Priming and memory of stress responses in organisms lacking a nervous system

Experience and memory of environmental stimuli that indicate future stress can prepare (prime) organismic stress responses even in species lacking a nervous system. The process through which such organisms prepare their phenotype for an improved response to future stress has been termed ‘priming’. However, other terms are also used for this phenomenon, especially when considering priming in different types of organisms and when referring to different stressors. Here we propose a conceptual framework for priming of stress responses in bacteria, fungi and plants which allows comparison of priming with other terms, e.g. adaptation, acclimation, induction, acquired resistance and cross protection. We address spatial and temporal aspects of priming and highlight current knowledge about the mechanisms necessary for information storage which range from epigenetic marks to the accumulation of (dormant) signalling molecules. Furthermore, we outline possible patterns of primed stress responses. Finally, we link the ability of organisms to become primed for stress responses (their ‘primability’) with evolutionary ecology aspects and discuss which properties of an organism and its environment may favour the evolution of priming of stress responses.     

Urea in Weaver Ant Feces: Quantification and Investigation of the Uptake and Translocation of Urea in <Emphasis Type="Italic">Coffea arabica</Emphasis>

Weaver ants are tropical insects that nest in tree canopies, and for centuries these ants have been used for pest control in tropical orchards. Trees hosting weaver ants might benefit not only from the pest protective properties of these insects but also an additional supply of nutrients from ant feces deposited on the leaves. In a recent study, we demonstrated that Coffea arabica plants hosting Oecophylla smaragdina weaver ants under laboratory conditions experienced enhanced nitrogen availability compared with plants grown without ants. Therefore, the aim of the present study was to further investigate the interactions of weaver ants with the host plants with respect to plant nutrition. Here, we report the identification and quantification of urea, a highly effective foliar nutrient present in the fecal depositions of O. smaragdina. Feces samples obtained from six O. smaragdina colonies were analyzed, and urea concentrations ranging from 1.98 to 31.05 μg/mg ant feces were detected. Subsequently, we investigated the uptake and translocation of ¹⁵N₂-urea in amounts corresponding to the estimated urea contribution via feces depositions on single host plant leaves under laboratory conditions. The results clearly demonstrated that fecal urea was not only assimilated but also translocated within the plant. This evidence strongly supports the hypothesis that the fecal urea of weaver ants is a source of nitrogen for the host trees. Thus, weaver ant feces likely contribute to an improved nutritional status of ant-hosting trees in tropical orchards, thereby adding value to the use of weaver ants for the biocontrol of insect pests.     

Regulatory hurdles for genome editing: process- vs. product-based approaches in different regulatory contexts

Novel plant genome editing techniques call for an updated legislation regulating the use of plants produced by genetic engineering or genome editing, especially in the European Union. Established more than 25 years ago and based on a clear distinction between transgenic and conventionally bred plants, the current EU Directives fail to accommodate the new continuum between genetic engineering and conventional breeding. Despite the fact that the Directive 2001/18/EC contains both process- and product-related terms, it is commonly interpreted as a strictly process-based legislation. In view of several new emerging techniques which are closer to the conventional breeding than common genetic engineering, we argue that it should be actually interpreted more in relation to the resulting product. A legal guidance on how to define plants produced by exploring novel genome editing techniques in relation to the decade-old legislation is urgently needed, as private companies and public researchers are waiting impatiently with products and projects in the pipeline. We here outline the process in the EU to develop a legislation that properly matches the scientific progress. As the process is facing several hurdles, we also compare with existing frameworks in other countries and discuss ideas for an alternative regulatory system.     

Euthanasia in Belgium: trends in reported cases between 2003 and 2013

Background:In 2002, the Belgian Act on Euthanasia came into effect, regulating the intentional ending of life by a physician at the patient’s explicit request. We undertook this study to describe trends in officially reported euthanasia cases in Belgium with regard to patients’ sociodemographic and clinical profiles, as well as decision-making and performance characteristics.Methods:We used the database of all euthanasia cases reported to the Federal Control and Evaluation Committee on Euthanasia in Belgium between Jan. 1, 2003, and Dec. 31, 2013 (n = 8752). The committee collected these data with a standardized registration form. We analyzed trends in patient, decision-making and performance characteristics using a χ² technique. We also compared and analyzed trends for cases reported in Dutch and in French.Results:The number of reported euthanasia cases increased every year, from 235 (0.2% of all deaths) in 2003 to 1807 (1.7% of all deaths) in 2013. The rate of euthanasia increased significantly among those aged 80 years or older, those who died in a nursing home, those with a disease other than cancer and those not expected to die in the near future (p < 0.001 for all increases). Reported cases in 2013 most often concerned those with cancer (68.7%) and those under 80 years (65.0%). Palliative care teams were increasingly often consulted about euthanasia requests, beyond the legal requirements to do so (p < 0.001). Among cases reported in Dutch, the proportion in which the person was expected to die in the foreseeable future decreased from 93.9% in 2003 to 84.1% in 2013, and palliative care teams were increasingly consulted about the euthanasia request (from 34.0% in 2003 to 42.6% in 2013). These trends were not significant for cases reported in French.Interpretation:Since legalization of euthanasia in Belgium, the number of reported cases has increased each year. Most of those receiving euthanasia were younger than 80 years and were dying of cancer. Given the increases observed among non–terminally ill and older patients, this analysis shows the importance of detailed monitoring of developments in euthanasia practice.In 2002, Belgium legalized euthanasia, defined as the intentional ending of life by a physician at the patient’s explicit request.^1,2 For a patient to be eligible for euthanasia, certain formal criteria for due care must be met.¹ These include a voluntary, well-considered, repeated and written request, expressed by a person with full mental capacity who is fully informed about his or her medical condition and the remaining therapeutic possibilities.¹ The person must be in a state of constant and unbearable physical or mental suffering that cannot be alleviated. Due care criteria for the procedure include an a priori consultation with a second independent physician, consultation with a third physician in cases where death is not expected in the foreseeable future and a posteriori reporting of the case for evaluation purposes.¹To safeguard due process and legal compliance and to enable societal control and evaluation, a mandatory notification procedure was built into the legislation.³ Physicians are required to report each case of euthanasia to the multidisciplinary Belgian Federal Control and Evaluation Committee on Euthanasia by completing and submitting a registration form within 4 working days after a death by euthanasia.^1,3 The evaluation committee reviews the form and determines whether euthanasia was performed in accordance with the legal requirements. Initially, only anonymous information is reviewed; where there is doubt about legality, the committee can revoke anonymity by majority decision and can ask the reporting physician for additional information. If the committee is of the opinion, based on a two-thirds majority, that the legal requirements were not fulfilled, the case is sent to the public prosecutor.^1,3 Although not mentioned in the Belgian law, physician-assisted suicide is treated as a form of euthanasia by the committee.⁴To facilitate societal control, the Federal Control and Evaluation Committee on Euthanasia is legally required to issue biennial reports of all reported cases,^1,3–9 providing basic statistics, an evaluation of the law and further recommendations. However, these statistics do not provide an overview of long-term trends. A more complete and thorough evaluation of case characteristics and analysis of trends is needed. In this way, adherence to the legal criteria can be evaluated, and developments in euthanasia practice that might raise concerns can be identified and addressed.Belgium has 2 main language communities: those who speak Dutch (roughly 60% of the population), who mainly live in Flanders, and those who speak French (about 40%), who mainly live in Wallonia. The Brussels-Capital Region is officially bilingual, but predominantly French-speaking. Several empirical studies have found differences in end-of-life practices, knowledge and attitudes between the regions and language communities, showing that Dutch-speaking physicians more often receive and grant euthanasia requests and are more inclined to adhere to legal safeguards.^10–14 The reports issued by the Federal Control and Evaluation Committee on Euthanasia show a striking disparity in euthanasia reporting between the 2 language communities.^4–9 Trends in the characteristics of reported cases and differences among them have not yet been studied.The committee’s reports have shown a continuing increase in the number of euthanasia cases.^4–9 The primary objective of this study was to examine changes in the number and incidence of euthanasia cases and the proportion of euthanasia cases relative to all deaths in Belgium up to and including 2013. The secondary objectives were to determine and report the sociodemographic and clinical characteristics of patients, the decision-making and performance characteristics of reported cases and the differences in trends in characteristics between cases reported in Dutch and cases reported in French.     

Multiparametric Functional MRI: A Tool to Uncover Subtle Changes following Allogeneic Renal Transplantation

PurposeTo investigate multiparametric functional MRI to characterize acute rejection in a murine allogeneic renal transplant model and evaluate the effect of novel therapeutics.ResultsDWI showed a significant diffusion restriction in allogeneic compared to syngeneic transplants (ADC: 0.63±0.08 vs. 1.29±0.12 mm²/s*10³) with decreasing diffusion restriction under therapy. DCE-MRI showed restored organ perfusion under Ciclosporin A alone and combination therapy (Plasma Flow: 43.43±12.49; 38.75±7.53ml/100ml/min) compared to syngeneic controls (51.03±12.49ml/100ml/min). Ex vivo analysis showed reduced monocytic infiltrates, attenuated levels of inflammatory cytokines under mNOX-E36 monotherapy with an additive effect of low dose Ciclosporin A. There was a significant (p<0.05) negative correlation between ADC and interstitial inflammation (r = -0.73) or macrophage infiltration (r = -0.81) and between organ perfusion and intimal arteritis (r = -0.63).ConclusionMultiparametric functional MRI is suited to detect renal allograft rejection in an experimental murine model and allows to characterize effects of immunosuppressive therapy alleviating acute rejection processes in allogeneic transplantation.     

Plant genome editing in the European Union—to be or not to be—a GMO

New plant-breeding techniques have been boosting plant breeding, since only a few years but already first promising products are pushing to the market. In contrast to this, in many countries, the current Directives regulating genetically modified organisms have been established more than 25 years ago, especially in the European Union being based on clear differentiation between transgenic plants and conventional breeding. Therefore, the question arises if these Directives are suitable to face the new challenge of genetic engineering or if there is a need for updated regulations.     

Naphthoquinone glycosides for bioelectroanalytical enumeration of the faecal indicator Escherichia coli

Microbial water quality monitoring for the presence of faecal indicator bacteria (FIB) is a mandatory activity in many countries and is key in public health protection. Despite technological advances and a need for methodological improvements, chromogenic and fluorogenic enzymatic techniques remain the mainstays of water quality monitoring for both public health agencies and regulated utilities. We demonstrated that bioelectroanalytical approaches to FIB enumeration are possible and can be achieved using commercially available enzyme‐specific resorufin glycosides, although these are expensive, not widely available or designed for purpose. Following this, we designed two naphthoquinone glycosides which performed better, achieving Escherichia coli detection in the range 5.0 × 10² to 5.0 × 10⁵ CFU ml^?1 22–54% quicker than commercially available resorufin glycosides. The molecular design of the naphthoquinone glycosides requires fewer synthetic steps allowing them to be produced for as little as US$50 per kg. Tests with environmental samples demonstrated the low tendency for abiotic interference and that, despite specificity being maintained between β‐glucuronidase and β‐galactosidase, accurate enumeration of E. coli in environmental samples necessitates development of a selective medium. In comparison to a commercially available detection method, which has U.S. Environmental Protection Agency (EPA) approval, our approach performed better at high organism concentrations, detecting 500 organisms in 9 h compared with 13.5 h for the commercial method. Bioelectroanalytical detection is comparable to current approved methods and with further development could result in improved detection times. A recent trend for low‐cost open‐source hardware means that automated, potentiostatically controlled E. coli detection systems could be constructed for less than US$100 per channel.     

Locus- and Site-Specific DNA Methylation of 19 kDa Zein Genes in Maize

An interesting question in maize development is why only a single zein gene is highly expressed in each of the 19-kDa zein gene clusters (A and B types), z1A2-1 and z1B4, in the immature endosperm. For instance, epigenetic marks could provide a structural difference. Therefore, we investigated the DNA methylation of the arrays of gene copies in both promoter and gene body regions of leaf (non-expressing tissue as a control), normal endosperm, and cultured endosperm. Although we could show that expressed genes have much lower methylation levels in promoter regions than silent ones in both leaf and normal endosperm, there was surprisingly also a difference in the pattern of the z1A and z1B gene clusters. The expression of z1B gene is suppressed by increased DNA methylation and activated with reduced DNA methylation, whereas z1A gene expression is not. DNA methylation in gene coding regions is higher in leaf than in endosperm, whereas no significant difference is observed in gene bodies between expressed and non-expressed gene copies. A median CHG methylation (25–30%) appears to be optimal for gene expression. Moreover, tissue-cultured endosperm can reset the DNA methylation pattern and tissue-specific gene expression. These results reveal that DNA methylation changes of the 19-kDa zein genes is subject to plant development and tissue culture treatment, but varies in different chromosomal locations, indicating that DNA methylation changes do not apply to gene expression in a uniform fashion. Because tissue culture is used to produce transgenic plants, these studies provide new insights into variation of gene expression of integrated sequences.     

Specific effects on enzyme activities upon ditution of petroselinum hortense cell cultures into water.

Large increases in the specific activities of phenylalanine ammonia-lyase (EC 4.3.1.5) and p-coumarate:CoA ligase (EC 6.2.1.-) occurred within a few hours after dilution of cultured Petroselinum hortense cells into water. No significant changes in the total amount of extractable protein and in the activities of chalcone isomerase (EC 5.5.1.6) and glutamate dehydrogenase (EC 1.4.1.2) were observed under the same conditions. The time course for the change in phenylalanine ammonia-lyase activity included a lag period of 2–3 h, a peak about 13 h after the onset of induction, and a subsequent period of rapid decline. The inducible amount of enzyme activity was greatly dependent upon the degree of dilution of the cells into water. Simultaneous induction by dilution and irradiation of the cells with white, fluorescent light resulted in an increase in the phenylalanine ammonia-lyase level of activity which exceeded that calculated from the sum of the separately induced levels. Consecutive inductions, first by dilution and then 5 h later by irradiation, each required a lag period of 2–3 h. Actinomycin D or cycloheximide were inhibitors of the induction. While the total protein-synthesizing capacity in vitro was not significantly influenced by the dilution of cells, analysis of the labeled products on polyacrylamide gels demonstrated small but significant changes in the radioactivity profiles. The results are consistent with the hypothesis that dilution of the cells into water reduces the concentration of one or more compounds of cellular origin, thereby stimulating the rate of de novo synthesis of a limited number of proteins.