Potential effects of oilseed rape expressing oryzacystatin-1 (OC-1) and of purified insecticidal proteins on larvae of the solitary bee Osmia bicornis

Despite their importance as pollinators in crops and wild plants, solitary bees have not previously been included in non-target testing of insect-resistant transgenic crop plants. Larvae of many solitary bees feed almost exclusively on pollen and thus could be highly exposed to transgene products expressed in the pollen. The potential effects of pollen from oilseed rape expressing the cysteine protease inhibitor oryzacystatin-1 (OC-1) were investigated on larvae of the solitary bee Osmia bicornis (= O. rufa). Furthermore, recombinant OC-1 (rOC-1), the Bt toxin Cry1Ab and the snowdrop lectin Galanthus nivalis agglutinin (GNA) were evaluated for effects on the life history parameters of this important pollinator. Pollen provisions from transgenic OC-1 oilseed rape did not affect overall development. Similarly, high doses of rOC-1 and Cry1Ab as well as a low dose of GNA failed to cause any significant effects. However, a high dose of GNA (0.1%) in the larval diet resulted in significantly increased development time and reduced efficiency in conversion of pollen food into larval body weight. Our results suggest that OC-1 and Cry1Ab expressing transgenic crops would pose a negligible risk for O. bicornis larvae, whereas GNA expressing plants could cause detrimental effects, but only if bees were exposed to high levels of the protein. The described bioassay with bee brood is not only suitable for early tier non-target tests of transgenic plants, but also has broader applicability to other crop protection products

QTL analysis for resistance to foliar damage caused by Thrips tabaci and Frankliniella schultzei (Thysanoptera: Thripidae) feeding in cowpea [Vigna unguiculata (L.) Walp.]

Three quantitative trait loci (QTL) for resistance to Thrips tabaci and Frankliniella schultzei were identified using a cowpea recombinant inbred population of 127 F2:8 lines. An amplified fragment length polymorphism (AFLP) genetic linkage map and foliar feeding damage ratings were used to identify genomic regions contributing toward resistance to thrips damage. Based on Pearson correlation analysis, damage ratings were highly correlated (r ≥ 0.7463) across seven field experiments conducted in 2006, 2007, and 2008. Using the Kruskall–Wallis and Multiple-QTL model mapping packages of MapQTL 4.0 software, three QTL, Thr-1, Thr-2, and Thr-3, were identified on linkage groups 5 and 7 accounting for between 9.1 and 32.1% of the phenotypic variance. AFLP markers ACC-CAT7, ACG-CTC5, and AGG-CAT1 co-located with QTL peaks for Thr-1, Thr-2, and Thr-3, respectively. Results of this study will provide a resource for molecular marker development and the genetic characterization of foliar thrips resistance in cowpea

Transgenic avidin maize is resistant to storage insect pests

Avidin is a glycoprotein found in chicken egg white, that sequesters the vitamin biotin. Here we show that when present in maize at levels of ≥100 p.p.m., avidin is toxic to and prevents development of insects that damage grains during storage. Insect toxicity is caused by a biotin deficiency, as shown by prevention of toxicity with biotin supplementation. The avidin maize is not, however, toxic to mice when administered as the sole component of their diet for 21 days. These data suggest that avidin expression in food or feed grain crops can be used as a biopesticide against a spectrum of stored-product insect pests

A study on trypsin, Aspergillus flavus and Bacillus sp. protease inhibitory activity in Cassia tora (L.) syn Senna tora (L.) Roxb. seed extract

<p>Abstract</p> <p>Background</p> <p>Proteases play an important role in virulence of many human, plant and insect pathogens. The proteinaceous protease inhibitors of plant origin have been reported widely from many plant species. The inhibitors may potentially be used for multiple therapeutic applications in viral, bacterial, fungal diseases and physiological disorders. In traditional Indian medicine system, <it>Cassia tora </it>(<it>Senna tora</it>) is reportedly effective in treatment of skin and gastrointestinal disorders. The present study explores the protease inhibitory activity of the above plant seeds against trypsin, <it>Aspergillus flavus </it>and <it>Bacillus </it>sp. proteases.</p> <p>Methods</p> <p>The crushed seeds of <it>Cassia tora </it>were washed thoroughly with acetone and hexane for depigmentation and defatting. The proteins were fractionated by ammonium sulphate (0-30, 30-60, 60-90%) followed by dialysis and size exclusion chromatography (SEC). The inhibitory potential of crude seed extract and most active dialyzed fraction against trypsin and proteases was established by spot test using unprocessed x-ray film and casein digestion methods, respectively. Electrophoretic analysis of most active fraction (30-60%) and SEC elutes were carried employing Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and Gelatin SDS-PAGE. Inhibition of fungal spore germination was studied in the presence of dialyzed active inhibitor fraction. Standard deviation (SD) and ANOVA were employed as statistical tools.</p> <p>Results</p> <p>The crude seeds' extract displayed strong antitryptic, bacterial and fungal protease inhibitory activity on x-ray film. The seed protein fraction 30-60% was found most active for trypsin inhibition in caseinolytic assay (P < 0.001). The inhibition of caseinolytic activity of the proteases increased with increasing ratio of seed extract. The residual activity of trypsin, <it>Aspergillus flavus </it>and <it>Bacillus </it>sp. proteases remained only 4, 7 and 3.1%, respectively when proteases were incubated with 3 mg ml^-1seed protein extract for 60 min. The inhibitory activity was evident in gelatin SDS-PAGE where a major band (~17-19 kD) of protease inhibitor (PI) was detected in dialyzed and SEC elute. The conidial germination of <it>Aspergillus flavus </it>was moderately inhibited (30%) by the dialyzed seed extract.</p> <p>Conclusions</p> <p><it>Cassia tora </it>seed extract has strong protease inhibitory activity against trypsin, <it>Aspergillus flavus </it>and <it>Bacillus </it>sp. proteases. The inhibitor in <it>Cassia tora </it>may attenuate microbial proteases and also might be used as phytoprotecting agent.</p

Overexpression of the aphid-induced serine protease inhibitor <i>CI2c </i>gene in barley affects the generalist green peach aphid, not the specialist bird cherry-oat aphid

<div><p>Aphids are serious pests in crop plants. In an effort to identify plant genes controlling resistance against aphids, we have here studied a protease inhibitor, CI2c in barley (<i>Hordeum vulgare</i> L.). The <i>CI2c</i> gene was earlier shown to be upregulated by herbivory of the bird cherry-oat aphid <i>(Rhopalosiphum padi</i> L.<i>)</i> in barley genotypes with moderate resistance against this aphid, but not in susceptible lines. We hypothesized that CI2c contributes to the resistance. To test this idea, cDNA encoding <i>CI2c</i> was overexpressed in barley and bioassays were carried out with <i>R</i>. <i>padi</i>. For comparison, tests were carried out with the green peach aphid (<i>Myzus persicae</i> Sulzer), for which barley is a poor host. The performance of <i>R</i>. <i>padi</i> was not different on the <i>CI2c</i>-overexpressing lines in comparison to controls in test monitoring behavior and fecundity. <i>M</i>. <i>persicae</i> preference was affected as shown in the choice test, this species moved away from control plants, but remained on the <i>CI2c</i>-overexpressing lines. <i>R</i>. <i>padi</i>-induced responses related to defense were repressed in the overexpressing lines as compared to in control plants or the moderately resistant genotypes. A putative susceptibility gene, coding for a β-1,3-glucanase was more strongly induced by aphids in one of the <i>CI2c</i>-overexpressing lines. The results indicate that the CI2c inhibitor in overexpressing lines affects aphid-induced responses by suppressing defense. This is of little consequence to the specialist <i>R</i>.<i>padi</i>, but causes lower non-host resistance towards the generalist <i>M</i>. <i>persicae</i> in barley.</p></div

Expression of two barley proteinase inhibitors in tomato promotes endogenous defensive response and enhances resistance to Tuta absoluta

[EN] Background: For as long as 350 million years, plants and insects have coexisted and developed a set of relationships which affect both organisms at different levels. Plants have evolved various morphological and biochemical adaptations to cope with herbivores attacks. However, Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae) has become the major pest threatening tomato crops worldwide and without the appropriated management it can cause production losses between 80 to 100%. Results: The aim of this study was to investigate the in vivo effect of a serine proteinase inhibitor (BTI-CMe) and a cysteine proteinase inhibitor (Hv-CPI2) from barley on this insect and to examine the effect their expression has on tomato defensive response. We found that larvae fed on the double transgenic plants showed a notable reduction in weight. Moreover, only 56% of the larvae reached the adult stage. The emerged adults showed wings deformities and reduced fertility. We also investigated the effect of proteinase inhibitors ingestion on the insect digestive enzymes. Our results showed a decrease in larval trypsin activity. Transgenes expression had no harmful effect on Nesidiocoris tenuis (Reuter) (Heteroptera: Miridae), a predator of Tuta absoluta, despite transgenic tomato plants attracted the mirid. We also found that barley cystatin expression promoted plant defense by inducing the expression of the tomato endogenous wound inducible Proteinase inhibitor 2 (Pin2) gene, increasing the production of glandular trichomes and altering the emission of volatile organic compounds. Conclusion: Our results demonstrate the usefulness of the co-expression of different proteinase inhibitors for the enhancement of plant resistance to Tuta absoluta.This work was partly supported by grants BIO2013-40747-R and AGL2014-55616-C3 from the Spanish Ministry of Economy and Competitiveness (MINECO)Hamza, R.; Pérez-Hedo, M.; Urbaneja, A.; Rambla Nebot, JL.; Granell Richart, A.; Gaddour, K.; Beltran Porter, JP.... (2018). Expression of two barley proteinase inhibitors in tomato promotes endogenous defensive response and enhances resistance to Tuta absoluta. BMC Plant Biology. 18. https://doi.org/10.1186/s12870-018-1240-6S18Oerke EC. Crop losses to pests. J Agric Sci. 2005;144(01):31.Jouanin L, Bonadé-Bottino M, Girard C, Morrot G, Giband M. Transgenic plants for insect resistance. Plant Sci. 1998;131(1):1–11.Markwick NP, Docherty LC, Phung MM, Lester MT, Murray C, Yao JL, Mitra DS, Cohen D, Beuning LL, Kutty-Amma S, et al. 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Expression of Multiple Resistance Genes Enhances Tolerance to Environmental Stressors in Transgenic Poplar (Populus × euramericana ‘Guariento’)

Commercial and non-commercial plants face a variety of environmental stressors that often cannot be controlled. In this study, transgenic hybrid poplar (Populus × euramericana ‘Guariento’) harboring five effector genes (vgb, SacB, JERF36, BtCry3A and OC-I) were subjected to drought, salinity, waterlogging and insect stressors in greenhouse or laboratory conditions. Field trials were also conducted to investigate long-term effects of transgenic trees on insects and salt tolerance in the transformants. In greenhouse studies, two transgenic lines D5-20 and D5-21 showed improved growth, as evidenced by greater height and basal diameter increments and total biomass relative to the control plants after drought or salt stress treatments. The improved tolerance to drought and salt was primarily attributed to greater instantaneous water use efficiency (WUEi) in the transgenic trees. The chlorophyll concentrations tended to be higher in the transgenic lines under drought or saline conditions. Transformed trees in drought conditions accumulated more fructan and proline and had increased Fv/Fm ratios (maximum quantum yield of photosystem II) under waterlogging stress. Insect-feeding assays in the laboratory revealed a higher total mortality rate and lower exuviation index of leaf beetle [Plagiodera versicolora (Laicharting)] larvae fed with D5-21 leaves, suggesting enhanced insect resistance in the transgenic poplar. In field trials, the dominance of targeted insects on 2-year-old D5-21 transgenic trees was substantially lower than that of the controls, indicating enhanced resistance to Coleoptera. The average height and DBH (diameter at breast height) of 2.5-year-old transgenic trees growing in naturally saline soil were 3.80% and 4.12% greater than those of the control trees, but these increases were not significant. These results suggested that multiple stress-resistance properties in important crop tree species could be simultaneously improved, although additional research is needed to fully understand the relationships between the altered phenotypes and the function of each transgene in multigene transformants

Guidelines for the use of chest radiographs in community-acquired pneumonia in children and adolescents

© 2017, The Author(s). National guidance from the United Kingdom and the United States on community-acquired pneumonia in children states that chest radiographs are not recommended routinely in uncomplicated cases. The main reason in the ambulatory setting is that there is no evidence of a substantial impact on clinical outcomes. However clinical practice and adherence to guidance is multifactorial and includes the clinical context (developed vs. developing world), the confidence of the attending physician, the changing incidence of complications (according to the success of immunisation programs), the availability of alternative imaging (and its relationship to perceived risks of radiation) and the reliability of the interpretation of imaging. In practice, chest radiographs are performed frequently for suspected pneumonia in children. Time pressures facing clinicians at the front line, difficulties in distinguishing which children require admission, restricted bed numbers for admissions, imaging-resource limitations, perceptions regarding risk from procedures, novel imaging modalities and the probability of other causes for the child’s presentation all need to be factored into a guideline. Other drivers that often weigh in, depending on the setting, include cost-effectiveness and the fear of litigation. Not all guidelines designed for the developed world can therefore be applied to the developing world, and practice guidelines require regular review in the context of new information. In addition, radiologists must improve radiographic diagnosis of pneumonia, reach consensus on the interpretive terminology that clarifies their confidence regarding the presence of pneumonia and act to replace one imaging technique with another whenever there is proof of improved accuracy or reliability