Thesis (DScAgric)--University of Stellenbosch, 1958. / OBJECTIVE. High economic production has ever been the aim and aspiration of the agriculturist and no less that of the fruit farmer. In striving towards this aim the latter has for a long time been at a disadvantage with regard to control of his nutritional programme. Even on naturally fertile soil, the question continually arises as to what the correct fertilizer treatment should be to maintain high productivity and how such a decision can be arrived at. A satisfactory answer to these questions could have been obtained from fertilizer trials if it was not such a difficult matter, in view of the extensive and long-term nature of such trials with fruit trees, to establish a sufficient number for each fruit species on different soil types and under different climatic conditions. Efforts to find a new approach to the problem have turned attention to the plant itself and its chemical make-up as affording the best index of its nutritional requirements. Intensive work in this direction has resulted in the evolution of a new tool in agriculture, the technique of diagnostic leaf analysis or 1Toliar diagnosis" as originally proposed by Lagatu and Maume in France and Thomas in u.s.A. A review of the literature is presented indicating the prodigous amount of research which has been applied to studies of the relationship between plant response and nutrient supply in terms of plant composition. Agriculturists have been quick to recognize the potentialities of leaf analysis as a practical guide in nutritional problems and advisory services based on foliar analysis have already been established for certain crops overseas. The experimental basis for formulating such a scheme for deciduous fruit in the Western Cape Province is provided by the factual evidence presented in this thesis.THE TECHNIQUE. The technique of diagnostic leaf analysis comprises sampling of leaves, preparation of sample for analysis and the analysis itself followed by interpretation of the analytical results by comparison with previously determined nutritional standards. Numerous factors were found to influence the final composition of the leaf sample as determined by analysis, such that strict adherence to a standardized procedure through all phases of sampling and preparation of leaf samples for analysis is required to eliminate or reduce errors likely to cause misleading interpretations. Experimental data are presented suggesting how the leaf sample should be selected on a tree and how it should be handled, cleaned, dried, ground and stored to reduce sampling and other errors. The final procedure as adopted eliminates most of the potential sources of experimental error but two unavoidable sources of e~ror remain to be accounted for, that due to tree variation and seasonal effect. The variation in leaf composition from tree to tree was found to be very considerable, so that aampling from a large enough group of trees (6 to 10) to reduce the error involved is essential in order to obtain leaf data which correctly reflects the nutrient status of the portion of the orchard concerned. Secondly, on the grounds of marked consistency found in different fruit species as to seasonal and year to year variation in mineral nutrient concentration, correction factors have been formulated and are suggested as a means of overcoming these sources of error. THEORETICAL BASIS. A diagnosis of the nutrient status in terms of the analytical results as finally determined is obtained by comparison of the data with previously established leaf composition standards of reference and by correct interpretation of the deviations from these standards. The theoretical basis for setting up these index values is discussed. The criterion used is based on the concept of Optimum Values which aaequately integrates the known relationships between plant response and nutrient supply in terms of internal nutrient concentration. A modification of this concept is proposed to the effect that for maximum growth and yield there exists an optimum range of nutrient concentrations with upper and lower limits for each of the functional elements, and that within this range the interrelationship between the individual nutrient elements is also optimal. Since no local fertilize~ trials with deciduous fruit trees are available and only one for grapes, data from highly productive plants in commercial orchards and vineyards were used to determine the upper and lower limits of the "optimum range", on the following premise. If leaf analysis data are available from a sufficient number of high performance orchards in different localities representing a wide range of nutrient supply and environment, the highest and lowest values obtained may be considered to represent a close approximation of the limits of the range required for optimum performance. It is contended that index values obtained in this way must be of practical value in assessing the nutrient status of fruit trees. It is further postulated that the lower limits for the micro-nutrients and even for magnesium may be justifiably adjusted according to the concentration levels associated with symptom expression. INDEX VALUES. The necessary data for determining standards of leaf composition were obtained from leaf analysis surveys of orchards and vineyards and from a grape fertilizer experiment in the Western Cape Province. Visual symptoms of prevailing nutritional disorders are described (supplemented by photographic illustrations) and their relation to leaf composition indicated. Tentative index values have been determined on the basis indicated for each fruit species, apple, pear, peach, apricot, plum, prune and grapes. These nutritional levels comprise upper and lower limits for the nutrients N, P, K, Ca, Mg 1 Mn 1 Fe and Cu, as well as the upper limits for B and Na. DIAGNOSTIC INTERPRETATIONS. Assessment of the nutrient status in terms of these index values suggests that many orchards and vineyards in the Western Cape Province, particularly prune, apricot and grapes, are suffering from malnutrition in some form and are likely to show a marked response to nutritional treatment as suggested by foliar diagnosis. The use of diagnostic leaf analysis constitutes an important advance in dealing with orohard problems in that an immediate decision is possible regarding nutrient status and related aspects such as selection of suitable sites for fertilizer trials and adjustment of the fertilizer programme. / AFRIKAANSE OPSOMMING: geen opsomming

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The shoot apex of rice, Oryza sativa CV Balilla was studied with a view to obtaining information about the meristematic activity of the different regions. The vegetative state only was studied and the rice was grown for 4-5 days at 30°C or 35°C under 4cm water in darkness, intermittent white light or continuous white, red, far-red or blue light. The part of the shoot apex studied was that above the level of the youngest leaf primordium and it was divided into six regions which took into account the tunica and corpus and regions of leaf initiation. The mitotic index was found every 2hrs. throughout 24hrs. in plants growing in continuous darkness, in a 12hr. day with white light of 10,000 lux and in continuous white light of 2,400 lux. The mitotic index varied little and no pronounced synchrony was evident under any of these conditions. The mitotic index of the flank and corpus regions was higher than that of the summit and sub-summit regions. The meristematic activity of the regions was investigated further using colchicine to accumulate metaphases. Colchicine blocks mitosis at metaphase, but does not affect the rate of entry of the cells into mitosis. The gradient of the increase in metaphases over 3-4 hours was used to calculate the cell-doubling time in each region and the effects of different environments on this were found. Cells of the summit region were found to divide at least 7 times more slowly than cells of the flanks and corpus and intermediate rates occurred in tie sub-summit regions. In 10,000 lux white light and at 30°C, the cell-doubling time for flanks and corpus averaged 13 hours which is shorter than any reported for equivalent regions in other plants. Cell-doubling times for the summit were over [illegible] hrs. In experiments where the intensity of white light varied during the growth of the seedlings a different relationship between the regions was found. The cells of the flanks and corpus divided more slowly and only about twice as fast as in the summit. The effect on cell-doubling times of excision on the shoot was studied and the immediate result was to speed up cell cycles in the leaf primordium and corpus. Excision has often been used to facilitate entry of a chemical into the shoot apex and it may have given a misleading picture of the situation in undisturbed shoot apices. Nuclear DNA synthesis in the regions was studied using tritiated-thymidine. In 1,000 lux continuous white light the flanks and corpus showed a gradual increase in labelling throughout the feeding which lasted for 72hrs. while the sub-summit and summit regions showed a negligible or low level of labelling in the same period of time. The level of DNA synthesis taking place in the regions thus paralleled that of meristematic activity found by using colchicine. The supply of tritiated thymidine to plants grown in continuous white light at 10,000 lux produced a non-linear low labelling pattern which could not be easily explained. Many attempts were made to overcome this lack of success involving excision, and surgery of various kinds, use of (6-³H) thymidine as well as (methyl-³H) thymidine and experimentation with red, far-red and blue light. However, none of those succeeded and so a pulse-labelling experiment was not attempted. The effect of an acute dose of X-rays was studied by finding the mitotic index immediately afterwards, andfrac12;hr. afterwards and 24hrs. afterwards and by studying the level of metaphases after colchicine in control and irradiated plants. A dose of 1, 2, 4 and 8 k.rad had a differential effect on the regions which is interpreted as a reflection of their different levels of meristematic activity. No stimulation of division of the summit was found which is the reverse of what has been found in the quiescent centre of roots after acute X-irradiation, although this region in often equated to the summit of the shoot. A study of cell pattern throughout a plastochron suggested on the basis that evidence of cell division and cell size are related, that meristematic activity in the tunica is highest in the flanks and that, as the activity progresses round the circumference of the apex in the formation of the leaf primordium, which sheaths the stem, cell division also takes place for a short distance toward the summit. In this way, preparation for formation of a leaf primordium occurs sometimes before it appears as a bump. The summit is considered to be carried forward passively and not to contribute a significant number of cells to the rest of the meristem. This interpretation supports the view that the summit is not the site of apical initials in higher plants as considered by the orthodox view. A study of the effect on cell kinetics of red and far-red light by using accumulation of metaphases with colchicine revealed a change in response to far-red light with the age of the seedling. Forty hrs. red and far-red light given to dark-grown 3andfrac12; day-old seedlings caused increased meristematic activity to take place in all regions but 16hrs. far-red light reduced activity and 16hrs. red light was stimulating. Forty hrs. far-red light after 2andfrac12; days dark gave high rates similar to 4andfrac12; day-old plants given 16hrs. far-red. Comparison of these rates on the basis of the age of the seedling at the end of the treatment revealed that plants which are 5 days old had similar rates after 16 or 40hrs. far-red, but plants which were 4 days old had higher rates after 40hrs. far-red than after 16hrs. far-red. Variation of response to far-red with age of the seedling has been found before and the link between phytochrome and leaf initiation is discussed. All regions of the apex, including the summit respond to light by a speeding-up of mitotic cycles but the summit is still at least seven times slower than the flanks and corpus. Finally, consideration is given to the significance of the results in rice and the mandeacute;ristandegrave;me d'attente controversy. Cell divisions in the summit may vary in importance in different apices but in the absence of the possibility of proving presence or absence of apical initials the controversy is put aside. The shift of growth balance in apices of plants subjected to different conditions is brought out and the importance of the summit to growth of the shoot apical meristem and the nature of its influence is discussed.

For the finite element (FE) simulation of relatively thin organs under complex dynamic loadings that are relevant in the biomedical engineering field, shell elements, compared to volume elements, have the potential to capture the whole thickness of the organ at once. Shell elements, are also known to feature efficiently large critical time steps, ensuring competitive computational times in dynamic structural analysis projects. As an improvement to the tools available for modelling and analysis, a new general nonlinear thick continuum-based (CB) shell FE embedded in an updated Lagrangian formulation and an explicit time integration scheme was recently developed. It can account for irregular and complex geometries, and hyper-elastic, large, nearly incompressible anisotropic 3D deformations characteristic of soft tissues. The original proof of concept was developed in MATLAB, which despite known advantages, is very slow. As a result, computational times, even for simple problems, have not been competitive. Therefore, the present work focused on re-writing the code in an efficient programming language with execution speed in mind in order to compete with the available elements which, in spite of having inferior capabilities, have better running times. In addition, a programming algorithm was needed to improve running time. Once it was implemented, the running time was reduced in half on a benchmark problem. Optimization was then exploited to introduce workarounds and design improvements that reduced running time further to 95% of its original value. The new version of the code was implemented in C++ and reached the goal of reducing running time while maintaining the expected functionality.

Prior to the commencement of this study in 1990, a number of reports had appeared in the literature describing the importance of insulin action during lactation in mammals (see Chapter 1). These studies investigated the changes in circulating insulin and glucagon concentrations during lactation, the relative numbers of insulin receptors in insulin-sensitive tissues, and glucose utilisation by these tissues. However, at that time, no information was available on the structure of the mammary insulin receptor. The rationale for undertaking this study was to characterise the structure of the rat mammary insulin receptor as a means of furthering our understanding of the role insulin plays during lactation.An initial requirement of this study was the development of a method for the convenient and inexpensive preparation of A14-tyrosyl[125I]iodoinsulin. A14-tyrosyl[125I]iodoinsulin displays binding characteristics which are virtually indistinguishable from the native hormone, which is a necessary requirement for tracers which are to be used in binding studies. In Chapter 2, I describe a method for the purification of A14-tyrosyl[125I]iodoinsulin from a mixture of iodinated insulin molecules which are produced following oxidation by chloramine-T in the presence of Na125iodine. In this method I employed disposable cartridges packed with a C18 support matrix to which the iodinated insulin molecules are readily adsorbed when in an aqueous solution.A 14-tyrosyl[125I]iodoinsulin absorbed most strongly to the C18 matrix and unwanted products were removed through a sequence of washes prior to the elution of the A14-tyrosyl[125I]iodoinsulin derivative using a buffer containing 50% (v/v) acetonitrile. This prodct was unambiguously shown to be A14-tyrosyl[125I]iodoinsulin by N-terminal amino acid sequencing. The quality of this radiolabel compared favourably with commercially ++ / available A14-tyrosyl[125I]iodoinsulin preparations both in terms of specific activity and stability upon storage at -20C. Furthermore, a modified method based on this protocol has been used in our and other laboratories for the isolation of other iodinated peptides with highly satisfactory results.I have established that the size of the a-subunit of the rat mammary insulin receptor is significantly diminished compared with the liver insulin receptor (125 kDa versus 130 kDa). This difference in size was present throughout all stages of lactation and was not due to proteolysis of a larger form. Furthermore, I demonstrated that both the mammary and liver insulin receptor a-subunits migrated equally on PAGE following treatment with neuraminidase, indicating that the apparent size difference may be accounted for by a variation in the extent of receptor sialation. Treatment of the mammary insulin receptor a-subunit with glycopeptidase F demonstrated that the size of the aglycoreceptor (100 kDa) was similar to that described for insulin receptors from other insulin-sensitive tissues.I characterised the distribution of mRNA encoding the two, naturally-occurring insulin receptor isoforms in mammary tissue throughout all stages of pregnancy and lactation. These insulin receptor isoforms differ due to the absence (IR-A) or presence (IR-B) of a 12 amino acid peptide, encoded by exon 11 of the insulin receptor gene, and located near the C-terminus of the insulin receptor a-subunit. Mammary tissue predominantly expressed IR-A mRNA in contrast to liver tissue, which almost exclusively expressed IRB mRNA. Furthermore, the ratio of IR-A to IR-B mRNA in mammary tissue changed significantly during the first week post-partum whilst the distribution of IR-A and IR-B mRNA in the liver remained constant throughout pregnancy and lactation. This difference in insulin receptor isoform ++ / expression between mammary and liver tissue also contributed to the estimated size difference between the insulin receptor a-subunits from these two tissues. In addition, I characterised the expression of IR-A and IR-B mRNA in several different tissues obtained from rats on day 14 of gestation through to 7 days post partum. I established that the splicing mechanism is functional at least as early as day 14 of gestation, suggesting a possible role for the preferential expression of a particular insulin receptor isoform during organogenesis. I observed that IR-A mRNA was the predominant isoform in all foetal tissue studied, and the proportion of this isoform declined as the animal matured. These changes were significant in cardiac muscle, kidney and most dramatic in the liver where the expression of IR-A mRNA changed from 53% in the 21 day old foetus (the day before parturition) to 13% in the 1 day old neonate. These results suggest that the splicing mechanism which generates the receptor isoforms is subject to acute hormonal and/or metabolic control.The current literature suggests that the carbohydrate moieties of the insulin receptor affects its affinity for insulin. Furthermore, the IR-A and IR-B isoforms have been shown to display a 2-fold difference in their insulin binding affinity when expressed in heterologous cell lines such at CHO cells or Rat-1 fibroblasts. Since both glycosylational and isoform distribution differences were evident between mammary and liver tissues, the insulin binding affinities of these receptors were compared. Estimates of the binding affinity parameters were performed at both 4 C and 37 C. At both temperatures the equilibrium binding constants for mammary and liver tissues were not significantly different suggesting that structural variations of the mammary insulin receptor had no effect on the insulin binding affinity under the ++ / conditions described in this study. Comparison of the 4 C and 37 C binding data showed that the mammary insulin receptor exhibited complex, temperature-dependent binding characteristics, similar to those previously described for the liver insulin receptor, and entirely consistent with the presence of a temperature-dependent regulatory protein that affects insulin binding.

Typescript (photocopy)

Graduation date: 2004

Graduation date: 1996

Inaug.-Diss.--Basel. / Includes bibliographical references.