Thesis (Ph.D.)--University of Adelaide, Dept. of Crop Protection, 1997. / Errata and addenda attached. Bibliography: leaves 195-211.

Thesis (Ph.D. (Botany)) - Rhodes University, 2008.

Doctor of Philosophy / Biochemistry and Molecular Biophysics Interdepartmental Program / Gerald R. Reeck / Armet is a bifunctional protein that is apparently universally distributed among multicellular animal species, vertebrate and invertebrate alike. A member of the Unfolded Protein Response, (UPR) Armet promotes survival in cells that are under endoplasmic-reticulum (ER) stress. I have carried out biophysical studies on human Armet looking for compounds that bind to Armet and hence could reduce its anti-apoptotic function, thus potentially joining the growing class of pro-apoptotic drugs. Performed primarily with 1H-15N HSQC NMR, ligand studies showed that approximately 60 of the 158 residues are potentially involved with binding. The 60 residues are distributed throughout both domains and the linker suggesting multi-domain interaction with the ligand. Circular dichroism studies showed heat denaturation in a two-step unfolding process with independent unfolding of both domains of Armet with Tm values near 68°C and 83 C with the C-terminal domain unfolding first, as verified by 1H-15N HSQC NMR measurements. I also provide the first identification of UPR transcripts in pea aphids, Acyrthosiphon pisum, the genetic model among aphids. I measured transcript abundance with hope of finding future transcriptional targets for pest mitigation. I identified 74 putative pea aphid UPR components, and all but three of the components have higher transcript levels in aphids feeding on plants than those that fed on diets. This activated UPR state is attributed to the need for saliva proteins for plant feeding. Because aphids are agriculturally significant pests, and saliva is pivotal to their feeding on host plants, genes that encode saliva proteins may be targets for pest mitigation. Here I have sought the aphid’s saliva proteome by combining results obtained in several laboratories by proteomic and transcriptomic approaches on several aphid species. With these data I constructed a tentative saliva proteome for the pea aphid by compiling, collating, and annotating the data from several laboratories. I used RNA-seq to verify the transcripts in pea aphid salivary glands, thus expanding the proposed saliva proteome from approximately 50 components to around 130 components, I found that transcripts of saliva proteins are upregulated during plant feeding compared to diet feeding.

No description available.

Penetration of sink as well as source leaves of wheat plants by the Russian wheat aphid, Diuraphis noxia (Mordvilko) was investigated using light, fluorescence and transmission electron techniques, to determine the feeding strategies adopted by the aphid in penetrating and successfully feeding from the phloem, and to assess the structural effects of the probing and feeding behaviour of D. noxia on the feeding sites. Examination of aphid-infested sink, as well as source leaf tissue, showed that D.noxia probed in cells of the vascular bundle more frequently than mesophyll cells. Within the vascular bundle, thin-walled sieve tubes were visited (probed) more than the other cells. In sink leaf material, 68 of 82 (83%) stylets and stylet tracks encountered during the examination of 1000 serial sections (from 5 different plants) terminated in thin-walled sieve tubes and only 14 (17%) in thick-walled sieve tubes. Thin-walled sieve tubes were visited more significantly than thick-walled sieve tubes. However, examination of the aphid.,.infested sink leaf on a per centimetre basis, from the tip of the leaf, revealed that thick-walled sieve tubes in the area closest to the tip (0-2cm from the tip) were as attractive to the aphid as were thin-walled sieve tubes, with no significant difference in the number of times thick- and thin-walled sieve tubes were probed in this area. Some 2-4cm from the tip however, thinwalled sieve tubes were significantly more probed and therefore more attractive than thick-walled sieve tubes. Examination of 2000 serial sections using aphid-infested source leaf tissue, showed that the thin-walled sieve tubes were significantly more probed than thickwalled sieve tubes, along the whole leaf, expressed as a total of all leaves, as well as on a per centimetre basis along the length of the leaf, with 212 (95%) of 222 terminations within the thin-walled sieve tubes and only 10 (5%) in thick-walledsieve tubes. The aphid probed the small vascular bundles (loading bundles) many more times than intermediate or large transport vascular bundles, in sink as well as source leaf. Of a total of 82 stylets and stylet tracks encountered in sink leaf tissue, 31 terminated in small vascular bundles ang the remaining 28 and 16 were located within large and intermediate vascular bundles respectively. In source leaf tissue 121 of 222 stylets and stylet tracks encountered were associated with small vascular bundles and only 58 tracks and 43 tracks with intermediate and large vascular bundles, respectively. The effect of sustained RWA feeding on the transport capacity was examined after the application of 5,6 carboxyfluoresceine diacetate (5,6-CFDA) in control (sink and source leaf tissue) and aphid-infested (source) wheat leaves, using fluorescence microscopy. After 3h acropetal longitudinal transport of 5,6-CF had occurred in sink leaves in longitudinal veins, as well as a lateral transfer via cross veins and subsequent unloading into mesophyll cells close to the tip of the leaf was observed. In control leaf tissue, the fluorescence front was detected up to about 5cm from the point of application and was only associated with the phloem and not unloaded. In contrast, aphid-infested leaf tissue showed very little 5,6-CF transport, being limited to 2cm or less from the point of application. Structural damage to the phloem in general and to the sieve tubes in particular within of control and infested wheat leaves was investigated using transmission electron microscopy (TEM). In addition, leaf strips were mounted in aniline blue to visualise callose deposition using the fluorescence microscopy. At the TEM level. infested leaf tissue showed various abnormalities, which included destruction of cell contents, membrane damage and subsequent loss of cell contents. TEM studies suggest severe osmotic shock resulted from the aphid's probing. Examination of leaf tissue using fluorescence microscopy showed that there was very little characteristic aniline blue-stained callose visible in control leaf tissue, other than the thin diffuse patches along the sieve plates and punctate spots associated with pore plasmodesmatal areas and plasmodesmatal aggregates. In contrast, the aphid-infested leaf tissue was heavily callosed, with callose deposited not only within the phloem tissue but also in neighbouring vascular parE:}nchyma cells as well. The data collectively suggest that D. noxia feeds preferentially within thin-walled sieve tubes, within the small longitudinal vascular bundles in sink , as well source leaf tissue. Based upon the data presented here the thin-walled sieve tubes in the wheat leaf appear to be more attractive to the aphid and that they are probably more functional in terms of transport system and unlo?lding in sink leaves. Aniline blue stained leaf material that had previously hosted large aphid colonies showed evidence of extensive callose deposits 24 to 36h after the aphids were removed, suggesting that the aphids caused severe mechanical damage to the vascular tissue and mesohyll cells as well. Damage (transient or more permanent) and the subsequent deposition of wound callose, disrupted phloem transport and hence the export of photoassimilate from the leaves.

The Russian wheat aphid (RWA) is a pest of cereals, such as wheat and barley. It feeds on these hosts by injecting saliva into the plants’ phloem tissue and consuming the mixture of saliva and photoassimilates. It has been proposed that the insect’s saliva contains elicitors or virulence factors, which cause the symptoms typically observed in susceptible wheat cultivars. These are leaf rolling, chlorotic streaking, a decrease in yield and death in cases of heavy infestation. In contrast, resistant plants display symptoms typical of defence responses, such as the formation of necrotic lesions and an increase in the expression of pathogenesis related proteins. But, most importantly, RWA feeding on these hosts does not result in their subsequent death. The objectives of the present study are thus to elucidate any putative virulence factors, present in insect saliva, that can result in the breakdown of resistance of cultivars and thus, lack of recognition and/or delayed onset of the plants’ defence responses. Thus, this thesis investigates the RWA on protein level to determine which components of these insects induce the different changes observed in the resistant and susceptible plants. Also, it examines whether or not the biotypes uniquely altered their elicitors in response to selective pressure. In Chapter 1 a brief introduction is presented on the Russian wheat aphid, its distribution and the effects of its feeding on resistant and susceptible wheat cultivars. In Chapter 2 a literature review provides insight on how the Russian wheat aphids feed and survive on wheat. It also outlines the control mechanisms which plants could employ to withstand attack from pests and pathogens. In Chapter 3 proteins were extracted from different parts of two Russian wheat aphid biotypes and separated on high pressure liquid chromatography (HPLC). Two biotypes were selected for the study to provide comparative information on the development of biotypes and/or their virulent elicitors. The presence of the potential elicitors was determined by examining the extent of leaf rolling chlorotic streaking/spots on injected plants’ leaves, determining the activity of defence related enzymes of the injected plants and visualizing the proteins extracted from these plants on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels. This was done in order to examine the plants on phenotypic, enzymatic and proteomic levels, which could confirm the results obtained on three different levels. It was found that resistant cultivars react similarly to the two biotypes, but that the RWA biotypes differ significantly on a protein level. Potential motivations for these variations are discussed. Results presented in this dissertation represent the outcomes of a study conducted from March 2005 to December 2006 in the Department of Genetics, Forestry and Agricultural Biotechnology Institute, University of Pretoria, under the supervision of Prof. A-M Botha-Oberholster. Chapter 3 is being prepared to be submitted for review in Insect Biochemistry and Molecular Biology. / Dissertation (MSc (Plant Genetics))--University of Pretoria, 2008. / Genetics / unrestricted

No description available.

No description available.

The populations of aphidophagous insects in low-spray and conventional programs were surveyed in Virginia apple orchards. Two sampling methods were utilized;aphid colony collection and beating tray collection. The low-spray blocks employed the use of mating disruption for control of the key direct pests, codling moth, Cydia pomonella (Linnaeus), and leafrollers. The abundance and diversity of aphid predators in low-spray mating disruption programs and conventionally controlled apple orchards was compared. Both sampling methods suggested that aphid predators were more plentiful in the mating disruption blocks. Several mating disruption blocks were found to accumulate significantly more predator-days and diversity than the matched control blocks. The reduction of insecticide input into the low-spray blocks may have allowed higher populations of aphidophagous insects to occur. Aphid populations in mating disruption blocks sometimes declined faster and had less resurgence than those in the conventionally controlled blocks. The tufted apple bud moth (TABM), Platynota idaeusalis (Walker), populations in four apple orchards in Winchester, Virginia, were compared for resistance to azinphosmethyl. These orchards were suspected by the growers to have resistant populations of TABM. Pheromone traps with insecticide incorporated into the adhesive were used to collect and test the moths. The results were not analyzed due to the low number of moths collected and high variability in mortality. / Master of Science

Dissertation (PhD)--University of Stellenbosch, 2001. / ENGLISH ABSTRACT: The basic biology of Eriosoma lanigerum (Hausmann) and its natural enemy, Aphelinus mali (Haldeman), was investigated in the Western Cape Province of South Africa. The first instar nymph can be identified by the absence of cornicles and the adult female by the presence of the vulva. Body length and distance between cornicles can be used to distinguish between the 2nd , 3rd and 4th instars. The development of E. lanigerum was negatively influenced by temperatures above 27DC. The net replacement rate (Ro) and intrinsic rate of increase (rm) peaked at 20DC. The theoretical lower and upper threshold temperatures for development were estimated at 4.48DC and 28.07DC, respectively. Crawlers of E. lanigerum migrated from the roots up into the trees during spnng to start new infestations. Colonies became visible from December and maximum colony numbers were reached from the end of summer until autumn, at which time most of the colonies were parasitised by A. mali. Colony numbers declined at the end of autumn after high parasitism and the appearance of winged E. lanigerum. The aphid overwintered on the tree. Chemical sprays, rain during spring and high temperature influenced population numbers negatively. E. lanigerum can be monitored by counting the unparasitised colonies in leaf axils of half of each of 25 trees per 2 hectare plot with 5 unparasitised colonies as the economic threshold. Sampling error was high at 40% but increasing the number of trees did not reduce it. Presence-absence sampling, which will reduce the time required for monitoring, did not seriously compromise the reliability of decisions regarding the necessity for intervention. Chemicals containing nitrogen usually sprayed for bitterpit control stimulated the settling of E. lanigerum crawlers on Granny Smith trees, while fruit weevil barriers for the control of Phlyctinus callos us Boh. limited crawler movement into the trees but did not prevent colonisation. All the postembryonic developmental stages of E. lanigerum were parasitised by A. mali. Complete parasitism of the population was never reached as younger instars sheltered under other aphids and mummified aphids. Development of A. mali was not influenced negatively by high temperatures. The minimum developmental temperature and number of degree days needed for development of the larval stage and emergence of the adult from the mummy were .6.72°C and 172.41°D, and 10.27°C and 109.89°D respectively. Mummies collected during early winter survived long periods of cold storage in postdiapause. The rrummum threshold temperature for postdiapause development of A. mali was ·10.15°C. Most chemicals tested against A. mali adults were highly toxic to the wasp, except endosulfan and two growth regulators, flufenoxuron and fenoxycarb. The mortality of adults exposed to the fungicides tested was low within the first 24 hours. The percentage emergence from the mummies was high for all chemicals tested, but more than 60% of the adults died soon after emergence from mummies treated with chlorpyrifos. Nearly 30% of the adults died soon after they emerged from carbaryl (XLR-Plus) and fenthion treated mummies. The growth regulators, flufenoxuron and fenoxycarb, did not influence fecundity adversely. / AFRIKAANSE OPSOMMING: Die basiese biologie van Enosoma lanigerum (Hausmann) en sy natuurlike vyand, Aphelinus mali (Haldeman), in die Weskaap Provinsie van Suid-Afrika isondersoek. Die eerste instar nimf kan aan die afwesigheid van kornikels en die volwassenes aan die teenwoordigheid van die vulva uitgeken word. Die liggaamslengte en afstand tussen die kornikels kan gebruik word om tussen instar 2, 3 en 4 te onderskei. Die ontwikkeling van E. lanigerum word nadelig deur temperature bo 27°C beïnvloed. Die netto vervangingstempo (Ro) en intrinsieke tempo van toename (rm) was die hoogste by 20°C. Die teoretiese minimum en maksimum temperatuur drempelwaardes vir ontwikkeling was 4.48°C en 28.07°C onderskeidelik. In die lente beweeg E. lanigerum kruipers op vanaf die wortels tot in die bome om nuwe kolonies te begin. Kolonies is sigbaar vanaf Desember en die hoogste koloniegetalle word aan die einde van die somer tot die herfs bereik, wanneer die meeste van die kolonies dan ook deur A. mali geparasiteer word. Teen laat-herfs neem koloniegetalle af as gevolg van hoë parasitisme en die verskyning van gevleuelde E. ,lanigerum. E. lanigerum oorwinter op die appelboom. Chemiese behandelings, reën gedurende die lente en hoë temperatuur beïnvloed koloniegetalle nadelig. E. lanigerum kan deur die aantal ongeparasiteerde kolonies in die blaaroksels van die helfte van 25 bome per 2 hektaar blok te tel, met 5 ongeparasiteerde kolonies as die ekonomiese drempelwaarde, gemonitor word. Die steekproefnemingsfout was hoog (40%), maar kon nie verminder word deur die aantal bome wat gemonitor is te verhoog nie. Steekproefneming, vir aan- of afwesigheid van kolonies, wat monitortyd sal verminder, het min invloed op die betroubaarheid van besluitnemings oor die noodsaaklikheid van bespuitings gehad. Stikstofbevattende chemikalieë, wat vir die beheer van bitterpit gespuit word, stimuleer vestiging van E. lanigerum kruipers op Granny Smith bome, terwyl snuitkewerversperrings, vir die beheer van Phlyctinus callosus Boh., die opwaartse beweging van kruipers in die bome beperk, maar nie kolonievorming van E. lanigerum verhoed nie. Alle postembrioniese ontwikkelingstadiums van E. lanigerum is deur A. mali geparasiteer. Totale parasitisme is nooit bereik nie, omdat jonger instars onder ander bloedluise en gemummifiseerde bloedluise skuil. Die ontwikkeling van A. mali word nie deur hoë temperature benadeel nie. Die minimum ontwikkelingstemperatuur en graaddae, nodig vir ontwikkeling van die larwale stadium en die verskyning van die volwassene uit die mummie, was 6.72°C met 172.4loD en 10.27°C met 109.89°D, onderskeidelik. Mummies wat vroeg in die winter versamel is, het lang periodes van koelopberging oorleef. Die minimum temperatuur drempelwaarde vir A. mali ontwikkeling in postdiapouse was 10.15°C. Die meeste van die chemikalieë wat getoets is, was hoogs toksies vir die volwasse wesp, behalwe endosulfan en die twee groeireguleerders, flufenoxuron en fenoxycarb. Die mortaliteit van volwassenes wat aan swamdoders blootgestel is, was laag binne die eerste 24 uur na blootstelling. Die persentasie uitkoms vanuit mummies was hoog vir al die chemikalieë wat getoets is, maar met chlorpyrifos het 60% van die volwassenes net na uitkoms doodgegaan. Ongeveer 30% van die volwassenes is dood na verskyning vanuit mummies wat met carbaryl (XLR-Plus) en fenthion gespuit is. Die groeireguleerders, flufenoxuron en fenoxycarb, het nie die vrugbaarheid van die parasiet merkbaar beïnvloed nie.