Turnover in local parasite populations temporarily favors host outcrossing over self‐fertilization during experimental evolution

The ubiquity of outcrossing in plants and animals is difficult to explain given its costs relative to self‐fertilization. Despite these costs, exposure to changing environmental conditions can temporarily favor outcrossing over selfing. Therefore, recurring episodes of environmental change are predicted to favor the maintenance of outcrossing. Studies of host–parasite coevolution have provided strong support for this hypothesis. However, it is unclear whether multiple exposures to novel parasite genotypes in the absence of coevolution are sufficient to favor outcrossing. Using the nematode Caenorhabditis elegans and the bacterial parasite Serratia marcescens, we studied host responses to parasite turnover. We passaged several replicates of a host population that was well‐adapted to the S. marcescens strain Sm2170 with either Sm2170 or one of three novel S. marcescens strains, each derived from Sm2170, for 18 generations. We found that hosts exposed to novel parasites maintained higher outcrossing rates than hosts exposed to Sm2170. Nonetheless, host outcrossing rates declined over time against all but the most virulent novel parasite strain. Hosts exposed to the most virulent novel strain exhibited increased outcrossing rates for approximately 12 generations, but did not maintain elevated levels of outcrossing throughout the experiment. Thus, parasite turnover can transiently increase host outcrossing. These results suggest that recurring episodes of parasite turnover have the potential to favor the maintenance of host outcrossing. However, such maintenance may require frequent exposure to novel virulent parasites, rapid rates of parasite turnover, and substantial host gene flow.     

Kinetic and docking studies of the interaction of quinones with the quinone reductase active site

NAD(P)H/quinone acceptor oxidoreductase type 1 (QR1) protects cells from cytotoxic and neoplastic effects of quinones though two-electron reduction. Kinetic experiments, docking, and binding affinity calculations were performed on a series of structurally varied quinone substrates. A good correlation between calculated and measured binding affinities from kinetic determinations was obtained. The experimental and theoretical studies independently support a model in which quinones (with one to three fused aromatic rings) bind in the QR1 active site utilizing a pi-stacking interaction with the isoalloxazine ring of the FAD cofactor.     

Light inhibition of leaf respiration as soil fertility declines along a post-glacial chronosequence in New Zealand: an analysis using the Kok method

Background and aims

Our study quantified variations leaf respiration in darkness (R _D) and light (R _L), and associated traits along the Franz Josef Glacier soil development chronosequence in New Zealand.

Methods

At six sites along the chronosequence (soil age: 6, 60, 150, 500, 12,000 and 120,000 years old), we measured rates of leaf R _D, R _L (using Kok method), light-saturated CO₂ assimilation rates (A), leaf mass per unit area (M _A), and concentrations of leaf nitrogen ([N]), phosphorus ([P]), soluble sugars and starch.

Results

The chronosequence was characterised by decreasing R _D, R _L and A, reduced [N] and [P] and increasing M _A as soil age increased. Light inhibition of R occurred across the chronosequence (mean inhibition = 16 %), resulting in ratios of R _L:A being lower than for R _D:A. Importantly, the degree of light inhibition differed across the chronosequence, being lowest at young sites and highest at old sites. This resulted in R _L:A ratios being relatively constant across the chronosequence, whereas R _D:A ratios increased with increasing soil age. Log-log R-A-M _A-[N] relationships remained constant along the chronosequence. By contrast, relationships linking rates of leaf R to [P] differed among leaves with low vs high [N]:[P] ratios. Slopes of log-log bivariate relationships linking R _L to A, M _A, [N] and [P] were steeper than that for R _D.

Conclusions

Our findings have important implications for predictive models that seek to account for light inhibition of R, and for our understanding of how environmental gradients impact on leaf trait relationships     

Antioxidant activity contributes to flavonol cardioprotection during reperfusion of rat hearts

The mechanism of flavonol-induced cardioprotection is unclear. We compared the protective actions of a flavonol that inhibits calcium utilization and has antioxidant activity, 3′,4′-dihydroxyflavonol (DiOHF); a flavonol that affects only calcium activity, 4′-OH-3′-OCH₃-flavonol (4′-OH-3′-OCH₃F); and a water-soluble flavonol with selective antioxidant activity, DiOHF-6-succinamic acid (DiOHF-6-SA), in isolated, perfused rat hearts. Hearts were subjected to global ischemia for 20 min followed by 30 min reperfusion and were treated with vehicle (0.05% DMSO), DiOHF, 4′-OH-3′-OCH₃F, or DiOHF-6-SA (all 10 μM, n = 5-8 per group). Flavonols were infused for 10 min before ischemia and during reperfusion. In vehicle-treated hearts, left-ventricular (LV) + dP/dt was reduced by 60% at the end of reperfusion compared to the preischemic level. Lactate dehydrogenase (LDH) release was elevated and endothelial NO synthase (eNOS) expression was lower in vehicle-treated hearts compared to shams. In comparison, DiOHF treatment improved LV function upon reperfusion, decreased LDH, and preserved eNOS expression. The antioxidant DiOHF-6-SA also preserved contractility, reduced LDH, and preserved eNOS expression. In contrast, hearts treated with 4′-OH-3′-OCH₃F showed a degree of contractile impairment similar to that of the vehicle group. DiOHF and DiOHF-6-SA also exerted cardioprotection when given only during reperfusion and not when administered only before ischemia. Flavonol-induced cardioprotection relies on antioxidant activity and is mainly exerted during reperfusion.     

Symbiont-mediated RNA interference in insects

RNA interference (RNAi) methods for insects are often limited by problems with double-stranded (ds) RNA delivery, which restricts reverse genetics studies and the development of RNAi-based biocides. We therefore delegated to insect symbiotic bacteria the task of: (i) constitutive dsRNA synthesis and (ii) trauma-free delivery. RNaseIII-deficient, dsRNA-expressing bacterial strains were created from the symbionts of two very diverse pest species: a long-lived blood-sucking bug, Rhodnius prolixus, and a short-lived globally invasive polyphagous agricultural pest, western flower thrips (Frankliniella occidentalis). When ingested, the manipulated bacteria colonized the insects, successfully competed with the wild-type microflora, and sustainably mediated systemic knockdown phenotypes that were horizontally transmissible. This represents a significant advance in the ability to deliver RNAi, potentially to a large range of non-model insects.     

Changes in Inositol 1,4,5-Trisphosphate and Inositol 1,3,4,5-Tetrakisphosphate Mass Accumulations in Cultured Adrenal Chromaffin Cells in Response to Bradykinin and Histamine

In previous studies it has been shown that both bradykinin and histamine increase the formation of 3H-labeled inositol phosphates in adrenal chromaffin cells prelabelled with [3H]inositol and that both these agonists stimulate release of catecholamines by a mechanism dependent on extracellular calcium. Here, we have used mass assays of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4] to investigate changes in levels of these two candidates as second messengers in response to stimulation with bradykinin and histamine. Bradykinin increased the mass of Ins(1,3,4,5)P4 despite the failure in earlier studies with [3H]inositol-labelled cells to observe a bradykinin-mediated increase in content of [3H]InsP4. Bradykinin elicited a very rapid increase in level of Ins(1,4,5)P3, which was maximal at 5-10 s and then rapidly decreased to a small but sustained elevation at 2 min. The bradykinin-elicited Ins(1,3,4,5)P4 response increased to a maximum at 30-60 s and at 2 min was still elevated severalfold above basal levels. Histamine, which produced a larger overall total inositol phosphate response in [3H]inositol-loaded cells, produced significantly smaller Ins(1,4,5)P3 and Ins(1,3,4,5)P4 responses compared with bradykinin. The bradykinin stimulation of Ins(1,4,5)P3 accumulation was partially dependent on a high (1.8 mM) extracellular Ca2+ concentration, whereas the Ins(1,3,4,5)P4 response was almost completely lost when the extracellular Ca2+ concentration was reduced to 100 nM. Changes in the inositol polyphosphate second messengers are compared with the time course of bradykinin-stimulated increases in free intracellular Ca2+ concentrations and noradrenaline release.     

Reversible inhibitors of regulators of G-protein signaling identified in a high-throughput cell-based calcium signaling assay

Regulator of G-protein signaling (RGS) proteins potently suppress G-protein coupled receptor (GPCR) signal transduction by accelerating GTP hydrolysis on activated heterotrimeric G-protein α subunits. RGS4 is enriched in the CNS and is proposed as a therapeutic target for treatment of neuropathological states including epilepsy and Parkinson's disease. Therefore, identification of novel RGS4 inhibitors is of interest. An HEK293-FlpIn cell-line stably expressing M₃-muscarinic receptor with doxycycline-regulated RGS4 expression was employed to identify compounds that inhibit RGS4-mediated suppression of M₃-muscarinic receptor signaling. Over 300,000 compounds were screened for an ability to enhance Gα_q-mediated calcium signaling in the presence of RGS4. Compounds that modulated the calcium response in a counter-screen in the absence of RGS4 were not pursued. Of the 1365 RGS4-dependent primary screen hits, thirteen compounds directly target the RGS-G-protein interaction in purified systems. All thirteen compounds lose activity against an RGS4 mutant lacking cysteines, indicating that covalent modification of free thiol groups on RGS4 is a common mechanism. Four compounds produce > 85% inhibition of RGS4-G-protein binding at 100 μM, yet are > 50% reversible within a ten-minute time frame. The four reversible compounds significantly alter the thermal melting temperature of RGS4, but not G-protein, indicating that inhibition is occurring through interaction with the RGS protein. The HEK cell-line employed for this study provides a powerful tool for efficiently identifying RGS-specific modulators within the context of a GPCR signaling pathway. As a result, several new reversible, cell-active RGS4 inhibitors have been identified for use in future biological studies.     

Clinical examinations on crab-eating macaques in mauritius

Clinical inspections were carried out in order to obtain sufficient information concerning the physiological characteristics and states of natural infection by various pathogenic agents in the population of crab-eating macaques (Macaca fascicularis) on Mauritius. The hematological and plasma biochemical values were within the normal ranges, showing no morbid signs. The intestinal parasitic appearance was so simplified that onlyStrongyloides andTrichuris were noted as helminthic findings. All of the monkeys examined were free from enteropathogenic organisms such asShigella andSalmonella, and were negative for measles, herpes simplex type 1, simian immunodeficiency virus/MAC, and SV5 antibodies. These data suggest that the macaques in Mauritius are considerably spared from natural infections by various pathogenic agents. This study was carried out under the humane conditions prescribed in “The Guidelines for the Study of Wild Primates and Use of Wild-born Primates” by the Primate Research Institute, Kyoto University, Japan.     

The use of artificial neural networks for the selection of the most appropriate formulation and processing variables in order to predict the in vitro dissolution of sustained release minitablets

The objective of this work was to apply artificial neural networks (ANNs) to examine the relative importance of various factors, both formulation and process, governing the in-vitro dissolution from enteric-coated sustained release (SR) minitablets. Input feature selection (IFS) algorithms were used in order to give an estimate of the relative importance of the various formulation and processing variables in determining minitablet dissolution rate. Both forward and backward stepwise algorithms were used as well as genetic algorithms. Networks were subsequently trained using the back propagation algorithm in order to check whether or not the IFS process had correctly located any unimportant inputs. IFS gave consistent rankings for the importance of the various formulation and processing variables in determining the release of drug from minitablets. Consistent ranking was achieved for both indices of the release process; ie, the time taken for release to commence through the enteric coat (T_lag) and that for the drug to diffuse through the SR matrix of the minitablet into the dissolution medium (T_90-10). In the case of the T_lag phase, the main coating parameters, along with the original batch blend size and the blend time with lubricant, were found to have most influence. By contrast, with the T_90-10 phase, the amounts of matrix forming polymer and direct compression filler were most important. In the subsequent training of the ANNs, removal of inputs regarded as less important led to improved network performance. ANNs were capable of ranking the relative importance of the various formulations and processing variables that influenced the release rate of the drug from minitablets. This could be done for all main stages of the release process. Subsequent training of the ANN verified that removal of less relevant inputs from the training process led to an improved performance from the ANN.