Dissertations / Theses on the topic 'Seedling resistance'

The increasing world population is putting pressure on global food production. Agriculture must meet these growing demands by increasing crop yields. One phenomenon which prevents seedling emergence and damages crop yield is soil crusting. Understanding of soil crusting and the factors which influence it is fundamental to ensuring good crop production. An instrument which will test soil crust strength in a novel way, mimicking seedling growth, may lead to pre-emptive agricultural soil management which could increase crop production. This work details the process of design, construction and testing of an ascending penetrometer to measure soil crust strength. The full design process is discussed from concept generation and evaluation, using experimental methods and a multi-criteria decision making tool, through to final design configuration, specification, manufacture and testing. Traditionally, soil penetrometers measure soil strength by forcing a probe from the surface of the soil into the bulk soil below. To more accurately measure the direct impedance a seedling would experience a device should measure impedance from the bulk soil upwards and into the soil crust, mimicking what a growing seedling would experience. Results prove that the manufactured ascending penetrometer with a force resolution of 0.01N and displacement resolution of 0.0004mm is capable of detecting differences in soil crusts. At these resolutions and accuracy to 0.1N and 0.1mm excellent repeatability was achieved. The machine is therefore a useful and realistic tool for quantitatively comparing soil crusts in soil. It is hoped that being able to compare soil crust strength will lead to improved soil management techniques.

[Abstract]Crown rot, caused by Fusarium pseudograminearum (Fpg), is an important soilborne disease of wheat and barley. The degree of crop damage depends on seasonal conditions. Typically, high moisture conditions early in the season encourage seedling infection from stubble residues. Moisture stress later in the season leads to the production of unfilled “whiteheads”. Current control relies on cultural practices and sowing of partially resistant varieties. In order to understand the nature of partial resistance, I have examined the patterns of disease symptom development and pathogen spread in susceptible and partially resistant tissues of both pot-grown wheat, barley and oat seedlings and field-grown inoculated wheat trials. Further research was conducted to determine whether differences in pathogenicity occur amongst a small subset of Australian Fpg isolates. Seedling experiments confirmed that differences in disease ratings between susceptible and partially resistant genotypes are detected in younger leaf sheaths of older seedlings. At later harvest times differences between these genotypes are not significant in older leaf sheaths. Re-isolation of Fpg from inoculated seedlings has shown that each tissue was infected later in partially resistant genotypes compared to susceptible ones with a significantly lower number of isolations recorded at each harvest time in 42 day old seedlings. Barley cultivars were rapidly infected by the pathogen and exhibited high levels of disease symptoms. By comparison levels of infection in oats were low compared to all other genotypes. No significant differences between genotypes were observed in coleoptile tissues, either in fungal colonisation or development of disease symptoms. Disease development in the subcrown internode varied between lines/cultivars but was not representative of the relative susceptibility of each genotype. The pathogen did not appear to invade plant tissue via the vascular system but rather spread directly across the stem from leaf sheath to leaf sheath. Field trials were designed to study disease symptom development and localisation of Fpg hyphae in all expanded tissues (excluding head and roots) in wheat genotypes of known susceptibility to crown rot. Plants were harvested at approximately fortnightly intervals throughout the growing season. The main effects and interactions of harvest, genotype and tiller on each plant part were examined with a detailed statistical analysis of differences seen in these factors between susceptible and partially resistant wheat genotypes, in two inoculated field trials. While differences between genotypes were mostly not significant at each harvest when disease rating or isolations from leaf sheath tissues were examined, important differences between susceptible and resistant genotypes were seen in disease developments and Fpg infections of stem tissue in field trials. Restriction of pathogen growth and symptom development was more pronounced in the tissues of 2-49 (possesses seedling resistance) than in the field resistant Sunco. At present, the mechanisms that lead to these resistance responses are unknown. The pathogenicity study aimed to determine whether 7 Fpg isolates and a mixed inoculum differed in ability to cause crown rot in 9 wheat genotypes ranging in susceptibility to this disease. Although a genotype*inoculum interaction was significant, there is no evidence of stable pathogenic races in the isolates examined in these experiments. The growth of all isolates was partially inhibited in a consistent manner on resistant genotypes when compared to very susceptible genotypes. These results confirm significant differences in the aggressiveness of Fpg isolates on wheat, evidenced by variation in mean disease severity between isolates growing on a range of host genotypes.

Genetic studies were carried out to determine the inheritance of unknown seedling resistance (USR) to leaf rust (caused by Puccinia hordei Otth.) in the barley cultivar Ricardo. In the greenhouse Ricardo/Gus F3 (187 lines) and BC1F2 (130 lines) populations based studies using an array of P. hordei pathotypes (pts), revealed that the USR in Ricardo was conferred by a single dominant gene, which was tentatively named RphRic. Bulk segregant analysis (BSA) of the F3 population using a multiplex-ready PCR technique mapped RphRic on chromosome 4H flanked by markers GBM1220 and GBM1003 at distances of 17.4 cM and 20.4 cM, respectively. Being the first gene for leaf rust resistance mapped on chromosome 4H, RphRic was catalogued as Rph21. Phenotyping of Ricardo/Peruvian (Rph2) F3 populations and genotyping of both parents using the Rph2-linked marker ITS1 confirmed the presence of Rph2 in Ricardo. The Ricardo/Gus F3 and BC1F2 populations segregated for the presence of an additional gene when tested under field conditions using the same pathotype (pt), 5457P+ (used in greenhouse). This uncharacterised adult plant resistance (APR) against P. hordei, found in Ricardo, appeared to be distinct from Rph20 when genotyped using a closely linked marker bPb-0837. Responses of 113 advanced breeding lines and cultivars of barley (Hordeum vulgare L. subsp. vulgare), along with the susceptible control genotype Gus, were assessed against P. hordei pts in the greenhouse at seedling and field at adult plant growth stages. The tests revealed the presence of APR in 68 lines, USR in 23 lines and the seedling resistance gene Rph3 in three lines. Marker bPb-0837 was present in 35 of the 68 lines carrying APR, which suggested that these 35 lines carry APR gene Rph20. The remaining 33 lines, which lacked the Rph20 linked marker, likely carry new sources of APR. Pedigree analysis of the 68 lines found to carry APR revealed that 32 were related to cultivar (cv.) Gull and to H. laevigatum, two were related to cv. Bavaria and one related to cvv. Manchuria and Taganrog. Ancestral pedigree analysis also revealed the common presence of cv. Diamant (X-ray mutant) in the parentage of lines likely carrying Rph20. The remaining 19 lines lacked detectable seedling resistance and were susceptible in the field at adult plant growth stages. Four international barley nurseries comprising 820 lines with 579 unique pedigrees were sourced from the International Centre for Agricultural Research in the Dry Areas (ICARDA) and analysed for resistance against isolates of P. hordei, P. graminis f. sp. tritici (Pgt) and barley grass stripe rust (BGYR). Overall analyses of the responses of 783 lines (excluding 37 missing lines) to P. hordei showed that 728 (93%) carried the major seedling resistance gene Rph3, five (0.65%) carried USR, six (0.75%) carried uncharacterised APR and 44 (5.6%) lines were susceptible at all growth stages. Of the six lines identified with uncharacterised APR, three likely carried Rph20 based on the presence of the Rph20-linked marker bPb-0837. Based on tests with several control genotypes, marker bPb-0837 was found to be more reliable than Ebmag0833 in detecting the presence of Rph20. All lines were resistant to Pgt pt 98-1,2,3,5,6 when tested as seedlings in the greenhouse. Out of the 783 lines tested, 164 produced immune responses, 284 produced resistant (1= to 3) responses and 335 produced mesothetic (X type) responses against pt 98-1,2,3,5,6. All but two 783 lines were highly resistant to BGYR in greenhouse tests, showing immune responses. The usefulness of 148 simple sequence repeats (SSRs) in revealing variability among Australian isolates of P. hordei were assessed. The markers comprised 71 developed for Pgt, 40 developed for P. triticina (Pt) and 37 developed for P. coronata f. sp. avenae (Pca). SSRs were tested across 22 pts of P. hordei from Australasia including one isolate of each of the control pathogens [Pt, Pgt, P. striformis f. sp. tritici (Pst), BGYR and P. graminis f. sp. avenae (Pga)]. Genotyping of P. hordei was also conducted with the PCR-fingerprinting primers M13 and (GACA)4. The SSRs developed from Pgt and Pt showed 100% cross amplification in P. hordei, while only nine Pca SSRs showed amplification in P. hordei. Of the 148 markers tested, only two Pgt SSRs (F4-15 and F7-22) were polymorphic. Both PCR-fingerprinting primers revealed polymorphisms among the isolates, with (GACA)4 generating the most informative fragments. Both SSRs and PCR fingerprinting markers out grouped the control pathogens Pt, Pgt, Pst, BGYR and Pga from the P. hordei pts. Polymorphic information content (PIC) values of SSR markers F4-15 and F7-22 were calculated as 0.50 and 0.55 respectively. Molecular analyses revealed evidence of clonal lineages among the P. hordei pts, supporting the hypothesis that some of the pts arose from mutational changes in the virulence of a founder pt. Sexual recombination within P. hordei in Australia on the alternate host Ornithogalum umbellatum may have resulted in some new pts with different virulence against Rph genes. This is the first study of Australasian pts of P. hordei using PCR-fingerprinting technique and SSR genotyping.

In wheat, stem rust is known to rapidly evolve new virulence to resistance genes. While more than 50 stem rust resistance (Sr) loci have been identified in wheat, only a few remain effective, particularly against the highly virulent race Ug99 (TTKSK race) and a mixture of durum-specific races. An association mapping (AM) study based on 183 durum wheat accessions was utilized to identify resistance loci for stem rust response in Ethiopia over four seasons and artificial inoculation with Ug99 (TTKSK race) and a mixture of durum-specific races under field conditions as well as in greenhouse test at seedling stage under controlled conditions for resistance to four highly virulent stem rust races: TRTTF, TTTTF, (TTKSK (Ug99) and JRCQC. The panel was profiled with 1,253 SSR and DArT markers. Twelve QTL-tagging markers were significant (P < 0.05) across three to four seasons. The role of Sr13, Sr9, Sr14, Sr17, and Sr28 was confirmed. Thirteen significant markers were in regions with no Sr genes/QTLs. The results under controlled conditions showed that 15, 20, 19 and 19 chromosome regions harbored markers that showed significant effects for races TRTTF, TTTTF, TTKSK and JRCQC, respectively. These genomic regions showed marker R2 values ranging from 1.13 to 8.34, 1.92 to 17.64, 1.75 to 23.12 and 1.51 to 15.33% for races TRTTF, TTTTF, TTKSK and JRCQC, respectively. The study demonstrates that stem rust resistance in durum wheat is governed in part by shared loci and in part by race-specific ones. The QTLs identified in this study through AM will be useful in the marker-assisted development of durum wheat cultivars with durable stem rust resistance.

In wheat, stem rust is known to rapidly evolve new virulence to resistance genes. While more than 50 stem rust resistance (Sr) loci have been identified in wheat, only a few remain effective, particularly against the highly virulent race Ug99 (TTKSK race) and a mixture of durum-specific races. An association mapping (AM) study based on 183 durum wheat accessions was utilized to identify resistance loci for stem rust response in Ethiopia over four seasons and artificial inoculation with Ug99 (TTKSK race) and a mixture of durum-specific races under field conditions as well as in greenhouse test at seedling stage under controlled conditions for resistance to four highly virulent stem rust races: TRTTF, TTTTF, (TTKSK (Ug99) and JRCQC. The panel was profiled with 1,253 SSR and DArT markers. Twelve QTL-tagging markers were significant (P < 0.05) across three to four seasons. The role of Sr13, Sr9, Sr14, Sr17, and Sr28 was confirmed. Thirteen significant markers were in regions with no Sr genes/QTLs. The results under controlled conditions showed that 15, 20, 19 and 19 chromosome regions harbored markers that showed significant effects for races TRTTF, TTTTF, TTKSK and JRCQC, respectively. These genomic regions showed marker R2 values ranging from 1.13 to 8.34, 1.92 to 17.64, 1.75 to 23.12 and 1.51 to 15.33% for races TRTTF, TTTTF, TTKSK and JRCQC, respectively. The study demonstrates that stem rust resistance in durum wheat is governed in part by shared loci and in part by race-specific ones. The QTLs identified in this study through AM will be useful in the marker-assisted development of durum wheat cultivars with durable stem rust resistance.

Doctor of Philosophy<br>Department of Agronomy<br>Tesfaye T. Tesso<br>The lack of effective post-emergence weed control options is often highlighted as one of the major factors behind dwindling acreage under sorghum (Sorghum bicolor (L.) Moench) in the United States. The discovery of herbicide resistance sources in wild sorghum population and subsequent efforts to incorporate them into cultivated sorghum was received with much optimism to change weed management practices in sorghum. As the development of the technology advances, especially of the Acetolactate synthase (ALS) resistance, concerns over the temporary interveinal chlorosis and reduced seedling vigor in some of the resistant families became heightened. This thesis research is designed to shed light on the genetic basis of seedling chlorosis and assess its impacts on yield potential. The study has three parts; the first part is focused on identifying the genetic causes and plant mechanisms associated with the chlorotic phenotype. ALS herbicide resistant sister-lines expressing normal and chlorotic phenotypes were analyzed via RNA sequencing at four time points during seedling growth. The study identified several variants of genes coding chloroplast precursors and those that cause epigenetic modifications. Once confirmed, genetic markers can be developed to track these gene variants in the breeding population and eliminate segregates genetically prone to chlorosis/yellowing. The second part of the study focuses on assessing the effect of ALS resistance associated chlorosis on agronomic and nutritional parameters of sorghum inbred lines. A set of ALS resistant lines expressing different levels of the chlorotic phenotype were evaluated in replicated field trials and laboratory methods. Results showed that interveinal chlorosis delays flowering but does not have negative effect on yield and nutritional parameters with and without herbicide treatment. The last part addresses whether there is any yield drag that may be associated with herbicide resistance traits and foliar interveinal chlorosis. For this, we synthesized a large set (182) of hybrids from ALS resistant, ACCase resistant and regular (susceptible) seed and pollinator parents. The hybrids were then evaluated in three sets at multiple locations during the 2014 and 2015 crop seasons along with commercial checks. The results revealed that resistance to both herbicides do not cause any drag to grain yield. The traits also do not have any negative impact on grain and nutritional quality of resistant hybrids.

Doctor of Philosophy<br>Genetics Interdepartmental Program<br>Bikram S. Gill<br>Leaf rust, caused by fungus Puccinia triticina, is an important foliar disease of wheat worldwide. Breeding for race-nonspecific resistant cultivars is the best strategy to combat this disease. Aegilops tauschii, D genome donor of hexaploid wheat, has provided resistance to several pests and pathogens of wheat. To identify potentially new adult plant resistance (APR) genes, 371 geographically diverse Ae. tauschii accessions were evaluated in field with leaf rust (LR) composite culture of predominant races. Accessions from Afghanistan only displayed APR whereas both seedling resistance and APR were common in the Caspian Sea region. Seventeen accessions with high APR were selected for production of synthetic hexaploid wheat (SHW), using ‘TetraPrelude’ and/or ‘TetraThatcher’ as tetraploid parents. Six SHWs were produced and evaluated for APR to LR and resistance to tan spot at seedling stage. Genetic analysis and mapping of APR introgressed from accession TA2474 was investigated in recombinant inbred lines (RIL) population derived from cross between SHW, TA4161-L3 and spring wheat cultivar, ‘WL711’. Genotyping-by-sequencing approach was used to genotype the RILs. Maximum disease severity (MDS) for LR was significantly correlated among all experiments and APR to LR was highly heritable trait in this population. Nine genomic regions significantly associated with APR to LR were QLr.ksu-1AL, QLr.ksu-1BS, QLr.ksu-1BL.1, QLr.ksu-1BL.2, QLr.ksu-2DS, QLr.ksu-2DL, QLr.ksu-5AL, QLr.ksu-5DL and QLr.ksu-6BL. Association of QLr.ksu-1BL.1 with marker Xwmc44 indicated this locus could be slow-rusting APR gene, Lr46/Yr29. QTLs detected on 2DS, 2DL and 5DL were contributed by TA4161-L3 and are novel, along with QLr.ksu-5AL. Tan spot, caused by necrotrophic fungus, Pyrenophora tritici-repentis, has recently emerged as a damaging disease of wheat worldwide. To identify QTLs associated with resistance to Race 1 of P. tritici-repentis, F[subscript]2:3 population derived from cross between SHW, TA4161-L1 and winter wheat cultivar, ‘TAM105’ was used. Two major effect QTLs, QTs.ksu-1AS.1 and QTs.ksu-7AS were significantly associated with tan spot resistance and contributed by TA4161-L1. QTs.ksu-7AS is a novel QTL and explained 17% of the phenotypic variation. Novel QTLs for APR to LR and tan spot identified in SHWs add new variation for broadening the gene pool of wheat and providing resources for breeding of durable resistant cultivars.

Herbicide resistance is evolving fast in Italy, especially in cropping systems characterised by low diversity in space and time. In these systems there is often no rotation of herbicides mode of action (i.e. durum wheat and rice monoculture). Resistance is also evolving faster in cross-pollinating and genetically variable species (i.e. Lolium spp.). In Italy about 100,000 ha have been estimated to suffer resistant problems, mainly in durum wheat (Lolium spp., Avena sterilis and Papaver rhoeas) and paddy rice (mostly Schoenoplectus mucronatus and Cyperus difformis). ACCase is an essential enzyme in fatty acid biosynthesis in eukaryotes and prokaryotes. In plants, two forms of ACCase have been identified - the first is located in the chloroplast, the primary site of plant fatty acid biosynthesis, and the second in the cytosol. The homomeric ACCase in the cytosol of nearly all plant species and the heteromeric ACCase in the chloroplasts of dicots are insensitive to APP, CHD and DEN herbicides. In contrast, the plastidic homomeric ACCase in nearly all grass species is herbicide-sensitive, and this is the basis for selective control of grass-weeds by ACCase herbicides. The aims of the research were: (1) to monitor throughout the country the situation of resistance to Acetyl-CoA Carboxylase (ACCase) in Lolium and Phalaris species following complaints to the agrochemical industries; (2) to develop a reliable Petri dish quick test to detect ACCase-inhibitors resistance in Lolium and Phalaris species; (3) to characterise clethodim and pinoxaden resistance in Lolium spp. using different experimental approaches; (4) agronomic and molecular characterisation of a few populations of P. paradoxa resistant to ACCase inhibitors. Through classic pot screenings, populations with different patterns and levels of resistance have been selected to be studied with different approaches: outdoor dose-response, at molecular level using molecular markers and sequencing and at physiological level through enzyme bioassay. As pot experiments are costly and time consuming, a rapid test in Petri dishes based on survival data has been developed for detecting resistance to ACCase-inhibitors. Screening results revealed that among 41 tested Lolium populations, 23 were resistant to clodinafop, 19 to sethoxydim and 15 to pinoxaden. Among the 17 P. paradoxa populations, 3 were resistant to clodinafop, and one to sethoxydim and pinoxaden. Resistance to pinoxaden is associated to a wide pattern of cross-resistance among ACCase-inhibitors. The Petri dish seed quick test has been developed in order to find a discriminating herbicide concentration between susceptible and resistant populations; a herbicide concentration ("comparing dose") that gives results comparable to the recommended field dose used in greenhouse pot experiments; and a concentration discriminating between strong and weak resistance. The "comparing doses" has been validated using other populations not included in the set up experiments. The "comparing dose" for Lolium spp. have been: clodinafop 1 ?M, pinoxaden 0.2 ?M, clethodim 0.2 ?M and sethoxydim 0.1 ?M; for P. paradoxa, clodinafop 0.1 ?M, pinoxaden 0.05 ?M and clethodim 0.1 ?M. The Petri dish seed quick test revealed to be reliable with the two species in screening large numbers of samples compared with the costly and time consuming pot experiment. The quick test improves ACCase-resistance detection and can be adopted as a routine facility. Molecular investigation identified five ACCase mutations (Ile1781Leu, Trp2027Cys, Ile2041Asn, Asp2078Gly and Cys2088Arg) and revealed 12 genotypes in 14 clethodim resistant Lolium populations. Pinoxaden resistance has been reported associated to Lolium population mutants for five ACCase mutations (Ile1781Leu, Ile2041Val/Asn, Asp2078Gly, Cys2088Arg and Gly2096Ala) with specific homozygous and heterozygous status among mutations. In P. paradoxa two mutations have been associated to pinoxaden resistance: Ile1781Val and Asp2078Gly, always recognised at the homozygous status. The research has established that resistance to ACCase herbicides depends on the specific resistant allele(s), on the homo/heterozygous status of plants for the specific resistant allele(s), and on the combinations of different resistant alleles, as well as herbicide rates. To understand and devise resistance management strategies, knowledge of all these factors coupled with field records of herbicide and agronomic techniques used is essential. The overall situation of grass resistance in Italian durum wheat crops indicates that there is no single chemical that can solve all resistance problems. Therefore all stakeholders should be aware that IWM (or better ICM) is needed to properly manage resistance in the field. It must also be clear that IWM or ICM requires a higher technological level coupled with a deeper knowledge of all components of the cropping system.

Some aspects of the presence of systemic endophytic fungi in agriculturally important New Zealand grasses were studied in relation to plant breeding. Seedling resistance to adult Argentine stem weevil feeding in perennial ryegrass, Italian ryegrass and tall fescue was found to be related to the presence of their respective Acremonium endophytes in the seed rather than to plant genetic resistance. In addition a study of perennial ryegrass revealed that this resistance was independent of endophyte viability. The seedling resistance conferred by the endophyte of Italian ryegrass was found to be beneficial for field establishment. This endophyte differs from that in perennial ryegrass and tall fescue in that it does not confer resistance to Argentine stem weevil on mature plants, but only on seedlings. The extent of plant genetic seedling tolerance to adult Argentine stem weevil feeding was limited to broad inter-specific differences, with tall fescue more tolerant than perennial ryegrass and both of these more tolerant than Italian ryegrass. This ranking corresponds with previous observations on feeding preference on mature plants. A study of factors affecting the concentration of endophyte mycelia in infected seed of perennial ryegrass revealed that plant genetic factors had little effect. The major factors studied were: 1) the endophyte concentration in the maternal parent plant directly influenced the endophyte concentration in the seed. 2) nitrogen fertilizer applications to a seed crop reduced the concentration of mycelia in the seed, with earlier applications having a greater effect. 3) application of the fungicide propiconazole (Tilt) to a seed crop reduced the endophyte concentration in the seed. 4) the endophyte concentration in the seed was found to directly influence the endophyte concentration in seedlings, six month old plants and that of seed harvested from a first year seed crop. As there have been no previous reports of tetraploid perennial ryegrass cultivars with endophyte an experiment was conducted to determine if these could be developed by the standard procedure of colchicine treatment. The results revealed that endophyte was retained following colchicine treatment.

Pine weevils cause major damage to newly planted conifer seedlings in reforestation areas. However, recent findings indicate that small (“mini”) seedlings, planted at the age of 7-10 weeks, are gnawed less by pine weevils than the larger, conventionally planted seedlings. Thus, it has been proposed that planting young conifer seedlings in clear-cut areas may reduce the damage caused by pine weevils. In attempts to determine why mini seedlings appear to be less damaged by pine weevils than “conventional” seedlings, the volatiles released by Norway spruce and Scots pine mini seedlings were investigated, since such chemicals are of great importance in herbivore-plant communication, inter alia acting as repellents, attractants or antifeedants. Volatiles from the seedlings were collected, separated and identified by solid phase microextraction (SPME) followed by gas chromatography-mass spectrometry. The results show that there are high levels of chemodiversity among both spruce and pine seedlings. Between-tissue and age-related variations in their emissions were also found. Norway spruce clones infested by mites were also examined to assess genotype- and pest-specific stress reactions of Norway spruce. Finally, the effects of certain spruce defense compounds on the behavior of the large pine weevil Hylobius abietis were examined.<br>QC 20100818

Les herbivores consomment les tissus végétaux et affectent la survie et la croissance des végétaux. Les ennemis naturels des herbivores peuvent réguler leurs populations et indirectement diminuer les dommages sur la végétation. Les relations entre plantes, herbivores et prédateurs sont liées aux caractéristiques intrinsèques des espèces végétales mais aussi influencées par la communauté végétale environnante. L'hypothèse de résistance par association stipule qu'une communauté végétale diversifiée entraîne une diminution des dégâts sur une plante cible par dilution, répulsion des herbivores et/ou favorisation des ennemis naturels. Inversement, les herbivores peuvent se concentrer sur la plante-cible dans une communauté végétale diversifiée (susceptibilité par association). L'objectif de cette thèse est d'évaluer 1) l'influence de la composition de la communauté végétale sur les herbivores et leurs dommages sur une plante-cible, 2) l'impact de la régulation des herbivores par les prédateurs et 3) les interactions entre herbivores, communauté végétale et prédateurs, et les conséquences pour les intensités d'herbivorie sur la plante-cible. Nous nous sommes focalisés sur les dommages causés par les insectes herbivores sur des plantules d'essences forestières feuillues (Betula pendula, Quercus robur et Q. ilex). La variété importante de communautés végétales, au niveau inter (essences dominantes) et intraparcellaire (présence, structure et composition), nous a permis de tester les effets du voisinage végétal sur les insectes herbivores et leurs dommages sur les plantules cibles. Les oiseaux insectivores sont les principaux prédateurs d'insectes et nous avons cherché à estimer les effets de leur exclusion sur les insectes et les conséquences pour les plantules, et ceci le long des gradients de composition du voisinage végétal. Enfin, le degré de spécialisation (spécialiste/généraliste) et le mode de vie (exo/endophyte) ont été caractérisés car ils conditionnent les réponses des insectes herbivores à la végétation voisine ainsi que la prédation avienne. Le voisinage végétal des plantules-cibles modifie la charge en herbivores et les dommages engendrés. Ces réponses sont liées à la spécialisation de l'insecte herbivore : la colonisation des plantules par les espèces spécialistes (mineuses de feuilles) est négativement affectée par la structure de la végétation locale (présence, recouvrement arbustif). Les dommages d'insectes généralistes augmentent avec la richesse spécifique des peuplements et dans les parcelles dominées par des espèces conspécifiques. Les abondances des insectes externes et leurs dommages sont régulés par les oiseaux insectivores mais cet effet indirect dépend de l'espèce de plantule-cible et de la composition de la communauté végétale. Les effets de cascade trophique sont les plus intenses dans les parcelles de pin maritime où les plantules constituent une ressource nouvelle pour les niveaux trophiques supérieurs. Plus localement, le voisinage végétal modifie la sensibilité des arthropodes à la prédation: les oiseaux insectivores bénéficient aux plantules lorsque les proies sont plus accessibles (végétation environnante supprimée).Pour conclure, une des principales richesses de ces résultats vient de la mise en évidence d'interactions très fortes existant entre les effets des oiseaux insectivores et la végétation environnante. Les effets indirects des prédateurs et directs de la végétation environnante sont très liés au degré de spécialisation des insectes phytophages et à leur mode de vie<br>Herbivory is thought to depend on intrinsic plant resistance traits and negatively affect survival and growth of plants. Predators can depress herbivore populations and thereby indirectly limit the consumption of primary producers. The surrounding vegetation is also expected to modify the interactions between a focal plant, its herbivores and their natural enemies. The surrounding vegetation may disrupt the colonisation and the consumption of a focal plant by insect herbivores, providing associational resistance. Increasing diversity of neighbouring vegetation can decrease resource availability or enhance herbivores control by predators. By contrast, associational susceptibility may occur with higher levels of herbivory in a focal plant in more diverse plant communities. We tested 1) the effect of the neighbouring vegetation on the levels of herbivory on seedlings, 2) the indirect impact of predators on herbivores and subsequent herbivory, and 3) the interactions between surrounding vegetation, focal plants, herbivores and predators. We examined the damage caused by insect herbivores on three native broadleaved species. We then compared the importance of taxonomic similarity between seedlings and canopy tree species and the structure and composition of surrounding vegetation at both large- and small-scales. Birds are vertebrate predators likely to limit damage to plants and facilitate plant growth by consuming herbivorous insects. We estimated their effects by excluding them from focal plants, and compared their interactive effects with vegetation diversity or removal. The effects of surrounding vegetation and avian predation were also disentangled on concealed- vs. external-feeding guilds (predation sensitivity) and specialist- vs. generalist-insects.Surrounding vegetation of focal seedlings influences the abundance of insect herbivores and damage. Specialist insects (leaf miners) are affected by the understorey vegetation close to oak seedlings, decreasing with its presence or structural diversity. The damage caused by generalist insects depend on the large-scale composition of plant communities, and increase with tree species richness and cover or on conspecific forest habitats. Exclusion of insectivorous birds affects insect herbivory in a species-specific manner, and also greatly varies with forest habitats and presence of vegetation around focal plants. The indirect effect of bird predation on leaf damage is observed on seedlings beneath noncongeneric canopy trees. The removal of local surrounding vegetation affects the top-down effect of insectivorous birds on insect herbivores, according to their sensitivity to predation and the accessibility of preys. To conclude, our studies provide experimental evidences of interactive effects between bird predation and neighbouring vegetation on insect herbivores and levels of herbivory. These indirect effects are strongly related to the specialisation of herbivores and to their sensitivity to bird predation

In this study, we have demonstrated that inoculation of bean seeds with non-pathogenic binucleate Rhizoctonia (np-BNR) at sowing protected bean seedlings from infection of R. solani. Using quantitative real-time RT-PCR (QRT-PCR), transcript levels of defense genes encoding 1,3-beta-glucanase (GLUC), phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS) in one-week old bean seedlings was monitored during np-BNR and R. solani interaction. The results revealed that protection effect of np-BNR correspond to a systemic suppression of these three defense genes' expression from significant higher level elicited by R. solani to the level of non-infected plants. This indicates that bio-protection by np-BNR isolates is not correlated to activation of these three defense genes' expression. Similar suppression was achieved for pre-colonization of bean seedlings with arbuscular mycorrhizal (AM) Glomus introradices on GLUC gene expression, although the AM fungus did not significantly reduce rot symptoms. Possible mechanisms implicated in down-regulation during plant-pathogen and np-BNR or AM interaction are discussed.

Thesis (MScAgric)--Stellenbosch University, 2012.<br>ENGLISH ABSTRACT: Botrytis cinerea Pers. Fr. [teleomorph Botryotinia fuckeliana (de Bary) Whetzel] causes serious losses of over 200 crops worldwide, including rooibos seedlings and pears. This pathogen is characterized by morphological, physiological and genetic diversity. The genetic diversity and population structure have not been investigated for B. cinerea populations in South Africa. Botrytis cinerea collected from rooibos seedlings and in pear orchards in the Western Cape of South Africa were investigated in the present study. The study was done with the aid of microsatellite markers, the amplification of mating type alleles MAT1-1 and MAT1-2 and determination of resistance towards various fungicides. Population dynamics was inferred and a similar picture emerged in both production systems. Botrytis cinerea annually causes severe losses of rooibos seedlings (Aspalathus linearis) in nurseries situated in the Clanwilliam region. Sampling was done in five nurseries and the cryptic species status of the isolates obtained was determined through restriction enzyme digestion of the Bc-hch gene. All but one (206 out of 207) of the isolates belonged to Group II or B. cinerea ‘sensu stricto’. Analysis of the B. cinerea Group II population, using seven microsatellite loci, was performed to assess the genetic population structure. Total gene diversity (H) was high, with a mean of 0.67. Two of the nurseries populations’ sample sizes were severely limited after clone correction, yet 100 genotypes were discerned among the 206 isolates genotyped. The percentage of maximal genotypic diversity (G) ranged between 16 and 68 for the five populations, with a total value of 17 for the 100 genotypes. One genotype, represented by 27 clones, was isolated from four nurseries. Relatively low but significant population differentiation was observed in total between nurseries (mean FST = 0.030, P = 0.001). The distribution of mating types MAT1-1 and MAT1-2 differed significantly from the ratio of 1:1 for the total population plus two of the nurseries’ populations. Three nursery populations had an equal mating type distribution. The index of association (IA) analyses suggests that the populations are asexually reproducing. Analysis of molecular variance (AMOVA) indicated that 97% of the total genetic variation is distributed within subpopulations. Fungicide resistance frequency against iprodione for 198 of the genotyped isolates displayed highly varying levels of resistance amongst the five nurseries. The mean total incidence of resistance towards iprodione was 43%, ranging from 0% to 81% for the five nurseries. Baseline sensitivity towards pyrimethanil yielded an average EC50 value of 0.096 mg/L. Botrytis cinerea isolates were collected from pear blossoms (Pyrus communis) in four orchards. Two orchards in the Ceres area and two in the Grabouw area were sampled from. A total of 181 isolates were collected from the four orchards. Incidence of blossom infection in the orchards ranged from 3% to 17%. Overall, there was a high incidence of isolates that had only the Boty transposable element (74%) compared to those harbouring both (Boty and Flipper), simultaneously (transposa, 24%). One isolate examined had the Flipper element only. Cryptic species status according to restriction enzyme digestion of the Bc-hch gene indicated that all the isolates belonged to Group II or B. cinerea ‘sensu stricto’. Analysis of the Group II population, through the use of seven microsatellite loci, was performed to assess the genetic population structure. Total gene diversity (H) was high, with a mean of 0.69 across all populations. Although two of the subpopulations displayed a high clonal proportion, overall 91 genotypes were discerned among the 181 isolates. The percentage of maximal genotypic diversity (G) ranged between 18 and 33 for the four populations, with a total value of 14 for the 91 genotypes. One genotype, represented by 27 clones, was isolated from all orchards. Moderate, but significant population differentiation was present in total among orchards (mean FST = 0.118, P = 0.001). The distribution of the mating types, MAT1-1 and MAT1-2, did not differ significantly from a 1:1 ratio for the total population as well as the subpopulations. Index of association (IA) analyses, on the other hand, suggests that the populations reproduce asexually. Analysis of molecular variance (AMOVA) indicated that 88% of the total genetic variation is distributed within subpopulations, 9% between subpopulations and only 3% between production areas. Fungicide resistance frequency against fenhexamid, iprodione and benomyl varied, with the highest levels of resistance present against benomyl and low levels of resistance seen towards iprodione and fenhexamid. In conclusion, this study has shown that there exist within the studied populations of B. cinerea, obtained from rooibos nurseries and pear orchards, an adaptive capacity to overcome current means of control. The use of population genetics to further our understanding of how plant pathogens interact and spread throughout a given environment is of cardinal importance in aiding the development of sustainable and integrated management strategies. Knowledge of the dispersal of B. cinerea in the two studied cropping systems has shed light on the inherent risk that it poses, and this together with knowledge of the levels of resistance that occurs should serve as an early warning to help divert possible loss of control in future.<br>AFRIKAANSE OPSOMMING: Botrytis cinerea Pers. Fr. [teleomorf Botryotinia fuckeliana (de Bary) Whetzel] veroorsaak ernstige verliese van meer as 200 gewasse wêreldwyd, insluitende rooibossaailinge en pere. Hierdie patogeen word deur morfologiese, fisiologiese, asook genetiese diversiteit gekenmerk. Die genetiese diversiteit en populasie-struktuur van B. cinerea populasies wat in Suid-Afrika voorkom, is nog nie ondersoek nie. Botrytis cinerea verkryg vanaf rooibossaailinge en in peerboorde in die Wes-Kaap van Suid-Afrika is ondersoek. Hierdie studie is met behulp van mikrosatellietmerkers, amplifikasie van die twee paringstipe gene (MAT1-1 en MAT1-2), asook die bepaling van weerstandsvlakke teenoor verskeie swamdoders, uitgevoer. Populasie-dinamika is afgelei en ‘n soortgelyke tendens is in beide produksie-sisteme waargeneem. Botrytis cinerea veroorsaak jaarliks ernstige verliese van rooibossaailinge (Aspalathus linearis) in kwekerye in die Clanwilliam-area. Monsters is in vyf kwekerye versamel en die kriptiese spesiestatus van die verkrygde isolate is deur restriksie-ensiemvertering van die Bc-hch geen bepaal. Almal behalwe een (206 uit 207) isolaat het aan Groep II of B. cinerea ‘sensu stricto’ behoort. Analise van die B. cinerea Groep II populasie, deur middel van sewe mikrosatellietmerkers, is uitgevoer om die genetiese populasiestruktuur te bepaal. Totale geendiversiteit (H) was hoog, met ‘n gemiddelde van 0.67. Alhoewel twee van die kwekerye se monstergrootte erg ingeperk is ná kloonverwydering, is daar nogtans 100 genotipes onder die 206 isolate wat geïsoleer is, waargeneem. Die persentasie van maksimale genotipiese diversiteit (G) het tussen 16 en 68, vir die vyf populasies, gewissel, met ‘n totaal van 17 vir die 100 genotipes. Een genotipe, verteenwoordig deur 27 klone, is uit vier kwekerye geïsoleer. Relatief lae dog noemenswaardige populasie-differensiasie is in totaal tussen kwekerye waargeneem (gem. FST = 0.030, P = 0.001). Die verspreiding van die twee paringstipes (MAT1-1 en MAT1-2) het beduidend verskil van ‘n 1:1 verhouding vir die totale populasie, asook twee van die kwekerye se populasies. Die drie oorblywende kwekerye se populasies het egter ‘n gelyke verdeling van die twee paringstipes getoon. Die indeks van assosiasie (IA) analises toon dat die populasies ongeslagtelik voortplant. Analise van molekulêre variasie (AMOVA) het aangedui dat 97% van die totale genetiese variasie binne die subpopulasies versprei is. Hoogs variërende vlakke van weerstand tussen die vyf kwekerye teenoor die swamdoder iprodioon, is vir die 198 isolate wat getoets is, gevind. Die totale gemiddelde frekwensie van weerstand teenoor iprodioon was 43%, wat tussen 0% en 81% vir die vyf kwekerye gevarieer het. Fondasie-vlak-sensitiwiteit vir pyrimethanil het ‘n gemiddelde EC50 waarde van 0.096 mg/L opgelewer. Botrytis cinerea isolate is ook vanuit peerbloeisels (Pyrus communis L.) vanuit vier boorde versamel, twee uit elk van die Ceres- en Grabouw-areas. In totaal is 181 isolate vanuit die vier boorde versamel. Die frekwensie van bloeiselinfeksie het tussen 3% en 17% gewissel. Oor die algemeen was daar ‘n hoë frekwensie van isolate wat slegs die Boty transponeerbare element teenwoordig gehad het (74%) in vergelyking met dié wat tegelykertyd beide (Boty en Flipper) teenwoordig gehad het. Een isolaat het slegs die Flipper element gehad. Bepaling van die kriptiese spesiestatus met behulp van restriksie-ensiemvertering van die Bc-hch geen het aangedui dat alle versamelde isolate tot Groep II of B. cinerea ‘sensu stricto’ behoort het. Analise van die Groep II populasie, deur middel van sewe mikrosatellietmerkers, is uitgevoer om genetiese populasie-struktuur te bepaal. Totale geendiversiteit (H) was hoog, met ‘n gemiddelde van 0.69 oor alle populasies. Alhoewel twee subpopulasies ‘n hoë klonale fraksie getoon het, is 91 genotipes tussen die 181 isolate wat verkry is, onderskei. Die persentasie van maksimale genotipiese diversiteit (G) het tussen 18 en 33 vir die vier populasies gewissel, met ‘n totale waarde van 14 vir die 91 genotipes. Een genotipe, verteenwoordig deur 27 klone, was in al vier boorde teenwoordig. Gematigde dog beduidende populasie differensiasie was in totaal tussen boorde teenwoordig (gem. FST = 0.118, P = 0.001). Die verspreiding van die paringstipes (MAT1-1 en MAT1-2) het nie betekenisvol van ‘n 1:1 verhouding vir die totale populasie, insluitende die subpopulasies, verskil nie. Indeks van assosiasie (IA) analises het egter aangedui dat die populasies ongeslagtelik voortplant. Analise van molekulêre variasie (AMOVA) het aangedui dat 88% van die totale genetiese variasie in subpopulasies te vinde was, 9% tussen subpopulasies en slegs 3% tussen produksie-areas. Frekwensie van swamdoder weerstandbiedendheid vir fenhexamid, iprodioon en benomyl het gewissel, met die hoogste vlakke teenoor benomyl waargeneem, maar baie lae vlakke teenoor fenhexamid en iprodioon. Samevattend het hierdie studie getoon dat die populasies van B. cinerea wat in hierdie twee produksie-sisteme, op rooibossaailinge en in peer boorde, ondersoek is, ‘n aanpasbaarheid toon om huidige metodes van beheer te oorkom. Die gebruik van populasiegenetika as ‘n hulpmiddel om ons kennis van patogeen-interaksies en -verspreiding te verbreed, is van kardinale belang in die ontwikkeling van geïntegreerde en volhoubare beheermaatreëls. Kennis van die verspreiding van B. cinerea in die bestudeerde gewasproduksiestelsels, werp lig op die inherente risiko wat dié patogeen inhou. Dít, tesame met kennis van die weerstandsvlakke wat voorkom, kan as ‘n vroegtydige waarskuwing dien ten einde moontlike verlies van beheer in die toekoms te help teenwerk.

Control of intracellular ion homeostasis is pivotal to plant salt tolerance. Plants have developed a number of mechanisms to keep ions at appropriate concentrations. Both transporters and channels on the plasma membrane play important roles in this function. Plant cyclic nucleotide-gated channels (CNGCs) in the plasma membrane are non-selective monovalent and divalent cation channels. So far, most studies on plant CNGCs have been conducted on heterologous systems. In planta, reverse genetic studies revealed the role of different CNGCs in cation uptake, transport and homeostasis. However, there is little information available about the functional characteristics of plant CNGCs. Among the 20 members of this protein family in Arabidopsis, only AtCNGC2 has been functionally identified as an ion channel; therefore, more functional characterization needs to be done on other members of this protein family. Several CNGCs were suggested to be involved in K+, Ca2+ and Na+ uptake and transport, but available information is scarce. This study investigated the relationship between CNGC10 and ion transport in Arabidopsis, with a particular emphasis on the involvement of CNGC10 in salt tolerance. Arabidopsis thaliana wild type (WT) and two AtCNGC10 antisense lines (A2 and A3) were used to characterise the impact of different level of salt stress on (i) root growth, ion concentration in tissues, ion fluxes across the root surface and intracellular ion concentration and pH at the seedling stage, and (ii) photosynthesis and ion concentration in tissues at the flowering stage. Plants of both antisense lines had higher K+ and lower Ca2+ and Mg2+ concentrations in shoots than WT plants when grown in non-salt control 1/4 Hoagland solution. Altered K+, Ca2+ and Mg2+ internal concentrations in AtCNGC10 antisense lines compared with WT plants under non-salt conditions indicated disturbed long distance ion transport, especially xylem loading/retrieval and/or phloem loading. The results of ion fluxes across the root surface also suggested that AtCNGC10 might be involved in transport of K+, Ca2+ and Mg2+ in tissue. Under sudden salt exposure, higher Na+ efflux and smaller K+ efflux in both antisense lines suggested that AtCNGC10 channels are involved in Na+ and K+ transport. The shoots of AtCNGC10 antisense lines A2 and A3 contained higher Na+ concentrations and significantly higher Na+/K+ ratios compared to WT, resulting in impaired photosynthesis and increased salt sensitivity in A2 and A3 than in WT plants. In contrast, seedlings of both antisense lines exposed to salt stress had lower shoot Na+/K+ ratios and longer roots than WT seedlings, indicating that A2 and A3 were more salt-tolerant than WT in the seedling stage, likely because growth is less dependent on photosynthesis in the seedling than in the flowering stage. These results suggested CNGC gene might play a different role during different developmental stages and in various plant organs.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior<br>Aiming to characterize the main transgenic soybeans cultivars used in Rio Grande do Sul, experiments were conducted during the 2007/2008 season, in Santa Maria/RS. In the field, the cultivars Relmo Anta RR 82, A 6001 RG, A 8000 RG, FUNDACEP 53 RR, FUNDACEP 54 RR, FUNDACEP 56 RR, BRS 244 RR, and CD 214 RR CD 219 RR, drilled at seeds rates of 250, 400 and 550 thousand seed ha-1 and at row spacing of 0.30, 0.45 and 0.60m. The variables were: grain yield, yield components, distribution of income in different strata of the plant, morphological characteristics, rate of soil covering and amount of light incident on the canopy. The experimental design was a randomized blocks, with 4 repetitions in trifatorial outline (9 cultivars x 3 seed rates x 3 row spacing). In laboratory, tests of peroxidase reaction had been carried through only using the tegument of the seeds or the entire seeds and also seedling color for 50 soybean cultivars. For the grain yield cultivars CD 214 RR and FUNDACEP 53 RR had the best results (3589 and 3530 kg ha-1, respectively). For the distribution of income in the plant strata, there is interaction between density of planting and row spacing, which indicate the trends for each cultivar. For the amount of light incident on the canopy, the interactions between the factors of planting density, row spacing, timing of the cycle of development and level of the canopy, combined with the architecture of each cultivar define the amount of light intercepted, after flowering. The morphological response of each cultivar is different for every situation tested, indicating the need for understanding the behavior of each material, to select the practices to maximize the yield. The use of the test of reaction of peroxidase used the entire seed can be used of so efficient form how much the traditionally used method (tegument). The identification of the seedling color may be an early way to identify varietal blend.<br>Com o objetivo de caracterizar as principais cultivares transgênicas de soja utilizadas no Rio Grande do Sul, foram realizados experimentos durante a safra agrícola 2007/2008, em Santa Maria/RS. Em campo, as cultivares Relmo Anta 82 RR, A 6001 RG, A 8000 RG, FUNDACEP 53 RR, FUNDACEP 54 RR, FUNDACEP 56 RR, BRS 244 RR, CD 214 RR e CD 219 RR foram submetidas às densidade de semeadura de 250, 400 e 550 mil semente ha-1 e aos espaçamentos entre linhas de 0,30, 0,45 e 0,60m. As variáveis avaliadas foram: rendimento de grãos, componentes do rendimento, distribuição do rendimento em diferentes estratos da planta, características morfológicas, taxa de cobertura do solo e quantidade de luz incidente no dossel. O delineamento experimental utilizado foi o de blocos ao acaso, com 4 repetições, no esquema trifatorial (9 cultivares x 3 densidades de semeadura x 3 espaçamentos entre linhas). Em laboratório, foram realizados testes de reação da peroxidase utilizando somente o tegumento das sementes ou as sementes inteiras e também coloração do hipocótilo para 50 cultivares de soja. Para o rendimento de grãos as cultivares CD 214 RR e FUNDACEP 53 RR apresentaram os melhores resultados (3589 e 3530 kg ha-1, respectivamente). Para a distribuição do rendimento nos estratos da planta, existe interação entre densidade de semeadura e espaçamento entre linhas, os quais indicam as tendências para cada cultivar. Para a quantidade de luz incidente no dossel, as interações entre os fatores densidade de semeadura, espaçamento entre linhas, época do ciclo de desenvolvimento e nível do dossel, aliados à arquitetura de cada cultivar definem a quantidade de luz interceptada, após o florescimento. A resposta morfológica de cada cultivar é diferenciada para todas as situações testadas, indicando a necessidade do conhecimento do comportamento de cada material, para a escolha das práticas que visem maximizar o rendimento de grãos. O uso do teste de reação da peroxidase utilizando a semente inteira pode ser utilizado de forma tão eficiente quanto o método tradicionalmente utilizado (tegumento). A identificação da coloração do hipocótilo pode ser uma forma precoce de identificar mistura varietal.

Cephalosporium stripe of wheat (Triticum aestivum), caused by the soilborne fungus Cephalosporium gramineum, results in significant yield reductions in dryland winter wheat crops of the U.S. Pacific Northwest. The development of resistant cultivars offers the best hope for disease control. Breeding for resistance is hampered by the long trial times inherent in screening adult plants, and by cultivar x environment interactions in field tests. The principal objective of this research was to develop and test a procedure for screening wheat seedlings in controlled environments for resistance to Cephalosporium stripe. Wheat seedlings were raised hydroponically in growth chambers, and the fungus was increased in large fermentation tanks. The seedlings were inoculated at about 12 days post-germination. Disease severity was assessed approximately seven days later using a chlorophyll meter to measure the symptoms of chlorosis and striping. In three trials, five soft white cultivars from the Pacific Northwest and four hard red cultivars from the Southern Great Plains with known levels of field resistance were tested with a Pacific Northwest fungal isolate. With one exception, chlorophyll readings ordered the cultivars appropriately, with moderately resistant cultivars ranking above susceptible cultivars. Three other moderately resistant cultivars from the Pacific Northwest also appeared in one or two trials, and were ranked properly by chlorophyll level. Chlorophyll levels of uninoculated plants were assayed to determine if differences in chlorophyll content were innate in the cultivars. The chlorophyll levels of uninoculated and inoculated seedling treatments were only significantly correlated when the cultivar Madsen, which ranks high both in resistance and in chlorophyll content, was included. In adult plants, flag-leaf chlorophyll level corresponded to intensity of Cephalosporium stripe symptoms where disease was present, and was independent of known field resistance in undiseased cultivars. The seedling screening technique was used to investigate pathogenic variability in C. gramineum. In two experiments, a total of eight cultivars from the Pacific Northwest and the Southern Great Plains were tested with three fungal isolates from each region. No evidence of virulence/vertical resistance was found. There was also no significant adaptation of isolates to greater virulence on cultivars from the same region.<br>Graduation date: 1998

Cotton seedling disease complex is caused by a number of pathogens inducing similar symptoms and can lead to thin, uneven stands that grow slowly and yield poorly. Preliminary work indicated that a set of high tannin cotton lines developed and released in 1989 by Texas A&M AgriLife Research, (Smith et al., 1990a, Smith et al., 1990b, Schuster et al., 1990) may possess increased resistance to disease. This evidence, along with additional studies that show a clear role of tannin in disease resistance, suggest that these high tannin cotton lines may prove useful in breeding for increased resistance to cotton seedling disease complex. High tannin cotton lines were screened for their resistance to Rhizoctonia solani and Pythium aphanidermatum. While no high tannin germplasm line was more resistant to R. solani than the resistant control, Tamcot SP 21, the potential for significant gains from selection was demonstrated. Fifteen high tannin lines expressed resistance to P. aphanidermatum equal to the resistant control, Tamcot Sphinx (El-Zik and Thaxton, 1996). This resistance was not shown to be correlated with tannin content, though it is still unclear whether or not this lack of correlation is real or due to limitations in measuring tannin in infected seedlings.

碩士<br>國立中興大學<br>農藝學系<br>88<br>Bentazon is usually used in weed control for rice and peanut crops. If the growth of rice, cv. TNG67, seedlings was affected and how the physiological response was influenced by bentazon, especially for the resistant- and the susceptible-lines among Japonica- and Indica-type rice, were investigated. According to the dose-response to bentazon of five growth characters including plant height, shoot fresh weight, total chlorophyll, chlorophyll a and chlorophyll b of TNG67 seedlings at the V3 stage, the effective concentration of leaf-applied bentazon causing 50% reduction of response (EC50) was determined. And further study on the physiological resistant mechanism existed in bentazon-resistant and -susceptible lines of rice under this concentration of bentazon was conducted. As a result of screening the bentazon-resistant and —susceptible lines of rice, lines M202 and FSK are referred to resistant- and susceptible-line, respectively, among Japonica-type rice, as well as the TCSW1 and IR36 are referred to resistant- and susceptible-line among Indica-type, respectively. The bentazon-resistant line, M202, and TNG67 were suffered from little or a slight visible injury, photosynthesis inhibition and accumulation of MDA after bentazon application compared to other lines. Further analysis of the physiological mechanism of resistance for the bentazon-resistant and -susceptible lines among Japonica- and Indica-type rice was conducted. Experimental results indicate that the resistant line of Japonica-type, M202 and TNG67, absorbed more 14C-bentazon, but most of 14C-isotopes absorbed either accumulated in bentazon-treated leaf or translocated out of the treated leaf to older leaves subsequently. It is also found that the phytotoxicity resulted from residual bentazon and its metabolite was slight in these two rice lines. The hydroxybentazon (bentazon-6-OH) could be detected in the bentazon-resistant rice lines, M202, but not in other rice lines, indicating that M202 and TNG67, especially for the former line had stronger ability to metabolite more bentazon. For the bentazon-susceptible lines, FSK and IR36, which absorbed less 14C-bentazon during treatment but most of the 14C-isotopes absorbed translocated out of treated leaf and accumulated in young leaves subsequently. More accumulation of bentazon at the younger leaf might result in death of rice seedlings. In addition, more residual bentazon and stronger phytotoxicity were also confirmed in these two bentazon-susceptible lines. And less polar metabolite was obtained. This result indicates that the bentazon-sensitivity of FSK and IR36 were due mainly to the lower ability of bentazon metabolism. Reviewing the experimental results of this study, it suggested that the bentazon-resistant mechanism existed in resistant lines, M202 and TNG67, is primarily result from the stronger metabolism of bentazon, especially the generation of bentazon-6-OH for the line M202. Besides, that the most of absorbed bentazon translocated to older leaves and/or deposited at the site of bentazon application also involved in the resistance mechanism.

Thesis (M.Sc. (Horticulture)) -- University of Limpopo, 2017.<br>Root-knot (Meloidogyne species) nematodes are economically destructive pathogens of over 3000 species, whereas others have resistance to Meloidogyne species. Wild watermelon (Cucumis africanus) and wild cucumber (Cucumis myriocarpus) are highly resistant to Meloidogyne species, particularly M. incognita and M. javanica. The two Cucumis species are used in inter-generic grafting with watermelon (Citrullus lanatus) as nematode resistant rootstocks. Also, the two Cucumis species are used in traditional medicine and in plant-parasitic nematode management as phytonematicides. The form of nematode resistance, which is essential in plant breeding, is not documented for the two Cucumis species. The objective of this study was to determine the form of nematode resistance in the two Cucumis species to M. incognita and M. javanica under greenhouse conditions. Four parallel experiments were each conducted under greenhouse conditions. Uniform six-week old Cucumis seedlings were transplanted into 250 ml polystyrene cups filled with 200 ml growing medium of steam-pasteurised fine sand. A week after transplanting, Cucumis seedlings were each infested by dispensing approximately 100 M. incognita second-stage juveniles (J2) or M. javanica J2 using a 20 ml plastic syringe by placing into 5-cm-deep furrow around the seedling stem and covered with growing medium. Treatments (periodic harvest intervals) were arranged in a randomised complete block design, replicated five times. Five seedlings from each experiment were harvested every second day, for 30 days, with stained roots being assessed for necrotic spot (suberised cells) number, giant cell number, proliferation of rootlet interference number and root gall number. Periodic harvest intervals were highly significant (P ≤ 0.01) on necrotic spot number, proliferation of rootlet interference number and root gall number in C. africanus-M. incognita relations, but were not significant for giant cell number. Treatments contributed 59, 64 and 50% in total treatment variation (TTV) of necrotic spot number, proliferation of rootlet interference number and root gall number, respectively. Harvest period had highly significant effects on necrotic spot number, giant cell number, proliferation of rootlet interference number and root gall number in C. africanus-M. javanica relations. Treatments contributed 55, 71, 63 and 59% in TTV of necrotic spot number, giant cell number, proliferation of rootlet interference number and root gall number, respectively. Periodic harvest intervals were significant (P ≤ 0.05) on giant cell number and highly significant on root gall number in C. myriocarpus-M. incognita relations. However, there were no significant treatment differences on necrotic spot number and proliferation of rootlet interference number. Treatments contributed 57 and 57% in TTV of root gall number and giant cell number, respectively. Harvest period had highly significant effects on giant cell number, proliferation of rootlet interference number and root gall number, but were not significant on necrotic spot number in C. myriocarpus-M. javanica relations. Treatments accounted for 67, 49 and 53% in TTV of giant cell number, proliferation of rootlet interference number and root gall number, respectively. In conclusion, the mechanism of resistance to M. incognita and M. javanica in both C. africanus and C. myriocarpus was post-infectional nematode resistance, which has attributes for introgression into commercial nematode-susceptible Cucumis cultivars.<br>Agricultural Research Council (ARC), National Research Foundation of South Africa; and the ARC-Universities Collaboration Centre for Smallholder Farmers

Identification of salt- resistant tree species and genotypes is needed for rehabilitation of lands affected by salinity in Mexico. This dissertation consists of four studies for analyzing the responses of Pinus leiophylla seedlings to salt. In the first study, resistance to salt stress was studied in six-month-old seedlings from eight different sources of seed collected from the areas with contrasting precipitation levels. Plants from the xeric areas were shorter and had smaller stem diameters but were less sensitive to salt stress and recovered faster from salt injury compared with the plants from mesic sites, suggesting that morphological and physiological adaptations to drought were helpful with salt stress resistance. In the second study, fascicle needle production and tissue ion accumulation were examined in NaCl-treated three-month-old seedlings from two populations of the xeric origin and two populations from the mesic areas. Seedlings from the xeric population of San Felipe developed fewer fascicles and had shorter needles compared with seedlings from the remaining three populations. NaCl treatment delayed the emergence of fascicles and reduced the fascicle needle production and needle length. However, the extent of needle injury and ion accumulation in shoots were lower in the San Felipe seedlings compared with the other studied populations. In the third study, the effects of branch pruning and seedling size on total transpiration and accumulation of Na+ and Cl- in tissues were examined. Total plant transpiration, as affected by plant size and branch pruning, was correlated with Na+ and Cl- needle concentrations and needle necrosis. Branch pruning reduced ion accumulation in the shoots and needle necrosis levels in short seedlings but not in the tall seedlings. In the fourth study, sprouting and physiological responses of 16 month-old-seedling to salt were examined. NaCl treatment concentrations of 100, 150, and 200 mM reduced gas exchange and root hydraulic conductance, caused needle injury and triggered sprouting of adventitious shoots. Sprouting from the upper parts of the main stem and lateral branches was three times greater with 100 and 150 mM NaCl compared with 200 mM NaCl treatment but, at the base of the stem, sprout numbers were similar for all NaCl treatments.

Two aspects of drought resistance were investigated on wet and dry ecotypes of three conifer species: 1) the relative importance of drought avoidance and drought tolerance mechanisms in resisting drought stress was assessed on Douglas-fir (Pseudotsuga menzieseii) and lodgepole pine (Pinus contorta) seedlings, and 2) the effects of drought on root hydraulic conductance and low temperature, on root water flow rates Were assessed on first-year seedlings of Douglas-fir, lodgepole pine and white spruce (Picea glauca). To study drought avoidance, Douglas-fir and lodgepole pine seedlings were grown in sealed containers in wet (522% water content) or dry (318% water content) peat/vermiculite soil in a factorial treatment design. Dry weights, water use, and root length were determined for seedlings at each of five harvests and stomatal conductance and shoot water potentials were measured during the last 12 weeks of the experiment. Lodgepole pine seedlings had greater dry matter production, water use, stomatal conductance and new root length than Douglas-fir seedlings. New root weight of lodgepole pine seedlings exceeded that of Douglas-fir seedlings during the last five weeks of the experiment, and specific root length of new roots was higher for lodgepole pine seedlings throughout the experiment. Douglas-fir seedlings showed higher water use efficiency (WUE) than lodgepole pine seedlings, although water uptake rates per unit of root dry weight showed little difference between species. Soil water treatment influenced specific root length of new roots, water uptake per unit of new root length, and WUE in Douglas-fir seedlings more than in lodgepole pine seedlings. To study drought tolerance, Douglas-fir and lodgepole pine seedlings were grown under drought and well-watered conditions. At each of three harvests a pressure-volume curve was produced for each seedling. Douglas-fir maintained a lower osmotic potential at full saturation [special characters omitted] and lower turgor loss point [special characters omitted] than lodgepole pine under both watering regimes,. Both species had lower [special characters omitted] when drought-stressed. Douglas-fir appears to be a more conservative species, maintaining low stomatal conductance and tolerating drought conditions, whereas lodgepole pine avoids drought by producing large amounts of roots to exploit the soil resource. To study root hydraulic conductance (Lproot) and water flow rates through roots (WFRR), water flow was measured through de-topped roots of Douglas-fir, lodgepole pine, and white spruce seedlings in a pressure chamber. In a drought experiment, seedlings were grown in sandy soil in a greenhouse under drought and well-watered conditions during their first growing season and, in a low temperature experiment, seedlings were grown in sandy soil in growth chambers at 25/20°C (day/night) and 15/10°C, In the drought experiment, water flow through roots was measured at three pressures. No differences in Lproot were found for Douglas-fir and white spruce seedlings grown under the two watering regimes, however, lodgepole pine seedlings had reduced Lproot when grown under drought conditions. Welk watered seedlings of lodgepole pine and white spruce had higher Lpr00t in 1989 than in 1990 whereas Douglas-fir seedlings had the same Lproot in both years. In the low temperature experiment, WFRR was measured at 1.0 MPa and temperatures of 20°C for 24 hours or 20°, 12°, and 4°C for 18, 15, and 15 hours respectively. At 20°C, white spruce seedlings had higher WFRR than the other two species. Lodgepole pine and white spruce seedlings grown in the 1S°/10°C growth chamber had higher WFRR than seedlings grown in the 25°/20°C growth chamber. Water flow rate decreased with temperature in all three species. After correcting for viscosity, all seedlings had lower WFRR with reduced temperature, except for Douglas-fir and white spruce seedlings grown at 15°/10°C which had the same WFRR at 20°C and 12°C. Therefore, Douglas-fir and white spruce seedlings were found to become less sensitive to low temperature (chilling) stress when pre-conditioned at low temperatures. In the drought and low temperature studies, dry weight biomass of white spruce was lowest but white spruce had a greater specific root length than lodgepole pine and Douglas-fir. In the drought study, biomass production in seedlings from wet ecotypes of each species was more reduced when drought-stressed than seedlings from dry ecotypes.<br>Graduate

Ph.D.<br>In plants, salicylic acid (SA) is a signaling molecule that regulates disease resistance responses such as systemic acquired resistance (SAR) and the hypersensitive response (HR), and has been implicated in both basal and acquired thermotolerance. It has also been shown that SA enhances heat-induced Hsp/Hsc70 accumulation in plants. In this study, temperature studies revealed that heat shock (HS) at 40 °C for 30 min significantly induced Hsp/Hsc70 accumulation in 3-week old tomato (UC82B) seedlings. Time- and dose-responsive studies showed that 0.1 mM SA for 17 hrs was unable to induce Hsp/Hsc70 but in combination with HS significantly (P > 0.001) potentiated this response. To investigate the mechanism of SA-mediated, heat-induced Hsp/Hsc70 potentiation, tomato seedlings were treated with either SA alone, HS or both, before analyses of hsp70 mRNA, Hsf DNA-binding and gene expression of hsp70, hsfAl, hsfA2 and hsfEll. SA alone established Hsf DNA-binding, but was not accompanied by increased Hsp70 accumulation or expression of hsp70 mRNA. SA had no significant effect on hsfA2 and hsf81 gene expression, but increased the basal levels of hsfAl. In heat-shocked plants, Hsf DNA-binding was enhanced, and increased hsfAl, hsfA2 and hsfB1 expression preceded accumulation of Hsp70. The combined treatment of SA and HS resulted in potentiated Hsf DNA-binding, enhanced expression of hsp70, hsfAl, hsfA2 and hsfB1, leading to potentiated levels of Hsp/Hsc70. Since increased hsp70 and hsf gene expression coincided with increased levels of Hsp70 accumulation, it is likely that the SA-mediated potentiation of Hsp70 is due to the ability of SA to regulate Hsfs during HS. This study therefore proposes a mechanism for the potentiation of Hsp70 by SA in the presence of heat, which might contribute to our understanding of the role SA plays in the heat shock response and thermotolerance.

碩士<br>國立臺灣大學<br>園藝學研究所<br>100<br>Myxosporium wilt, caused by the pathogen Nalanthamala psidii (Sawada et Kurosawa) Schroers & M. J. Wingf. (≡Myxosporium psidii Sawada et Kurosawa), belongs to one of systemical fungal diseases. This disease has been the most severe and destructive disease of guava (Psidium guajava) cultivation in Taiwan since the 1970s due to the development of pruning techniques for nonseasonal production. The pruning techniques result in numerous wounded sites and increase chances for the invasion of this pathogen. No chemical could prevent or cure this disease and no other solution is available except to burn and remove infected trees from the orchards immediately as the symptoms ocurred. Therefore, breeding for disease resistance is requied. The objectives for the present studies were to: (1) study the effectiveness of inoculation methods for guava seedlings; (2) screening seedlings for N. psidii resistance; (3) evaluate the pathogenicity of N. psidii culture filtrate to seedlings. In the present study, pitiching and root trimming severely injured seedlings and caused a lot portion of seedlings died. Therefore, the effectiveness of the two inoculation methods could not be compared; and thus, the effects of wounding method, seedling stage and inoculum concentration on inoculation of N. psidii could not be determined. Among 2760 seedlings we screened after 3 inoculations within 2 years, 132 puataive resistant plants were obtained. Half of them were ‘Pei-pa’ open-pollinated (OP) progenies. After wounding branches of ‘Pei-pa’ hybrid and OP seedlings of other lines were inoculated with agar plugs of N. psidii Mp-02 isolate, wilting symptoms of most plants only occurred above the wounds. The observation period, 2 months, was not long enough to determine their resistance/susceptibility. However, pathogenicity of N. psidii culture filtrate to guava seedlings could not be confirmed.

碩士<br>國立中興大學<br>植物病理學系所<br>98<br>Twelve isolates of Alternaria spp. were isolated from various crops in Taiwan. Those fungi were respectively grown in liquid culture media for 5-7 days and harvested their mycelia. The mycelial samples were pulverized in liquid nitrogen, and then homogenized with precooled Tris-buffer, estimated the quantity of their total protein by the Bradford method, bovine serum albumin (BSA) solution was used as a standard, the absorbance value equal to 2 µg BSA/ml. Spraying the mycelial extracts obtained from Alternaria spp. on Chinese cabbage seedlings inoculated with Rhizoctonia solani AG-4, APR01-TP and ACO01-TP, were able to reduce 33.33% disease incidence of Rhizoctonia damping-off of Chinese cabbage seedlings. Dipping germinated Chinese cabbage seeds with each of mycelial extracts obtained from 12 isolates of Alternaria spp., ATA01-TP, ABR03-TP, ALY01-TP, AOR01-TP and APR01-TP was respctively able to promote the extension of Chinese cabbage roots as well as to enhance development of their lateral roots. Especially, the mycelial extract of APR01 was more effective in stimulating the growth of plants at 36.56%. APR01 was identified as Alternaria tenuissima according to its conidial morphology, conidigenous cells, and ITS sequencing. In greenhouse tests, spraying the mycelial extract from A. tenuissima APR01 once or thrice on Chinese cabbage seedlings grown on soil infested with R. solani AG-4 RST-04, the fresh weight of Chinese cabbage seedlings were heavier up to 38.3% and 48% compared to untreated control. Inoculating mycelial discs of R. solani AG-4 RST-04 on Chinese cabbage leaves after sparying mycelial extract of A. tenuissima APR01, the expansion of the lesion size were slower compared to the control. It was proved that the mycelial extract of A. tenuissima APR01 could not inhibit mycelial growth of R. solani AG-4 RST-04. To estimate the activity of phenylalanine ammonia lyase (PAL) and peroxidase (POD) in Chinese cabbage seedlings after spraying the mycelial extract of A. tenuissima APR01 and inoculating R. solani AG-4 RST-04, the enzyme activity of the PAL and POD in treated plants were higher than the control. These results suggested that there were some elements existed in mycelial extract of isolate APR01 were able to induce Chinese cabbage seedlings resistant against the pathogen. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was performed for mycelial extract of A. tenuissima APR01 and the bands were displayed on polyacrylamide gel stained with CBR. Mycelial extract of A. tenuissima APR01 was heated at 50 ℃ water bath and lost its biological activity. The effective component of the mycelial extract of A. tenuissima APR01 was not ultra-filtrated in the membrane tubing with molecular weight cut-off of 10,000. Therefore, the major activity of mycelial extract of A. tenuissima APR01 is a protein with > 10 kilodalton of molecular weight.

Plants and microorganisms interact with each other constantly, with some interactions being mutually beneficial and others being detrimental to the plants. The features of the organisms involved in such interactions will determine the characteristics of individual pathosystems. Plants respond readily to pathogen attacks, regardless of the pathosystem; furthermore, variation in the resistance to pathogens within species is common and well documented in many plant species. The variability in pathogen resistance is at the core of genetic improvement programs for disease resistance. True resistance to pathogens in plants is a genetically determined and complex trait that can involve both constitutive and induced mechanisms at different levels of organization. The complexity of this phenomenon makes the study of compatible plant - pathogen interactions challenging, and typically, disease resistance studies focus on specific aspects of a pathosystem, such as field resistance, anatomical or physiological features of resistant plants, or molecular mechanisms of resistance. The Thuja sp. - Didymascella thujina (E.J. Durand) Maire interaction is an important pathosystem in western North America, which has been studied for more than five decades. Western redcedar (Thuja plicata Donn ex D. Don) is very susceptible to cedar leaf blight (D. thujina), a biotroph that affects the tree at all stages, although seedlings are the most sensitive to the pathogen. The characteristics of the Thuja sp. - D. thujina interaction, the wealth of information on the pathosystem and the excellent Thuja sp. genetic resources available from the British Columbia Ministry of Forests, Lands, Natural Resource Operations and Rural Development make this interaction an ideal system to advance the study of disease resistance mechanisms in conifers. This Doctoral project presents a comprehensive investigation of the constitutive and induced resistance mechanisms against D. thujina in T. plicata, Thuja standishii (Gord.) Carrière and a Thuja standishii x plicata hybrid at the phenotypic and gene expression levels, undertaken with the objective of exploring the resistance mechanisms against the biotroph in these conifers. The project also aimed to establish base knowledge for the future development of markers for marker-assisted breeding of T. plicata. The investigations included a combination of histological, chemical and next generation sequencing (NGS) methodologies. NGS data were analyzed, in addition to the traditional clustering analyses, with cutting edge machine learning methods, including grade of membership analysis, dynamic topic modelling and stability selection analysis. The studies were progressively more controlled to narrow the focus on the resistance mechanisms to D. thujina in Thuja sp. Histological characteristics related to D. thujina resistance in Thuja sp. were studied first, along with the relationship between climate of origin and disease resistance. The virulence of D. thujina was also documented early in this project. Chemical and gene expression constitutive and induced responses to D. thujina infection in T. plicata seedlings were studied next. T. plicata clonal lines were then comprehensively studied to shed light on the mechanisms behind known physiologically determined resistance. A holistic investigation of the resistance mechanisms to D. thujina in T. standishii, T. plicata and a T. standishii x plicata hybrid explored the possibility of a gene-for-gene resistance model. Thirty-five T. plicata families were screened during the four field seasons carried out between 2012 and 2015, totalling more than 1,400 seedlings scored for D. thujina severity. Thirteen of those families were used in the five studies performed during the program, along with two T. plicata seedling lines self-pollinated for five generations and three T. plicata clonal lines. One T. standishii clonal line, and one T. standishii x plicata clone were also investigated during the program. A total of 16 histological and anatomical characteristics were studied in more than 750 samples, and more than 270 foliar samples were analyzed for 60 chemical and nutritional compounds. Almost one million transcriptomic sequences in four individually assembled reference transcriptomes were examined during the program. The results of the project support the variability in the resistance to D. thujina in T. plicata, as well as the higher resistance to the pathogen in plants originating from cooler and wetter environments. The data collected also depicted the existence of age-related resistance in T. plicata, and confirmed the full resistance to the disease in T. standishii. Western redcedar plants resistant and susceptible to D. thujina showed constitutive differences at the phenotypic and gene expression levels. Resistant T. plicata seedlings had thicker cuticles, constitutively higher concentrations of sabinene, alpha-thujene, and higher levels of expression of NBS-LRR disease resistance proteins. Resistant clones of T. plicata and T. standishii had higher expression levels of bark storage proteins and of dirigent proteins. Plants from all ages, species and resistance classes studied that were infected with D. thujina showed the accumulation of aluminum in the foliage, and increased levels of sequences involved in cell wall reinforcement. Additional responses to D. thujina infection in T. plicata seedlings included the downregulation of some secondary metabolic pathways, whereas pathogenesis-related proteins were upregulated in clonal lines of T. plicata. The comprehensive approach used here to study the Thuja sp. - D. thujina pathosystem could be applied to other compatible plant-pathogen interactions.<br>Graduate<br>2020-08-31