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Telomerase is a nonclassical DNA polymerase that uses its integral RNA as a template to synthesize telomeric repeats onto chromosome ends. The molecular mechanism of telomerase is unique and involves a translocation step after the synthesis of each telomeric repeat. To directly measure the enzymatic turnover of substrate and the efficiency of the translocation step we have extended our two- color single molecule fluorescence coincidence method (Anal.Chem. 2003, 75, 1664-1670). The method employs Cy5-dATP incorporation into a DNA primer that has been prelabeled with a reference fluorophore. Measurements are performed in the single molecule regime and products, which necessarily have both fluorophores, are excited by two independent lasers, and give rise to coincident events. By counting the number of coincident events and using the coincidence detection efficiency, it is possible to determine the number of the extended products generated by attomole quantities of telomerase, without separation or the use of PCR or radioactivity. Histograms of the logarithms of the ratios of the Cy5 to the reference fluorophore fluorescence can be used to determine the length distribution of the products and hence the enzyme processivity. The mean processivity obtained from the single molecule fluorescence coincidence assay is 0.32 ± 0.04, in good agreement with the value of 0.37 ± 0.05 derived from the direct radioactive assay approach. The function of the alignment domain of human telomerase RNA in sustaining catalytic activity in vitro has been reevaluated using this method. Together with our previous results (Nucleic Acids Res. 2002, 30, 4470-4480) these experiments identify the essential residues in the alignment domain of human telomerase RNA that contribute to the activity and processivity of telomerase. [ABSTRACT FROM AUTHOR]

The oldest and most metal-poor Milky Way stars form a kinematically hot halo, which motivates the two major formation scenarios for our galaxy: extended hierarchical accretion and rapid collapse. RR Lyrae stars are excellent tracers of old and metal-poor populations. We measured the kinematics of 43 RR Lyrae stars in the inner regions of the nearby Large Magellanic Cloud (LMC) galaxy. The velocity dispersion equals 53 ± 10 kilometers per second, which indicates that a kinematically hot metal-poor old halo also exists in the LMC. This result suggests that our galaxy and smaller late-type galaxies such as the LMC have similar early formation histories. [ABSTRACT FROM AUTHOR]

The European Union's commitment to increase recycling and recovery rates of municipal solid waste requires significant changes in current waste management. Local governments are developing various strategies for treating the organic fraction of municipal waste (biowaste) via composting. Community composting centres (CCC), green waste collection, treatment points and community gardens are some of these new approaches. Population density and spatial distribution, together with the existence of community green areas, determine the location of the various infrastructures for recycling local biowaste. The composting process consumes high amounts of bulking agent (BA) necessary to provide the structure that allows, amongst other uses, biowaste aeration and microbial surface colonisation. Shredded green waste from parks, gardens and households can be used as BA in community composting and home composting. In this study, a total of 46 compost samples obtained from CCC with two types of handling were analysed: 22 samples treated by vertical flow (VF) and 24 samples treated by horizontal flow (HF). The HF model allowed better use of the volume of modular composting units and the VF model required less effort and time for the CCC operator. Mature, stable and high-nutrient-content composts were obtained with both models. These composts met the legal requirements to be used as an organic amendment, and they can be delivered to the participants or used in community gardens in the municipality., (© 2023. The Author(s).)

Purpose: To evaluate the additive effect of Transcranial Direct Current Stimulation (tDCS) associated with multi-component training (MT) on the functional capacity (FC) of older adults and to assess whether these effects remain after the end of training. The secondary objectives were to evaluate the locomotion capacity, balance, functional independence, and quality of life and correlate them with functional capacity., Methodology: Twenty-eight older adults were randomized into two groups: experimental (MT associated with active tDCS - a-tDCS) and control (MT associated with sham tDCS - s-tDCS). The FC was measured by the Glittre-ADL test, locomotion capacity by the 6-minute walk test, balance by the BESTest, functional independence by the FIM, and quality of life by the WHQOL. The assessments were performed pre-, post-intervention, and 30-day follow-up., Results: There was a significant decrease in the time to the Glittre-ADL test when comparing the a-tDCS and s-tDCS groups after the interventions (139.77 ± 21.62, 205.10 ± 43.02, p < .001) and at the 30-day follow-up (142.74 ± 17.12, 219.55 ± 54.05, p < .001), respectively. There was a moderate correlation between FC and locomotion capacity and balance., Conclusions: The addition of tDCS potentiated the results of MT to impact FC, maintaining the positive results longer. Locomotion and balance influenced the improvement of functional capacity.

While heart failure is a primary cause of death for many in-transit-loss (ITL) pigs, the underlying cause of these deaths is not known. Cardiomyopathies are considered a common cause of heart failure in humans and often have a genetic component. The objective of this study was to determine if genes associated with cardiomyopathies could be identified in ITL pigs. Samples from the hearts of pigs that died during transport to an abattoir in Ontario, Canada were collected and genotyped along with samples from pigs that did not die during transport (ILT hearts: n = 149; non-ITL/control hearts: n = 387). Genome-wide analyses were carried out on each of the determined phenotypes (gross cardiac lesions) using a medium density single nucleotide polymorphism (SNP) chip and 500 kb windows/regions for analysis, with 250 kb regions of overlap. The distribution derived by a multidimensional scaling (MDS) analysis of all phenotypes demonstrated a lack of complete separation between phenotypes of affected and unaffected animals, which made diagnosis difficult. Although genetic differences were small, a few genes associated with dilated cardiomyopathy (DCM) and arrhythmogenic right ventricular cardiomyopathy (ARVM) were identified. In addition, multiple genes associated with cardiac arrhythmias and ventricular hypertrophy were identified that can possibly result in heart failure. The results of this preliminary study did not provide convincing evidence that a single, heritable cardiomyopathy is the cause of heart failure in ITL pigs., (Copyright and/or publishing rights held by the Canadian Veterinary Medical Association.)

Ultrahot giant exoplanets receive thousands of times Earth's insolation 1,2 . Their high-temperature atmospheres (greater than 2,000 kelvin) are ideal laboratories for studying extreme planetary climates and chemistry 3-5 . Daysides are predicted to be cloud-free, dominated by atomic species 6 and much hotter than nightsides 5,7,8 . Atoms are expected to recombine into molecules over the nightside 9 , resulting in different day and night chemistries. Although metallic elements and a large temperature contrast have been observed 10-14 , no chemical gradient has been measured across the surface of such an exoplanet. Different atmospheric chemistry between the day-to-night ('evening') and night-to-day ('morning') terminators could, however, be revealed as an asymmetric absorption signature during transit 4,7,15 . Here we report the detection of an asymmetric atmospheric signature in the ultrahot exoplanet WASP-76b. We spectrally and temporally resolve this signature using a combination of high-dispersion spectroscopy with a large photon-collecting area. The absorption signal, attributed to neutral iron, is blueshifted by -11 ± 0.7 kilometres per second on the trailing limb, which can be explained by a combination of planetary rotation and wind blowing from the hot dayside 16 . In contrast, no signal arises from the nightside close to the morning terminator, showing that atomic iron is not absorbing starlight there. We conclude that iron must therefore condense during its journey across the nightside.

The hydrocarbon residue in petroleum product storage tanks is waste generated in large quantities that must be properly managed to reduce its risk to the environment. By comparing the effect of two organic cosubstrates, the aim of our research is to determine the feasibility of composting as a bioremediation method for the treatment of the solid phase of the hydrocarbon residue. For this purpose, four treatments of the pollutant waste were established in triplicate: waste only; waste with bulking agent (1:2); waste with fish sludge and bulking agent (1:2:6); and waste with municipal sewage sludge and bulking agent (1:2:6). The composting system consisted of 12 reactors with a capacity of 30 L, each equipped with aeration and temperature control. Both at the beginning and the end of the experiment (20 days), we evaluated the physicochemical parameters, the structure of the microbial community through phospholipid fatty acid analysis, and the total petroleum hydrocarbon content (TPH). Treatments with cosubstrates maintained thermophilic temperatures, during 14 and 8 days in fish and municipal sludge respectively, while in the controls mesophilic conditions were maintained. The incorporation of fish sludge decreased TPH present in the initial mixture by 39.5%. The municipal sludge treatment resulted in a lower of temperatures and a TPH decrease close to 23.9%. In the control treatments, there was a slight TPH decrease, mainly due to the forced ventilation. Although, both composting treatments with cosubstrates proved adequate for the bioremediation of residue from hydrocarbon storage tanks, fish sludge presented best bioremediation conditions. Municipal sewage sludge provided a bioaugmentation effect due to its rich diversity and microbial biomass. Fish sludge could have biostimulant and surfactant effect producing an aliphatic mixture of pollutant waste with the nutritional requirements to promote the development of fungal communities., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

In-transit losses of market hogs represent a small proportion of all market-weight pigs shipped in a year. This suggests that individual pig factors may be a significant cause of in-transit losses along with more traditionally considered environmental and transport factors. An investigation was performed to determine whether cardiac pathology and heart weights were associated with pigs that did or did not die during transport to an abattoir. The hearts from 70 pigs that died in-transit to one Ontario abattoir and 388 pigs that arrived alive were collected and examined. Hearts from pigs that died during transport demonstrated greater frequencies of cardiac lesions ( P < 0.05). These included hypertrophy of ventricle walls (Left: 97% vs. 64%; Right: 86% vs. 57%), dilation of ventricle chambers (Left: 79% vs. 0.5%; Right: 100% vs. 5%), and dilation of the pulmonary artery and aorta (59% vs. 1.5%). Total heart weight to body weight ratios were increased (3.6 vs. 3.3 g/kg) and left ventricle plus septum weight over right ventricle weight ratio was decreased in pigs that died during transport over non-in-transit loss pigs (2.5 vs. 2.8; P < 0.05). This may indicate reduced cardiac function in hogs that died during transport. Pigs with reduced cardiac function would have exercise intolerance and be more susceptible to death during transport due to the increased cardiac workload required during sorting, loading, and transport of the pigs to the abattoir. Further research to quantify cardiac function in pigs with cardiac lesions or abnormal heart weight ratios is warranted., (© The Author(s) 2018. Published by Oxford University Press on behalf of the American Society of Animal Science.)

Clinically healthy pigs used in research are assumed to have normal cardiac structure and function. Subclinical cardiac abnormalities may adversely affect the responses being measured in these experiments. The gross and histologic lesions observed in hearts collected from a Canadian abattoir between 2012 and 2015 indicated an unexpectedly high prevalence of cardiac abnormalities: 75% (297/396) of the hearts examined had such lesions. The ratios of total heart weight to body weight and of right ventricle weight to body weight were significantly greater for the hearts with lesions than for the hearts with no lesions, which suggests that cardiac remodeling, particularly hypertrophy, had occurred. The large percentage of hearts with cardiac remodeling from asymptomatic market pigs demonstrates an increased probability that subclinical cardiac abnormalities may exist in research pigs, especially those accessed through commercial channels. Researchers should be aware of this likelihood if subclinical cardiac abnormalities could adversely affect their experimental findings.

Fonseca Alves, DJ, Bartholomeu-Neto, J, Júnior, ER, Ribeiro Zarricueta, BS, Nóbrega, OT, and Córdova, C. Walking speed, risk factors, and cardiovascular events in older adults-systematic review. J Strength Cond Res 31(11): 3235-3244, 2017-It is important that new clinical measures can identify risk factors and predict cardiovascular events. Although the walking speed (WS) test is a potential candidate, consolidating data from multiple studies is required to determine comparative references. We examined the associations of WS measures with markers of cardiovascular risk and with cardiovascular events in noninstitutionalized subjects older than 60 years. A systematic review of observational studies was conducted using MEDLINE and SCOPUS from inception of the databases to December 2014, aiming at studies that evaluated WS as the primary outcome (usual or maximal pace) within a distance ≤20 m associated with cardiovascular health. All 15 included studies (29,845 subjects) reported significant associations of WS with different cardiovascular risk factors (coronary artery calcification, C-reactive protein, hypertension, diabetes, and intima-media thickness) and occurrence of cardiovascular events (peripheral artery disease, stroke, and mortality). Approximately 80% of the studies used a distance ≤6 m and WS at usual pace. There was high heterogeneity in the risk thresholds established by different studies. Our results suggest usefulness of the WS test as a tool for cardiovascular risk stratification in older adults. However, the variation in speed thresholds and diversity of protocols among studies suggest caution when generalizing results to different older adult populations. Because the WS test is a simple, cheap, and safe tool to administer, we make suggestions for its standardization in future studies.

This research evaluates, through microbial dynamics, the use of earthworms Eisenia andrei for maturation of pre-composted pig manure in comparison with maturation under static conditions and with vermicomposting of fresh pig manure. Therefore, two substrates were used (fresh and pre-composted pig manure) and four treatments were developed: fresh manure vermicomposting, control of fresh manure without earthworms, pre-composting followed by vermicomposting and static maturation of pre-composted manure. In order to determine the microbial dynamics, the enzymatic activities and profiles of phospholipid fatty acids (PLFAs) were evaluated over a 112-days period. Physicochemical and biological parameters of the obtained products were also analyzed. The presence of earthworms significantly reduced (p<0.05) microbial biomass and all the microbial groups (Gram+bacteria, Gram-bacteria, and fungi) in both substrates. The enzymatic activities (cellulase, β-glucosidase and acid phosphatase) behaved in a significantly distinctive manner (p<0.05) depending on the treatment. Microbial communities had significant correlations (p<0.05) with hydrolytic activities during static maturation of pre-composted manure. This indicates a direct effect of microbiota evolution on the degradative processes; however, complex earthworm-microbiota interactions were established in the presence of E. andrei. After earthworms' removal from vermicompost of fresh substrate at 70day, an increase in Gram + (4.4 times), Gram - (3.8 times) and fungi (2.8 times) were observed and, although the vermicompost achieved quality values, it is necessary to optimize the vermicompost aging phase period to improve the stability. Static maturation presented stability on microbial dynamics that indicated a slow degradation of organic compounds so that, maturation of pre-composted manure through vermicomposting is better option., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

A new family of copper(I) complexes of general formula [Cu(dppe)(NN)] + have been synthesized and fully characterized, with dppe=1.2-bis(diphenylphosphino)ethane and NN representing several bidentate heteroaromatic ligands: 2,2'-bipy=2.2'-bipyridine (1), Me 2 bpy=4.4'-dimethyl-2,2'-bipyridine (2), dpytz=3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine (3), dpp=2.3-bis(2-pyridyl)pyrazine (4), and the metallaligand [Ru(η 5 -C 5 H 5 )(PPh 3 )(dpp)] + (5), yielding the bimetallic copper(I)-ruthenium(II) complex [Cu(dppe)(μ-dpp)Ru(η 5 -C 5 H 5 )(PPh 3 )] 2+ (6). The single crystal structures of complexes (2) and (4) were determined by X-ray diffraction studies. All the complexes exhibit high cytotoxicity against the human cancer cells A2780 and MCF7 with IC 50 values far lower than those found for the antitumor drug cisplatin in the same cell lines and even surpassing cisplatin resistance in the A2780cisR cells. They display IC 50 values on the human embryonic kidney HEK293 non-tumoral cells of the same order of magnitude as those found for the tumoral cells. In the ovarian cells the compounds induce rapid production of reactive oxygen species (ROS) probably through mitochondrial pathways. According to the results reported here, these compounds can be considered as prospective antitumoral agents that deserve further evaluation., (Copyright © 2017 Elsevier Inc. All rights reserved.)

In general, in composting facilities the active, or intensive, stage of the process is done separately from the maturation stage, using a specific technology and time. The pre-composted material to be matured can contain enough biodegradable substrates to cause microbial proliferation, which in turn can cause temperatures to increase. Therefore, not controlling the maturation period during waste management at an industrial level can result in undesired outcomes. The main hypothesis of this study is that controlling the maturation stage through turning provides one with an optimized process when compared to the static approach. The waste used was sludge from a seafood-processing plant, mixed with shredded wood (1:2, v/v). The composting system consists of an intensive stage in a 600L static reactor, followed by maturation in triplicate in 200L boxes for 112 days. Two tests were carried out with the same process in reactor and different treatments in boxes: static maturation and turning during maturation when the temperature went above 55°C. PLFAs, organic matter, pH, electrical conductivity, forms of nitrogen and carbon, hydrolytic enzymes and respiratory activity were periodically measured. Turning significantly increased the duration of the thermophilic phase and consequently increased the organic-matter degradation. PCA differentiated significantly the two treatments in function of tracking parameters, especially pH, total carbon, forms of nitrogen and C/N ratio. So, stability and maturity optimum values for compost were achieved in less time with turnings. Whereas turning resulted in microbial-group stabilization and a low mono/sat ratio, static treatment produced greater variability in microbial groups and a high mono/sat ratio, the presence of more degradable substrates causes changes in microbial communities and their study during maturation gives an approach of the state of organic-matter degradation. Obtaining quality compost and optimizing the composting process requires using turning as a control mechanism during maturation., Competing Interests: The authors have declared that no competing interests exist.

During composting, facilities usually exert greater control over the bio-oxidative phase of the process, which uses a specific technology and generally has a fixed duration. After this phase, the material is deposited to mature, with less monitoring during the maturation phase. While there has been considerable study of biological parameters during the thermophilic phase, there is less research on the stabilization and maturation phase. This study evaluates the effects of the type of starting material on the evolution of microbial dynamics during the maturation phase of composting. Three waste types were used: sludge from the fish processing industry, municipal sewage sludge and pig manure, each independently mixed with shredded pine wood as bulking agent. The composting system for each waste type comprised a static reactor with capacity of 600L for the bio-oxidative phase followed by stabilization and maturation phase in triplicate 200L boxes for 112days. Phospholipid fatty acids, enzyme activities and physico-chemical parameters were measured throughout the maturation phase. The evolution of the total microbial biomass, Gram + bacteria, Gram - bacteria, fungi and enzymatic activities (β-glucosidase, cellulase, protease, acid and alkaline phosphatase) depended significantly on the waste type (p<0.001). The predominant microbial community for each waste type remained present throughout the maturation process, indicating that the waste type determines the microorganisms that are able to develop at this stage. While fungi predominated during fish sludge maturation, manure and municipal sludge were characterized by a greater proportion of bacteria. Both the structure of the microbial community and enzymatic activities provided important information for monitoring the composting process. More attention should be paid to the maturation phase in order to optimize composting., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Municipal sewage sludge is a waste with high organic load generated in large quantities that can be treated by biodegradation techniques to reduce its risk to the environment. This research studies vermicomposting and vermicomposting after composting of sewage sludge with the earthworm specie Eisenia andrei. In order to determine the effect that earthworms cause on the microbial dynamics depending on the treatment, the structure and activity of the microbial community was assessed using phospholipid fatty acid analysis and enzyme activities, during 112days of vermicomposting of fresh and composted sewage sludge, with and without earthworms. The presence of earthworms significantly reduced microbial biomass and all microbial groups (Gram+ bacteria, Gram- bacteria and fungi), as well as cellulase and alkaline phosphatase activities. Combined composting-vermicomposting treatment showed a lesser development of earthworms, higher bacterial and fungal biomass than vermicomposting treatment and greater differences, compared with the control without earthworms, in cellulase, β-glucosidase, alkaline and acid phosphatase. Both treatments were suitable for the stabilization of municipal sewage sludge and the combined composting-vermicomposting treatment can be a viable process for maturation of fresh compost., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Porcine epidemic diarrhea virus was first diagnosed in Ontario in January of 2014. An outbreak investigation was conducted and it was hypothesized that feed containing spray-dried porcine plasma contaminated with the virus was a risk factor in the introduction and spread of the disease in Ontario.

Telomerase is a nonclassical DNA polymerase that uses its integral RNA as a template to synthesize telomeric repeats onto chromosome ends. The molecular mechanism of telomerase is unique and involves a translocation step after the synthesis of each telomeric repeat. To directly measure the enzymatic turnover of substrate and the efficiency of the translocation step we have extended our two-color single molecule fluorescence coincidence method (Anal.Chem. 2003, 75, 1664-1670). The method employs Cy5-dATP incorporation into a DNA primer that has been prelabeled with a reference fluorophore. Measurements are performed in the single molecule regime and products, which necessarily have both fluorophores, are excited by two independent lasers, and give rise to coincident events. By counting the number of coincident events and using the coincidence detection efficiency, it is possible to determine the number of the extended products generated by attomole quantities of telomerase, without separation or the use of PCR or radioactivity. Histograms of the logarithms of the ratios of the Cy5 to the reference fluorophore fluorescence can be used to determine the length distribution of the products and hence the enzyme processivity. The mean processivity obtained from the single molecule fluorescence coincidence assay is 0.32 +/- 0.04, in good agreement with the value of 0.37 +/- 0.05 derived from the direct radioactive assay approach. The function of the alignment domain of human telomerase RNA in sustaining catalytic activity in vitro has been reevaluated using this method. Together with our previous results (Nucleic Acids Res. 2002, 30, 4470-4480) these experiments identify the essential residues in the alignment domain of human telomerase RNA that contribute to the activity and processivity of telomerase.

Objective: To determine the cause of an outbreak of Escherichia coli 0157:H7 related to animal exposures so that further transmission could be prevented., Design: Description of laboratory investigations and a case control study., Setting: Agricultural pavilion at an annual fair in Ontario., Population: People with laboratory evidence of E coli 0157:H7 (seven people) and others with diarrhea (155 people) who called the health unit following a media release were interviewed. Animals that were accessed most frequently by the public in the agriculture pavilion were tested for E coli 0157:H7. In the case control study, a case was defined as someone with laboratory confirmed E coli 0157:H7, or someone who developed severe or bloody diarrhea two to eight days after attending the agricultural pavilion at the fair (61 people). A convenience sample of people who attended the agricultural pavilion but did not develop diarrhea was selected as the control group (89 people)., Interventions: Human and animal E coli 0157:H7 specimens were subtyped. Cases and controls were interviewed using a standardized questionnaire., Results: Subtyping of the seven human isolates of E coli 0157:H7 revealed five that were of an extremely uncommon phage type. Three samples from goats and one from sheep at the petting zoo in the agricultural pavilion were of this same phage type. The case control study also implicated goats (odds ratio [OR] 3.65; 95% CI 1.63 to 8.52) and sheep (OR 2.94; 95% CI 1.33 to 6.57) from the petting zoo., Conclusions: Results of this investigation suggest strongly that the goats and sheep from the petting zoo were the source of this outbreak of E coli 0157:H7.