No description available.

Weed seeds contaminating cattle feed can pass unaffected through the animal's digestive tract and may germinate when manure is returned to land as fertilizer. This investigation was undertaken to determine the effects of anaerobic fermentation of raw manure on the viability of Johnsongrass and Fall Panicum seeds which may be present in the waste. Dairy waste containing Johnsongrass and Fall Panicum seeds were subjected to batch and continuously-fed anaerobic fermentation processes in laboratory-scale digesters. The effect of influent solids concentration (4% and 6%) and retention time (15 days and 20 days) on seed viability was observed. The experiments were carried out in the mesophilic temperature range [35°C±1ºC]. Fall Panicum seeds were found to be less resistant to anaerobic digestion than Johnsongrass seeds. Greater seed destruction was achieved in 20-day digesters than in the 15-day digesters. The influent solids concentrations did not have significant effect on seed viability. By applying the Schafer-Chilcote seed population model, it was observed that the fermentation process was more effective in destroying non-dormant seeds than dormant seeds. / Master of Science

Guayule (Parthenium argentatum Gray) is one of two major plants in the world grown for natural rubber and therefore, is of potential importance to the U.S. One area of research interest relates to methods of enhancing production of viable seeds. Therefore, studies were conducted in 1984, 1985, and 1986, in Tucson, Arizona to determine the effect of honey bee pollination on: seed set; seed weight; total seed yield; percentage of seed germination; yield and percentage of rubber and resin content. This experiment involved four guayule cultivars and four pollination treatments: plants caged with bees; plants caged without bees; plants open-pollinated; and plants individually covered with Delnet bags. All four cultivars responded positively to honey bee pollination. Plots with bees produced at least 195% more seeds than plots from which bees were excluded. However, there were no qualitative differences in the seed weights between the treatments. The percentage of seed germination from plots serviced by bees was significantly greater (65%) than from plots without bees (50%). Highest seed germination rates were obtained when seeds were collected in May (80%) and September (76%). June, July, and August seed collections resulted in lower seed germination rates (40%, 26%, and 63%, respectively). The plots in which bees were present gave a higher rubber yield (323 kg/ha) than plots without bees (255 kg/ha). However, there were no quantitative differences in the percentage of resin content between the treatments. These studies demonstrated that (1) honey bees can increase seed yield, seed germination, and rubber content in guayule, and (2) seeds produced during summer months had poorer germination rates and lower yields.

Typescript (photocopy). / Digitized by Kansas Correctional Industries

Bibliography: leaves 155-182. Most agricultural crop seeds are 'orthodox' ie. desiccation is a necessary feature of their complete life cycle. Low moisture content lengthens the storage life of orthodox seeds. Recalcitrant seeds, which do not tolerate low moisture content, lose viability in dry storage. The thesis studies the role of sugars in desiccation tolerance in developing seeds and investigates other mechanisms which might be involved in desiccation tolerance and desiccation sensitivity.

published_or_final_version / Botany / Master / Master of Philosophy

No description available.

Three questions are considered in the context of the possible effects of seedassociated mycoflora, typified by Fusarium moniliforme, during hydrated storage of recalcitrant seeds of the tropical species, Avicennia marina. These are: 1) whether fungal infection reduces storage lifespan; 2) whether seeds become more susceptible to fungal attack during storage and whether they posses defence mechanisms that might suppress fungal proliferation in hydrated storage (production of antifungal compounds and 13-1,3-glucanase (EC 3.2.1.39) and chitinase (EC 3.2.1.14)] and 3) whether it is possible to discriminate ultrastructurally between inherent deteriorative changes and those that are fungally-induced. 1) The data indicate unequivocally that if fungal activity is curtailed, then the hydrated storage lifespan of A. marina seeds can be considerably extended. 2) When inoculated immediately with F. moniliforme, newly harvested seeds were extremely susceptible to the adverse effects of the fungus, while seeds that had been wet-stored for 4 days showed a considerably heightened resilience to the effects of the fungus prior to inoculation. The enhanced resilience, although declining, persisted in seeds stored hydrated for up to 10 days prior to inoculation, being lost after 12 days. This finding was supported by significant increase in 13-1,3-glucanase and chitinase and in antifungal compound production during 10 days of wet storage. After 14 days of wetstorage, seeds become more susceptible to the effects of fungusthanthose in the newly harvested condition. 3) The resilience of seeds that had been stored in the short-term was associated with ultrastructural changes indicative of enhanced metabolic activity associated with the onset of germination (e.g. increase in vacuolation, well-developed mitochondria and endomembrane system [ER and Golgi bodies]). However, with sustained stress associated with wet-storage IV conditions, the seeds became increasingly badly affected by the fungus, showing some ultrastructural fungally-induced abnormalities (e.g. nuclear lobing, presence of lipid bodies and prevalence of Golgi bodies that had many associated vesicles) and a decrease in 13-1,3-glucanase and chitinase activity. It is suggested that the decreased susceptibility of A. marina seeds during short-term storage relies on the ability to create an antifungal environment prior to infection (through synthesis and accumulation of pre-formed and induced antifungal compounds and antifungal enzymes), which would also be an effective strategy during germination in the natural environment. / Thesis (Ph.D.)-University of KwaZulu-Natal, Durban, 2004

Achieving long-term storage of germplasm is critical for the conservation of plant biodiversity. Seed storage practices require that degradative reactions causing ageing be limited. By reducing the water content, cytoplasmic viscosity is increased to levels that minimise deteriorative reactions. Reducing the storage temperature additionally increases the storage lifespan by further reducing the rate at which such deleterious processes occur. Two broad categories of seeds can be distinguished based on their storage behaviour. Orthodox seeds are desiccation-tolerant; generally shed in the dry state and are metabolically quiescent. Such seeds are usually stored at low water contents (e.g. 5%), and their high cytoplasmic viscosity prevents freezing damage during cooling to subzero temperatures. On the other hand, desiccation-sensitive (recalcitrant) seeds do not undergo a maturation-drying phase, they are metabolically active at shedding, and sensitive to extreme or prolonged drying. Accordingly, recalcitrant seeds cannot be stored under conventional conditions because they do not survive drying to low water contents and are damaged by sub-zero temperatures, even when dried to the lowest water content tolerated. Therefore, procedures that facilitate harmless drying and cooling to low temperatures are required to achieve long-term storage of recalcitrant germplasm. Recalcitrant seeds that are dried rapidly can attain relatively lower water contents without injury. However, these seeds are usually large and this limits the drying rates that can be achieved even under favourable conditions. Isolating embryonic axes from the rest of the seed facilitates faster drying, and a consequent reduction in the water content at which damage occurs. In axes of many species, the level of drying attained before lethal desiccation damage occurs is sufficient to limit freeZing damage during cryogenic exposure and facilitate survival in vitro. However, many others are damaged when dried to water contents that preclude freezing, and also are killed if cooled to sub-zero temperatures at higher water contents. In such instances, the window of permissible water contents leading to survival may be small or nonexistent. A basic premise explored in this thesis is that by restricting the growth of intracellular ice crystals using increasingly rapid cooling rates, the range of permissible water contents can be widened, facilitating survival of axes at higher water contents. An overview of the problems associated with the long-term storage of recalcitrant germplasm, and the rationale behind such rapid cooling approach are presented in Chapter 1 of the present thesis. Subsequent chapters report investigations on the effects of variables required to dry and cryopreserve embryonic axes with minimum damage, in keeping with this approach. Collectively, those studies aimed at establishing a robust cryopreservation procedure for the conservation of recalcitrant germplasm with broad applicability across species. The approach presently adopted entailed manipulating the water content of excised axes using rapid drying to discrete water content ranges, and also using different methods to cool axes to cryogenic temperatures at various rates. The calorimetric properties of water in axes were investigated for Camellia sinensis (L.) O. Kuntze using differential scanning calorimetry (DSC). For all species, the effect of any drying or cooling treatment tested was determined by assessing the survival of axes in vitro, which provided the most reliable indicator of cellular damage. Additionally, the effects of different treatments upon the structural and functional integrity of axes were assessed using light and electron microscopy as well as measurement of electrolyte leakage. The studies undertaken are presented in a similar sequence to that in which they took place during the course of the experimental phase of this work. These are summarised below. Partial drying plays a pivotal role in the approach developed, and microscopy has contributed towards increasing present understanding of desiccation damage. Microscopy was used to determine the effects of drying rate upon the ultrastructure of recalcitrant axes. It was necessary to find reliable protocols to prepare specimens for light and electron microscopy that did not alter the architecture of the cells in the dry state. Freeze-substitution and conventional aqueous fixation were compared in Chapter 2 using variously dried material from three species. The results obtained revealed that an unacceptably high extent of artefactual rehydration occurs during aqueous fixation, and highlight the need for anhydrous processing of dehydrated samples. Significantly, that study also revealed that many cellular events commonly associated with desiccation damage (e.g. withdrawal, tearing and/or vesiculation of the plasmalemma) are not seen in freeze-substituted preparations, and are likely artefacts of aqueous fixation. Freeze-substitution was used subsequently (Chapter 3) to assess the effects of slow drying (2 - 3 days) or rapid drying (min) upon the survival of embryonic axes of jackfruit (Artocarpus heterophyllus Lamk.) Results confirmed the beneficial effects of rapid drying, and also provided insights into ultrastructural changes and probable causes underlying cellular damage that occur during a drying/rehydration cycle. Efforts subsequently focused on determining the effect of cooling rate upon survival of recalcitrant axes at various water contents. The study on embryonic axes of recalcitrant camellia sinensis (tea; Chapter 4) tested the hypothesis that rapid cooling facilitates survival of axes at higher water content by restricting the growth of ice crystals to within harmless dimensions. The presence of sharp peaks in DSC melting thermograms was indicative of decreased survival in vitro. These peaks were attributed to the melting of ice crystals sufficiently large to be detected by DSC as well as to cause lethal damage to axes. Increasing the cooling rate from 10°C min-1 to that attained by forcibly plunging naked axes into sub-cooled nitrogen increased the upper limit of water content facilitating survival in vitro from c. 0.4 to 1.1 - 1.6 g H20 g-1 (dry mass [dmb]). Subsequent studies tested whether a similar trend occurred in other recalcitrant species cooled under similar conditions. In order to investigate further the relationship between water content, cooling rate and survival it was necessary to achieve cooling rates reproducibly, and to quantify these reliably. The plunging device required to achieve rapid cooling, and instruments required to measure the cooling rates attained, are described in Chapter 5. That study investigated the effects of cryogen type, depth of plunge and plunging velocity on the cooling rates measured by thermocouples either bare or within tissues of similar size and water content as encountered in cryopreservation experiments. This plunger was used in subsequent studies to achieve the fastest cooling conditions tested. Favourable cooling conditions were selected, and the efficacy of this procedure to cryopreserve relatively large axes was tested (Chapter 6) using embryonic axes of horse chestnut (Aesculus hippocastanum L.) Axes at water contents above c. 0.75 g g-1 could not be cooled faster than c. 60°C S-1, but cooling rates of axes below this water content increased markedly with isopentane, and to a lesser extent with subcooled nitrogen. Axes were killed when cooled at water contents above 1.0 g g-1 but survived fully (albeit abnormally) when cooled in isopentane between 1.0 and 0.75 g g-1. Complete survival and increasingly normal development was attained at water contents below 0.75 g g-1, especially if isopentane was used. The study on horse chestnut axes emphasised that water content and cooling rate are co-dependent during non-equilibrium cooling. Accordingly, that study could not determine whether survival at lower water contents increased because of the corresponding increase in cooling rates measured, or because of the higher cytoplasmic viscosity that resulted from drying. That uncertainty was addressed by the study discussed in Chapter 7, using axes of the trifoliate orange (Poncirus trifoliata [L.] RAF.) That study investigated the effect of cytoplasmic viscosity upon survival of axes cooled and warmed at different rates. Survival and normal development was high at lower water contents, and seemingly independent of cooling rate at about 0.26 g g-1. At higher water contents the range of cooling rates facilitating survival became narrower and displaced towards higher cooling rates. This study revealed two conspicuous inconsistencies that questioned the beneficial effect of rapid cooling. Firstly, the fastest cooling rates did not necessarily facilitate the highest survival. Secondly, survival of fully hydrated axes was higher when cooled under conditions that encouraged - rather than restricted - the growth of intracellular ice crystals. These inconsistencies were explored further using embryonic axes of silver maple (Acer saccharinum L.). Freeze-fracture replicas and freeze-substitution techniques provided valuable insights into the way in which ice crystals were distributed in cells cooled using different methods at rates ranging between 3.3 and 97°C S-1. Extensive intracellular freezing was common to all treatments. Unexpectedly, fully hydrated axes not only survived cryogenic exposure, but many axes developed normally when cooled using the relatively slower methods (77 and 3.3°C S-1) if warming was rapid. The most conspicuous ultrastructural difference between plunge cooling and the relatively slower methods was the exclusion of ice from many intracellular compartments in the latter. It is possible that even the fastest warming cannot prevent serious cellular damage if ice crystals form within such 'critical' compartments. It is proposed that the intracellular location of ice is a stronger determinant of survival that the size attained by ice crystals. The study of A. saccharinum also investigated the recovery of axes cooled fully hydrated either rapidly (97°C S-1) or slowly (3.3°C S-1). This facet of the study showed that cell lysis became apparent immediately after warming only where damage was most extensive. In other cells damage became apparent only after 2.5 to 6 h had elapsed, thus cautioning against inferring survival from the ultrastructural appearance of cells immediately after warming. Microscopy enabled cell repair as well as the pattern of growth of cryopreserved tissues to be appraised at the cellular, tissue and organ levels. Similar studies are required to understand further the nature of freezing damage, and how those events affect cell function. The salient trends observed in previous chapters are brought together in Chapter 9. / Thesis (Ph.D.)-University of Natal, Durban, 2002.

Desiccation-sensitive seeds show differential viability characteristics during drying at different rates. A number of studies have demonstrated that rapid dehydration permits survival to lower water contents than does slower desiccation. The aim and objective of the present study was to test the hypothesis which states that rapid drying of desiccation-sensitive seeds removes water sufficiently fast to reduce the accumulation of metabolic damage. In addition, the hypothesis that wet storage subjects desiccation-sensitive seeds to mild, but increasingly severe, water stress causing oxidative damage if additional water is not supplied, was tested. In the present study, axes of germinating orthodox seeds of Pisum sativum and newlyshed recalcitrant counterparts of Quercus robur, Strychnos madagascariensis, Trichilia emetica, Trichilia dregeana and Avicennia marina were subjected to rapid or slow drying or wet storage. For those species where more than one harvest was investigated, differences were observed in water contents at shedding. For all the species studied, the dehydration rate could be described by an exponential and a modified inverse function for both desiccation regimes, and the water content remained constant with wet storage. The level of tetrazolium staining and germination percentage of axes decreased sharply drying and hydrated storage such that the marked decline took place at lower water contents upon rapid than slow dehydration. The conductivity of electrolyte leachate increased progressively during desiccation and moist storage of axes of all species investigated. Greater membrane leakage occurred upon slow, than rapid dehydration in axes of all species studied. Activities of respiratory enzymes which have a potentially regulatory role in glycolysis, phosphofructokinase (PFK), or the tricarboxylic acid cycle, malate dehydrogenase (MDH), and levels of the oxidized form of the coenzyme, nicotinamide adenine dinucleotide (NAD), of the enzymes of the electron transport chain, NADH dehydrogenases ofNADH-ubiquinone (coenzyme Q) reductase (complex I) and NADHcytochrome c reductase (complex IV), were monitored in the present investigation. v In addition, the role of free radical activity in the form of lipid peroxidation, which has been implicated in loss of viability in seeds, was examined by assaying the levels of hydroperoxides. The involvement of the free radical processing enzymes, superoxide dismutase (SOD), catalase (CAT) and glutathione reductase (GR), and the antioxidant, ascorbic acid (AsA), was also ascertained. The activity of PFK in axes of P. sativum remained constant during drying and wet storage. However, PFK activity increased as rapid dehydration and hydrated storage of Q. robur axes proceeded. In contrast, the activity of PFK in axes of Q. robur decreased during slow desiccation. Similarly, PFK activity was reduced upon drying, and moist storage, of T. dregeana axes such that higher activity of PFK was seen during rapid than slow dehydration. The activity ofPFK inA. marina axes also declined upon desiccation. The activity ofMDH in axes of P. sativum was also unchanged during drying and wet storage. However, an increase in MDH activity was recorded in Q. robur axes during dehydration and hydrated storage such that the activity of MDH was higher upon slow than rapid desiccation. In contrast, MDH activity in axes of T. dregeana decreased as drying proceeded. Similarly, the activity of J\.1DH declined during dehydration and moist storage of A. marina axes. An increase in the level of NAD occurred in axes of P. sativum during drying. In contrast, a decrease in NAD levels was seen upon dehydration and wet storage of Q. robur axes such that the level of NAD was higher upon rapid than slow desiccation. There was an enhancement of the level of NAD in axes of T. dregeana during hydrated storage. Conversely, NAD levels declined during drying ofA. marina axes. A decrease in the level of hydroperoxides in axes of P. sativum was seen as rapid drying proceeded. In contrast, hydroperoxide levels increased during wet storage of P. sativum axes. Similarly, the levels of hydroperoxides were enhanced upon dehydration and hydrated storage of Q. robur axes such that they were higher in axes during slow desiccation compared to those dried rapidly. Conversely, the hydroperoxide level in axes of T. dregeana was reduced upon rapid dehydration. In contrast, an elevation of the level of hydroperoxides was observed during moist storage. The levels of hydroperoxides remained constant as desiccation and wet storage ofA. marina axes proceeded. vi The activity of SOD in axes of P. sativum decreased during rapid drying. In contrast, SOD activity increased upon slow dehydration and wet storage ofP. sativum axes. There was a decline in the activity of SOD in Q. robur axes during slow desiccation. Similarly, SOD activity was diminished upon drying of axes of T. dregeana. The activity ofSOD in T. dregeana axes was enhanced during hydrated storage. An elevation in SOD activity also took place during rapid dehydration and moist storage of axes ofA. marina. The activity of CAT did not change during drying of axes of P. sativum. However, a decrease in CAT activity in Q. robur axes was seen upon slow dehydration and wet storage. Similarly, the activity of CAT declined as desiccation of axes of T. dregeana proceeded. In contrast, CAT activity inA. marina axes increased during slow drying. Whereas the activity of GR in axes of P. sativum increased during drying and wet storage, GR activity decreased in A. marina axes upon all treatments such that the activity ofGR was higher during rapid than slow dehydration. GR activity also declined upon slow desiccation and hydrated storage ofaxes of Q. robur. Similarly, the activity of GR in T. dregeana axes was reduced during moist storage. Finally, a decrease in the level of AsA in axes of P. sativum took place during drying. Nonetheless, dehydration and wet storage of Q. robur axes were associated with no siginificant change in AsA levels. There was also a decline in the level of AsA in axes of T. dregeana as rapid desiccation proceeded. Similarly, a reduction in AsA level occurred upon slow drying ofaxes ofA. marina. The results presented here are consistent with the observation that drying and wet storage adversely affected the respiratory enzymes, PFK, MDH and NADH dehydrogenase. It is suggested that the resultant metabolic imbalance led to more leakage of electrons from the mitochondrial electron transport chain than normal, and through lipid peroxidation increased levels of hydroperoxides. In addition, dehydration and hydrated storage may depress the activities of free radical processing enzymes, SOD, CAT and GR and levels of antioxidant, AsA. This phenomenon was less pronounced during rapid, in comparison to slow, desiccation and moist storage. However, it appears that the above biochemical events are overtaken by physical damage at higher water contents in the highly recalcitrant seeds. It was concluded that the differential effects of VII the drying rate and wet storage on responses of desiccation-sensitive seeds varies with tissue, harvest, species and the degree of desiccation sensitivity. / Thesis (Ph.D.)-University of KwaZulu-Natal, 2004.