Dissertations / Theses on the topic 'Pyrethrum'

O desenvolvimento da agricultura, a modernização dos meios dos cultivos, buscando atender a um mercado exigente, tem-se baseado no uso de insumos agrícolas para garantir a produção de alta qualidade e de quantidade. Tais culturas demandam o uso de pesticidas, cujos resíduos, principalmente em hortaliças e frutas, são motivo de preocupação com a saúde dos consumidores e de operários que necessitam retornar às culturas tratadas com esses agrotóxicos. Os objetivos deste estudo foram: a) estudar o comportamento dos resíduos de deltametrina, de diferentes formulações, em frutos, folhas e solo; b) avaliar os resíduos deslocáveis do inseticida nas folhas, como ponto de partida para estudos de exposição ocupacional; c) correlacionar os teores de deltametrina nos frutos com o controle da broca-das-cucurbitáceas e d) correlacionar os resíduos nos frutos com o limite máximo de resíduo (LMR) e o intervalo de segurança (período de carência) estabelecidos pela legislação. Os tratamentos foram: a) testemunha; b) três aplicações da formulação em concentrado emulsionável Decis 25 CE na dosagem de 30 mL p.c.100 L-1 de água (0,75 g i.a. deltametrina.100 L-1 de água); c) três aplicações da formulação em concentrado emulsionável Decis Ultra 100 CE na dosagem de 7,5 mL p.c.100 L-1 de água (0,75 g i.a. deltametrina.100 L-1 de água); e d) três aplicações da formulação em suspensão concentrada Decis 200 SC na dosagem de 3,75 mL de p.c.100 L-1 de água (0,75 g i.a. deltametrina 100 L-1 de água). As amostras de fruto e de solo foram colhidas a (-1); zero; 1; 3; 5 e 7 dias após a última aplicação; as amostras de folhas e as de para estudos de resíduos deslocáveis nestas foram tomadas a (-1); zero; 1; 3; 5; 7 e 14 dias. O método analítico constou da extração dos resíduos de deltametrina de acetato de etila, limpeza dos extratos por técnica de cromatografia de permeação em gel (GPC), com eluição feita com uma mistura de acetato de etila/ciclohexano. A determinação quantitativa por cromatografia de gás. Para a avaliação da eficiência do inseticida, foram feitas seis levantamentos de infestação, determinando-se a porcentagem de frutos brocados. Os resultados indicaram que os resíduos de deltametrina no fruto, embora em baixos níveis, encontravam-se acima do LMR (0,03 mg.kg-1), mesmo um dia após o término do intervalo de segurança (2 dias) para as formulações do inseticida. Nas folhas, os resíduos resultantes das aplicações da formulação SC foram sempre maiores do que ambas CE (10-20 vezes), sendo de 15-80 vezes mais altas do que nos frutos. No solo, os resíduos foram também baixos (0,01-0,05 mg.kg-1), e semelhantes nas três formulações, entretanto, com baixa dissipação. Os resíduos deslocáveis de deltametrina nas folhas foram muito elevados quando de aplicações em SC, comparadas com as formulações CE (4-20 vezes), com valores de meias-vida de 2,8; 3,8 e 32,2 dias, respectivamente para as formulações Decis 25 CE, Decis Ultra 100 CE e Decis 200 SC. O inseticida, nas formulações em que foi aplicado, foi eficiente no controle da praga durante todo o período de avaliação.<br>The agriculture development, the modernization of cultivation means, trying to meet a demanding market, has been based on the use of agricultural supplies to assure a high quality and quantity production. Such crops demand the use of pesticides, whose residues, especially in fruit and vegetable production, raise worries about the consumers’ health and of the workers who need to return to the treated crop with these pesticides. The objectives of this study were: a) to study the behavior of deltamethrin residues applied in different formulations in fruits, leaves and soil; b) to evaluate the dislodgeable residues of the insecticide in leaves, as a starting point for occupational exposure studies; c) to correlate deltamethrin concentration in fruit with the pickleworm control and d) to correlate residues in fruit with the maximum residue level (MRL) and the safety interval established by the legislation. The treatments were: a) control ; b) three applications of the emulsifiable concentrate formulation Decis 25 CE.100-1 at the dosage of 30 mL of c.p.100 L-1 water (0.75 g a.i. deltamethrin.100L-1 water); c) three applications of the emulsifiable formulation of Decis Ultra 100 CE at the dosage of 7.5 mL of p.c.100 L-1 water (0.75 g a.i. deltamethrin.100. L-1 water) and d) three applications of the concentrate suspension formulation Decis 200 SC at the dosage of 3.75 mL of c.p.100.L-1 water (0.75 g a.i. deltamethrin 100L-1 water). The fruit and soil samples were taken at (-1); zero; 1; 3; 5 and 7 days after the last application; leaf samples and also those for the studies of dislodgeable residues on them were collected at (-1); zero; 1; 3; 5; 7 and 14 days after the last application. The analytical method consisted of the extraction of deltamethrin residues in ethyl acetate, clean-up of the extracts by gel permeation chromatography (GPC), with elutition made with a mixture of ethyl acetate/cyclohexane. The quantitative determination was made by gas chromatograph. To evaluate the insecticide efficacy six infestation surveys were carried out, based on the percentage of infested fruit . The results indicate that the deltamethrin residues in fruit, though in low levels, were found higher than the MRL (0.03 mg.kg-1), for the three insecticide formulations, even a day after the end of the safety interval (2 days). In the leaves the resulting residues of the applications of SC formulation were always bigger than both CE (10-20 times), being 15-80 times higher in the fruits. In the soil, the residues were also low (0.01-0.05 mg kg-1), and similar in the three formulations, however with low dissipation. The dislodgeable residues of deltamethrin in leaves were higher in SC as compared to both CE (4-20 times), with half-life values of 2.8; 3.8 and 32.2 days, respectively for the formulation Decis 25 CE, Decis Ultra 100 CE and Decis 200 SC. The insecticide in the formulations applied was efficient in the pest control during the whole evaluation period.

Drying technology is being more and more used in chemical industry, food industry, processing agricultural products, and fibber industry. In the majority of processing industries, drying is carried out for one or more reasons: 1)To offer ease dealing in further processing. For example; drying sweet chrysanthemum in order to increase sweet agents. It can be instead of the cane sugar. The sweet taste is nearly 200 times of that of the cane sugar. Generally, after picking up the fresh leaves of the sweet chrysanthemum, it is necessary to send them to drying equipment at the temperature about 80 C° ---100 C°. With the help of the drying their fresh leaves, their volume will be reduced to one of seventh or one of eighth of fresh leaves volume. Then it is through chemical processing them to get sweet ingredients. Finally, these sweet ingredients are dried at the temperature nearly 140 C° to get the final white products. 2) To supply the final product with satisfactory moisture, such as tea, tobacco. The moisture of fresh tea leaves is about 70%. After drying the final moisture of tea is about 6 %. 3) To preserve the products during storage. For example, drying rice, corn, and other agricultural products are to impede the enzyme to oxidation of these agricultural products, because the final lower moisture of the dried agricultural can limit the enzyme to grow. 4) To avoid the presence of the moisture which may lead to corrosion as in drying some metal products. The drying process includes moisture migration and evaporation. The heat and mass transfer are always key controls for any drying processes in designing the dryer. The drying energy efficiency and drying cost should be considered firstly. Currently high density industrial drying technology is developing quickly based on a deeper understanding of the drying process and the character of wet objects. The purpose of the paper is to analyse the drying process, including heat and mass transfer, moisture movement and moisture evaporation, and the air flow situation in the drying chamber through discussing the practical drying pyrethrum. A moisture moving model in capillaries is set up to study the moisture movement inside the pyrethrum, which will influence the drying rate because the moisture in the wet pyrethrum exists mainly in the capillaries. A mathematical model of Heat and Mass transfer has been developed to explaining the drying principle and to design a suitable dryer for drying pyrethrum. Psychrometric chart and other charts are introduced to understand the change of warm air conditions during drying pyrethrum. The drying curve predicts the moisture changes in the wet materials and the characteristics of the dryer. Analysis of the air flow patterns in the air duct leads to improvement in the air velocity distribution in the drying chamber. After studying the whole drying theory, the forced convection multiple continuous dryer is recommended for drying pyrethrum. In this dryer, the heat energy necessary for moisture evaporation is supplied through the convection of warm air. The vapour is carried away by the warm air. The continuous drying will result in higher drying efficiency. This dryer offers some outstanding features as follows: (1) This dryer offers a larger output for a given floor area. (2) The drying conditions can be easily controlled. (3) Higher heat efficiency can be obtained in this dryer due to the continuous drying process. (4) Automatic loading and discharging save some labour costs. (5) Compared with other dryers, lower operation costs and lower manufacture costs are obtained in selecting this dryer. High density drying should produce the top quality dried products at the lowest cost. In drying plants or agricultural products, the top drying quality has some specification as follows. (1) The final moisture contents of the dried products should be in equilibrium with that of the air in which it is to be stored so that further changes in moisture are small. This conditions is called the equilibrium moisture content. For most organic plants, the EMC is in the ranges 9-15% of oven dry weight. (2) The oxidation of ingredients should be lower than 15% by weight. (3) The broken volume should be lower than 15% by the total volume. (4) Odour and colour should be maintained. For example drying tobacco, the colour should be the golden yellow. For drying some vegetables, the natural green colour should be kept during the drying process. It is hoped that this study may lead to improved drying technology in drying Chinese tea, tobacco, vegetables, mushrooms, agricultural products and food.

Pyrethrum oil sourced from Tanacetum cinerariifolium is the main commercial source of the natural pesticide, pyrethrum oil with the active components being pyrethrin esters. Pyrethrum oil also contains sesquiterpene lactones (STLs) with pyrethrosin being the major STL. Following the refining process, pyrethrins are diluted from 80% to 53% with paraffinic naphthenic (LPA) carrier oil and pyrethrosin, which makes up 0.2-0.4% of the final product, is less soluble in the diluted oil and precipitates during storage. These crystals have been reported to block the nozzles during insecticide application. In addition, it has been reported that exposure of humans to pyrethrosin elicits an allergenic response. The solubility of pyrethrosin in pyrethrin oil diluted with a carrier oil is a major factor that compromises the quality of pyrethrin oil. This study focuses on two aspects: pyrethrosin solubility in stored pyrethrin oil in the presence of carrier oils and the investigation of the biosynthetic pathway for the production of STLs. Firstly, pyrethrum STLs were characterised and quantified using different techniques, including X-ray crystallography, polarised light microscopy and nuclear magnetic resonance (NMR). The precipitate was found to be pyrethrosin, despite the tentative identification of seven other STLs in the oil including cyclo-pyrethrosin, β-cyclo-pyrethrosin, dihydro-β-cyclopyrethrosin, C7-C8 guaianolides, tatridin A and dihydro-tatridin A and C7-C8 eudesmanolide, chrysanin. A method for quality control prior to distribution of the commercial product using polarised micoscopy was developed. Quantitative analytical methods developed in this study to analyse pyrethrum oil firstly trialled the use of liquid chromatography, as the pyrethrin esters are thermally labile however, coupling this with detection using Mass Spectrometry (MS) with Electrospray Ionisation (ESI) was non quantitative as the esters were found to degrade in the electrospray ionisation (ESI) chamber at high temperatures (400°C), whilst pyrethrosin condensed at lower temperatures (120°C, 200°C). Photo Diode Array (PDA) detection was also unsuitable for the simultaneous analyses of pyrethrosin and pyrethrin esters as co-eluting peaks required the specificity of Single Ion Monitoring (SIM). Similarly, when analysed by Gas Chromatography with detection by Flame Ionisation Detection (GC/FID), degraded pyrethrin I coeluted with pyrethrosin, resulting in an elevated baseline on the GC chromatogram. An optimal temperature gradient for GC was established which minimised the degradation of pyrethrin esters allowing for the quantification of pyrethrosin in the trials undertaken in this study. With a view to selectively remove pyrethrosin from pyrethrum oil prior to distribution, the application of silica and octadecane columns were trialled. Silica was found to effectively retain 94% solubilised pyrethrosin in reverse phase chromatography using hexane as the mobile phase while normal phase chromatography using octadecane as the matrix was less effective. Centrifugation was a superior method for removing precipitated pyrethrosin crystals when compared to filtration (0.45μm). The precipitation of pyrethrosin from oil is facilitated in the industry by storage of refined oil, diluted with carrier oil at -10°C, for a period of months, incurring significant costs to commercial operations. This study established the rate of precipitation under different storage conditions in several time series trials. The agitation of oil during storage to provide nucleation sites only slightly increased the rate of crystallisation. The study showed that the majority of the crystals were formed within the first 10 days at -10°C. Subsequent trials confirmed that the rate of precipitation was independent of storage temperature and predominantly occurred within the first 5 to 10 days of storage at both -10°C and 4°C. Larger-scale storage trials of pyrethrin oil at -10°C, 4°C, and room temperature showed that no significant difference in the rate of crystal formation was found for up to 90 days between the treatments. The blockage of spray nozzles by crystals could be alleviated by keeping pyrethrosin in solution. As such, the solubility of pyrethrosin in pyrethrin oil that had been diluted with different carrier oils was established. The solubility of pyrethrosin in mineral oil was found to be 9.4, 6.6, 5.93, 5.27, 5.1, 4.4 and 3.4 mg/g in 100, 90, 85, 80, 73, 70 and 55% diluted oil, respectively. A faster rate of precipitation was recorded for increasing amounts of carrier oil. The use of Isopar-M oil, mineral oil, and canola oil as alternative carrier oils were trialled at dilutions of 100, 80, 70, 55, 45, and 38 % relative to pyrethrum oil. All of the oils formed a partition except canola oil. Pyrethrosin concentration remained constant over time in all of the different dilution ratios of canola oil in pyrethrum oil. The localisation of STLs in the trichomes of pyrethrum flowers was confirmed by the selective extraction of the lactones by dipping the complete pyrethrum flower head in DCM. Only trace levels of esters were co-extracted, with the exception of cinerin 1. Environmental Scanning Electron Microscope (ESEM) images confirmed that the trichomes were emptied of oil without damaging the epidermis. An alternative to the removal of pyrethrosin from pyrethrum oil was to selective breed out the genetics for the production of STLs. Two seed lines named ‘Evil’ and ‘Virtuous’ had been selectively bred to produce high and low levels of pyrethrosin, respectively. Examples of each were propagated, vernalised and the flowers screened. The lowest level of pyrethrosin was 0.06%, detected in Virtuous, which was 24 times less than that recorded for Evil at 1.42%. Peaks tentatively identified as cyclopyrethrosin and dihydro-B-cyclopyrethrosin, on average, were more concentrated in Virtuous plants. ESEM was used to determine the size, density, and distribution of trichomes in both seed lines. On average, there was no significant difference in the trichomes number, although the spatial distance between organelles was found to be significantly different between the two seedlines. The genetic difference between Evil and Virtuous presented the opportunity to investigate the biosynthetic pathway of STLs. Three enzymes, TcGAS, TcGAO, and TcCOS, previously identified as providing the precursors to STLs, were compared by analysing the Gene Expression Ratio (GER) of these enzymes. The entire methodology of GER analysis was optimised for Tasmanian pyrethrum seedlines. Primers to allow for DNA amplification by Polymerase Chain Reaction (PCR) were designed along with the primers for two reference genes TcGAPDH and TcActin7, using a nucleotide sequence database GenBank of National Centre for Biotechnology Information (NCBI), and a public database for primer design Primer 3 of Free Software Foundation USA. The PCR program was optimised for primers specificity for all five genes. The amplified DNA of TcGAS, TcGAO, and TcGAPDH showed a single band on the gel with the expected number of base pairs (bp) when referenced to a ‘ladder’ of known gene fragment sizes, however, a double band was observed for TcCOS, and this may have been due to the formation of primer dimers. Real-time quantitative PCR (RT-qPCR) was used to establish a DNA standard curve with good linear regression efficiency of TcGAS (efficiency:1.00, R\(^2\)=0.9995), TcGAO (efficiency:0.97, R\(^2\)=0.9996), TcGAPDH (efficiency:1.01, R\(^2\)=0.9997), and TcCOS (efficience:0.99, R\(^2\)=0.9994). Melt curves analysis confirmed the purity of DNA for all the genes except for TcCOS, which showed a double peak. Each gene was sequenced, and the BLAST database showed the TcGAPDH sequence matched 93% with the related species Helianthus annuus. The sequences for TcGAS, TcGAO, and TcCOS matched 100%, 97%, and 100% respectively with Tanacetum cinerariifolium and TcActin7 matched 96% with Chrysanthemum morifolium, which is a plant species closely related to pyrethrum in Asteraceae. The relative rate of expression for the three genes in Evil and Virtuous was compared using the technique known as GER analysis. The RNA of six Biological Replicates (BRPS) from stage 3 flowers of each seedline were extracted. The RNA Integrity Number (RIN) for each BRP was >7, confirming the quality of the RNA. The Reverse Transcriptase qPCR (RT-qPCR) of TcGAPDH showed good Cq differences for both seedlines but the melt curve of the cDNA from the Evil seedline presented two peaks rather than a single DNA amplicon. Sequence analysis of the TcGAPDH revealed that the two peaks of the amplicon were due to nucleotide degeneracy of the sequence in Evil BRPS. In GER analysis by RT-qPCR, the cDNA segments of the 12 BRPS were amplified, and the expression ratio of each gene was calculated against two reference genes using the Relative Expression Software Tools (REST) method. The results showed TcGAS expression is down-regulated (p=0.025), while TcGAO (p=0.59) and TcCOS (p=0.14) were at the same level in Virtuous relative to Evil plants. The down-regulated expression of TcGAS correlated with lower pyrethrosin concentrations but did not correlate with the overall increased levels of other STLs, which were higher in Virtuous compared to Evil. Down-regulation of TcGAS, and the implied decrease in the biosynthesis of germacrene A, a precursor to STLs early in the biosynthetic pathway, may provide for the redirection of resources to other secondary metabolites in Virtuous plants such as flavonoids or other minor bioactives. This study provides knowledge to reconsider existing processes for the production of quality pyrethrum oil in terms of pyrethrosin solubility and storage. Investigations undertaken into the biosynthetic pathway for STLs contributes to the understanding of enzyme activity to inform pyrethrum breeding programs.

Pyrethrum is a perennial daisy that is predominantly out-crossed. It is grown commercially in Tasmania for pyrethrins, which are extracted from the flowers and are used in the preparation of insecticides. A pyrethrum breeding program was conducted by the University of Tasmania from 1978 to 1998, with the aim of developing varieties suitable for Tasmanian growing and production systems. The primary selection character was pyrethrins yield, and the program consisted of population improvement through recurrent selection for two indirect selection characters; visually-estimated flower yield and the UV-assay of pyrethrins at a specific maturity stage (open but not over-blown). Varieties, consisting of either clones or single crosses between two clones, were selected from each successive recurrent selection generation. This study was undertaken to evaluate the selection methods used for the University's breeding program from the period of 1985 to 1998, and to assess the impact on breeding methods of the change in the mid-1990's from the establishment of the crop by clonal splits planted at a contant spacing (0.5 m) to establishment by direct-drilling at a somewhat higher average plant density. Data records for the two main selection characters were analysed in order to identify genetic changes in the breeding population. This indicated that there had been a genetic gain in UV-assays but not in flower yield. The efficiency of direct selection for first year yields in the Univeristy's base population was compared with indices of the indirect selection characters and yield components. A trial was established to obtain estimates of genetic parameters to construct selection indices and predict selection responses. Estimated heritability was moderate for pyrethrins yeld (0.26-0.39) and for the yield components of pyrethrins content (0.24-0.34) and flower yield (0.17-0.30), and low for percent dry matter content (0.00-0.15). Estimates were also obtained for two product quality characters, both derived from the ratio of the six different esters that form the active product in the pyrethrum extract. Heritability for these character was moderate to high. The effect of planting density on selection for pyrethrins yield was also investigated. The index of the two indirect selection characters was the most efficient method for single-plant selection in the planting densities used commercially from 1980 to 1995. However, an index of component characters was more efficient for the densities currently used in commercial areas. There was evidence for a genotype-density interaction for flower and pyrethrins yield but not for percentage pyrethrins or the ratios of pyrethrins esters. The potential to reduce the length of the period of obligate vegetative growth through breeding was also assessed. There was evidence for significant level of additive genetic control for this character and it was concluded that the vegetative period could be reduced by recurrent selection. The performance of new synthetic varieties were evaluated and the relative merits of varieties derived from a single cross of two clones and polycross of several clones were discussed.

This study investigated the morphological and physiological changes associated with flower initiation and development in pyrethrum, Tanacetum cinerariaefolium L. Detailed morphological descriptions of vegetative and floral apices have been given and a scale of reproductive developmental stages based on these descriptions was proposed. It was shown that each stage of apical development was associated with a narrow range of apical diameters. The irreversible commitment to floral development was observed to occur when the first involucral bract was initiated on the apical dome and this point was characterised by a critical apical size. The apical diameter at this stage was always approximately 220 pm. A juvenile-like condition was described for pyrethrum seedlings, tissue culture explants and vegetatively divided splits. During the period of juvenile-like growth the plants were not competent to respond to normally inductive treatments. The juvenile-like phase lasted until the plants had reached a minimum size or stage of development, but did not depend on chronological age. The attainment of meristem competence was associated with the release of lateral buds from apical dominance. Terminal meristems were never observed to initiate flowers. Axillary meristems became competent to flower a short time after being released from apical dominance, while older axillary meristems were observed to lose their competence. It was noted that the loss of competence to flower of lateral shoot meristems occurred after the release from apical dominance of new axillary buds on each lateral shoot. The effects of the following environmental conditions on flowering were examined in a series of experiments; vernalisation, daylength, day temperature and photon flux. The major environmental requirement for flower initiation in pyrethrum was found to be a period of low night temperature or vernalisation. While flowering occurred eventually under 'non-inductive' conditions through an autonomous induction process, vemalising conditions were required to stimulate rapid flower initiation and development. Plants displayed a quantitative response to vernalisation as longer periods under vernalising conditions resulted in larger numbers of flowers, longer flower stems and more rapid flower initiation and development. Night temperatures of less than 18 °C were required to provide the vernalisation stimulus, with two weeks at 6°C or three weeks at 12°C demonstrated to be the minimum vernalisation requirement under short days and day temperatures of 20- 30°C. Both day temperature and photon flux density conditions were shown to modify the response to vemalisation. Low photon flux density conditions (350 gmol.m -2.s1 or less) retarded flower initiation regardless of day temperature. High day temperatures combined with low photon flux resulted in a devemalisation-like effect where the plants were incapable of responding to otherwise inductive vemalising conditions. A true devernalisation effect was also demonstrated under these conditions with the vemalisation stimulus being reversed by a later high temperature / low photon flux treatment Daylength had a quantitative effect on both flower initiation and development, with both processes promoted by long days. The inhibitory effect of short days was thought to be mediated through reduced assimilate supply and not via the phyochrome reactions. It was concluded that pyrethrum is a day-neutral species as its daylength reaction was due to the daily light integral and not to photoperiod. Autoradiography was used to follow the distribution of 14C photosynthate during flower initiation and development under 'inductive' and 'non-inductive' conditions. This method was also used to study the effect of devemalising conditions on assimilate partitioning. The terminal shoot apex and young developing leaves were the main sinks for assimilates under 'non-inductive' conditions. The sink strength of the axillary shoots in 'inductive' conditions was observed to increase prior to the end of evocation and they became the dominant sink for radiolabelled assimilates as floral development progressed. 'Devemalising' conditions reduced the sink strength of the axillary buds, or prevented the translocation of assimilates to them. Radioimmunoassays were performed to quantify the changes in plant growth regulator concentrations in mature leaf samples under 'inductive', 'non-inductive' and 'devernalising' conditions. Vernalising conditions stimulated an increase in the concentration of gibberellins while 'devernalising' conditions resulted in a reduction in the concentration of gibberellins to levels below that of unvemalised plants. The concentration of the auxin indolylacetic acid declined under vernalising conditions while sdevernalising' conditions prevented this decline. No evidence was found during this study of a role for the cytokinins or abscisic acid in the flowering of pyrethrum. However a possible role for these hormones could not be ruled out as all the assays were performed on mature leaf samples and as such would not have detected any localised fluxes of hormones in other plant organs. Two cultivation techniques for manipulating the flowering behaviour of pyrethrum in the field were examined. The first involved the application of growth retardants to reduce flower stem height and the degree of lodging at harvest. 'EL-500' and 'Cultar' at rates equivalent to 5 Kg active ingredient per hectare or above were shown to significantly reduce flower stem height and lodging. The growth retardants also reduced flower yield if applied during the period of flower initiation. The most effective control of lodging, without reducing flower yield, was found to result from application of the growth retardant immediately prior to the period of maximum stem extension, which under Tasmanian field conditions occurs in October. The growth retardant 'Culiar' reduced the gibberellin concentration in treated plants, and the effects of the growth retardant were reversible by application of gibberellin A3. The effect of trimming on the flowering of plants in the field was also examined. Multiple trimming treatments were shown to be the best strategy for promoting vegetative growth during the normal flowering period. Slashing plants to a height of approximately three centimetres above ground level during October and November, when the inductive vernalising conditions no longer prevail, significantly reduced the flower yield and increased the yield of vegetatively divided splits. This is of economic value in field nurseries where multiplication of planting stock is achieved through vegetative division of nursery stock. The process(es) of flower initiation and development were discussed in light of the experiments detailed above.

At the time this research project was initiated, the Tasmanian pyrethrum industry was attempting to establish crops by sowing rather than by planting of 'splits' or seedling plugs. This thesis investigated plant density and sowing times required for maximum yield. Studies were also conducted to improve chances for successful crop establishment from seed. That work investigated environmental requirements for germination and aspects of seed quality and seed production. Previous studies have examined the influence of density on yield of pyrethrins, but none had been conducted in sown trials, in cool temperate environments, over a wide range of plant densities. Furthermore, the influence of plant density on components of yield had not been intensively investigated. This work identified that maximum yield was achieved in the first season at between 16 and 39 plants/m2 and at or above 16 plants/m2 in the second season following establishment. Yield was a function of dry flower yield rather than due to change in percentage of pyrethrins in the flowers. Higher flower yield was associated with greater above- ground dry matter production. The yield component which increased with plant density was number of flowering tillers/m 2 . Yield components decreasing with density included number of flowering tillers/plant, flowers/tiller and dry weight/flower. Other aspects of changing plant morphology and development with density investigated included, crop height, mean flower maturity and plant survival. The recommendation to industry was to aim for a plant population of above 16 and below 39 plants/m2 . Yields achieved in this work were unprecedented in the pyrethrum agronomy literature. No reported studies have examined the influence of time of sowing on pyrethrins yield. Field studies showed that sowings later than mid-November led to significant reductions in yield in the first flower harvest. Yield reductions were associated with decreased dry matter production/plant and flowering tillers/plant. Later sowings resulted in plants failing to flower or in significant reductions in the extent of flowering. Sowing earlier than mid-November resulted in no significant increases in yield. As pyrethrum crops have not been traditionally established by sowing, only scant information was available on requirements for germination, or seed quality. A field study and three laboratory trials investigated the influence of temperature and seed quality on germination and emergence. Results demonstrated that rate and final emergence varied significantly at different times of year. In general, the proportion of viable seed sown that emerged and survived was very low. Both rate of emergence and final emergence percentage were associated with temperature. Laboratory investigation of germination percentage, rate and uniformity of germination of a seed lot under a range of constant temperatures confirmed the previously reported findings relating to germination characteristics of this temperate species. Unexpected though was the high proportion of dead seed found at temperatures only several degrees higher than the temperature for optimal germination rate. Six seed lots were subsequently germinated at low, medium and high temperatures which provided some insight into the seed death phenomenon as well as documenting the range of behaviours from different seed lots. An explanation for differences in germination behaviour involving seed maturity was proposed for different seed lots and this was tested in a subsequent study. Finally, the effects of an 18 month storage period on the germination characteristics seed lots was investigated in a laboratory trial. There was little change in most germination parameters except for time to complete germination which increased in all seed lots after storage and uniformity of germination which decreased. Changes were assumed to be due to satisfaction of an after ripening requirement. Laboratory studies investigated the influence of seed mass on various germination and seedling development characteristics. Variation in seed mass within seed lots was identified but this had little impact on rate of germination or other germination parameters. However, a following, study revealed that heavier seedlings emerged from seeds that germinated earliest within seed lots. Furthermore, heavier seedlings demonstrated faster development than did lighter seedlings. Although seedling mass was found to be associated with rate of germination within seed lots, this factor failed • to explain significant differences in mean time to germinate between seed lots. It was subsequently recommended to industry that cleaning on a size or mass basis could improve seed quality. The influence of harvest date on seed quality and quantity were investigated. Results indicated that losses of larger achenes from the harvested flowers were occurring with lateness of harvest. Characteristics of the seed including mean seed mass, proportion of viable seed in the sample, germination percentage and rate of germination were found to vary considerably with harvest date. Data generated in this work were found to be consistent with the model which proposed that variability in germination parameters was due largely due to maturity of seed at harvest. A following study revealed that relative position on the capitulum also had a profound influence on germination parameters of the seed. The recommendation for industry emanating from the harvest date work was that flowers should be cut at a field capitulum moisture of 25%. Finally, variation in seed quality and quantity was evaluated both within and between capitula. This study revealed that small flower heads produced fewer and smaller seed than larger heads. Outer achenes, regardless of whether they came from large or small capitula were found to germinate more rapidly than inner achenes. Peripheral achenes weighed more than central achenes and gave rise to heavier seedlings. The methods and results in this study will serve as a valuable source of information for agronomists and plant breeders working on improvement of pyrethrum production. The investigations on seed quality and seed production provide a sound base for future efforts to improve seed quality and crop establishment. The findings presented provide the Tasmanian pyrethrum industry with critical information with respect to target plant densities, sowing times, seed quality and seed production. It is expected that implementation of findings from the study will prove to be pivotal in continued industry profitability and expansion.

The Tasmanian pyrethrum industry has been operating on a commercial basis for 23 years and is now the second largest producer of pyrethrum (Tanacetum cinerariaefolium L.) for natural insecticides in the world. The industry, with a annual farm gate value in excess of $7 million, makes a significant contribution to the rural economy of Tasmania (Australia). Currently, production involves 120 contracted growers and 1300 hectares of land. To maintain its position as a world leader in the production of pyrethrum, the Tasmanian industry must continue to improve production technologies and efficiencies. An emerging area of concern is the management of weeds, with some weed species in pyrethrum being particularly difficult to control. This includes weeds that are commonly found in vegetable crops which are grown in rotation with pyrethrum as well as relatively uncommon species; such as Anthriscus caucalis and Torilis nodosa. This study investigated the biology and control of both A. caucalis and T. nodosa which belong to the Apiaceae family. A morphological examination highlighted easily distinguishable characteristics for the identification of these relatively unfamiliar species. Anthriscus caucalis seedlings are identifiable by their tri-pinnate compound leaves which are glabrous on top with scattered hairs beneath. The fruit of A. caucalis is ovoid in shape and 2.5 to 3.5mm in size with distinguishing hooked spines and a short beak. The pedicels have a ring of hairs at the top. Torilis nodosa seedlings are identifiable by their deeply bi-pinnate compound leaves and narrow linear lobes. The fruit of T. nodosa is ovoid in shape and 2.5 to 3mm in diameter and composed of 2 distinct dimorphic mericarps. The outer mericarp has barbellate spines with the inner mericarp tuberculate. A survey of pyrethrum crops revealed that the occurrence of these species was high, occurring in 30% of pyrethrum crops, with A. caucalis being the more prevalent species. As pyrethrum is a perennial crop which can be grown for up to five years, it was also found that the frequency of occurrence of Apiaceae species increased with increasing crop age. Investigations into the germination characteristics of A. caucalis and T. nodosa revealed that A. caucalis possessed an innate seed dormancy which was overcome by seed scarification and dry storage at 20°C. Torilis nodosa displayed no innate seed dormancy. Both species, A. caucalis and T. nodosa, were found to behave predominantly as winter annuals, germinating in autumn and over wintering as small rosettes. Studies indicated that T. nodosa has a transient to short term persistent seedbank, while A. caucalis has a short to long term persistent seedbank. Anthriscus caucalis was found to undergo rapid vegetative stem development during late winter early spring with flowering commencing during mid spring. Seed maturation occurred in early summer. Tori/is nodosa was found to produce procumbent stems in mid to late spring and flower approximately 6 weeks later than A. caucalis with seed maturation occurring in mid to late to summer. Studies into the chemical control of A. caucalis and T. nodosa identified a small number of herbicides with potential for use in pyrethrum. Applications of dimethenamid at 3.6 kg/ha provided the most selective pre emergent control for both species, while clomazone applied at 120.0 g/ha provided very effective control of T. nodosa. Imazamox applied at 34 g/ha provided significant post emergent control of both species with excellent selectivity for use in pyrethrum. A number of other herbicides were identified as having activity on A. caucalis and T. nodosa, however lower levels of selectivity limited their potential adoption for use in pyrhethrum. This thesis has provided significant information to the pyrethrum industry on the biology and competitive nature of the relatively unknown weed species A. caucalis and T. nodosa. The thesis has also provided immediate short term weed control strategies and enhanced the weed management options available for the industry.

Pyrethrum is a major crop in Tasmania, Australia. The lack of predictability in flower dry matter (DM) production and therefore crop yield is one of the biggest challenges for the industry, with large variation in dry matter yield currently occurring between production sites and seasons. If the industry is to expand, consistency in production is required to ensure crop production matches the volumes contracted for delivery to the market. Knowledge of the main factors affecting DM accumulation under commercial production conditions is required to develop crop management strategies capable of delivering consistent, high yields. The specific objectives of the studies reported in this thesis were to investigate the effect of: (i) light interception; (ii) plant density; and (iii) water stress on DM yield and pyrethrin content of pyrethrum. The indirect measurement of light interception in pyrethrum using crop green leaf area and volume was used to assess the relationship with flower DM yield and pyrethrin content. Strong relationships were identified within sites and seasons, but not across all seasons and sites assessed in the study. As there was no clear relationship between light intercepted and pyrethrin yield, other factors could be involved. It was hypothesised that differences could be related to variation in rate of crop development, plant density and drought stress, which can occur during the later stages of flower development in Tasmania. Plant density was shown to have a large effect on DM partitioning in a field trail conducted in north-western Tasmania in 2008-09. The relatively large leaf, stem and flower DM yield per plant at low (6 plants/m2) plant densities was compensated by the increase in plant numbers at high (44 plants/m2) densities. Variation in plant density is therefore likely to affect light interception. There was, however, no difference in the concentration of pyrethrin, the active ingredient extracted from the pyrethrum flowers, with plant density. No significant differences in DM accumulation were found between pyrethrum genotypes and there were no density x genotype interactions. Drought stress was investigated by conducting pot trials under controlled conditions. Treatments included 3, 4 and 5 day watering intervals applied at early flower maturity stage 2 (FMS 2), mid (FMS 4) and late (FMS 6) flowering stages for either short (10 days) and long (20 days) durations. Plants watered on a 3 day interval tended to have lower (less negative) leaf water potential and displayed no visual signs of water stress. The FMS stage that water stress treatment was applied affected plant and flower dry matter yield, but watering interval or duration of stress treatment had no significant effect. Plants produced greater DM when drought stress was imposed during early compared with late flowering. Plant pyrethrin content, measured as pyrethrin concentration in the harvested flowers, was not affected by any of the drought treatments. Consequently, it was concluded that plants grown in field conditions are likely to be able to recover from mild drought stress, particularly during early flowering, but yield will be reduced if plants are stressed at later flowering stages. This thesis has provided information to the pyrethrum industry on plant growth in response to crop management strategies and has highlighted the complexity of processes determining crop dry matter and pyrethrin yields.

Pyrethrum (Tanacatum cinerariifolium) is cultivated worldwide and in southern Australia for the extraction of insecticidal esters or pyrethrins contained within the achenes of flowers. Producing a significant proportion of the worlds botanical pyrethrins, Australian crops may suffer reduced yields from annual flower disease epidemics caused by pathogenic fungi Botrytis cinerea and Sclerotinia sclerotiorum. Little is known regarding (i) the epidemiology of flower blights caused by B. cinerea and S. sclerotiorum in pyrethrum, (ii) the efficacy of current control methods, (iii) whether there is evidence of fungicide resistance in the fungal population, (iv) whether alternative fungicides can provide improved control over those currently used, (v) whether other fungi could be involved in annual flower disease epidemics and, (vi) the loss in flower yield and pyrethrin assay from flower diseases. A survey of the incidence of B. cinerea and S. sclerotiorum in flowers was undertaken in ten commercial pyrethrum crops in one year. Flowers from non-fungicide treated areas in commercial crops were periodically sampled throughout flowering prior to being surface sterilised and incubated under high humidity to encourage fungal growth. B. cinerea and S. sclerotiorum were prevalent, with both occurring in flowers from all 10 crops. The mean incidence of B. cinerea in flowers sampled between 10-11 December across all crops was 56%, significantly higher (P = 0.024) than S. sclerotiorum (29.4%), while between 16-18 December mean incidence of B. cinerea was 75.7%, again significantly higher (P = 0.026) than incidence of S. sclerotiorum (51.8%) at this time. The main means of managing flower blights of pyrethrum is currently through a fungicide program over flowering involving tebuconazole and carbendazim. The efficacy of the flowering fungicide program for controlling B. cinerea and S. sclerotiorum flower blights and promoting benefits in terms of increased yield was evaluated in nontreated and fungicide treated plots in 10, 10 and 17 commercial pyrethrum crops during the flowering period over three years, respectively. In each of two years, fungicide treatment resulted in a mean incidence of B. cinerea near to harvest which bordered on being significantly lower (0.05 < P < 0.1) than no treatment. In the third year, fungicide treatment significantly (P = 0.038) reduced mean fungal incidence from 75% to 53.3% in nontreated close to harvest. The incidence of Sclerotinia sclerotiorum in flowers from nontreated was significantly higher (P < 0.001) than from fungicide treated in year one (62.8% and 29.2% respectively) and again significantly higher (P = 0.019) in year two (26% and 8.1% respectively). Across all fields, fungicide applications in year one resulted in no significant yield improvement. During year two, mean pyrethrin yield (across all sites) was significantly increased from 90.4kg/ha in nontreated areas to 108.4 kg/ha in fungicide treated plots (P = 0.032) and during the final year fungicide treatment significantly increased dry flower yield by 216 kg/ha (P = 0.018) in comparison to nontreated. However, while fungal incidence was often reduced within individual fields or across all sites by fungicides, few measurable benefits in yield quantity or quality were measured. In vitro sensitivity testing of 49 and 96 isolates of B. cinerea and S. sclerotiorum respectively to fungicides commercially used for control was conducted. A proportion of B. cinerea isolates appeared sensitive to carbendazim at low concentrations i.e. 39% had EC50 less than 1.05 μg a.i./ml. However a high proportion (61%) of B. cinerea isolates had EC50 values to carbendazim greater than 100 μg a.i./ml. This provided evidence for potential resistance amongst the population of B. cinerea and may explain the inconsistency of yield responses from the fungicide program. By comparison, 96.9% of S. sclerotiorum isolates had EC50 values <2.4 μg a.i./ml to carbendazim with a mean of 0.5 μg a.i./ml. However, a small number (3.1%) of S. sclerotiorum isolates were highly insensitive to carbendazim with EC50 of >1000 μg a.i./ml. All isolates of B. cinerea and S. sclerotiorum were sensitive to tebuconazole at low concentrations with mean EC50 of 0.64 and 0.18 μg a.i./ml respectively. Alternative fungicides for flower disease control were evaluated in replicated field trials to determine suitability for inclusion into the flowering fungicide program as replacements for the potentially ineffective and now deregistered fungicide carbendazim. Boscalid and iprodione showed greater benefits in terms of yield from statistical analysis than other treatments, and significantly reduced fungal incidence of flowers equal to, or better than, commercial treatments. The mean incidence of B. cinerea from nontreated flowers sampled on 4 December of 30% was significantly (P = 0.007) higher than boscalid (8.5%) and iprodione (10%). Mean incidence of S. sclerotiorum of flowers from nontreated plots sampled on 4 December was 26.5%, and significantly (P <.001) higher than boscalid (7%) and iprodione (12%). In vitro fungicide sensitivity of 46 isolates of B. cinerea and S. sclerotiorum to iprodione indicated no evidence of reduced sensitivity with mean EC50 (and maximum) values of 1.62 (8.48 μg a.i./ml) and 0.19 (0.61 μg a.i./ml) respectively. Sclerotinia minor, a previously undocumented pathogen of pyrethrum flowers, was consistently isolated from diseased, surface-sterilised pyrethrum flowers over multiple years. Fungal identity was confirmed with morphological, genetic and phylogenetic evaluation. Occurrence in-field of mature, sporulating, apothecia of S. minor were documented; while sclerotia of 8 of 10 isolates conditioned in the laboratory successfully underwent carpogenic germination in vitro demonstrating the ability of endemic isolates of S. minor to produce air borne inoculum necessary to achieve flower infection in pyrethrum and other crops. The relative sensitivity of isolates of S. minor to the fungicides carbendazim, tebuconazole and iprodione was evaluated. Mean (and maximum) EC50 values of S. minor were 1.92 (2.62 μg a.i/ml) for carbendazim, 0.1 (0.13 μg a.i./ml) for tebuconazole and 0.3 (1.23 μg a.i/ml) for iprodione. The effect of B. cinerea and S. sclerotiorum flower inoculation on measured yield attributes and pyrethrum flower development were evaluated with glasshouse studies. Inoculation of immature flowers with Botrytis cinerea resulted in highly significant reductions in dry weight (P <.001) and pyrethrin yield (P <.001) of mature flowers compared to non inoculated. Inoculation with S. sclerotiorum in two of four varieties resulted in significantly higher flower development stages after 21 days (P = 0.021; P <.001) and significantly faster flower senescence (P = 0.001; P = 0.008) in comparison to non inoculated. Inoculation of flowers with ascospores of S. minor led to symptoms of flower disease indistinguishable from those of S. sclerotiorum flower blight, significantly lower fresh flower weights in replicated experiments (P = <.001; 0.032) and significantly higher developmental stage of flowers (P = <.001) after only two weeks in comparison to non inoculated. These studies indicated that flower infection with B. cinerea and S. minor could significantly reduce flower yield and pyrethrin yields. Furthermore, significantly faster flower development; senescence and desiccation may result from flower infection with S. sclerotiorum and S. minor. The completion of Koch’s postulates additionally confirmed pathogenicity toward pyrethrum flowers and demonstrated pure cultures of S. minor could be reisolated from flowers inoculated on living host plants. The study has provided new knowledge in the epidemiology of known flower blights of pyrethrum caused by B. cinerea and S. sclerotiorum. Furthermore the study has described for the first time S. minor as a pathogen of pyrethrum flowers and documented a rare example of carpogenic germination of sclerotia being involved in the epidemiology of a disease caused by S. minor. The study has also provided insights into the effect of flower blights on pyrethrum yield and into improving the management of flower diseases by fungicides.

Tan spot is one of the most significant foliar diseases of pyrethrum (Tanacetum cinerariifolium) in Tasmania, Australia. It is associated with tan coloured, necrotic lesions on leaves, stems and flower buds which result in a loss of green leaf area. Severe outbreaks can lead to plant death and termination of crops. Phylogenetic and morphological analysis of the causal pathogen, originally described as Microsphaeropsis tanaceti, resulted in reclassification of the pathogen as two closely related species of Didymella, namely D. tanaceti and D. rosea. Molecular differences between the two species were evident for four of the five genome regions examined. Furthermore, morphological differences in culture pigmentation and conidial size allowed rapid identification of the two species. To elucidate explanations for the shift from a minor disease in 2001 to one of the most frequent and severe diseases of pyrethrum, the reproductive strategy and population structure of the dominant tan spot pathogen, D. tanaceti were investigated. Two first harvest fields in each of two regions, were intensively sampled in July/August 2012, using two 50 m transects in each field, to provide field populations of D. tanaceti for analysis. Tan spot incidence in these fields was high, with 325 isolates obtained from 800 sampling units. Analysis of the structure and arrangement of mating-type (MAT) genes identified a putative heterothallic mode of reproduction for D. tanaceti, with either a single MAT1-1-1 or MAT1-2-1 gene occurring in D. tanaceti isolates. A multiplex MAT-specific PCR assay was developed and validated. This assay was utilised to quantify the number of D. tanaceti isolates of each MAT gene within the field populations. Isolates with a MAT1-1-1 gene occurred in equal frequencies with isolates containing a MAT1-2-1 gene. Significant spatial structure of isolates of each MAT gene in the fields was absent. Additionally, two haplotypes of the MAT1-1-1 gene, sharing 99.6% sequence homology, were identified. Isolates of the two haplotypes were differentiated using a restriction enzyme digest of the MAT1-1-1 amplicon. Within the field populations, haplotype I was dominant, occurring in 95.6% of isolates with a MAT1-1-1 gene. These results suggested that the occurrence of a cryptic sexual reproduction cycle in Tasmanian pyrethrum fields could not be dismissed. However, in vitro crosses between compatible isolates failed to produce ascospores, suggesting that specific environmental conditions and temporal requirements were necessary. To assess the genetic structure of D. tanaceti, a set of polymorphic microsatellite (SSR) markers were identified based on sequence data obtained from genomic sequencing. The D. tanaceti field populations were genotyped using eight SSR markers, with alleles determined by high resolution melt analysis. Within the 317 isolates genotyped, 127 multilocus genotypes were identified, with 82 represented by a single isolate. Furthermore, high average genetic and genotypic diversity were identified within the field populations. However, genotypic spatial structure between regions, fields and within transects in each field were absent. Similar frequencies of alleles were observed for each marker in each of the fields. Evidence of high levels of genotypic migration between fields was also detected. These results suggested that individuals within these populations may have originated from a common genetic source, such as an alternative host or seed crop. Linkage disequilibrium was detected within fields. Thus, long distance dispersal of inoculum via ascospores may have only minimally contributed to disease incidence in these fields. Control of tan spot has relied, mostly, on the use of boscalid, a succinate dehydrogenase inhibitor fungicide. Boscalid has been used widely in pyrethrum for control of multiple pathogens. To initiate the characterisation of the molecular mechanisms associated with the observed boscalid resistant phenotypes, the succinate dehydrogenase subunit B (SDHB) gene of D. tanaceti was sequenced. The SDHB gene sequences of isolates with varied in vitro growth responses to boscalid (different resistant phenotypes) were compared. The results revealed that a decreased sensitivity to boscalid was associated with the substitution of a highly conserved histidine residue at codon 277 with either tyrosine (H277Y) or arginine (H277R). These two substitutions have been shown to cause boscalid resistance in other fungal species. In addition, an isoleucine to valine (I279V) substitution occurred at codon 279, but was not correlated with a decreased sensitivity to boscalid. Both the H277Y and H277R substitutions were associated with isolates exhibiting moderately resistant (EC₅₀ 0.5 ‒ 5.0 μg a.i/mL), resistant (EC₅₀ 5.0 ‒ 50.0 μg a.i/mL) and highly resistant (EC₅₀ 50.0 ‒ 250.0 μg a.i/mL) phenotypes. No isolates with a boscalid susceptible phenotype (EC₅₀ 0.0 ‒ 0.5 μg a.i/mL) were associated with these substitutions. However, the association of isolates with a WT SDHB gene for each of the moderately resistant, resistant and highly resistant phenotypes restricted the ability to correlate the H277Y and H277R substitutions with the resistant phenotypes. This indicated that mutations in the succinate dehydrogenase subunit C (SDHC) and D (SDHD) genes, or other regions may also occur in D. tanaceti. To evaluate the extent of SDHB gene mutations in the 2012 field populations, a high resolution melt analysis assay was developed and its ability to identify mutations in codons 277 and 279 of the SDHB gene verified. The majority of D. tanaceti isolates within the field populations contained a mutation in the SDHB gene. The H277Y substitution was the most dominant, occurring in 52.3% of isolates, while the H277R substitution occurred in 9.3%. Overall this thesis has identified and characterised the pathogens associated with tan spot of pyrethrum in Australia; establishing them as two species (D. tanaceti and D. rosea) from the genus Didymella. It has developed a collection of SSR markers and assays for the rapid identification of mating-type for use in future studies. Furthermore, it has initiated the characterisation of the molecular mechanisms associated with boscalid resistance and developed an assay for the rapid identification of SDHB alleles. Moreover, it has provided a greater understanding of population biology and structure of the dominant tan spot pathogen; D. tanaceti by the characterisation of intensively sampled field populations. While the specific reasons for the rapid increase in tan spot incidence and severity remain unclear, this study has identified possible factors which could be associated with it. Despite evidence against a frequent sexual cycle in the field populations, the high genotypic diversity within populations suggests D. tanaceti individuals have a high adaptive ability. The adaptive ability of the pathogen population was demonstrated by the development of insensitivity to the fungicide boscalid. However, it may have also provided D. tanaceti individuals with a competitive advantage (e.g. increased virulence) over other pyrethrum pathogens. Furthermore, the decreased efficiency of disease control from boscalid, due to the moderately high incidence of fungicide resistance, has undoubtedly played a significant role in increasing disease incidence. Thus, efficient long term control of the disease will need to be delivered from an integrated approach, incorporating methods to decrease inoculum loads and strategies based on alternation of fungicides in different resistance groupings with regular evaluation of the pathogen population for fungicide resistance.

The indiscriminate use of pesticides in the control of agricultural and public health pests has led to the development of insecticide resistance in many insect pest species, posing a large threat to the continued success of chemical control measures. Resistance towards pesticides can be due to a number of mechanisms, but enhanced detoxification of the insecticide by metabolic enzymes, such as esterases, glutathione-S-transferases (GSTs) or monooxygenases, plays a large role in insecticide resistance. Synergists, such as piperonyl butoxide (PBO) can be used to overcome metabolic insecticide resistance. Synergists are capable of inhibiting enzymes involved in insecticide resistance, thus restoring a level of susceptibility. By virtue of its enzyme inhibitory qualities, PBO is often used to enhance the efficacy of pyrethrum (an organic insecticide). In some countries, however, PBO is no longer certified for organic use and this study explored the use of natural plant oils or extracts as potential organically-certifiable pyrethrum synergists. From a commercial viewpoint, an effective natural synergist could potentially allow development of new pyrethrum formulations into higher value niche markets, such as organic agriculture. Synergised pyrethrum could also offer a new control option against insects resistant to synthetic pesticides in mainstream agriculture. The efficacy of various natural plant oils and extracts as pyrethrum synergists were examined for synergistic potential towards pyrethrum with several agriculturally significant insect pest species, Helicoverpa armigera, Frankliniella occidentalis, Myzus persicae, Meligethes aeneus and Musca domestica, a serious public health pest, using discriminating dose bioassays. PBO proved to be the most effective pyrethrum synergist in all species tested, however, a number of natural plant oils also showed efficacy. The most promising compounds were dill apiole oil and parsley seed oil, synergising pyrethrum in all the species tested. Dill apiole oil and parsley seed oil contain the methylenedioxyphenol ring structure that is characteristic of PBO. Enzyme inhibition studies were used to examine the ability of the natural plant oils and extracts to inhibit the enzymes involved in metabolic insecticide resistance (esterases, GSTs and monooxygenases). In general, inhibition of esterases and GSTs was not correlated to synergism of pyrethrum in vivo. However, in H. armigera and M. persicae, with esterase mediated pyrethroid resistance, PBO, dill apiole oil and parsley seed oil inhibited non-specific esterases. In M. aeneus and M. domestica, where resistance to pyrethroids has been linked to monooxygenases, PBO, dill apiole oil and parsley seed oil showed significant inhibition of monooxygenases. However, not all compounds that inhibited esterases or monooxygenases synergised pyrethrum. The lack of correlation between enzyme inhibition and synergism could be due to a number of factors, including the ability of the synergist to penetrate the insect cuticle, speed of distribution of the synergist through the insect, metabolism of the compounds in the insect and affinity for the target site.

Determination of the critical level of training data, and investigation of targeting the time of image acquisition to specific crop growth cycles, will increase the efficiency of remote sensing data analysis, for recognition of poppies and pyrethrum. The objective of this project, was to determine whether the amount of training data (critical sample size) contributed significantly to classification, using three methods of analysis of two season's data for, poppy and pyrethrum crops, on the North West Coast of Tasmania and to investigate the timing of image acquisition. Distinction between class types was not an objective of the study. Eight Landsat 5 TM, two SPOT XS and three SPOT XI images were acquired between 02 July 1997 and 28 March 1999. Observations of the spectral response of poppies showed that, Landsat TM bands one, two, three and five in November and January, provided significantly different, peak pixel values for the poppy crop. SPOT XS band two in February also provided peak pixel values for poppies. The spectral response of pyrethrum indicated that an increase in pixel value for the January Landsat TM data in bands five and seven was distinct, as was the peak in SPOT XI band three during December. A principal component analysis, (PCA) was carried out separately on each image. For all Landsat TM imagery, over 98.482% of variance was contained within the first three principal components. Similarly, for all SPOT XI data, over 99.189% of variance was contained within the first three principal components. When SPOT XS data was analysed, the first and second components accounted for over 98.909% data variance. The merged spectral response patterns generated from the automatic internal average relative reflectance (AIARR), normalised difference vegetation index (NDVI) and PCA images by an unsupervised, iterative self-organising data analysis technique (ISODATA) for the poppy and pyrethrum AOIs, provided the input for a supervised classification. As the AIARR, NDVI and PCA data were normally distributed, and the spectral response patterns were parametric, a maximum likelihood parametric decision rule was selected. The amount of training data had a significant effect on the contribution to classification for the three analysis methods, over two season's data for each of the crop types. To achieve a classification accuracy of 90% for poppies, the acquisition of Landsat data in either November or January, required a PCA with 80% of the total poppy area used as training (calibration) data. To achieve a classification accuracy of 96% for poppies, the acquisition of SPOT XI data in either October or December, required a NDVI analysis with 50% of the total poppy area used as training (calibration) data. For pyrethrum, a classification accuracy of 80% was achieved by acquiring imagery in the post harvest and dormancy stage (late February to October), using a PCA method and 90% of the total amount of data available for training. When imagery was acquired in late December or early January, using 40% of the total amount of pyrethrum data for training contributed, on average, to the classification of 87% of the crop, when analysed using the NDVI method. The findings of this research showed that the choice of the training set (quality and quantity) had an influence on the success of a classification approach as well as the choice of image analysis technique. Timely acquisition of imagery was shown to be required to achieve a satisfactory level of contribution to classification from training data of poppies and pyrethrum.

Pyrethrum (Tanacetum cinerarafolium) is a perennial plant grown commercially in Tasmania, Australia for production of insecticidal pyrethrins. Tasmania produces over 30 % of the world's supply of natural pyrethrins. Ray blight disease caused by the fungal pathogen Phoma ligulicola var. inoxydabilis, is a major limiting factor of pyrethrum production in Tasmania. It causes dieback of emerging stems in spring, and flower disease over summer. Most of the information regarding this pathogen is based on studies of the variety that causes ray blight on chrysanthemum, P. ligulicola var. ligulicola. The objectives of this project were to characterise the biological, cultural and genetic variability, and investigate the reproductive nature of the P. ligulicola var. inoxydabilis from pyrethrum. This study also aimed to confirm the species identity and determine the variety of Phoma exigua from diseased pyrethrum plants. Assessment of morphological and cultural characteristics showed considerable variability among P. ligulicola isolates from pyrethrum (n = 116) and chrysanthemum (n =5). Due to the observed variability and divergence from published descriptions of the two P. ligulicola varieties it was not possible to reliably differentiate between the two varieties on the basis of morphological criteria alone. The most notable discrepancy was that P. ligulicola var. inoxydabilis isolates did not produce the teleomorph (Didymella ligulicola var. inoxydabilis) in culture. The two P. ligulicola varieties were differentiated on the basis of the presence of metabolite E and minor divergence in the ribosomal DNA region of the internal transcribed spacer (ITS) region. Phylogenetic analysis of the translation elongation factor (EF1-α), ITS and glyceraldehyde-3-phoshpate dehydrogenase (G3PD) regions confirmed the identity of P. ligulicola and P. exigua isolates to species level, and suggested that the P. ligulicola var. inoxydabilis population from pyrethrum is largely clonal. The EF1-α and G3PD sequences did not clearly differentiate between P. ligulicola varieties, the host from which they were isolated or the geographic location from which they were sourced. Phoma ligulicola var. inoxydabilis isolates from pyrethrum (n = 111) were tested in vitro for sensitivity to the DMI fungicide difenoconazole and had a low mean EC₅₀ of 0.136 μg a.i./ml. The continuous sensitivity distributions detected for the P. ligulicola populations indicate that resistance development for this pathosystem is likely to be a gradual process. The mean EC₅₀ of four P. exigua isolates was 50 times greater than for P. ligulicola var. inoxydabilis isolates. A PCR assay for determination of mating type detected the HMG motif in the five P. ligulicola var. ligulicola isolates and in one of the 116 P. ligulicola var. inoxydabilis isolates. Individual isolates of P. ligulicola var. ligulicola produced perithecia and ascospores on leaf amended agar, but none were detected for P. ligulicola var. inoxydabilis. These results indicate that the former variety was homothallic, and the latter variety was heterothallic. The pathogenicity of 18 P. ligulicola and two P. exigua isolates to a single variety of pyrethrum was characterised in two separate experiments by inoculating detached leaves in vitro and glass-house grown plants. In the detached leaf assay significant differences were detected between isolates in lesion growth. There was a positive correlation between the lesion growth in the detached leaf assay and disease incidence in the greenhouse trial for both Phoma species. This is the first report that P. exigua var. exigua is pathogenic to commercial pyrethrum. Symptoms produced on eight week old pyrethrum plants in the greenhouse, inoculated with P. exigua conidial suspensions, were mid-brown leaf lesions up to 5 mm long, and occasional stem lesions.