Foliage of Tropical Trees and Shrubs and Their Secondary Metabolites Modify In Vitro Ruminal Fermentation, Methane and Gas Production without a Tight Correlation with the Microbiota.

Ruminants, mainly cattle, contribute to greenhouse gases (GHG) emissions as methane (CH4) is produced by ruminal fermentation. Hence, various anti-methanogenic feed strategies have been studied, including the use of plants with secondary metabolites. This study evaluated in vitro ruminal fermentation metrics, microbial composition by digital droplet PCR (ddPCR) and the CH4 production of the foliage of several tropical trees and shrubs: Leucaena leucocephala, Moringa oleifera, Albizia lebbeck, Enterolobium cyclocarpum, Piscidia piscipula, Brosimum alicastrum, Lysiloma latisiliquum, Guazuma ulmifolia, Cnidoscolus aconitifolius, Gliricidia sepium and Bursera simaruba, using Cynodon plectostachyus grass as control. The results showed a wide variation in the chemical composition of the foliage, as well as in the ruminal microbiota. The crude protein (CP) content ranged from 11 to 25%, whereas the content of condensed tannins (CT) and saponins (S) was from 0.02 to 7%, and 3.2 to 6.6%, respectively. The greatest dry matter degradability (DMD) after 72 h was 69% and the least 35%, the latter coinciding with the least gas production (GP). A negative correlation was found between the CT and CH4 production, also between protozoa and fungi with the SGMT group of archaea. We concluded that the foliage of some tropical trees and shrubs has a high nutritional value and the potential to decrease CH4 production due to its CT content.

Study of racial profile of the native Guajolote (Meleagris gallopavo gallopavo) in two regions of Mexico: morphometric characterization.

The present study aimed at the morphometric characterization of the native Guajolote reared in two regions of Mexico using multivariate analysis techniques. Data from a total of 362 unrelated native Guajolotes (257 males and 105 females) were used: of these, 160 were from the Central region and 202 were from the Southeast region. The birds were also grouped according to age, as youngs (≤ 8 months; n = 150) and adults (≥ 9 months; n = 212). The body weight (BW) and nine morphometric measurements-chest circumference (CC), body length (BL), body height (BH), neck length (NL), peak length (PL), shank length (SL), shank diameter (SD), wing length (WL) and wing width (WW)-were measured. There were significant differences (p < 0.05) due to the effect of region, sex and age for most of the morphometric measurements evaluated. There was a high percentage of positive and significant correlations (p < 0.001; p < 0.01) between the variables. In each region, three principal components were extracted that represented more than 75% of the accumulated variation among the variables. The most discriminating morphometric measurements between populations were WW, PL and NL. The Mahalanobis distance between the males and females of the two populations was 37.457 and 29.310 (p < 0.001), respectively. This differentiation can contribute to the definition of the phenotypic standard of this poultry genetic resource for its official recognition as a breed, as well as in the orientation of its genetic improvement programs in the future.

Assessment of serum total 25-hydroxyvitamin D assays for Vitamin D External Quality Assessment Scheme (DEQAS) materials distributed at ambient and frozen conditions.

The Vitamin D External Quality Assessment Scheme (DEQAS) distributes human serum samples four times per year to over 1000 participants worldwide for the determination of total serum 25-hydroxyvitamin D [25(OH)D)]. These samples are stored at -40 °C prior to distribution and the participants are instructed to store the samples frozen at -20 °C or lower after receipt; however, the samples are shipped to participants at ambient conditions (i.e., no temperature control). To address the question of whether shipment at ambient conditions is sufficient for reliable performance of various 25(OH)D assays, the equivalence of DEQAS human serum samples shipped under frozen and ambient conditions was assessed. As part of a Vitamin D Standardization Program (VDSP) commutability study, two sets of the same nine DEQAS samples were shipped to participants at ambient temperature and frozen on dry ice. Twenty-eight laboratories participated in this study and provided 34 sets of results for the measurement of 25(OH)D using 20 ligand binding assays and 14 liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods. Equivalence of the assay response for the frozen versus ambient DEQAS samples for each assay was evaluated using multi-level modeling, paired t-tests including a false discovery rate (FDR) approach, and ordinary least squares linear regression analysis of frozen versus ambient results. Using the paired t-test and confirmed by FDR testing, differences in the results for the ambient and frozen samples were found to be statistically significant at p < 0.05 for four assays (DiaSorin, DIAsource, Siemens, and SNIBE prototype). For all 14 LC-MS/MS assays, the differences in the results for the ambient- and frozen-shipped samples were not found to be significant at p < 0.05 indicating that these analytes were stable during shipment at ambient conditions. Even though assay results have been shown to vary considerably among different 25(OH)D assays in other studies, the results of this study also indicate that sample handling/transport conditions may influence 25(OH)D assay response for several assays.

Interlaboratory comparison of 25-hydroxyvitamin D assays: Vitamin D Standardization Program (VDSP) Intercomparison Study 2 - Part 2 ligand binding assays - impact of 25-hydroxyvitamin D2 and 24R,25-dihydroxyvitamin D3 on assay performance.

An interlaboratory comparison study was conducted by the Vitamin D Standardization Program (VDSP) to assess the performance of ligand binding assays (Part 2) for the determination of serum total 25-hydroxyvitamin D [25(OH)D]. Fifty single-donor samples were assigned target values for concentrations of 25-hydroxyvitamin D2 [25(OH)D2], 25-hydroxyvitamin D3 [25(OH)D3], 3-epi-25-hydroxyvitamin D3 [3-epi-25(OH)D3], and 24R,25-dihydroxyvitamin D3 [24R,25(OH)2D3] using isotope dilution liquid chromatography-tandem mass spectrometry (ID LC-MS/MS). VDSP Intercomparison Study 2 Part 2 includes results from 17 laboratories using 32 ligand binding assays. Assay performance was evaluated using mean % bias compared to the assigned target values and using linear regression analysis of the test assay mean results and the target values. Only 50% of the ligand binding assays achieved the VDSP criterion of mean % bias ≤ |± 5%|. For the 13 unique ligand binding assays evaluated in this study, only 4 assays were consistently within ± 5% mean bias and 4 assays were consistently outside ± 5% mean bias regardless of the laboratory performing the assay. Based on multivariable regression analysis using the concentrations of individual vitamin D metabolites in the 50 single-donor samples, most assays underestimate 25(OH)D2 and several assays (Abbott, bioMérieux, DiaSorin, IDS-EIA, and IDS-iSYS) may have cross-reactivity from 24R,25(OH)2D3. The results of this interlaboratory study represent the most comprehensive comparison of 25(OH)D ligand binding assays published to date and is the only study to assess the impact of 24R,25(OH)2D3 content using results from a reference measurement procedure.

Assessment of serum total 25-hydroxyvitamin D assay commutability of Standard Reference Materials and College of American Pathologists Accuracy-Based Vitamin D (ABVD) Scheme and Vitamin D External Quality Assessment Scheme (DEQAS) materials: Vitamin D Standardization Program (VDSP) Commutability Study 2.

An interlaboratory study was conducted through the Vitamin D Standardization Program (VDSP) to assess commutability of Standard Reference Materials® (SRMs) and proficiency testing/external quality assessment (PT/EQA) samples for determination of serum total 25-hydroxyvitamin D [25(OH)D] using ligand binding assays and liquid chromatography-tandem mass spectrometry (LC-MS/MS). A set of 50 single-donor serum samples were assigned target values for 25-hydroxyvitamin D2 [25(OH)D2] and 25-hydroxyvitamin D3 [25(OH)D3] using reference measurement procedures (RMPs). SRM and PT/EQA samples evaluated included SRM 972a (four levels), SRM 2973, six College of American Pathologists (CAP) Accuracy-Based Vitamin D (ABVD) samples, and nine Vitamin D External Quality Assessment Scheme (DEQAS) samples. Results were received from 28 different laboratories using 20 ligand binding assays and 14 LC-MS/MS methods. Using the test assay results for total serum 25(OH)D (i.e., the sum of 25(OH)D2 and 25(OH)D3) determined for the single-donor samples and the RMP target values, the linear regression and 95% prediction intervals (PIs) were calculated. Using a subset of 42 samples that had concentrations of 25(OH)D2 below 30 nmol/L, one or more of the SRM and PT/EQA samples with high concentrations of 25(OH)D2 were deemed non-commutable using 5 of 11 unique ligand binding assays. SRM 972a (level 4), which has high exogenous concentration of 3-epi-25(OH)D3, was deemed non-commutable for 50% of the LC-MS/MS assays.

Chemical Perturbation of Chloroplast-Related Processes Affects Circadian Rhythms of Gene Expression in Arabidopsis: Salicylic Acid Application Can Entrain the Clock.

The plant circadian system reciprocally interacts with metabolic processes. To investigate entrainment features in metabolic-circadian interactions, we used a chemical approach to perturb metabolism and monitored the pace of nuclear-driven circadian oscillations. We found that chemicals that alter chloroplast-related functions modified the circadian rhythms. Both vitamin C and paraquat altered the circadian period in a light-quality-dependent manner, whereas rifampicin lengthened the circadian period under darkness. Salicylic acid (SA) increased oscillatory robustness and shortened the period. The latter was attenuated by sucrose addition and was also gated, taking place during the first 3 h of the subjective day. Furthermore, the effect of SA on period length was dependent on light quality and genotype. Period lengthening or shortening by these chemicals was correlated to their inferred impact on photosynthetic electron transport activity and the redox state of plastoquinone (PQ). Based on these data and on previous publications on circadian effects that alter the redox state of PQ, we propose that the photosynthetic electron transport and the redox state of PQ participate in circadian periodicity. Moreover, coupling between chloroplast-derived signals and nuclear oscillations, as observed in our chemical and genetic assays, produces traits that are predicted by previous models. SA signaling or a related process forms a rhythmic input loop to drive robust nuclear oscillations in the context predicted by the zeitnehmer model, which was previously developed for Neurospora. We further discuss the possibility that electron transport chains (ETCs) are part of this mechanism.

Physiological and Genetic Dissection of Sucrose Inputs to the Arabidopsis thaliana Circadian System.

Circadian rhythms allow an organism to synchronize internal physiological responses to the external environment. Perception of external signals such as light and temperature are critical in the entrainment of the oscillator. However, sugar can also act as an entraining signal. In this work, we have confirmed that sucrose accelerates the circadian period, but this observed effect is dependent on the reporter gene used. This observed response was dependent on sucrose being available during free-running conditions. If sucrose was applied during entrainment, the circadian period was only temporally accelerated, if any effect was observed at all. We also found that sucrose acts to stabilize the robustness of the circadian period under red light or blue light, in addition to its previously described role in stabilizing the robustness of rhythms in the dark. Finally, we also found that CCA1 is required for both a short- and long-term response of the circadian oscillator to sucrose, while LHY acts to attenuate the effects of sucrose on circadian period. Together, this work highlights new pathways for how sucrose could be signaling to the oscillator and reveals further functional separation of CCA1 and LHY.

AKIN10 activity as a cellular link between metabolism and circadian-clock entrainment in Arabidopsis thaliana.

AKIN10, the catalytic subunit of the Snf1 (sucrose non-fermenting 1)-related kinase 1 (SnRK1) complex, acts as an energy sensor in plants. We showed that AKIN10-induced expression affects the pace of the circadian clock and particularly the phase of expression of GIGANTEA (GI). The AKIN10 effect on period length required TIME FOR COFFEE (TIC), a circadian-clock component with developmental and metabolic roles. Here we expand on the possible interactions between AKIN10, whose activity is involved in transcriptional reprogramming, and clock elements GI and TIC. We hypothesize how they could participate in clock entrainment through a metabolic signal derived from carbon pools and starch metabolism. Additionally, we consider further the role of cellular energy status to the clock through the formation of a hypothetical protein complex. We also demonstrate the role of AKIN10, but not its sequence-related kinase AKIN11, on clock periodicity. Altogether we present a model of action of these elements in metabolic-related clock entrainment.

The metabolic sensor AKIN10 modulates the Arabidopsis circadian clock in a light-dependent manner.

Plants generate rhythmic metabolism during the repetitive day/night cycle. The circadian clock produces internal biological rhythms to synchronize numerous metabolic processes such that they occur at the required time of day. Metabolism conversely influences clock function by controlling circadian period and phase and the expression of core-clock genes. Here, we show that AKIN10, a catalytic subunit of the evolutionarily conserved key energy sensor sucrose non-fermenting 1 (Snf1)-related kinase 1 (SnRK1) complex, plays an important role in the circadian clock. Elevated AKIN10 expression led to delayed peak expression of the circadian clock evening-element GIGANTEA (GI) under diurnal conditions. Moreover, it lengthened clock period specifically under light conditions. Genetic analysis showed that the clock regulator TIME FOR COFFEE (TIC) is required for this effect of AKIN10. Taken together, we propose that AKIN10 conditionally works in a circadian clock input pathway to the circadian oscillator.

The GI-CDF module of Arabidopsis affects freezing tolerance and growth as well as flowering.

Plants monitor and integrate temperature, photoperiod and light quality signals to respond to continuous changes in their environment. The GIGANTEA (GI) protein is central in diverse signaling pathways, including photoperiodic, sugar and light signaling pathways, stress responses and circadian clock regulation. Previously, GI was shown to activate expression of the key floral regulators CONSTANS (CO) and FLOWERING LOCUS T (FT) by facilitating degradation of a family of CYCLING DOF FACTOR (CDF) transcriptional repressors. However, whether CDFs are implicated in other processes affected by GI remains unclear. We investigated the contribution of the GI-CDF module to traits that depend on GI. Transcriptome profiling indicated that mutations in GI and the CDF genes have antagonistic effects on expression of a wider set of genes than CO and FT, whilst other genes are regulated by GI independently of the CDFs. Detailed expression studies followed by phenotypic assays showed that the CDFs function downstream of GI, influencing responses to freezing temperatures and growth, but are not necessary for proper clock function. Thus GI-mediated regulation of CDFs contributes to several processes in addition to flowering, but is not implicated in all of the traits influenced by GI.

TIME FOR COFFEE is an essential component in the maintenance of metabolic homeostasis in Arabidopsis thaliana.

Plants often respond to environmental changes by reprogramming metabolic and stress-associated pathways. Homeostatic integration of signaling is a central requirement for ensuring metabolic stability in living organisms. Under diurnal conditions, properly timed rhythmic metabolism provides fitness benefits to plants. TIME FOR COFFEE (TIC) is a circadian regulator known to be involved in clock resetting at dawn. Here we explored the mechanism of influence of TIC in plant growth and development, as initiated by a microarray analysis. This global profiling showed that a loss of TIC function causes a major reprogramming of gene expression that predicts numerous developmental, metabolic, and stress-related phenotypes. This led us to demonstrate that this mutant exhibits late flowering, a plastochron defect, and diverse anatomical phenotypes. We further observed a starch-excess phenotype and altered soluble carbohydrate levels. tic exhibited hypersensitivity to oxidative stress and abscisic acid, and this was associated with a striking resistance to drought. These phenotypes were connected to an increase in total glutathione levels that correlated with a readjustment of amino acids and polyamine pools. By comparatively analyzing our transcriptomic and metabolomic data, we concluded that TIC is a central element in plant homeostasis that integrates and coordinates developmental, metabolic, and environmental signals.

[Implementation of diabetic retinopathy screening using digital retinography in primary care]. / Implantación del cribado de retinopatía diabética mediante retinografía digital en atención primaria.

OBJECTIVE: To improve early detection and monitoring of diabetic retinopathy (DR) in two primary care centers (PCC) using a non-mydriatic retinal camera (NMRC). DESIGN: Prospective longitudinal descriptive study. LOCATION: Area 11 Primary Care, Madrid. PARTICIPANTS: Adult patients with diabetes (DM) without funduscopic examination in the last two years (FO). INTERVENTIONS: Implementation process of screening for DR by NMRC in PCC. Digital photography of the retina made by nursing staff. Interpretation of retinal images by an ophthalmologist. MEASUREMENTS: Number of DM with FO. Number of patients with DR. Metabolic control in the last two years (HbA1c). STUDY PERIOD: September 2009-September 2011. RESULTS: DM increased from 2850 to 3357. The proportion of patients with FO increased from 6.7% (95% CI: 5.7-7.6%) to 32.4% (95% CI: 30.8-34.0%) (P<.001). The prevalence of DR increased from 3.85% (95% CI: 3.14-4.58%) to 4.3% (95% CI: 3.59-4.99%). The percentage of patients with FO and DR decreased from 60% (95% CI: 52.77-67.23%) to 14% (95% CI: 11.87-16.09%) (P<.001). The proportion of DM with HbA1c increased from 21.9% (95% CI: 20.36-23.43%) to 52.4% (95% CI: 50.69-54.10%) (P<.001). The mean HbA1c in patients with DR and without DR was 7.8 (95% CI: 7.50-8.06) and 7.1 (IC95% CI: 7.08-7.22) (P<.001), respectively. CONCLUSIONS: NMRC in PCC is accompanied by an increase in the number of diabetics with FO examination. The implementing of DR screening is possible if an NMRC is available, along with available resources, the motivation of the professionals involved and management commitment. A longer follow-up is required to determine its impact on referral rates to ophthalmology service, and to only refer those with dubious images.

TIME FOR COFFEE represses accumulation of the MYC2 transcription factor to provide time-of-day regulation of jasmonate signaling in Arabidopsis.

Plants are confronted with predictable daily biotic and abiotic stresses that result from the day-night cycle. The circadian clock provides an anticipation mechanism to respond to these daily stress signals to increase fitness. Jasmonate (JA) is a phytohormone that mediates various growth and stress responses. Here, we found that the circadian-clock component TIME FOR COFFEE (TIC) acts as a negative factor in the JA-signaling pathway. We showed that the tic mutant is hypersensitive to growth-repressive effects of JA and displays altered JA-regulated gene expression. TIC was found to interact with MYC2, a key transcription factor of JA signaling. From this, we discovered that the circadian clock rhythmically regulates JA signaling. TIC is a key determinant in this circadian-gated process, and as a result, the tic mutant is defective in rhythmic JA responses to pathogen infection. TIC acts here by inhibiting MYC2 protein accumulation and by controlling the transcriptional repression of CORONATINE INSENSITIVE1 in an evening-phase-specific manner. Taken together, we propose that TIC acts as an output component of the circadian oscillator to influence JA signaling directly.