Quantitative PCR assay of sewage-associated Bacteroides markers to assess sewage pollution in an urban lake in Dhaka, Bangladesh

This paper aimed to assess the magnitude of sewage pollution in an urban lake in Dhaka, Bangladesh by using Quantitative PCR (qPCR) of sewage-associated Bacteroides HF183 markers. PCR was also used for the quantitative detection of ruminant wastewater-associated CF128 markers along with the enumeration of traditional fecal indicator bacteria, namely, enterococci. The number of enterococci in lake water samples ranged from 1.1 x 104 to 1.9 x 105 CFU/100 ml of water. From the 20 water samples tested, 14 (70%) and 7 (35%) were PCR positive for the HF183 and CF128 markers, respectively. The numbers of the HF183 and CF128 markers in lake water samples were 3.9 x 104 to 6.3 × 107 and 9.3 x 103 to 6.3 x 105 genomic units (GU)/100 ml of water, respectively. The high numbers of enterococci and the HF183 markers indicate sewage pollution and potential health risks to those who use the lake water for non-potable purposes such as bathing and washing clothes. This is the first study that investigated the presence of microbial source tracking (MST) markers in Dhaka, Bangladesh where diarrhoeal diseases is one of the major causes of childhood mortality. The molecular assay as used in this study can provide valuable information on the extent of sewage pollution, thus facilitating the development of robust strategies to minimise potential health risks.

Exploring retinal and functional markers of diabetic neuropathy

Diabetic peripheral neuropathy (DPN) is one of the most debilitating complications of diabetes. DPN is a major cause of foot ulceration and lower limb amputation. Early diagnosis and management is a key factor in reducing morbidity and mortality. Current techniques for clinical assessment of DPN are relatively insensitive for detecting early disease or involve invasive procedures such as skin biopsies. There is a need for less painful, non-invasive and safe evaluation methods. Eye care professionals already play an important role in the management of diabetic retinopathy; however recent studies have indicated that the eye may also be an important site for the diagnosis and monitoring of neuropathy. Corneal nerve morphology has been shown to be a promising marker of diabetic neuropathy occurring elsewhere in the body, and emerging evidence tentatively suggests that retinal anatomical markers and a range of functional visual indicators could similarly provide useful information regarding neural damage in diabetes – although this line of research is, as yet, less well established. This review outlines the growing body of evidence supporting a potential diagnostic role for retinal structure and visual functional markers in the diagnosis and monitoring of peripheral neuropathy in diabetes.

Identification of novel markers for uncomplicated lower genital tract infections and upper genital tract pathology due to Chlamydia trachomatis.

Background: Untreated Chlamydia trachomatis infections in women can result in disease sequelae such as salpingitis and pelvic inflammatory disease (PID), ultimately culminating in tubal occlusion and infertility. Whilst nucleic acid amplification tests can effectively diagnose uncomplicated lower genital tract (LGT) infections, they are not suitable for diagnosing upper genital tract (UGT) pathological sequelae. As a consequence, this study aimed to identify serological markers that can, with a high degree of sensitivity and specificity, discriminate between LGT infections and UGT pathology. Methods: Plasma was collected from 73 women with a history of LGT infection, UGT pathology due to C. trachomatis or no serological evidence of C. trachomatis infection. Western blotting was used to analyse antibody reactivity against extracted chlamydial proteins. Sensitivity and specificity of differential markers were also calculated. Results: Four antigens (CT157, CT423, CT727 and CT396) were identified and found to be capable of discriminating between the infection and disease sequelae state. Sensitivity and specificity calculations showed that our assay for diagnosing LGT infection had a sensitivity and specificity of 75% and 76% respectively, whilst the assay for identifying UGT pathology demonstrated 80% sensitivity and 86% specificity. Conclusions: The use of these assays could potentially facilitate earlier diagnoses in women suffering UGT pathology due to C. trachomatis.

Reinforced hydrogels for silicone copolymer delivery for scar remediation

Hypertrophic scars are formed by collagen overproduction in wounded areas and often occur in victims of severe burns. There are several methods for hypertrophic scar remediation and silicone gel therapy is one of the more successful methods. Research by others has shown that the activity of these gels may be due to migration of amphiphilic silicone oligomers from the gel and into the dermis, down-regulating production of collagen by fibroblasts. Normal silicone oil (PDMS) does not produce the same effect on fibroblasts. The main purpose of this project is the introduction of a particular amphiphilic silicone rake copolymer into an appropriate network which can absorb and release the silicone copolymer on the scarred area. Hydrogels are polymeric crosslinked networks which can swell in water or a drug solution, and gradually release the drug when applied to the skin. The application of gel enhances the effectiveness of the therapy, reduces the period of treatment and can be comfortable for patients to use. Polyethylene glycol (PEG) based networks have been applied in this research, because the amphiphilic silicone rake copolymer to be used as a therapy has polyethylene oxide (PEO) as a side chain. These PEO side chains have very similar chemical structure to a PEG gel chain so enhancing both the compatibility and the diffusion of the amphiphilic silicone rake copolymer into and out of the gel. Synthesis of PEG-based networks has been performed by two methods: in situ silsesquioxane formation as crosslink with a sol-gel reaction under different conditions and UV curing. PEG networks have low mechanical properties which is a fundamental limitation of the polymer backbone. For mechanical properties enhancement, composite networks were synthesized using nano-silica with different surface modification. The chemical structure of in situ silsesquioxane in the dry network has been examined by Solid State NMR, Differential Scanning Calorimetry (DSC) and swelling measurements in water. Mechanical properties of dry networks were tested by Dynamic Mechanical Thermal Analysis (DMTA) to determine modulus and interfacial interaction between silica and the network. In this way a family of self-reinforced networks has been produced that have been shown to absorb and deliver the active amphiphilic silicone- PEO rake copolymer.

Scaling digital radiographs for templating in total hip arthroplasty using conventional acetate templates independent of calibration markers

We describe a scaling method for templating digital radiographs using conventional acetate templates independent of template magnification without the need for a calibration marker. The mean magnification factor for the radiology department was determined (119.8%, range117%-123.4%). This fixed magnification factor was used to scale the radiographs by the method described. 32 femoral heads on postoperative THR radiographs were then measured and compared to the actual size. The mean absolute accuracy was within 0.5% of actual head size (range 0 to 3%) with a mean absolute difference of 0.16mm (range 0-1mm, SD 0.26mm). Intraclass Correlation Coefficient (ICC) showed excellent reliability for both inter and intraobserver measurements with ICC scores of 0.993 (95% CI 0.988-0.996) for interobserver measurements and intraobserver measurements ranging between 0.990-0.993 (95% CI 0.980-0.997).

Corneal markers of diabetic neuropathy

Diabetic neuropathy is a significant clinical problem that currently has no effective therapy, and in advanced cases, leads to foot ulceration and lower limb amputation. The accurate detection, characterization and quantification of this condition are important in order to define at-risk patients, anticipate deterioration, monitor progression, and assess new therapies This review evaluates novel corneal methods of assessing diabetic neuropathy. Two new non-invasive corneal markers have emerged, and in cross-sectional studies have demonstrated their ability to stratify the severity of this disease. Corneal confocal microscopy allows quantification of corneal nerve parameters and non-contact corneal esthesiometry, the functional correlate of corneal structure, assesses the sensitivity of the cornea. Both these techniques are quick to perform, produce little or no discomfort for the patient, and are suitable for clinical settings. Each has advantages and disadvantages over traditional techniques for assessing diabetic neuropathy. Application of these new corneal markers for longitudinal evaluation of diabetic neuropathy has the potential to reduce dependence on more invasive, costly, and time-consuming assessments, such as skin biopsy.

Neuropsychological development in children with early and continuously treated phenylketonuria : association with biochemical markers

PKU is a genetically inherited inborn error of metabolism caused by a deficiency of the enzyme phenylalanine hydroxylase. The failure of this enzyme causes incomplete metabolism of protein ingested in the diet, specifically the conversion of one amino acid, phenylalanine, to tyrosine, which is a precursor to the neurotransmitter dopamine. Rising levels of phenylalanine is toxic to the developing brain, disrupting the formation of white matter tracts. The impact of tyrosine deficiency is not as well understood, but is hypothesized to lead to a low dopamine environment for the developing brain. Detection in the newborn period and continuous treatment (a low protein phe-restricted diet supplemented with phenylalanine-free protein formulas) has resulted in children with early and continuously treated PKU now developing normal I.Q. However, deficits in executive function (EF) are common, leading to a rate of Attention Deficit Hyperactivity Disorder (ADHD) up to five times the norm. EF worsens with exposure to higher phenylalanine levels, however recent research has demonstrated that a high phenylalanine to tyrosine ratio (phenylalanine:tyrosine ratio), which is hypothesised to lead to poorer dopamine function, has a more negative impact on EF than phenylalanine levels alone. Research and treatment of PKU is currently phenylalanine-focused, with little investigation of the impact of tyrosine on neuropsychological development. There is no current consensus as to the veracity of tyrosine monitoring or treatment in this population. Further, the research agenda in this population has demonstrated a primary focus on EF impairment alone, even though there may be additional neuropsychological skills compromised (e.g., mood, visuospatial deficits). The aim of this PhD research was to identify residual neuropsychological deficits in a cohort of children with early and continuously treated phenylketonuria, at two time points in development (early childhood and early adolescence), separated by eight years. In addition, this research sought to determine which biochemical markers were associated with neuropsychological impairments. A clinical practice survey was also undertaken to ascertain the current level of monitoring/treatment of tyrosine in this population. Thirteen children with early and continuously treated PKU were tested at mean age 5.9 years and again at mean age 13.95 years on several neuropsychological measures. Four children with hyperphenylalaninemia (a milder version of PKU) were also tested at both time points and provide a comparison group in analyses. Associations between neuropsychological function and biochemical markers were analysed. A between groups analysis in adolescence was also conducted (children with PKU compared to their siblings) on parent report measures of EF and mood. Minor EF impairments were evident in the PKU group by age 6 years and these persisted into adolescence. Life-long exposure to high phenylalanine:tyrosine ratio and/or low tyrosine independent of phenylalanine were significantly associated with EF impairments at both time points. Over half the children with PKU showed severe impairment on a visuospatial task, and this was associated only with concurrent levels of tyrosine in adolescence. Children with PKU also showed a statistically significant decline in a language comprehension task from 6 years to adolescence (going from normal to subnormal), this deficit was associated with lifetime levels of phenylalanine. In comparison, the four children with hyperphenylalaninemia demonstrated normal function at both time points, across all measures. No statistically significant differences were detected between children with PKU and their siblings on the parent report of EF and mood. However, depressive symptoms were significantly correlated with: EF; long term high phe:tyr exposure; and low tyrosine levels independent of phenylalanine. The practice survey of metabolic clinics from 12 countries indicated a high level of variability in terms of monitoring/treatment of tyrosine in this population. Whilst over 80% of clinics surveyed routinely monitored tyrosine levels in their child patients, 25% reported treatment strategies to increase tyrosine (and thereby lower the phenylalanine:tyrosine ratio) under a variety of patient presentation conditions. Overall, these studies have shown that EF impairments associated with PKU provide support for the dopamine-deficiency model. A language comprehension task showed a different trajectory, serving a timely reminder that non-EF functions also remain vulnerable in this population; and that normal function in childhood does not guarantee normal function by adolescence. Mood impairments were associated with EF impairments as well as long term measures of phenylalanine:tyrosine and/or tyrosine. The implications of this research for enhanced clinical guidelines are discussed given varied current practice.

Psycho-markers of weight loss. The roles of TFEQ Disinhibition and Restraint in exercise-induced weight management

Eating behaviour traits, namely Disinhibition and Restraint, have the potential to exert an effect on food intake and energy balance. The effectiveness of exercise as a method of weight management could be influenced by these traits. Fifty eight overweight and obese participants completed 12-weeks of supervised exercise. Each participant was prescribed supervised exercise based on an expenditure of 500 kcal/session, 5 d/week for 12-weeks. Following 12-weeks of exercise there was a significant reduction in mean body weight (-3.26 ± 3.63 kg), fat mass (FM: -3.26 ± 2.64 kg), BMI (-1.16 ± 1.17 kg/m2)and waist circumference (WC: -5.0 ± 3.23 cm). Regression analyses revealed a higher baseline Disinhibition score was associated with a greater reduction in BMI and WC, while Internal Disinhibition was associated with a larger decrease in weight, %FM and WC. Neither baseline Restraint or Hunger were associated with any of the anthropometric markers at baseline or after 12-weeks. Furthermore, after 12-weeks of exercise, a decrease in Disinhibition and increase in Restraint were associated with a greater reduction in WC, whereas only Restraint was associated with a decrease in weight. Post-hoc analysis of the sub-factors revealed a decrease in External Disinhibition and increase in Flexible Restraint were associated with weight loss. However, an increase in Rigid Restraint was associated with a reduction in %FM and WC. These findings suggest that exercise-induced weight loss is more marked in individuals with a high level of Disinhibition. These data demonstrate the important roles that Disinhibition and Restraint play in the relationship between exercise and energy balance.

Pre-clinical evaluation of amphiphilic silicone oligomers for scar remediation

The formation of hypertrophic scars is a frequent outcome of wound repair and often requires further therapy with treatments such as silicone gel sheets (SGS; Perkins et al., 1983). Although widely used, knowledge regarding SGS and their mechanism of action on hypertrophic scars is limited. Furthermore, SGS require consistent application for at least twelve hours a day for up to twelve consecutive months, beginning as soon as wound reepithelialisation has occurred. Preliminary research at QUT has shown that some species of silicone present in SGS have the ability to permeate into collagen gel skin mimetics upon exposure. An analogue of these species, GP226, was found to decrease both collagen synthesis and the total amount of collagen present following exposure to cultures of cells derived from hypertrophic scars. This silicone of interest was a crude mixture of silicone species, which resolved into five fractions of different molecular weight. These five fractions were found to have differing effects on collagen synthesis and cell viability following exposure to fibroblasts derived from hypertrophic scars (HSF), keloid scars (KF) and normal skin (nHSF and nKF). The research performed herein continues to further assess the potential of GP226 and its fractions for scar remediation by determining in more detail its effects on HSF, KF, nHSF, nKF and human keratinocytes (HK) in terms of cell viability and proliferation at various time points. Through these studies it was revealed that Fraction IV was the most active fraction as it induced a reduction in cell viability and proliferation most similar to that observed with GP226. Cells undergoing apoptosis were also detected in HSF cultures exposed to GP226 and Fraction IV using the Tunel assay (Roche). These investigations were difficult to pursue further as the fractionation process used for GP226 was labour-intensive and time inefficient. Therefore a number of silicones with similar structure to Fraction IV were synthesised and screened for their effect following application to HSF and nHSF. PDMS7-g-PEG7, a silicone-PEG copolymer of low molecular weight and low hydrophilic-lipophilic balance factor, was found to be the most effective at reducing cell proliferation and inducing apoptosis in cultures of HSF, nHSF and HK. Further studies investigated gene expression through microarray and superarray techniques and demonstrated that many genes are differentially expressed in HSF following treatment with GP226, Fraction IV and PDMS7-g-PEG7. In brief, it was demonstrated that genes for TGFβ1 and TNF are not differentially regulated while genes for AIFM2, IL8, NSMAF, SMAD7, TRAF3 and IGF2R show increased expression (>1.8 fold change) following treatment with PDMS7-g-PEG7. In addition, genes for αSMA, TRAF2, COL1A1 and COL3A1 have decreased expression (>-1.8 fold change) following treatment with GP226, Fraction IV and PDMS7-g-PEG7. The data obtained suggest that many different pathways related to apoptosis and collagen synthesis are affected in HSF following exposure to PDMS7-g-PEG7. The significance is that silicone-PEG copolymers, such as GP226, Fraction IV and PDMS7-g-PEG7, could potentially be a non-invasive substitute to apoptosis-inducing chemical agents that are currently used as scar treatments. It is anticipated that these findings will ultimately contribute to the development of a novel scar therapy with faster action and improved outcomes for patients suffering from hypertrophic scars.

Markers of chronic disease in offenders in a high secure correctional centre in Queensland

This study aimed to gauge the presence of markers of chronic disease, as a basis for food and nutrition policy in correctional facilities. One hundred and twenty offenders, recruited from a Queensland Correctional Centre, provided informed consent and completed both dietary interviews and physical measurements. Mean age of the sample was 35.5 ± 12 years (range = 19–77 yrs); mean age of the total population (n = 945) was 32.8 ± 10 years (range = 19–80 yrs). Seventy-nine participants also provided fasting blood samples. The mean body mass index (BMI) was 27 ± 3.5 kg/m2; 72% having a BMI > 25 kg/m2. Thirty-three percent were classified overweight or obese using waist circumference (mean = 92 ± 10 cm). Mean blood pressure measurement was systolic = 130 ± 14 mmHg and diastolic = 73 ± 10 mmHg. Twenty-four percent were classified as hypertensive of whom three were on antihypertensive medication. Eighteen percent had elevated triglycerides, and 40% unfavourable total cholesterol to HDL ratios. Homeostatic Model Assessment (HOMA scores) were calculated from glucose and insulin. Four participants were insulin resistant, two of whom had known diabetes. Metabolic syndrome, based on waist circumference (adjusted for ethnicity), blood lipids, blood pressure and plasma glucose indicated that 25% (n = 20) were classified with metabolic syndrome. Eighty-four percent (n = 120) reported some physical activity each day, with 51 percent participating ≥two times daily. Fifty-four percent reported smoking with an additional 20% having smoked in the past. Findings suggest that waist circumference rather than weight and BMI only should be used in this group to determine weight status. The data suggest that markers of chronic disease are present and that food and nutrition policy must reflect this. Further analysis is being completed to determine relevant policy initiatives.

Isolation and characterisation of novel microsatellite and mitochondrial DNA markers for the Eastern Water Dragon (Physignathus lesueurii)

Habitat fragmentation as a result of urbanisation is a growing problem for native lizard species. The Eastern Water Dragon (Physignathus lesueurii) is a social arboreal agamid lizard, native to Australia. This species represents an ideal model species to investigate the effect of urbanisation because of their prominent abundance in the urban landscape. Here we describe the isolation and characterisation of a novel set of 74 di-, tri-, and tetramicrosatellites from which 18 were selected and optimised into two multiplexes. The 18 microsatellites generated a total 148 alleles across the two populations. The number of alleles per locus varied from 2 to 18 alleles and measures of Ho and He varied from 0.395 to 0.877 and from 0.441 to 0.880, respectively. We also present primers for four novel mitochondrial DNA (mtDNA) markers. The combined length of the four mtDNA marker pairs was 2,528 bp which included 15 nucleotides changes. In comparison to threatened species, which are generally characterised by small population sizes, the Eastern Water Dragon represents an ideal model species to investigate the effect of urbanisation on their behavioural ecology and connectivity patterns among populations.