Validity and Reliability of the Student Refractive Error and Eyeglasses Questionnaire-Revised Version in Myopic Children.

SIGNIFICANCE: Multiple vision-related quality of life (VRQol) instruments exist, but questionnaires designed specifically for myopic children that are appropriate for assessing the impact of refractive error are rare. PURPOSE: This study aimed to assess the validity and reliability of the Student Refractive Error and Eyeglasses Questionnaire - Revised (SREEQ-R) in school-aged children with myopia in the United Kingdom. METHODS: Community optometrists in the United Kingdom invited children up to the age of 18 years presenting for an eye examination with current or previous reported use of eyeglasses for myopia to complete the SREEQ-R, which consists of 20 specific items divided into two sections each with three response categories. The "without glasses" section relates to perceptions of uncorrected vision/not wearing glasses, and the "with glasses" relates to corrected vision/wearing glasses. Rasch analysis was used to explore the psychometric performance (content, construct validity, and reliability) of the questionnaire items and scale using Winsteps software (Winsteps.com. Portland, OR). RESULTS: A total of 125 eligible children with a mean ± standard deviation age of 12.7 ± 2.9 years completed the SREEQ-R. All items fit the Rasch model and were retained, and the scale was found to be unidimensional. All children and item infit and outfit mean square statistics fell within the recommended fit criteria. As per the Rasch analysis, the person reliability coefficients were 0.84 and 0.91, whereas item reliabilities were 0.99 and 0.80 for the without glasses and with glasses sections, respectively. The internal consistency for the SREEQ-R was good; Cronbach α values were 0.84 for without glasses and 0.91 for with glasses. CONCLUSIONS: The SREEQ-R had satisfactory validity and reliability evidence. Construct validity of the scale was supported to measure the impact of uncorrected and corrected refractive error on vision-related quality of life in myopic school-aged children in the United Kingdom. The SREEQ-R could be used in future studies to evaluate vision-related quality of life in children with myopia.

Atg2, Atg9 and Atg18 in mitochondrial integrity, cardiac function and healthspan in Drosophila.

In yeast, the Atg2-Atg18 complex regulates Atg9 recycling from phagophore assembly site during autophagy; their function in higher eukaryotes remains largely unknown. In a targeted screening in Drosophila melanogaster, we show that Mef2-GAL4-RNAi-mediated knockdown of Atg2, Atg9 or Atg18 in the heart and indirect flight muscles led to shortened healthspan (declined locomotive function) and lifespan. These flies displayed an accelerated age-dependent loss of cardiac function along with cardiac hypertrophy (increased heart tube wall thickness) and structural abnormality (distortion of the lumen surface). Using the Mef2-GAL4-MitoTimer mitochondrial reporter system and transmission electron microscopy, we observed significant elongation of mitochondria and reduced number of lysosome-targeted autophagosomes containing mitochondria in the heart tube but exaggerated mitochondrial fragmentation and reduced mitochondrial density in indirect flight muscles. These findings provide the first direct evidence of the importance of Atg2-Atg18/Atg9 autophagy complex in the maintenance of mitochondrial integrity and, regulation of heart and muscle functions in Drosophila, raising the possibility of augmenting Atg2-Atg18/Atg9 activity in promoting mitochondrial health and, muscle and heart function.

The Gene Expression Program for the Formation of Wing Cuticle in Drosophila.

The cuticular exoskeleton of insects and other arthropods is a remarkably versatile material with a complex multilayer structure. We made use of the ability to isolate cuticle synthesizing cells in relatively pure form by dissecting pupal wings and we used RNAseq to identify genes expressed during the formation of the adult wing cuticle. We observed dramatic changes in gene expression during cuticle deposition, and combined with transmission electron microscopy, we were able to identify candidate genes for the deposition of the different cuticular layers. Among genes of interest that dramatically change their expression during the cuticle deposition program are ones that encode cuticle proteins, ZP domain proteins, cuticle modifying proteins and transcription factors, as well as genes of unknown function. A striking finding is that mutations in a number of genes that are expressed almost exclusively during the deposition of the envelope (the thin outermost layer that is deposited first) result in gross defects in the procuticle (the thick chitinous layer that is deposited last). An attractive hypothesis to explain this is that the deposition of the different cuticle layers is not independent with the envelope instructing the formation of later layers. Alternatively, some of the genes expressed during the deposition of the envelope could form a platform that is essential for the deposition of all cuticle layers.

ChtVis-Tomato, a genetic reporter for in vivo visualization of chitin deposition in Drosophila.

Chitin is a polymer of N-acetylglucosamine that is abundant and widely found in the biological world. It is an important constituent of the cuticular exoskeleton that plays a key role in the insect life cycle. To date, the study of chitin deposition during cuticle formation has been limited by the lack of a method to detect it in living organisms. To overcome this limitation, we have developed ChtVis-Tomato, an in vivo reporter for chitin in Drosophila. ChtVis-Tomato encodes a fusion protein that contains an apical secretion signal, a chitin-binding domain (CBD), a fluorescent protein and a cleavage site to release it from the plasma membrane. The chitin reporter allowed us to study chitin deposition in time lapse experiments and by using it we have identified unexpected deposits of chitin fibers in Drosophila pupae. ChtVis-Tomato should facilitate future studies on chitin in Drosophila and other insects.

The Drosophila planar polarity gene multiple wing hairs directly regulates the actin cytoskeleton.

The evolutionarily conserved frizzled/starry night (fz/stan) pathway regulates planar cell polarity (PCP) in vertebrates and invertebrates. This pathway has been extensively studied in the Drosophila wing, where it is manifested by an array of distally pointing cuticular hairs. Using in vivo imaging we found that, early in hair growth, cells have multiple actin bundles and hairs that subsequently fuse into a single growing hair. The downstream PCP gene multiple wing hairs (mwh) plays a key role in this process and acts to antagonize the actin cytoskeleton. In mwh mutants hair initiation is not limited to a small region at the distal edge of pupal wing cells as in wild type, resulting in multiple hairs with aberrant polarity. Extra actin bundles/hairs are formed and do not completely fuse, in contrast to wild type. As development proceeded additional hairs continued to form, further increasing hair number. We identified a fragment of Mwh with in vivo rescue activity and that bound and bundled F-actin filaments and inhibited actin polymerization in in vitro actin assays. The loss of these activities can explain the mwh mutant phenotype. Our data suggest a model whereby, prior to hair initiation, proximally localized Mwh inhibits actin polymerization resulting in polarized activation of the cytoskeleton and hair formation on the distal side of wing cells. During hair growth Mwh is found in growing hairs, where we suggest it functions to promote the fusion of actin bundles and inhibit the formation of additional actin bundles that could lead to extra hairs.

Measuring and mitigating agricultural greenhouse gas production in the US Great Plains, 1870-2000.

The Great Plains region of the United States is an agricultural production center for the global market and, as such, an important source of greenhouse gas (GHG) emissions. This article uses historical agricultural census data and ecosystem models to estimate the magnitude of annual GHG fluxes from all agricultural sources (e.g., cropping, livestock raising, irrigation, fertilizer production, tractor use) in the Great Plains from 1870 to 2000. Here, we show that carbon (C) released during the plow-out of native grasslands was the largest source of GHG emissions before 1930, whereas livestock production, direct energy use, and soil nitrous oxide emissions are currently the largest sources. Climatic factors mediate these emissions, with cool and wet weather promoting C sequestration and hot and dry weather increasing GHG release. This analysis demonstrates the long-term ecosystem consequences of both historical and current agricultural activities, but also indicates that adoption of available alternative management practices could substantially mitigate agricultural GHG fluxes, ranging from a 34% reduction with a 25% adoption rate to as much as complete elimination with possible net sequestration of C when a greater proportion of farmers adopt new agricultural practices.

Implications of Observed and Simulated Soil Carbon Sequestration for Management Options in Corn-based Rotations.

Managing cropping systems to sequester soil organic C (SOC) improves soil health and resilience to changing climate. Perennial crops, no-till planting, manure, and cover crops can add SOC; however, their impacts have not been well documented in the northeastern United States. Our objectives were (i) to monitor SOC from a bioenergy cropping study in Pennsylvania that included a corn ( L.)-soybean [ (L.) Merr.]-alfalfa ( L.) rotation, switchgrass ( L.), and reed canarygrass ( L.); (ii) to use the CQESTR model to predict SOC sequestration in the bioenergy crops (with and without projected climate change); and (iii) to use CQESTR to simulate influence of tillage, manure, cover cropping, and corn stover removal in typical dairy forage (silage corn-alfalfa) or grain corn-soybean rotations. Over 8 yr, measured SOC increased 0.4, 1.1, and 0.8 Mg C ha yr in the bioenergy rotation, reed canarygrass, and switchgrass, respectively. Simulated and measured data were significantly correlated ( < 0.001) at all depths. Predicted sequestration (8-14 Mg C ha over 40 yr) in dairy forage rotations was much larger than with corn-soybean rotations (-4.0-0.6 Mg C ha over 40 yr), due to multiple years of perennial alfalfa. No-till increased sequestration in the simulated dairy forage rotation and prevented a net loss of C in corn-soybean rotations. Simulations indicated limited impact of cover crops and manure on long-term SOC sequestration. The low solids content of liquid dairy manure is the likely reason for the less-than-expected impact of manure. Overall, simulations suggest that inclusion of alfalfa provides the greatest potential for SOC sequestration with a typical Pennsylvania crop rotation.

The proteins encoded by the Drosophila Planar Polarity Effector genes inturned, fuzzy and fritz interact physically and can re-pattern the accumulation of "upstream" Planar Cell Polarity proteins.

The frizzled/starry night pathway regulates planar cell polarity in a wide variety of tissues in many types of animals. It was discovered and has been most intensively studied in the Drosophila wing where it controls the formation of the array of distally pointing hairs that cover the wing. The pathway does this by restricting the activation of the cytoskeleton to the distal edge of wing cells. This results in hairs initiating at the distal edge and growing in the distal direction. All of the proteins encoded by genes in the pathway accumulate asymmetrically in wing cells. The pathway is a hierarchy with the Planar Cell Polarity (PCP) genes (aka the core genes) functioning as a group upstream of the Planar Polarity Effector (PPE) genes which in turn function as a group upstream of multiple wing hairs. Upstream proteins, such as Frizzled accumulate on either the distal and/or proximal edges of wing cells. Downstream PPE proteins accumulate on the proximal edge under the instruction of the upstream proteins. A variety of types of data support this hierarchy, however, we have found that when over expressed the PPE proteins can alter both the subcellular location and level of accumulation of the upstream proteins. Thus, the epistatic relationship is context dependent. We further show that the PPE proteins interact physically and can modulate the accumulation of each other in wing cells. We also find that over expression of Frtz results in a marked delay in hair initiation suggesting that it has a separate role/activity in regulating the cytoskeleton that is not shared by other members of the group.

A novel MitoTimer reporter gene for mitochondrial content, structure, stress, and damage in vivo.

Mitochondrial dysfunction plays important roles in many diseases, but there is no satisfactory method to assess mitochondrial health in vivo. Here, we engineered a MitoTimer reporter gene from the existing Timer reporter gene. MitoTimer encodes a mitochondria-targeted green fluorescent protein when newly synthesized, which shifts irreversibly to red fluorescence when oxidized. Confocal microscopy confirmed targeting of the MitoTimer protein to mitochondria in cultured cells, Caenorhabditis elegans touch receptor neurons, Drosophila melanogaster heart and indirect flight muscle, and mouse skeletal muscle. A ratiometric algorithm revealed that conditions that cause mitochondrial stress led to a significant shift toward red fluorescence as well as accumulation of pure red fluorescent puncta of damaged mitochondria targeted for mitophagy. Long term voluntary exercise resulted in a significant fluorescence shift toward green, in mice and D. melanogaster, as well as significantly improved structure and increased content in mouse FDB muscle. In contrast, high-fat feeding in mice resulted in a significant shift toward red fluorescence and accumulation of pure red puncta in skeletal muscle, which were completely ameliorated by voluntary wheel running. Hence, MitoTimer allows for robust analysis of multiple parameters of mitochondrial health under both physiological and pathological conditions and will be highly useful for future research of mitochondrial health in multiple disciplines in vivo.

Dusky-like functions as a Rab11 effector for the deposition of cuticle during Drosophila bristle development.

The morphogenesis of Drosophila sensory bristles is dependent on the function of their actin and microtubule cytoskeleton. Actin filaments are important for bristle shape and elongation, while microtubules are thought to mediate protein and membrane trafficking to promote growth. We have identified an essential role for the bristle cuticle in the maintenance of bristle structure and shape at late stages of bristle development. We show that the small GTPase Rab11 mediates the organized deposition of chitin, a major cuticle component in bristles, and disrupting Rab11 function leads to phenotypes that result from bristle collapse rather than a failure to elongate. We further establish that Rab11 is required for the plasma membrane localization of the ZP domain-containing Dusky-like (Dyl) protein and that Dyl is also required for cuticle formation in bristles. Our data argue that Dyl functions as a Rab11 effector for mediating the attachment of the bristle cell membrane to chitin to establish a stable cuticle. Our studies also implicate the exocyst as a Rab11 effector in this process and that Rab11 trafficking along the bristle shaft is mediated by microtubules.

Integrating biorefinery and farm biogeochemical cycles offsets fossil energy and mitigates soil carbon losses.

Crop residues are potentially significant sources of feedstock for biofuel production in the United States. However, there are concerns with maintaining the environmental functions of these residues while also serving as a feedstock for biofuel production. Maintaining soil organic carbon (SOC) along with its functional benefits is considered a greater constraint than maintaining soil erosion losses to an acceptable level. We used the biogeochemical model DayCent to evaluate the effect of residue removal, corn stover, and wheat and barley straw in three diverse locations in the USA. We evaluated residue removal with and without N replacement, along with application of a high-lignin fermentation byproduct (HLFB), the residue by-product comprised of lignin and small quantities of nutrients from cellulosic ethanol production. SOC always decreased with residue harvest, but the decrease was greater in colder climates when expressed on a life cycle basis. The effect of residue harvest on soil N2O emissions varied with N addition and climate. With N addition, N2O emissions always increased, but the increase was greater in colder climates. Without N addition, N2O emissions increased in Iowa, but decreased in Maryland and North Carolina with crop residue harvest. Although SOC was lower with residue harvest when HLFB was used for power production instead of being applied to land, the avoidance of fossil fuel emissions to the atmosphere by utilizing the cellulose and hemicellulose fractions of crop residue to produce ethanol (offsets) reduced the overall greenhouse gas (GHG) emissions because most of this residue carbon would normally be lost during microbial respiration. Losses of SOC and reduced N mineralization could both be mitigated with the application of HLFB to the land. Therefore, by returning the high-lignin fraction of crop residue to the land after production of ethanol at the biorefinery, soil carbon levels could be maintained along with the functional benefit of increased mineralized N, and more GHG emissions could be offset compared to leaving the crop residues on the land.

Quality Control and Reproducibility in M-Mode, Two-Dimensional, and Speckle Tracking Echocardiography Acquisition and Analysis: The CARDIA Study, Year 25 Examination Experience.

INTRODUCTION: Few large studies describe quality control procedures and reproducibility findings in cardiovascular ultrasound, particularly in novel techniques such as speckle tracking echocardiography (STE). We evaluate the echocardiography assessment performance in the Coronary Artery Risk Development in Young Adults (CARDIA) study Year 25 (Y25) examination (2010-2011) and report findings from a quality control and reproducibility program conducted to assess Field Center image acquisition and reading center (RC) accuracy. METHODS: The CARDIA Y25 examination had 3475 echocardiograms performed in 4 US Field Centers and analyzed in a RC, assessing standard echocardiography (LA dimension, aortic root, LV mass, LV end-diastolic volume [LVEDV], ejection fraction [LVEF]), and STE (two- and four-chamber longitudinal, circumferential, and radial strains). Reproducibility was assessed using intraclass correlation coefficients (ICC), coefficients of variation (CV), and Bland-Altman plots. RESULTS: For standard echocardiography reproducibility, LV mass and LVEDV consistently had CV above 10% and aortic root below 6%. Intra-sonographer aortic root and LV mass had the most robust values of ICC in standard echocardiography. For STE, the number of properly tracking segments was above 80% in short-axis and four-chamber and 58% in two-chamber views. Longitudinal strain parameters were the most robust and radial strain showed the highest variation. Comparing Field Centers with echocardiography RC STE readings, mean differences ranged from 0.4% to 4.1% and ICC from 0.37 to 0.66, with robust results for longitudinal strains. CONCLUSION: Echocardiography image acquisition and reading processes in the CARDIA study were highly reproducible, including robust results for STE analysis. Consistent quality control may increase the reliability of echocardiography measurements in large cohort studies.

dusky-like is required to maintain the integrity and planar cell polarity of hairs during the development of the Drosophila wing.

The cuticular hairs and sensory bristles that decorate the adult Drosophila epidermis and the denticles found on the embryo have been used in studies on planar cell polarity and as models for the cytoskeletal mediated morphogenesis of cellular extensions. ZP domain proteins have recently been found to be important for the morphogenesis of both denticles and bristles. Here we show that the ZP domain protein Dusky-like is a key player in hair morphogenesis. As is the case in bristles, in hairs dyl mutants display a dramatic phenotype that is the consequence of a failure to maintain the integrity of the extension after outgrowth. Hairs lacking dyl function are split, thinned, multipled and often very short. dyl is required for normal chitin deposition in hairs, but chitin is not required for the normal accumulation of Dyl, hence dyl acts upstream of chitin. A lack of chitin however, does not mimic the dyl hair phenotype, thus Dyl must have other targets in hair morphogenesis. One of these appears to be the actin cytoskeleton. Interestingly, dyl mutants also display a unique planar cell polarity phenotype that is distinct from that seen with mutations in the frizzled/starry night or dachsous/fat pathway genes. Rab11 was previously found to be essential for Dyl plasma membrane localization in bristles. Here we found that the expression of a dominant negative Rab11 can mimic the dyl hair morphology phenotype consistent with Rab11 also being required for Dyl function in hairs. We carried out a small directed screen to identify genes that might function with dyl and identified Chitinase 6 (Cht6) as a strong candidate, as knocking down Cht6 function led to weak versions of all of the dyl hair phenotypes.

The LTAR Common Experiment at Upper Chesapeake Bay: Integrated.

Corn (Zea mays) crops harvested as grain in autumn do not provide opportunity for cover crop establishment, which may be remedied by interseeding cover crops into growing corn. Grazing cover crops after corn grain harvest could provide added revenues and increase nutrient cycling in the system while providing additional ecosystem services. However, tradeoffs between cash crop productivity and cover crop inclusion, and use as grazed forage, are not fully understood. This 4-year Long-Term Agroecosystem Research Integrated Common Experiment project evaluated the effect of interseeding cereal rye (Secale cereale) into corn for grazing after corn grain harvest on corn grain yield and late-season grazing. Cereal rye was interseeded into corn in early June. After corn grain harvest, six paddocks at each location were randomly allotted to grazed (GRAZ) or not grazed (NG). The GRAZ paddocks were grazed with beef cattle in late autumn and again in early spring if regrowth allowed. Paddocks were flown with an unmanned aerial system (UAS) to characterize spatial forage yield and quality. Cereal rye provided an additional 20-30 grazing days in the autumn for 24 beef cows on 4.8 ha. Early spring growth shows potential to provide even greater forage yields than autumn, but growth is less dependable. Corn grain yields did not decrease except in 2019 (dry year) when yields were 40% lower. There were no significant differences in soil health indicators between GRAZ and NG paddocks. The UAS shows promise as a tool for monitoring forage yield and quality and optimizing grazing management.

The LTAR Cropland Common Experiment in the Upper Chesapeake Bay.

Dairy production is a key agricultural enterprise in the Upper Chesapeake Bay (UCB) basin, where phosphorous (P) and nitrogen (N) loading contribute to eutrophication. Import of forages and grains and application of mineral fertilizers contribute to nutrient imbalances in the basin. The UCB Long-Term Agroecosystem Research Cropland Common Experiment aims to evaluate diverse crop rotations that minimize the need for imported feed, maximize year-round living cover, and reduce nutrient losses. UCB's plot-scale experiment was established in 2018, incorporating an ongoing cropping system study that was established by the Pennsylvania State University in 2010. An alternative dairy cropping rotation (including silage and grain corn [Zea mayes L.], alfalfa [Medicago sativa L.]/orchardgrass [Dactylis glomerata L.] mix, winter rye silage [Secale cereale L.], and sorghum-sudangrass [Sorghum × drummondii (Steud.) Millsp. & Chas]) that employed manure injection, integrated pest management, and less frequent manure application was compared to a prevailing, conventionally managed silage corn-alfalfa rotation with higher manure application rates. A field-scale experiment was established in 2019 to monitor alternative production practices (manure injection and avoidance of neonicotinoid seed treatment) and prevailing practices in three fields on a commercial dairy farm. Findings suggest that crop rotation diversification, manure injection, and integrated pest management have the potential to increase the economic and environmental sustainability of dairy cropping systems, but long-term evaluation is needed for confirmation.

Life cycle inventory of Miscanthus production on a commercial farm in the US.

There has been considerable interest in use of Miscanthus (Miscanthus x giganteus) as a feedstock for bioenergy production due to its potential to reduce greenhouse gas emissions associated with cellulosic feedstock production and more recently for alternative uses as a biomass crop. To date, data on Miscanthus production in the US has been based on small scale research plots due to the lack of commercial scale production fields. Research plot yields are often much higher than commercial fields for a variety of reasons including reduced spatial variability and location on better quality farmland. The objectives of this study were to quantify the inputs for production of Miscanthus at the commercial farm scale, evaluating methods to characterize fuel use for establishment and management of Miscanthus production and using satellite data to characterize spatial yield variation of production fields. We logged energy use on agricultural machinery from Miscanthus production planted on more than 1000 ha of land and modeled N2O emissions and changes in soil carbon using DayCent. Although fuel use was higher for land preparation in fields with perennial vegetation, fuel to harvest Miscanthus dominated greenhouse gas (GHG) emissions (>90%) from agriculture machinery for crop management. The N2O emissions and changes in soil carbon were the largest source and sink of GHG emissions associated with Miscanthus production, respectively. Although ~ 50% of the established lands had Miscanthus yields < 5 Mg/ha, yields needed to be > 5 Mg/ha for ΔSOC to be positive. Given the large impact of yield on ΔSOC, net GHG for Miscanthus production with yields of 5 to 25 Mg/ha ranged ~130 to -260 kg CO2e/Mg biomass. Use of both energy use for Miscanthus harvest and satellite imagery were good methods to characterize spatial variability of commercial production fields. This demonstrates the potential to use this within field yield data to better understand factors driving subfield yield variability and use of satellite data to quantify early yield predictions.

Densified biomass can cost-effectively mitigate greenhouse gas emissions and address energy security in thermal applications.

Regional supplies of biomass are currently being evaluated as feedstocks in energy applications to meet renewable portfolio (RPS) and low carbon fuel standards. We investigate the life cycle greenhouse gas (GHG) emissions and associated abatement costs resulting from using densified switchgrass for thermal and electrical energy. In contrast to the large and positive abatement costs for using biomass in electricity generation ($149/Mg CO(2)e) due to the low cost of coal and high feedstock and power plant operation costs, abatement costs for replacing fuel oil with biomass in thermal applications are large and negative (-$52 to -$92/Mg CO(2)e), resulting in cost savings. Replacing fuel oil with biomass in thermal applications results in least cost reductions compared to replacing coal in electricity generation, an alternative that has gained attention due to RPS legislation and the centralized production model most often considered in U.S. policy. Our estimates indicate a more than doubling of liquid fuel displacement when switchgrass is substituted for fuel oil as opposed to gasoline, suggesting that, in certain U.S. locations, such as the northeast, densified biomass would help to significantly decarbonize energy supply with regionally sourced feedstock, while also reducing imported oil. On the basis of supply projections from the recently released Billion Ton Report, there will be enough sustainably harvested biomass available in the northeast by 2022 to offset the entirety of heating oil demand in the same region. This will save NE consumers between $2.3 and $3.9 billion annually. Diverting the same resource to electricity generation would cost the region $7.7 billion per year. While there is great need for finding low carbon substitutes for coal power and liquid transportation fuels in the U.S., we argue that in certain regions it makes cost- (and GHG mitigation-) effective sense to phase out liquid heating fuels with locally produced biomass first.