Doctoral Degrees (Horticultural Science)

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Aspects of avocado fruit growth and development : towards understanding the 'Hass' small fruit syndrome.
(1997) Moore-Gordon, Clive Scott.; Cowan, Ashton Keith.; Wolstenholme, B. Nigel.
Persea americana Mill. cv. Hass is predisposed towards producing a high proportion of undersized fruit. Reasons for phenotypically small 'Hass' fruit are obscure, but it does appear to be aggravated by adverse growing conditions. A detailed study of the metabolic control of avocado fruit growth was carried out to determine the underlying physiological reasons for the appearance of the 'Hass' small fruit phenotype. Furthermore, the application of a mulch was evaluated as a possible management strategy to increase 'Hass' fruit size. Anatomical and morphological comparisons were made between normal and small 'Hass' fruit in an attempt to characterise the 'Hass' small fruit phenotype. Small fruit always contained a degenerate seed coat and fruit size was closely correlated with seed size. Kinetic analysis of changes in cell number and size during fruit development revealed that growth was limited by cell number in phenotypically small fruit. Analysis of endogenous isopentenyladenine (iP) and abscisic acid (ABA) revealed that ABA concentration was negatively correlated with size of similarly aged fruit. Calculation of the iP:ABA ratio showed a linear relationship with increasing fruit size. Qualitative and quantitative differences in mesocarp sterol composition were observed between normal and phenotypically small fruit. Both the normal and small-fruit phenotypes were used to probe the interaction between end-products of isoprenoid biosynthesis and activity of mesocarp 3-hydroxy-3- methylglutaryl coenzyme A reductase (HMGR) in the metabolic control of avocado fruit growth. In phenotypically small fruit, a 70% reduction in microsomal HMGR activity was associated with a substantial rise in mesocarp ABA concentration at all stages of development. Application of mevastatin, a competitive inhibitor of HMGR, via the pedicel reduced growth of phenotypically normal fruit and increased mesocarp ABA concentration. These effects were reversed by co-treatment of fruit with either mevalonate, iP or the synthetic cytokinin (CK) analogue, N-(2-chloro-4-pyridyl)-N-phenylurea, but were unaffected by gibberellic acid. Likewise, in vivo application of ABA reduced fruit growth and HMGR activity, and accelerated abscission at all stages of development, effects that were reversed by co-treatment with iP. In contrast, the effect of sterols on mevastatin-induced inhibition of fruit growth was temporally different. Application of either stigmasterol or cholesterol during phase I caused a decline in growth, accelerated fruit abscission and exacerbated the effects of mevastatin whereas during phase II and III, stigmasterol reversed inhibition of fruit growth. Stigmasterol did not however, reverse the inhibitory effect of mevastatin on HMGR activity - presumably as a result of mevastatin-induced increased endogenous ABA. It was therefore concluded that ABA accumulation downregulates mesocarp HMGR activity and that in situ CK biosynthesis modulates the effect of ABA during phase I of fruit growth whereas, both CK and sterols perform this function during the later stages to sustain the developmental programme. The effect of an altered CK:ABA ratio on solute allocation, cell-to-cell communication and plasmodesmatal structure was investigated in 'Hass' avocado fruits to determine the relationship between a change in hormone balance and expression of phenotypically small fruit. Exogenous application of ABA induced early seed coat senescence and retarded fruit growth, and these effects were negated in fruit co-injected with ABA and iP. The underlying physiological mechanisms associated with ABA-induced retardation of 'Hass' avocado fruit growth included: diminution of mesocarp and seed coat plasmodesmatal branching; gating of mesocarp and seed coat plasmodesmata by deposition of apparently proteinaceous material in the neck region; abolishment of the electrochemical gradient between mesocarp and seed coat parenchyma; and arrest of cell-to-cell chemical communication. In addition, solute allocation in ABA-treated fruit resembled closely that of phenotypically small fruit confirming that elevated ABA concentration had contributed to the decline in postphloem symplastic continuity. In a field trial in the KwaZulu-Natal midlands, root growth was substantially increased throughout three seasons by the application of a coarse composted pinebark mulch. Mulching resulted in a significant 6.6% increase in mean fruit mass, in spite of 14.7% more fruits per tree. The combined effect was a 22.6% increase in overall yield. Differences in productivity between treatments closely correlated to levels of bark carbohydrate reserves. Data collated during this study to suggest that mulching at least partly ameliorated tree stress included: a reduction in the incidence of premature seed coat senescence and pedicel ring-neck, both of which are considered to be advanced symptoms of the stress syndrome; a lowering of mean foliage temperatures; and a reduction in the degree of photoinhibition during the heat of the day.
Plant hormone homeostasis and the control of 'hass' avocado fruit size.
(2002) Taylor, Nicolette Jane.; Cairns, Andrew Lawrence Patrick.; Cowan, Ashton Keith.; Van Staden, Johannes.
The 'Hass' avocado produces two distinct phenotypically different populations of fruit, i.e. normal and small fruit. The small fruit variant is characterized by early seed coat senescence that results in arrested growth, due to dramatically reduced cell cycle activity. This system has been used to study the metabolic control of fruit growth for two reasons. Firstly, the 'Hass' avocado is a major export crop in South Africa and unmarketable small fruit cost the industry millions of rands per season. Secondly, in the absence of evergreen tree-crop mutants with which to dissect controlling mechanisms contributing to the control of final fruit size, the 'Hass' avocado and its small fruit variant provides an ideal system to investigate the physiology, biochemistry and molecular biology of fruit growth in subtropical species. A detailed study was conducted to probe the contribution of hormones in the control of final fruit size by comparing and contrasting tissue distribution and content of hormones in developing 'Hass' avocado and its small fruit variant. In addition the proposal that changes in hormone homeostasis occur as a result of differences in the allocation of the molybdenum cofactor (MoCo) and changes in the activity of xanthine dehydrogenase (XDH) and the aldehyde oxidases (AO) involved in abscisic acid (ABA) and indole-3-acetic acid (IAA) metabolism was evaluated. Activity of XDH, xanthoxal (XAN) oxidase, indole acetaldehyde (IA-aid) oxidase and cytokinin oxidase (CKOX) was related to tissue content and composition of IAA and ABA. Comparisons between normal and small fruit revealed that under conditions where CKOX is elevated, the increased adenine produced inhibits XDH activity, which leads to elevated activity of the AOs involved in ABA and possibly IAA biosynthesis as a result of increased MoCo allocation to these enzymes. Further analyses revealed that both cytokinin (CK) and auxin elevates CKOX activity and that adenine and CK do indeed inhibit XDH activity, which leads to increased AO activity. In addition, application of CK to normal fruit increased IAA in mesocarp tissue but reduced IAA content of seed tissue and reduced ABA in mesocarp tissue but had no effect on ABA in seed tissue. Cytokinin oxidase therefore contributes to the regulation of ABA and IAA metabolism during plant organ growth by modulating the activity of XDH. Low XDH and lA-aid oxidase activity together with high XAN oxidase and CKOX activity early in fruit development combine to reduce both elongation and radial growth, which results in the appearance of the 'Hass' small fruit phenotype. This event was associated with high ABA and low IAA in seed tissue of small fruit, but high ABA and IAA in seed coat and mesocarp tissue of these fruit. Thus, whilst low IAA in seed tissue is associated with reduced growth the . reverse is true in seed coat and mesocarp tissue where high IAA retards tissue growth. Calculation of CK/ABA and CK/IAA ratios revealed that a decrease in these ratios was found in mesocarp tissue of small fruit. However, in seed tissue of small fruit both IAA and ABA were decreased relative to CK. The maintenance of the correct hormonal balance in avocado fruit thus ensures the continuation of cell division cycle activity, with any changes responsible for the high incidence of a small fruit variant in the 'Hass' avocado.
Post-phloem transport and metabolism of sucrose in avocado.
(2001) Cripps, Ryan F.; Smith, Michael Trevor.; Cowan, Ashton Keith.
In South Africa, and in several other sub/tropical countries, the avocado represents a commercially important crop. Very little is currently understood about the metabolism of sugars in this fruit. The variety 'Hass' is a popular cultivar that is grown extensively in South Africa. However this. cultivar has a tendency to produce two distinct fruit phenotypes: a normal sized variant and a small, undersized variant. Current literature suggests that the small fruit phenotype is characterised by an elevated abscisic acid (ABA) to cytokinin ratio and altered isoprenoid metabolism. The results presented in the current investigation represent the findings from a detailed study into the metabolism and transport of sugars in 'Hass' fruit in an attempt to characterise solute allocation in developing avocado fruit. Furthermore, the activities of sugar metabolising enzymes, routes of solute movement and polyphenolic contents of normal, small and ABA-treated fruit were compared and contrasted to evaluate the potential role of ABA in the induction and expression of the small fruit phenotype. The enzymes invertase, sucrose synthase (SSy) and sucrose phosphate synthase are involved in the metabolism of sucrose (Suc) and, hence, phloem unloading, post-sieve element transport and fruit growth. Although not the major sugar present, Suc was found in avocado phloem sap, and the enzymology for its metabolism was shown to exist in avocado fruit. It appears that sink strength is established during early fruit growth by high acid invertase activity, especially during the period of rapid cell division. As fruit growth progresses the activity of SSy and an enzyme responsible for the oxidation of perseitol (tentatively termed perseitol dehydrogenase) increases, suggesting that these enzymes play an important role in the supply of carbon during the linear phase of fruit growth. All Suc metabolising enzyme activity diminishes as the fruit approaches maturity. With the exception of SSy (in the cleavage direction), all enzymes assayed showed a general increase in relative rates of activity in small and ABA-treated fruit. Similarly, ABA-treatment of seed coat discs in vitro resulted in the elevation of insoluble and soluble acid invertase, SSy (in the synthesis direction), and sucrose phosphate synthase activity. Furthermore, both small and ABA-treated fruit were characterised by elevated total soluble sugars, glucose and fructose levels. These observations suggest that altered sugar metabolism, as a consequence of changes in endogenous ABA levels, may contribute to the occurrence of the small fruit. The seed coat represents an import link between the seed, the mesocarp and the parental plant tissues. Loss of seed coat and endosperm integrity accompanied fruit maturation and a reduction in the movement of solutes into the seed. An increase in polyphenolics in the seed coat tissue seemed critical in this reduced movement. Both the small and ABA-treated fruit were characterised by early senescence of the seed coat, which was accompanied by both a loss of transport into and out of the seed and premature maturation of the fruit. This premature seed coat senescence appeared similar to programmed cell death in tissues exposed to stress or elevated reactive oxygen species, stimuli that are often accompanied by elevated ABA levels. Callose was localised to the plasmodesmata and is proposed to play a role in the gating of, and hence movement through, these pores. Small fruit were characterised by a loss of symplastic continuity, as represented by fewer plasmodesmata, and reduced callose degradation. Comparison of callose content and rates of synthesis suggest that ABA-treatment, similarly, reduces callose catabolism. The association of ABA with both the premature senescence of the seed coat and a reduction in symplastic continuity, and, hence, a reduction in solute transport, further cements the potential role of ABA in the occurrence of the small fruit phenotype.

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Browsing Doctoral Degrees (Horticultural Science) by Author "Cowan, Ashton Keith."