Development of a Rep-inducible, BBTV-based expression system in banana

Banana bunchy top is regarded as the most important viral disease of banana, causing significant yield losses worldwide. The disease is caused by Banana bunchy top virus (BBTV), which is a circular ssDNA virus belonging to the genus Babuvirus in the family Nanoviridae. There are currently few effective control strategies for this and other ssDNA viruses. “In Plant Activation” (InPAct) is a novel technology being developed at QUT for ssDNA virus-activated suicide gene expression. The technology exploits the rolling circle replication mechanism of ssDNA viruses and is based on a unique “split” gene design such that suicide gene expression is only activated in the presence of the viral Rep. This PhD project aimed to develop a BBTV-based InPAct system as a suicide gene strategy to control BBTV. The BBTV-based InPAct vector design requires a BBTV intergenic region (IR) to be embedded within an intron in the gene expression cassette. To ensure that the BBTV IR would not interfere with intron splicing, a TEST vector was initially generated that contained the entire BBTV IR embedded within an intron in a β-glucuronidase (GUS) expression vector. Transient GUS assays in banana embryogenic cell suspensions indicated that cryptic intron splice sites were present within the IR. Transcript analysis revealed two cryptic intron splice sites in the Domain III sequence of the CR-M within the IR. Removal of the CR-M from the TEST vector resulted in an enhancement of GUS expression suggesting that the cryptic intron splice sites had been removed. An InPAct GUS vector was subsequently generated that contained the modified BBTV IR, with the CR-M (minus Domain III) repositioned within the InPAct cassette. Using transient histochemical and fluorometric GUS assays in banana embryogenic cells, the InPAct GUS vector was shown to be activated in the presence of the BBTV Rep. However, the presence of both BBTV Rep and Clink was shown to have a deleterious effect on GUS expression suggesting that these proteins were cytotoxic at the levels expressed. Analysis of replication of the InPAct vectors by Southern hybridisation revealed low levels of InPAct cassette-based episomal DNA released from the vector through the nicking/ligation activity of BBTV Rep. However, Rep-mediated episomal replicons, indicative of rolling circle replication of the released circularised cassettes, were not observed. The inability of the InPAct cassette to be replicated was further investigated. To examine whether the absence of Domain III of the CR-M was responsible, a suite of modified BBTV-based InPAct GUS vectors was constructed that contained the CR-M with the inclusion of Domain III, the CR-M with the inclusion of Domain III and additional upstream IR sequence, or no CR-M. Analysis of replication by Southern hybridisation revealed that neither the presence of Domain III, nor the entire CR-M, had an effect on replication levels. Since the InPAct cassette was significantly larger than the native BBTV genomic components (approximately 1 kb), the effect of InPAct cassette size on replication was also investigated. A suite of size variant BBTV-based vectors was constructed that increased the size of a replication competent cassette to 1.1 kbp through to 2.1 kbp.. Analysis of replication by Southern hybridisation revealed that an increase in vector size above approximately 1.5 - 1.7 kbp resulted in a decrease in replication. Following the demonstration of Rep-mediated release, circularisation and expression from the InPAct GUS vector, an InPAct vector was generated in which the uidA reporter gene was replaced with the ribonuclease-encoding suicide gene, barnase. Initially, a TEST vector was generated to assess the cytotoxicity of Barnase on banana cells. Although transient assays revealed a Barnase-induced cytotoxic effect in banana cells, the expression levels were sub-optimal. An InPAct BARNASE vector was generated and tested for BBTV Rep-activated Barnase expression using transient assays in banana embryogenic cells. High levels of background expression from the InPAct BARNASE vector made it difficult to accurately assess Rep-activated Barnase expression. Analysis of replication by Southern hybridisation revealed low levels of InPAct cassette-based episomal DNA released from the vector but no Rep-mediated episomal replicons indicative of rolling circle replication of the released circularised cassettes were again observed. Despite the inability of the InPAct vectors to replicate to enable high level gene expression, the InPAct BARNASE vector was assessed in planta for BBTV Rep-mediated activation of Barnase expression. Eleven lines of transgenic InPAct BARNASE banana plants were generated by Agrobacterium-mediated transformation and were challenged with viruliferous Pentalonia nigronervosa. At least one clonal plant in each line developed bunchy top symptoms and infection was confirmed by PCR. No localised lesions were observed on any plants, nor was there any localised GUS expression in the one InPAct GUS line challenged with viruliferous aphids. The results presented in this thesis are the first study towards the development of a BBTV-based InPAct system as a Rep-activatable suicide gene expression system to control BBTV. Although further optimisation of the vectors is necessary, the preliminary results suggest that this approach has the potential to be an effective control strategy for BBTV. The use of iterons within the InPAct vectors that are recognised by Reps from different ssDNA plant viruses may provide a broad-spectrum resistance strategy against multiple ssDNA plant viruses. Further, this technology holds great promise as a platform technology for the molecular farming of high-value proteins in vitro or in vivo through expression of the ssDNA virus Rep protein.

Older patients request more information : a survey of use of written patient education materials in general practice

Objectives: To determine GPs' reported use of written education materials with older patients and older patients' reported receipt of these materials. To determine GPs' and older patients' perceptions of written materials.---------- Method: Using self-report questionnaires, two populations were surveyed; a randomised sample of 50 GPs (29 males and 21 females) practising in Brisbane's southern suburbs and a convenience sample of 188 older community-dwelling people (aged over 64 years).----------- Results: All GPs reported using written materials with patients, although 28% had not given any to the Last 10 patients. This increased to 46% when patients were older. Twenty percent of patients wanted more written information from their GP, while some GPs believed that older patients preferred verbal information and gave out written information only when they perceived patient interest. All GPs reported giving written materials at the time of consultation and over two thirds discussed the content with patients. Just over 50% of patients reported receiving written information from GPs in the Last six months and only hall of these again discussed it directly with their GP. Overall, patients were more positive than GPs about the value of written education materials.---------- Conclusions: Older patients' desire for written information may be better met if they are more assertive in requesting this of GPs and GPs may better serve their patients' needs if they make written information more readily available to them. Better access to materials and more financial incentives to give them out might also increase GPs' use of written materials.

The condition, development and suggestions of policy of the highway express freight transportation system in China

The highway express freight transportation (HEFT) is a new transportation organization form separated from the common freight transportation with economic development and incessant adjustment of highway transportation structure in China. At present, the phenomenon of inadaptability still exists in the HEFT system of China, from foundation structure like highways, parking lots and stations to transportation equipments and transportation organizing. In order to develop the HEFT system more rationally and effectively, we should start with the structure of the system, conform the resources existing, and consummate the freight transport system. In due course, relevant policies and measures to supervise, lead and support are necessary and important. This paper analyzes the existing problems of HEFT system in our country, based on its characteristics, development situation and adaptability, and presents the policy and measures of promoting and leading the development of the HEFT system.

The genesis of science education research in Australasia

Although the sciences were being taught in Australian schools well before the Second World War, the only evidence of research studies of this teaching is to be found in the report, published by ACER in 1932 of Roy Stanhope’s survey of the teaching of chemistry in New South Wales and a standardized test he had developed. Roy Stanhope was a science teacher with a research masters degree in chemistry. He had won a scholarship to go to Stanford University for doctoral studies, but returned after one year when his scholarship was not extended. He went on to be a founder in 1943 of the Australian Science Teachers Association (ASTA), which honours this remarkable pioneer through its annual Stanhope Oration. In his retirement Stanhope undertook a comparative study of science

Introduction : science education research in Australasia : an overview

This chapter overviews the major themes of research reviewed and justifies the selection of topics.

Parents’ choice of early childhood education service in Hong Kong : a pilot study

The introduction of a voucher scheme for early childhood education in Hong Kong has resulted in significant changes in the field. This paper reports data from a pilot study that aimed at gaining a deeper understanding of how parents chose early childhood education service following the introduction of a voucher scheme in Hong Kong. Eight-six Chinese parents with children aged three participated in interviews and focus group discussions. This group of parents had just gone through the process of selecting a kindergarten or nursery for their child for the school year of 2007-2008. Parents from a range of socioeconomic circumstances and educational levels who had selected non-profit kindergartens and nurseries in public and private housing estates participated. Results showed that what parents looked in their choice of service matched closely with how they defined quality. As evidenced in the study, parents’ changing views on quality shared a great deal of resemblance with the specific notion of quality being heavily promoted by recent reform policy. The findings pointed to the complex interactions of policy, choice and practices of early childhood education. The new voucher scheme is intensifying the governing of the self and the field, the impact of which can be worrying.

Practical Transformations and Transformational Practices : Globalization, Postmodernism, and Early Childhood Education : Advances in Early Education and Day Care

Both traditional and progressive curricula are inadequate for the task of responding to the economic, political, social, and cultural changes that have occurred as a result of globalization. This book documents some of the ongoing work occurring in early childhood settings that is aimed at improving, and ultimately transforming, early childhood practice in these changed and changing times. The authors do not simply critique developmental approaches or the increasing standardization of the field. Instead, they describe how they are playing around with postmodern ideas in practice and developing unique approaches to the diverse educational circumstances that confront early childhood educators. Whether it is preparing teachers, using materials, or developing policies, each chapter provides readers with possibilities for enacting pedagogies that are responsive to the contemporary circumstances shaping the lives of young children.

Aligning school reform and teacher education reform in the middle years : an Australian case study

This paper discusses the development of a new Bachelor of Education (Middle Years of Schooling) at The University of Queensland, Australia. The middle years of schooling have increasingly been the focus of education reform initiatives in Australia, but this has not been accompanied by significant increases in the number of teacher education institutions offering specialised middle schooling-level teacher preparation programmes. Considering the rapidly changing social and economic context and the emergent state of middle schooling in Australia, the programme represented a conceptual and practical opportunity and challenge for The University of Queensland team. Working collaboratively, the team sought to design a teacher education preservice programme that was both responsive and generative: that is, responsive to local school contexts and to current education research and reform at national and international levels; and generative of cutting-edge theories and practices associated with middle schooling, teachers' work, and teacher education. This paper focuses on one component of the Middle Years of Schooling Teacher Education programme at The University of Queensland; namely, the practicum. We first present the underlying principles of the practicum programme and then examine "dilemmas" that emerged early in the practicum. These issues and tensions were associated with the ideals of "middle years" philosophy and the pragmatics of school reform associated with that approach. Within this context, we explore what it means to be both responsive and generative, and describe how we as teacher educators negotiated the extremes these terms implied.

Mainstreaming sustainability into pre-service teacher edcuation in Australia

There is a growing interest in and support for education for sustainability in Australian schools. Australian Government schemes such as the Australian Sustainable Schools Initiative (AuSSI), along with strategies such as Educating for a Sustainable Future: A National Environmental Education Statement for Australian Schools(NEES(Australian Government and Curriculum Corporation (2005) and Living Sustainably: The Australian Government’s National Action Plan for Education for Sustainability (Australian Government 2009), recognise the need and offer support for education for sustainability in Australian schools. The number of schools that have engaged with AuSSI indicates that this interest also exists within Australian schools. Despite this, recent research indicates that pre-service teacher education institutions and programs are not doing all they can to prepare teachers for teaching education for sustainability or for working within sustainable schools. The education of school teachers plays a vital role in achieving changes in teaching and learning in schools. Indeed, the professional development of teachers in education for sustainability has been identified as ‘the priority of priorities’. Much has been written about the need to ‘reorient teacher education towards sustainability’. Teacher education is seen as a key strategy that is yet to be effectively utilised to embed education for sustainability in schools. Mainstreaming sustainability in Australian schools will not be achieved without the preparation of teachers for this task. The Mainstreaming Sustainability model piloted in this study seeks to engage a range of stakeholder organisations and key agents of change within a system to all work simultaneously to bring about a change, such as the mainstreaming of sustainability. The model is premised on the understanding that sustainability will be mainstreamed within teacher education if there is engagement with key agents of change across the wider teacher education system and if the key agents of change are ‘deeply’ involved in making the change. The model thus seeks to marry broad engagement across a system with the active participation of stakeholders within that system. Such a systemic approach is a way of bringing together diverse viewpoints to make sense of an issue and harness that shared interpretation to define boundaries, roles and relationships leading to a better defined problem that can be acted upon more effectively. Like action research, the systemic approach is also concerned with modelling change and seeking plausible solutions through collaboration between stakeholders. This is important in ensuring that outcomes are useful to the researchers/stakeholders and the system being researched as it creates partnerships and commitments to the outcomes by stakeholder participants. The study reported on here examines whether the ‘Mainstreaming Sustainability’ model might be effective as a means to mainstream sustainability in pre-service teacher education. This model, developed in an earlier study, was piloted in the Queensland teacher education system in order to examine its effectiveness in creating organisational and systemic change. The pilot project in Queensland achieved a number of outcomes. The project: • provided useful insights into the effectiveness of the Mainstreaming Sustainability model in bringing about change while also building research capacity within the system • developed capacities within the teacher education community: o developing competencies in education for sustainability o establishing more effective interactions between decision-makers and other stakeholders o establishing a community of inquiry • changed teaching and learning approaches used in participating teacher education institutions through: o curriculum and resource development o the adoption of education for sustainability teaching and learning processes o the development of institutional policies • improved networks within the teacher education system through: o identifying key agents of change within the system o developing new, and building on existing, partnerships between schools, teacher education institutions and government agencies • engaged relevant stakeholders such as government agencies and non-government organisations to understand and support the change Our findings indicate that the Mainstreaming Sustainability model is able to facilitate organisational and systemic change – over time – if: • the individuals involved have the conceptual and personal capacities needed to facilitate change, that is, to be a key agent of change • stakeholders are engaged as participants in the process of change, not simply as ‘interested parties’ • there is a good understanding of systemic change and the opportunities for leveraging change within systems. In particular, in seeking to mainstream sustainability in pre-service teacher education in Queensland it has become clear that one needs to build capacity for change within participants such as knowledge of education for sustainability, conceptual skills in systemic thinking, action research and organisational change, and leadership skills. It is also of vital importance that key agents of change – those individuals who are ‘hubs’ within a system and can leverage for change across a wide range of the system – are identified and engaged with as early as possible. Key agents of change can only be correctly identified, however, if the project leaders and known participants have clearly identified the boundary to their system as this enables the system, sub-system and environment of the system to be understood. Through mapping the system a range of key organisations and stakeholders will be identified, including government and nongovernment organisations, teacher education students, teacher education academics, and so on. On this basis, key agents of change within the system and sub-system can be identified and invited to assist in working for change. A final insight is that it is important to have time – and if necessary the funding to ‘buy time’ – in seeking to bring about system-wide change. Seeking to bring about system-wide change is an ambitious project, one that requires a great deal of effort and time. These insights provide some considerations for those seeking to utilise the Mainstreaming Sustainability model to bring about change within and across a pre-service teacher education system.

Queensland Emergency Medical System : a structural and organizational model for the emergency medical system in Australia

Objective: The emergency medical system (EMS) can be defined as a comprehensive, coordinated and integrated system of care for patients suffering acute illness and injury. The aim of the present paper is to describe the evolution of the Queensland Emergency Medical System (QEMS) and to recommend a strategic national approach to EMS development. Methods: Following the formation of the Queensland Ambulance Service in 1991, a state EMS committee was formed. This committee led the development and approval of the cross portfolio QEMS policy framework that has resulted in dynamic policy development, system monitoring and evaluation. This framework is led by the Queensland Emergency Medical Services Advisory Committee. Results: There has been considerable progress in the development of all aspects of the EMS in Queensland. These developments have derived from the improved coordination and leadership that QEMS provides and has resulted in widespread satisfaction by both patients and stakeholders. Conclusions: The strategic approach outlined in the present paper offers a model for EMS arrangements throughout Australia. We propose that the Council of Australian Governments should require each state and Territory to maintain an EMS committee. These state EMS committees should have a broad portfolio of responsibilities. They should provide leadership and direction to the development of the EMS and ensure coordination and quality of outcomes. A national EMS committee with broad representation and broad scope should be established to coordinate the national development of Australia's EMS.

Implementing an employee performance management system in a nonprofit organization

Nonprofit organizations are not exempt from the imperatives of employee attraction, retention, and motivation. As competition for staff, donors, and funding increases, the need to manage employee performance will continue to be a critical human resource management issue. This article outlines a study of the introduction of a performance management system in an Australian nonprofit organization and analyzes its design and implementation. It explores how performance management can be introduced and used effectively within a nonprofit environment to benefit staff and the organization. However, the use of performance management is not without its challenges, and the research also identified initial employee resistance and a resulting initial spike in labor turnover. However, findings indicate that if nonprofit organizations are willing to undertake consultation with staff and ensure that the organization's specific context, values, and mission are reflected in the performance management system, it can be a useful tool for managers and a direct benefit to employees.