It has been postulated that the absence of human and cattle schistosomiasis in parts of southern Africa where lechwe antelope (Kobus leche) occur is a consequence of an immunologically-mediated protection induced by repeated exposure to the cercariae of Schistosoma margrebowiei and S. Leiperi, which are common parasites of these animals. The aim of the studies described was the development of animal models in which to investigate this hypothesis. The infection characteristics of the antelope schistosomes in BALB/c mice and Mastomys Coucha were assessed. Both schistosome species reached full patency in these hosts, although S. Margrebowiei infections deteriorated rapidly in M.Coucha. While they differed markedly in terms of egg production rates and preferred sites of tissue egg deposition, both species caused severe hepatosplenomegaly and portal hypertension in the mouse model. Modulation of the granulomatous responses to ova in the tissues was demonstrated. Mice harbouring mature antelope schistosome infections displayed strong partial resistance to challenge infections with both homologous parasites and the human schistosome, S. Mansoni. However, the failure of challenge parasites to become established was considered to be due largely to changes in the portal-hepatic vasculature resulting from egg-induced immunopathology. Resistance to S. Mansoni challenge did not develop in mice infected with radiation-attenuated cercariae of the antelope schistosomes. The suitability of rats and guinea pigs as alternative models was assessed. Worm recoveries from rats were low and there was no evidence of egg-deposition. Worm yields from the guinea pig were relatively high, but sexual development was poor and short-lived. Since excretion of S. Margrebowiei eggs has occasionally been reported from humans, and since the guinea pig supports full sexual maturation of S. Mansoni, this animal appeared to provide a particularly appropriate model for the present investigation. However, repeated exposure of guinea pigs to cercariae of the antelope schistosomes, over a period of 24 weeks, failed to induce significant resistance to S. Mansoni challenge infection. The need for further experimental and field studies is discussed. An area in the Okavango Delta (Ngamiland, Botswana) has been identified as a possible site for field work. / Thesis (M.Sc.)-University of Natal, Durban, 1992.

The wood anatomy, hydraulic properties and leaf water status of five sub-tropical plant species were studied. The specimens studied were growing in a private, irrigated garden. Consequently, any differences in xylem anatomy would be a result of phylogenetic and not environmental factors. Podocarpus latifolius, being a gymnosperm, had only narrow, short tracheids as the conducting conduits. The size of the vessels of the four angiosperms increased in the following order: Tecomaria capensis, followed by Cinnamomum camphora, Trichilia dregeana and finally Barringtonia racemosa had the widest vessels. T. capensis and T. dregeana had the shortest vessel lengths. Those of C. camphora were slightly longer and the vessels of B. racemosa were the longest. Measurements of hydraulic conductance (Kh), twig specific conductivity (TSC) and xylem specific conductivity (XSC) followed similar trends to those of conduit diameters. The measurements of leaf specific conductivity (LSC) , however, did not follow these trends. T. dregeana, which had a far higher Kh than P. latifolius, did not have a significantly different LSC. This is because the twigs of T. dregeana supported a far greater leaf area than did the twigs of P. latifolius. There was also no significant difference in LSC among T. capensis, C. camphora and B. racemosa, although their LSC's were all significantly higher than those of P. latifolius and T. dregeana. The consequence of efficient xylem anatomy thus seems to be, not only a greater supply of water to the leaf but also, and perhaps more importantly, it allows a greater leaf area to be produced. The length of the vessels was also shown to have a large effect on the hydraulic conductivity of the twigs. The Kh values measured on excised twigs were found to range between 40% and 87% of the Kh calculated using the Hagen- Poiseuille equation. Conduit size distributions were also found to be important in calculating the Kh. The more efficent xylem anatomy of B. racemosa resulted in little decrease in plant water potential even with large increases in transpiration. P. latifolius on the other hand showed a considerable decrease in leaf water potential with just a slight increase in transpiration. The other three species showed decreases in leaf water potential inter-mediate to these two extremes. The inferred root-to-leaf condutivity, shown by the inverse of the slopes of the water potential versus transpiration curves, were lower than the LSC measurements taken on excised twigs in the laboratory. The difference between the inferred and the measured LSC's could give an indication of resistances such as those within the root and at the soil-root interface. / Thesis (M.Sc.)-University of Natal, Durban, 1991.

In vitro cell and tissue culture systems were developed for sugarcane in order to utilise current transformation techniques to introduce genes to South African sugarcane varieties, which would be difficult, if not impossible to achieve in conventional breeding programmes. Embryogenic calli were initiated in the dark from stem explants of sugarcane varieties NCo376 and N13, on a MS medium containing sucrose (20-50 g/l), 2,4-D (2-4 mg/l), casein (1 g/l), inositol (100 mg/l) and agar (9g/l). After 2 months the somatic embryos were cultured in a light/dark photoperiod for a further 2 months. The best combination of sucrose and 2,4-D for callus initiation, and subsequent plant regeneration, was 20 g/l and 2 mg/l, respectively. Plant yields ranged from 16 to 36 plants per gram fresh weight callus, and the yields were not significantly increased by the addition of activated charcoal to the regeneration medium. When plantlets reached a height of 10 cm, they were transferred to autoclaved soil in pots, hardened-off and placed in the glasshouse. Suspension cultures were initiated from friable NCo376 calli in liquid MS medium shaken at 100 rev/min in the dark at 27°C, and were subcultured every 3-7 days. Protoplasts from various sources (leaf, calli and suspension cultures) were obtained after enzymatic digestion in cellulase (20-30 g/l), macerozyme (0,2 g/l), hemicellulase (5 g/l), and sorbitol (0,55 M) in a calcium and magnesium salt solution. Protoplasts cultured for 48 h resulted in a loss in viability of 84%. The potential of the seed as a recipient for direct gene uptake was investigated, as this eliminated the need for in vitro culture and plant regeneration. Uptake of [3H] pBR322 DNA by seeds was demonstrated, and seeds with the testa removed exhibited higher initial uptake rates than those with intact seed coats. However, transient expression, using the GUS reporter gene (coding for bacterial B-glucuronidase) carried on plasmid pBI221, could not be conclusively shown using the histochemical GUS assay, due to GUS activity generated by either microbial contamination or endogenous plant GUS activity. Neither microwaving to eradicate contaminants nor the addition of methanol (20%) to the GUS incubation buffer were successful in overcoming positive results observed in control seeds. An alternative approach to sugarcane transformation, using PEG-mediated DNA uptake and subsequent transient expression of GUS by protoplasts was investigated, but microbial contamination was a persistant problem and no positive results were observed. Further examination and elimination of endogenous contamination is required before transformation studies can be continued. / Thesis (M.Sc.)-University of Natal, Durban, 1992.

The technique of restriction fragment length polymorphism (RFLP) analysis, of chloroplastic and genomic DNA, was investigated as a means of identifying eucalypt species and cultivars which are morphologically indistinguishable from one another. In order to resolve chloroplast DNA (cpDNA) RFLPs, a method was developed to extract high yields of intact chloroplasts from Eucalyptus grandis S/N M6. Starch contamination was reduced by incubation of saplings in the dark for 48 h prior to extraction and watering with a solution containing 370 mM Na-phosphate and 296 mM KN03. Optimal chloroplast yields (25 ug chlorophyll/g fresh mass) were obtained by chopping leaf material, using a vertical homogenizer, in a buffer containing 350 mM sorbitol, 50 mM tris-HCL and 5 mM EDTA, 0.1 % (w/v) bovine serum albumin, 0.15 % (w/v) 2-mercaptoethanol, 2 mM L-ascorbic acid and 1 mM MgCI2 followed by washing of leaf pieces in a buffer containing only sorbitol, tris-HCL and EDTA. When these chloroplasts were used in an "in-organelle" DNA digestion procedure, polymorphisms were observed between the cpDNA profiles resolved for E. grandis S/N M6 and that of an outgroup species (spinach). However, the developed chloroplast extraction technique could not be used to obtain chloroplasts from various other eucalypt species, probably as a result of variability in the material at an ultrastructural or biochemical level. For the analysis of genomic DNA RFLPs, a DNA extraction procedure was optimized for use with various eucalypt species and cultivars. This included the development of a purifcation technique during which DNA was ammonium acetate-ethanol precipitated and subjected to mini-dialysis. Following Dra I restriction of DNA, the extract was electrophoresed and Southern blotted onto both nylon and nitrocellulose membranes. These were probed with a Hind-III restricted sample of the multilocus plasmid probe pV47-2. This probe was labelled using 32p as well as a non-radioactive labelling substance digoxygenin (DIG). Hybridization conditions, including the composition of the hybridization buffer, were optimized for use with these labels, and DNA RFLPs (fingerprints) were resolved for the eucalypt species E. grandis and E. macarthurii and cultivars of E. grandis (S/N M6, TAG 5 and TAG 14). An average of 8.5 bands were detected with 32p and 5.0 fragments with DIG. All the species and cultivars fingerprinted with the 32P-label could be distinguished from one another. However, as a result of the reduced sensitivity of the DIG system, two of the E. grandis cultivars, S/N M6 and TAG 5, could not be differentiated. It is concluded that the latter system would be most suitable for incorporation into a routine eucalypt screening programme, although it is suggested that the colourimetric detection assay, used in this study to resolve DNA bands, be replaced by a more sensitive one. / Thesis (M.Sc)-University of Natal, Durban, 1993.

For the South African forestry industry, the patula pine (Pinus patula) is the most commercially important softwood species. A pine clonal programme has yet to be fully implemented in this country and at present much effort is being made to establish clonal plantings of selected trees. In order to accomplish this, it is essential that satisfactory commercially viable propagation technologies be developed for this species. This study examined the possibilities and constraints of three different in vitro systems for mass propagation of rare and important P. patula material. Seed germination and sterilisation techniques were developed for adventitious bud and somatic embryogenesis experimentation. Adventitious buds were initiated from excised 'mature P. patula embryos cultured on LM medium containing 5 mg 1-1 BA. Although, between 50 and 60% of the embryo explants produced adventitious buds, only 3-5 buds per explant actually developed further to form distinct shoots. The adventitious shoots elongated slowly (±8 mm in 2 months) on LM medium, containing 10 g 1-1 activated charcoal. Axillary buds were induced on 10 week-old juvenile shoots, after the development of an effective surface sterilisation procedure, using 0.02% HgCL2. The effect of removing the apex and trimming the needles on bud induction was significant. Dwarf shoots elongated at a rate of 25 mm in 5 weeks. Rooting studies conducted on juvenile P. patula shoots indicated that the most effective treatment was wounding the shoot base and placing the shoot in composted bark growing medium, under a greenhouse mist regime. Rooting percentages were low (50%). Included in this study is the first successful production of somatic pro-embryos from mature Pinus patula embryos. Calli were produced on LM induction medium containing 2 mg 1-1 2,4-D. Cultures were first placed in the dark for 4 weeks and then transferred to a 16 h photoperiod for a further 2 weeks, after which Stage 1 embryogenic cells were observed. When calli were placed on LM maturation medium, containing 12 mg 1-1 ABA, for a further 6-8 weeks, pro-embryo structures (maximum of 7 pro-embryos per callus) were detected embedded In the callus mass. Hence, investigations into the development of protocols for the micropropagation of Pinus patula, were undertaken. Two major constraints for applying in vitro techniques to the commercial production of pine were identified: the poor yield of shoots and pro-embryos and the length of time taken for plantlets to be produced. This study, however, provides some fundamental knowledge and background work required by tree breeders who wish to implement biotechnological techniques in the selection and improvement of P. patula genotypes. / Thesis (M.Sc.)-University of Natal, Durban, 1993.

In Africa, the grain sorghum (Sorghum bicolor (L.) Moench), is malted to provide the most important ingredient in brewing, malt, which is used primarily for the production of traditional (opaque) sorghum beer. Malting is the germination of cereal grain in moist air under controlled conditions, the primary objective being to promote the development of hydrolytic enzymes which are not present in the ungerminated grain. The malting process can be physically split into three distinct unit operations (viz. steeping, germination and drying). To date, little attention has been given to optimising the conditions of steeping for sorghum. The effects of different steeping variables (time, temperature and aeration) on the quality (in terms of diastatic power (amylase activity), free amino nitrogen and hot water extract) of sorghum malt for brewing were investigated. Malt quality was found to increase with steeping time, over the range 16-40 hours and the optimum steeping temperature was found to be in the range 25 to 30°C. Aeration during steeping appeared to be necessary to maximise the malt quality, particularly when steeping was conducted for long periods at high temperatures. Of particular significance was the observation that final sorghum malt quality was highly significantly correlated (p<0.01) with grain moisture content at steep-out (the end of the imbibition period). When steeping conditions based on these findings were used, a germination temperature of 25-30°C was found to be optimal for sorghum malt quality. As with steep-out moisture, green malt (grain after the specified germination time) moisture content was correlated Significantly (p<0.01) with final sorghum malt quality. The finding that sorghum malt quality is related to steepout moisture content was given further substance when it was shown that the stimulatory effect on sorghum malt quality of steeping sorghum in a dilute solution of alkali, actually increases the amount of water taken up during steeping probably because the alkali disrupted the pericarp cell wall structure of the grain. Barley malting practices have taken advantage of the knowledge that the exogenous application of gibberellic acid can enhance the synthesis of the critically important malt hydrolytic enzyme, a-amylase. To date, literature on the effect of exogenous application of gibberellic acid on sorghum malt quality has been inconclusive; with reports both of no effects, and of positive effects, on amylase activity. To elucidate the possible control mechanisms involved in sorghum germination, a combined HPLC-radioimmunoassay technique was used to determine the levels of selected plant growth regulators from the groups auxin, cytokinins, gibberellins and abscisic acid in sorghum at various stages of germination. Levels of gibberellic acid were low throughout germination. During germination the levels of the other plant growth regulators declined, but a peak in cytokinins followed the first visible signs of root protrusion. The high level of the germination inhibitor and gibberellic acid antagonist, abscisic acid, in the germ (embryo inclusive of scutellum) portion of the mature non-germinated grains was noteworthy. Based on these findings, it was determined that sorghum malt quality could in fact be improved significantly by the application of exogenous gibberellic acid. However, this was effective only if it was administered during the end of steeping or at the beginning of the germination step. By optimising the conditions of steeping and germination and by steeping in dilute NaOH or in gibberellic acid not only should it be possible to enhance the quality of sorghum malt, it should be possible to reduce the time required to obtain the specific quality, thereby offering a saving to the sorghum maltster in terms of operation costs and enhancing the total throughput possible from the malting plant. / Thesis (Ph.D.)-University of Natal, Durban, 1997.

The zygotic germplasm of plant species producing desiccation-sensitive seeds can be conserved in the long-term only by cryopreservation. Usually the embryonic axis is excised from the cotyledons and is used as the explant for cryopreservation as it is small and provides a large surface area:volume ratio. However the shoot of the axis of most species studied does not develop after excision, with the result that survival after cryopreservation is often recorded as callus production or simply explant enlargement and/or greening. Thus, besides explant size, factors such as in vitro regeneration techniques, physical injury induced upon excision and developmental status of the seed could compromise the success of cryopreservation. This study investigated the effect of the factors mentioned above, with particular attention to the developmental status of the seeds on explant in vitro development (section 3.1), response to dehydration (section 3.2) and cryopreservation of the desiccation-sensitive embryonic axes (section 3.3) of two species: Trichilia dregeana, T. emetica and embryos of a third, Strychnos gerrardii. For all three species, investigations were conducted on the embryonic axes/embryos excised from mature seeds immediately after fruit harvesting and from mature seeds stored under hydrated conditions for different periods (in order to achieve different degrees of development). In addition, preliminary studies were carried out on axes of T. dregeana to assess whether generation of reactive oxygen species (ROS) occurs in response to wounding upon axis excision (section 3.4). Excised embryonic axes of T. dregeana and T. emetica did not develop shoots in vitro unless the explants included attached cotyledonary segments. Following the development associated with short-term storage, however, the excised axes could develop shoots after complete cotyledon excision. The embryos from the (endospermous) seeds of S. gerrardii which included the paper-thin cotyledons, developed normally in vitro, with percentage germination increasing with seed storage time. For all three species, in vitro axis germination was promoted when activated charcoal was included in the germination medium, regardless of the developmental stage of the seeds. When dehydrated to approximately 0.3 g H2O g-1 dry mass (g g-1), embryonic axes from all three species failed to develop shoots even though a minimum of 50% produced roots in all cases. Hence, shoot production was shown to be more sensitive to desiccation than was root production. Furthermore, the sensitivity of the shoot apical meristem to desiccation was not ameliorated with seed storage for T. dregeana and T. emetica, but did decrease for S. gerrardii when seeds were stored for 6 – 8 weeks. The application of certain cryoprotectants did facilitate production of shoots after dehydration by a few axes of both Trichilia spp. For T. dregeana explants, combination of glycerol and sucrose allowed for 10% of the axes to retain the ability for shoot production after dehydration while for T. emetica explants, the combination of DMSO and glycerol (10 - 20% shoot production after dehydration) was best. The efficacy of the cryoprotectants was not influenced by storage period. The provision of cryoprotectants still needs to be tested for S. gerrardii. Survival of subsequent cryopreservation of T. dregeana and S. gerrardii explants was best achieved with rapid cooling in nitrogen slush, with the cooling procedure for T. emetica explants still to be optimized. The highest post-cryopreservation survival of T. dregeana axes was achieved when seeds had been stored for three months, while the seed storage period did not affect post-thaw survival of the axes of T. emetica or S. gerrardii. A small proportion of S. gerrardii explants only, produced shoots after cryopreservation, whereas the surviving embryonic axes of T. dregeana and T. emetica regenerated only as non-embryogenic callus. Although callus production is less desirable than successful seedling establishment, it has the potential for micropropagation if embryogenicity can be induced. Ultrastructural examination of the shoot apical meristem of T. dregeana after a 3-d recovery period, following excision, revealed considerable cellular derangement, although damage of individual organelles could not be resolved microscopically. Preliminary studies on T. dregeana involving a colorimetric assay using epinephrine, confirmed the generation of ROS in response to wounding associated with axis excision. Reactive oxygen species generated appeared to persist over prolonged periods rather than occurring only as a single oxidative burst. Hence, ROS production at the wound site could be the primary factor contributing to lack of shoot development. Axes immersed in the anti-oxidant, ascorbic acid (AsA) immediately after excision, showed lower ROS production and 10% shoot development when cultured in vitro, indicating that the oxidative burst coincident with, and after excision might be counteracted if immediate ROS production can be adequately quenched. Future investigations should aim to identify the specific ROS associated with wounding and optimize an anti-oxidant treatment(s) that will facilitate shoot development. Thus, the successful cryopreservation of the germplasm of the species tested, and others producing recalcitrant seeds, depends on a spectrum of species-specific factors, some still to be elucidated. / Thesis (M.Sc.)-University of KwaZulu-Natal, Westville, 2007.

Pigeon Valley Park, on Durban's Berea Ridge, is an approximately 10-ha remnant of coastal forest, which is totally surrounded by suburban housing and roads. As such it is ideal as a study area for investigating the applicability of the MacArthur-Wilson Theory of Island Biogeography (1963,1967) and Diamond's (1975) geometric reserve-design principles to fragmented Coastal Forests in Durban. This study began in January 1989 and the results are reported as at October 1992. Field notes from as far back as 1981 were used to augment the findings of the current work. Research focused on the forest-dwelling, Natal Robin Cossypha natalensis, and territory mapping showed that the reserve supports up to 53 individuals during the breeding season. An annotated checklist and its comparison to historical and regional checklists revealed where localised extinctions may have occurred, and thus identifies a set of coastal forest species which are susceptible to habitat fragmentation. Work on two potential dispersal corridors for bird movement into and out of the valley showed that the reserve is not yet fully isolated to most species which are currently present. On the other hand, there are some forest species which have isolated populations at Pigeon Valley Park, as well as others which do not seem able to establish and maintain viable populations. A set of 'indicator', forest bird species which are susceptible to habitat fragmentation, is defined. Practical management suggestions with the aim of increasing the long-term viability of the area as an avifaunal preserve, are presented. / Thesis (M.Sc.)-University of Natal, Durban,1992.

Although Eucalyptus species are indigenous to Australia, they have been successfully cultivated in South Africa, where they are mainly used in the mining and paper industries. With the explosion in these industries it has been necessary to increase Eucalyptus plantations, often into areas which experience frost and chilling temperatures. To combat this, high yielding Eucalyptus species able to cope with these conditions would be desirable. The ability to rank species according to their chilling tolerance will enhance decisions as to the suitability of species for use in the field. To this aim, two biochemical and two biophysical parameters were chosen to investigate and characterise Eucalyptus nitens, E. smithii, E. macarthurii, E. grandis and E. grandis x nitens (GNI026). Ranking of these species in terms of chilling tolerance did not appear possible using the data from the biochemical parameters (proline concentration and glutathione reductase activity), but the biophysical parameters (fluorescence characteristics and onset of temperature of melt) gave results comparable to those obtained in field trials using these species. Fluorescence characteristics were particularly useful in assessing the chilling tolerance of the species in question. Fluorescence is the recommended technique for further studies as it is relatively inexpensive, rapid, does not require destructive sampling and can be used in both the laboratory and field. / Thesis (M.Sc.)-University of Natal, 1993.

This study investigated aspects of the biology of Sarpa salpa, such as reproduction, age and growth, and mortality, which are necessary for an assessment of the status of this species off the KwaZulu-Natal (KZN) Coast. The importance of S. salpa to the shore-based fishery in KZN was evaluated using Natal Parks Board shore patrol data. These data were validated by analysing preliminary results of an independent shore-angling survey along the KZN Coast. Despite differences in the catch composition and catch rates between the two analyses, both data sources highlighted the importance of S. salpa to the shore-based fishery in KZN. Shore-based catches were markedly seasonal coinciding with the breeding season of the species. The species in KZN is targeted primarily to provide a supplementary source of animal protein. An investigation of the reproductive biology of S. salpa indicated a protracted spawning period for the species. Size at 50 percent maturity for combined sexes was attained at 145 mm fork length. The sex ratio in shore-based catches was 1:1.6 in favour of males. A frequency distribution by size indicated that males dominated the smaller size classes while females dominated the larger size classes. Detailed histological examination of gonadal development showed that S. salpa has the potential for protandrous sex change. An age and growth study based on the examination of whole otoliths indicated that S. salpa was relatively fast-growing and a maximum age of six years was recorded for the species. One opaque band was laid down per year. This was validated by marginal increment analysis and by an oxytetracycline labelling experiment using captive fish. Growth in S. salpa was described by a Von Bertalanffy growth function: Lage (mm FL) = 224mm(1 -e-o.55 year-1(age+o.51years)) The natural mortality rate (M = 0.6 year-1) was derived using Pauly's equation and the current fishing mortality (F) rate was estimated at 0.8 year-1. The current status of S. salpa in the shore-based fishery was assessed by determining the effects of F and age-at-capture on the yield- and spawner biomass-per-recruit. Current levels of fishing pressure on S. salpa appeared to be appropriate for utilisation of the stock off the KZN South Coast. In terms of management, S. salpa appears to be in no need of any restrictive measures at present. / Thesis (M.Sc.)-University of Natal, Durban, 1995.