Production of the baculovirus-expressed dengue virus glycoprotein NS1 can be improved dramatically with optimised regimes for fed-batch cultures and the addition of the insect moulting hormone, 20-Hydroxyecdysone

A perennial problem in recombinant protein expression is low yield of the product of interest. A strategy which has been shown to increase the production of baculovirus-expressed proteins is to utilise fed-batch cultures. One disadvantage of this approach is the time-consuming task of optimising the feeding strategy. Previously, a statistical optimisation routine was applied to develop a feeding strategy that increased the yield of beta-Galactosidase (beta-Gal) by 2.4-fold (Biotechnol. Bioeng, 59 (1998) 178). This involves the single addition of nutrient concentrates (amino acids, lipids. glucose and yeastolate ultrafiltrate) into Sf9 cell cultures grown in SF900II medium. In this study, it is demonstrated that this optimised fed-batch strategy developed for a high-yielding intracellular product beta-Gal could be applied successfully to a relatively low-yielding glycosylated and secreted product such as the dengue virus glycoprotein NS1. Optimised batch infections yielded 4 mug/ml of NS1 at a peak cell density of 4.2 x 10(6) cells/ml. In contrast. optimised fed-batch infections exhibited a 3-fold improvement in yield, with 12 mug ml of NS1 produced at a peak cell density of 11.3 x 10(6) cells/ml. No further improvements in yield were recorded when the feed volumes were doubled and the peak cell density was increased to 23 x 10(6) cells/ml, unless the cultures were stimulated by the addition of 4 mug/ml of 20-Hydroxyecdysone (an insect moulting hormone). In this case, the NS1 yield was increased to 20 mug/ml. which was nearly 5-fold higher than optimised batch cultures. (C) 2002 Elsevier Science B.V. All rights reserved.

Tillering in grain sorghum over a wide range of population densities: Identification of a common hierarchy for tiller emergence, leaf area development and fertility

Most studies of tiller development have not related the physiological and morphological features of each calm to its subsequent fertility. This introduced problems when trying to account for the effects of tillering on yield in crop models. The objective of this study was to detect the most likely early determinants of tiller fertility in sorghum by identifying hierarchies for emergence, fertility and grain number of tillers over a wide range of assimilate availabilities. Emergence, phenology, leaf area development and dry weight partitioning were quantified weekly for individual tillers and main culms of tillering and uniculm plants grown at one of four densities, from two to 16 plants m(-2). For a given plant in any given density, the same tiller hierarchy applied for emergence of tillers, fertility of the emerged tillers and their subsequent grain number. These results were observed over a range of tiller fertility rates (from 7 to 91%), fertile tiller number per plant at maturity (from 0.2 to 4.7), and tiller contribution to grain yield (from 5 to 78%). Tiller emergence was most probably related to assimilate supply and light quality. Development, fertility and contribution to yield of a specific tiller were highly dependent on growing conditions at the time of tiller emergence, particularly via early leaf area development of the tiller, which affected its subsequent leaf area accumulation. Assimilate availability in the main culm at the time of tiller emergence was the most likely early determinant of subsequent tiller fertility in this study. (C) 2002 Annals of Botany Company.

Tillering in grain sorghum over a wide range of population densities: Modelling dynamics of tiller fertility

The prediction of tillering is poor or absent in existing sorghum crop models even though fertile tillers contribute significantly to grain yield. The objective of this study was to identify general quantitative relationships underpinning tiller dynamics of sorghum for a broad range of assimilate availabilities. Emergence, phenology, leaf area development and fertility of individual main calms and tillers were quantified weekly in plants grown at one of four plant densities ranging from two to 16 plants m(-2). On any given day, a tiller was considered potentially fertile (a posteriori) if its number of leaves continued to increase thereafter. The dynamics of potentially fertile tiller number per plant varied greatly with plant density, but could generally be described by three determinants, stable across plant densities: tiller emergence rate aligned with leaf ligule appearance rate; cessation of tiller emergence occurred at a stable leaf area index; and rate of decrease in potentially fertile tillers was linearly related to the ratio of realized to potential leaf area growth. Realized leaf area growth is the measured increase in leaf area, whereas potential leaf area growth is the estimated increase in leaf area if all potentially fertile tillers were to continue to develop. Procedures to predict this ratio, by estimating realized leaf area per plant from intercepted radiation and potential leaf area per plant from the number and type of developing axes, are presented. While it is suitable for modelling tiller dynamics in grain sorghum, this general framework needs to be validated by testing it in different environments and for other cultivars. (C) 2002 Annals of Botany Company.

Development of a generic crop model template in the cropping system model APSIM

The Agricultural Production Systems slMulator, APSIM, is a cropping system modelling environment that simulates the dynamics of soil-plant-management interactions within a single crop or a cropping system. Adaptation of previously developed crop models has resulted in multiple crop modules in APSIM, which have low scientific transparency and code efficiency. A generic crop model template (GCROP) has been developed to capture unifying physiological principles across crops (plant types) and to provide modular and efficient code for crop modelling. It comprises a standard crop interface to the APSIM engine, a generic crop model structure, a crop process library, and well-structured crop parameter files. The process library contains the major science underpinning the crop models and incorporates generic routines based on physiological principles for growth and development processes that are common across crops. It allows APSIM to simulate different crops using the same set of computer code. The generic model structure and parameter files provide an easy way to test, modify, exchange and compare modelling approaches at process level without necessitating changes in the code. The standard interface generalises the model inputs and outputs, and utilises a standard protocol to communicate with other APSIM modules through the APSIM engine. The crop template serves as a convenient means to test new insights and compare approaches to component modelling, while maintaining a focus on predictive capability. This paper describes and discusses the scientific basis, the design, implementation and future development of the crop template in APSIM. On this basis, we argue that the combination of good software engineering with sound crop science can enhance the rate of advance in crop modelling. Crown Copyright (C) 2002 Published by Elsevier Science B.V. All rights reserved.

Future contributions of crop modelling - from heuristics and supporting decision making to understanding genetic regulation and aiding crop improvement

Crop modelling has evolved over the last 30 or so years in concert with advances in crop physiology, crop ecology and computing technology. Having reached a respectable degree of acceptance, it is appropriate to review briefly the course of developments in crop modelling and to project what might be major contributions of crop modelling in the future. Two major opportunities are envisioned for increased modelling activity in the future. One opportunity is in a continuing central, heuristic role to support scientific investigation, to facilitate decision making by crop managers, and to aid in education. Heuristic activities will also extend to the broader system-level issues of environmental and ecological aspects of crop production. The second opportunity is projected as a prime contributor in understanding and advancing the genetic regulation of plant performance and plant improvement. Physiological dissection and modelling of traits provides an avenue by which crop modelling could contribute to enhancing integration of molecular genetic technologies in crop improvement. Crown Copyright (C) 2002 Published by Elsevier Science B.V. All rights reserved.

The Three-dimensional Solution Structure of NaD1, a New Floral Defensin from Nicotiana alata and its Application to a Homology Model of the Crop Defense Protein alfAFP

NMR spectroscopy and simulated annealing calculations have been used to determine the three-dimensional structure of NaD1, a novel antifungal and insecticidal protein isolated from the flowers of Nicotiana alata. NaD1 is a basic, cysteine-rich protein of 47 residues and is the first example of a plant defensin from flowers to be characterized structurally. Its three-dimensional structure consists of an a-helix and a triple-stranded anti-parallel beta-sheet that are stabilized by four intramolecular disulfide bonds. NaD1 features all the characteristics of the cysteine-stabilized up motif that has been described for a variety of proteins of differing functions ranging from antibacterial insect defensins and ion channel-perturbing scorpion toxins to an elicitor of the sweet taste response. The protein is biologically active against insect pests, which makes it a potential candidate for use in crop protection. NaD1 shares 31% sequence identity with alfAFP, an antifungal protein from alfalfa that confers resistance to a fungal pathogen in transgenic potatoes. The structure of NaD1 was used to obtain a homology model of alfAFP, since NaD1 has the highest level of sequence identity with alfAFP of any structurally characterized antifungal defensin. The structures of NaD1 and alfAFP were used in conjunction with structure - activity data for the radish defensin Rs-AFP2 to provide an insight into structure-function relationships. In particular, a putative effector site was identified in the structure of NaD1 and in the corresponding homology model of alfAFP. (C) 2002 Elsevier Science Ltd. All rights reserved.

Towards the on-farm conservation of the assassin bug Pristhesancus plagipennis (Walker) (Hemiptera : Reduviidae) during winter using crop plants as refuges

Habitat instability associated with seasonal crop succession in broad-acre farming systems presents a problem for the conservation and utilisation of beneficial insects in annual field crops. The present paper describes two experiments used to measure the potential of seven plant species to be utilised as winter refuges to support and conserve the predatory bug Pristhesancus plagipennis (Walker). In the first experiment, replicated plots of canola (Brassica napus ), red salvia (Salvia coccinea ), niger (Guizotia abyssinica ), linseed (Linum usitatissimum ), lupins (Lupinus angustifolius ), and lucerne (Medicago falcata ) were planted in a randomized experiment during Autumn 1998. Upon crop establishment, adults and nymphs of P. plagipennis were released into treatment plots and their numbers were assessed, along with those of their potential prey, throughout the ensuing winter months. Post-release sampling suggested that canola and niger retained a proportion of adult P. plagipennis , while niger, lucerne and canola retained some nymphs. The other plant species failed to support P. plagipennis nymphs and adults postrelease. In the second experiment, niger was compared with two lines of sunflower (Helianthus annus ). Both sunflower lines harboured significantly higher (P < 0.05) densities of P. plagipennis nymphs than did niger. The more successful refuge treatments (sunflower, niger and canola) had an abundance of yellow flowers that were attractive to pollinating insects, which served as supplementary prey on which P. plagipennis were observed to feed. Sunflower and niger also supported high densities of the prey insect Creontiades dilutus (Stal) and provided protective leafy canopies which supplied shelter during the winter months. The potential and limitations for using each plant species as a winter refuge to retain P. plagipennis during winter are discussed.

Yield responses to ethephon for unshaken and mechanically shaken macadamia

Ethephon promotes fruit abscission and accelerates harvest of macadamia, Macadamia integrifolia (Proteaceae), but has limited use due to concerns that associated abscission of inner-canopy leaves may reduce subsequent yield and nut quality. Yield and quality were monitored for 2 years following ethephon application to both unshaken and mechanically shaken trees of the late-abscising cultivar, A16. Nut quality was not adversely affected in subsequent seasons, but effects on yield varied. In 3 of 6 experiments, ethephon reduced yield in the year after application. However, in 4 of the 6 experiments, 2 years of ethephon application greatly elevated yield in the third year. This was not a compensating recovery from low second-year yield, as third-year yield of trees that received only 1 ethephon treatment did not differ from yield of control trees. Ethephon-assisted harvest remains feasible for macadamia, although further work is warranted given the potential risks and considerable benefits for subsequent yield. Inner canopy defoliation, resulting from ethephon use, could represent a canopy management technique for dense-canopy fruit trees.

Genotype and environment effects on dynamics of harvest index during grain filling in sorghum

An approach based on a linear rate of increase in harvest index (141) with time after anthesis has been used as a simple means-to predict grain growth and yield in many crop simulation models. When applied to diverse situations, however, this approach has been found to introduce significant error in grain yield predictions. Accordingly, this study was undertaken to examine the stability of the HI approach for yield prediction in sorghum [Sorghum bicolor (L.) Moench]. Four field experiments were conducted under nonlimiting water. and N conditions. The experiments were sown at times that ensured a broad range in temperature and radiation conditions. Treatments consisted of two population densities and three genotypes varying in maturity. Frequent sequential harvests were used to monitor crop growth, yield, and the dynamics of 111. Experiments varied greatly in yield and final HI. There was also a tendency for lower HI with later maturity. Harvest index dynamics also varied among experiments and, to a lesser extent, among treatments within experiments. The variation was associated mostly with the linear rate of increase in HI and timing of cessation of that increase. The average rate of HI increase was 0.0198 d(-1), but this was reduced considerably (0.0147) in one experiment that matured in cool conditions. The variations found in IN dynamics could be largely explained by differences in assimilation during grain filling and remobilization of preanthesis assimilate. We concluded that this level of variation in HI dynamics limited the general applicability of the HI approach in yield prediction and suggested a potential alternative for testing.

Evaluating plant breeding strategies by simulating gene action and dryland environment effects

Functional genomics is the systematic study of genome-wide effects of gene expression on organism growth and development with the ultimate aim of understanding how networks of genes influence traits. Here, we use a dynamic biophysical cropping systems model (APSIM-Sorg) to generate a state space of genotype performance based on 15 genes controlling four adaptive traits and then search this spice using a quantitative genetics model of a plant breeding program (QU-GENE) to simulate recurrent selection. Complex epistatic and gene X environment effects were generated for yield even though gene action at the trait level had been defined as simple additive effects. Given alternative breeding strategies that restricted either the cultivar maturity type or the drought environment type, the positive (+) alleles for 15 genes associated with the four adaptive traits were accumulated at different rates over cycles of selection. While early maturing genotypes were favored in the Severe-Terminal drought environment type, late genotypes were favored in the Mild-Terminal and Midseason drought environment types. In the Severe-Terminal environment, there was an interaction of the stay-green (SG) trait with other traits: Selection for + alleles of the SG genes was delayed until + alleles for genes associated with the transpiration efficiency and osmotic adjustment traits had been fixed. Given limitations in our current understanding of trait interaction and genetic control, the results are not conclusive. However, they demonstrate how the per se complexity of gene X gene X environment interactions will challenge the application of genomics and marker-assisted selection in crop improvement for dryland adaptation.

Simultaneous FT-NIR and ESR analyses to yield propagation rate coefficients for polymerization of methyl methacrylate based monomers

A hyphenated instrumental approach has been used to obtain reliable values for the propagation rate coefficients as a function of conversion for polymerizations of methyl methacrylate (MMA) and a mixture of MMA and ethyleneglycol dimethacrylate (EGDMA) with a 1:1 concentration of double bonds, from near the onset of the Trommsdorf region into the glass region. ESR spectroscopy was used to measure the radical concentration while FT-NIR fibre-optic spectroscopy was employed to measure instantaneously the double-bond concentration within the temperature-controlled cavity of the ESR instrument during polymerization. The advantage of this approach to the measurement of the rate coefficient is that it is equally applicable to branching and linear polymerizations. For the polymerization of methyl methacrylate, the values of the rate coefficient at the lowest conversions at which reliable values could be obtained were in agreement with recently reported values obtained by the PLP-SEC method. For the lowest conversions, the values obtained were 403 1 mol(-1) s(-1) at 306 K for MMA and 5201 mol(-1) s(-1) at 310 K for a 1:1 mixture of MMA and EGDMA. (C) 2003 Society of Chemical Industry.

Crescimento, produção e rendimento dos clones da variedade ‘Conilon Vitória’ em condições de déficit hídrico e irrigado

O Estado do Espírito Santo é o maior produtor de grãos de café conilon do Brasil. Devido aos trabalhos na seleção de plantas produtivas, os avanços na produtividade desta cultura são mais pronunciados com a utilização da técnica da irrigação. A utilização da irrigação na cafeicultura possibilita a adoção de novas tecnologias de plantio, expansão da área produtiva, uniformização da produção diminuindo os grãos chochos e mal granados, além de mitigar os problemas da deficiência hídrica nos períodos críticos da cultura. Diante do exposto, objetivou-se com este estudo, avaliar o crescimento, a produção e o rendimento dos treze clones da variedade de café ‘Conilon Vitória’, em condições de déficit hídrico e irrigado. O experimento foi instalado no ano de 2010 no Instituto Federal do Espírito Santo, Campus Alegre, em esquema de parcelas subdivididas 3 x 13, sendo nas parcelas o manejo de irrigação em três níveis (irrigado, irrigado com déficit hídrico de 50% e sem irrigação) e nas subparcelas os clones da variedade ‘Conilon Vitória’ em treze níveis, em um delineamento em blocos casualizados, com três repetições. Cada parcela experimental foi constituída por cinco plantas úteis. O manejo da irrigação adotado foi via clima, onde a ETo foi estimada por meio da equação de Hargreaves e Samani (1985). Os dados foram submetidos à análise de variância e as médias comparadas pelo teste de Tukey para comparações entre o fator manejo e Scott-Knott para comparações entre o fator clone. O crescimento dos ramos das plantas foi diferenciado em relação ao manejo de irrigação aplicado. Plantas sem irrigação apresentaram maiores picos de crescimento. Plantas irrigadas apresentaram menores picos de crescimento dos ramos, porém, com maior período de crescimento. A produtividade média de plantas irrigadas foi cerca de 474,5% superior a plantas sem irrigação. A produtividade média de plantas irrigadas com déficit hídrico de 50% foi cerca de 329,6 % superior a plantas sem irrigação. O rendimento entre plantas irrigadas e irrigadas com déficit hídrico de 50% não diferiu. Já plantas sem irrigação apresentaram menor rendimento em todos os clones. A irrigação influenciou no tamanho dos grãos, plantas irrigadas apresentaram maiores percentagens de grãos retidos em peneiras 13 e superiores. Já plantas sem irrigação apresentaram maiores percentagens de grãos retidos na fundagem.

Cobertura permanente de solo com leguminosas herbáceas perenes no cultivo de bananeiras no médio Vale do Jequitinhonha

A região da Caatinga é caracterizada pelas altas temperaturas durante o ano e má distribuição das chuvas. Em virtude desses fatores de clima regional, tem-se a necessidade de adoção por práticas que elevem à eficiência e sustentabilidade agrícola local. Assim, objetivou-se avaliar a aptidão de leguminosas herbáceas perenes como cobertura permanente de solo no cultivo de bananeira. Foram conduzidos três experimentos, para avaliação das leguminosas, utilizou-se um delineamento experimental em blocos casualizados, com quatro repetições, em esquema de parcelas subdivididas no espaço e para algumas variáveis, sub-subdividida no tempo, sendo: Fator “A” os dois diferentes ambientes de plantio: municípios de Itaobim/MG e Virgem da Lapa/MG; Fator “B”, nas subparcelas, dois manejos de cobertura do solo e para algumas variáveis, três manejos, constituídos pelas leguminosas: cudzu tropical (Pueraria phaseoloides) calopogônio (Calopogonium mucunoides) e solo descoberto (solo capinado); Fator “C” épocas de coleta de dados. Para avaliação das bananeiras, foram dois experimentos em blocos casualizados, com quatro repetições, em esquema de parcelas sudivididas no espaço, sendo: nas parcelas, fator “A” constituído por três manejos de cobertura do solo, pelas leguminosas: cudzu tropical e o calopogônio, e solo descoberto (solo capinado); fator “B”, nas subparcelas, plantas de bananeiras em três idades morfofisiológicas (diferentes ciclos e tamanhos); Para algumas variáveis que foram submetidas a coletas periódicas, utilizou-se o esquema de parcelas subsubdivididas no tempo, acrescentando-se o fator “C”, datas das coletas nas sub-subparcelas, tendo como referência os dias após semeadura (DAS) das leguminosas. Foram avaliadas as seguintes variáveis: taxa de cobertura do solo; potencial de deposição de folhas e a ciclagem de nutrientes; capacidade de inibição da vegetação espontânea; conservação da temperatura e promoção da retenção de umidade do solo. Também foi avaliado o crescimento vegetativo e produtividade das bananeiras. Como resultados principais, notou-se que as leguminosas proporcionaram eficiente cobertura do solo, o calopogônio apresentou o maior acúmulo de N, P, K, e Ca, via deposição de material senescente, tal como maior inibição das plantas espontâneas nos pomares de bananeiras. Essa cobertura também promoveu uma eficiente redução da temperatura do solo, possibilitando menor variação térmica nas camadas de maior concentração radicular da bananeira, e consequentemente, obtendo maior acúmulo de umidade no solo. As bananeiras cultivadas sobre coberturas vivas de solo apresentaram aumento gradativo no crescimento e peso de cacho. Os resultados reforçam o potencial uso dessas espécies na fruticultura, principalmente em regiões de severas restrições hídricas, como forma de adubação e otimização de diversos processos biológicos em seu ambiente de cultivo.

Crescimento, produção e rendimento dos clones da variedade ‘Conilon Vitória’ em condições de déficit hídrico e irrigado

O Estado do Espírito Santo é o maior produtor de grãos de café conilon do Brasil. Devido aos trabalhos na seleção de plantas produtivas, os avanços na produtividade desta cultura são mais pronunciados com a utilização da técnica da irrigação. A utilização da irrigação na cafeicultura possibilita a adoção de novas tecnologias de plantio, expansão da área produtiva, uniformização da produção diminuindo os grãos chochos e mal granados, além de mitigar os problemas da deficiência hídrica nos períodos críticos da cultura. Diante do exposto, objetivou-se com este estudo, avaliar o crescimento, a produção e o rendimento dos treze clones da variedade de café ‘Conilon Vitória’, em condições de déficit hídrico e irrigado. O experimento foi instalado no ano de 2010 no Instituto Federal do Espírito Santo, Campus Alegre, em esquema de parcelas subdivididas 3 x 13, sendo nas parcelas o manejo de irrigação em três níveis (irrigado, irrigado com déficit hídrico de 50% e sem irrigação) e nas subparcelas os clones da variedade ‘Conilon Vitória’ em treze níveis, em um delineamento em blocos casualizados, com três repetições. Cada parcela experimental foi constituída por cinco plantas úteis. O manejo da irrigação adotado foi via clima, onde a ETo foi estimada por meio da equação de Hargreaves e Samani (1985). Os dados foram submetidos à análise de variância e as médias comparadas pelo teste de Tukey para comparações entre o fator manejo e Scott-Knott para comparações entre o fator clone. O crescimento dos ramos das plantas foi diferenciado em relação ao manejo de irrigação aplicado. Plantas sem irrigação apresentaram maiores picos de crescimento. Plantas irrigadas apresentaram menores picos de crescimento dos ramos, porém, com maior período de crescimento. A produtividade média de plantas irrigadas foi cerca de 474,5% superior a plantas sem irrigação. A produtividade média de plantas irrigadas com déficit hídrico de 50% foi cerca de 329,6 % superior a plantas sem irrigação. O rendimento entre plantas irrigadas e irrigadas com déficit hídrico de 50% não diferiu. Já plantas sem irrigação apresentaram menor rendimento em todos os clones. A irrigação influenciou no tamanho dos grãos, plantas irrigadas apresentaram maiores percentagens de grãos retidos em peneiras 13 e superiores. Já plantas sem irrigação apresentaram maiores percentagens de grãos retidos na fundagem.