My dissertation and research focused on the effects of insecticides on squash bugs, its egg parasitoid, and pollinators in the production of cucurbits in Virginia. Plants in the cucumber family are dependent on insect pollination for successful fruit set, and are also susceptible to plant eating insects. Squash bugs are capable of transmitting cucurbit yellow vine decline, and their feeding can cause significant wilt and death in many varieties. To control for squash bug and other pests, growers commonly combine the application of broad-spectrum insecticides with the frequently applied prophylactic fungicides. Broad-spectrum insecticide applications are known to have negative effects on natural enemy populations, are capable of promoting insecticide resistance, and can have negative effects on pollinators if care in their use is not taken. Squash bugs have several natural enemies, but their predominant egg parasitoid is most effective at reducing damaging populations. The scelionid wasp Gryon pennsylvanicum Ashmead, is a prevalent egg parasitoid in Virginia and can be negatively affected by the application of broad-spectrum insecticides. Through survey efforts I found that G. pennsylvanicum is widely distributed throughout Virginia and is capable of high rates of egg parasitism (>90%). This is contrary to the 20% level previously assumed for the East Coast. I explored the effects of narrow-spectrum insecticides on the fate of the egg parasitoids, those developing in the host egg and emerged adults of G. pennsylvanicum. Contact assays showed that the insecticides λ-cyhalothrin and sulfoxaflor had caused high adult parasitoid mortality. As new insecticides get registered for use there is often concern about their effect on pollinators, specifically the European honey bee Apis mellifera L. I evaluated the use of large flight cages as a method to measure the sub-lethal effects of narrow-spectrum insecticides to honey bees, as a means to qualify risk. The method utilizes small colonies of honey bees (with stores of nectar and pollen) and their feeding at a treated sucrose solution after being trained to a feeder in an enclosed arena. This choice-test style behavioral experiment shows promise in qualifying the risks associated with insecticide exposure in the field. In the case of pyrifluquinazon, colonies repeatedly choose to avoid feeding at tainted feeders even after training with no other outside sources of food present. Further researching the sub-lethal behavioral effects that insecticides have on bees in a colony can help us better qualify their risk. / Ph. D.

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Yield differences resulting from magnesium, nitrogen, and micronutrients application were small. The incidence of crown blight was spotty and not related to fertilizer treatment.

Continuing studies on cucurbit viruses have reinforced previous conclusions that these diseases are the most important factors in cantaloup crown blight. Epidemiological studies of the several viruses concerned are concentrated on ascertaining the sources of virus for cantaloup infection and developing methods for the mathematical analysis of linear spread of virus. New methods of identification of the viruses are being developed through the use of serology. This involves biophysical studies of the viruses in the development of purification techniques. Purification viruses must be obtained before antisera can be developed.

Current Arizona cultural practices for cantaloup are not compatable with mechanized harvesting equipment now being developed for cantaloup. Mechanical harvesters will require a uniformly sloped bed, preferably flat, at harvest time. Experiments thus far indicate that this condition can be met by reshaping the conventional bed after the plants have emerged.

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Malgré l’importance agronomique des cucurbitacées au Venezuela et le fort impact des maladies virales sur la production, le pathosystème viral y a été peu étudié. Cinq virus ont été décrits par des travaux souvent anciens: le cucumovirus Cucumber mosaic virus (CMV), les potyvirus Papaya ringspot virus (PRSV), Zucchini yellow mosaic virus (ZYMV) et Watermelon mosaic virus (WMV), le comovirus Squash mosaic virus (SqMV), et un begomovirus partiellement caractérisé, le Melon chlorotic mosaic virus (MeCMV). Pour lutter contre ces agents pathogènes, il est nécessaire de bien connaître le pathosystème viral présent localement. Nous avons donc caractérisé les principaux virus provoquant des maladies sur ces cultures dans le pays, en vue de comprendre l’évolution du pathosystème et de développer des méthodes de lutte adaptées. Nos études épidémiologiques ont montré que le begomovirus MeCMV représente la principale menace sur melon et pastèque, les potyvirus ZYMV et PRSV étant la principale menace sur courge. Les isolats Vénézuéliens de ZYMV se sont révélés génétiquement homogènes et biologiquement très variables comme cela a été observé dans d'autres régions du monde. La résistance au ZYMV conférée par le gène Zym chez le melon PI 414723 est surmontée par certains isolats, alors que le concombre TGM représente une source stable de résistance au ZYMV. Les types W et P de PRSV sont présents au Venezuela, mais seul le PRSV-W été trouvé sur cucurbitacées cultivées et sauvages. Une autre souche virale, initialement appelée PRSV-T et détectée au Venezuela, constitue une espèce différente du PRSV d’après ses propriétés moléculaires et biologiques établies dans ce travail. / In Venezuela, cucurbits viruses are among de major constraints for cucurbit production. Five viruses have been described infecting cucurbits in the country: Cucumber mosaic virus (CMV, Cucumovirus), Papaya ringspot virus (PRSV, Potyvirus), Zucchini yellow mosaic virus (ZYMV, Potyvirus), Squash mosaic virus (SqMV, Comovirus), and Melon chlorotic mosaic virus (MeCMV, Begomovirus).The current frequency and impact of these viruses is Venezuela is not well known. In this work, the major cucurbit viruses were identified and characterized in order to estimate the viral pathosystem affecting cucurbit production in the country. The begomovirus MeCMV appears to be the major constraint for melon and watermelon production, while the potyviruses ZYMV and PRSV were the most important viruses infecting squash crops in this survey. Molecular characterisation of ZYMV isolates revealed a low genetic diversity of this virus in Venezuela. In contrast, ZYMV isolates were biologically variable as observed in several countries worldwide. Two types of PRSV, P and W, are present in Venezuela. PRSV-W is the only type naturally infecting cucurbits in Venezuela. Another type of PRSV, formerly referred as PRSV-T, was detected. Its molecular and biological characterisation revealed that it is indeed a new species related to but distinct from PRSV. Therefore, the name zucchini tigré mosaic virus (ZTMV) is proposed for this virus.

Memorial apresentado à Faculdade de Ciências Agrônomicas, UNESP, Câmpus de Botucatu, para fins de concurso público para obtenção do título de Livre-Docente na disciplina "Hortaliças de Frutos" junto ao Departamento de Produção Vegetal / Resumo: Seleção recorrente é um dos métodos mais eficientes para o melhoramento de espécies alógamas, especialmente quando progênies S1 são utilizadas. Considerando-se que abobrinha geralmente não apresenta perda de vigor pela endogamia, este método pode ser adequado para o melhoramento desta espécie. Neste trabalho foram realizados experimentos com o objetivo de avaliar a eficiência da seleção recorrente em abobrinha 'Piramoita'. Foram realizados três ciclos de seleção recorrente a partir da cultivar Piramoita (população P0), com avaliação e seleção de progênies S1. Novas populações foram obtidas com a recombinação de plantas das progênies selecionadas, utilizando-se sementes remanescentes. No primeiro ciclo foram avaliadas 74 progênies e selecionadas 14, no segundo foram avaliadas 60 e selecionadas 10 progênies e no terceiro ciclo foram avaliadas 77 e selecionadas 12 progênies. Foram obtidas populações melhoradas após um (PI), dois (PII) e três (PIII) ciclos de seleção recorrente. Quatro populações (P0, PI, PII e PIII) foram avaliadas em um delineamento em blocos ao acaso, com oito repetições e cinco plantas por parcela. Em todos os experimentos foram avaliadas as seguintes características: número de frutos total e comercial por planta, % de frutos comerciais, produção, em massa, de frutos total e comercial por planta e a massa média de fruto comercial. Foram observados aumentos de produção lineares significativos ao longo dos ciclos de seleção. Para número de frutos total e comercial e produção, em massa, total e comercial foram obtidos aumentos com a população PIII, comparativamente a população inicial, de 32, 63, 24 e 57%, respectivamente. A massa média de fruto comercial não foi afetada pela seleção recorrente. Conclui-se que a seleção recorrente foi eficiente para melhorar a abobrinha 'Piramoita'... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Recurrent selection is one of the most efficient methods to improve alogamous crops, especially when S1 progenies are utilized. Since squash usually does not have inbreeding depression, this method may be adequate for squash improvement. In this study, experiments were set up to evaluate recurrent selection efficiency in 'Piramoita' squash. Three cycles were made from cultivar Piramoita (population P0), with evaluation and selection of S1 progenies and recombination of plants from selected progenies, using remained seeds. In the first cycle, 74 progenies were evaluated and 14 were selected, in the second 60 progenies were evaluated and 10 were selected and in the third cycle 77 progenies were evaluated and 12 were selected. Improved populations were obtained after one (PI), two (PII) and three (PIII) cycles of recurrent selection. Four populations (P0, PI, PII and PIII) were evaluated in a randomized block design, with eight replicates and five plants per plot. In all experiments the following characteristics were evaluated: total and commercial number of fruits per plant, total and commercial fruit yield, percentage of commercial fruit and commercial average fruit weight. Linear regressions of yield, number and weight of fruits over selection cycles were obtained. Total and commercial number and weight of fruits per plant were improved from P0 to PIII by 32, 63, 24 and 57%, respectively. Commercial average fruit weight was not affected by selection cycles. It is concluded that recurrent selection was efficient to improve 'Piramoita' squash, regarding to total fruit yield and, mainly, to commercial fruit yield. S1 progenies obtained from improved population were also superior to progenies obtained from the original one.

This thesis aimed to investigate several fundamental and perplexing questions relating to the phloem loading and transport mechanisms of <i>Cucurbita maxima</i>, by combining metabolomic analysis with cell biological techniques. This putative symplastic loading species has long been used for experiments on phloem anatomy, phloem biochemistry, phloem transport physiology and phloem signalling. Symplastic loading species have been proposed to use a polymer trapping mechanism to accumulate RFO (raffinose family oligosaccharides) sugars to build up high osmotic pressure in minor veins which sustains a concentration gradient that drives mass flow. However, extensive evidence indicating a low sugar concentration in their phloem exudates is a long-known problem that conflicts with this hypothesis. Previous metabolomic analysis shows the concentration of many small molecules in phloem exudates is higher than that of leaf tissues, which indicates an active apoplastic loading step. Therefore, in the view of the phloem metabolome, a symplastic loading mechanism cannot explain how small molecules other than RFO sugars are loaded into phloem. <br><br> Most studies of phloem physiology using cucurbits have neglected the possible functions of vascular architecture in phloem transport. It is well known that there are two phloem systems in cucurbits with distinctly different anatomical features: central phloem and extrafascicular phloem. However, mistaken conclusions on sources of cucurbit phloem exudation from previous reports have hindered consideration of the idea that there may be important differences between these two phloem systems. <br><br> The major results are summarized as below:<br> 1) O-linked glycans in <i>C.maxima</i> were structurally identified as beta-1,3 linked glucose polymers, and the composition of glycans in cucurbits was found to be species-specific. Inter-species grafting experiments proved that these glycans are phloem mobile and transported uni-directionally from scion to stock.<br> 2) As indicated by stable isotopic labelling experiments, a considerable amount of carbon is incorporated into small metabolites in phloem exudates. However, the incorporation of carbon into RFO sugars is much faster than for other metabolites.<br> 3) Both CO2 labelling experiments and comparative metabolomic analysis of phloem exudates and leaf tissues indicated that metabolic processes other than RFO sugar metabolism play an important role in cucurbit phloem physiology.<br> 4) The underlying assumption that the central phloem of cucurbits continuously releases exudates after physical incision was proved wrong by rigorous experiments including direct observation by normal microscopy and combined multiple-microscopic methods. Errors in previous experimental confirmation of phloem exudation in cucurbits are critically discussed.<br> 5) Extrafascicular phloem was proved to be functional, as indicated by phloem-mobile carboxyfluorescein tracer studies. Commissural sieve tubes interconnect phloem bundles into a complete super-symplastic network.<br> 6) Extrafascicular phloem represents the main source of exudates following physical incision. The major transported metabolites by these extrafacicular phloem are non-sugar compounds including amino acids, O-glycans, amines.<br> 7) Central phloem contains almost exclusively RFO sugars, the estimated amount of which is up to 1 to 2 molar. The major RFO sugar present in central phloem is stachyose. <br> 8) Cucurbits utilize two structurally different phloem systems for transporting different group of metabolites (RFO sugars and non-RFO sugar compounds). This implies that cucurbits may use spatially separated loading mechanisms (apoplastic loading for extrafascicular phloem and symplastic loading for central phloem) for supply of nutrients to sinks. <br> 9) Along the transport systems, RFO sugars were mainly distributed within central phloem tissues. There were only small amounts of RFO sugars present in xylem tissues (millimolar range) and trace amounts of RFO sugars in cortex and pith. The composition of small molecules in external central phloem is very different from that in internal central phloem.<br> 10) Aggregated P-proteins were manually dissected from central phloem and analysed by both SDS-PAGE and mass spectrometry. Partial sequences of peptides were obtained by QTOF <i>de novo</i> sequencing from trypsin digests of three SDS-PAGE bands. None of these partial sequences shows significant homology to known cucurbit phloem proteins or other plant proteins. This proves that these central phloem proteins are a completely new group of proteins different from those in extrafascicular phloem. The extensively analysed P-proteins reported in literature to date are therefore now shown to arise from extrafascicular phloem and not central phloem, and therefore do not appear to be involved in the occlusion processes in central phloem. / Phloem transportiert ein ausgedehntes Spektrum an Molekülen zwischen Pflanzenorganen, um Wachstum und Entwicklung zu koordinieren. Folglich ist eine umfassende und unvoreingenommene Metabolom-Analyse notwendig, um unser Verständnis über den Transport von Stoffwechselprodukten sowie über Phloemtransport zu vertiefen. Phloemexsudate von Kürbispflanzen werden unter Verwendung der Metabolom-Analyse analysiert. Bei diesen Pflanzen wird angenommen, dass sie symplastische Beladungswege verwenden, um Photoassmilate als Ausgangsschritt des Phloemtransportes zu konzentrieren. Zwei neue Familien Callose-verwandter Substanzen, 1,3-Overknüpfte Glycane, sowie eine Reihe anderer kleinerer Metabolite werden in den Phloemexsudaten detektiert. Metabolom-Daten und physiologische Experimente widersprechen früher berichtetem Verständnis des Phloemexsudationsprozesses in Kürbispflanzen. Folglich bestätigt sich der Phloemexsudationsprozeß durch Kombination unterschiedlicher mikroskopischer Techniken. Kürbispflanzen besitzen zwei Phloemsysteme mit eindeutigen anatomischen Eigenschaften. Es zeigt sich, daß Phloemexsudate in Kürbissen hauptsächlich vom extrafaszikulären Phloem, nicht vom zentralen Phloem, stammen. In den letzten Jahrzehnten wurde gewöhnlich mißverstanden, daß Phloemexsudate vom zentralen Phloem stammen. Die eindeutigen metabolischen Profile der unterschiedlichen Phloemsysteme, die durch Metabolom-Analysen in der räumlichen Auflösung beobachtet werden, bestätigen die unterschiedlichen physiologischen Funktionen der zwei unterschiedlichen Phloemsysteme: das zentrale Phloem transportiert hauptsächlich Zucker, während das extrafaszikuläre Phloem ein ausgedehntes Spektrum von Metaboliten transportiert. Es kann auch ein unterschiedliches metabolisches Profil kleiner Moleküle zwischen internem und externem zentralem Phloem beobachtet werden. Von Strukturproteinen des zentralen Phloems wurden auch Proben genommen und mittels Massenspektrometrie analysiert. Diese Proteine erweisen sich als neuartige Proteine, die sich zu denen im extrafaszikulären Phloem unterscheiden. Dies bestätigt ferner den Funktionsunterschied der unterschiedlichen Phloemsysteme in Kürbispflanzen. Basierend auf diesen neuartigen Entdeckungen des Phloem-Metaboloms und dem vorhergehenden Wissen über den Phloemtransport in Kürbispflanzen, wird ein neues Modell vorgeschlagen, um den Mechanismus des Phloemtransports in der symplastischen Beladung zu verstehen.<br>