Wet-dry cycling extends seed persistence by re-instating antioxidant capacity

Seeds in the field experience wet-dry cycling that is akin to the well-studied commercial process of seed priming in which seeds are hydrated and then re-dried to standardise their germination characteristics. To investigate whether the persistence (defined as in situ longevity) and antioxidant capacity of seeds are influenced by wet-dry cycling, seeds of the global agronomic weed Avena sterilis ssp. ludoviciana were subjected to (1) controlled ageing at 60% relative humidity and 53.5°C for 31 days, (2) controlled ageing then priming, or (3) ageing in the field in three soils for 21 months. Changes in seed viability (total germination), mean germination time, seedling vigour (mean seedling length), and the concentrations of the glutathione (GSH) / glutathione disulphide (GSSG) redox couple were recorded over time. As controlled-aged seeds lost viability, GSH levels declined and the relative proportion of GSSG contributing to total glutathione increased, indicative of a failing antioxidant capacity. Subjecting seeds that were aged under controlled conditions to a wet-dry cycle (to −1 MPa) prevented viability loss and increased GSH levels. Field-aged seeds that underwent numerous wet-dry cycles due to natural rainfall maintained high viability and high GSH levels. Thus wet-dry cycles in the field may enhance seed longevity and persistence coincident with re-synthesis of protective compounds such as GSH.

Characterization of the mitochondrial active-site serine protein LACTB: filaments in the mitochondrial intermembrane space

Mitochondria have evolved from endosymbiotic alpha-proteobacteria. During the endosymbiotic process early eukaryotes dumped the major component of the bacterial cell wall, the peptidoglycan layer. Peptidoglycan is synthesized and maintained by active-site serine enzymes belonging to the penicillin-binding protein and the β-lactamase superfamily. Mammals harbor a protein named LACTB that shares sequence similarity with bacterial penicillin-binding proteins and β-lactamases. Since eukaryotes lack the synthesis machinery for peptidoglycan, the physiological role of LACTB is intriguing. Recently, LACTB has been validated in vivo to be causative for obesity, suggesting that LACTB is implicated in metabolic processes. The aim of this study was to investigate the phylogeny, structure, biochemistry and cell biology of LACTB in order to elucidate its physiological function. Phylogenetic analysis revealed that LACTB has evolved from penicillin binding-proteins present in the bacterial periplasmic space. A structural model of LACTB indicates that LACTB shares characteristic features common to all penicillin-binding proteins and β-lactamases. Recombinat LACTB protein expressed in E. coli was recovered in significant quantities. Biochemical and cell biology studies showed that LACTB is a soluble protein localized in the mitochondrial intermembrane space. Further analysis showed that LACTB preprotein underwent proteolytic processing disclosing an N-terminal tetrapeptide motif also found in a set of cell death-inducing proteins. Electron microscopy structural studies revealed that LACTB can polymerize to form stable filaments with lengths ranging from twenty to several hundred nanometers. These data suggest that LACTB filaments define a distinct microdomain in the intermembrane space. A possible role of LACTB filaments is proposed in the intramitochondrial membrane organization and microcompartmentation. The implications of these findings offer novel insight into the evolution of mitochondria. Further studies of the LACTB function might provide a tool to treat mitochondria-related metabolic diseases.

Estrogen Induction Of Biotin-Binding Protein In Immature Chicks - Kinetics, Hormonal Specificity And Modulation

Employing a specific radioimmunoassay for quantification, the kinetics of estrogen-induced elevation in the plasma concentration of biotin-binding protein (BBP) in immature male chicks was investigated. A single injection of the steroid hormone enhanced the plasma BBP content several-fold at 6 h, reaching peak levels around 48 h and declining thereafter. A 2-fold amplification of the response was evident during secondary stimulation with the hormone. The magnitude of the response was hormonal dose-dependent while the initial lag phase and the time of peak protein accumulation were unaltered within the hormonal doses tested. The circulatory half-life of the specific protein in normal and estrogenized birds was 10 h. Hyperthyroidism markedly decreased the hormonal response while the opposite effect was seen during hypothyroidism. The antiestrogens E- and Z-clomiphene citrate effectively blocked the protein induction whereas progesterone, either alone or in combination with estrogen, was ineffective in modulating the induction. Cycloheximide administration drastically inhibited the inductive response. The above observations clearly suggest that the genes corresponding to the two isofunctional proteins of chicken egg, viz. BBP and avidin, are differentially regulated.

Estrogen modulation of riboflavin carrier protein in the bonnet monkey (Macaca radiata)

Circulatory concentrations of riboflavin carrier protein (RCP) were quantitated in bonnet macaques by employing a heterologous radioimmunoassay involving 125I-labelled chicken RCP and its antiserum. The levels of monkey RCP in the serum seem to be governed by the estrogenic status of the animals. An increase in concentration of serum estradiol in the adult females during the menstrual cycle and early pregnancy could be correlated with enhanced serum RCP levels. Estadiol-17β administered to both immature female and male monkeys, specifically brought about elevated levels of RCP with a slower time course of response in males than in females. These results could be a reflection of a more rapid decline of both circulatory estrogen and RCP concentrations in male serum. Repeated administration of estradiol-17β to male animals led to prolonged elevated levels of RCP following estrogen administration. Thus, it would appear that the evolutionary conservation of RCPs from the aves to the primates encompasses not only their physicochemical similarities but also extends to the estrogenic modulation of their elaboration.

Seed production of Stylosanthes guyanensis. 1. Growth and development.

The pattern of growth and development of seed crops of stylo (Stylosanthes guyanensis) was derived from measurements made on experimental and commercial crops in north Queensland. The three cultivars Cook, Endeavour, and Schofield differed appreciably only in the timetable of their development. Each had distinct successive phases of vegetative and reproductive development culminating in total annual seed production of 700-800 kg ha-1 from a healthy closed canopy, the main recorded cause of reduced production being the disease Botrytis sp. In a healthy crop of Cook, the peak quantity of standing seed represented almost 90 per cent of the total accountable seed, and the rise to and decline from this peak proceeded at rates of the order of 3-4 per cent per day. It is deduced that, although there appears to be little potential for either increase in overall production or improvement in synchronization or retention characteristics beyond that currently attained by a closed canopy of healthy plants, there is scope for an increase in the efficiency of recovery of standing seed. Maximum recovery will be achieved through attention to choice of time of harvest, presentation of a minimum amount of extraneous vegetation to the harvester, and improvement in harvester separation.

Seed production of Stylosanthes guyanensis. 2. The consequences of defoliation.

The consequences of defoliation on seed production of stylo (Stylosanthes guyanensis) were examined in field experiments at Walkamin in north Queensland. The practical aim of defoliation is to present a level uncompacted crop canopy to the harvester without a reduction in the quantity of seed carried at harvest ripeness. It was concluded that the latest date at which defoliation is reasonably certain to achieve its objectives is about four weeks before first flower initiation. In north Queensland, this means late February for cvv. Cook and Endeavour and early April for cv. Schofield. The results suggest that development of the population of individual shoots must be synchronized to produce the highest peaks of standing seed; that this is best achieved by ensuring that a closed crop canopy with a ceiling shoot population exists at the time of first flower initiation; and that poorly synchronized shoot development is a consequence of defoliating too late and a cause of reduced seed production.

Seed production of Stylosanthes guyanensis. 3. Effects of pre-harvest desiccation.

Strips within commercial crops of Stylosanthes guyanensis in the Mareeba district of north Queensland were sprayed with diquat 4, 6 and 10 days before harvest and compared with unsprayed strips. Pre-harvest desiccation made combine harvesting easier, but did not increase harvest yield. Where seed formation and maturation was still possible, desiccation prevented this without substantially increasing the loss of seed to the ground; increased harvest efficiency was thus offset by a diminished quantity of standing seed. However, where there was little or no further potential for seed development, diquat had virtually no effect on the quantity of standing seed or harvest efficiency. It was concluded that the results warranted neither recommendation nor further evaluation of preharvest desiccation of S. guyanensis seed crops.

Effects of stage of defoliation on seed production and growth of Stylosanthes humilis.

Stylosanthes humilis swards grown at Brisbane in irrigated boxes were defoliated (about 60 per cent removal of tops) at floral initiation, first flower appearance, or advanced flowering stages ; seed yield was 45, 16, and 14 per cent respectively of seed yield in undefoliated swards. Decreased yields were primarily due to poor seed set of florets, were also associated with reduced inflorescence density and floret number per inflorescence, and occurred despite increases (in some defoliation treatments) in seed size, leaf growth rate, and differentiation of leaves and branches. Total seasonal plant growth was independent of defoliation treatment.

Phenotypic and genotypic variation within populations of kikuyu (Pennisetum clandestinum) in Australia.

Experiments at 2 sites in subtropical eastern Australia investigated the variation in agronomic attributes, quality and genetic structure existing within: naturally-occurring populations of kikuyu ( Pennisetum clandestinum) from within Australia; selections produced from the treatment of Whittet seed with mutagenic chemicals; and available cultivars. Runners were collected from coastal areas extending from Western Australia to the Atherton Tableland in north Queensland. One experiment evaluated 10 mutagenic selections and 4 cultivars in a lattice design and the other evaluated 12 ecotypes and 3 cultivars in a randomised block design. The experimental unit was single plants, which were sown on a 1.5 m grid into a weed-free seed-bed (Mutdapilly) or a killed kikuyu stand (Wollongbar), both of which were kept clear of weeds and other kikuyu plants for the duration of the experiments. Foliage height, forage production and runner yield were assessed. Leaf material was analysed for concentrations of crude protein (CP), acid detergent fibre (ADF) and neutral detergent fibre (NDF) and for in vitro dry matter digestibility (IVDDM) in autumn, winter and spring. DNA was extracted from each plant in the ecotype comparison and subjected to a modified DAF (DNA amplification fingerprinting) analysis to determine the level of genetic relatedness. In the first experiment, none of the mutagenic lines derived from Whittet yielded significantly more or was more digestible than commercial Whittet material, although some selections were superior to the other commercial kikuyu cultivars, Noonan and Crofts, and 'common' kikuyu. However, there were significant differences in plant height and runner expansion. In the second experiment, significant differences in plant height, foliage yield, runner development, and leaf CP, ADF, NDF and IVDDM concentrations were demonstrated between the ecotypes, mutagenic selections and cultivars. There was a 4- to 6-fold difference in plant yield and a 6- to 10-fold difference in runner production between the ecotypes at the 2 sites. Quality of the leaf ranged from 200 to 270 g/kg (CP), from 700 to 770 g/kg (IVDDM), from 170 to 250 g/kg (ADF) and from 470 to 550 g/kg (NDF). Improvements in quality and agronomic attributes were not mutually exclusive. Genetic fingerprint analysis of the kikuyu lines indicated that they formed 2 broad groupings. Most of the regional ecotypes were grouped with 'common' kikuyu as represented by the material collected from Wollongbar, and the Beechmont, Atherton Tableland and Gympie ecotypes were grouped with the registered cultivars Whittet, Noonan and Crofts. Two lines produced by mutagenesis from Whittet remained closely linked to Whittet. These results suggest that there was variation between populations of kikuyu in yield, quality and genetic diversity but that mutagenesis by treating seed with sodium azide and diethylene sulphide did not achieve a significant change in the digestibility of leaf over cv. Whittet.

Baker's yeast expressing the Japanese encephalitis virus envelope protein on its cell surface: induction of an antigen-specific but non-neutralizing antibody response

Live recombinant Saccharomyces cerevisiae yeast expressing the envelope antigen of Japanese encephalitis virus (JEV) on the outer mannoprotein layer of the cell wall were examined for their ability to induce antigen-specific antibody responses in mice. When used as a modelantigen, parenteral immunization of mice with surface-expressing GFP yeast induced a strong anti-GFP antibody response in the absence of adjuvants. This antigen delivery approach was then used for a more stringent system, such as the envelope protein of JEV, which is a neurotropic virus requiring neutralizing antibodies for protection.Although 70% of cells were detected to express the total envelope protein on the surface by antibodies raised to the bacterially expressed protein, polyclonal anti-JEV antibodies failed to react with them. In marked contrast, yeast expressing the envelope fragments 238-398, 373-399 and 373-500 in front of a Gly-Ser linker were detected by anti-JEV antibodies as well as a monoclonal antibody but not by antibodies raised to the bacterially expressed protein. Immunization of mice with these surface-expressing recombinants resulted in a strong antibody response. However, the antibodies failed to neutralize the virus, although the fragments were selected based on neutralizing determinants.

Utilisation of local ingredients in commercial pig feeds

The long-term competitiveness of the both the Vietnamese feed and pig production industries are constrained and under pressure whilst the industry is dependent on the use of imported feed ingredients in diets for animal production. These cost pressures are a result of import taxes, transport costs, currency fluctuations and feed supply limitations. By undertaking studies on available resources which are currently under-utilised and with potential as local feeds, we can prove their suitability for use as feedstuffs in pig diets and as replacements for imported feed ingredients. In undertaking this process we can lower feeding costs for pig production in Vietnam by the use of local feeds which are cheaper, generate new industries in Vietnam harvesting or processing these feeds and increase the incomes of Vietnamese workers who are involved in producing these by-products. Our project has shown that rubber seed, when processed correctly to lower the hydrogen cyanide content, is a safe and suitable protein meal feedstuff for use in pig diets with the potential to replace significant quantities of imported soybean and fishmeal in Vietnamese pig diets as long as diets are balanced for any amino acid shortfalls. Our peanut studies have shown that use of binders can help alleviate pig production problems with aflatoxin content in peanut meals. Further work is needed to characterise the fate of the bound aflatoxin to see if there is any meat residue risk. Cassava residue is a resultant by-product from starch extraction in both large and small cassava processing factories. Sub-samples from these two mill types were collected and evaluated for residue HCN. Analyses has shown that the processing and sun drying results in a product with relatively consistent low HCN content. Chemical analyses also reveal that significant residual starch also remains in this by-product. Digestibility studies and pig performance feeding studies have shown that cassava residue can be included in diets at 30% with no adverse effect, although the higher fibre content of this product means that strategically, cassava residue is more suitably used in finisher and sow diets. Research has examined the digestible energy content of a number of sunflower meal types available in Australia and identified major differences in their energy value based on processing, additionally, amino acid analysis has shown a significantly lower lysine content than previous reported. We also examined the digestible energy content of a number of Australian stylo forage legume harvest batches and identified the differences in their energy value based on age/harvest time of the forage legume. Analysis results of various stylo cuts showed that the early cut stylo has a higher starch content and lower fibre fraction content than observed in late and recut stylo which were allowed to grow longer. As a result the faecal digestible energy content was higher for the early cut stylo than for the subsequent cut stylo material which had been allowed to become woody. The results have shown that feeding of stylo meal does provide some nutritive value to the pig with increased energy and nitrogen supply, with a portion of the nitrogen presented which the pig is able to retain. Based on nutrient and fibre content stylo could have a useful role in sow feeding and satiety under non-stall housing situations. With increasing Vietnamese investment in rubber production seen with larger areas under plantations the amounts of rubber seed available for animal feeding will grow significantly over the next 15 years and the importance of the by-product ie rubber seed meal as a protein source in diets for Vietnamese pigs.

Cost effective formulation of vegetable-protein based diets

The project will provide information on the use of phytase in sorghum based diets so that producers and nutrionists will have confidence in vegetable protein based diets.