Can natural forest expansion contribute to Europe's restoration policy agenda? An interdisciplinary assessment.

Natural forest expansion (NFE), that is, the establishment of secondary forest on non-forested land through natural succession, has substantially contributed to the widespread expansion of forests in Europe over the last few decades. So far, EU policies have largely neglected the potential of NFE for meeting policy objectives on restoration. Synthesising recent interdisciplinary research, this paper assesses the challenges and opportunities of NFE in view of contributing to European forest and ecosystem restoration. Specifically, we discuss the potential for supporting climate change mitigation and adaptation, biodiversity conservation, and forestry and economic use, summarize the current knowledge about societal perceptions and the policymaking on NFE, and make policy recommendations to better use the potential of NFE. We conclude that NFE has the potential to contribute to the European restoration policy agenda if local contexts and possible trade-offs are properly considered.

A demo-genetic model shows how silviculture reduces natural density-dependent selection in tree populations.

Biological production systems and conservation programs benefit from and should care for evolutionary processes. Developing evolution-oriented strategies requires knowledge of the evolutionary consequences of management across timescales. Here, we used an individual-based demo-genetic modelling approach to study the interactions and feedback between tree thinning, genetic evolution, and forest stand dynamics. The model combines processes that jointly drive survival and mating success-tree growth, competition and regeneration-with genetic variation of quantitative traits related to these processes. In various management and disturbance scenarios, the evolutionary rates predicted by the coupled demo-genetic model for a growth-related trait, vigor, fit within the range of empirical estimates found in the literature for wild plant and animal populations. We used this model to simulate non-selective silviculture and disturbance scenarios over four generations of trees. We characterized and quantified the effect of thinning frequencies and intensities and length of the management cycle on viability selection driven by competition and fecundity selection. The thinning regimes had a drastic long-term effect on the evolutionary rate of vigor over generations, potentially reaching 84% reduction, depending on management intensity, cycle length and disturbance regime. The reduction of genetic variance by viability selection within each generation was driven by changes in genotypic frequencies rather than by gene diversity, resulting in low-long-term erosion of the variance across generations, despite short-term fluctuations within generations. The comparison among silviculture and disturbance scenarios was qualitatively robust to assumptions on the genetic architecture of the trait. Thus, the evolutionary consequences of management result from the interference between human interventions and natural evolutionary processes. Non-selective thinning, as considered here, reduces the intensity of natural selection, while selective thinning (on tree size or other criteria) might reduce or reinforce it depending on the forester's tree choice and thinning intensity.

Seed predation-induced Allee effects, seed dispersal and masting jointly drive the diversity of seed sources during population expansion.

The environmental factors affecting plant reproduction and effective dispersal, in particular biotic interactions, have a strong influence on plant expansion dynamics, but their demographic and genetic consequences remain an understudied body of theory. Here, we use a mathematical model in a one-dimensional space and on a single reproductive period to describe the joint effects of predispersal seed insect predators foraging strategy and plant reproduction strategy (masting) on the spatio-temporal dynamics of seed sources diversity in the colonisation front of expanding plant populations. We show that certain foraging strategies can result in a higher seed predation rate at the colonisation front compared to the core of the population, leading to an Allee effect. This effect promotes the contribution of seed sources from the core to the colonisation front, with long-distance dispersal further increasing this contribution. As a consequence, our study reveals a novel impact of the predispersal seed predation-induced Allee effect, which mitigates the erosion of diversity in expanding populations. We use rearrangement inequalities to show that masting has a buffering role: it mitigates this seed predation-induced Allee effect. This study shows that predispersal seed predation, plant reproductive strategies and seed dispersal patterns can be intermingled drivers of the diversity of seed sources in expanding plant populations, and opens new perspectives concerning the analysis of more complex models such as integro-difference or reaction-diffusion equations.

Infection with Influenzavirus A in a murine model induces epithelial bronchial lesions and distinct waves of innate immune-cell recruitment.

Introduction: Inflammatory lesions after Influenza A viruses (IAV) are potential therapeutic target for which better understanding of post-infection immune mechanisms is required. Most studies to evaluate innate immune reactions induced by IAV are based on quantitative/functional methods and anatomical exploration is most often non-existent. We aimed to study pulmonary damage and macrophage recruitment using two-photon excitation microscopy (TPEM) after IAV infection. Methods: We infected C57BL/6 CD11c+YFP mice with A/Puerto Ricco/8/34 H1N1. We performed immune cell analysis, including flow cytometry, cytokine concentration assays, and TPEM observations after staining with anti-F4/80 antibody coupled to BV421. We adapted live lung slice (LLS) method for ex-vivo intravital microscopy to analyze cell motility. Results: TPEM provided complementary data to flow cytometry and cytokine assays by allowing observation of bronchial epithelium lesions and spreading of local infection. Addition of F4/80-BV421 staining allowed us to precisely determine timing of recruitment and pulmonary migration of macrophages. Ex-vivo LLS preserved cellular viability, allowing us to observe acceleration of macrophage motility. Conclusion: After IAV infection, we were able to explore structural consequences and successive waves of innate immune cell recruitment. By combining microscopy, flow cytometry and chemokine measurements, we describe novel and precise scenario of innate immune response against IAV.

Masting is uncommon in trees that depend on mutualist dispersers in the context of global climate and fertility gradients.

The benefits of masting (volatile, quasi-synchronous seed production at lagged intervals) include satiation of seed predators, but these benefits come with a cost to mutualist pollen and seed dispersers. If the evolution of masting represents a balance between these benefits and costs, we expect mast avoidance in species that are heavily reliant on mutualist dispersers. These effects play out in the context of variable climate and site fertility among species that vary widely in nutrient demand. Meta-analyses of published data have focused on variation at the population scale, thus omitting periodicity within trees and synchronicity between trees. From raw data on 12 million tree-years worldwide, we quantified three components of masting that have not previously been analysed together: (i) volatility, defined as the frequency-weighted year-to-year variation; (ii) periodicity, representing the lag between high-seed years; and (iii) synchronicity, indicating the tree-to-tree correlation. Results show that mast avoidance (low volatility and low synchronicity) by species dependent on mutualist dispersers explains more variation than any other effect. Nutrient-demanding species have low volatility, and species that are most common on nutrient-rich and warm/wet sites exhibit short periods. The prevalence of masting in cold/dry sites coincides with climatic conditions where dependence on vertebrate dispersers is less common than in the wet tropics. Mutualist dispersers neutralize the benefits of masting for predator satiation, further balancing the effects of climate, site fertility and nutrient demands.

Calcium Phosphate Cements Combined with Blood as a Promising Tool for the Treatment of Bone Marrow Lesions.

The solid phase of a commercial calcium phosphate (Graftys® HBS) was combined with ovine or human blood stabilized either with sodium citrate or sodium heparin. The presence of blood delayed the setting reaction of the cement by ca. 7-15 h, depending on the nature of the blood and blood stabilizer. This phenomenon was found to be directly related to the particle size of the HBS solid phase, since prolonged grinding of the latter resulted in a shortened setting time (10-30 min). Even though ca. 10 h were necessary for the HBS blood composite to harden, its cohesion right after injection was improved when compared to the HBS reference as well as its injectability. A fibrin-based material was gradually formed in the HBS blood composite to end-up, after ca. 100 h, with a dense 3D organic network present in the intergranular space, thus affecting the microstructure of the composite. Indeed, SEM analyses of polished cross-sections showed areas of low mineral density (over 10-20 µm) spread in the whole volume of the HBS blood composite. Most importantly, when the two cement formulations were injected in the tibial subchondral cancellous bone in a bone marrow lesion ovine model, quantitative SEM analyses showed a highly significant difference between the HBS reference versus its analogue combined with blood. After a 4-month implantation, histological analyses clearly showed that the HBS blood composite underwent high resorption (remaining cement: ca. 13.1 ± 7.3%) and new bone formation (newly formed bone: 41.8 ± 14.7%). This was in sharp contrast with the case of the HBS reference for which a low resorption rate was observed (remaining cement: 79.0 ± 6.9%; newly formed bone: 8.6 ± 4.8%). This study suggested that the particular microstructure, induced by the use of blood as the HBS liquid phase, favored quicker colonization of the implant and acceleration of its replacement by newly formed bone. For this reason, the HBS blood composite might be worth considering as a potentially suitable material for subchondroplasty.

Common microgeographical selection patterns revealed in four European conifers.

Microgeographical adaptation occurs when the effects of directional selection persist despite gene flow. Traits and genetic loci under selection can then show adaptive divergence, against the backdrop of little differentiation at other traits or loci. How common such events are and how strong the selection is that underlies them remain open questions. Here, we discovered and analysed microgeographical patterns of genomic divergence in four European and Mediterranean conifers with widely differing life-history traits and ecological requirements (Abies alba MIll., Cedrus atlantica [Endl.] Manetti, Pinus halepensis Mill. and Pinus pinaster Aiton) by screening pairs from geographically close forest stands sampled along steep ecological gradients. We inferred patterns of genomic divergence by applying a combination of divergence outlier detection methods, demographic modelling, Approximate Bayesian Computation inferences and genomic annotation to genomic data. Surprisingly for such small geographical scales, we showed that selection is strong in all species but generally affects different loci in each. A clear signature of selection was systematically detected on a fraction of the genome, of the order of 0.1%-1% of the loci depending on the species. The novel modelling method we designed for estimating selection coefficients showed that the microgeographical selection coefficient scaled by population size (Ns) was 2-30. Our results convincingly suggest that selection maintains within-population diversity at microgeographical scales in spatially heterogeneous environments. Such genetic diversity is likely to be a major reservoir of adaptive potential, helping populations to adapt under fluctuating environmental conditions.

Importance of interindividual interactions in eco-evolutionary population dynamics: The rise of demo-genetic agent-based models.

The study of eco-evolutionary dynamics, that is of the intertwinning between ecological and evolutionary processes when they occur at comparable time scales, is of growing interest in the current context of global change. However, many eco-evolutionary studies overlook the role of interindividual interactions, which are hard to predict and yet central to selective values. Here, we aimed at putting forward models that simulate interindividual interactions in an eco-evolutionary framework: the demo-genetic agent-based models (DG-ABMs). Being demo-genetic, DG-ABMs consider the feedback loop between ecological and evolutionary processes. Being agent-based, DG-ABMs follow populations of interacting individuals with sets of traits that vary among the individuals. We argue that the ability of DG-ABMs to take into account the genetic heterogeneity-that affects individual decisions/traits related to local and instantaneous conditions-differentiates them from analytical models, another type of model largely used by evolutionary biologists to investigate eco-evolutionary feedback loops. Based on the review of studies employing DG-ABMs and explicitly or implicitly accounting for competitive, cooperative or reproductive interactions, we illustrate that DG-ABMs are particularly relevant for the exploration of fundamental, yet pressing, questions in evolutionary ecology across various levels of organization. By jointly modelling the effects of management practices and other eco-evolutionary processes on interindividual interactions and population dynamics, DG-ABMs are also effective prospective and decision support tools to evaluate the short- and long-term evolutionary costs and benefits of management strategies and to assess potential trade-offs. Finally, we provide a list of the recent practical advances of the ABM community that should facilitate the development of DG-ABMs.

Interactions between microenvironment, selection and genetic architecture drive multiscale adaptation in a simulation experiment.

When environmental conditions differ both within and among populations, multiscale adaptation results from processes at both scales and interference across scales. We hypothesize that within-population environmental heterogeneity influences the chance of success of migration events, both within and among populations, and maintains within-population adaptive differentiation. We used a simulation approach to analyse the joint effects of environmental heterogeneity patterns, selection intensity and number of QTL controlling a selected trait on local adaptation in a hierarchical metapopulation design. We show the general effects of within-population environmental heterogeneity: (i) it increases occupancy rate at the margins of distribution ranges, under extreme environments and high levels of selection; (ii) it increases the adaptation lag in all environments; (iii) it impacts the genetic variance in each environment, depending on the ratio of within- to between-populations environmental heterogeneity; (iv) it reduces the selection-induced erosion of adaptive gene diversity. Most often, the smaller the number of QTL involved, the stronger are these effects. We also show that both within- and between-populations phenotypic differentiation (Q ST ) mainly results from covariance of QTL effects rather than QTL differentiation (F STq ), that within-population QTL differentiation is negligible, and that stronger divergent selection is required to produce adaptive differentiation within populations than among populations. With a high number of QTL, when the difference between environments within populations exceeds the smallest difference between environments across populations, high levels of within-population differentiation can be reached, reducing differentiation among populations. Our study stresses the need to account for within-population environmental heterogeneity when investigating local adaptation.

Combination of biocompatible hydrogel precursors to apatitic calcium phosphate cements (CPCs): Influence of the in situ hydrogel reticulation on the CPC properties.

In the field of bone regenerative medicine, injectable calcium phosphate cements (CPCs) are used for decades in clinics, as bone void fillers. Most often preformed polymers (e.g., hyaluronic acid, collagen, chitosan, cellulose ethers…) are introduced in the CPC formulation to make it injectable and improve its cohesion. Once the cement has hardened, the polymer is simply trapped in the CPC structure and no organic subnetwork is present. By contrast, in this work a CPC was combined with organic monomers that reticulated in situ so that a continuous biocompatible 3D polymeric subnetwork was formed in the CPC microstructure, resulting in a higher permeability of the CPC, which might allow to accelerate its in vivo degradation. Two options were investigated depending on whether the polymer was formed before the apatitic inorganic network or concomitantly. In the former case, conditions were found to reach a suitable rheology for easy injection of the composite. In addition, the in situ formed polymer was shown to strongly affect the size, density, and arrangement of the apatite crystals formed during the setting reaction, thereby offering an original route to modulate the microstructure and porosity of apatitic cements.

Partitioning heterozygosity in subdivided populations: Some misuses of Nei's decomposition and an alternative probabilistic approach.

Nei's decomposition of total expected heterozygosity in subdivided populations into within- and between-subpopulation components, HS and DST , respectively, is a classical tool in the conservation and management of genetic resources. Reviewing why this is not a decomposition into independent terms of within- and between-subpopulation gene diversity, we illustrate how this approach can be misleading because it overemphasizes the within-subpopulation component compared to Jost's nonadditive decomposition based on gene diversity indices. Using probabilistic partitioning of the total expected heterozygosity into independent within- and between-subpopulation contributions, we show that the contribution of the within-subpopulation expected heterozygosity to the total expected heterozygosity is not HS , as suggested by Nei's decomposition, but HS /s, with s being the number of subpopulations. Finally, we compare three possible approaches of decomposing total heterozygosity in subdivided populations (i.e., Nei's decomposition, Jost's approach, and probabilistic partitioning) with regard to independence between terms and sensitivity to unequal subpopulation sizes. For the conservation and management of genetic resources, we recommend using probabilistic partitioning and Jost's differentiation parameter rather than Nei's decomposition.

In vivo resorption of injectable apatitic calcium phosphate cements: Critical role of the intergranular microstructure.

The in vivo resorption rate of two injectable apatitic calcium phosphate cements used in clinics (Graftys® HBS and NORIAN®) was compared, using a good laboratory practice (GLP) study based on an animal model of critical-sized bone defect. To rationalize the markedly different biological properties observed for both cements, key physical features were investigated, including permeability and water-accessible porosity, total porosity measured by mercury intrusion and gravimetry, and microstructure. Due to a different concept for creating porosity between the two cements investigated in this study, a markedly different microstructural arrangement of apatite crystals was observed in the intergranular space, which was found to significantly influence both the mechanical strength and in vivo degradation of the two calcium phosphate cements.

Spatial and temporal patterns of a pulsed resource dynamically drive the distribution of specialist herbivores.

Patterns and drivers of the spatio-temporal distribution of herbivores are key elements of their ecological and evolutionary impacts on plant populations. Herbivore spatial distributions may be influenced by increased (RCH: resource concentration hypothesis) or decreased (RDH: resource dilution hypothesis) resource densities, but the effect of temporal variations in resource densities on such distributions remains poorly documented. We used a survey of a masting tree species and its seed predators in Southeastern France to address the effect of a host's pulsed resource on the spatio-temporal distributions of highly specialized insect herbivores feeding on seeds. Variations in both resource and seed predator densities were assessed by estimating seed production and seed infestation rates in focus trees during 10 consecutive years. We found increasing seed infestation rates with decreasing host tree densities in years of low seed production, indicating a RDH pattern of seed predators. However, such pattern was not persistent in years of high seed production during which seed infestation rates did not depend on host tree densities. We showed that temporal variations in resource density can lead to transience of seed predator spatial distribution. This study highlights how predictions of plant-herbivore interactions in natural ecosystems may rely on temporal components underlying RCH and RDH hypotheses.

A Triple Amino Acid Substitution at Position 88/94/95 in Glycoprotein GP2a of Type 1 Porcine Reproductive and Respiratory Syndrome Virus (PRRSV1) Is Responsible for Adaptation to MARC-145 Cells.

The Meat Animal Research Center-145 (MARC-145) cell line has been proven to be valuable for viral attenuation regarding vaccine development and production. Cell-adaptation is necessary for the efficient replication of porcine reproductive and respiratory syndrome virus (PRRSV) in these cells. Multiple sequence analysis revealed consistent amino acid substitutions in GP2a (V88F, M94I, F95L) of MARC-145 cell-adapted strains. To investigate the putative effect of these substitutions, mutations at either position 88, 94, 95, and their combinations were introduced into two PRRSV1 (13V091 and IVI-1173) infectious clones followed by the recovery of viable recombinants. When comparing the replication kinetics in MARC-145 cells, a strongly positive effect on the growth characteristics of the 13V091 strain (+2.1 log10) and the IVI-1173 strain (+1.7 log10) compared to wild-type (WT) virus was only observed upon triple amino acid substitution at positions 88 (V88F), 94 (M94I), and 95 (F95L) of GP2a, suggesting that the triple mutation is a determining factor in PRRSV1 adaptation to MARC-145 cells.

Porcine Alveolar Macrophage-like cells are pro-inflammatory Pulmonary Intravascular Macrophages that produce large titers of Porcine Reproductive and Respiratory Syndrome Virus.

Lung inflammation is frequently involved in respiratory conditions and it is strongly controlled by mononuclear phagocytes (MNP). We previously studied porcine lung MNP and described a new population of cells presenting all the features of alveolar macrophages (AM) except for their parenchymal location, that we named AM-like cells. Herein we showed that AM-like cells are macrophages phagocytosing blood-borne particles, in agreement with a pulmonary intravascular macrophages (PIM) identity. PIM have been described microscopically long time ago in species from the Laurasiatheria superorder such as bovine, swine, cats or cetaceans. We observed that PIM were more inflammatory than AM upon infection with the porcine reproductive and respiratory syndrome virus (PRRSV), a major swine pathogen. Moreover, whereas PRRSV was thought to mainly target AM, we observed that PIM were a major producer of virus. The PIM infection was more correlated with viremia in vivo than AM infection. Finally like AM, PIM-expressed genes were characteristic of an embryonic monocyte-derived macrophage population, whose turnover is independent of bone marrow-derived hematopoietic precursors. This last observation raised the interesting possibility that AM and PIM originate from the same lung precursor.

Towards Identification of the Substrates of ATP-Binding Cassette Transporters.

Most ATP-binding cassette (ABC) proteins function in transmembrane transport, and plant genomes encode a large number of ABC transporters compared with animal or fungal genomes. These transporters have been classified into eight subfamilies according to their topology and phylogenetic relationships. Transgenic plants and mutants with altered ABC transporter expression or function have contributed to deciphering the physiological roles of these proteins, such as in plant development, responses to biotic and abiotic stress, or detoxification activities within the cell. In agreement with the diversity of these functions, a large range of substrates (e.g. hormones and primary and secondary metabolites) have been identified. We review in detail transporters for which substrates have been unambiguously identified. However, some cases are far from clear, because some ABC transporters have the ability to transport several structurally unrelated substrates or because the identification of their substrates was performed indirectly without any flux measurement. Various heterologous or homologous expression systems have been used to better characterize the transport activity and other biochemical properties of ABC transporters, opening the way to addressing new issues such as the particular structural features of plant ABC transporters, the bidirectionality of transport, or the role of posttranslational modifications.

The Nicotiana tabacum ABC transporter NtPDR3 secretes O-methylated coumarins in response to iron deficiency.

Although iron is present in large amounts in the soil, its poor solubility means that plants have to use various strategies to facilitate its uptake. In this study, we show that expression of NtPDR3/NtABCG3, a Nicotiana tabacum plasma-membrane ABC transporter in the pleiotropic drug resistance (PDR) subfamily, is strongly induced in the root epidermis under iron deficiency conditions. Prevention of NtPDR3 expression resulted in N. tabacum plants that were less tolerant to iron-deficient conditions, displaying stronger chlorosis and slower growth than those of the wild-type when not supplied with iron. Metabolic profiling of roots and root exudates revealed that, upon iron deficiency, secretion of catechol-bearing O-methylated coumarins such as fraxetin, hydroxyfraxetin, and methoxyfraxetin to the rhizosphere was compromised in NtPDR3-silenced plants. However, exudation of flavins such as riboflavin was not markedly affected by NtPDR3-silencing. Expression of NtPDR3 in N. tabacum Bright Yellow-2 (BY-2) cells resulted in altered intra- and extracellular coumarin pools, supporting coumarin transport by this transporter. The results demonstrate that N. tabacum secretes both coumarins and flavins in response to iron deficiency and that NtPDR3 plays an essential role in the plant response to iron deficiency by mediating secretion of O-methylated coumarins to the rhizosphere.

Design and properties of a novel radiopaque injectable apatitic calcium phosphate cement, suitable for image-guided implantation.

An injectable purely apatitic calcium phosphate cement (CPC) was successfully combined to a water-soluble radiopaque agent (i.e., Xenetix® ), to result in an optimized composition that was found to be as satisfactory as poly(methyl methacrylate) (PMMA) formulations used for vertebroplasty, in terms of radiopacity, texture and injectability. For that purpose, the Xenetix dosage in the cement paste was optimized by injection of the radiopaque CPC in human cadaveric vertebrae under classical PMMA vertebroplasty conditions, performed by interventional radiologists familiar with this surgical procedure. When present in the cement paste up to 70 mg I mL-1 , Xenetix did not influence the injectability, cohesion, and setting time of the resulting composite. After hardening of the material, the same observation was made regarding the microstructure, mechanical strength and alpha-tricalcium phosphate to calcium deficient apatite transformation rate. Upon implantation in bone in a small animal model (rat), the biocompatibility of the Xenetix-containing CPC was evidenced. Moreover, an almost quantitative release of the contrast agent was found to occur rapidly, on the basis of in vitro static and dynamic quantitative studies simulating in vivo implantation. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2786-2795, 2018.

A straightforward approach to enhance the textural, mechanical and biological properties of injectable calcium phosphate apatitic cements (CPCs): CPC/blood composites, a comprehensive study.

Two commercial formulations of apatitic calcium phosphate cements (CPCs), Graftys® Quickset (QS) and Graftys® HBS (HBS), similar in composition but with different initial setting time (7 and 15min, respectively), were combined to ovine whole blood. Surprisingly, although a very cohesive paste was obtained after a few minutes, the setting time of the HBS/blood composite dramatically delayed when compared to its QS analogue and the two blood-free references. Using solid state NMR, scanning electron microscopy and high frequency impedance measurements, it was shown that, in the particular case of the HBS/blood composite, formation of a reticulated and porous organic network occurred in the intergranular space, prior to the precipitation of apatite crystals driven by the cement setting process. The resulting microstructure conferred unique biological properties to this material upon implantation in bone defects, since its degradation rate after 4 and 12weeks was more than twice that for the three other CPCs, with a significant replacement by newly formed bone. STATEMENT OF SIGNIFICANCE: A major challenge in the design of bone graft substitutes is the development of injectable, cohesive, resorbable and self-setting calcium phosphate cement (CPC) that enables rapid cell invasion with initial mechanical properties as close as bone ones. Thus, we describe specific conditions in CPC-blood composites where the formation of a 3D clot-like network can interact with the precipitated apatite crystals formed during the cement setting process. The resulting microstructure appears more ductile at short-term and more sensitive to biological degradation which finally promotes new bone formation. This important and original paper reports the design and in-depth chemical and physical characterization of this groundbreaking technology.

Emission of volatile organic compounds from petunia flowers is facilitated by an ABC transporter.

Plants synthesize a diversity of volatile molecules that are important for reproduction and defense, serve as practical products for humans, and influence atmospheric chemistry and climate. Despite progress in deciphering plant volatile biosynthesis, their release from the cell has been poorly understood. The default assumption has been that volatiles passively diffuse out of cells. By characterization of a Petunia hybrida adenosine triphosphate-binding cassette (ABC) transporter, PhABCG1, we demonstrate that passage of volatiles across the plasma membrane relies on active transport. PhABCG1 down-regulation by RNA interference results in decreased emission of volatiles, which accumulate to toxic levels in the plasma membrane. This study provides direct proof of a biologically mediated mechanism of volatile emission.