A semiochemical view of the ecology of the seed beetle Acanthoscelides obtectus Say (Coleoptera: Chrysomelidae, Bruchinae).

The dried bean beetle, Acanthoscelides obtectus, is an economically important pest of stored legumes worldwide. Tracking the human-aided dispersion of its primary hosts, the Phaseolus vulgaris beans, it is now widespread in most bean-growing areas of the tropics and subtropics. In temperate regions where it can only occasionally overwinter in the field, A. obtectus proliferates in granaries, having multiple generations a year. Despite its negative impact on food production, no sensitive detection or monitoring tools exist, and the reduction of local populations still relies primarily on inorganic insecticides as fumigating agents. However, in the quest to produce more nutritious food more sustainably and healthily, the development of environmentally benign crop protection methods is vital against A. obtectus. For this, knowledge of the biology and chemistry of both the host plant and its herbivore will underpin the development of, among others, chemical ecology-based approaches to form an essential part of the toolkit of integrated bruchid management. We review the semiochemistry of the mate- and host-finding behaviour of A. obtectus and provide new information about the effect of seed chemistry on the sensory and behavioural ecology of host acceptance and larval development.

Progress on reducing acrylamide levels in potato crisps in Europe, 2002 to 2019.

European Snacks Association (ESA) data on acrylamide in potato crisps from 2002 to 2019 (99704 observations) were analysed. Acrylamide levels have plateaued since 2011, although the lowest mean so far was attained in 2018 at 353 ± 2.7 ng g-1: a 54% reduction since 2002. The 85th, 90th and 95th quantiles did show evidence of continued downward progress, the 90th quantile being lower than the 750 ng g-1 European Benchmark Level from 2017 to 2019. A smaller dataset from the European Food Safety Authority (2124 observations) for 2011-2018 was also analysed. The yearly means were higher than those of the ESA data but showed a fall in average acrylamide from 715 ± 40.5 ng g-1 in 2015 to 505 ± 28.5 ng g-1 in 2018, as well as steep falls in the 85th, 90th and 95th quantiles. Nevertheless, even the 85th quantile remained above the 750 ng g-1 Benchmark Level. The ESA data showed a reduction in the proportion of samples with acrylamide exceeding 750 ng g-1, from over 40% in 2002 to 7.75% in 2019. Seasonality was evident, with highest acrylamide levels from November to May. Crisp type had little effect except that thicker types had a higher proportion of samples containing >750 ng g-1 acrylamide. Analysis of the region of origin in Europe of the final product revealed improvements in the east and north. Geographical factors combined with seasonality continued to be problematic but was also an aspect in which progress was most evident. The findings show that improvements have been made in reducing the number of samples with very high levels of acrylamide, but do not suggest that mean acrylamide levels could be reduced substantially below where they have been since 2011, or that levels could be kept consistently below the current Benchmark Level.

Mapping sites of gibberellin biosynthesis in the Arabidopsis root tip.

Root elongation depends on the action of the gibberellin (GA) growth hormones, which promote cell production in the root meristem and cell expansion in the elongation zone. Sites of GA biosynthesis in the roots of 7-d-old Arabidopsis thaliana seedlings were investigated using tissue-specific GA inactivation in wild-type (Col-0) or rescue of GA-deficient dwarf mutants. Tissue-specific GA depletion was achieved by ectopic expression of the GA-inactivating enzyme AtGA2ox2, which is specific for C19 -GAs, and AtGA2ox7, which acts on C20 -GA precursors. In addition, tissue-specific rescue of ga20ox triple and ga3ox double mutants was shown. Furthermore, GUS reporter lines for major GA20ox, GA3ox and GA2ox genes were used to observe their expression domains in the root. The effects of expressing these constructs on the lengths of the root apical meristem and cortical cells in the elongation zone confirmed that roots are autonomous for GA biosynthesis, which occurs in multiple tissues, with the endodermis a major site of synthesis. The results are consistent with the early stages of GA biosynthesis within the root occurring in the meristematic region and indicate that the penultimate step of GA biosynthesis, GA 20-oxidation, is required in both the meristem and elongation zone.

Foliar N Application at Anthesis Stimulates Gene Expression of Grain Protein Fractions and Alters Protein Body Distribution in Winter Wheat (Triticum aestivum L.).

The major components of wheat storage proteins are gliadins and glutenins, and as they contribute differently to baking quality, a balanced mixture of these components is essential. The application of foliar nitrogen (N) at anthesis is a common practice to improve protein concentration and composition. The aim of this study was to investigate the effects of a foliar N application at anthesis on storage protein gene expression during grain development and on the distribution of protein concentration and protein body size within the grain. In this experiment, an additional N application at anthesis stimulated the expression of genes of the majority of storage proteins when the N supply was low. Furthermore, it led to higher protein concentrations in the subaleurone layers, while in the center of the lobes, the protein concentrations were decreased. These changes will affect the protein recovery in white flours, as proportionally more protein might be lost during milling processes.

Molecular quantification of Plasmodium parasite density from the blood retained in used RDTs.

Most malaria-endemic countries are heavily reliant upon rapid diagnostic tests (RDT) for malaria case identification and treatment. RDT previously used for malaria diagnosis can subsequently be used for molecular assays, including qualitative assessment of parasite species present or the carriage of resistance markers, because parasite DNA can be extracted from the blood inside the RDT which remains preserved on the internal components. However, the quantification of parasite density has not previously been possible from used RDT. In this study, blood samples were collected from school-age children in Western Kenya, in the form of both dried blood spots on Whatman filter paper, and the blood spot that is dropped into rapid diagnostic tests during use. Having first validated a robotic DNA extraction method, the parasite density was determined from both types of sample by duplex qPCR, and across a range of densities. The methods showed good agreement. The preservation of both parasite and human DNA on the nitrocellulose membrane inside the RDT was stable even after more than one year's storage. This presents a useful opportunity for researchers or clinicians wishing to gain greater information about the parasite populations that are being studied, without significant investment of resources.

Identification of a non-host semiochemical from tick-resistant donkeys (Equus asinus) against Amblyomma sculptum ticks.

Amblyomma sculptum is a tick affecting animal and human health across Argentina, Bolivia, Paraguay and Brazil. Donkeys, Equus asinus, are known to be resistant to A. sculptum, suggesting that they can produce non-host tick semiochemicals (allomones), as already demonstrated for some other vertebrate host/pest interactions, whereas horses, Equus caballus, are considered as susceptible hosts. In this study, we tested the hypothesis that donkeys produce natural repellents against A. sculptum, by collecting sebum from donkeys and horses, collecting the odour from sebum extracts, and identifying donkey-specific volatile compounds by gas chromatography (GC) and coupled GC-mass spectrometry (GC-MS). From the complex collected blends, five main compounds were identified in both species. Hexanal, heptanal and (E)-2-decenal were found predominantly in donkey extracts, whilst ethyl octanoate and ethyl decanoate were found predominantly in horse extracts. One compound, (E)-2-octenal, was detected exclusively in donkey extracts. In Y-tube olfactometer bioassays 36 different A. sculptum nymphs were tested for each extract, compound and concentration. The dry sebum extracts and the compounds identified in both species induced neither attraction nor repellency. Only (E)-2-octenal, the donkey-specific compound, displayed repellency, with more nymphs preferring the arm containing the solvent control when the compound was presented in the test arm across four concentrations tested (p < 0.05, Chi-square test). A combination of a tick attractant (ammonia) and (E)-2-octenal at 0.25 M also resulted in preference for the control arm (p < 0.05, Chi-square test). The use of semiochemicals (allomones) identified from less-preferred hosts in tick management has been successful for repelling brown dog ticks, Rhipicephalus sanguineus sensu lato from dog hosts. These results indicate that (E)-2-octenal could be used similarly to interfere in tick host location and be developed for use in reducing A. sculptum numbers on animal and human hosts.

Host shift induces changes in mate choice of the seed predator Acanthoscelides obtectus via altered chemical signalling.

The mechanisms of host shift in phytophagous insects are poorly understood. Among the many proposed processes involved, sexual selection via semiochemicals has recently been suggested. This hypothesizes that sexual communication using pheromones is modified as a result of development on a new host, and such plant-induced phenotypic divergence in mate recognition cues can lead to reproductive isolation between host lines. We tested this hypothesis on Acanthoscelides obtectus, an oligophagous bruchid of Phaseolus vulgaris beans worldwide, which also develops in acceptable non-hosts, such as chickpea (Cicer arietinum L.). Male sex pheromone blends of the bean, chickpea and chickpea/bean host lines during artificially induced host shifts showed different composition. Bean-reared females did not distinguish between blends, whereas chickpea and chickpea/bean females preferred the chickpea male pheromone. However, electrophysiological (EAG) responses to male odour of antennae of the three female host lines were similar, all preferring bean-reared males. Egg-laying choice tests revealed a uniform preference for bean seeds across female host lines, even after multiple generations, whereas larvae did not distinguish between bean and chickpea seeds. We conclude that the development of divergent chemical signalling systems during host shifts does not facilitate the evolution of host races in A. obtectus, because oviposition preferences remain unaffected.

Final grain weight is not limited by the activity of key starch-synthesising enzymes during grain filling in wheat.

Since starch is by far the major component of the mature wheat grain, it has been assumed that variation in the capacity for starch synthesis during grain filling can influence final grain weight. We investigated this assumption by studying a total of 54 wheat genotypes including elite varieties and landraces that were grown in two successive years in fields in the east of England. The weight, water content, sugars, starch, and maximum catalytic activities of two enzymes of starch biosynthesis, ADP-glucose pyrophosphorylase and soluble starch synthase, were measured during grain filling. The relationships between these variables and the weights and starch contents of mature grains were analysed. Final grain weight showed few or no significant correlations with enzyme activities, sugar levels, or starch content during grain filling, or with starch content at maturity. We conclude that neither sugar availability nor enzymatic capacity for starch synthesis during grain filling significantly influenced final grain weight in our field conditions. We suggest that final grain weight may be largely determined by developmental processes prior to grain filling. Starch accumulation then fills the grain to a physical limit set by developmental processes. This conclusion is in accord with those from previous studies in which source or sink strength has been artificially manipulated.

Dry matter losses and quality changes during short rotation coppice willow storage in chip or rod form.

This study compares dry matter losses and quality changes during the storage of SRC willow as chips and as rods. A wood chip stack consisting of approximately 74 tonnes of fresh biomass, or 31 tonnes dry matter (DM) was built after harvesting in the spring. Three weeks later, four smaller stacks of rods with an average weight of 0.8 tonnes, or 0.4 tonnes DM were built. During the course of the experiment temperature recorders placed in the stacks found that the wood chip pile reached 60 °C within 10 days of construction, but the piles of rods remained mostly at ambient temperatures. Dry matter losses were calculated by using pre-weighed independent samples within the stacks and by weighing the whole stack before and after storage. After 6 months the wood chip stack showed a DM loss of between 19.8 and 22.6%, and mean losses of 23.1% were measured from the 17 independent samples. In comparison, the rod stacks showed an average stack DM loss of between 0 and 9%, and between 1.4% and 10.6% loss from the independent samples. Analysis of the stored material suggests that storing willow in small piles of rods produces a higher quality fuel in terms of lower moisture and ash content; however, it has a higher fine content compared to storage in chip form. Therefore, according to the two storage methods tested here, there may be a compromise between maximising the net dry matter yield from SRC willow and the final fine content of the fuel.

Plasmodium-associated changes in human odor attract mosquitoes.

Malaria parasites (Plasmodium) can change the attractiveness of their vertebrate hosts to Anopheles vectors, leading to a greater number of vector-host contacts and increased transmission. Indeed, naturally Plasmodium-infected children have been shown to attract more mosquitoes than parasite-free children. Here, we demonstrate Plasmodium-induced increases in the attractiveness of skin odor in Kenyan children and reveal quantitative differences in the production of specific odor components in infected vs. parasite-free individuals. We found the aldehydes heptanal, octanal, and nonanal to be produced in greater amounts by infected individuals and detected by mosquito antennae. In behavioral experiments, we demonstrated that these, and other, Plasmodium-induced aldehydes enhanced the attractiveness of a synthetic odor blend mimicking "healthy" human odor. Heptanal alone increased the attractiveness of "parasite-free" natural human odor. Should the increased production of these aldehydes by Plasmodium-infected humans lead to increased mosquito biting in a natural setting, this would likely affect the transmission of malaria.

Temperature and nitrogen supply interact to determine protein distribution gradients in the wheat grain endosperm.

Gradients exist in the distribution of storage proteins in the wheat (Triticum aestivum) endosperm and determine the milling properties and protein recovery rate of the grain. A novel image analysis technique was developed to quantify both the gradients in protein concentration, and the size distribution of protein bodies within the endosperm of wheat plants grown under two different (20 or 28 °C) post-anthesis temperatures, and supplied with a nutrient solution with either high or low nitrogen content. Under all treatment combinations, protein concentration was greater in the endosperm cells closest to the aleurone layer and decreased towards the centre of the two lobes of the grain, i.e. a negative gradient. This was accompanied by a decrease in size of protein bodies from the outer to the inner endosperm layers in all but one of the treatments. Elevated post-anthesis temperature had the effect of increasing the magnitude of the negative gradients in both protein concentration and protein body size, whilst limiting nitrogen supply decreased the gradients.

Isolation and identification of floral attractants from a nectar plant for the dried bean beetle, Acanthoscelides obtectus (Coleoptera: Chrysomelidae, Bruchinae).

BACKGROUND: The response of virgin females of the legume pest Acanthoscelides obtectus (Coleoptera: Bruchidae) to headspace extracts of volatiles collected from flowers of a nectar plant, Daucus carota, was investigated using behaviour (four-arm olfactometry) and coupled gas chromatography-electroantennography (GC-EAG). RESULTS: Odours from inflorescences were significantly more attractive to virgin female beetles than clean air. Similarly, a sample of volatile organic compounds (VOCs) collected by air entrainment (dynamic headspace collection) was more attractive to beetles than a solvent control. In coupled GC-EAG experiments with beetle antennae and the VOC extract, six components showed EAG activity. Using coupled GC-mass spectrometry (GC-MS) and GC peak enhancement with authentic standards, the components were identified as α-pinene (S:R 16:1), sabinene, myrcene, limonene (S:R 1:3), terpinolene and (S)-bornyl acetate. Females preferred the synthetic blend of D. carota EAG-active volatiles to the solvent control in bioassays. When compared directly, odours of D. carota inflorescences elicited stronger positive behaviour than the synthetic blend. CONCLUSION: This is the first report of behaviourally active volatiles linked to pollen location for A. obtectus, and development of the six-component blend is being pursued, which could underpin the design of semiochemical-based field management approaches against this major pest of stored products. © 2018 Society of Chemical Industry.

Wheat receptor-kinase-like protein Stb6 controls gene-for-gene resistance to fungal pathogen Zymoseptoria tritici.

Deployment of fast-evolving disease-resistance genes is one of the most successful strategies used by plants to fend off pathogens1,2. In gene-for-gene relationships, most cloned disease-resistance genes encode intracellular nucleotide-binding leucine-rich-repeat proteins (NLRs) recognizing pathogen-secreted isolate-specific avirulence (Avr) effectors delivered to the host cytoplasm3,4. This process often triggers a localized hypersensitive response, which halts further disease development 5 . Here we report the map-based cloning of the wheat Stb6 gene and demonstrate that it encodes a conserved wall-associated receptor kinase (WAK)-like protein, which detects the presence of a matching apoplastic effector6-8 and confers pathogen resistance without a hypersensitive response 9 . This report demonstrates gene-for-gene disease resistance controlled by this class of proteins in plants. Moreover, Stb6 is, to our knowledge, the first cloned gene specifying resistance to Zymoseptoria tritici, an important foliar fungal pathogen affecting wheat and causing economically damaging septoria tritici blotch (STB) disease10-12.

Effects of variety, year of cultivation and sulphur supply on the accumulation of free asparagine in the grain of commercial wheat varieties.

Free asparagine concentration, which is the determining factor for acrylamide-forming potential in cereals, was measured in grain from wheat grown in field trials in the United Kingdom in 2011-2012 and 2012-2013. There were 25 varieties in 2012 and 59 in 2013, with eleven present in both trials. The trials were split-plot, with half of each plot supplied with sulphur and the other half not. The varietal means (mmol per kg) for free asparagine in the sulphur-fed wheat ranged from 1.521 to 2.687 in 2011-2012 and 0.708 to 11.29 in 2012-2013. Eight varieties were identified as having consistently low free asparagine concentration. There was a differential response of varieties to sulphur, and much higher levels of free asparagine in 2012-2013 versus 2011-2012. Given the short commercial lifespan of some wheat varieties, it is concluded that information on free asparagine concentration should be made available when a variety is launched.

The early inflorescence of Arabidopsis thaliana demonstrates positional effects in floral organ growth and meristem patterning.

KEY MESSAGE: Linear modelling approaches detected significant gradients in organ growth and patterning across early flowers of the Arabidopsis inflorescence and uncovered evidence of new roles for gibberellin in floral development. Most flowering plants, including the genetic model Arabidopsis thaliana, produce multiple flowers in sequence from a reproductive shoot apex to form a flower spike (inflorescence). The development of individual flowers on an Arabidopsis inflorescence has typically been considered as highly stereotypical and uniform, but this assumption is contradicted by the existence of mutants with phenotypes visible in early flowers only. This phenomenon is demonstrated by mutants partially impaired in the biosynthesis of the phytohormone gibberellin (GA), in which floral organ growth is retarded in the first flowers to be produced but has recovered spontaneously by the 10th flower. We presently lack systematic data from multiple flowers across the Arabidopsis inflorescence to explain such changes. Using mutants of the GA 20-OXIDASE (GA20ox) GA biosynthesis gene family to manipulate endogenous GA levels, we investigated the dynamics of changing floral organ growth across the early Arabidopsis inflorescence (flowers 1-10). Modelling of floral organ lengths identified a significant, GA-independent gradient of increasing stamen length relative to the pistil in the wild-type inflorescence that was separable from other, GA-dependent effects. It was also found that the first flowers exhibited unstable organ patterning in contrast to later flowers and that this instability was prolonged by exogenous GA treatment. These findings indicate that the development of individual flowers is influenced by hitherto unknown factors acting across the inflorescence and also suggest novel functions for GA in floral patterning.

Acrylamide levels in potato crisps in Europe from 2002 to 2016.

European manufacturers' data on acrylamide in potato crisps from 2002 to 2016 were analysed. A previous study showed a 53% reduction in mean acrylamide levels from 763 ng g-1 in 2002 to 358 ng g-1 in 2011. Analysis of data from the longer period showed that since 2011 there has been a levelling off, with the mean level for 2016 being 412 ng g-1 (still a 46% reduction from 2002), suggesting that the most effective acrylamide reduction measures had been devised and implemented by 2011. There were similar trends in the 90th and 95th quantile values, with the 90th quantile values being below 1000 ng g-1 (the European Commission's current 'Indicative Value' for acrylamide in potato crisps) since 2010. The proportion of samples with acrylamide above 2000 ng g-1 fell from 4.8% in 2002 to 0.6% in 2016. Acrylamide levels showed marked seasonal variability, being highest in the first half of the year when potatoes were being used from storage, and lowest from July to September when potatoes were being harvested. Acrylamide levels were higher in thicker types of crisp in the early years of the study, but this difference disappeared in the later years, suggesting that manufacturers had acted to reduce acrylamide formation in these products. Higher values for acrylamide were recorded in north and east Europe than in the south and west up to 2013. Levels in the north and east declined in recent years, but remained higher in the north than in the other regions. The manufacturers' data were compared with a much smaller dataset provided by the European Food Safety Authority (EFSA). Levels of acrylamide in the EFSA dataset were consistently higher than in the manufacturers' data, possibly due to uneven sampling through the year and the seasonality of acrylamide levels.

Odours of Plasmodium falciparum-infected participants influence mosquito-host interactions.

Malaria parasites are thought to influence mosquito attraction to human hosts, a phenomenon that may enhance parasite transmission. This is likely mediated by alterations in host odour because of its importance in mosquito host-searching behaviour. Here, we report that the human skin odour profile is affected by malaria infection. We compared the chemical composition and attractiveness to Anopheles coluzzii mosquitoes of skin odours from participants that were infected by Controlled Human Malaria Infection with Plasmodium falciparum. Skin odour composition differed between parasitologically negative and positive samples, with positive samples collected on average two days after parasites emerged from the liver into the blood, being associated with low densities of asexual parasites and the absence of gametocytes. We found a significant reduction in mosquito attraction to skin odour during infection for one experiment, but not in a second experiment, possibly due to differences in parasite strain. However, it does raise the possibility that infection can affect mosquito behaviour. Indeed, several volatile compounds were identified that can influence mosquito behaviour, including 2- and 3-methylbutanal, 3-hydroxy-2-butanone, and 6-methyl-5-hepten-2-one. To better understand the impact of our findings on Plasmodium transmission, controlled studies are needed in participants with gametocytes and higher parasite densities.

Effects of Cultivar and Nitrogen Nutrition on the Lipid Composition of Wheat Flour.

Despite being minor components of flour, wheat (Triticum aestivum L.) lipids contribute to grain processing. They are particularly important for bread making, where they adsorb to the surface of gas bubbles formed during the proving stage of bread making, stabilizing the gas cells and improving gas retention within the dough. This contributes to the volume and texture of the loaf. However, little is understood about how their amount, composition, and properties vary in response to genotype (G), environment (E) or G × E interactions. Six wheat lines were therefore grown at three levels of nitrogen supply at Rothamsted Research, and 48 lipid species across six lipid classes were identified and quantified in white flour using electrospray ionization-tandem triple quadrupole mass spectrometry (ESI-MS/MS). This showed clear differences in the contents and compositions of lipids between cultivar as well as effects of nitrogen fertilization, which would be expected to have impacts on the processing properties of the samples.

cis-Jasmone Elicits Aphid-Induced Stress Signalling in Potatoes.

Elicitation of plant defense signaling that results in altered emission of volatile organic compounds (VOCs) offers opportunities for protecting plants against arthropod pests. In this study, we treated potato, Solanum tuberosum L., with the plant defense elicitor cis-jasmone (CJ), which induces the emission of defense VOCs and thus affects the behavior of herbivores. Using chemical analysis, electrophysiological and behavioral assays with the potato-feeding aphid Macrosiphum euphorbiae, we showed that CJ treatment substantially increased the emission of defense VOCs from potatoes compared to no treatment. Coupled GC-electroantennogram (GC-EAG) recordings from the antennae of M. euphorbiae showed robust responses to 14 compounds present in induced VOCs, suggesting their behavioral role in potato/aphid interactions. Plants treated with CJ and then challenged with M. euphorbiae were most repellent to alate M. euphorbiae. Principal component analysis (PCA) of VOC collections suggested that (E)-2-hexenal, (E,E)-4,8,12-trimethyl-1,3,7,11-tridecatetraene (TMTT), (E)-ß-farnesene, (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), methyl salicylate (MeSA), CJ, and methyl benzoate (MeBA) were the main VOCs contributing to aphid behavioral responses, and that production of TMTT, (E)-ß-farnesene, CJ, and DMNT correlated most strongly with aphid repellency. Our findings confirm that CJ can enhance potato defense against aphids by inducing production of VOCs involved in aphid-induced signalling.