The resistance of intra-amoebal grown legionella pneumophila

Survival studies were conducted on Legionella pneumophila cells that had been grown intracellulary in Acanthamoeba polyphaga and then exposed to polyhexamethylene biguanide (PHMB), benzisothiazolone (BIT), 5-chloro-N-methylisothiazolone (CMIT) and tetradecyltrimethyl ammonium bromide (TTAB). Susceptibilities were also determined for L.pneumophila grown under nutrient sufficient and iron-, nitrogen- and phosphate-depleted conditions, in a chemically defined medium. BIT was relatively ineffective against cells grown under iron-depletion; in contrast iron-depleted conditions increased the susceptibilities of cells to PHMB, TTAB and CMIT. Cells grown under phosphate-depletion showed a marked increase in sensitivity towards all the biocides. Conversely, the activities of all four biocides were greatly reduced against L.pneumophila grown in amoebae. To study the physiological basis for the increased resistance of intra-amoebal grown legionella, the surface properties of the cells were examined by studying outer membrane proteins (OMs), lipopolysaccharides and cellular fatty acids. Intra-amoebal grown legionella were found to differ in several respects compared to cells grown in vitro; they contained a novel 15-kDal OM protein and a monosaturated straight-chain fatty acid (18:19). These compounds were also found in abundant quantities in the host amoeba. Intra-amoebal grown legionella contained more LPS bands than did in vitro grown organisms and were less susceptible to protease K digestion. Cells grown under phosphate depletion were markedly sensitive to protease K digestion and contained lower levels of LPS. Immunoblot analysis of intra-amoebal grown legionella with anti-acanthamoebal serum revealed that both the surface of the bacteria and sarkosyl extracted OMs contained amoebal proteins. These findings suggest that the 15-kDal OM protein is likely to be of amoebal origin and binds tightly to the OM of the bacterium. It is proposed that disruption of amoebal membranes, as a result of intra-amoebal infection liberates macromolecules, including a 15-kDal polypeptide, a major constituent of the membrane, which associates closely with the surface of the legionellae. Thus L.pneumophila which have extraneous membrane material bound to their surface may respond differently to biocide inactivation, as these macromolecules may act as a penetration barrier to such agents. This phenomenon could contribute to the recalcitrance of legionellae in water systems.

Chloride resistance and durability of cement paste and concrete containing metakaolin

Metakaolin (MK), a calcined clay, was included as a partial cement replacement material, at up to 20% by weight of binder, in cement pastes and concrete, and its influence on the resistance to chloride ingress investigated. Reductions in effective chloride diffusion coefficients through hardened cement paste were obtained for binary blends and by combining OPC, MK and a second cement replacement material of pulverised fuel ash or ground granulated blast furnace slag. Steady state oxygen diffusion measurements through hardened cement pastes measured using an electrochemical cell showed that the interaction between charged species and the pore surfaces is a major factor in determining chloride diffusion rate. Rheology of the binder, particularly at high MK replacement levels, was found to have a dramatic influence on the diffusion performance of cement pastes. It was concluded that plasticising admixtures are essential for adequate dispersion of MK in cement pastes. Chloride concentration profile analysis of the concrete cylinders, exposed to sodium chloride solution for one year, was employed to obtain apparent chloride diffusion coefficients for concrete specimens. MK was found to reduce the depth of chloride penetration into concrete when compared with that of unblended mixes. Corrosion rate and corrosion potential measurements were taken on steel bars embedded in concrete exposed to a saline environment under conditions of cyclic wetting and drying. The initiation time for corrosion was found to be significantly longer for MK blended mixes than for plain OPC systems. The aggregate-paste interfacial zone of MK blended systems was investigated by steady state diffusion of chloride ions through mortar containing glass beads as model aggregate. For the model aggregate specimens tested the work confirmed the hypothesis that properties of the bulk paste are the controlling factors in ionic diffusion through mortar.

Smoothing a Discrete Hazard Function for the Number of Patients Colonized with Methicillin-Resistance Staphylococcus Aureus in an Intensive Care Unit

A new method for estimating the time to colonization of Methicillin-resistant Staphylococcus Aureus (MRSA) patients is developed in this paper. The time to colonization of MRSA is modelled using a Bayesian smoothing approach for the hazard function. There are two prior models discussed in this paper: the first difference prior and the second difference prior. The second difference prior model gives smoother estimates of the hazard functions and, when applied to data from an intensive care unit (ICU), clearly shows increasing hazard up to day 13, then a decreasing hazard. The results clearly demonstrate that the hazard is not constant and provide a useful quantification of the effect of length of stay on the risk of MRSA colonization which provides useful insight.

Exploring the resistance : an Australian perspective on educating for sustainability in early childhood

Climate change and sustainability are issues of global significance. While other education sectors have implemented education for sustainability for many years, the early childhood sector has been slow to take up this challenge. This position paper poses the question: Why has this sector been so slow to engage with sustainability? Explanations are proposed based on a review of research literature and the authors’ long engagement in seeking to bring early childhood education and education for sustainability together. The imperative is for the early childhood sector to engage in education for sustainability without delay and to ‘get active’ for a sustainable future.

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.

Resistance training for obese, type 2 diabetic adults: A review of the evidence

In both developed and developing countries, increased prevalence of obesity has been strongly associated with increased incidence of type 2 diabetes mellitus (T2DM) in the adult population. Previous research has emphasized the importance of physical activity in the prevention and management of obesity and T2DM, and generic exercise guidelines originally developed for the wider population have been adapted for these specific populations. However, the guidelines traditionally focus on aerobic training without due consideration to other exercise modalities. Recent reviews on resistance training in the T2DM population have not compared this modality with others including aerobic training, or considered the implications of resistance training for individuals suffering from both obesity and T2DM. In short, the optimal mix of exercise modalities in the prescription of exercise has not been identified for it benefits to the metabolic, body composition and muscular health markers common in obesity and T2DM. Similarly, the underlying physical, social and psychological barriers to adopting and maintaining exercise, with the potential to undermine the efficacy of exercise interventions, have not been addressed in earlier reviews. Because it is well established that aerobic exercise has profound effects on obesity and T2DM risk, the purpose of this review was to address the importance of resistance training to obese adults with T2DM.

Glycaemic load is associated with insulin resistance in older Australian women

Background: Diets with a high postprandial glycemic response may contribute to long-term development of insulin resistance and diabetes, however previous epidemiological studies are conflicting on whether glycemic index (GI) or glycemic load (GL) are dietary factors associated with the progression. Our objectives were to estimate GI and GL in a group of older women, and evaluate cross-sectional associations with insulin resistance. Subjects and Methods: Subjects were 329 Australian women aged 42-81 years participating in year three of the Longitudinal Assessment of Ageing in Women (LAW). Dietary intakes were assessed by diet history interviews and analysed using a customised GI database. Insulin resistance was defined as a homeostasis model assessment (HOMA) value of >3.99, based on fasting blood glucose and insulin concentrations. Results: GL was significantly higher in the 26 subjects who were classified as insulin resistant compared to subjects who were not (134±33 versus 114±24, P<0.001). In a logistic regression model, an increment of 15 GL units increased the odds of insulin resistance by 2.09 (95%CI 1.55, 2.80, P<0.001) independently of potential confounding variables. No significant associations were found when insulin resistance was assessed as a continuous variable. Conclusions: Results of this cross-sectional study support the concept that diets with a higher GL are associated with increased risk of insulin resistance. Further studies are required to investigate whether reducing glycemic intake, by either consuming lower GI foods and/or smaller serves of carbohydrate, can contribute to a reduction in development of insulin resistance and long-term risk of type 2 diabetes.