Encapsulation and release of a bacterial carotenoid from hydrogel matrix: Characterization,kinetics and antioxidant study

Various natural polymers with hydrophilic properties have been used to form hydrogels for the encapsulation and delivery of nutrients and drugs in food and pharmaceutical industries. Among them, chitosan (ChiHG)‐ and alginate (AlgHG)‐ based hydrogels have been extensively explored for delivery of several nutraceuticals in recent years. Release of natural canthaxanthin (CX) obtained from Dietzia maris NITD (accession number: HM151403) has been investigated with emphasis on biomedical applications. Significant changes (P < 0.05) in degree of swelling and moisture content (% dry basis) were found after encapsulation of bacterial canthaxanthin (BCX), but the gel content remained unchanged. BCX encapsulation efficiency was calculated to be 55.92% and 60.45% in ChiHG and AlgHG, respectively. A noticeable change in heat of fusion d melting point was recorded in ChiHG and AlgHG after BCX encapsulation. Swelling and BCX release from gel matrix was performed under two different pH (1.2 and 7.4). The results showed that swelling of hydrogel and BCX release was facilitated at higher pH (7.4) than acidic pH (1.2). With regard to the release kinetics data, it was found that BCX is released from bothand AlgHG in a diffusion transport method. In addition, antioxidant activity of BCX encapsulated hydrogels was found significantly higher (P < 0.001) in terms of DPPH, ABTS, nitrite, hydroxyl radical scavenging and reducing power assay. These results indicated that BCX can be successfully encapsulated into a polymeric hydrogel to obtain a dynamic biomaterial that may be used in drug delivery applications in future.     

Estimators for QST and coalescence times

Comparisons of to can provide insights into the evolutionary processes that lead to differentiation, or lack thereof, among the phenotypes of different groups (e.g., populations, species), and these comparisons have been performed on a variety of taxa, including humans. Here, I show that for neutrally evolving (i.e., by genetic drift, mutation, and gene flow alone) quantitative characters, the two commonly used estimators have somewhat different interpretations in terms of coalescence times, particularly when the number of groups that have been sampled is small. A similar situation occurs for estimators. Consequently, when observations come from only a small number of groups, which is not an unusual situation, it is important to match estimators appropriately when comparing to .     

Purging putative siblings from population genetic data sets: a cautionary view

Interest has surged recently in removing siblings from population genetic data sets before conducting downstream analyses. However, even if the pedigree is inferred correctly, this has the potential to do more harm than good. We used computer simulations and empirical samples of coho salmon to evaluate strategies for adjusting samples to account for family structure. We compared performance in full samples and sibling‐reduced samples of estimators of allele frequency (), population differentiation () and effective population size (). Results: (i) unless simulated samples included large family groups together with a component of unrelated individuals, removing siblings generally reduced precision of and ; (ii) based on the linkage disequilibrium method was largely unbiased using full random samples but became increasingly upwardly biased under aggressive purging of siblings. Under nonrandom sampling (some families over‐represented), using full samples was downwardly biased; removing just the right ‘Goldilocks’ fraction of siblings could produce an unbiased estimate, but this sweet spot varied widely among scenarios; (iii) weighting individuals based on the inferred pedigree (to produce a best linear unbiased estimator, BLUE) maximized precision of when the inferred pedigree was correct but performed poorly when the pedigree was wrong; (iv) a variant of sibling removal that leaves intact small sibling groups appears to be more robust to errors in inferences about family structure. Our results illustrate the complex challenges posed by presence of family structure, suggest that no single optimal solution exists and argue for caution in adjusting population genetic data sets for the presence of putative siblings without fully understanding the consequences.     

Integrated process development of a reactive extraction concept for itaconic acid and application to a real fermentation broth

Itaconic acid (IA) has a high potential to be used as a bio‐based platform chemical and its biocatalytic production via fermentation has significantly improved within the last decade. Additionally downstream processing using reactive extraction (RE) was described, potentially enabling a more efficient sustainable bioprocess producing IA. The bottleneck to overcome is the connection of up‐ and downstream processing, caused by lack of biocompatibility of the RE systems and direct application to fermentation broth. Within this study, a biocompatible RE system for IA is defined (pH dependency, extraction mechanism) and used for direct application to a fermentation broth. By optimizing the biocatalyst, the production medium, and the extraction system in an integrated approach, it was possible to define critical parameters that enabled a tuning of the overall bioprocess. With an extraction yield of Y_IA = 0.80 ± 0.03, IA could be produced as sole carboxylic acid ( = 0.490 mol/kg^aq) using a RE system consisting of ethyl oleate as organic solvent and tri‐n‐octylamine as extractant ( = 0.6 mol/kg^org). This work is a proof of concept and demonstrates that by joint consideration of up‐ and downstream processing, optimized bioprocesses can be developed.     

High‐throughput identification of protein mutant stability computed from a double mutant fitness landscape

The effect of a mutation on protein stability is traditionally measured by genetic construction, expression, purification, and physical analysis using low‐throughput methods. This process is tedious and limits the number of mutants able to be examined in a single study. In contrast, functional fitness effects can be measured in a high‐throughput manner by various deep mutational scanning tools. Using protein GB 1, we have recently demonstrated the feasibility of estimating the mutational stability effect ( G) of single‐substitution based on the functional fitness profile of all double‐substitutions. The principle is to identify genetic backgrounds that have an exhausted stability margin. The functional effect of an additional substitution on these genetic backgrounds can then be used to compute the mutational G based on the biophysical relationship between functional fitness and thermodynamic stability. However, to identify such genetic backgrounds, the approach described in our previous study required a benchmark dataset, which is a set of known mutational G. In this study, a benchmark‐independent approach is developed. The genetic backgrounds of interest are identified using k‐means clustering with the integration of structural information. We further demonstrated that a reasonable approximation of G can also be obtained without taking structural information into account. In summary, this study describes a novel method for computing G from double‐substitution functional fitness profiles alone, without relying on any known mutational G as a benchmark.     

Summary goodness‐of‐fit statistics for binary generalized linear models with noncanonical link functions

Generalized linear models (GLM) with a canonical logit link function are the primary modeling technique used to relate a binary outcome to predictor variables. However, noncanonical links can offer more flexibility, producing convenient analytical quantities (e.g., probit GLMs in toxicology) and desired measures of effect (e.g., relative risk from log GLMs). Many summary goodness‐of‐fit (GOF) statistics exist for logistic GLM. Their properties make the development of GOF statistics relatively straightforward, but it can be more difficult under noncanonical links. Although GOF tests for logistic GLM with continuous covariates (GLMCC) have been applied to GLMCCs with log links, we know of no GOF tests in the literature specifically developed for GLMCCs that can be applied regardless of link function chosen. We generalize the Tsiatis GOF statistic originally developed for logistic GLMCCs, (), so that it can be applied under any link function. Further, we show that the algebraically related Hosmer–Lemeshow () and Pigeon–Heyse (J²) statistics can be applied directly. In a simulation study, , , and J² were used to evaluate the fit of probit, log–log, complementary log–log, and log models, all calculated with a common grouping method. The statistic consistently maintained Type I error rates, while those of and J² were often lower than expected if terms with little influence were included. Generally, the statistics had similar power to detect an incorrect model. An exception occurred when a log GLMCC was incorrectly fit to data generated from a logistic GLMCC. In this case, had more power than or J².     

pKa predictions for proteins,RNAs, and DNAs with the Gaussian dielectric function using DelPhi pKa

We developed a Poisson‐Boltzmann based approach to calculate the values of protein ionizable residues (Glu, Asp, His, Lys and Arg), nucleotides of RNA and single stranded DNA. Two novel features were utilized: the dielectric properties of the macromolecules and water phase were modeled via the smooth Gaussian‐based dielectric function in DelPhi and the corresponding electrostatic energies were calculated without defining the molecular surface. We tested the algorithm by calculating values for more than 300 residues from 32 proteins from the PPD dataset and achieved an overall RMSD of 0.77. Particularly, the RMSD of 0.55 was achieved for surface residues, while the RMSD of 1.1 for buried residues. The approach was also found capable of capturing the large shifts of various single point mutations in staphylococcal nuclease (SNase) from ‐cooperative dataset, resulting in an overall RMSD of 1.6 for this set of pKa's. Investigations showed that predictions for most of buried mutant residues of SNase could be improved by using higher dielectric constant values. Furthermore, an option to generate different hydrogen positions also improves predictions for buried carboxyl residues. Finally, the calculations on two RNAs demonstrated the capability of this approach for other types of biomolecules. Proteins 2015; 83:2186–2197. © 2015 Wiley Periodicals, Inc.     

Effective size of density‐dependent populations in fluctuating environments

Reliable estimates of effective population size are of central importance in population genetics and evolutionary biology. For populations that fluctuate in size, harmonic mean population size is commonly used as a proxy for (multi‐) generational effective size. This assumes no effects of density dependence on the ratio between effective and actual population size, which limits its potential application. Here, we introduce density dependence on vital rates in a demographic model of variance effective size. We derive an expression for the ratio in a density‐regulated population in a fluctuating environment. We show by simulations that yearly genetic drift is accurately predicted by our model, and not proportional to as assumed by the harmonic mean model, where N is the total population size of mature individuals. We find a negative relationship between and N. For a given N, the ratio depends on variance in reproductive success and the degree of resource limitation acting on the population growth rate. Finally, our model indicate that environmental stochasticity may affect not only through fluctuations in N, but also for a given N at a given time. Our results show that estimates of effective population size must include effects of density dependence and environmental stochasticity.     

Sensitivity analysis of effective population size to demographic parameters in house sparrow populations

The ratio between the effective and the census population size, , is an important measure of the long‐term viability and sustainability of a population. Understanding which demographic processes that affect most will improve our understanding of how genetic drift and the probability of fixation of alleles is affected by demography. This knowledge may also be of vital importance in management of endangered populations and species. Here, we use data from 13 natural populations of house sparrow (Passer domesticus) in Norway to calculate the demographic parameters that determine . Using the global variance‐based Sobol’ method for the sensitivity analyses, we found that was most sensitive to demographic variance, especially among older individuals. Furthermore, the individual reproductive values (that determine the demographic variance) were most sensitive to variation in fecundity. Our results draw attention to the applicability of sensitivity analyses in population management and conservation. For population management aiming to reduce the loss of genetic variation, a sensitivity analysis may indicate the demographic parameters towards which resources should be focused. The result of such an analysis may depend on the life history and mating system of the population or species under consideration, because the vital rates and sex–age classes that is most sensitive to may change accordingly.     

Contemporary effective population and metapopulation size (Ne and meta‐Ne): comparison among three salmonids inhabiting a fragmented system and differing in gene flow and its asymmetries

We estimated local and metapopulation effective sizes ( and meta‐) for three coexisting salmonid species (Salmo salar, Salvelinus fontinalis, Salvelinus alpinus) inhabiting a freshwater system comprising seven interconnected lakes. First, we hypothesized that might be inversely related to within‐species population divergence as reported in an earlier study (i.e., F_ST: S. salar> S. fontinalis> S. alpinus). Using the approximate Bayesian computation method implemented in ONeSAMP, we found significant differences in () between species, consistent with a hierarchy of adult population sizes (). Using another method based on a measure of linkage disequilibrium (LDNE: ), we found more finite values for S. salar than for the other two salmonids, in line with the results above that indicate that S. salar exhibits the lowest among the three species. Considering subpopulations as open to migration (i.e., removing putative immigrants) led to only marginal and non‐significant changes in , suggesting that migration may be at equilibrium between genetically similar sources. Second, we hypothesized that meta‐ might be significantly smaller than the sum of local s (null model) if gene flow is asymmetric, varies among subpopulations, and is driven by common landscape features such as waterfalls. One ‘bottom‐up’ or numerical approach that explicitly incorporates variable and asymmetric migration rates showed this very pattern, while a number of analytical models provided meta‐ estimates that were not significantly different from the null model or from each other. Our study of three species inhabiting a shared environment highlights the importance and utility of differentiating species‐specific and landscape effects, not only on dispersal but also in the demography of wild populations as assessed through local s and meta‐s and their relevance in ecology, evolution and conservation.     

speed‐ne: Software to simulate and estimate genetic effective population size (Ne) from linkage disequilibrium observed in single samples

The genetic effective population size, N_e, can be estimated from the average gametic disequilibrium () between pairs of loci, but such estimates require evaluation of assumptions and currently have few methods to estimate confidence intervals. speed‐ne is a suite of matlab computer code functions to estimate from with a graphical user interface and a rich set of outputs that aid in understanding data patterns and comparing multiple estimators. speed‐ne includes functions to either generate or input simulated genotype data to facilitate comparative studies of estimators under various population genetic scenarios. speed‐ne was validated with data simulated under both time‐forward and time‐backward coalescent models of genetic drift. Three classes of estimators were compared with simulated data to examine several general questions: what are the impacts of microsatellite null alleles on , how should missing data be treated, and does disequilibrium contributed by reduced recombination among some loci in a sample impact . Estimators differed greatly in precision in the scenarios examined, and a widely employed estimator exhibited the largest variances among replicate data sets. speed‐ne implements several jackknife approaches to estimate confidence intervals, and simulated data showed that jackknifing over loci and jackknifing over individuals provided ~95% confidence interval coverage for some estimators and should be useful for empirical studies. speed‐ne provides an open‐source extensible tool for estimation of from empirical genotype data and to conduct simulations of both microsatellite and single nucleotide polymorphism (SNP) data types to develop expectations and to compare estimators.     

Effects of chronic elevated nitrate concentrations on the structure and function of river biofilms

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Deep soil carbon dynamics are driven more by soil type than by climate: a worldwide meta‐analysis of radiocarbon profiles

The response of soil carbon dynamics to climate and land‐use change will affect both the future climate and the quality of ecosystems. Deep soil carbon (>20 cm) is the primary component of the soil carbon pool, but the dynamics of deep soil carbon remain poorly understood. Therefore, radiocarbon activity (C), which is a function of the age of carbon, may help to understand the rates of soil carbon biodegradation and stabilization. We analyzed the published C contents in 122 profiles of mineral soil that were well distributed in most of the large world biomes, except for the boreal zone. With a multivariate extension of a linear mixed‐effects model whose inference was based on the parallel combination of two algorithms, the expectation–maximization (EM) and the Metropolis–Hasting algorithms, we expressed soil C profiles as a four‐parameter function of depth. The four‐parameter model produced insightful predictions of soil C as dependent on depth, soil type, climate, vegetation, land‐use and date of sampling (). Further analysis with the model showed that the age of topsoil carbon was primarily affected by climate and cultivation. By contrast, the age of deep soil carbon was affected more by soil taxa than by climate and thus illustrated the strong dependence of soil carbon dynamics on other pedologic traits such as clay content and mineralogy.     

The influence of copper on alkaline protease stability toward autolysis and thermal inactivation

Alkaline proteases are one of the most important group of enzymes that are indispensable in a number of different industrial sectors. In this work, the effect of copper ions (Cu²⁺) was investigated for improving the thermostability and hydrolytic performance of Bacillus clausii GMBE 42 alkaline protease at different temperatures (45–65°C). Maximal residual activity was observed in the presence of 5 mM CuCl₂. The enzyme was thermoinactivated according to first‐order kinetics. A stabilization effect caused by copper ions was the result of a decrease in both autolysis and thermoinactivation rates. Thermodynamic analysis of the thermoinactivation process showed that E_a,i, ΔG_i, and ΔH_i values of the enzyme were higher in the presence of copper ions, but there was no measurable change in ΔS_i values. These results show the thermostabilizing potential of copper ions on the enzyme. Lower K_m values and higher k_cat and k_cat/K_m values were obtained in the presence of copper ions, which is an indication of the nonessential activation of the enzyme by copper ions. Thermodynamic analysis of casein hydrolysis showed that in presence of copper ions E_a, ΔG^≠, ΔH^≠, , and values of enzyme were lower, but there was no change in ΔS^≠ values. This is so far the first study that investigates the effect of cations on the basic catalytic and thermodynamic properties of an alkaline serine protease, which may be used to remove protein wastes from various industries such as food and leather processing.     

Different fates of deposited and in a temperate forest in northeast China: a 15N tracer study

Increasing atmospheric reactive nitrogen (N) deposition due to human activities could change N cycling in terrestrial ecosystems. However, the differences between the fates of deposited and are still not fully understood. Here, we investigated the fates of deposited and , respectively, via the application of ¹⁵NH₄NO₃ and NH₄¹⁵NO₃ in a temperate forest ecosystem. Results showed that at 410 days after tracer application, most was immobilized in litter layer (50 ± 2%), while a considerable amount of penetrated into 0–5 cm mineral soil (42 ± 2%), indicating that litter layer and 0–5 cm mineral soil were the major N sinks of and , respectively. Broad‐leaved trees assimilated more ¹⁵N under NH₄¹⁵NO₃ treatment compared to under ¹⁵NH₄NO₃ treatment, indicating their preference for –N. At 410 days after tracer application, 16 ± 4% added ¹⁵N was found in aboveground biomass under treatment, which was twice more than that under treatment (6 ± 1%). At the same time, approximately 80% added ¹⁵N was recovered in soil and plants under both treatments, which suggested that this forest had high potential for retention of deposited N. These results provided evidence that there were great differences between the fates of deposited and , which could help us better understand the mechanisms and capability of forest ecosystems as a sink of reactive nitrogen.     

Nest occurrence and survival of King Rails in fire‐managed coastal marshes in North Carolina and Virginia

Prescribed burning maintains marsh habitat, but its impact on breeding King Rails (Rallus elegans) is poorly understood. This practice may serve as a means to enhance populations of a species whose numbers are declining in the southeastern United States. We used call‐broadcast surveys and nest searches to categorize the state of occupied plots by the presence or non‐presence of nesting activity in the Back Bay region, North Carolina and Virginia, in 2010. We also used nest video surveillance to estimate nest survival in 2009 and 2010. The probabilities that a surveyed plot was occupied (¹) and contained an active nest (²) were higher in recently burned marsh plots (0–1 year‐since‐burn [YSB]) than in plots with ≥2 YSB at Mackay Island and Back Bay National Wildlife Refuges (NWRs). Highest probabilities were recorded in 0–1 YSB plots at Mackay Island NWR ( = 0.96 ± 0.04, = 0.75 ± 0.18), and the lowest in ≥2 YSB plots at Back Bay NWR ( = 0.21 ± 0.10, = 0.03 ± 0.04). Nest survival from egg laying to hatching (31 d) was 0.48 (95% confidence interval [CI] = 0.06–0.83), with an estimated 0.79 (95% CI = 0.29–0.96) survival rate for the incubation stage (21 d). These nest survival estimates for King Rails in fire‐managed marshes were similar to estimates for other populations. Measures of vegetation cover, proxies for concealment, did not differ between nest sites and unused sites, even within recently burned marshes. This lack of differences in vegetation structure suggests that regrowth occurs rapidly during the period between burning (winter months) and the onset of reproduction (late April). Thus, recently burned marshes may benefit nesting King Rails by providing nest concealment. In addition, burned marshes may enhance availability of many invertebrates. Although we found that the probability that surveyed plots contained active nests was higher in recently burned marsh plots, estimates of fledging success are needed before marsh burns can be considered an effective means of fostering population growth.     

The role of a cytosolic superoxide dismutase in barley–pathogen interactions

Reactive oxygen species (ROS), including superoxide ( / ) and hydrogen peroxide (H₂O₂), are differentially produced during resistance responses to biotrophic pathogens and during susceptible responses to necrotrophic and hemi‐biotrophic pathogens. Superoxide dismutase (SOD) is responsible for the catalysis of the dismutation of / to H₂O₂, regulating the redox status of plant cells. Increased SOD activity has been correlated previously with resistance in barley to the hemi‐biotrophic pathogen Pyrenophora teres f. teres (Ptt, the causal agent of the net form of net blotch disease), but the role of individual isoforms of SOD has not been studied. A cytosolic CuZnSOD, HvCSD1, was isolated from barley and characterized as being expressed in tissue from different developmental stages. HvCSD1 was up‐regulated during the interaction with Ptt and to a greater extent during the resistance response. Net blotch disease symptoms and fungal growth were not as pronounced in transgenic HvCSD1 knockdown lines in a susceptible background (cv. Golden Promise), when compared with wild‐type plants, suggesting that cytosolic / contributes to the signalling required to induce a defence response to Ptt. There was no effect of HvCSD1 knockdown on infection by the hemi‐biotrophic rice blast pathogen Magnaporthe oryzae or the biotrophic powdery mildew pathogen Blumeria graminis f. sp. hordei, but HvCSD1 also played a role in the regulation of lesion development by methyl viologen. Together, these results suggest that HvCSD1 could be important in the maintenance of the cytosolic redox status and in the differential regulation of responses to pathogens with different lifestyles.     

Characterization of the superoxide anion radical scavenging activity by tetracycline antibiotics in aprotic media

The tetracycline family antibiotics are widely used as human and veterinary treatments. The drugs are effective as antibiotics and also show antimicrobial and non‐microbial action. However, the antioxidant properties of tetracyclines have not been characterized in aprotic media. To better understand their biological functions, the in vitro superoxide anion radical () scavenging activities of tetracycline, chlortetracycline, oxytetracycline, doxycycline and methacycline were characterized, along with a very efficient scavenger, tiron, in dimethyl sulphoxide (DMSO), using ultra‐weak chemiluminescence (CL). We found that tetracycline, chlortetracycline and doxycycline efficiently inhibited CL from the ‐generating system at concentration levels of 0.02–1.0 mmol/L. Methacycline and oxytetracycline were the scavengers at concentration levels of 0.01–0.1 mmol/L, whereas when their concentration was lowered the drugs were capable of generating , leading to CL enhancement. For all the data obtained in this study, the scavenging activity for the compounds tested decreased in the following order: tetracycline > doxycycline > chlortetracycline > tiron methacycline > oxytetracycline. These results indicate that the tetracycline drugs directly alter redox chemistry in aprotic media. Copyright © 2011 John Wiley & Sons, Ltd.     

Superoxide anion radical scavenging property of catecholamines

The direct effect of the four catecholamines (adrenaline, noradrenaline, dopamine and isoproterenol) on superoxide anion radicals () was investigated. The reaction between 18‐crown‐6‐ether and potassium superoxide in dimethylsulfoxide was used as a source of . The reactivity of catecholamines with was examined using chemiluminescence, reduction of nitroblue tetrazolium and electron paramagnetic resonance spin‐trapping techniques. 5,5‐Dimethyl‐1‐pyrroline‐N‐oxide was included as the spin trap. The results showed that the four catecholamines were effective and efficient in inhibiting chemiluminescence accompanying the potassium superoxide/18‐crown‐6‐ether system in a dose‐dependent manner over the range 0.05–2 mm in the following order: adrenaline > noradrenaline > dopamine > isoproterenol, with, IC₅₀ = 0.15 ± 0.02 mm 0.21 ± 0.03 mm , 0.27 ± 0.03 mm and 0.50 ± 0.04 mm , respectively. The catecholamines examined also exhibited a strong scavenging effect towards when evaluated this property by the inhibition of nitroblue tetrazolium reduction (56–73% at 1 m concentration). A very similar capacity of scavenging was monitored in the 5,5‐dimethyl‐1‐pyrroline‐N‐oxide spin‐trapping assay. The results suggest that catecholamines tested may involve a direct effect on scavenging radicals. Copyright © 2013 John Wiley & Sons, Ltd.