Master of Science / Department of Entomology / John C. Reese / The soybean aphid, Aphis glycines is documented to have arrived in North America in mid 2000 and has ever since established itself as a formidable pest of soybeans, with the capacity to cause immense crop losses. This formidable pest with its complex life cycle and habits represents a current threat to soybean production. Host plant resistance is a promising avenue that can offer considerable control over the soybean aphid problem. Antibiosis being the most effective host plant resistance category, this study was aimed at attempting to understand the effects of induction on the antibiosis levels of soybeans. In the first set of experiments, different soybean genotypes and two soybean aphid biotypes were tested to comprehend if and how the genotypes and biotypes affected the survival and reproduction of the aphid. The experiments revealed mixed results that can be attributed to the genotypes tested and the biotypes used. While some genotypes showed no significant changes due to previous infestation, K1621 suggested signs of induced resistance to biotype 1 and PI567301B showed induced resistance to biotype 2, while K1639 pointed towards induced susceptibility to biotype 2. A follow up feeding behavior study with Electrical Penetration Graph (EPG) technique was carried out on PI567301B to elucidate if the induced resistance was tissue-specific, which could affect the feeding behavior of the aphid (biotype 2); but the results showed no appreciable differences in the feeding behavior of the aphids on clean vs. infested plants. Induced response studies shed light on how plants respond to herbivory and help us identify how changes in plant physiology affect the various herbivores that visit it for food and shelter. This knowledge can thus be applied to the development of superior varieties of crops that can defend themselves better against recurring infestations.

Doctor of Philosophy / Department of Entomology / R. Jeff Whitworth / The Hessian fly, Mayetiola destructor (Say), has been a historically significant pest of wheat in Kansas. However, it has been 60+ years since research has been conducted examining the flies’ activity throughout the year. Results of pheromone trapping in 4 counties in Kansas shows that Hessian fly (HF) males are actively flying in the fall, at least 1 month after the historical fly-free dates. Therefore, the Hessian Fly-Free Date is no longer valid and should be referred to as the Best Pest Management Date. Using pheromones for fall and spring trapping also indicated that HF is more active throughout the spring than previously thought, with almost continuous fly emergence and numerous emergence peaks in both spring and fall. The use of resistant wheat cultivars has been adapted to protect seedling plants from HF larval feeding in the fall. However, it is unknown if these cultivars are still providing protection after winter vernalization. Greenhouse trials indicated that ‘Armour’, a cultivar considered intermediately resistant, remains resistant under infestation levels of 1 fly/tiller but significant seed weight losses occured under infestations of 3 flies/tiller. In the field, Armour did not provide protection post-vernalization, with plants containing similar numbers of flaxseeds (pupae) as the susceptible cultivar, ‘Fuller’, and having significant losses of culm height (cm), number of spikelets/spike, number of seeds/spike, and seed weight (grams) when infested. ‘Duster’, a cultivar considered highly resistant, appeared to provide resistance to HF larval feeding in both the greenhouse and the field, and even produced significantly heavier seeds when infested with 3 flies/tiller in the greenhouse. These results suggest that post-vernalization screening should be conducted on all HF resistant cultivars to determine if each continues to provide protection. Little information is available showing if and how HF larval feeding on more mature wheat (Feekes 7-10), post-vernalization, impact plants, aside from lodging. Greenhouse and field infestations of a susceptible cultivar, Fuller, showed that significant losses of culm height (cm), number of seeds/spikelet, and seed weight will result from as few as 1 larva /culm. Yield losses averaged 0.13g/spike (65 kg/ha) compared to non-infested plants.

Master of Science / Department of Entomology / James R. Nechols / Temperature has an important indirect impact on pest populations. Direct effects occur, but also may result from temperature-induced changes in plant quality, including the expression of host plant resistance traits. Therefore, I examined both indirect and direct effects of temperature on biotype 1 soybean aphids (SBA), Aphis glycines, on a Rag1-resistant soybean variety and compared the effects with a susceptible variety to gain a better understanding of how temperature impacts SBA. Four aphid responses were evaluated: preimaginal development, survival to adulthood, number of progeny produced, and adult longevity. In the first experiment, I grew soybean seedlings to the V-0 stage at 25°C and then conditioned them for 0, 3 or 5 days at 20° or 30°C before infesting with a single first instar SBA at each of the two experimental temperatures. Based on previous literature for SBA, I hypothesized that conditioning plants at the lower temperature would cause resistance to break down and that longer exposure would exacerbate the effect. Results showed that conditioning soybeans to 20°C significantly reduced SBA survival, and the effect on survival increased with longer conditioning. Conditioning plants to 30°C had no significant effect on SBA survival. However, estimated population growth decreased as conditioning time increased at 30°C and this effect was also observed at 20°C. Thus, plant resistance may have increased at both temperatures. The second experiment compared SBA responses, including population growth, at four temperatures (15, 20, 25, and 30⁰C) on a Rag1-resistant and susceptible soybean variety. I predicted that SBA fitness would be lower at all temperatures on resistant soybeans, but the magnitude of differences between cultivars would not be uniform across temperatures. Results indicated that both temperature (highest and lowest) and plant resistance detrimentally affected SBA fitness. There was also a significant interaction between the two variables with respect to SBA survival. Survival was lower and development rates were slower on the resistant cultivar. SBA required more degree-days to develop on resistant soybeans compared to the susceptible cultivar. This information will aid soybean producers in implementing a cost-efficient IPM strategy involving Rag1 resistant soybeans to combat SBA under a range of temperatures.

Doctor of Philosophy / Department of Entomology / Ming-Shun Chen / C. Michael Smith / The Hessian fly, Mayetiola destructor (Say), is a major pest of wheat, and is controlled mainly through deploying fly-resistant wheat cultivars. This study investigated five M. destructor populations collected from Texas, Louisiana, and Oklahoma, where infestation by Hessian fly has been high in recent years. Eight resistance genes including H12, H13, H17, H18, H22, H25, H26, and Hdic, were found to be highly effective against all tested M. destructor populations in this region, conferring resistance to 80% or more of plants containing one of these resistant genes. The frequency of biotypes virulent to resistant genes ranged from 0 to 45%. A logistic regression model was established to predict biotype frequencies based on the correlation between the percentages of susceptible plants obtained in a virulence test. In addition to the virulence test, the log-odds of virulent biotype frequencies were determined by a traditional approach to predict the logistic regression model. Characterization of a bacterial artificial chromosome (BAC) clone identified a gene encoding a protein with sequence similarity to bacterial levanases. Blast searching with the levanase-like protein identified 14 levanase/inulase-like genes or gene fragments. In this study, we determined the expression levels of these genes in different developmental stages and different tissues of 3-d old larvae of M. destructor. Sequence analysis revealed that six genes encode full length proteins, three were truncated at the 5’ end, and five truncated at the 3’ end. Sequences of putative proteins showed approximately 42% similarities to bacterial levanases or inulases, and 36% similarity to fungal levanases or inulases. No sequence similarities were found with any known animal or plant proteins. Comparative analysis of sequences among 14 levanase/inulase-like genes revealed that positions for intron/exon boundaries are conserved among different genes even though the length of each intron and exon varied among different genes. The expression patterns of the levanase/inulase-like genes were different among developmental stages and larval tissues of M. destructor. Interestingly, three genes presented alternative splicing bands in different developmental stages, and two genes exhibited splicing bands in different tissues of 3 d old M. destructor. This study would be useful for future studies of the characterization and function of levanase/inulase-like genes of these enzymes in plant-insect interactions.

O presente trabalho teve como objetivo avaliar o efeito do silício aplicado em diferentes cultivares de cana-de-açúcar sobre alguns aspectos biológicos e comportamentais da cigarrinha (Mahanarva fimbriolata) e sobre a produtividade e a qualidade da cana-de-açúcar, em condições de laboratório e campo. Em laboratório, as cultivares SP79-1011 e SP80-1816 (resistentes à cigarrinha) e SP81-3250 (suscetível) foram cultivadas em solo arenoso ou argiloso sem e com adubação silicatada (800 kg·ha-1 de silicato de potássio). Em outro experimento, as cultivares SP81-3250 e SP79-1011, mantidas em vasos com e sem adição de silício, foram expostas a adultos da cigarrinha e, após diferentes períodos (0, 12, 24 e 36h), foi determinada a concentração de compostos fenólicos solúveis totais (CFST) nas plantas. Nesses dois experimentos, os insetos foram provenientes de criação mantida em laboratório. O experimento de campo foi realizado na Usina Guaíra, em Guaíra, SP e os tratamentos foram constituídos de 16 cultivares com e sem aplicação de silício (800 kg·ha-1 de silicato de cálcio e magnésio). A avaliação da infestação de cigarrinhas (ninfas e adultos) foi realizada quinzenalmente. Na época da colheita da cana, plantas foram coletadas para a análise dos parâmetros tecnológicos. Em laboratório, o silício absorvido e acumulado na planta causou um acréscimo na mortalidade de ninfas e, dependendo da cultivar, este elemento também proporcionou aumento na duração da fase ninfal e decréscimo na longevidade de machos e fêmeas. A cultivar SP79-1011 foi a que teve o melhor desempenho na maioria dos parâmetros analisados, tendo apresentado maior teor de silício foliar e causado maior mortalidade de ninfas e menor longevidade de fêmeas. O período de pré-oviposição, a fecundidade e a viabilidade dos ovos não foram afetados pelo silício nas plantas ou pela cultivar utilizada. O valor de CFST não diferiu entre plantas tratadas e não tratadas com silício. Após as primeiras horas de exposição à cigarrinha, a cultivar resistente não só produziu maior concentração de compostos fenólicos, mas, quando tratada com silício, respondeu mais rapidamente ao ataque. A adubação silicatada, no campo, diminuiu o número de ninfas, não interferiu na população de adultos e aumentou os valores de Brix% e Pol do caldo, assim como o teor de Fibra% cana. Por outro lado, houve variação na produtividade em função das cultivares, constatando-se os maiores valores nas cultivares SP84-1431 e SP80-3280. / The present work aimed to study the effect of silicon applied in different cultivars of sugarcanes in biological and behavioral aspects of the spittlebug (Mahanarva fimbriolata) and in the yield and quality of the cane. Studies were conducted in the laboratory and on the field. In laboratory the cultivars SP79-1011 and SP80-1816 (resistant to spittlebug) and SP81-3250 (susceptible) were grown in sandy or clay soil with and without silicon fertilization (800 kg·ha-1 of potassium silicate). In another experiment, the cultivars SP81-3250 and SP79-1011 were kept in pots with and without silicon addition, and were exposed to spittlebug adults throughout different periods (0, 12, 24 and 36h), and the concentration of soluble phenolic compounds in plants were quantified. In both experiments the insects were obtained from a laboratory rearing. The field experiment was conducted at Guaíra Sugarcane Mill, in Guaíra, São Paulo. Treatments consisted of 16 cultivars with and without silicon application (800 kg·ha-1 of calcium and magnesium silicate). The evaluation of spittlebug infestation (nymphs and adults) was performed fortnightly. At the time of sugarcane harvest, plants were collected to carry out the analysis of technological parameters. In the laboratory experiment, the silicon absorbed and accumulated in the plant caused an increase in nymphs mortality and, depending on variety, this element also provided an increase in the duration of the nymphal stage and decreased longevity of males and females. The SP79-1011 cultivar showed the best performance in most parameters, presenting higher silicon content in leaves and causing increased mortality of nymphs and shorter female longevity. The pre-oviposition period, fecundity and egg viability were not affected by the silicon in plants or the cultivar used. The value of soluble phenolic compounds did not differ between plants treated and not treated with silicon. After the first hours of exposure to spittlebug, the resistant cultivar not only produced a higher concentration of phenolic compounds, but when treated with silicon responded more quickly to the attack. Silicon fertilization in the field reduced the number of nymphs but did not affect the adult population and increased values of Brix% and Pol% from cane juice, and the sugarcane fiber content. Moreover, there was change in yield among cultivars with the highest values for SP84-1431 and SP80-3280.

Este estudo foi realizado com o objetivo de identificar genótipos de feijão Phaseolus vulgaris resistentes aos carunchos Acanthoscelides obtectus e Zabrotes subfasciatus, bem como avaliar o efeito associado desses genótipos resistentes com inseticidas de origem vegetal. Para tanto, foram utilizados acessos de P. vulgaris do Banco de Germoplasma do Instituto Agronômico de Campinas e inseticidas comerciais de origem botânica. No screening inicial, amostras dos genótipos foram infestadas com cada uma das espécies de bruquídeos separadamente, avaliando-se o número de insetos emergidos aos 50 dias após a infestação. Dos 49 genótipos testados contra A. obtecus, não houve emergência naqueles com números de acesso 525, 584 e 615, podendo ser considerados os mais resistentes. Em relação a Z. subfasciatus, os genótipos com números de acesso 2, 35, 251, 570, 583, 584, 610, 621, 634, 816, 818 e 819 se mostraram mais resistentes entre os 185 avaliados. Destes, os genótipos portadores de arcelina 583, 584, 816, 818 e 819, além de 570 e 610, foram selecionados como os mais promissores para os testes subseqüentes, juntamente com a variedade Bolinha, que foi utilizada como controle de suscetibilidade. Não foi observada correlação entre as características morfoagronômicas dos genótipos de P. vulgaris e a sua resistência às espécies de bruquídeos avaliadas, indicando que a resistência a estas pragas não está associada às características da flor, vagem, semente e fenologia dos genótipos. A massa de mil sementes, que é indicativa da origem dos genótipos, foi um dos descritores analisados, mostrando também que a resistência de P. vulgaris a A. obtectus e a Z. subfasciatus não está relacionada à origem dos genótipos. Em testes de livre escolha e de confinamento, avaliou-se o efeito dos genótipos selecionados no screening, juntamente com a variedade Bolinha, sobre o comportamento e biologia de Z. subfasciatus. Verificou-se que a avaliação da preferência de Z. subfasciatus por genótipos de P. vulgaris em teste de livre escolha pode ser feita com 1 dia após a infestação e que \'Bolinha\', apesar de ser suscetível a Z. subfasciatus e favorecer o seu desenvolvimento, apresenta antixenose para oviposição em relação à praga. Nos testes realizados, os genótipos contendo arcelina tenderam a ser mais resistentes que os demais sem essa proteína, sendo que os seus efeitos sobre Z. subfasciatus incluíram o aumento da mortalidade no período de desenvolvimento, alongamento desse período e redução do peso de adultos emergidos, mantendo-se, de certa forma, estáveis ao longo de duas gerações da praga. A resistência conferida pela arcelina revelou ser do tipo antibiose, tendo como causas a impropriedade nutricional e a ação no metabolismo do inseto. Com relação aos inseticidas botânicos, foram testados 3 produtos comerciais, sendo 2 à base de azadiractina e um à base de rotenona. Entre estes, o produto que mais afetou o desenvolvimento de Z. subfasciatus foi NeemPro®, derivado de nim (Azadirachta indica), o qual apresentou efeito ovicida e prolongou a duração do período de desenvolvimento de Z. subfasciatus. Frente a isto, avaliou-se o efeito associado de NeemPro® com o genótipo resistente portador de arcelina 818 sobre alguns parâmetros biológicos de Z. subfasciatus. Verificou-se que os efeitos mais severos sobre Z. subfasciatus foram causados pelo genótipo resistente, independentemente do inseticida à base de nim e que o uso associado de ambos não provoca efeito aditivo ou sinérgico, não sendo recomendado para o manejo de Z. subfasciatus. / This research was carried out to identify Phaseolus vulgaris genotypes resistant to the bean weevils Acanthoscelides obtectus and Zabrotes subfasciatus, as well as to evaluate the effect of these genotypes in association with botanical insecticides. To reach this objective, P. vulgaris accessions from the Germplasm Bank of Instituto Agronômico de Campinas and commercial insecticides from botanical origin were tested. In the initial screening, samples of bean genotypes were infested with the weevil species separately and the number of adults emerged at the 50th day after infestation was evaluated. There was no A. obtectus emergence in genotypes 525, 584 and 615, among the 49 ones screened against this pest. In relation to Z. subfasciatus, genotypes with accession numbers 2, 35, 251, 570, 583, 584, 610, 621, 634, 816, 818 and 819 showed themselves resistant among 185 screened ones. The arcelin-containing genotypes 583, 584, 816, 818 and 819, plus 570 and 610 (both lacking this protein), were selected as the most promising for additional evaluations. Bolinha variety was also used as the susceptible standard. No correlation between morpho-agronomical characteristics of the P. vulgaris genotypes and their resistance to the weevils was observed, indicating that resistance to these two pests is not associated to genotypes flower, pod and seed characters or plant phenology. The mass of 1000 seeds, which indicates the origin of genotypes, was one of the used descriptors, showing that P. vulgaris resistance to A. obtectus and Z. subfasciatus is not related to genotypes origin too. In free- and no-choice tests, it was evaluated the effect of the screened genotypes on Z. subfasciatus behavior and biology, compared to \'Bolinha\'. It was verified that in free-choice tests, the evaluation of Z. subfasciatus preference for P. vulgaris genotypes can be done 1 day after infestation. Despite being susceptible to Z. subfasciatus and supporting its development, \'Bolinha\' holds antixenosis for oviposition in relation to the pest. In the bioassays carried out, genotypes containing arcelin tended to be more resistant than those lacking this protein and their effects on Z. subfasciatus include increasing of the mortality in the developmental period, enlargement of this period and reduction in adult weight, also being stable during two pest generations. The resistance provided by arcelin revealed itself to be antibiosis, by acting as antinutrients and also as antimetabolics. In relation to botanical insecticides, 3 commercial products, 2 of them based on azadirachtin and 1 based on rotenone, were evaluated. The insecticide NeemPro®, extracted from neem (Azadirachta indica) was the only one significantly causing ovicidal effect and enlarging Z. subfasciatus developmental period. Based on these results, the associated effect of NeemPro® and the resistant arcelin-containing P. vulgaris genotype 818 on some Z. subfasciatus biological parameters was evaluated. It was verified that the most severe effects on Z. subfasciatus were caused by the resistant bean genotype, independently of the neem based insecticide. The associated use of these two control methods no results in additive or synergistc effect and is not recommended for the management of Z. subfasciatus.

Visando avaliar os efeitos do tempo e da temperatura de armazenamento de grãos de feijoeiro, Phaseolus vulgaris L., sobre a manifestação da resistência ao caruncho Acanthoscelides obtectus (Say, 1831), realizaram-se ensaios com e sem chance de escolha sob condições de laboratório. Em casa-de-vegetação avaliou-se a preferência para oviposição e os danos causados pelo caruncho em vagens e grãos. Os genótipos utilizados nos ensaios foram Arc.1S, Arc.3S, Arc.5S, Carioca Pitoco, Ipa 6, Porrillo 70, Onix, Arc.1, Arc.2, Arc.3 e Arc.4. Em teste com chance de escolha utilizando-se grãos de diferentes idades, observou-se que os genótipos selvagens Arc.1S e Arc.5S expressaram antibiose e/ou não-preferência para alimentação como mecanismos de resistência a A. obtectus; os genótipos Arc.1 e Arc.2 apresentaram somente antibiose; já Arc.3S expressou não-preferência para oviposição e baixos níveis de antibiose. Em teste sem chance de escolha, os genótipos Arc.1S, Arc.3S, Arc.5S, Arc.1 e Arc.2 expressaram antibiose. Em teste onde avaliou-se a influência do tamanho de grãos na manifestação da resistência, os genótipos selvagens Arc.1S, Arc.3S e Arc.5S expressaram antibiose e/ou não-preferência para alimentação; Arc.1 e Arc.2 expressaram antibiose. No estudo com grãos armazenados sob diferentes temperaturas observou-se antibiose em grãos de Arc.1S e Arc.2, independentemente da temperatura em que estiveram confinados; já com Arc.1, notou-se que os efeitos antibióticos de seus grãos foram reduzidos à medida que a temperatura de armazenamento se elevou. Constatou-se também que 25°C não é a melhor temperatura para discriminar genótipos em estudos de resistência, comparativamente à 20°C e 30°C. Em casa-de-vegetação observou-se que as vagens e grãos do genótipo Arc.4 foram mais preferidos para oviposição e alimentação por A. obtectus em relação aos demais materiais; por sua vez Arc.1, Arc.5S, Ipa 6, Arc.3S e Porrillo 70 revelaram-se menos preferidos para oviposição e alimentação pelo caruncho. / Aiming to evaluate the effects of the time and temperatures of storage bean grains Phaseolus vulgaris L. on the expression of the resistance to the bean weevil Acanthoscelides obtectus (Say, 1831), free and no choice tests were accomplished under laboratory conditions. The preference for oviposition and damages caused by the weevil in pods and grains were evaluated under greenhouse conditions. The genotypes used in the assays were Arc.1S, Arc.3S, Arc.5S, Carioca Pitoco, Ipa 6, Porrillo 70, Onix, Arc.1, Arc.2, Arc.3 and Arc.4. In free choice test using grains of different ages, the wild genotypes Arc.1S and Arc.5S expressed antibiosis and/or no-preference for feeding as resistance mechanisms to A. obtectus; the genotypes Arc.1 and Arc.2 presented only antibiosis; Arc.3S expressed no-preference for oviposition and low antibiosis levels. In no-choice test, the genotypes Arc.1S, Arc.3S, Arc.5S, Arc.1 and Arc.2 expressed antibiosis. In test where the influence of the size of grains was evaluated in the expression of resistance, the wild genotypes Arc.1S, Arc.3S and Arc.5S expressed antibiosis and/or no-preference for feeding; Arc.1 and Arc.2 expressed antibiosis. In the study with grains stored under different temperatures, antibiosis was observed in grains of Arc.1S and Arc.2, independently of the confinement temperature; in Arc.1, it was observed that the antibiotic effects of its grains were reduced as the storage temperature increased. It was also verified that 25°C was not the best temperature to discriminate genotypes in studies of resistance, comparatively to 20°C and 30°C. In greenhouse the pods and grains of Arc.4 were the most preferred for oviposition and feeding by A. obtectus, comparing to the other materials; Arc.1, Arc.5S, Ipa 6, Arc.3S and Porrillo 70 were the least preferred for oviposition and feeding by the bean weevil.

Most plants are capable of mounting resistance responses to various pathogen attacks. For a hypersensitive response (HR) to occur, a dominant or semi-dominant resistance (R) plant gene is required to recognize a dominant avirulence (Avr) factor of the pathogen. Three types of Arabidopsis thaliana, Dijon-17 (Di-17), Dijon-3 (Di-3), and Columbia-0 (Col-0), are significant in understanding the genetics of Turnip crinkle virus (TCV) resistance. It has been shown that three genes are needed for successful resistance to TCV in A. thaliana: the dominant R gene HRT, the recessive gene rrt, and a third gene, TIP. Crosses of Di-17 and Di-3 plants, and crosses of Di-3 and Col-0 plants are being analyzed to determine the genotype of the F1 progeny. Using cleaved amplified polymorphic sequence (CAPS) markers, it is possible to determine the genotype of the progeny compared to the wild-type parents at the HRT and TIP loci. Additionally, protein analysis tools will be employed to compare the Di-3 and Di-17 TIP alleles to determine if there are any significant differences in the protein.

During post-alighting behaviour gravid female turnip root fly, D.jloralis, select a plant for oviposition predominantly during the initial landing phase; the cabbage root fly, D.radicum, also utilise the leaf resting phase. The post-alighting behaviour exhibited by D.radicum and D.floralis infers that oviposition site selection is primarily based upon positive stimuli present on the leaf surface. Ranking of four genotypes of plants for antixenotic resistance to oviposition by D.radicum and D.jloralis was found to be the same for both fly species, tested in the laboratory (swede cv Doon Major, most susceptible; kale cv Fribor, most resistant) and varied x80 (D.floralis) and x5 (D.radicum). Field experiments showed that oviposition (which was dominated by D.radicum) varied x2 between plant genotypes (swede cv Doon Major, most susceptible; swede cv GRL aga, most resistant). Testing of Brassica leaf surface extracts, applied to surrogate plants, indicated that leaf surface chemicals strongly influence the site of oviposition of D.floralis. Methanol soluble polar compounds are the most stimulatory element for D.floralis and a fraction which contained aliphatic glucosinolates stimulated oviposition strongly although glucosinolates were not the primary oviposition stimulant. Collaborative experiments indicate that "CIF" (cabbage identification factor) is probably present in this fraction. The concentrations of Brassica root sugars are generally reduced by the damage of both D.radicum and D.floralis and appear to influence larval development. The percentages of plant fibre and lignin in the roots of Brassicas rise following the damage of D.floralis. The concentrations of individual glucosinolates in Brassica roots arc radically altered by the damage of D.floralis and D.radicum. D.floralis damage resulted in a rise in the concentration of aromatic glucosinolates and a fall in the concentration of aliphatic glucosinolates. D.radicum damage generally resulted in an elevated concentration of both aliphatic and aromatic glucosinolates. There was no clear evidence that glucosinolatc profiles were associated with different levels of antibiotic resistance to D.radicum and D.floralis. GRL aga (SCRI breeding line) was consistently resistant to the oviposition and larval feeding of D.radicum and D.floralis both in the laboratory and in the field. It was shown that the use of end-of-season chemical analysis to assess the influence of plant chemistry on insect development or host plant resistance in field experiments and the use of damage indexes based on the percentage of a plant root damaged by D.radicum may be flawed.

The soil-borne filamentous fungus Trichoderma virens is a biocontrol agent with a well known ability to produce antibiotics, parasitize pathogenic fungi and induce systemic resistance in plants. Here we report the identification, purification and characterization of an elicitor secreted by T. virens; a small protein designated Sm1 (small protein 1). Confrontation and disk assays demonstrated that Sm1 lacks toxic activity against plants and microbes. Native, purified Sm1 triggers production of reactive oxygen species in rice (Oryza sativa) and cotton (Gossypium hirsutum), and induces the expression of defense related genes both locally and systemically in cotton. Gene expression analysis revealed that SM1 is expressed throughout fungal development and is transcriptionally regulated by nutrient conditions and the presence of a host plant. When T. virens was co-cultured with cotton in an axenic hydroponic system, SM1 expression and secretion of the protein was significantly higher than when the fungus was grown alone. These results indicate that Sm1 is involved in plant-Trichoderma recognition and the induction of resistance by activation of plant defense mechanisms. Following the cloning of SM1, strains disrupted in or over-expressing SM1 were generated. Targeted gene disruption revealed that SM1 was not involved in fungal development. Expression of defense related genes in cotton and maize (Zea mays) was induced locally and systemically following colonization by T. virens in the hydroponic system. Low levels of expression of cotton or maize defense genes were found when seedlings were grown with a T. virens strain disrupted in SM1, ssupporting the Sm1-elicitor hypothesis. Additionally, unique proteins in T.virens-cotton/maize interaction were identified. Thus, the induction of defense responses in two agriculturally important crops appears to be microbially mediated. Functional analysis of a cell wall degrading enzyme, beta-1,6-glucananse (Tv-bgn3) from T. virens, demonstrated involvement of this enzyme indirectly in mycoparasitic activity of T. virens. Protein extracts from the strain disrupted in TV-BGN3 displayed reduced capability to inhibit growth of Pythium ultimum as compared to the wild-type. Additionally, protein extracts from the strains co-expressed with TV-BGN2 (beta-1,3-glucananse) from T. virens showed a significantly increased capability to inhibit growth of P. ultimum and Rhizoctonia solani hyphae.