The development of Lactococcus lactis as an antimicrobial agent

Non-pathogenic lactic acid bacteria are economically important Gram-positive bacteria used extensively in the food industry. Due to their “generally regarded as safe” status, certain species from the genera Lactobacillus and Lactococcus are also considered desirable as candidates for the production and secretion of recombinant proteins, particular those with therapeutic applications. The hypothesis examined by this thesis is that Lactococcus lactis can be modified to be an effective antimicrobial agent. Therefore, the aims of this thesis were to investigate the optimisation of the expression, secretion and/or activities of potential heterologous antimicrobial proteins by the model lactic acid bacterium, Lactococcus lactis subsp. cremoris MG1363. L. lactis strains were engineered to express and secrete the recombinant CyuC surface protein from Lactobacillus reuteri BR11, and a fusion protein consisting of CyuC and lysostaphin using the Sep promoter and secretion signal. CyuC has been characterised as a cystine-binding protein, but has also been demonstrated to have fibronectin binding activity. Lysostaphin is a bacteriolytic enzyme with specific activity against the Gram-positive pathogen, Staphylococcus aureus. These modified L. lactis strains were then investigated to see if they had the ability to inhibit the adhesion of S. aureus to host extracellular matrix (ECM) proteins. It was observed that the cell extracts of the L. lactis strain with the vector only (pGhost9:ISS1) was able to inhibit the adhesion of S. aureus to fibronectin, whilst the cell extracts of the L. lactis strain expressing lysostaphin was able to inhibit adhesion to keratin. Finally, this thesis has identified specific lactococcal genes that affect the secretion of lysostaphin through the use of random transposon mutagenesis. Ten mutants with higher lysostaphin activity contained insertions in four different genes encoding: (i) an uncharacterised putative transmembrane protein (llmg_0609), (ii) an enzyme catalysing the first step in peptidoglycan biosynthesis (murA2), (iii) a homolog of the oxidative defence regulator (trmA), and (iv) an uncharacterised putative enzyme involved in ubiquinone biosynthesis (llmg_2148). The higher lysostaphin activity observed in these mutants was found to be due to higher amounts of lysostaphin being secreted. The findings of this thesis contribute to the development of this organism as an antimicrobial agent and also to our understanding of L. lactis genetics.

In vivo performance of melimine as an antimicrobial coating for contact lenses in models of CLARE and CLPU

Purpose: One strategy to minimize bacteria-associated adverse responses such as microbial keratitis, contact lens–induced acute red eye (CLARE), and contact lens induced peripheral ulcers (CLPUs) that occur with contact lens wear is the development of an antimicrobial or antiadhesive contact lens. Cationic peptides represent a novel approach for the development of antimicrobial lenses.---------- Methods: A novel cationic peptide, melimine, was covalently incorporated into silicone hydrogel lenses. Confirmation tests to determine the presence of peptide and anti-microbial activity were performed. Cationic lenses were then tested for their ability to prevent CLPU in the Staphylococcus aureus rabbit model and CLARE in the Pseudomonas aeruginosa guinea pig model. ---------- Results: In the rabbit model of CLPU, melimine-coated lenses resulted in significant reductions in ocular symptom scores and in the extent of corneal infiltration (P < 0.05). Evaluation of the performance of melimine lenses in the CLARE model showed significant improvement in all ocular response parameters measured, including the percentage of eyes with corneal infiltrates, compared with those observed in the eyes fitted with the control lens (P ≤ 0.05). ---------- Conclusions: Cationic coating of contact lenses with the peptide melimine may represent a novel method of prevention of bacterial growth on contact lenses and consequently result in reduction of the incidence and severity of adverse responses due to Gram-positive and -negative bacteria during lens wear.

Sensitivity of Dot-ELISA on Nitrocellulose Membranes in Comparison with ELISA on Polystyrene Plates for the Detection of Four Plant Viruses

ABSTR.4CT Senitivity of dot-immunobindinding ELf SA on nitrocellulose membrane (DotELISA)was compared with double-antibody sandwich ELISA (DAS-ELlSA) on polystyrene plates for the detection of bean yellow mosaic virus (BYMV), broad bean stain virus (WMV-2). Dot-ELISA was 2 and 1O times more sensitive than DAS-ELISA for the detection of BBSV and WMV-2, respectively, whereas DAS-ELISA was more sensitive than Dot-ELiSA for {he detection of BYMV. Both techniques were equally sensitive for the detection of BYDV. Using one day instead uf the two-day procedure, the four viruses were still detectable and the ralative sensitivity of both techniques remained the same.

Survey of Viruses Affecting Faba Bean in Six Arab Countries

A field survey of faba bean (Vicia [aba L.) for viruses in six Arab countries showed the presence of nine viruses. Bean leaf roll virus (BLRV), bean yellow mosaic virus (BYMV), broad bean mottle virus (BBMV) and to a lesser extent broad bean stain virus (BBSV) were the most common. When testing with ELISA 789 samples with symptoms suggestive of virus infection collected from Egypt, Lebanon, Morocco, Sudan, Syria and Tunisia, BBMV was detected in 203 samples, BBSV in 151, broad bean true mosaic virus (BBTMV) in 7, broad bean wilt virus (BBWV) in 47, BYMV in 314, cucumber mosaic virus (CMV) in 96, pea enation mosaic virus (PEMV) in 31, and pea seed-borne mosaic virus (PSbMV) in 49 samples. Identity of selected field isolates was confirmed by electron microscopy and host reaction studies. In a yield experiment, infection with BYMV, BBMV and BBSV 11 weeks after sowing (pre-flowering) led to 81, 54 and 84% yield loss, respectively. Inoculation with the same viruses 15 weeks after sowing (flowering) and 20 weeks after sowing (pod setting) led to 56, 84 and 18%, and 39, 37 and 18% yield loss, respectively.

Immunohistochemical analysis of structural changes in collagen for the assessment of osteoarthritis

Collagen fibrillation within articular cartilage (AC) plays a key role in joint osteoarthritis (OA) progression and, therefore, studying collagen synthesis changes could be an indicator for use in the assessment of OA. Various staining techniques have been developed and used to determine the collagen network transformation under microscopy. However, because collagen and proteoglycan coexist and have the same index of refraction, conventional methods for specific visualization of collagen tissue is difficult. This study aimed to develop an advanced staining technique to distinguish collagen from proteoglycan and to determine its evolution in relation to OA progression using optical and laser scanning confocal microscopy (LSCM). A number of AC samples were obtained from sheep joints, including both healthy and abnormal joints with OA grades 1 to 3. The samples were stained using two different trichrome methods and immunohistochemistry (IHC) to stain both colourimetrically and with fluorescence. Using optical microscopy and LSCM, the present authors demonstrated that the IHC technique stains collagens only, allowing the collagen network to be separated and directly investigated. Fluorescently-stained IHC samples were also subjected to LSCM to obtain three-dimensional images of the collagen fibres. Changes in the collagen fibres were then correlated with the grade of OA in tissue. This study is the first to successfully utilize the IHC staining technique in conjunction with laser scanning confocal microscopy. This is a valuable tool for assessing changes to articular cartilage in OA.

Predictive factors of contact lens case contamination

PURPOSE: To examine the relationship between contact lens (CL) case contamination and various potential predictive factors. METHODS: 74 subjects were fitted with lotrafilcon B (CIBA Vision) CLs on a daily wear basis for 1 month. Subjects were randomly assigned one of two polyhexamethylene biguanide (PHMB) preserved disinfecting solutions with the corresponding regular lens case. Clinical evaluations were conducted at lens delivery and after 1 month, when cases were collected for microbial culture. A CL care non-compliance score was determined through administration of a questionnaire and the volume of solution used was calculated for each subject. Data was examined using backward stepwise binary logistic regression. RESULTS: 68% of cases were contaminated. 35% were moderately or heavily contaminated and 36% contained gram-negative bacteria. Case contamination was significantly associated with subjective dryness symptoms (OR 4.22, CI 1.37–13.01) (P<0.05). There was no association between contamination and subject age, ethnicity, gender, average wearing time, amount of solution used, non-compliance score, CL power and subjective redness (P>0.05). The effect of lens care system on case contamination approached significance (P=0.07). Failure to rinse the case with disinfecting solution following CL insertion (OR 2.51, CI 0.52–12.09) and not air drying the case (OR 2.31, CI 0.39–13.35) were positively correlated with contamination; however, did not reach statistical significance. CONCLUSIONS: Our results suggest that case contamination may influence subjective comfort. It is difficult to predict the development of case contamination from a variety of clinical factors. The efficacy of CL solutions, bacterial resistance to disinfection and biofilm formation are likely to play a role. Further evaluation of these factors will improve our understanding of the development of case contamination and its clinical impact.

Dynamic compression improves biosynthesis of human zonal chondrocytes from osteoarthritis patients

Objective: We hypothesize that chondrocytes from distinct zones of articular cartilage respond differently to compressive loading, and that zonal chondrocytes from osteoarthritis (OA) patients can benefit from optimized compressive stimulation. Therefore, we aimed to determine the transcriptional response of superficial (S) and middle/deep (MD) zone chondrocytes to varying dynamic compressive strain and loading duration. To confirm effects of compressive stimulation on overall matrix production, we subjected zonal chondrocytes to compression for 2 weeks. Design: Human S and MD chondrocytes from osteoarthritic joints were encapsulated in 2% alginate, pre-cultured, and subjected to compression with varying dynamic strain (5, 15, 50% at 1 Hz) and loading duration (1, 3, 12 h). Temporal changes in cartilage-specific, zonal, and dedifferentiation genes following compression were evaluated using quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR). The benefits of long-term compression (50% strain, 3 h/day, for 2 weeks) were assessed by measuring construct glycosaminoglycan (GAG) content and compressive moduli, as well as immunostaining. Results: Compressive stimulation significantly induced aggrecan (ACAN), COL2A1, COL1A1, proteoglycan 4 (PRG4), and COL10A1 gene expression after 2 h of unloading, in a zone-dependent manner (P < 0.05). ACAN and PRG4 mRNA levels depended on strain and load duration, with 50% and 3 h loading resulting in highest levels (P < 0.05). Long-term compression increased collagen type II and ACAN immunostaining and total GAG (P < 0.05), but only S constructs showed more PRG4 stain, retained more GAG (P < 0.01), and developed higher compressive moduli than non-loaded controls. Conclusions: The biosynthetic activity of zonal chondrocytes from osteoarthritis joints can be enhanced with selected compression regimes, indicating the potential for cartilage tissue engineering applications. © 2012 Osteoarthritis Research Society International.

Molecular characterisation of environmental enterococci derived from water samples and assessment of associated public health hazards

Enterococci are versatile Gram-positive bacteria that can survive under extreme conditions. Most enterococci are non-virulent and found in the gastrointestinal tract of humans and animals. Other strains are opportunistic pathogens that contribute to a large number of nosocomial infections globally. Epidemiological studies demonstrated a direct relationship between the density of enterococci in surface waters and the risk of swimmer-associated gastroenteritis. The distribution of infectious enterococcal strains from the hospital environment or other sources to environmental water bodies through sewage discharge or other means, could increase the prevalence of these strains in the human population. Environmental water quality studies may benefit from focusing on a subset of Enterococcus spp. that are consistently associated with sources of faecal pollution such as domestic sewage, rather than testing for the entire genus. E. faecalis and E. faecium are potentially good focal species for such studies, as they have been consistently identified as the dominant Enterococcus spp. in human faeces and sewage. On the other hand enterococcal infections are predominantly caused by E. faecalis and E. faecium. The characterisation of E. faecalis and E. faecium is important in studying their population structures, particularly in environmental samples. In developing and implementing rapid, robust molecular genotyping techniques, it is possible to more accurately establish the relationship between human and environmental enterococci. Of particular importance, is to determine the distribution of high risk enterococcal clonal complexes, such as E. faecium clonal complex 17 and E. faecalis clonal complexes 2 and 9 in recreational waters. These clonal complexes are recognized as particularly pathogenic enterococcal genotypes that cause severe disease in humans globally. The Pimpama-Coomera watershed is located in South East Queensland, Australia and was investigated in this study mainly because it is used intensively for agriculture and recreational purposes and has a strong anthropogenic impact. The primary aim of this study was to develop novel, universally applicable, robust, rapid and cost effective genotyping methods which are likely to yield more definitive results for the routine monitoring of E. faecalis and E. faecium, particularly in environmental water sources. To fullfill this aim, new genotyping methods were developed based on the interrogation of highly informative single nucleotide polymorphisms (SNPs) located in housekeeping genes of both E. faecalis and E. faecium. SNP genotyping was successfully applied in field investigations of the Coomera watershed, South-East Queensland, Australia. E. faecalis and E. faecium isolates were grouped into 29 and 23 SNP profiles respectively. This study showed the high longitudinal diversity of E. faecalis and E. faecium over a period of two years, and both human-related and human-specific SNP profiles were identified. Furthermore, 4.25% of E. faecium strains isolated from water was found to correspond to the important clonal complex-17 (CC17). Strains that belong to CC17 cause the majority of hospital outbreaks and clinical infections globally. Of the six sampling sites of the Coomera River, Paradise Point had the highest number of human-related and human-specific E. faecalis and E. faecium SNP profiles. The secondary aim of this study was to determine the antibiotic-resistance profiles and virulence traits associated with environmental E. faecalis and E. faecium isolates compared to human pathogenic E. faecalis and E. faecium isolates. This was performed to predict the potential health risks associated with coming into contact with these strains in the Coomera watershed. In general, clinical isolates were found to be more resistant to all the antibiotics tested compared to water isolates and they harbored more virulence traits. Multi-drug resistance was more prevalent in clinical isolates (71.18% of E. faecalis and 70.3 % of E. faecium) compared to water isolates (only 5.66 % E. faecium). However, tetracycline, gentamicin, ciprofloxacin and ampicillin resistance was observed in water isolates. The virulence gene esp was the most prevalent virulence determinant observed in clinical isolates (67.79% of E. faecalis and 70.37 % of E. faecium), and this gene has been described as a human-specific marker used for microbial source tracking (MST). The presence of esp in water isolates (16.36% of E. faecalis and 19.14% of E. faecium) could be indicative of human faecal contamination in these waterways. Finally, in order to compare overall gene expression between environmental and clinical strains of E. faecalis, a comparative gene hybridization study was performed. The results of this investigation clearly demonstrated the up-regulation of genes associated with pathogenicity in E. faecalis isolated from water. The expression study was performed at physiological temperatures relative to ambient temperatures. The up-regulation of virulence genes demonstrates that environmental strains of E. faecalis can pose an increased health risk which can lead to serious disease, particularly if these strains belong to the virulent CC17 group. The genotyping techniques developed in this study not only provide a rapid, robust and highly discriminatory tool to characterize E. faecalis and E. faecium, but also enables the efficient identification of virulent enterococci that are distributed in environmental water sources.

Risk control in transmission system expansion planning with wind generators

With the advent of large-scale wind farms and their integration into electrical grids, more uncertainties, constraints and objectives must be considered in power system development. It is therefore necessary to introduce risk-control strategies into the planning of transmission systems connected with wind power generators. This paper presents a probability-based multi-objective model equipped with three risk-control strategies. The model is developed to evaluate and enhance the ability of the transmission system to protect against overload risks when wind power is integrated into the power system. The model involves: (i) defining the uncertainties associated with wind power generators with probability measures and calculating the probabilistic power flow with the combined use of cumulants and Gram-Charlier series; (ii) developing three risk-control strategies by specifying the smallest acceptable non-overload probability for each branch and the whole system, and specifying the non-overload margin for all branches in the whole system; (iii) formulating an overload risk index based on the non-overload probability and the non-overload margin defined; and (iv) developing a multi-objective transmission system expansion planning (TSEP) model with the objective functions composed of transmission investment and the overload risk index. The presented work represents a superior risk-control model for TSEP in terms of security, reliability and economy. The transmission expansion planning model with the three risk-control strategies demonstrates its feasibility in the case study using two typical power systems

Link discovery for Chinese/English cross-language web information retrieval

Nowadays people heavily rely on the Internet for information and knowledge. Wikipedia is an online multilingual encyclopaedia that contains a very large number of detailed articles covering most written languages. It is often considered to be a treasury of human knowledge. It includes extensive hypertext links between documents of the same language for easy navigation. However, the pages in different languages are rarely cross-linked except for direct equivalent pages on the same subject in different languages. This could pose serious difficulties to users seeking information or knowledge from different lingual sources, or where there is no equivalent page in one language or another. In this thesis, a new information retrieval task—cross-lingual link discovery (CLLD) is proposed to tackle the problem of the lack of cross-lingual anchored links in a knowledge base such as Wikipedia. In contrast to traditional information retrieval tasks, cross language link discovery algorithms actively recommend a set of meaningful anchors in a source document and establish links to documents in an alternative language. In other words, cross-lingual link discovery is a way of automatically finding hypertext links between documents in different languages, which is particularly helpful for knowledge discovery in different language domains. This study is specifically focused on Chinese / English link discovery (C/ELD). Chinese / English link discovery is a special case of cross-lingual link discovery task. It involves tasks including natural language processing (NLP), cross-lingual information retrieval (CLIR) and cross-lingual link discovery. To justify the effectiveness of CLLD, a standard evaluation framework is also proposed. The evaluation framework includes topics, document collections, a gold standard dataset, evaluation metrics, and toolkits for run pooling, link assessment and system evaluation. With the evaluation framework, performance of CLLD approaches and systems can be quantified. This thesis contributes to the research on natural language processing and cross-lingual information retrieval in CLLD: 1) a new simple, but effective Chinese segmentation method, n-gram mutual information, is presented for determining the boundaries of Chinese text; 2) a voting mechanism of name entity translation is demonstrated for achieving a high precision of English / Chinese machine translation; 3) a link mining approach that mines the existing link structure for anchor probabilities achieves encouraging results in suggesting cross-lingual Chinese / English links in Wikipedia. This approach was examined in the experiments for better, automatic generation of cross-lingual links that were carried out as part of the study. The overall major contribution of this thesis is the provision of a standard evaluation framework for cross-lingual link discovery research. It is important in CLLD evaluation to have this framework which helps in benchmarking the performance of various CLLD systems and in identifying good CLLD realisation approaches. The evaluation methods and the evaluation framework described in this thesis have been utilised to quantify the system performance in the NTCIR-9 Crosslink task which is the first information retrieval track of this kind.

Ureaplasma parvum serovar 3 multiple banded antigen size variation after chronic intra-amniotic infection/colonization

Ureaplasma species are the microorganisms most frequently associated with adverse pregnancy outcomes. The multiple banded antigen (MBA), a surface-exposed lipoprotein, is a key virulence factor of ureaplasmas. The MBA demonstrates size variation, which we have shown previously to be correlated with the severity of chorioamnion inflammation. We aimed to investigate U. parvum serovar 3 pathogenesis in vivo, using a sheep model, by investigating: MBA variation after long term (chronic) and short term (acute) durations of in utero ureaplasma infections, and the severity of chorioamnionitis and inflammation in other fetal tissues. Inocula of 2x107 colony-forming-units (CFU) of U. parvum serovar 3 (Up) or media controls (C) were injected intra-amniotically into pregnant ewes at one of three time points: day 55 (69d Up, n=8; C69, n=4); day 117 (7d Up, n=8; C7, n=2); and day 121 (3d Up, n=8; C3, n=2) of gestation (term=145-150d). At day 124, preterm fetuses were delivered surgically. Samples of chorioamnion, fetal lung, and umbilical cord were: (i) snap frozen for subsequent ureaplasma culture, and (ii) fixed, embedded, sectioned and stained by haematoxylin and eosin stain for histological analysis. Selected fetal lung clinical ureaplasma isolates were cloned and filtered to obtain cultures from a single CFU. Passage 1 and clone 2 ureaplasma cultures were tested by western blot to demonstrate MBA variation. In acute durations of ureaplasma infection no MBA variants (3d Up) or very few MBA variants (7d Up) were present when compared to the original inoculum. However, numerous MBA size variants were generated in vivo (alike within contiguous tissues, amniotic fluid and fetal lung, but different variants were present within chorioamnion), during chronic, 69d exposure to ureaplasma infection. For the first time we have shown that the degree of ureaplasma MBA variation in vivo increased with the duration of gestation.

Metabolic and ecological study of environmental pentose utilizing bacteria (E-PUB)

Lignocellulosic materials, such as sugar cane bagasse, a waste product of the sugarcane processing industry, agricultural residues and herbaceous crops, may serve as an abundant and comparatively cheap feedstock for largescale industrial fermentation, resulting in the production of marketable end-products. However, the complex structure of lignocellulosic materials, the presence of various hexose and pentose sugars in the hemicellulose component, and the presence of various compounds that inhibit the organisms selected for the fermentation process, all constitute barriers that add to the production costs and make full scale industrial production economically less feasible. The work presented in this thesis was conducted in order to screen microorganisms for ability to utilize pentose sugars derived from the sugar mill industrial waste. A large number of individual bacterial strains were investigated from hemi-cellulose rich material collected at the Proserpine and Maryborough sugar mills, notably soil samples from the mill sites. The research conducted to isolation of six pentose-capable Gram-positive organisms from the actinomycetes group by using pentose as a sole carbon source in the cultivation process. The isolates were identified as Corynebacterium glutamicum, Actinomyces odontolyticus, Nocardia elegans, and Propionibacterium freudenreichii all of which were isolated from the hemicellulose-enriched soil. Pentose degrading microbes are very rare in the environment, so this was a significant discovery. Previous research indicated that microbes could degrade pentose after genetic modification but the microbes discovered in this research were able to naturally utilize pentose. Six isolates, identified as four different genera, were investigated for their ability to utilize single sugars as substrates (glucose, xylose, arabinose or ribose), and also dual sugars as substrates (a hexose plus a pentose). The results demonstrated that C. glutamicum, A. odontolyticus, N. elegans, and P. freudenreichii were pentose-capable (able to grow using xylose or other pentose sugar), and also showed diauxie growth characteristics during the dual-sugar (glucose, in combination with xylose, arabinose or ribose) carbon source tests. In addition, it was shown that the isolates displayed very small differences in growth rates when grown on dual sugars as compared to single sugars, whether pentose or hexose in nature. The anabolic characteristics of C. glutamicum, A. odontolyticus, N. elegans and P. freudenreichii were subsequently investigated by qualitative analysis of their end-products, using high performance liquid chromatography (HPLC). All of the organisms produced arginine and cysteine after utilization of the pentose substrates alone. In addition, P. freudenreichii produced alanine and glycine. The end-product profile arising from culture with dual carbon sources was also tested. Interestingly, this time the product was different. All of them produced the amino acid glycine, when grown on a combination substrate-mix of glucose with xylose, and also glucose with arabinose. Only N. elegans was able to break down ribose, either singly or in combination with glucose, and the end-product of metabolism of the glucose plus ribose substrate combination was glutamic acid. The ecological analysis of microbial abundance in sugar mill waste was performed using denaturing gradient gel electrophoresis (DGGE) and also the metagenomic microarray PhyloChip method. Eleven solid samples and seven liquid samples were investigated. A very complex bacterial ecosystem was demonstrated in the seven liquid samples after testing with the PhyloChip method. It was also shown that bagasse leachate was the most different, compared to all of the other samples, by virtue of its richness in variety of taxa and the complexity of its bacterial community. The bacterial community in solid samples from Proserpine, Mackay and Maryborough sugar mills showed huge diversity. The information found from 16S rDNA sequencing results was that the bacterial genera Brevibacillus, Rhodospirillaceae, Bacillus, Vibrio and Pseudomonas were present in greatest abundance. In addition, Corynebacterium was also found in the soil samples. The metagenomic studies of the sugar mill samples demonstrate two important outcomes: firstly that the bagasse leachate, as potentially the most pentose-rich sample tested, had the most complex and diverse bacterial community; and secondly that the pentose-capable isolates that were initially discovered at the beginning of this study, were not amongst the most abundant taxonomic groups discovered in the sugar mill samples, and in fact were, as suspected, very rare. As a bioprospecting exercise, therefore, the study has discovered organisms that are naturally present, but in very small numbers, in the appropriate natural environment. This has implications for the industrial application of E-PUB, in that a seeding process using a starter culture will be necessary for industrial purposes, rather than simply assuming that natural fermentation might occur.

Effect of CT electron density identification on Monte Carlo simulations of radiotherapeutic treatments

Introduction: The accurate identification of tissue electron densities is of great importance for Monte Carlo (MC) dose calculations. When converting patient CT data into a voxelised format suitable for MC simulations, however, it is common to simplify the assignment of electron densities so that the complex tissues existing in the human body are categorized into a few basic types. This study examines the effects that the assignment of tissue types and the calculation of densities can have on the results of MC simulations, for the particular case of a Siemen’s Sensation 4 CT scanner located in a radiotherapy centre where QA measurements are routinely made using 11 tissue types (plus air). Methods: DOSXYZnrc phantoms are generated from CT data, using the CTCREATE user code, with the relationship between Hounsfield units (HU) and density determined via linear interpolation between a series of specified points on the ‘CT-density ramp’ (see Figure 1(a)). Tissue types are assigned according to HU ranges. Each voxel in the DOSXYZnrc phantom therefore has an electron density (electrons/cm3) defined by the product of the mass density (from the HU conversion) and the intrinsic electron density (electrons /gram) (from the material assignment), in that voxel. In this study, we consider the problems of density conversion and material identification separately: the CT-density ramp is simplified by decreasing the number of points which define it from 12 down to 8, 3 and 2; and the material-type-assignment is varied by defining the materials which comprise our test phantom (a Supertech head) as two tissues and bone, two plastics and bone, water only and (as an extreme case) lead only. The effect of these parameters on radiological thickness maps derived from simulated portal images is investigated. Results & Discussion: Increasing the degree of simplification of the CT-density ramp results in an increasing effect on the resulting radiological thickness calculated for the Supertech head phantom. For instance, defining the CT-density ramp using 8 points, instead of 12, results in a maximum radiological thickness change of 0.2 cm, whereas defining the CT-density ramp using only 2 points results in a maximum radiological thickness change of 11.2 cm. Changing the definition of the materials comprising the phantom between water and plastic and tissue results in millimetre-scale changes to the resulting radiological thickness. When the entire phantom is defined as lead, this alteration changes the calculated radiological thickness by a maximum of 9.7 cm. Evidently, the simplification of the CT-density ramp has a greater effect on the resulting radiological thickness map than does the alteration of the assignment of tissue types. Conclusions: It is possible to alter the definitions of the tissue types comprising the phantom (or patient) without substantially altering the results of simulated portal images. However, these images are very sensitive to the accurate identification of the HU-density relationship. When converting data from a patient’s CT into a MC simulation phantom, therefore, all possible care should be taken to accurately reproduce the conversion between HU and mass density, for the specific CT scanner used. Acknowledgements: This work is funded by the NHMRC, through a project grant, and supported by the Queensland University of Technology (QUT) and the Royal Brisbane and Women's Hospital (RBWH), Brisbane, Australia. The authors are grateful to the staff of the RBWH, especially Darren Cassidy, for assistance in obtaining the phantom CT data used in this study. The authors also wish to thank Cathy Hargrave, of QUT, for assistance in formatting the CT data, using the Pinnacle TPS. Computational resources and services used in this work were provided by the HPC and Research Support Group, QUT, Brisbane, Australia.

Very sensitive fiber Bragg grating accelerometer using transverse forces with an easy over-range protection and low cross axial sensitivity

The first fiber Bragg grating (FBG) accelerometer using direct transverse forces is demonstrated by fixing the FBG by its two ends and placing a transversely moving inertial object at its middle. It is very sensitive because a lightly stretched FBG is more sensitive to transverse forces than axial forces. Its resonant frequency and static sensitivity are analyzed by the classic spring-mass theory, assuming the axial force changes little. The experiments show that the theory can be modified for cases where the assumption does not hold. The resonant frequency can be modified by a linear relationship experimentally achieved, and the static sensitivity by an alternative method proposed. The principles of the over-range protection and low cross axial sensitivity are achieved by limiting the movement of the FBG and were validated experimentally. The sensitivities 1.333 and 0.634 nm/g were experimentally achieved by 5.29 and 2.83 gram inertial objects at 10 Hz from 0.1 to 0.4 g (g = 9.8 m/s 2), respectively, and their resonant frequencies were around 25 Hz. Their theoretical static sensitivities and resonant frequencies found by the modifications are 1.188 nm/g and 26.81 Hz for the 5.29 gram one and 0.784 nm/g and 29.04 Hz for the 2.83 gram one, respectively.

Effectiveness of cysteine proteases on protein/pigment film removal

Objective: Theaflavin (TF) from the black tea can react to human salivary proline-rich proteins (PRPs) to form stains on exposed dental surfaces. Here, we employed a model of protein/pigment film using TF and dephosphorylated bovine b-casein (Db-CN), which has an extended conformation, similar to that of salivary PRPs, on a sensor surface to assess the efficacy of cysteine proteases (CPs) including papain, stem bromelain, and ficin, on removing TF bound to Db-CN and the control TF readsorption on the residual substrate surfaces was also measured. Methods: The protein/pigment complex film was built by using a quartz crystal microbalance with dissipation (QCM-D). The efficacies of CPs were assessed by Boltzman equation model. The surface details were detected by grazing angle infrared spectroscopy spectra, atomic force microscopy images, and contact angles. Results: The efficacy order of CPs on hydrolyzing protein/pigment complex film is ficin > papain > bromelain. The results from grazing angle infrared spectroscopy spectra, atomic force microscopy images, and contact angles demonstrated that TF bound on the Db- CN was effectively removed by the CPs, and the amount of TF readsorption on both the residual film of the Db-CN/TF and the Db-CN was markedly decreased after hydrolysis. Conclusion: This study indicates the potential application of the CPs for tooth stain removal and suggests that these enzymes are worthy of further investigation for use in oral healthcare.