Nanostructural differences in pectic polymers isolated from strawberry fruits with low expression levels of pectate lyase or polygalacturonase genes

Our research group has obtained transgenic strawberry plants expressing antisense sequences of either a pectate lyase (APEL lines) [1] or a polygalacturonase gene (APG lines) [2]. Both genes encode ripening-specific endo-pectinases with a common target, deesterified homogalacturonans, but each enzyme act by a different mechanism and pH range. Ripe fruits from both transgenic genotypes were significantly firmer than control, being APG fruits on average 25% firmer than APEL fruits. Cell wall analysis of both transgenic genotypes indicated that pectin fractions extracted with CDTA and sodium carbonate were significantly modified in transgenic fruits [2,3]. To gain insight in the role of these pectinases in pectin disassembly during ripening, CDTA and Na2CO3 pectins have been analyzed by atomic force microscopy (AFM). APEL and APG CDTA pectins had similar contour lengths but both were significantly longer than control. Similarly, APG carbonate chains were longer than control, showing APEL carbonate chains an intermediate length. Furthermore, transgenic pectins displayed a more complex branching pattern and a higher number of micellar aggregates, especially in the sodium carbonate fractions of APG samples. Acid hydrolysis of carbonate pectins reduced the number of micellar aggregates. AFM analyses confirm that the inhibition of both pectinases reduces pectin disassembly, and also suggest that each pectinase acts on specific pectin domains. Particularly, polygalacturonase silencing induces more significant pectin modifications, nicely correlated with the firmer phenotype of APG fruits, than the down-regulation of pectate lyase

Estimating HIV Medication Adherence and Persistence: Two Instruments for Clinical and Research Use

Antiretroviral therapy (ART) requires lifelong daily oral therapy. While patient characteristics associated with suboptimal ART adherence and persistence have been described in cohorts of HIV-infected persons, these factors are poor predictors of individual medication taking behaviors. We aimed to create and test instruments for the estimation of future ART adherence and persistence for clinical and research applications. Following formative work, a battery of 148 items broadly related to HIV infection and treatment was developed and administered to 181 HIV-infected patients. ART adherence and persistence were assessed using electronic monitoring for 3 months. Perceived confidence in medication taking and self-reported barriers to adherence were strongest in predicting non-adherence over time. Barriers to adherence (e.g., affordability, scheduling) were the strongest predictors of non-adherence, as well as 3- and 7-day non-persistence. A ten-item battery for prediction of these outcomes (www.med.unc.edu/ncaidstraining/adherence/for-providers) and a 30-item battery reflective of underlying psychological constructs can help identify and study individuals at risk for suboptimal ART adherence and persistence

Nanostructural changes in cell wall pectins during strawberry fruit ripening assessed by atomic force microscopy

Rapid loss of firmness occurs during strawberry (Fragaria × ananassa Duch) ripening, resulting in a short shelf life and high economic losses. The disassembly of cell walls is considered the main responsible for fruit softening, being pectins extensively modified during strawberry ripening (Paniagua et al. 2014). Atomic force microscopy allows the analysis of individual polymer chains at nanostructural level with a minimal sample preparation (Morris et al., 2001). The main objective of this research was to compare pectins of green and red ripe strawberry fruits at the nanostructural level to shed light on structural changes that could be related to softening. Cell walls from strawberry fruits were extracted and fractionated with different solvents to obtain fractions enriched in a specific component. The yield of cell wall material, as well as the amount of the different fractions, decreased in ripe fruits. CDTA and Na2CO3 fractions underwent the largest decrements, being these fractions enriched in pectins supposedly located in the middle lamella and primary cell wall, respectively. Uronic acid content also decreased significantly during ripening in both pectin fractions, but the amount of soluble pectins, those extracted with phenol:acetic acid:water (PAW) and water increased in ripe fruits. Monosaccharide composition in CDTA and Na2CO3 fractions was determined by gas chromatography. In both pectin fractions, the amount of Ara and Gal, the two most abundant carbohydrates, decreased in ripe fruits. The nanostructural characteristics of CDTA and Na2CO3 pectins were analyzed by AFM. Isolated pectic chains present in the CDTA fraction were significantly longer and more branched in samples from green fruits than those present in samples obtained from red fruit. In spite of slight differences in length distributions, Na2CO3 samples from unripe fruits displayed some longer chains at low frequency that were not detected in ripe fruits. Pectin aggregates were more frequently observed in green fruit samples from both fractions. These results support that pectic chain length and the nanostructural complexity of the pectins present in CDTA and Na2CO3 fractions diminish during strawberry fruit development, and these changes, jointly with the loss of neutral sugars, could contribute to the solubilization of pectins and fruit softening. Paniagua et al. (2014). Ann Bot, 114: 1375-1383 Morris et al. (2001). Food Sci Tech 34: 3-10 This research was supported by FEDER EU Funds and the Ministerio de Educación y Ciencia of Spain (grant reference AGL2011-24814)Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Unravelling the nanostructure of strawberry fruit pectins by atomic force microscopy

Atomic force microscopy (AFM) allows the analysis of individual polymers at nanostructural level with a minimal sample preparation. This technique has been used to analyse the pectin disassembly process during the ripening and postharvest storage of several fleshy fruits. In general, pectins analysed by AFM are usually visualized as isolated chains, unbranched or with a low number of branchs and, occasionally, as large aggregates. However, the exact nature of these structures is unknown. It has been suggested that pectin aggregates represent a mixture of rhamnonogalacturonan I and homogalacturonan, while isolated chains and their branches are mainly composed by polygalacturonic acid. In order to gain insight into the nature of these structures, sodium carbonate soluble pectins from ripe strawberry (Fragaria x ananassa, Duch.) fruits were subjected to enzymatic digestion with endo-Polygalacturonase M2 from Aspergillus aculeatus, and the samples visualized by AFM at different time intervals. Pectins isolated from control, non-transformed plants, and two transgenic genotypes with low level of expression of ripening-induced pectinase genes encoding a polygalacturonase (APG) or a pectate lyase (APEL) were also included in this study. Before digestion, isolated pectin chains from control were shorter than those from transgenic fruits, showing number-average (LN) contour length values of 73.2 nm vs. 95.9 nm and 91.4 nm in APG and APEL, respectively. The percentage of branched polymers was significantly higher in APG polyuronides than in the remaining genotypes, 33% in APG vs. 6% in control and APEL. As a result of the endo-PG treatment, a gradual decrease in the main backbone length of isolated chains was observed in the three samples. The minimum LN value was reached after 8 h of digestion, being similar in the three genotypes, 22 nm. By contrast, the branches were not visible after 1.5-2 h of digestion. LN values were plotted against digestion time and the data fitted to a first-order exponential decay curve, obtaining R2 values higher than 0.9. The half digestion time calculated with these equations were similar for control and APG pectins, 1.7 h, but significantly higher in APEL, 2.5 h, indicating that these polymer chains were more resistant to endo-PG digestion. Regarding the pectin aggregates, their volumes were estimated and used to calculate LN molecular weights. Before digestion, control and APEL samples showed complexes of similar molecular weights, 1722 kDa, and slightly higher than those observed in APG samples. After endo-PG digestion, size of complexes diminished significantly, reaching similar values in the three pectin samples, around 650 kDa. These results suggest that isolated polymer chains visualized by AFM are formed by a HG domain linked to a shorter polymer resistant to endo-PG digestion, maybe xylogalacturonan or RG-I. The silencing of the pectate lyase gene slightly modified the structure and/or chemical composition of polymer chains making these polyuronides more resistant to enzymatic degradation. Similarly, polygalacturonic acid is one of the main component of the aggregates.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

The Economic and Fiscal Impact of a Microgrid in Downtown Cleveland, Ohio

This report relates the results of an investigation into market conditions for a proposed microgrid in downtown Cleveland, Ohio, as well the potential for additional jobs, income, and tax revenues that might accompany such an enterprise. Power interruptions have been estimated to cost commercial and industrial customers more than $100 billion each year in the United States.1 Because microgrids can reduce or eliminate power disruptions, the proposed microgrid could position Cleveland to capture growth among those industries that experience relatively greater losses when power outages occur. This includes momentary interruptions, which account for a “substantial portion”2 of such costs. The improved quality, reliability, resiliency, and security associated with a Cleveland microgrid could offer a locational advantage in attracting companies for which a power interruption is particularly costly. Access to clean, distributed generation is also an attribute that is of significant interest to commercial end users

The Economic and Fiscal Impact of a Microgrid in Downtown Cleveland, Ohio

This report relates the results of an investigation into market conditions for a proposed microgrid in downtown Cleveland, Ohio, as well the potential for additional jobs, income, and tax revenues that might accompany such an enterprise. Power interruptions have been estimated to cost commercial and industrial customers more than $100 billion each year in the United States.1 Because microgrids can reduce or eliminate power disruptions, the proposed microgrid could position Cleveland to capture growth among those industries that experience relatively greater losses when power outages occur. This includes momentary interruptions, which account for a “substantial portion”2 of such costs. The improved quality, reliability, resiliency, and security associated with a Cleveland microgrid could offer a locational advantage in attracting companies for which a power interruption is particularly costly. Access to clean, distributed generation is also an attribute that is of significant interest to commercial end users

Metal ions and graphene-based compounds as alternative treatment options for burn wounds infected by antibiotic-resistant Pseudomonas aeruginosa.

Burn infections caused by Pseudomonas aeruginosa pose a major complication in wound healing. This study aimed to determine the antimicrobial effect of metal ions, graphene (Gr), and graphene oxide (GO), individually and in combination, against the planktonic and biofilm states of two antimicrobially resistant clinical strains of P. aeruginosa each with different antibiotic resistance profiles. Minimum inhibitory, minimum bactericidal, and fractional inhibitory concentrations were performed to determine the efficacy of the metal ions and graphene composites individually and their synergy in combination. Crystal violet biofilm and XTT assays measured the biofilm inhibition and metabolic activity, respectively. Molybdenum, platinum, tin, gold, and palladium ions exhibited the greatest antimicrobial activity (MIC = 7.8-26.0 mg/L), whilst GO and Gr demonstrated moderate-to-no effect against the planktonic bacterial cells, irrespective of their antibiograms. Biofilms were inhibited by zinc, palladium, silver, and graphene. In combination, silver-graphene and molybdenum-graphene inhibited both the planktonic and biofilm forms of the bacteria making them potential candidates for development into topical antimicrobials for burns patients infected with antibiotic-resistant P. aeruginosa

Sediment supply and barrier dynamics as driving mechanisms of Holocene coastal change for the southern North Sea basin

The combined effects of climate change and human impact lead to regional and local coastal responses that pose major challenges for the future resilience of coastal landscapes, increasing the vulnerability of communities, infrastructure and nature conservation interests. Using the Suffolk coast, southeast England, as a case study, we investigate the importance of sediment supply and barrier dynamics as driving mechanisms of coastal change throughout the Holocene. Litho-, bio- and chronostratigraphic methods are used to decipher the mechanisms of coastal change from the record preserved within coastal stratigraphy. Results suggest that local coastal configuration and sediment supply were the most influential in determining coastal change during the mid- and late Holocene, against a background control of sea-level rise. The importance of sedimentological and morphological factors in shaping Holocene coastal changes in the southern North Sea basin must therefore be considered when using the database of evidence from this region as an analogue for future change under accelerated sea-level rise

On the role of abnormal DL(CO) in ex-smokers without airflow limitation: symptoms, exercise capacity and hyperpolarised helium-3 MRI

BACKGROUND: The functional effects of abnormal diffusing capacity for carbon monoxide (DLCO) in ex-smokers without chronic obstructive pulmonary disease (COPD) are not well understood. OBJECTIVE: We aimed to evaluate and compare well established clinical, physiological and emerging imaging measurements in ex-smokers with normal spirometry and abnormal DLCO with a group of ex-smokers with normal spirometry and DLCO and ex-smokers with Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage I COPD. METHODS: We enrolled 38 ex-smokers and 15 subjects with stage I COPD who underwent spirometry, plethysmography, St George\u27s Respiratory Questionnaire (SGRQ), 6 min Walk Test (6MWT), x-ray CT and hyperpolarised helium-3 ((3)He) MRI. The 6MWT distance (6MWD), SGRQ scores, (3)He MRI apparent diffusion coefficients (ADC) and CT attenuation values below -950 HU (RA950) were evaluated. RESULTS: Of 38 ex-smokers without COPD, 19 subjects had abnormal DLCO with significantly worse ADC (p=0.01), 6MWD (p=0.008) and SGRQ (p=0.01) but not RA950 (p=0.53) compared with 19 ex-smokers with normal DLCO. Stage I COPD subjects showed significantly worse ADC (p=0.02), RA950 (p=0.0008) and 6MWD (p=0.005), but not SGRQ (p=0.59) compared with subjects with abnormal DLCO. There was a significant correlation for (3)He ADC with SGRQ (r=0.34, p=0.02) and 6MWD (r=-0.51, p=0.0002). CONCLUSIONS: In ex-smokers with normal spirometry and CT but abnormal DLCO, there were significantly worse symptoms, 6MWD and (3)He ADC compared with ex-smokers with normal DLCO, providing evidence of the impact of mild or early stage emphysema and a better understanding of abnormal DLCO and hyperpolarised (3)He MRI in ex-smokers without COPD

Chemistry and Kinematics of the Late-Forming Dwarf Irregular Galaxies Leo A, Aquarius, and Sagittarius DIG

We present Keck/DEIMOS spectroscopy of individual stars in the relatively isolated Local Group dwarf galaxies Leo A, Aquarius, and the Sagittarius dwarf irregular galaxy. The three galaxies—but especially Leo A and Aquarius—share in common delayed star formation histories (SFHs) relative to many other isolated dwarf galaxies. The stars in all three galaxies are supported by dispersion. We found no evidence of stellar velocity structure, even for Aquarius, which has rotating H i gas. The velocity dispersions indicate that all three galaxies are dark-matter-dominated, with dark-to-baryonic mass ratios ranging from 4.4_(-0.8)^(+1.0) (SagDIG) to 9.6_(-1.8)^(+2.5) (Aquarius). Leo A and SagDIG have lower stellar metallicities than Aquarius, and they also have higher gas fractions, both of which would be expected if Aquarius were further along in its chemical evolution. The metallicity distribution of Leo A is inconsistent with a closed or leaky box model of chemical evolution, suggesting that the galaxy was pre-enriched or acquired external gas during star formation. The metallicities of stars increased steadily for all three galaxies, but possibly at different rates. The [α/Fe] ratios at a given [Fe/H] are lower than that of the Sculptor dwarf spheroidal galaxy, which indicates more extended SFHs than Sculptor, consistent with photometrically derived SFHs. Overall, the bulk kinematic and chemical properties for the late-forming dwarf galaxies do not diverge significantly from those of less delayed dwarf galaxies, including dwarf spheroidal galaxies