Clinical significance of Mycobacterium asiaticum isolates in Queensland, Australia

Mycobacterium asiaticum was first reported as a cause of human disease in 1982, with only a few cases in the literature to date. This study aims to review the clinical significance of M. asiaticum isolates in Queensland, Australia. A retrospective review (1989 to 2008) of patients with M. asiaticum isolates was conducted. Data were collected through the Queensland TB Control Centre database. Disease was defined in accordance with the American Thoracic Society criteria. Twenty-four patients (13 female) had a positive culture of M. asiaticum, many residing around the Tropic of Capricorn. M. asiaticum was responsible for pulmonary disease (n = 2), childhood lymphadenitis (n = 1), olecranon bursitis (n = 1), 6 cases of possible pulmonary disease, and 2 possible wound infections. Chronic lung disease was a risk factor for pulmonary infection, and wounds/lacerations were a risk factor for extrapulmonary disease. Extrapulmonary disease responded to local measures. Pulmonary disease responded to ethambutol-isoniazid-rifampin plus pyrazinamide for the first 2 months in one patient, and amikacin-azithromycin-minocycline in another patient. While M. asiaticum is rare in Queensland, there appears to be an environmental niche. Although often a colonizer, it can be a cause of pulmonary and extrapulmonary disease. Treatment of pulmonary disease remains challenging. Extrapulmonary disease does not mandate specific nontuberculous mycobacterium (NTM) treatment.

Empirically validated interventions for adult disorders

The movement toward evidence-based practice in psychology and medicine should offer few problems in cognitive-behavior therapies because it is consistent with the principles by which they have been developed and disseminated. However, the criteria for assessing empirical status, including the heavy emphasis on manualized treatments, need close examination. A possible outcome of the evidence-based movement would be to focus on the application of manualized treatments in both training and clinical practice; problems with that approach are discussed. Commitment to evidence-based treatment should also include comparisons between psychological and pharmacological interventions, so that rational health care decisions can be made. Psychologists should not be afraid of following the evidence, even when it supports treatments that are not cognitive-behavioral in stated orientation. Such results should be taken as an opportunity for theoretical development and new empirical inquiry rather than be a cause for concern.

The manipulation of apoptosis-related genes to generate resistance to Fusarium wilt and water stress in banana

Bananas are susceptible to a diverse range of biotic and abiotic stresses, many of which cause serious production constraints worldwide. One of the most destructive banana diseases is Fusarium wilt caused by the soil-borne fungus, Fusarium oxysporum f. sp. cubense (Foc). No effective control strategy currently exists for this disease which threatens global banana production. Although disease resistance exists in some wild bananas, attempts to introduce resistance into commercially acceptable bananas by conventional breeding have been hampered by low fertility, long generation times and association of poor agronomical traits with resistance genes. With the advent of reliable banana transformation protocols, molecular breeding is now regarded as a viable alternative strategy to generate disease-resistant banana plants. Recently, a novel strategy involving the expression of anti-apoptosis genes in plants was shown to result in resistance against several necrotrophic fungi. Further, the transgenic plants showed increased resistance to a range of abiotic stresses. In this thesis, the use of anti-apoptosis genes to generate transgenic banana plants with resistance to Fusarium wilt was investigated. Since water stress is an important abiotic constraint to banana production, the resistance of the transgenic plants to water stress was also examined. Embryogenic cell suspensions (ECS) of two commercially important banana cultivars, Grand Naine (GN) and Lady Finger (LF), were transformed using Agrobacterium with the anti-apoptosis genes, Bcl-xL, Bcl-xL G138A, Ced-9 and Bcl- 2 3’ UTR. An interesting, and potentially important, outcome was that the use of anti-apoptosis genes resulted in up to a 50-fold increase in Agrobacterium-mediated transformation efficiency of both LF and GN cells over vector controls. Regenerated plants were subjected to a complete molecular characterisation in order to detect the presence of the transgene (PCR), transcript (RT-PCR) and gene product (Western blot) and to determine the gene copy number (Southern blot). A total of 36 independently-transformed GN lines (8 x Bcl-xL, 5 x Bcl-xL G138A, 15 x Ced-9 and 8 x Bcl-2 3’ UTR) and 41 independently-transformed LF lines (8 x Bcl-xL, 7 x BclxL G138A, 13 x Ced-9 and 13 x Bcl-2 3’ UTR) were identified. The 41 transgenic LF lines were multiplied and clones from each line were acclimatised and grown under glasshouse conditions for 8 weeks to allow monitoring for phenotypic abnormalities. Plants derived from 3 x Bcl-xL, 2 x Ced-9 and 5 x Bcl-2 3’ UTR lines displayed a variety of aberrant phenotypes. However, all but one of these abnormalities were off-types commonly observed in tissue-cultured, non-transgenic banana plants and were therefore unlikely to be transgene-related. Prior to determining the resistance of the transgenic plants to Foc race 1, the apoptotic effects of the fungus on both wild-type and Bcl-2 3’ UTR-transgenic LF banana cells were investigated using rapid in vitro root assays. The results from these assays showed that apoptotic-like cell death was elicited in wild-type banana root cells as early as 6 hours post-exposure to fungal spores. In contrast, these effects were attenuated in the root cells of Bcl-2 3’ UTR-transgenic lines that were exposed to fungal spores. Thirty eight of the 41 transgenic LF lines were subsequently assessed for resistance to Foc race 1 in small-plant glasshouse bioassays. To overcome inconsistencies in rating the internal (vascular discolouration) disease symptoms, a MatLab-based computer program was developed to accurately and reliably assess the level of vascular discolouration in banana corms. Of the transgenic LF banana lines challenged with Foc race 1, 2 x Bcl-xL, 3 x Ced-9, 2 x Bcl-2 3’ UTR and 1 x Bcl-xL G138A-transgenic line were found to show significantly less external and internal symptoms than wild-type LF banana plants used as susceptible controls at 12 weeks post-inoculation. Of these lines, Bcl-2 3’ UTR-transgenic line #6 appeared most resistant, displaying very mild symptoms similar to the wild-type Cavendish banana plants that were included as resistant controls. This line remained resistant for up to 23 weeks post-inoculation. Since anti-apoptosis genes have been shown to confer resistance to various abiotic stresses in other crops, the ability of these genes to confer resistance against water stress in banana was also investigated. Clonal plants derived from each of the 38 transgenic LF banana plants were subjected to water stress for a total of 32 days. Several different lines of transgenic plants transformed with either Bcl-xL, Bcl-xL G138A, Ced-9 or Bcl-2 3’ UTR showed a delay in visual water stress symptoms compared with the wild-type control plants. These plants all began producing new growth from the pseudostem following daily rewatering for one month. In an attempt to determine whether the protective effect of anti-apoptosis genes in transgenic banana plants was linked with reactive oxygen species (ROS)-associated programmed cell death (PCD), the effect of the chloroplast-targeting, ROS-inducing herbicide, Paraquat, on wild-type and transgenic LF was investigated. When leaf discs from wild-type LF banana plants were exposed to 10 ìM Paraquat, complete decolourisation occurred after 48 hours which was confirmed to be associated with cell death and ROS production by trypan blue and 3,3-diaminobenzidine (DAB) staining, respectively. When leaf discs from the transgenic lines were exposed to Paraquat, those derived from some lines showed a delay in decolourisation, suggesting only a weak protective effect from the transgenes. Finally, the protective effect of anti-apoptosis genes against juglone, a ROS-inducing phytotoxin produced by the causal agent of black Sigatoka, Mycosphaerella fijiensis, was investigated. When leaf discs from wild-type LF banana plants were exposed to 25 ppm juglone, complete decolourisation occurred after 48 hours which was again confirmed to be associated with cell death and ROS production by trypan blue and DAB staining, respectively. Further, TdT-mediated dUTP nick-end labelling (TUNEL) assays on these discs suggested that the cell death was apoptotic. When leaf discs from the transgenic lines were exposed to juglone, discs from some lines showed a clear delay in decolourisation, suggesting a protective effect. Whether these plants are resistant to black Sigatoka is unknown and will require future glasshouse and field trials. The work presented in this thesis provides the first report of the use of anti-apoptosis genes as a strategy to confer resistance to Fusarium wilt and water stress in a nongraminaceous monocot, banana. Such a strategy may be exploited to generate resistance to necrotrophic pathogens and abiotic stresses in other economically important crop plants.

Behaviour and design of cold-formed steel compression members at elevated temperatures

Cold-formed steel members have been widely used in residential, industrial and commercial buildings as primary load bearing structural elements and non-load bearing structural elements (partitions) due to their advantages such as higher strength to weight ratio over the other structural materials such as hot-rolled steel, timber and concrete. Cold-formed steel members are often made from thin steel sheets and hence they are more susceptible to various buckling modes. Generally short columns are susceptible to local or distortional buckling while long columns to flexural or flexural-torsional buckling. Fire safety design of building structures is an essential requirement as fire events can cause loss of property and lives. Therefore it is essential to understand the fire performance of light gauge cold-formed steel structures under fire conditions. The buckling behaviour of cold-formed steel compression members under fire conditions is not well investigated yet and hence there is a lack of knowledge on the fire performance of cold-formed steel compression members. Current cold-formed steel design standards do not provide adequate design guidelines for the fire design of cold-formed steel compression members. Therefore a research project based on extensive experimental and numerical studies was undertaken at the Queensland University of Technology to investigate the buckling behaviour of light gauge cold-formed steel compression members under simulated fire conditions. As the first phase of this research, a detailed review was undertaken on the mechanical properties of light gauge cold-formed steels at elevated temperatures and the most reliable predictive models for mechanical properties and stress-strain models based on detailed experimental investigations were identified. Their accuracy was verified experimentally by carrying out a series of tensile coupon tests at ambient and elevated temperatures. As the second phase of this research, local buckling behaviour was investigated based on the experimental and numerical investigations at ambient and elevated temperatures. First a series of 91 local buckling tests was carried out at ambient and elevated temperatures on lipped and unlipped channels made of G250-0.95, G550-0.95, G250-1.95 and G450-1.90 cold-formed steels. Suitable finite element models were then developed to simulate the experimental conditions. These models were converted to ideal finite element models to undertake detailed parametric study. Finally all the ultimate load capacity results for local buckling were compared with the available design methods based on AS/NZS 4600, BS 5950 Part 5, Eurocode 3 Part 1.2 and the direct strength method (DSM), and suitable recommendations were made for the fire design of cold-formed steel compression members subject to local buckling. As the third phase of this research, flexural-torsional buckling behaviour was investigated experimentally and numerically. Two series of 39 flexural-torsional buckling tests were undertaken at ambient and elevated temperatures. The first series consisted 2800 mm long columns of G550-0.95, G250-1.95 and G450-1.90 cold-formed steel lipped channel columns while the second series contained 1800 mm long lipped channel columns of the same steel thickness and strength grades. All the experimental tests were simulated using a suitable finite element model, and the same model was used in a detailed parametric study following validation. Based on the comparison of results from the experimental and parametric studies with the available design methods, suitable design recommendations were made. This thesis presents a detailed description of the experimental and numerical studies undertaken on the mechanical properties and the local and flexural-torsional bucking behaviour of cold-formed steel compression member at ambient and elevated temperatures. It also describes the currently available ambient temperature design methods and their accuracy when used for fire design with appropriately reduced mechanical properties at elevated temperatures. Available fire design methods are also included and their accuracy in predicting the ultimate load capacity at elevated temperatures was investigated. This research has shown that the current ambient temperature design methods are capable of predicting the local and flexural-torsional buckling capacities of cold-formed steel compression members at elevated temperatures with the use of reduced mechanical properties. However, the elevated temperature design method in Eurocode 3 Part 1.2 is overly conservative and hence unsuitable, particularly in the case of flexural-torsional buckling at elevated temperatures.