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<p>Kukuruzni plamenca (<em>Ostrinia nubilalis</em>, Hbn.) je vrsta umerenog klimatskog područja&nbsp;<br />koja niske zimske&nbsp; temperature preživljava ulaskom u dijapauzu u stadijumu gusenice.&nbsp;Dijapauza i otpornost na hladnoću (cold hardiness,&nbsp; eng.), kod mnogih insekatskih vrsta često&nbsp;predstavljaju esencijalne, tesno povezane komponente strategije preživljavanja delovanja&nbsp;niske temperature, kao &scaron;to je to slučaj i kod kukuruznog plamenca&nbsp; <em>Ostrinia&nbsp; nubilalis</em>, Hbn. U&nbsp;okviru grupe insekata otpornih na hladnoću, svrstava se u insekte otporne na zamrzavanje&nbsp;(freeze tolerant,&nbsp; eng.) jer su&nbsp; sposobne da prežive ekstracelularno zamrzavanje telesnih&nbsp;tečnosti.</p><p>Iako se o osnovnim strukturnim i metaboličkim de&scaron;avanjima u dijapauzi insekata već dosta&nbsp;zna, jo&scaron; uvek postoje brojne dileme o ovom vidu preživljavanja insekata u nepovoljnim&nbsp;uslovima sredine, posebno sa aspekta izloženosti niskim zimskim temperaturama i&nbsp;mehanizmima krioprotekcije. Zbog toga je&nbsp; cilj ove doktorske disertacije&nbsp; bio&nbsp; da se sa aspekta&nbsp;biofizičkih osobina lipida i sastava masnih kiselina: &nbsp;odrede biofizičke osobine (temperature&nbsp;faznog prelaza) ukupnih lipida nedijapauzirajućih i dijapauzirajućih gusenica tokom dijapauze&nbsp;(početak, sredina i kraj),&nbsp; kao i u kontrolisanim uslovima hlađenja dijapauzirajućih gusenica;&nbsp;zatim da se&nbsp; odredi zastupljenost pojedinih klasa lipida u hemolimfi i masnom telu&nbsp;nedijapauzirajućih i dijapauzirajućih gusenica;&nbsp; da se&nbsp; utvrde promene sastava masnih kiselina&nbsp;strukturne (polarne) i rezervne (nepolarne)&nbsp; frakcije&nbsp; lipida celih gusenica&nbsp; tokom dijapauze i&nbsp;pod uticajem različitih temperatura u eksperimentalnim uslovima; kao i da se&nbsp; odredi tkivna&nbsp;specifičnost, hemolimfe i masnog tela, masnih kiselina ukupnih lipida&nbsp; i&nbsp; biofizičke&nbsp; osobine&nbsp;ukupnih lipida masnog tela nedijapauzirajućih i dijapauzirajućih gusenica tokom dijapauze,&nbsp;kao i u kontrolisanim uslovima hlađenja dijapauzirajućih gusenica.&nbsp;&nbsp;</p><p>U analizi lipida kukuruznog plamenca <em>Ostrinia nubilalis</em>, Hbn. kori&scaron;ćene su sledeće tehnike:&nbsp;tankoslojna hromatografija (TLC) sa skenirajućom denzitometrijom, za razdvajanje i analizu&nbsp;klasa lipida;&nbsp; zatim&nbsp; gasno-masena hromatografija (GC-MS), za analizu masnih kiselina kao i&nbsp;diferencijalna skenirajuća kalorimetrija (DSC&nbsp; &ndash;&nbsp; differential scanning calorimetry,&nbsp; eng.), za&nbsp;termalnu analizu ekstrahovanih lipida.</p><p>Kod&nbsp; gusenica <em>Ostrinia&nbsp; nubilalis</em>&nbsp;sastav&nbsp; masnih kiselina ukupnih lipida analiziranih tkiva i&nbsp;lipidnih frakcija značajno se razlikuje između&nbsp; ova dva fiziolo&scaron;ka stanja, stanja dijapauze i&nbsp;nedijapauze.&nbsp; Dijapauza indukuje značajno povećanje sadržaja mononezasićenih masnih&nbsp;kiselina, pre svega palmitoleinske (16:1n-7) i oleinske masne kiseline (18:1n-9) uz&nbsp;istovremeno smanjenje sadržaja zasićenih i&nbsp; polinezasićenih masnih kiselina, pre svega&nbsp;palmitinske (16:0) i linolne masne kiseline (18:2n-6).&nbsp; Temperature faznog prelaza ukupnih&nbsp;lipida celih gusenica i masnog tela&nbsp; značajno su niže kod dijapauzirajućih gusenica&nbsp; <em>O.&nbsp;nubilalis</em>,&nbsp; usled &nbsp;značajnog povećanja&nbsp; stepena nezasićenosti masnih kiselina u njihovom&nbsp;sastavu. Značajno niže temperature&nbsp; faznog prelaza lipida dijapauzirajućih gusenica&nbsp; <em>O.<br />nubilalis</em>&nbsp; rezultat su povećanja stepena nezasićenosti njihovih masnih kiselina. Možemo&nbsp;zaključiti da su&nbsp; promene u sastavu&nbsp; masnih kiselina lipida indukovane dijapauzom važna&nbsp;komponenta preživljavanja niskih temperatura kod&nbsp; <em>O. nubilalis</em>&nbsp; Hbn. jer se&nbsp; veliki deo lipida&nbsp;održava u tečnom stanju čak i na temperaturama daleko ispod 0&deg;C, &scaron;to je neophodno za&nbsp;održavanje njihove fluidnosti odnosno funkcionalnosti.</p> / <p>European corn borer (<em>Ostrinia nubilalis</em>,&nbsp; Hbn.)&nbsp; inhabiting temperate regions&nbsp; that&nbsp;survive low winter temperatures as a diapausing fifth instar larva.&nbsp; Diapausing larvae are cold&nbsp;hardy and freeze tolerant as they are able to survive the extracellular formation of ice crystals.&nbsp;Winter survival for many insect species living in seasonally cold environments includes a&nbsp;radical remodeling of metabolism. This implies entry into a developmentally arrested,&nbsp;hypometabolic state of winter diapause, which in <em>O. nubilalis</em>&nbsp; and&nbsp; many&nbsp; other&nbsp; insect species&nbsp;of these habitats is closely related to their cold hardiness.</p><p>Although the basic structural and metabolic events consider insect diapause is already known,&nbsp;there are still many doubts about the survival of insects in an unfavorable environment,&nbsp;particularly in terms of&nbsp; exposure to low winter temperatures &nbsp;and mechanisms of&nbsp;cryoprotection. Therefore, the aim of this thesis in terms of biophysical properties of lipids&nbsp;and fatty acid compositions,&nbsp; was to: i) to determine &nbsp;the biophysical properties (phase&nbsp;transition temperatures) of the total lipids of non-diapausing&nbsp; larvae&nbsp; and diapausing larvae&nbsp;during diapause (early diapause, mid diapause and termination of diapause), as well as under&nbsp;controlled cooling conditions during different phases of diapause; ii) to determine the fatty&nbsp;acid compositions of structural (polar) and storage (non-polar)&nbsp; total&nbsp; lipid fractions of non-diapausing&nbsp; larvae&nbsp; and diapausing larvae, as well as under the influence of different&nbsp;<br />temperatures during diapause in the controled experimental conditions; iii)&nbsp; to determine the&nbsp;lipid classes&nbsp; composition of total lipids extracted from the haemolymph and fat body of non-diapausing and diapausing larvae;&nbsp; iiii) to determine&nbsp; the haemolymph and fat body total lipid&nbsp;fatty acid compositions&nbsp; as well as the biophysical properties of the fat body total lipids of&nbsp;non-diapausing larvae&nbsp; and diapausing larvae during diapause as well as in terms of controlled&nbsp;cooling conditions. Thus, our research was focused on the non-diapausing (ND) as well as the&nbsp;diapausing larvae&nbsp; (D) that&nbsp; were exposed to low temperatures during different phases&nbsp; of&nbsp;diapause.</p><p>During analysis of European corn borer&nbsp; <em>O. nubilalis</em>&nbsp; lipids, the following techniques was&nbsp;used: thin&nbsp; layer&nbsp; chromatography (TLC) with scanning densitometry, for the separation and&nbsp;quantification&nbsp; of lipid classes; gas-mass chromatography (GC-MS ) for the&nbsp; analysis of the&nbsp;fatty acid composition&nbsp; of lipids and&nbsp; the differential scanning calorimetry (DSC), for the&nbsp;thermal analysis of the extracted lipids.</p><p>The fatty acid compositions&nbsp; and biophysical properties of lipids differed markedly between&nbsp;non-diapausing and diapausing larvae&nbsp; of&nbsp; <em>O. nubilalis</em>. Diapause was associated with a&nbsp;dramatic increase&nbsp; mainly the proportions of palmitoleic acid (16:1n-7) and oleic acid (18:1n-9), with concurrent reductions in palmitic acid (16:0) and linoleic acid (18:2n-6). The increase&nbsp;in&nbsp; the level of unsaturation of&nbsp; lipids,&nbsp; which caused a marked shift in their phase transitions to&nbsp;lower temperatures,&nbsp; was triggered&nbsp; by diapause rather than low temperatures.&nbsp; Adjustments of&nbsp;fatty acid compositions are likely to be an important component of winter diapause &nbsp;mechaisms, possibly maintaining the fluidity of cell membranes, and the functionality &nbsp;of the&nbsp;organism during lower winter temperatures.&nbsp; We conclude that&nbsp; <em>O. nubilalis &nbsp;</em>undergoes&nbsp;remodeling of fatty acid profiles of lipids as an element of its overwintering physiology which&nbsp;may improve the ability to harden during diapause.</p>

Mon sujet de thèse consistait à étudier les mécanismes de détection des phytoecdystéroïdes chez la Pyrale du maïs, Ostrinia nubilalis Hübner. Des études préalables, ont montré en effet que la Pyrale du maïs détectait ces produits, chez les larves, grâce à des organes gustatifs. En outre, la présence des phytoecdystéroïdes dans l'alimentation inhibait la prise de nourriture. Ces composés ont donc un double aspect, hormones car ils induisent la mue et antiappétents car ils sont détectés par des cellules gustatives et inhibent l'alimentation.<br />Nous avons précisé les modalités d'action de ces composés sur les cellules gustatives. Dans un premier temps, en précisant le spectre de sensibilité des récepteurs gustatifs. A l'aide de 20 composés différents, nous avons montré que les récepteurs gustatifs avaient une sensibilité (1) différente des récepteurs hormonaux et (2) qu'ils différaient selon le type des sensilles gustatives. Ces résultats suggèrent que d'autres molécules que l'hormone de mue (la 20-hydroxyecdysone ou 20E) peuvent avoir un effet antiappétent plus grand. <br />Dans un deuxième temps, nous avons cherché à mettre en relation la sélectivité de l'alimentation avec la sensibilité aux ecdystéroïdes en mesurant l'activité humorale induite dans les cellules gustatives chez des larves de différentes espèces d'Ostrinia qui ont un régime alimentaire différent (oligophage ou sélectif versus polyphage ou peu sélectif). Les résultats de cette étude a permis de montrer que la sélectivité du régime alimentaire n'a pas de relation directe avec la sensibilité aux ecdystéroïdes. <br />Dans un troisième temps, nous avons tenté d'évaluer si les ecdystéroïdes présentés de manière externe par rapport aux organes gustatifs avaient le même effet que lorsqu'ils étaient injectés dans l'animal. En effet, les animaux proches de la mue cessent de s'alimenter et nous voulions vérifier si le pic hormonal interne ne modifiait pas la sensibilité gustative ce qui aurait pu diminuer la motivation alimentaire. Les expériences menées se sont faites en trois étapes : (1) par une mesure de la sensibilité gustative au cours du stade larvaire sur des insectes alimentés en milieu artificiel ou des feuilles naturelles (2) par une exposition des sensilles à une solution de 20E ou solution saline de la larve (3) par une injection de 20E ou de solution saline dans la larve. Les résultats obtenus montrent que la sensibilité évolue au cours du temps et qu'il est impossible d'enregistrer l'activité électrique des neurones gustatifs en fin de stade, probablement pour des problèmes techniques. L'exposition des sensilles est un protocole traumatique qui donne des résultats intéressants, dans la mesure où l'activité des cellules gustatives semble inhibée. L'injection de 20E montre clairement que le taux d'ecdystéroïdes interne n'a pas d'incidence sur la sensibilité mais module par contre l'alimentation.<br />Enfin, dans une dernière partie, nous avons tenté d'élucider par quelle cellule sont perçus les ecdystéroïdes et si ce sont les mêmes cellules qui détectent toutes les substances antiappétentes. Les protocoles utilisés ont consisté à présenter des substances en mélange et à exposer les cellules à l'un des composés au préalable. Les résultats obtenus étaient en faveur de l'hypothèse qui prévoit que les phytoecdystéroïdes excitent le récepteur aux antiappétents. <br />Les phytoecdystéroïdes jouent un rôle de métabolites secondaires de défense, au moins à l'égard de la Pyrale du maïs.

In den letzten 20 Jahren hat sich der Maiszünsler (<i>Ostrinia nubilalis HÜBNER</i>), aus der Schmetterlingsfamilie der Pyralidae oder Zünsler, zum bedeutendsten tierischen Schädling des Maises (<i>Zea mays</i>) entwickelt. Eine Möglichkeit den Befall des Maiszünslers abzuwenden, bietet der Anbau von <i>Bacillus thuringiensis</i>-Mais (Bt-Mais). Mit Hilfe der Gentechnik wurden Gene des Bakteriums <i>Bacillus thuringiensis</i> übertragen, die einen für Fraßinsekten giftigen Wirkstoff bilden, wodurch die Pflanzen während der kompletten Vegetation vor den Larven des Maiszünslers geschützt sind.<br><br> Ziel des vorliegenden Projektes war es, in einer 3-jährigen Studie die Auswirkungen des großflächigen Anbaus von Bt-Mais auf die ökologische Situation und den Handlungsrahmen des integrierten Pflanzenschutzes komplex zu untersuchen. Dazu wurden in Betrieben im Oderbruch, das als permanentes Befallsgebiet des Maiszünslers gilt, in den Jahren 2002 bis 2004 jährlich zwei Felder mit jeweils einer Bt-Sorte und einer konventionellen Sorte angelegt. Zusätzlich wurden biologische und chemische Maiszünsler-Bekämpfungsvarianten geprüft.<br><br> Durch verschiedene Methoden wie Bonituren, Ganzpflanzenernten, Bodenfallenfänge und Beobachtungen des Wahlverhaltens von (Flug-)insekten konnten Aussagen zum Vorkommen von Insekten und Spinnentieren getroffen werden, wobei hierfür Daten aus Untersuchungen der Jahre 2000 und 2001 im Oderbruch ergänzend herangezogen werden konnten. Durch Ertragsmessungen, Energie- und Qualitätsermittlungen, sowie Fusarium- und Mykotoxinanalysen konnte der Anbau von Bt-Mais als neue Alternative zur Bekämpfung des Maiszünslers bewertet werden.<br><br> Bezüglich des Auftretens von Insekten und Spinnentieren wurden im Mittel der fünfjährigen Datenerhebung beim Vergleich der Bt-Sorte zur konventionellen Sorte, mit Ausnahme der fast 100 %igen Bekämpfung des Maiszünslers, keine signifikanten Unterschiede festgestellt. Hierfür wurde ein besonderes Augenmerk auf Thripse, Wanzen, Blattläuse und deren Fraßfeinde, sowie mittels Bodenfallenfängen auf Laufkäfer und Spinnen gerichtet.<br><br> Die erwarteten ökonomischen Vorteile wie etwa Ertragsplus oder bessere Nährstoff- und Energiegehalte durch geringeren Schaden beim Anbau von Bt-Mais als Silomais blieben in den Untersuchungsjahren aus. Allerdings zeigten Fusarium- und Mykotoxinanalysen eine geringere Belastung des Bt-Maises, was möglicherweise auf den geringeren Schaden zurückzuführen ist, da beschädigte Pflanzen für Fusarium und Mykotoxine anfälliger sind.<br><br> Desweiteren konnten erste methodische Ansätze für ein auf EU-Ebene gefordertes, den Anbau von Bt-Mais begleitendes Monitoring, erarbeitet werden. So konnten Vorschläge für geeignete Methoden, deren Umfang sowie des Zeitpunktes der Durchführungen gemacht werden. / In the last 20 years the European corn borer (<i>Ostrinia nubilalis</i>, <i>Pyralidae</i>) has become the most important pest in maize (<i>Zea mays</i>). One of a couple of possibilities to reduce the infestation by the European corn borer is the cultivation of <i>Bacillus thuringiensis</i> maize (Bt maize). Genetic engineering transmitted genes from <i>Bacillus thuringiensis</i>, which produce a substance that is toxic to feeding insects and thus protect plants against the larvae of the European corn borer during the whole vegetation.<br><br> The present project is a 3-year study to identify the effects of Bt maize growing on the ecological situation and the possibilities of integrated plant protection. From 2002 to 2004, two fields in the Oderbruch region, where Ostrinia nubilalis occurs, were each planted with Bt maize and a conventional maize variety every year. Furthermore, a biological and a chemical strategy against the European corn borer were verified.<br><br> Different methods like counts, harvest of whole plants, pitfall traps and observation the landing behaviour of flying insects were used to determine the abundance of insects and spiders. Furthermore, we could use additional data from studies obtained in the Oderbruch region in 2000 and 2001. The determination of yield, quality and energy content of the crops as well as of the degree of Fusarium infection and contamination by mycotoxins led to the conclusion that the cultivation of Bt maize is a new alternative strategy to control the European corn borer.<br><br> The average occurrence of insects and spiders did not differ significantly between Bt maize and the conventional variety in the 5 years of data recording. The only exception is the almost total control of the European corn borer. Attention was especially paid to thrips, bugs, aphids and their feeding enemies and using ground traps to ground beetles and spiders.<br><br> The expected economic benefits like increased yield or nutrient and energy content of the crop as a result of a minimized damage to silage Bt maize were not achieved in the years under investigation. However, the analysis of Fusarium and mycotoxins indicated a lower exposure of Bt maize, which may result from a lower damage caused by Ostrinia nubilalis, and damaged plants are more susceptible to Fusarium and mycotoxins.<br><br> Furthermore, we developed a first methodological approach for the monitoring procedure of Bt maize growing required by the EU. We have made proposals on appropriate methods, their extent as well as the optimum time of their application.

Separate field studies were started in fall 2005, which continued through fall 2007, to investigate the effect of different levels of European corn borer tunneling on yield in corn grown for grain and to predict spring infestation levels of early season soil insects, specifically white grubs (Coleoptera: Scarabaeidae) and wireworms (Coleoptera: Elateridae) in cornfields. In the first study, model variables included corn growth stage and larvae per plant. In both years of this study, larvae per plant had a significant effect on grain yield. Grain yield was reduced by 13.1 and 3.65% in plants infested with four larvae per plant in 2006 and 2007, respectively. For 2006, linear regression models provided average percent yield loss per larva per plant at 4.1, 6.8, and 1.8% during late vegetative (V12), early silking (R1), and blister (R2) growth stages, respectively. Economic injury levels (EILs) were calculated based on average percent yield reductions across each growth stage and year. In the second study, no significant differences were detected in both fall and spring between two sampling methods after correcting for differences in sampling volume. Strong correlations were observed between fall and spring grub densities in both years. In 2006, fields with grub densities above the spring nominal threshold had significantly greater stand and yield in the Poncho 1250 (1.25 mg clothianidin / kernel) treatment when compared to the Poncho 250 (0.25 mg clothianidin / kernel) and untreated plots. This information was used to develop fall EILs and economic thresholds for white grubs. / Master of Science in Life Sciences

Double-cropping of corn (Zea mays L.) for grain following the harvest of a small grain crop has been under evaluation in Virginia as an alternative cropping strategy (Brann and Pitman, 1997). To assess the potential constraints on late planted corn imposed by insects and diseases, double-cropped corn was evaluated in field experiments in Montgomery County, Virginia from 1998 to 2000. Factors included two near-isoline hybrids (NK4640 and NK4640Bt), insecticides at planting (tefluthrin in all years, 1998-2000; and imidacloprid in 1999 and 2000), and fungicide treatments (azoxystrobin or propiconazole). Response variables included yield, moisture at harvest, grain test weight, damage by European corn borer (Ostrinia nubilalis), damage by corn earworm (Heliothis zea), disease progress curves for gray leaf spot Cercospora zeae-maydis), and number of plants exhibiting virus symptoms. The Bt hybrid performed significantly better than the non-Bt hybrid for yield and test weight in double-cropped corn in 1998 and 2000, but not in 1999. A spatially referenced site suitability analysis was performed for full season and double-cropped corn in Virginia using weighted abiotic factors and constraints. Thornthwaite potential evapotranspiration (PET) and PET minus precipitation were used to identify areas of the state having a lower average moisture deficit during the silking months for double-cropped corn compared to full-season corn. It is concluded that double-cropped corn production is a viable option in Virginia where abiotic factors are not constraining, particularly growing season length and moisture availability during the sensitive stages of development. / Master of Science

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