 4 days . The length of the CBB life cycle can be slowed and accelerated depending on average temperature ; the developmental times used here are based on 25 C rearing conditions . Offspring sex ratio is skewed toward females, ranging from 1:5 to 1:494 . Since males are flightless, mating occurs between siblings within the natal cherry. Fertilized females then disperse to colonize other cherries, though multigenerational oviposition within the natal cherry is possible. The prolonged maturation of the coffee crop allows continual reproduction, with 2–8 CBB generations feasible in a single season if environmental conditions and food availability be favorable . With the removal of cherries during harvest, adult CBB will enter diapause in coffee cherries that remain on the plant or fall to the ground .

Flies were constantly provided with water and artificial diet that served as both a food source and an oviposition medium. Before their use in experiments, all flies were allowed to mate for 8 d in mixed-sex cages. Some small fruit varieties were numbered since this information is proprietary.This trial was conducted in Oxnard, California, USA on highbush blueberry plots during 2020. Plants were irrigated with three drip stakes per plot ten times a day for ten-minute intervals delivering 1.1 liters of water per hour. Screenhouses were fully enclosed with screen material to prevent insects from entering. There were three 70 m x 5 m screenhouses with GUM or UTC treatment randomly assigned to the north or south end of each screenhouse for a total of 6 plots. Within each screenhouse, treatment plots contained twelve plants in two rows, and plots were separated by 45 m. one-hundred flies were released in each plot four times, once per week. Three GUM deployment plots were compared with three UTC plots. GUM dispensers were installed in every other plant with irrigation stakes placed directly through the pads. The GUM application was completed on 14 April. Plots were sampled every seven days from 14 April to 12 May. One sample consisted of 50 berries.Ten field trials were conducted from September to November 2020 across multiple coastal production regions in California, USA , at different ranches and on multiple varieties being grown under high tunnels. Each location was a replicate consisting of two plots and were randomly assigned at each ranch to GUM or to UTC. Plots within a ranch received similar irrigation, fertilizer, flower buckets wholesale and insecticides. Each plot received a minimum of four spinosad sprays timed 7-10 days apart during the cropping period and based on monitoring trends from fruit collections.

Additional peroxyacetic acid applications were applied at 2-3 day intervals after each spinosad application, followed by a C. subtsugae application 1-2 d after each peroxyacetic acid application. Throughout the experimental periods, GUM dispensers were distributed evenly throughout each plot and replaced every 21 days. GUM dispensers were staked directly under the drip line in soil plots, and irrigation stakes were placed directly through the dispenser in substrate plantings. Six fruit samples were collected from each treatment plot every week for 4 to 12 weeks. Samples were collected at least 2 m from each edge of the tunnel as well as from the center of the tunnel approximately 20- 30 m from the edge of the tunnel and at ~0.75 m from the ground. Each sample consisted of 50 berries. Sample berries were incubated at room temperature for 2-4 days to allow for larval growth and facilitate detection. Samples were evaluated by crushing fruit and submerging them in a saltwater solution . The crushed fruit solution was then poured into a tray where D. suzukii larvae subsequently floated to the top of the solution and were counted.The buildup of D. suzukii populations was modeled under four scenarios i.e.; no intervention , GUM only, insecticide , and GUM and insecticide . The model parameters were obtained from experimental work and iterations of the model have been used in previous studies . Recorded D. suzukii population levels and weather data were used as model inputs. Outputs from the model were directly compared with D. suzukii infestation data of the blueberry field trial 1 . This trial was selected because of its relatively long duration and is most suitable for describing population build-up. Ambient temperature influences the fecundity rates, mortality rates, and maturation delays of the four principal life stages . The simulations were based on daily mean temperature data recorded at Aurora, Oregon, USA, between June and September 2020. We assumed that the flies had access to unlimited fruit and that no other factors affected population dynamics .

Parameter values, including for fecundity rates, mortality rates, and maturation delays, were obtained from laboratory experiments on blueberry . The simulations were initialized on 30 June with a population composed equally of adult males and females. The model simulations track relative population densities, and the initial adult density was chosen so that the simulated egg density matched the final eggs/berry in the UTC treatment. The GUM dispensers were assumed to reduce D. suzukii fecundity by 49%, according to data from Tait et al. . Insecticide induced mortality rates caused by GS were calculated from laboratory data . Spinosad was used as insecticide model. The effects of GUM and GS were assumed to start on 9 July in accordance with the model design. Details on the model and on how the GS and GUM treatments were implemented can be found in the Supplementary Material. The simulations were implemented using Wolfram Mathematica 13.0 . The code for the simulations is available online1 .The current study supports findings from previous laboratory and small-scale field cage trials. Here we show through field collected and modeled data that food-grade gum use can reduce D. suzukii fruit damage . The aim of this work was to acquire detailed knowledge about limitations of food-grade gum in a range of commercial cropping systems including blueberry, blackberry, cherry, raspberry, strawberry, and winegrape. These studies were conducted in two key production regions i.e., California and Oregon in the USA. The overall results supported initial findings and provided additional evidence that this tool can reduce D. suzukii crop damage especially when applied together with the grower standard. Both field-collected data and model simulations indicates that there is a synergistic effect of food-grade gum when used in combination with a conventional insecticide. For most of the experiments , field plots receiving the food-grade gum resulted in either numerical or statistical differences in D. suzukii damage compared to untreated control plots. This was not recorded for the cherry, strawberry, and blackberry trials. Reasonable hypothesis about these data are discussed below. In trials where D. suzukii infestations were measured in buffer plots , there was evidence of a reduction in damage, but not at the same level as in plots treated by the food-grade gum. Overall, considering all the trials, crop damage was reduced up to 78% over a period of up to 21 days post application of the food-grade gum. The results from the current study indicate that the food-grade gum can be used in combination with standard insecticides , and in some cases as a stand-alone treatment to reduce the infestation level of D. suzukii. Similar reductions in D. suzukii damage were reported under laboratory and controlled semi-field conditions , suggesting that the food-grade gum resulted in lower damage due to oviposition. These findings support earlier results where the effects of semiochemical volatiles emanating from the food-grade gum resulted in significant behavioral changes . In several trials, data lower oviposition and fruit infestation in the presence of the food-grade gum under field conditions. Reasons of why in multiple trials a statistical difference was not reached, can be explained by multiple parameters observed by scientists and growers such as animals removing the cottons pads, water-irrigation issues, and wind. These factors are addressed in a future publication . In the Hood River cherry trial, constant windy conditions may have resulted in dispersion of volatiles, flower harvest buckets ultimately resulting in impacts that were less pronounced. There is little doubt that efficiency of the food-grade gum can vary depending on production conditions and crop . Host preference of D.suzukii was ranked 4th for cherry, followed by blueberry and winegrape . Such differences in host preference should be considered when applying food-grade gum.

Synthetic blends can be less attractive compared to the actual fruit; thus, additional adjustments may be required to minimize egg-laying in the fruit. Results showed that the application of the food-grade gum in grape shows clear impacts to protect berries from D. suzukii attack. Considering the vulnerability of several winegrape cultivars towards D. suzukii and the encouraging results collected, we have reasons to believe that the food-grade gum can be a useful tool for the winegrape production. For the food-grade gum applications in blueberry in open field experiments, the infestation rate for the food-grade gum and grower standard were 70% and 85% lower than that for untreated control respectively, with the food-grade gum treatment resulted in a significantly lower infestation rate compared with the control. Open and semi-field experiments conducted in California provide similar outcomes to those in Oregon. Blueberry experiments conducted in California within a screenhouse provided 45.5% egg reduction. There were sequential applications with differing timing and the results indicated that early applications resulted in lower egg reductions . A potential hypothesis for this phenomenon could be related to environmental conditions including temperature and humidity that could significantly change the emission of plant volatiles . Egg reduction in raspberry and blackberry varied from 42-90% and 24-70% respectively. Two cultivars of raspberry have been subjected to the trial and in both cases there was reduction in egg infestation. For blackberry the same cultivar has been evaluated but in three different farms. Results were consistent between the different locations. For strawberry, in several cases results showed numerically increased larval levels compared in the food-grade gum treatments. A potential hypothesis for this phenomenon could be related to either unreported production practices or environmental conditions that could significantly change the emission of plant volatiles or the food-grade gum. Other reasons that can justify the negative results, range from lack of irrigation to rodents removing food-grade gum within a day of placement . The trial run in Watsonville, California, showed a numerical reduction of eggs when the foodgrade gum was applied as standalone and in combination with pesticide. As discussed previously, multiple factors may have impacted the trial. Meta-analysis to determine differences between food-grade gum and untreated control and mean larvae resulted in a highly significant difference. Despite the non-statistical significant results gotten in multiple trials, the meta-analysis showed that by analyzing together all the trials, the food-grade gum has a significant positive effect on protecting fruits from D. suzukii infestation. The data originated clearly indicate that the presence of the food-grade gum substrate is a valid approach to keep D. suzukii away from berries. This analysis seems extremely valuable because it provides a general idea of how, overall, the use of the new tool has the potential to bring benefits to the small berry industries all over the world. For this study, the initial D. suzukii adult densities were fitted to match the untreated control treatment. The relatively similar trends displayed between simulations and real data suggest that model assumptions are close to representative of treatments. Simulation outputs however differed slightly from the field data in the earlier phase of the season. The simulations suggest an earlier buildup of D. suzukii populations compared to the sudden increase of infestation in the field trial. A reason for this difference could be that the model output was compared with the experimental data by assuming that the simulated egg population is proportional to the mean number of eggs found per fruit in the experiments. This assumption is reasonable for constant fruit levels, but the availability of ripe fruit in the trials were not constant. Under commercial field conditions, fruit is harvested every 7-10 days for this cultivar. This means that less susceptible fruit is available directly after every harvest event, likely negatively impacting D. suzukii population levels. Therefore, a high availability of ripe fruit in the middle portion of the experiment likely resulted in fewer eggs laid per berry compared to later in the season when fruit are less available. These differences in ovipositional resources likely resulted in the sudden increase in recorded infestation levels towards the latter portion of the experiment. Future work should focus on these relationships of pest population level and crop availability to determine risk. Finally, data collected under different environmental conditions over periods ranging from 10 to 60 days do not appear to impact the efficacy of the food-grade gum. Treated fruits were less damaged by D. suzukii. Additional factors such as active distance, commercial field longevity and improved formulation will result in additional improvements and future adoption.The Berry phase has played significant roles in many aspects of physics, ranging from atoms to molecules to condensed-matter systems.

Though there is a paucity of human research linking endogenous measures of dopamine function with reinforcement learning, dopaminergic drugs modulate RPE-like fMRI signals . Beyond reinforcement learning, dopamine has been linked to a multitude of cognitive processes thought to support complex, goal-directed decision-making such as episodic memory , working memory , flexibility , and valuation . Therefore, it would be reasonable to expect that deficits in dopamine function would negatively impact decision-making, and possibly through multiple paths. Aging is accompanied by alterations in multiple components of the dopamine system, including loss of dopamine-producing neurons in the substantia nigra , and losses in dopamine receptors and transporters . However, there is accumulating evidence from in vivo PET imaging in humans indicating that dopamine changes in aging are more heterogenous than previously thought. In this review, we focus on three aspects of intra and inter individual variability and consider how they may obscure evidence of systematic changes in decision-making with age. First, declines in the dopamine system vary substantially across individuals . Second, pre and post synaptic components of the dopamine system may decline at different rates and in different directions . Third, declines in the dopamine system may be spatially heterogeneous . We provide examples for how these factors may affect decision-making processes relying on reinforcement learning as well as goal-directed processes thought to rely on working memory . We posit that incorporating in vivo imaging to account for intraindividual and inter individual variability in dopamine function may explain some of the null or conflicting age effects in the decision sciences.Perhaps reflecting the complex relationship between dopamine function and aging, black plastic plant pots wholesale there is surprisingly little consensus on the nature of age-related changes in value-based decision making using laboratory-based tasks.

For example, studies in animal models strongly implicate dopamine in risk-taking . However, meta-analyses of tasks assessing risk-taking in young and older adults found no effect of age , or small effects indicating greater risk aversion in older adults when potential financial gains are at stake . Previous discussions of the mixed effects in the decision making literature have emphasized how variability in task framing and difficulty have profound effects on performance in older adults and can potentially alter the direction of observed age differences . For example, a recent meta-analysis of the Iowa gambling task suggests risk aversion in aging may develop progressively over the course of a single experimental session . Therefore, older adults may appear more risk seeking or more risk averse depending on a given task’s demands on learning. While previous discussions have brought to light the importance of between-study task differences in interpreting inconsistencies in the direction of reported age-group differences, here we emphasize the ways in which inter individual variability in older adults preclude the identification of systematic age-group differences within a single study. One limitation of previous studies is the absence of in vivo assessment of dopamine function using methods such as PET. PET imaging has been critical for clarifying essential questions in cognitive aging. For example, relevant to the field of Alzheimer’s disease research, PET imaging is being used to resolve conflicting accounts of the pathological mechanisms affecting memory. Recent PET findings have demonstrated preferential relationships between the accumulation of tau and memory . Similarly, dopamine PET imaging has been central for resolving controversies regarding the neural basis of cognitive training gains and mechanisms of transfer.

Backman and colleagues have demonstrated 5 weeks of working memory training increases striatal dopamine release during performance of the training task as well during performance on untrained working memory tasks . Incorporation of neurochemical and neuropathological quantitation allows for unique insights into the mechanisms underlying cognitive decline or enhancement in humans that are not possible using fMRI, electroencephalography, or structural imaging alone. Here, we discuss how the addition of in vivo dopamine measures to behavioral and structural and functional imaging studies will be useful for organizing the range of age effects reported in the decision sciences. We first provide background on in vivo dopamine imaging, and describe the strengths and limitations of these methods. We next identify three sources of inter individual and intra individual variability in age-related changes in brain dopamine, which have been revealed through PET imaging. Using specific examples to illustrate how these sources of variability can produce inconsistent age group effects, we propose ways in which in vivo imaging can clarify the neural basis for these findings. Finally, we address the possibility that changes in dopamine function join with age-related alterations in affective attention to increase inter individual variability in decision-making performance. We suggest that age-related changes in affective attention influence decision-making, and may, at times, oppose the effects of altered dopamine function on performance. We propose that accounting for interactions between dopamine and affective attention will be useful for explaining apparent noise in decision-making performance between individuals and between tasks.In this section, we briefly review methods for in vivo dopamine imaging in humans, which we hope provides useful background information for our discussion of how these methods can bolster our understanding of decision-making in aging.

PET imaging allows for the assessment of multiple components of dopamine function in vivo in animal models and humans. Here, we focus our review on PET imaging methods, though similar principles apply to SPECT imaging. Radiotracers have been developed that target dopamine receptors, transporters, and enzymes involved in dopamine synthesis . Commonly, PET imaging is conducted while subjects are not cognitively engaged in a specific task, but are in baseline resting conditions. Examples of how dopamine PET is paired with simultaneous cognitive task performance is described below. In a typical experiment, a subject is injected with a single bolus of the radiotracer and undergoes imaging over the course of 60–90 minutes. Kinetic modeling is applied to the data to provide a single whole-brain image . This image provides a static snapshot of the occupancy of specific dopamine receptors or transporters, or enzymatic function underlying dopamine synthesis capacity within an individual . Similarly, neuromelanin-sensitive MR approaches provide a static snapshot of the health of the nigral dopamine system , though relationships between MR and PET measures have not yet been established . Region of interest analyses can test how these measures vary across individuals and correlate with specific behaviors or other neural measures. For imaging receptors and transporters, radiotracers that act as competitive agonists or antagonists are used. It is worth noting that while most tracers give good quantitation in the striatum where the concentration of dopamine targets is high, fewer tracers allow for measurement in regions such as thalamus, amygdala, hippocampus, and cortex, where concentrations may be 10-fold less. Therefore, higher affinity tracers for D1 and D2/3 receptors must be used for research aimed at delineating contributions of cortical dopamine in decision-making. Tracers targeting receptors and transporters are characterized as “reversible,” meaning they bind to their target but can also dissociate until they reach a steady state during which the flux of tracer tissue binding equals the flux of dissociation back into blood. Calculations of non-displaceable binding potential are common for assessing individual difference in the availability of dopamine receptors and transporters . In a given region of interest, BPND reflects the density and affinity of the targeted receptor or transporter. However, as the tracer is in competition with endogenous dopamine to bind to its target, BPND is also sensitive to individual differences in the concentration of endogenous dopamine. Therefore,BPND comprises both the density/affinity of the receptor/transporter of interest as well as the concentration of competing dopamine particularly for lower affinity tracers. Thus, there is no “pure” PET measure of dopamine receptor density or transporter density. There are established PET methods for assessing dopamine release within individual subjects, which capitalize on the competitive displacement of radiotracers by endogenous dopamine . Specifically, decreases in receptor BPND accompany increases in extracellular dopamine concentration, which has been validated by simultaneous microdialysis . Due to slow tracer kinetics, black plastic plant pots bulk current PET imaging methods do not allow for event-related measurement of dopamine release for single trials. Therefore, direct comparison with phasic dopamine release afforded by fast scan cyclic voltammetry in animal models is untenable. In humans, PET measures of dopamine release reflect changes in extracellular dopamine across 10–60 minutes, depending on study design.

One approach is to collect two PET scans per subject and compare baseline BPND with BPND during task performance , or following administration of a drug that increases synaptic dopamine concentration by blocking dopamine reuptake or stimulating release . A second approach is to measure alteration in dopamine release across a single session. These protocols are somewhat more onerous and usually require constant infusion of the radiotracer, but have demonstrated increases in dopamine release associated with cognitive task performance . In addition to PET imaging, it is possible to assess dopaminergic function in vivo using neuromelanin-sensitive MR. This approach enables the visualization of monoaminergic nuclei in the substantia nigra pars compacta and locus ceruleus . Neuromelanin’s binding to iron and copper facilitates the visualization of neuromelanin-rich regions in using MR approaches . Its sequestration of heavy metals is likely neuroprotective, though after neuronal death, the release of toxins into extracellular space may be detrimental . Supporting the validity of this measure for assessing individual differences in the integrity of substantia nigra dopaminergic function, there is evidence that neuromelanin MR signal is reduced in Parkinson’s disease and distinguishes healthy controls from people with schizophrenia and depression . Consistent with PET dopamine synthesis findings, healthy aging is associated with elevation of neuromelanin . Individual differences in neuromelanin MR signal in healthy aging have been linked to variability in reward learning , memory performance , and fMRI activation during encoding . To date, there has been little investigation characterizing relationships between neuromelanin MR signal and dopamine PET measures within subject in young or older adults. One study reported a positive relationship between neuromelanin signal and D2/3 BPND in VTA/substantia nigra, but failed to find a relationship with dopaminesynthesis capacity . However, this study may have been underpowered , and used L-[β− 11C]DOPA to measure dopamine synthesis capacity, which complicates data analysis compared to the fluoro-l-mtyrosine PET measure of dopamine synthesis capacity . While the neuromelanin MR imaging approach is still under active development , it represents a low-cost and easily implemented way to approximate individual differences in the integrity of neurochemical systems relevant to cognition.In vivo imaging has the advantage of providing a within-subject, continuous measure of dopamine function that can be used to assess individual differences and offers a perspective of which brain regions may be preferentially associated with specific aspects of cognitive performance. What has emerged in the study of aging is the observation that changes are not monotonic. Different components of the dopamine system appear to change at different rates, in different directions, or not at all. Further, age-related changes in dopamine receptors may be spatially nonuniform. Below, we summarize these findings, which together speak to the limitations of experimental approaches that do not seek to account for such heterogeneity in the effects of aging on the neural systems supporting decision-making. A recent meta-analysis examining age-related changes in dopamine PET measures found consistent evidence that D1 and D2/3 BPND decline with aging . Though the studies examined in this meta-analysis were cross-sectional rather than longitudinal, the magnitude of age-related reductions was illustrated by the estimation of the percent reduction per decade of life. D1 receptors declined at a rate of ~14% per decade while D2/3 receptors and the dopamine transporter declined at a rate of 8%–9% per decade. These estimations derived from human PET imaging studies are generally consistent with, though in some cases are slightly higher than quantitation from postmortem human tissue and nonhuman animal studies . While receptor BPND is lower in older adults relative to young, studies consistently reveal substantial inter-individual variability in D1 and D2/3 BPND in older adults. For example, D2/3 BPND is relatively preserved in people who are more physically active . Such interindividual variability appears to be relevant to cognition, as it correlated with differences in psychomotor function , executive function , and memory in older adults. While dopamine receptor BPND declines in aging, there is accumulating evidence that dopamine synthesis capacity is elevated in older adults .

The mobile phase consisted of isocratic elution with acetonitrile:water at a flow rate of 1.0 ml/min with a run time of 22 min. Standard solutions of 10 mg/L of D-glucose, D-fructose, Dsucrose, and D-raffinose were injected to obtain the retention time for each compound, and detection was conducted by RID. Sugar standards were purchased from VWR International . Sugar concentration of each sample was determined by comparison of the peak area and retention time with standard sample curves. Starch content of the roots, shoots, and leaves was conducted using the Starch Assay Kit SA-20 in accordance with the manufacturer’s instructions. Briefly, pellets of different tissues were dissolved in 1 ml DMSO and incubated for 5 min in a water bath at 100◦C. Starch digestion commenced with the addition of 10 µl α-amylase and then incubated in boiling water for another 5 min. Then, the ddH2O was added to a total volume of 5 ml. Next, 500 µl of the above sample and 500 µl of starch assay reagent were mixed and incubated for 15 min at 60◦C. Negative controls with the starch assay reagent blank, sample blank, and glucose assay reagent blank and positive controls with starch from wheat and corn were performed. Reaction started with the incubation of 500 µl of each sample and 1 ml of glucose assay reagent at 37◦C and was stopped with the addition of 1 ml of 6 M sulfuric acid after 30 min. The reaction was followed with a Cary 100 Series UVV is Spectrophotometer and starch content was expressed as percent of starch per tissue dried weight.In spite of the warming trends recorded for the study area within the two growing seasons covered by this study, 30 planter pot the plant water status recorded in both growing seasons was optimal for grapevine growth as indicated by the midday SWP and the gs . Thus, seasonal integrals of SWP ranged between -0.8 and -1.1 MPa, while gs ranged between 150 and 250 mmol m−2 s −1 , in accordance to the midday SWP and gs values considered as well-watered conditions .

Moreover, water status of the grapevines subjected to less applied water amount never reached values lower than -1.5 MPa for SWP and/or 50 mmol m−2 s −1 for gs , which have been reported to impair grapevine performance and berry ripening . As Keller et al. reported before, in warmer years, 100% ETc treatment may suffer from mild water deficit. Thus, under our experimental conditions, at the end of the season, especially in 2020, grapevines reached SWP values to ca. -1.2 MPa; however, they are not sufficient to impair grapevine physiology and metabolism in warm climates . Previous studies highlighted that plant water status is closely related to leaf gas exchange parameters . Thus, low values of SWP were related to decreased gs likely because plants subjected to mild to moderate water deficit close their stomata as an early response to water scarcity to diminish water loss and carbon assimilation . Accordingly, in both growing seasons, a higher SWP promoted increased stomatal conductance and, consequently, net carbon assimilation rates in grapevines subjected to 100% ETc. AN and gs peaked around veraison and then declined in all the treatments similar to several studies conducted in a warm climate before . Thus, previous studies have pointed out that limited photosynthetic performance, hence lower gs and AN values, may be triggered by passive or active signals . Nevertheless, AN in 50% ETc treatment was not severely decreased presumably by increases in WUE, which have been related to improvements in stomatal sensitivity to water loss and vapor pressure despite the hormonal signaling from roots to shoots . Likewise, Tortosa et al. suggested that differences in WUE between Tempranillo grapevine clones were more explanatory of the variations in carbon assimilation rather than a different stomatal control. Finally, it is worth mentioning that WUE was significantly lower in the driest and hotter growing season regardless of the irrigation treatment as previously reported . Regarding intrinsic WUE , no effect due to growing conditions was observed in contrast to previous studies on vines subjected to mild water stress .

The water deficits applied in this study were from moderate to severe based on SWP values; thus, it is expected that the vegetative and reproductive growth of vines will be impacted accordingly. Thus, in previous studies, higher water deficits resulted in reductions of yield and berry size . The reduction in berry mass has been associated with the inhibition of cell expansion and the diminution of inner mesocarp cell sap . The detrimental effects of 25% ETc were reported previously, suggesting that this applied water amount may not be adequate for hot climates with very little or no summer precipitation . Vegetative growth was also impaired by water deficits applied in this study, as indicated in the decrease of leaf and root dry bio-masses measured in 25 and 50% ETc treatments. Diminution of root growth under water stress has been related to the loss of cell turgor and increased penetration resistance of dried soils . In addition, a recent study suggested that the loss of leaves could decrease the supply of carbohydrates and/or growth hormones to meristematic regions, thereby inhibiting growth . In accordance with previous studies, severe water deficits led to lower shoot to root ratio because root growth is generally less affected than shoot growth in drought-stressed grapevines . Given that grapevine vegetative growth occurs soon after bud break in springtime, our results corroborated the crucial role of water availability during that period on vine development, physiological performance, and yield components reported in previous studies . Thus, irrigation of grapevines during summer could not be sufficient to fulfill water requirements when rainfall has been scarce in spring , and precipitation amounts prior to bud break result in cascading effects for the rest of the growing season that cannot be overcome with supplemental irrigation .The allocation of NSC varied between organs for which roots accounted 30%, shoots 25%, and leaves 40% of the whole plant NSCs at harvest, slightly differing from those reported for several fruit trees but similar to theworks in grapevine . The NSC composition was highly dependent on the grapevine organs, with starch being the main NSC in the roots and shoots.

Previous studies reported that roots accumulated the largest amounts of starch in plastids, namely amyloplasts, which is fundamental to allow rapid vegetative development during the next spring . Our results also corroborated with this finding. Our results indicated that, apart from fruits, SS were mainly accumulated in the leaves at harvest, which accounted for about 90% of the total leaf NSC. Thus, the allocation of NSCs in different organs allowed the plants to persist when respiration rate was higher than photo assimilation in annual events, but also aided in responding to abiotic stresses such as drought . Our results indicated that plants that received 100% ETc had higher NSC content. Similarly, a previous study with potted grapevines reported increased starch and SS contents in the leaves from the grapevines with higher leaf area to fruit ratio that were well-watered . In shoots, sucrose and raffinose proportions were higher in 50 and 100% ETc treatments compared with 25% ETc. As a great part of the shoot biomass is vascular tissue, this may suggest an increase in NSC translocation in these treatments. Although sucrose is the main sugar for carbon translocation through the phloem into the sink tissues, recent research highlighted the roles of other sugars, such as raffinose, in carbon translocation and storage . On the other hand, plastic growers pots previous research reported less NSC accumulation in grapevine canes under carbon starvation at a low leaf to fruit ratios, suggesting that sucrose may control starch accumulation through adjustment of the sink strength . Furthermore, Rossouw et al. also highlighted the role of raffinose toward root carbohydrate source functioning in grapevines with significantly lower leaf to fruit ratio due to defoliation from carbon starvation . When the photosynthetic supply of carbohydrates is limited, remobilization from perennial tissues can provide an alternative carbon source . Thus, previous research conducted on potted grapevines reported a concurrent starch remobilization from roots with a rapid berry sugar accumulation . Conversely, under our experimental conditions , no effect of water deficits on NSC remobilization from roots to berries was observed despite the decreased leaf to fruit ratio. Likewise, Keller et al. did not observe higher amounts of sugars in berries from field-grown Cabernet Sauvignon subjected to 25% ETc compared with 70 or 100% ETc under field conditions.Under our experimental conditions, yield per plant was strongly related to shoot, leaf, and root BM. Similarly, Field et al. found that grapevines with the lowest shoot growth rate before veraison had significantly less fruit set than the other treatments, attributing these effects to the restoration of root carbohydrate reserves that occurred at the same time. Grapevines subjected to 25% ETc had reduced photo assimilates due to lower AN in both seasons resulting in less NSC in the source leaves available for new growth and exported to sinks. This resulted in a general lower plantBM . Contrarily, grapevines subjected to 100% ETc had higher photo assimilation rates throughout the course of the study that led to higher SS and starch content and, consequently, to the improvement of BM and, therefore, higher harvest index. Therefore, the reduced growth rate of both sink and source organs in response to water deficits indicated that the availability of carbon is a major growth constraint. The yield per plant of 50% ETc was lower than 100% ETc, but not as low as 25% ETc. However, canopy BM was greatly reduced in both 50% ETc and 25% ETc compared with 100% ETc. Accordingly, Field et al. reported that grapevine grown under warm soil conditions favored shoot and fruit development over carbohydrate reserve accumulation. In contrast, Candolfi-Vasconcelos et al. reported that a lower leaf area to fruit ratio increased the translocation of carbohydrates from permanent structures to reproductive organs to support grape ripening. The shoot to root ratio revealed a positive relationship with the total BM, leaf and root NSC, and N contents. Thus, the distribution of biomass relies on the C:N ratio as highlighted by the negative relationship between shoot to root and the sucrose:nitrogen ratios. Similarly, a linear relationship between NSC and root to shoot ratio in grapevines grown under stressful conditions was previously reported . From a molecular point of view, the alterations of source:sink ratios led to transcriptional adjustments of genes involved in starch metabolism, including the upregulation of VvGPT1 and VvNTT for lower leaf area to fruit ratios . Furthermore, enhanced root biomass in 100% ETc likely resulted from higher sugar content in the roots as our data supported. It was recently reported that increases in root elongation and hexose contents were due to the VvSWEET4 overexpression, a gene implied as a grapevine response to abiotic stress . Similarly, Medici et al. reported up- or downregulation of the genes encoding hexose transporters in grapevines subjected to water deficits corroborating this result. Therefore, although some genes may be expressed under water deficit, lack of carbon accumulation impaired the growth. The relationship between root to shoot ratio and plant nitrogen content was previously reported for grapevines, suggesting that dry matter partitioning is largely a function of the internal status of the plants . We found decreased N content in grapevines facing water deficits, which resulted in a decrease of total BM. Similarly, Romero et al. reported reductions in leaf nitrogen content when vines were subjected to water deficits. These authors suggested that nutrient uptake may be reduced due to deficits in soil water profile, and the slow root growth under these conditions consequently inhibited grapevine growth. In our study, N content was strongly related to photosynthetic pigments. Accordingly, previous studies reported lower leaf N and leaf chlorophyll in deficit-irrigated grapevines, suggesting quantitative losses in the photosynthetic apparatus and/or damage to the biochemical photosynthetic machinery, decreasing photosynthetic capacity as corroborated with the lower NSC leaf content with water deficits. Finally, molecular research over the last decades has suggested the important regulatory functions of sucrose and N metabolites in metabolism at the cellular and subcellular levels and/or in gene expression patterns, giving new insights into how plants may modulate over a longer period its growth and biomass allocation in response to fluctuating environmental conditions .

Few studies have conducted a thorough evaluation of below ground vine biomass in vineyards, although Elderfield did estimate that fine roots contributed 20–30% of total NPP and that C was responsible for 45% of that dry matter. More recently, Brunori et al. studied the capability of grapevines to efficiently store C throughout the growing season and found that root systems contributed to between 9 and 26% of the total vine C fixation in a model Vitis vinifera sativa L. cv Merlot/berlandieri rupestris vineyard. The results of our study provide a utilitarian analysis of C storage in mature wine grape vines, including above and below ground fractions and annual vs. perennial allocations. Such information constitutes the basic unit of measurement from which one can then estimate the contribution of wine grapes to C budgets at multiple scales— fruit, plant or vineyard level—and by region, sector, or in mixed crop analyses. Our study builds on earlier research that focused on the basic physiology, development and allocation of biomass in vines. Previous research has also examined vineyard-level carbon at the landscape level with coarser estimates of the absolute C storage capacity of vines of different ages, as well as the relative contribution of vines and woody biomass in natural vegetation in mixed vineyard-wild land landscapes. The combination of findings from those studies, together with the more precise and complete carbon-by-vine structure assessment provided here, mean that managers now have access to methods and analytical tools that allow precise and detailed C estimates from the individual vine to whole-farm scales. As carbon accounting in vineyard landscapes becomes more sophisticated, blueberry in container widespread and economically relevant, such vineyard-level analyses will become increasingly important for informing management decisions.

The greater vine-level measuring precision that this study affords should also translate into improved scaled-up C assessments . In California alone, for example, there are more than 230,000 ha are planted in vines. Given that for many, if not most of those hectares, the exact number of individual vines is known, it is easy to see how improvements in vine-level measuring accuracy can have benefits from the individual farmer to the entire sector. Previous efforts to develop rough allometric woody biomass equations for vines notwithstanding, there is still a need to improve our precision in estimating of how biomass changes with different parameters. Because the present analysis was conducted for 15 year old Cabernet vines, there is now a need for calibrating how vine C varies with age, varietal and training system. There is also uncertainty around the influence of grafting onto rootstock on C accumulation in vines. As mentioned in the methods, the vines in this study were not grafted—an artifact of the root-limiting duripan approximately 50 cm below the soil surface. The site’s location on the flat, valley bottom of a river floodplain also means that its topography, while typical of other vineyard sites per se, created conditions that limit soil depth, drainage and decomposition. As such, the physical conditions examined here may differ significantly from more hilly regions in California, such as Sonoma and Mendocino counties. Similarly, the lack of a surrounding natural vegetation buffer at this site compared to other vineyards may mean that the ecological conditions of the soil communities may or may not have been broadly typical of those found in other vineyard sites. Thus, to the extent that future studies can document the degree to which such parameters influence C accumulation in vines or across sites, they will improve the accuracy and utility of C estimation methods and enable viticulturists to be among the first sectors in agriculture for which accurate C accounting is an industry wide possibility.

The current study was also designed to complement a growing body of research focusing on soil-vine interactions. Woody carbon reserves and sugar accumulation play a supportive role in grape quality, the main determinant of crop value in wine grapes. The extent to which biomass production, especially in below ground reservoirs, relates to soil carbon is of immediate interest for those focused on nutrient cycling, plant health and fruit production, as well as for those concerned with C storage. The soil-vine interface may also be the area where management techniques can have the highest impact on C stocks and harvest potential. We expect the below ground estimates of root biomass and C provided here will be helpful in this regard and for developing a more thorough understanding of below ground C stores at the landscape level. For example, Williams et al. estimated this component to be the largest reservoir of C in the vineyard landscape they examined, but they did not include root biomass in their calculations. Others have assumed root systems to be ~30% of vine biomass based on the reported biomass values for roots, trunk, and cordons. With the contribution of this study, the magnitude of the below ground reservoir can now be updated.Grapes are the most valuable fruit crop in the United States, valued at over $6.5 billion annually , but climate change is projected to reduce grape production and quality . Climate affects grape quality by impacting the concentration of sugars, organic acids, and secondary compounds . The climatic conditions producing the highest quality wine cause the berries to reach optimal ratios between sugar and acid concentrations and maximum concentrations of pigment, aroma, and flavor compounds simultaneously . Hot temperatures accelerate sugar accumulation, forcing growers to harvest earlier, before berries reach optimal flavor development, to avoid the high alcohol content and insipid wine flavor from excessive sugar to acid ratios .

Harvest dates have shifted earlier historically, and climate models predict further acceleration of ripening . Growers can partly compensate through management practices, such as trimming canopies or using shade clothes to reduce the ratio of sugar supply to demand , though these practices are costly and increasingly ineffective in the face of climate change . Planting existing cultivars or developing new cultivars with slower sugar accumulation are promising alternative strategies to mitigate these climate change impacts, but these efforts have been hindered by uncertainty around the plant traits controlling sugar accumulation . Grape cultivars vary in berry maturation and sugar accumulation rates, and in their response to abiotic stress, but the main anatomical and physiological mechanisms driving these differences remain unknown . Multiple physiological processes influence berry sugar accumulation and its responses to climate, including photosynthesis, long-distance sugar transport, and local transport and metabolism in the berries . However, the relative importance of these factors in regulating sugar concentrations and fruit growth is debated . Photosynthetic responses to heat and water stress could impact cultivar differences in accumulation rates by affecting the sugar supply for ripening . Further, sugar is transported from the photosynthesizing leaves to the berries through the sugar-conducting vascular tissue – the phloem. At the onset of ripening , the berries significantly accelerate sugar accumulation by initiating active sugar unloading from the phloem, making the phloem the primary pathway for water and resource influx into the berries . The importance of phloem transport to ripening suggests that phloem traits could be important drivers of cultivar differences in sugar accumulation, and that modifying phloem traits to slow sugar accumulation under hot conditions could help mitigate the impacts of climate change on wine quality. However, plastic planters bulk the main traits controlling sugar accumulation in grape remain unclear . The rate of phloem transport is determined by both the hydraulic resistance to the flow of sugar sap, and the activity and kinetics of water and sugar transporters in the sources, sinks, and along the transport pathway . Modeling studies suggest that increasing the hydraulic resistance of the phloem reduces sugar export to the sinks . Therefore, selecting grape cultivars with lower total phloem conductance could decelerate sugar accumulation and improve the synchronization of sugar accumulation with flavor development under hotter conditions. However, a higher hydraulic resistance can make the phloem more susceptible to declines or even complete failures in transport under severe water stress . Thus, we expect cultivars that produce high-quality wine in hot, dry conditions to exhibit phloem hydraulic resistances that slow berry sugar accumulation while avoiding phloem failure. The phloem transport pathway is composed of individual sugar-conducting cells with porous end walls stacked to form conduits . The anatomy of the transport pathway, including the total cross-sectional area of sieve tubes in plant organs, lumen area of individual sieve tubes, and porosity of the sieve plates, significantly impacts pathway resistance . Plants with a greater cross-sectional area dedicated to phloem , sieve tubes with wider lumen areas , and larger and more abundant pores in the sieve plates are expected to have a lower hydraulic resistance .

Total phloem cross-sectional area in the shoots has been found to vary between several grape cultivars , and a greater cross-sectional phloem area has been linked to faster sugar accumulation in the fruit in other crop species . However, the variation of phloem structural traits across cultivars adapted to a diverse range of climatic conditions and the relationship of these traits to sugar accumulation is largely unknown for grapevines. Establishing these anatomical links could allow breeders to modify sugar accumulation by selecting for phloem traits, instead of management practices that can negatively impact the fruit zone environment or yield . In this study, we used a common garden experiment to evaluate the links between phloem anatomy and sugar accumulation across 18 winegrape cultivars typically grown in climatically diverse grape growing regions. We assessed phloem and xylem vascular anatomy in leaf petioles and midveins and berry pedicels, to capture hydraulic resistance along the long-distance transport pathway. We also measured maximum berry sugar accumulation rates in the post-veraison ripening period to capture the greatest capacity for sugar transport . We predicted that traits that reduce hydraulic resistance, including larger total cross-sectional phloem areas, larger mean lumen areas for individual sieve tubes, and more porous sieve plates would increase maximum sugar accumulation rates. We also predicted that cultivars typically grown in hotter wine regions would have traits that increase hydraulic resistance, as an adaptation to increase wine quality by reducing the rate of sugar accumulation. In addition, we measured photosynthesis and vine water stress to compare the impacts of phloem anatomy, vine carbon supply, and vine water status on sugar accumulation rates. Overall, our goals were to determine the most influential traits for sugar accumulation in grape berries and evaluate the role of phloem anatomy in adapting grape cultivars to a wide range of different climates.After 7 days in FAA, the light microscopy samples were soaked in 50% ethanol for 5 mins and then stored in 70% ethanol in preparation for paraffin embedding. Samples were first infiltrated with paraffin by using an Autotechnicon Tissue Processor to treat samples with the following sequence of solutions: 70%, 85%, 95%, 100% ethanol, 1 ethanol:1 toluene, 100% toluene , and paraffin wax , each for 1 hour. The infiltrated samples were then embedded into paraffin blocks with a Leica Histo-Embedder , and allowed to cool. A rotary microtome was then used to make 7μm-thick cross-sections for leaf laminas, petioles, and berry pedicels. Pedicel cross-sections were sampled from the receptacle and petiole and midvein cross-sections were sampled near the interface of the lamina and petiole. After the cross-sections were imaged, pedicels for four cultivars were remelted from their wax molds, oriented longitudinally and sectioned again at 7μm to obtain sieve element lengths. Sections were stained using a 1% aniline blue and 1% safranin solution following a modified staining procedure . Sections were then viewed under bright field or florescence microscopy using a Leica DM4000B microscope and a DFC7000T digital camera . Each pedicel , midvein , and petiole section was then measured for total phloem and xylem cross-sectional area using ImageJ software, by manually selecting relevant tissue areas. Safranin stained the secondary cell walls of the xylem red and phloem cell walls were stained blue by aniline blue. The phloem area measurements included sieve tubes and phloem fibers and parenchyma , and xylem area measurements included xylem vessels, fibers, and parenchyma.The pedicel electron microscopy samples were processed following Mullendore . Briefly, samples were thawed at room temperature, washed in DI water, and cut into 1 mm cross sections with a fresh double-sided razor blade.

Shade cloths can be used to cover the entire canopy and thus affect a large-scale change in vineyard microclimate, or they can be applied directly to the fruiting zone to localize their impacts to berry microclimate. Previous studies have reported that shade cloths are effective in reducing maximum berry temperatures, though they have used different methods for quantifying these effects. Mart´ınez-Luscher et al. ¨ reported that 40% black shade cloths covering the fruiting zone of the canopy reduced cluster temperature by 3.7◦C during the warmest time of the day, while Greer found that 70% shade cloths covering the grapevines reduced canopy temperature by an average of 4.6◦C throughout the day. Similarly, thin shade cloths and plastic films covering the grapevines were shown by Rana et al. to reduce midday berry temperatures by 2◦C and 6◦C below air temperature, respectively. While it is clear that shade cloth can be effective in reducing overall berry temperature, a higher degree of control of berry temperature may be desired. In certain instances, it may be beneficial to reduce berry temperature by a defined margin to avoid negative trade-offs, while also balancing temperatures between opposing sides of the vine. However, many interacting variables are likely to influence the efficacy of shade cloth, such as row orientation, row spacing, trellis type, and topography. Due to the large number of important variables affecting berry temperature, it can be difficult to generalize the relatively small number of experimental results that are only able to explore a few variable combinations in order to predict the effect of shade cloth for a given vineyard system. Crop models provide the potential for generalizing the results of field experiments to predict the outcomes of proposed management strategies for a specific site or climate scenario. Such models could allow for optimization of the design or management of vineyards to mitigate elevated berry temperatures under current or future climates, growing berries in containers given that a large number of simulations can be efficiently performed to cover a wide parameter space.

Previous work has developed models of spherical fruit temperature, including one study that simulated the effect of hail nets on apple temperature. In a recent study, a 3D model was developed and validated that accurately simulated the spatial and temporal temperature fluctuations of grape berries in vineyards with different climates, topographies, and trellises.However, the current version of this model is not able to represent the effects of shade cloth on canopy and berry temperature. The overall goal of this work was to enable model-based evaluation and optimization of strategies for grape berry temperature control using fruit zone shade cloth. With this goal in mind, specific objectives of this study were to: 1) develop a physically-based 3D model of grape berry temperature that incorporates the effect of shade cloth, 2) generate an experimental data set against which the model can be validated, and 3) quantify the interacting effects of different strategies for excessive berry temperature mitigation such as altered row orientation, row spacing, topography, and shade cloth density.The model of grape berry temperature was based on the 3D model described in Ponce de Leon´ and Bailey, and modified to include the effects of shade cloth. A brief description of the overall model is provided below, with a focus on novel additions associated with shade cloth. The model was developed within the Helios modeling framework and has been validated based on field measurements of berry temperature between veraison to harvest. The computational domain consists of 3D geometric elements that fully resolve the spatial structure of the plants, berries, and shade cloth . The berries were represented by 3D tessellated spheres composed of triangular elements, the ground surface by a planar grid of rectangular elements, the woody tissues by a cylindrical mesh of triangular elements, and the leaves by planar rectangles masked to the shape of leaves using the transparency channel of a PNG image.

The shade cloth was formed by rectangles masked to the shape of a grid using the transparency channel of a PNG image, where “holes” in the shade cloth were created based on a grid of transparent pixels.Radiation transfer was simulated using a backward-ray-tracing approach that ensures each geometric element is adequately sampled for both short-wave and long-wave radiation. The model launches a large number of rays from each geometric element to simulate the various modes of radiation transfer, including emission, reflection and transmission based on the radiative properties of each element. When a ray encounters a transparent pixel on an element masked by a PNG image, the ray continues with no interaction. This allows for an efficient fully-resolved representation of shade cloth with a large number of holes. To eliminate domain edge effects, periodic lateral boundaries can be enabled that effectively creates an infinitely repeating vine geometry in the horizontal. For berries, the latent flux term was assumed to be zero since latent cooling is typically considered negligible after veraison. For this study, heat released from metabolic activity ´ within the fruit was assumed to be low enough to be neglected. The heat transfer coefficient from fruit, ground, and leaf surfaces to the atmosphere, h, are specified as described in Ponce de Leon and Bailey . Previous field tests suggested an important role of berry heat storage in accurately representing temperature dynamics, and laboratory and field tests have shown that the chosen values for Cp and rhoA result in berry dynamic temperature responses in close agreement with measurements.To calculate Rn, the model requires specification of the incoming radiation flux, which was generated following the REST2 model of Gueymard, which itself requires site longitude, latitude, offset from UTC, atmospheric pressure, air temperature, atmospheric turbidity coefficient, relative humidity, and Julian day of the year.

The incoming radiation was split into short-wave radiation and long-wave radiation. The short-wave radiation was assumed to be partitioned between PAR and NIR bands, 47% and 53%, respectively. The incoming diffuse long-wave radiation flux from the sky was estimated using the model of Prata. The input parameters to calculate Rn are the surface reflectivity and transmissivity for PAR and NIR bands, and the surface emissivity for long-wave radiation for each element in the domain. The measured values for berries, leaves and ground are reported in Ponce de Leon´ and Bailey and listed in Table 4.1. For all simulations, ambient diffuse solar radiation was neglected and all solar energy was collimated in the direction of the sun. The number of direct rays sampled on each element was set to 500 rays. The radiation scattering depth was chosen to be 3, and it was verified that using larger values would have a negligible impact on results.To evaluate the model accuracy, the time series of experimental measurements and simulated temperatures for berries furthest west were compared. Model agreement with field measurements was quantified using the error metrics normalized root mean squared error – normalized by the difference between maximum and minimum measured data, the coefficient of determination , and the index of agreement. To analyze the modeled berry temperature results in the “simulation experiment”, we took the average temperature of five berries in each of five exterior clusters on both sides of the vine. The selected number of berries and clusters represent a typical experimental sampling strategy . For comparison purposes, the total canopy and cluster daily light interception was calculated by integrating the light interception fluxes over the day. The daily light interception was calculated per vine area for the canopy and per berry surface area for the clusters To compare the effect of shade cloth density on berry temperatures in the simulated vineyard designs, we used indices based on the duration of elevated temperature and the intensity . The 35◦C temperature threshold was chosen because it has been reported to correspond to the approximate onset of unfavorable effects of elevated temperature on grape composition. Furthermore, to identify conditions that effectively balance berry temperature between opposing sides of the vine, we constructed a conditional inference tree with the variables listed in Table 4.2. The conditional inference tree was constructed using the “party” package of R statistical software. The conditional inference tree is a tree-based classification algorithm that performs binary recursive partitioning of data into groups containing observations with similar values. Conditional inference trees are similar to decision trees, blueberry containers the main difference being that each node in the conditional inference trees uses a significance test of independence to select a predictor variable rather than selecting the predictor variable that maximizes the information measure. In the conditional inference tree, predictor variables are circled and ranked and the ones at the top have the highest correlation with the response variable. The algorithm stops if the null hypothesis of independence is not rejected, however, for visualization purposes, the maximum depth of the decision tree was set to 4.

The magnitude of the wind speed varied across the sampling period, but overall Unilateral tended to have greater wind speeds compared to Goblet . During the day, the wind in the fruiting zone tended to come from the east while at night the wind tended to come from the west for both trellis systems . Since the row orientation of Goblet and Unilateral was N-S and the vines were on an east-facing slope, the wind coming from the east tended to be of greater magnitude. For both Goblet and Unilateral, the relative humidity significantly decreased on 25 Sept. 2019, likely due to the increase in air temperature. Overall, during the sampling period, Unilateral tended to be slightly less humid compared to Goblet. The relative reduction in maximum berry temperatures due to the shade cloth in Goblet and Unilateral was consistent across the sampling period, which included a wide range of ambient air temperatures . Pulp temperatures of more than 40◦C were measured on 25 Sept. 2019 for unshaded berries in both trellis systems. For both Goblet and Unilateral, the berry temperature under the shade cloth was close to the air temperature in the morning, while air temperature differences between berries under the shade cloths increased in the afternoon. The peak in measured elevated temperatures happened at different times in Goblet and Unilateral , primarily due to the influence of the ratio between plant height and row spacing on berry exposure. In the evening, under low-light conditions, the During the sampling period, average daily maximum temperatures of the west-facing berries under the shade cloths were 1.6◦C and 2.7◦C higher than the daily maximum air temperature in Goblet and Unilateral, respectively. The control west-facing berries with no shade cloth were up to 8.2◦C and 5.9◦C above the daily maximum air temperature in Goblet and Unilateral, respectively . The relative reduction in maximum berry temperatures due to the shade cloth in Goblet and Unilateral was consistent across the sampling period, which included a wide range of ambient air temperatures . Pulp temperatures of more than 40◦C were measured on 25 Sept. 2019 for unshaded berries in both trellis systems. For both Goblet and Unilateral, the berry temperature under the shade cloth was close to the air temperature in the morning, while air temperature differences between berries under the shade cloths increased in the afternoon. The peak in measured elevated temperatures happened at different times in Goblet and Unilateral , primarily due to the influence of the ratio between plant height and row spacing on berry exposure. In the evening, under low-light conditions, the berry temperatures were similar to the air temperatures, while at night, radiative cooling likely caused the pulp temperature to fall below the air temperatures. Under low light and at night, the temperature differences among treatments were small .The model was validated by using the experimental measurements to determine whether the modeled berry temperatures appropriately responded to the shade cloth relative to the control . The model reproduced the magnitude of the berry temperature increase over air temperature reasonably well for both Goblet and Unilateral. The largest source of error appeared to be due to the transition period when berry sun exposure began, where the time of the simulated maximum temperature increase tended to happen earlier than that of the measurement . This mismatch could be due to slight inaccuracies in determining the position of each berry and leaf.Figures 4.7, 4.8, and 4.6 summarize the results of the simulation experiment in which the effect of different row orientations, row spacing, and slope aspects on berry temperature were evaluated.

A survey in Poland and China found that strawberries had the highest frequency of multiple pesticide residues. In the current study, insecticides were highly prevalent in vegetables and fruits . Some insecticides appeared one time and others were detected several times. Chlorantraniliprole is one of the insecticides detected one time only in apple. This insecticide is one of the diamide insecticides that are widely used against a variety of insect pests due to their selectivity and low mammalian toxicity. Tian et al. determined diamide insecticides in mushrooms and found that these insecticides can be effectively analyzed using HPLC-MS/MS with LOD and LOQ of 0.05 and 5 ug kg−1 , respectively, and recovery rates ranging from 73.5–110.2%. On the other hand, chlorpyrifos is an insecticide that was detected several times in the tested vegetable and fruit samples. Although this insecticide is recommended in Egypt against almond worms in cotton and termites in buildings according to the approved recommendations for agricultural pest control , it was detected in vegetables and fruits collected from farmers’ markets . This insecticide is no longer approved by European Commission due to harmful effects on different organs. In spite of that, it is still detected in a high percentage in many samples of fruits and vegetables, which is consistent with the results obtained in this study. Fungicides were detected in fruits in a higher percentage than in vegetables . The fungicide pyraclostrobin was detected in grapes only . The dissipation rate of this fungicide was studied in strawberry in Egypt when treated with the recommended field rate. It was found that 82% of this fungicide degraded within 14 days of treatment with a half-life of 5 days. In contrast, container growing raspberries the fungicide thiophanate-methyl was detected in four vegetable samples and in six fruit samples as recorded in Tables 1 and 2.

As this fungicide is widely used in the control of a variety of pathogens pre- and post-harvest, it was detected in many vegetable and fruit samples, herbal medicine, raisins, salmon, beebread, and also in cow and human milk. Samples of cucumber and apples were found to have ≥15 pesticide residues . Chlorpyrifos and lamda-cyhalothrin were detected in more than 15 samples with some values higher than MRLs. In our study, pesticide residues exceeding MRLs in vegetables and fruits were 41 and 39%, respectively. Other studies showed the same results, i.e., in Mwanja et al., pesticide residues were detected in 63.3% of the tested vegetable and fruit samples with residue levels exceeding MRLs of the codex Alimentarius in cabbage, tomato, and orange samples. Further, in the study of Hamed et al., residues of pesticides in apples and grapes from Egypt were determined and they reported that 12.7 and 16.4% of pesticide residues exceeded the MRLs, which was slightly lower than what we found in the current study . Consistent with our findings, a study conducted by Parveen et al. in Pakistan reported that pesticide residues in apple and grape samples exceeded MRLs with 28 and 20%, respectively. Estimation of pesticide residues in imported food is necessary to know about food safety. A study in the United Kingdom for monitoring levels of pesticide residues in imported foods from different countries showed that 51.3% of Egypt samples, compared to 77% , 68.3% , 55.1% , 46.1% , and 45.7% contained detectable pesticide residues. They recorded that India, Kenya, Brazil, Egypt, Chile, and the United States were countries with residue levels exceeding MRLs in 18.1%, 11.4%, 7.8%, 5.1%, 3.2%, and 2%, respectively. In the same context, Osaili et al. determined pesticide residues in samples of imported vegetables to the United Arab Emirates.

They found that 30.5% pesticide residues higher than MRLs in total imported samples and found 14% of the Egyptian samples compared to 47%, 33%, 13%, and 43% from India, United Kingdom, China, and Sri Lanka, respectively, contained residues higher than MRLs. The results of monitoring pesticide residues in fruits and vegetables showed that some samples had residues that exceed the MRL standard, which may lead to risks when consuming food contaminated with these pesticides. In addition, some pesticides do not have corresponding residual limits, which make it difficult for farmers to safely use these pesticides and for the government to monitor their use. Therefore, identification of acute and chronic dietary risks is necessary to assess the risks associated with consuming vegetables or fruits that contain pesticide residues above the MRLs. In this regard, Chu et al. evaluated the risks of food exposure to 26 insecticides on strawberries and found that despite the presence of high detection rates for these residues, they showed risks of acute and chronic exposure at a level of less than 100%. In our results of risk assessment, residues of lambda-cyhalothrin, fipronil, dimothoate, and omethoate were found to have acute or chronic risks in consumers in the case of consuming 100 or 200 gm day−1 of spinach, zucchini, kaki, and strawberry, respectively . In line with our findings, the results of Tao et al., 2021 showed that the fungicide carbendazim had a risk quotient value of 2.9 in wheat flour samples, indicating an unacceptable dietary risk. Furthermore, Tankiewicz and Berg showed that pesticides of lambda-cyhalothrin in courgettes, captan in apples and cucumbers, dimethoate in cour-gettes, and linuron in carrots exceeded the MRLs and pose a health risk. In an Indian study conducted by Sinha et al., they stated that excessive application of pesticides on grapes cause adverse health effects in developing countries as grapes and apples are contaminated with different classes of pesticides including organophosphate, which cause high health risks for consumers.

In this context, Javeres et al. showed that the prolonged exposure to insecticides could lead to physiological disorders including high blood pressure, hyperglycemia, overweight or dyslipidemia, which may cause metabolic syndrome and other chronic diseases. For these adverse effects, it is important in each country to monitor pesticide residues in food for food safety and human health.Tomato is the world’s second largest vegetable crop rich in nutrients. Tomato fruit development includes three stages. The first stage is characterized by an increase in cell number and starch accumulation, followed by cell enlargement with starch degradation and soluble sugar accumulation in the second stage. Fruit ripening is the last stage, associated with the accumulation of soluble sugars, carotenoids, organic acids, and volatile organic compounds in fruits. The chlorophyll accumulation and photosynthetic activity of green fruits influence the nutritional components and flavor of ripening tomato fruits. Some genes have been reported to affect chlorophyll accumulation, chloroplast development and fruit quality. As negative regulators, DE-ETIOLATED 1/high pigment 2 and UV-DAMAGED DNA-BINDING PROTEIN 1/ high pigment 1 are involved in chloroplast formation and chlorophyll accumulation in tomato fruits. The tomato GOLDEN2-LIKE transcription factors SlGLK1 and SlGLK2 play an important role in chloroplast formation and chlorophyll accumulation.Evidence suggests that the SlGLK2 gene is predominantly expressed in fruits and that the latitudinal gradient of SlGLK2 expression influences the production of unevenly colored tomato fruits. Over expression of the APRR2- LIKE gene, the closest homolog of SlGLK2, increased the size and number of chloroplasts and enhanced chlorophyll accumulation in green tomato fruits. TKN2 and TKN4, two Class I KNOTTED1-LIKE HOMEOBOX proteins, act as transcriptional activators of SlGLK2 and APRR2-LIKE genes to promote chloroplast development in tomato fruits. BEL1-LIKE HOMEODOMAIN11 also plays an important role in chlorophyll synthesis and chloroplast development in tomato fruits. The ripening of tomato is mainly regulated by the ethylene pathway and many transcription factors. In the ethylene bio-synthetic pathway, S-adenosylmethionine synthetase catalyzes the reaction of ATP and methionine to form S-adenosyl-L-methionine. 1-Aminocyclopropane-1-carboxylic acid synthase and ACC oxidase catalyze the conversion of SAM to ACC and of ACC to ethylene, respectively. The MADS box gene RIPENING INHIBITOR controls the early phase of ripening and ethylene production via transcriptional regulation of ACSs and ACOs. The other ripening regulators affecting ethylene production also include the NAC transcription factor NOR, the SQUAMOSA PROMOTER BINDING protein CNR, blueberries in pots the ethylene response factor ERF B3, the AP2/ERF member AP2a, and several MADS box proteins, such as TDR4/SlFUL1, SlFUL2, SlMADS1, TAGL1, and TAG1.

Auxin is an important phytohormone involved in flower fertilization, fruit setting, fruit initiation and development. Auxin is also essential in the regulation of cell division and expansion, controlling final fruit size. Auxin modulates plant development through transcriptional regulation of auxin-responsive genes, which is primarily mediated by two gene families: the short-lived nuclear protein Aux/IAA family and auxin response factors. Most ARFs have an N-terminal DNAbinding domain required for transcriptional regulation of auxin response genes, a middle region functioning as a repression domain or activation domain , and a C-terminal dimerization domain involved in the formation of homodimers or heterodimers. ARFs can act as either an activator or a repressor of the transcription of auxin-responsive genes. Numerous studies have indicated that ARFs are involved in many tomato developmental processes. SlARF4 negatively regulates chlorophyll accumulation and starch biosynthesis in tomato fruit. Our previous study showed that SlARF10 positively regulated chlorophyll accumulation via direct activation of the expression of SlGLK1. Downregulation of ARF6 and ARF8 by over expression of Arabidopsis microRNA167 results in the failure of pollen germination on the stigma surface and/or growth through the style in tomato. However, the function of SlARF6 in the regulation of fruit development is still not well understood. In this study, SlARF6A was found to be involved in photosynthesis, sugar accumulation and fruit development in tomato. Our data demonstrate that SlARF6A plays an important role in the regulation of fruit quality and development.The SlARF6A gene has an open reading frame of 2608 bp encoding a putative protein of 869 amino acids. Amino acid sequence analysis revealed that, like SlARF7 and SlARF8, which have typical conserved ARF domains, SlARF6A protein also contained B3-DNA, ARF, and AUX/IAA binding domains . A phylogenetic tree was constructed to gain insight into the phylogenetic relationship among ARF proteins in Arabidopsis and tomato. ARFs were divided into four major classes: I, II, III, and VI. SlARF6A along with SlARF6B and AtARF6 were grouped into subclass IIa and are closely related to AtARF8 and SlARF8 , indicating possible functional similarity among them. To determine the expression pattern of SlARF6A in planta, a transcriptional fusion was constructed between the SlARF6A promoter and the GUS reporter gene. GUS staining in the transgenic tomato plants was detected in leaves, stems, buds, flowers, and fruits at different developmental stages, an indication of the ubiquitous expression of SlARF6A in all tissues tested. The GUS staining was weak in the early fruits at 2 and 4 days post anthesis but became strong at 8, 30 and 45 DPA , suggesting possible roles of SlARF6A in the development of tomato fruits. To examine its subcellular localization in plants, SlARF6A was fused to GFP and transferred into tobacco protoplasts. Fluorescence microscopy analysis revealed that SlARF6A was specifically localized in the nuclei . A GAL4-responsive reporter system in yeast was employed to reveal the transcriptional activity of SlARF6A. SlARF6A was fused to GAL4-BD to form a pGBKT7-SlARF6A fusion plasmid and subsequently transformed into yeast. Yeast transformants harboring the pGBKT7-SlARF6A construct grew well in the medium lacking Trp, His, and Ade , while the yeasts transformed with pGBKT7 vector alone could not . Assessing transcriptional activity revealed that SlARF6A is a transcriptional activator.To elucidate the physiological significance of the SlARF6A gene in fruit development, upregulated and downregulated transgenic lines corresponding to independent transformation events were generated in tomato plants. qRT-PCR was used to evaluate the expression level of SlARF6A in all transgenic lines. Compared with thelevel in the wild type , the expression level of SlARF6A was decreased in RNAi 2 and 6 plants but increased in OE-4 and 6 plants . It is noteworthy that altered SlARF6A expression led to a dramatic change in chlorophyll accumulation in transgenic lines. Compared with WT plants, the OE-SlARF6A plants had dark-green fruits at the green fruit stage, whereas the RNAi-SlARF6A plants had light-green fruits . The impact of altered SlARF6A expression on chlorophyll accumulation was analyzed by measuring the chlorophyll content in fruits and leaves. The SlARF6A overexpression lines possessed greater accumulation of chlorophyll in the fruits at immature green, mature green, breaker, and orange stages, whereas the RNAi lines had lower chlorophyll accumulation in the fruits at immature green and mature green stages than the WT plants .

However, using 9 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 occuring 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 , while 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 U.S., Brazil, and Italy when compared to 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 lab populations from Japan and Hawaii have half as much pairwise diversity as the wild South Korean population, consistent with as mall lab 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, plant pots with drainage which can occur during a population shrinkage, while 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 be statistically different except for Western U.S. 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 one 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 U.S. 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 geographic 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 U.S. 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 U.S., 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 U.S., we were surprised to discover a lack of structure on a finer scale, either based on latitude or simple geographic distance, given the large number of loci analyzed. In a similar study using 3,484 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 U.S. 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 U.S. 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 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 U.S., 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 U.S., European, Asian, and Brazilian populations varied between 8.86% to 9.02%. However, we were surprised to see that our Italian and Irish samples had much higher values of Fst compared to 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, plastic plants pots we find that our treemix and migration results largely coincide with the proposed invasion pathway inferred from microsatellites , as well as a recent pre-print that reanalyzed invasion pathways with pooled sequencing data . We see that European and U.S./Brazilian populations form two distinct clades, 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 U.S., then the Eastern U.S. and Brazil. Additionally, in the Western U.S., 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 U.S./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 U.S. 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 U.S. 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 outgroup . 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 ofinvasive 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 U.S. 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 non-variable 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.Tuta absoluta is a worldwide economic pest of tomatoes and other solanaceous crops. A member of the gelechiid family, this moth lays eggs on the above-ground portion of the plant, where the hatched larvae will spend their lives creating “mines” throughout the plant tissue before pupating and emerging as adults . At a reproduction rate of up to 10 generations per year, untreated infestations will eventually result in complete death of the plant, leading to up to 100% agricultural loss. Although a large effort has been made to develop and implement integrated pest management programs across different world regions , typical treatments have included heavy use of a variety of insecticides , leading to the rapid appearance of insecticide resistance. As tomatoes represent a massive economic industry, with an estimated 252 million metric tons of tomatoes harvested in 2020 , there is a serious need to understand the invasive biology of this insect and to develop tools for detection and prevention. Tuta absoluta was originally detected in Peru in 1917 but was not recorded as an agricultural pest until the 1960s and 70s when it was discovered in tomato fields in Chile, Argentina, and Venezuela; by the 1990s it was widespread across South America. In 2006, Tuta absoluta appeared in Spain ; since then it has rapidly colonized Europe, Asia, and Africa. It is generally believed that the Peruvian highlands is the ancestral home of T. absoluta, and that the rapid colonization to the rest of Latin America was due to the introduction of Tuta absoluta by human transport of contaminated fruit, although few studies have confirmed this .