Inheritance of Resistance to Rice Tungro Spherical Virus in a Near-Isogenic Line Derived from Utri Merah and in Rice Cultivar TKM6

Resistance to rice virus diseases is an important requirement in many Southeast Asian rice breeding programs. Inheritance of resistance to rice tungro spherical virus (RTSV) in TW5, a near-isogenic line derived from Indonesian rice cultivar Utri Merah, was compared to that in TKM6, an Indian rice cultivar. Both TKM6 and Utri Merah are cultivars resistant to RTSV infections. Crosses were made between TKM6 and TN1, a susceptible cultivar, and between TW5 and TN1, and F3 lines were evaluated for their resistance to RTSV using two RTSV inoculum sources and a serological assay (ELISA). In TKM6, the resistance to the mixture of RTSV-V + RTBV inoculum source was controlled by a single recessive gene, whereas in TW5, the resistance was controlled by two recessive genes. A single recessive gene, however, controlled the resistance in TW5 when another RTSV variant, RTSV-VI, was used, suggesting that the resistance in TW5 depends on the nature of the RTSV inoculum used. RT-PCR, sequence, and phylogenetic analyses confirmed that RTSV-VI inoculum differs from RTSV-V inoculum and accurate phenotyping of the resistance to RTSV requires the use of a genetic marker.

Development of a high-level gene expression system for the production of bioplastics in sugarcane

Despite various approaches, the production of biodegradable plastics such as polyhydroxybutyrate (PHB) in transgenic plants has met with limited success due largely to low expression levels. Even in the few instances where high levels of protein expression have been reported, the transgenic plants have been stunted indicating PHB is phytotoxic (Poirier 2002). This PhD describes the application of a novel virus-based gene expression technology, termed InPAct („In Plant Activation.), for the production of PHB in tobacco and sugarcane. InPAct is based on the rolling circle replication mechanism by which circular ssDNA viruses replicate and provides a system for controlled, high-level gene expression. Based on these features, InPAct was thought to represent an ideal system to enable the controlled, high-level expression of the three phb genes (phbA, phbB and phbC) required for PHB production in sugarcane at a preferred stage of plant growth. A Tobacco yellow dwarf virus (TbYDV)-based InPAct-phbA vector, as well as linear vectors constitutively expressing phbB and phbC were constructed and different combinations were used to transform tobacco leaf discs. A total of four, eight, three and three phenotypically normal tobacco lines were generated from discs transformed with InPAct-phbA, InPAct-phbA + p1300-TaBV P-phbB/phbC- 35S T, p1300-35S P-phbA-NOS T + p1300-TaBV P-phbB/phbC-35S T and InPAct-GUS, respectively. To determine whether the InPAct cassette could be activated in the presence of the TbYDV Rep, leaf samples from the eight InPActphbA + p1300-TaBV P-phbB/phbC-35S T plants were agroinfiltrated with p1300- TbYDV-Rep/RepA. Three days later, successful activation was indicated by the detection of episomes using both PCR and Southern analysis. Leaf discs from the eight InPAct-phbA + p1300-TaBV P-phbB/phbC-35S T transgenic plant lines were agroinfiltrated with p1300-TbYDV-Rep/RepA and leaf tissue was collected ten days post-infiltration and examined for the presence of PHB granules. Confocal microscopy and TEM revealed the presence of typical PHB granules in five of the eight lines, thus demonstrating the functionality of InPActbased PHB production in tobacco. However, analysis of leaf extracts by HPLC failed to detect the presence of PHB suggesting only very low level expression levels. Subsequent molecular analysis of three lines revealed low levels of correctly processed mRNA from the catalase intron contained within the InPAct cassette and also the presence of cryptic splice sites within the intron. In an attempt to increase expression levels, new InPAct-phb cassettes were generated in which the castorbean catalase intron was replaced with a synthetic intron (syntron). Further, in an attempt to both increase and better control Rep/RepA-mediated activation of InPAct cassettes, Rep/RepA expression was placed under the control of a stably integrated alc switch. Leaf discs from a transgenic tobacco line (Alc ML) containing 35S P-AlcR-AlcA P-Rep/RepA were supertransformed with InPAct-phbAsyn or InPAct-GUSsyn using Agrobacterium and three plants (lines) were regenerated for each construct. Analysis of the RNA processing of the InPAct-phbAsyn cassette revealed highly efficient and correct splicing of the syntron, thus supporting its inclusion within the InPAct system. To determine the efficiency of the alc switch to activate InPAct, leaf material from the three Alc ML + InPAct-phbAsyn lines was either agroinfiltrated with 35S P-Rep/RepA or treated with ethanol. Unexpectedly, episomes were detected not only in the infiltrated and ethanol treated samples, but also in non-treated samples. Subsequent analysis of transgenic Alc ML + InPAct-GUS lines, confirmed that the alc switch was leaky in tissue culture. Although this was shown to be reversible once plants were removed from the tissue culture environment, it made the regeneration of Alc ML + InPAct-phbsyn plant lines extremely difficult, due to unintentional Rep expression and therefore high levels of phb expression and phytotoxic PHB production. Two Alc ML + InPAct-phbAsyn + p1300-TaBV P-phbB/phbC-35S T transgenic lines were able to be regenerated, and these were acclimatised, alcohol-treated and analysed. Although episome formation was detected as late as 21 days post activation, no PHB was detected in the leaves of any plants using either microscopy or HPLC, suggesting the presence of a corrupt InPAct-phbA cassette in both lines. The final component of this thesis involved the application of both the alc switch and the InPAct systems to sugarcane in an attempt to produce PHB. Initial experiments using transgenic Alc ML + InPAct-GUS lines indicated that the alc system was not functional in sugarcane under the conditions tested. The functionality of the InPAct system, independent of the alc gene switch, was subsequently examined by bombarding the 35S Rep/RepA cassette into leaf and immature leaf whorl cells derived from InPAct-GUS transgenic sugarcane plants. No GUS expression was observed in leaf tissue, whereas weak and irregular GUS expression was observed in immature leaf whorl tissue derived from two InPAct- GUS lines and two InPAct-GUS + 35S P-AlcR-AlcA P-GUS lines. The most plausible reason to explain the inconsistent and low levels of GUS expression in leaf whorls is a combination of low numbers of sugarcane cells in the DNA replication-conducive S-phase and the irregular and random nature of sugarcane cells bombarded with Rep/RepA. This study details the first report to develop a TbYDV-based InPAct system under control of the alc switch to produce PHB in tobacco and sugarcane. Despite the inability to detect quantifiable levels of PHB levels in either tobacco or sugarcane, the findings of this study should nevertheless assist in the further development of both the InPAct system and the alc system, particularly for sugarcane and ultimately lead to an ethanol-inducible InPAct gene expression system for the production of bioplastics and other proteins of commercial value in plants.

Development of a high-speed sensing and detection system for automatic removal of packages with leaky seals from a high-speed food processing and packaging line

Contamination of packaged foods due to micro-organisms entering through air leaks can cause serious public health issues and cost companies large amounts of money due to product recalls, consumer impact and subsequent loss of market share. The main source of contamination is leaks in packaging which allow air, moisture and microorganisms to enter the package. In the food processing and packaging industry worldwide, there is an increasing demand for cost effective state of the art inspection technologies that are capable of reliably detecting leaky seals and delivering products at six-sigma. The new technology will develop non-destructive testing technology using digital imaging and sensing combined with a differential vacuum technique to assess seal integrity of food packages on a high-speed production line. The cost of leaky packages in Australian food industries is estimated close to AUD $35 Million per year. Contamination of packaged foods due to micro-organisms entering through air leaks can cause serious public health issues and cost companies large sums of money due to product recalls, compensation claims and loss of market share. The main source of contamination is leaks in packaging which allow air, moisture and micro-organisms to enter the package. Flexible plastic packages are widely used, and are the least expensive form of retaining the quality of the product. These packets can be used to seal, and therefore maximise, the shelf life of both dry and moist products. The seals of food packages need to be airtight so that the food content is not contaminated due to contact with microorganisms that enter as a result of air leakage. Airtight seals also extend the shelf life of packaged foods, and manufacturers attempt to prevent food products with leaky seals being sold to consumers. There are many current NDT (non-destructive testing) methods of testing the seal of flexible packages best suited to random sampling, and for laboratory purposes. The three most commonly used methods are vacuum/pressure decay, bubble test, and helium leak detection. Although these methods can detect very fine leaks, they are limited by their high processing time and are not viable in a production line. Two nondestructive in-line packaging inspection machines are currently available and are discussed in the literature review. The detailed design and development of the High-Speed Sensing and Detection System (HSDS) is the fundamental requirement of this project and the future prototype and production unit. Successful laboratory testing was completed and a methodical design procedure was needed for a successful concept. The Mechanical tests confirmed the vacuum hypothesis and seal integrity with good consistent results. Electrically, the testing also provided solid results to enable the researcher to move the project forward with a certain amount of confidence. The laboratory design testing allowed the researcher to confirm theoretical assumptions before moving into the detailed design phase. Discussion on the development of the alternative concepts in both mechanical and electrical disciplines enables the researcher to make an informed decision. Each major mechanical and electrical component is detailed through the research and design process. The design procedure methodically works through the various major functions both from a mechanical and electrical perspective. It opens up alternative ideas for the major components that although are sometimes not practical in this application, show that the researcher has exhausted all engineering and functionality thoughts. Further concepts were then designed and developed for the entire HSDS unit based on previous practice and theory. In the future, it would be envisaged that both the Prototype and Production version of the HSDS would utilise standard industry available components, manufactured and distributed locally. Future research and testing of the prototype unit could result in a successful trial unit being incorporated in a working food processing production environment. Recommendations and future works are discussed, along with options in other food processing and packaging disciplines, and other areas in the non-food processing industry.

On-line film distribution : Its history and global complexion

‘Nobody knows anything’, said William Goldman of studio filmmaking. The rule is ever more apt as we survey the radical changes that digital distribution, along with the digitisation of production and exhibition, is wreaking on global film circulation. Digital Disruption: Cinema Moves On-line helps to make sense of what has happened in the short but turbulent history of on-line distribution. It provides a realistic assessment of the genuine and not-so-promising methods that have been tried to address the disruptions that moving from ‘analogue dollars’ to ‘digital cents’ has provoked in the film industry. Paying close attention to how the Majors have dealt with the challenges – often unsuccessfully – it focuses as much attention on innovations and practices outside the mainstream. Throughout, it is alive to, and showcases, important entrepreneurial innovations such as Mubi, Jaman, Withoutabox and IMDb. Written by leading academic commentators that have followed the fortunes of world cinema closely and passionately, as well as experienced hands close to the fluctuating fortunes of the industry, Digital Disruption: Cinema Moves On-line is an indispensable guide to great changes in film and its audiences.

Assembly line balancing the comparison of COMSOAL and MSNSH technique in Motorcycle manufacturing company

Today’s highly competitive market influences the manufacturing industry to improve their production systems to become the optimal system in the shortest cycle time as possible. One of most common problems in manufacturing systems is the assembly line balancing problem. The assembly line balancing problem involves task assignments to workstations with optimum line efficiency. The line balancing technique, namely “COMSOAL”, is an abbreviation of “Computer Method for Sequencing Operations for Assembly Lines”. Arcus initially developed the COMSOAL technique in 1966 [1], and it has been mainly applied to solve assembly line balancing problems [6]. The most common purposes of COMSOAL are to minimise idle time, optimise production line efficiency, and minimise the number of workstations. Therefore, this project will implement COMSOAL to balance an assembly line in the motorcycle industry. The new solution by COMSOAL will be used to compare with the previous solution that was developed by Multi‐Started Neighborhood Search Heuristic (MSNSH), which will result in five aspects including cycle time, total idle time, line efficiency, average daily productivity rate, and the workload balance. The journal name “Optimising and simulating the assembly line balancing problem in a motorcycle manufacturing company: a case study” will be used as the case study for this project [5].

Optimization of chimeric HIV-1 virus-like particle production in a baculovirus-insect cell expression system

A baculovirus-insect cell expression system potentially provides the means to produce prophylactic HIV-1 virus-like particle (VLP) vaccines inexpensively and in large quantities. However, the system must be optimized to maximize yields and increase process efficiency. In this study, we optimized the production of two novel, chimeric HIV-1 VLP vaccine candidates (GagRT and GagTN) in insect cells. This was done by monitoring the effects of four specific factors on VLP expression: these were insect cell line, cell density, multiplicity of infection (MOI), and infection time. The use of western blots, Gag p24 ELISA, and four-factorial ANOVA allowed the determination of the most favorable conditions for chimeric VLP production, as well as which factors affected VLP expression most significantly. Both VLP vaccine candidates favored similar optimal conditions, demonstrating higher yields of VLPs when produced in the Trichoplusia ni Pro insect cell line, at a cell density of 1 × 106 cells/mL, and an infection time of 96 h post infection. It was found that cell density and infection time were major influencing factors, but that MOI did not affect VLP expression significantly. This work provides a potentially valuable guideline for HIV-1 protein vaccine optimization, as well as for general optimization of a baculovirus-based expression system to produce complex recombinant proteins. © 2009 American Institute of Chemical Engineers.

Optimising preventive maintenance strategy for production Lines

Preventive Maintenance (PM) is often applied to improve the reliability of production lines. A Split System Approach (SSA) based methodology is presented to assist in making optimal PM decisions for serial production lines. The methodology treats a production line as a complex series system with multiple (imperfect) PM actions over multiple intervals. The conditional and overall reliability of the entire production line over these multiple PM intervals are hierarchically calculated using SSA, and provide a foundation for cost analysis. Both risk-related cost and maintenance-related cost are factored into the methodology as either deterministic or random variables. This SSA based methodology enables Asset Management (AM) decisions to be optimised considering a variety of factors including failure probability, failure cost, maintenance cost, PM performance, and the type of PM strategy. The application of this new methodology and an evaluation of the effects of these factors on PM decisions are demonstrated using an example. The results of this work show that the performance of a PM strategy can be measured by its Total Expected Cost Index (TECI). The optimal PM interval is dependent on TECI, PM performance and types of PM strategies. These factors are interrelated. Generally, it was found that a trade-off between reliability and the number of PM actions needs to be made so that one can minimise Total Expected Cost (TEC) for asset maintenance.

Procuring OSM : base-line models of off-site manufacture business processes in Australia

A pressing cost issue facing construction is the procurement of off-site pre-manufactured assemblies. In order to encourage Australian adoption of off-site manufacture (OSM), a new approach to underlying processes is required. The advent of object oriented digital models for construction design assumes intelligent use of data. However, the construction production system relies on traditional methods and data sources and is expected to benefit from the application of well-established business process management techniques. The integration of the old and new data sources allows for the development of business process models which, by capturing typical construction processes involving OSM, provides insights into such processes. This integrative approach is the foundation of research into the use of OSM to increase construction productivity in Australia. The purpose of this study is to develop business process models capturing the procurement, resources and information flow of construction projects. For each stage of the construction value chain, a number of sub-processes are identified. Business Process Modelling Notation (BPMN), a mainstream business process modelling standard, is used to create base-line generic construction process models. These models identify OSM decision-making points that could provide cost reductions in procurement workflow and management systems. This paper reports on phase one of an on-going research aiming to develop a proto-type workflow application that can provide semi-automated support to construction processes involving OSM and assist in decision-making in the adoption of OSM thus contributing to a sustainable built environment.

Traits of fear resistance and susceptibility in an advanced intercross line

Genetic variability in the strength and precision of fear memory is hypothesised to contribute to the etiology of anxiety disorders, including post-traumatic stress disorder. We generated fear-susceptible (F-S) or fear-resistant (F-R) phenotypes from an F8 advanced intercross line (AIL) of C57BL/6J and DBA/2J inbred mice by selective breeding. We identified specific traits underlying individual variability in Pavlovian conditioned fear learning and memory. Offspring of selected lines differed in the acquisition of conditioned fear. Furthermore, F-S mice showed greater cued fear memory and generalised fear in response to a novel context than F-R mice. F-S mice showed greater basal corticosterone levels and hypothalamic corticotrophin-releasing hormone (CRH) mRNA levels than F-R mice, consistent with higher hypothalamic-pituitary-adrenal (HPA) axis drive. Hypothalamic mineralocorticoid receptor and CRH receptor 1 mRNA levels were decreased in F-S mice as compared with F-R mice. Manganese-enhanced magnetic resonance imaging (MEMRI) was used to investigate basal levels of brain activity. MEMRI identified a pattern of increased brain activity in F-S mice that was driven primarily by the hippocampus and amygdala, indicating excessive limbic circuit activity in F-S mice as compared with F-R mice. Thus, selection pressure applied to the AIL population leads to the accumulation of heritable trait-relevant characteristics within each line, whereas non-behaviorally relevant traits remain distributed. Selected lines therefore minimise false-positive associations between behavioral phenotypes and physiology. We demonstrate that intrinsic differences in HPA axis function and limbic excitability contribute to phenotypic differences in the acquisition and consolidation of associative fear memory. Identification of system-wide traits predisposing to variability in fear memory may help in the direction of more targeted and efficacious treatments for fear-related pathology. Through short-term selection in a B6D2 advanced intercross line we created mouse populations divergent for the retention of Pavlovian fear memory. Trait distinctions in HPA-axis drive and fear network circuitry could be made between naïve animals in the two lines. These data demonstrate underlying physiological and neurological differences between Fear-Susceptible and Fear-Resistant animals in a natural population. F-S and F-R mice may therefore be relevant to a spectrum of disorders including depression, anxiety disorders and PTSD for which altered fear processing occurs.

A broad-scale analysis of links between coastal fisheries production and mangrove extent : a case-study for northeastern Australia

The paradigm that mangroves are critical for sustaining production in coastal fisheries is widely accepted, but empirical evidence has been tenuous. This study showed that links between mangrove extent and coastal fisheries production could be detected for some species at a broad regional scale (1000s of kilometres) on the east coast of Queensland, Australia. The relationships between catch-per-unit-effort for different commercially caught species in four fisheries (trawl, line, net and pot fisheries) and mangrove characteristics, estimated from Landsat images were examined using multiple regression analyses. The species were categorised into three groups based on information on their life history characteristics, namely mangrove-related species (banana prawns Penaeus merguiensis, mud crabs Scylla serrata and barramundi Lates calcarifer), estuarine species (tiger prawns Penaeus esculentus and Penaeus semisulcatus, blue swimmer crabs Portunus pelagicus and blue threadfin Eleutheronema tetradactylum) and offshore species (coral trout Plectropomus spp.). For the mangrove-related species, mangrove characteristics such as area and perimeter accounted for most of the variation in the model; for the non-mangrove estuarine species, latitude was the dominant parameter but some mangrove characteristics (e.g. mangrove perimeter) also made significant contributions to the models. In contrast, for the offshore species, latitude was the dominant variable, with no contribution from mangrove characteristics. This study also identified that finer scale spatial data for the fisheries, to enable catch information to be attributed to a particular catchment, would help to improve our understanding of relationships between mangroves and fisheries production.

Dancing as Knowledge Production/Dancer as Researcher in Practice-led Research

This chapter discusses research undertaken for a PhD in dance, highlighting the oscillating and ambivalent nature of practice-led research methods. Holding an unusual position within the field of practice-led research in dance, the researcher adopted the role of dancer within four creative processes led by four choreographers. Although instigating and leading this research, the author produced new knowledge of dancing practices whilst being directed creatively by the choreographers in the research environment. In line with many definitions of practice-led research, the methods used were emergent from the research arena and responsive to the requirements of the project. Deeply imbricated in the research environment, the researcher used her embodied self as a research tool, as the one who participated, discovered and recorded. This outsider/insider role of researcher/participant produced a series of meta-positions, leading to a mixed-modal outcome that encompassed live performance, a movement treatise and a written thesis.

Renewable energy distribution in public spaces: Analyzing the case of Ballast Point Park in Sydney, using a triple bottom line approach

As cities are rapidly developing new interventions against climate change, embedding renewable energy in public spaces is an important strategy. However, most interventions primarily include environmental sustainability while neglecting the social and economic interrelationships of electricity production. Although there is a growing interest in sustainability within environmental design and landscape architecture, public spaces are still awaiting viable energy-conscious design and assessment interventions. The purpose of this paper is to investigate this issue in a renowned public space—Ballast Point Park in Sydney—using a triple bottom line (TBL) case study approach. The emerging factors and relationships of each component of TBL, within the context of public open space, are identified and discussed. With specific focus on renewable energy distribution in and around Ballast Point Park, the paper concludes with a general design framework, which conceptualizes an optimal distribution of onsite electricity produced from renewable sources embedded in public open spaces.

Cytokine mRNA expression and IFN-γ production of immunised nematode resistant and susceptible lambs against natural poly-generic challenge

IL-2, IL-4 and IFN-γ mRNA expression, and production of IFN-γ was examined in mesenteric lymph node cells (MLNC) and CD4+ enriched T cell populations of nematode resistant (R) and susceptible (S) line lambs by use of RT-PCR and ELISA. Five R and S line lambs that were immunised by repeated oxfendazole-abbreviated infections and 5 non-immunised R and S line lambs were used. All lambs grazed nematode infected pasture for 107 days. Immunisation enhanced the resistant status in both R and S lambs. MLNC obtained from slaughtered animals were stimulated with Con A or T. colubriformis specific antigen. Non-stimulated MLNC of immunised lambs expressed higher levels of IL-4 mRNA and lower levels of IL-2 mRNA than non-immunised lambs. MLNC of immunised R and S line lambs stimulated with antigen for 24 h expressed detectable amounts of IL-4 mRNA that was not seen in non-immunised controls. CD4+ T cell enriched cell populations of immunised R and S lambs and non-immunised R lambs expressed moderate to high levels of IL-4 mRNA. Con A stimulated MLNC of immunised R and S lambs expressed high levels IFN-γ mRNA and produced high amounts of IFN-γ. Lower levels were present in non-immunised controls. The results indicate that R line lambs and immunised S line lambs respond to natural nematode challenge with a predominating IL-4 cytokine response when compared to non-immunised S lambs.

Evaluation of the most effective measures of infectivity of a precocious line of Eimeria acervulina in poultry

Coccidiosis is an economically important parasitic disease of chickens that, in Australia, is caused by seven species of the genus Eimeria.1 The disease has traditionally been controlled by prophylactic drugs, but vaccination with attenuated lines of the parasites2–4 is rapidly gaining acceptance world wide. Live Eimeria vaccines are produced in batches which are not frozen and have a limited shelf life. The per cent infectivity of vaccine seed stocks and the vaccines produced from them must therefore be accurately monitored using standardised dose dependant assays to ensure that shelf life, quality control and vaccine release specifications are met. Infectivity for the chicken host cannot readily be determined by microscopic observation of oocysts or sporocyst hatching.5 Dose dependent parameters such as body weight gain, feed conversion ratio, visual lesion scores, mortality, oocysts production, clinical symptoms and microscopic lesion counts could be used as measures of infectivity.6–11 These parameters show significant dose dependant effects with field strains, but lines of vaccine parasites that have been selected for precocious development with associated reduced virulence and reproductive capability may not have the same effect.3,4 The aim of this trial was to determine which parameters provide the most effective measures of infective dose in birds inoculated with a precocious vaccine strain.

Genetic differentiation analysis for the identification of complementary parental pools for sorghum hybrid breeding in Ethiopia

Key message The potential for exploiting heterosis for sorghum hybrid production in Ethiopia with improved local adaptation and farmers preferences has been investigated and populations suitable for initial hybrid development have been identified. Abstract Hybrids in sorghum have demonstrated increased productivity and stability of performance in the developed world. In Ethiopia, the uptake of hybrid sorghum has been limited to date, primarily due to poor adaptation and absence of farmer’s preferred traits in existing hybrids. This study aimed to identify complementary parental pools to develop locally adapted hybrids, through an analysis of whole genome variability of 184 locally adapted genotypes and introduced hybrid parents (R and B). Genetic variability was assessed using genetic distance, model-based STRUCTURE analysis and pair-wise comparison of groups. We observed a high degree of genetic similarity between the Ethiopian improved inbred genotypes and a subset of landraces adapted to lowland agro-ecology with the introduced R lines. This coupled with the genetic differentiation from existing B lines, indicated that these locally adapted genotype groups are expected to have similar patterns of heterotic expression as observed between introduced R and B line pools. Additionally, the hybrids derived from these locally adapted genotypes will have the benefit of containing farmers preferred traits. The groups most divergent from introduced B lines were the Ethiopian landraces adapted to highland and intermediate agro-ecologies and a subset of lowland-adapted genotypes, indicating the potential for increased heterotic response of their hybrids. However, these groups were also differentiated from the R lines, and hence are different from the existing complementary heterotic pools. This suggests that although these groups could provide highly divergent parental pools, further research is required to investigate the extent of heterosis and their hybrid performance.