The transformation of banana with potential virus resistance genes

One approach to reducing the yield losses caused by banana viral diseases is the use of genetic engineering and pathogen-derived resistance strategies to generate resistant cultivars. The development of transgenic virus resistance requires an efficient banana transformation method, particularly for commercially important 'Cavendish' type cultivars such as 'Grand Nain'. Prior to this study, only two examples of the stable transformation of banana had been reported, both of which demonstrated the principle of transformation but did not characterise transgenic plants in terms of the efficiency at which individual transgenic lines were generated, relative activities of promoters in stably transformed plants, and the stability of transgene expression. The aim of this study was to develop more efficient transformation methods for banana, assess the activity of some commonly used and also novel promoters in stably transformed plants, and transform banana with genes that could potentially confer resistance to banana bunchy top nanovirus (BBTV) and banana bract mosaic potyvirus (BBrMV). A regeneration system using immature male flowers as the explant was established. The frequency of somatic embryogenesis in male flower explants was influenced by the season in which the inflorescences were harvested. Further, the media requirements of various banana cultivars in respect to the 2,4-D concentration in the initiation media also differed. Following the optimisation of these and other parameters, embryogenic cell suspensions of several banana (Musa spp.) cultivars including 'Grand Nain' (AAA), 'Williams' (AAA), 'SH-3362' (AA), 'Goldfinger' (AAAB) and 'Bluggoe' (ABB) were successfully generated. Highly efficient transformation methods were developed for both 'Bluggoe' and 'Grand Nain'; this is the first report of microprojectile bombardment transformation of the commercially important 'Grand Nain' cultivar. Following bombardment of embryogenic suspension cells, regeneration was monitored from single transfom1ed cells to whole plants using a reporter gene encoding the green fluorescent protein (gfp). Selection with kanamycin enabled the regeneration of a greater number of plants than with geneticin, while still preventing the regeneration of non-transformed plants. Southern hybridisation confirmed the neomycin phosphotransferase gene (npt II) was stably integrated into the banana genome and that multiple transgenic lines were derived from single bombardments. The activity, stability and tissue specificity of the cauliflower mosaic virus 358 (CaMV 35S) and maize polyubiquitin-1 (Ubi-1) promoters were examined. In stably transformed banana, the Ubi-1 promoter provided approximately six-fold higher p-glucuronidase (GUS) activity than the CaMV 35S promoter, and both promoters remained active in glasshouse grown plants for the six months they were observed. The intergenic regions ofBBTV DNA-I to -6 were isolated and fused to either the uidA (GUS) or gfjJ reporter genes to assess their promoter activities. BBTV promoter activity was detected in banana embryogenic cells using the gfp reporter gene. Promoters derived from BBTV DNA-4 and -5 generated the highest levels of transient activity, which were greater than that generated by the maize Ubi-1 promoter. In transgenic banana plants, the activity of the BBTV DNA-6 promoter (BT6.1) was restricted to the phloem of leaves and roots, stomata and root meristems. The activity of the BT6.1 promoter was enhanced by the inclusion of intron-containing fragments derived from the maize Ubi-1, rice Act-1, and sugarcane rbcS 5' untranslated regions in GUS reporter gene constructs. In transient assays in banana, the rice Act-1 and maize Ubi-1 introns provided the most significant enhancement, increasing expression levels 300-fold and 100-fold, respectively. The sugarcane rbcS intron increased expression about 10-fold. In stably transformed banana plants, the maize Ubi-1 intron enhanced BT6.1 promoter activity to levels similar to that of the CaMV 35S promoter, but did not appear to alter the tissue specificity of the promoter. Both 'Grand Nain' and 'Bluggoe' were transformed with constructs that could potentially confer resistance to BBTV and BBrMV, including constructs containing BBTV DNA-1 major and internal genes, BBTV DNA-5 gene, and the BBrMV coat protein-coding region all under the control of the Ubi-1 promoter, while the BT6 promoter was used to drive the npt II selectable marker gene. At least 30 transgenic lines containing each construct were identified and replicates of each line are currently being generated by micropropagation in preparation for virus challenge.

Swept frequency biompedance analysis for the determination of body water compartments

Bioelectrical impedance analysis, (BIA), is a method of body composition analysis first investigated in 1962 which has recently received much attention by a number of research groups. The reasons for this recent interest are its advantages, (viz: inexpensive, non-invasive and portable) and also the increasing interest in the diagnostic value of body composition analysis. The concept utilised by BIA to predict body water volumes is the proportional relationship for a simple cylindrical conductor, (volume oc length2/resistance), which allows the volume to be predicted from the measured resistance and length. Most of the research to date has measured the body's resistance to the passage of a 50· kHz AC current to predict total body water, (TBW). Several research groups have investigated the application of AC currents at lower frequencies, (eg 5 kHz), to predict extracellular water, (ECW). However all research to date using BIA to predict body water volumes has used the impedance measured at a discrete frequency or frequencies. This thesis investigates the variation of impedance and phase of biological systems over a range of frequencies and describes the development of a swept frequency bioimpedance meter which measures impedance and phase at 496 frequencies ranging from 4 kHz to 1 MHz. The impedance of any biological system varies with the frequency of the applied current. The graph of reactance vs resistance yields a circular arc with the resistance decreasing with increasing frequency and reactance increasing from zero to a maximum then decreasing to zero. Computer programs were written to analyse the measured impedance spectrum and determine the impedance, Zc, at the characteristic frequency, (the frequency at which the reactance is a maximum). The fitted locus of the measured data was extrapolated to determine the resistance, Ro, at zero frequency; a value that cannot be measured directly using surface electrodes. The explanation of the theoretical basis for selecting these impedance values (Zc and Ro), to predict TBW and ECW is presented. Studies were conducted on a group of normal healthy animals, (n=42), in which TBW and ECW were determined by the gold standard of isotope dilution. The prediction quotients L2/Zc and L2/Ro, (L=length), yielded standard errors of 4.2% and 3.2% respectively, and were found to be significantly better than previously reported, empirically determined prediction quotients derived from measurements at a single frequency. The prediction equations established in this group of normal healthy animals were applied to a group of animals with abnormally low fluid levels, (n=20), and also to a group with an abnormal balance of extra-cellular to intracellular fluids, (n=20). In both cases the equations using L2/Zc and L2/Ro accurately and precisely predicted TBW and ECW. This demonstrated that the technique developed using multiple frequency bioelectrical impedance analysis, (MFBIA), can accurately predict both TBW and ECW in both normal and abnormal animals, (with standard errors of the estimate of 6% and 3% for TBW and ECW respectively). Isotope dilution techniques were used to determine TBW and ECW in a group of 60 healthy human subjects, (male. and female, aged between 18 and 45). Whole body impedance measurements were recorded on each subject using the MFBIA technique and the correlations between body water volumes, (TBW and ECW), and heighe/impedance, (for all measured frequencies), were compared. The prediction quotients H2/Zc and H2/Ro, (H=height), again yielded the highest correlation with TBW and ECW respectively with corresponding standard errors of 5.2% and 10%. The values of the correlation coefficients obtained in this study were very similar to those recently reported by others. It was also observed that in healthy human subjects the impedance measured at virtually any frequency yielded correlations not significantly different from those obtained from the MFBIA quotients. This phenomenon has been reported by other research groups and emphasises the need to validate the technique by investigating its application in one or more groups with abnormalities in fluid levels. The clinical application of MFBIA was trialled and its capability of detecting lymphoedema, (an excess of extracellular fluid), was investigated. The MFBIA technique was demonstrated to be significantly more sensitive, (P<.05), in detecting lymphoedema than the current technique of circumferential measurements. MFBIA was also shown to provide valuable information describing the changes in the quantity of muscle mass of the patient during the course of the treatment. The determination of body composition, (viz TBW and ECW), by MFBIA has been shown to be a significant improvement on previous bioelectrical impedance techniques. The merit of the MFBIA technique is evidenced in its accurate, precise and valid application in animal groups with a wide variation in body fluid volumes and balances. The multiple frequency bioelectrical impedance analysis technique developed in this study provides accurate and precise estimates of body composition, (viz TBW and ECW), regardless of the individual's state of health.

Overhead electricity line connector tester

Assessment of the condition of connectors in the overhead electricity network has traditionally relied on the heat dissipation or voltage drop from existing load current (50Hz) as a measurable parameter to differentiate between satisfactory and failing connectors. This research has developed a technique which does not rely on the 50Hz current and a prototype connector tester has been developed. In this system a high frequency signal is injected into the section of line under test and measures the resistive voltage drop and the current at the test frequency to yield the resistance in micro-ohms. From the value of resistance a decision as to whether a connector is satisfactory or approaching failure can be made. Determining the resistive voltage drop in the presence of a large induced voltage was achieved by the innovative approach of using a representative sample of the magnetic flux producing the induced voltage as the phase angle reference for the signal processing rather than the phase angle of the current, which can be affected by the presence of nearby metal objects. Laboratory evaluation of the connector tester has validated the measurement technique. The magnitude of the load current (50Hz) has minimal effect on the measurement accuracy. Addition of a suitable battery based power supply system and isolated communications, probably radio and refinement of the printed circuit board design and software are the remaining development steps to a production instrument.

Frequency Domain Characterization Of Sliding Mode Control Of An Inverter Used In DSTATCOM Application

In this paper, the commonly used switching schemes for sliding mode control of power converters is analyzed and designed in the frequency domain. Particular application of a distribution static compensator (DSTATCOM) in voltage control mode is investigated in a power distribution system. Tsypkin's method and describing function is used to obtain the switching conditions for the two-level and three-level voltage source inverters. Magnitude conditions of carrier signals are developed for robust switching of the inverter under carrier-based modulation scheme of sliding mode control. The existence of border collision bifurcation is identified to avoid the complex switching states of the inverter. The load bus voltage of an unbalanced three-phase nonstiff radial distribution system is controlled using the proposed carrier-based design. The results are validated using PSCAD/EMTDC simulation studies and through a scaled laboratory model of DSTATCOM that is developed for experimental verification

Raising the Standards: An Integrated Approach to Promoting Professional Standards and Avoiding Professional Criminality

Crime in the Professions critically examines the nature and extent of crime and deviance in the professions and how it should be dealt with. The increasing professionalization of the work force and the changes in the way in which professionals carry out their work, such as through the use of computing and communications technology, have created new opportunities for professionals to break the law. Looking in detail at the nature and extent of crime committed by professionals such as doctors, accountants and nurses the book offers some innovative solutions on preventing and controlling professional crime. In addition to examining the nature and extent of crime in the professions, it also addresses some critical issues of regulation for the future, concerning new and emerging professional groups and issues relating to emerging technologies.

Development and validation of a novel approach to work sampling : a study of nurse practitioner work patterns

Objectives: This methodological paper reports on the development and validation of a work sampling instrument and data collection processes to conduct a national study of nurse practitioners’ work patterns. ---------- Design: Published work sampling instruments provided the basis for development and validation of a tool for use in a national study of nurse practitioner work activities across diverse contextual and clinical service models. Steps taken in the approach included design of a nurse practitioner-specific data collection tool and development of an innovative web-based program to train and establish inter rater reliability of a team of data collectors who were geographically dispersed across metropolitan, rural and remote health care settings. ---------- Setting: The study is part of a large funded study into nurse practitioner service. The Australian Nurse Practitioner Study is a national study phased over three years and was designed to provide essential information for Australian health service planners, regulators and consumer groups on the profile, process and outcome of nurse practitioner service. ---------- Results: The outcome if this phase of the study is empirically tested instruments, process and training materials for use in an international context by investigators interested in conducting a national study of nurse practitioner work practices. ---------- Conclusion: Development and preparation of a new approach to describing nurse practitioner practices using work sampling methods provides the groundwork for international collaboration in evaluation of nurse practitioner service.

A ‘cost-per-wear’ model based on contact lens replacement frequency

Purpose: The aim was to construct and advise on the use of a cost-per-wear model based on contact lens replacement frequency, to form an equitable basis for cost comparison. ---------- Methods: The annual cost of professional fees, contact lenses and solutions when wearing daily, two-weekly and monthly replacement contact lenses is determined in the context of the Australian market for spherical, toric and multifocal prescription types. This annual cost is divided by the number of times lenses are worn per year, resulting in a ‘cost-per-wear’. The model is presented graphically as the cost-per-wear versus the number of times lenses are worn each week for daily replacement and reusable (two-weekly and monthly replacement) lenses.---------- Results: The cost-per-wear for two-weekly and monthly replacement spherical lenses is almost identical but decreases with increasing frequency of wear. The cost-per-wear of daily replacement spherical lenses is lower than for reusable spherical lenses, when worn from one to four days per week but higher when worn six or seven days per week. The point at which the cost-per-wear is virtually the same for all three spherical lens replacement frequencies (approximately AUD$3.00) is five days of lens wear per week. A similar but upwardly displaced (higher cost) pattern is observed for toric lenses, with the cross-over point occurring between three and four days of wear per week (AUD$4.80). Multifocal lenses have the highest price, with cross-over points for daily versus two-weekly replacement lenses at between four and five days of wear per week (AUD$5.00) and for daily versus monthly replacement lenses at three days per week (AUD$5.50).---------- Conclusions: This cost-per-wear model can be used to assist practitioners and patients in making an informed decision in relation to the cost of contact lens wear as one of many considerations that must be taken into account when deciding on the most suitable lens replacement modality.

A collaborative approach to home care delivery for older clients: Perspectives of home care providers

A collaborative approach to home care (HC) delivery for older clients has taken centre stage (Nies, 2006). In Finland, public home help and home health care services have been combined to form the home care unit, whose goal is to provide a collaborative approach to care delivery through cooperation and sharing of responsibilities. In this model, the general practitioner (GP), home care nurses (HCN) and home help workers (HHW) care for shared clients. GPs and HCNs provide health care, such as monitoring of clients’ health status, and HHWs assist with personal care tasks such as dressing, washing and meal preparation. As the needs of older clients are multiple, collaboration is needed as one professional group cannot take sole responsibility (Nies, 2006). This paper reports on a study undertaken to examine home care unit care providers’ perspectives of the collaborative approach to HC delivery for older clients.

A new approach for finding an appropriate combination of texture parameters for classification

This paper suggests an approach for finding an appropriate combination of various parameters for extracting texture features (e.g. choice of spectral band for extracting texture feature, size of the moving window, quantization level of the image, and choice of texture feature etc.) to be used in the classification process. Gray level co-occurrence matrix (GLCM) method has been used for extracting texture from remotely sensed satellite image. Results of the classification of an Indian urban environment using spatial property (texture), derived from spectral and multi-resolution wavelet decomposed images have also been reported. A multivariate data analysis technique called ‘conjoint analysis’ has been used in the study to analyze the relative importance of these parameters. Results indicate that the choice of texture feature and window size have higher relative importance in the classification process than quantization level or the choice of image band for extracting texture feature. In case of texture features derived using wavelet decomposed image, the parameter ‘decomposition level’ has almost equal relative importance as the size of moving window and the decomposition of images up to level one is sufficient and there is no need to go for further decomposition. It was also observed that the classification incorporating texture features improves the overall classification accuracy in a statistically significant manner in comparison to pure spectral classification.

Statistical modeling to determine resonant frequency information present in combustion chamber pressure signals

A statistical modeling method to accurately determine combustion chamber resonance is proposed and demonstrated. This method utilises Markov-chain Monte Carlo (MCMC) through the use of the Metropolis-Hastings (MH) algorithm to yield a probability density function for the combustion chamber frequency and find the best estimate of the resonant frequency, along with uncertainty. The accurate determination of combustion chamber resonance is then used to investigate various engine phenomena, with appropriate uncertainty, for a range of engine cycles. It is shown that, when operating on various ethanol/diesel fuel combinations, a 20% substitution yields the least amount of inter-cycle variability, in relation to combustion chamber resonance.

Focus-score weighted super-resolution for uncooperative iris recognition at a distance and on the move

Uncooperative iris identification systems at a distance and on the move often suffer from poor resolution and poor focus of the captured iris images. The lack of pixel resolution and well-focused images significantly degrades the iris recognition performance. This paper proposes a new approach to incorporate the focus score into a reconstruction-based super-resolution process to generate a high resolution iris image from a low resolution and focus inconsistent video sequence of an eye. A reconstruction-based technique, which can incorporate middle and high frequency components from multiple low resolution frames into one desired super-resolved frame without introducing false high frequency components, is used. A new focus assessment approach is proposed for uncooperative iris at a distance and on the move to improve performance for variations in lighting, size and occlusion. A novel fusion scheme is then proposed to incorporate the proposed focus score into the super-resolution process. The experiments conducted on the The Multiple Biometric Grand Challenge portal database shows that our proposed approach achieves an EER of 2.1%, outperforming the existing state-of-the-art averaging signal-level fusion approach by 19.2% and the robust mean super-resolution approach by 8.7%.