MR image-based measurement of rates of change in volumes of brain structures. Part 1: Method and validation

A detailed analysis procedure is described for evaluating rates of volumetric change in brain structures based on structural magnetic resonance (MR) images. In this procedure, a series of image processing tools have been employed to address the problems encountered in measuring rates of change based on structural MR images. These tools include an algorithm for intensity non-uniforniity correction, a robust algorithm for three-dimensional image registration with sub-voxel precision and an algorithm for brain tissue segmentation. However, a unique feature in the procedure is the use of a fractional volume model that has been developed to provide a quantitative measure for the partial volume effect. With this model, the fractional constituent tissue volumes are evaluated for voxels at the tissue boundary that manifest partial volume effect, thus allowing tissue boundaries be defined at a sub-voxel level and in an automated fashion. Validation studies are presented on key algorithms including segmentation and registration. An overall assessment of the method is provided through the evaluation of the rates of brain atrophy in a group of normal elderly subjects for which the rate of brain atrophy due to normal aging is predictably small. An application of the method is given in Part 11 where the rates of brain atrophy in various brain regions are studied in relation to normal aging and Alzheimer's disease. (C) 2002 Elsevier Science Inc. All rights reserved.

Post-discharge surveillance: can patients reliably diagnose surgical wound infections?

Post-discharge surgical wound infection surveillance is an important part of many infection control programs. It is frequently undertaken by patient self-assessment, prompted either by a telephone or postal questionnaire. To assess the reliability of this method, 290 patients were followed for six weeks post-operatively. Their wounds were photographed and also covertly assessed for signs of infection by two experienced infection control nurses (ICNs). Patients also responded to a postal questionnaire seeking evidence of infection at both week four and week six post-surgery. Correlation between the patient's assessment of their wound and the ICNs diagnosis was poor (r = 0. 37) with a low positive predictive value (28.7%), although negative predictive value was high (98.2%). Assessment of photos for signs of infection by two experienced clinicians also correlated poorly with the ICNs diagnosis of infection (r = 0.54). The patient's recall of prescription of an antibiotic by their general practitioner (GP) for wound infection during the postoperative period correlated best with the ICNs diagnosis (r = 0.76). This latter measure, particularly when confirmed by the GP in those patients reporting an infection, appears to provide the most valid and resource efficient marker of post-discharge surgical wound infection.

Influence of planting date, method of handling and seed size on supersweet sweet corn seed quality

A supersweet sweet corn hybrid, Pacific H5, was planted at weekly intervals (P-1 to P-5) in spring in South-Eastern Queensland. All plantings were harvested at the same time resulting in immature seed for the last planting (P-5). The seed was handled by three methods: manual harvest and processing (M-1), manual harvest and mechanical processing (M-2) and mechanical harvest and processing (M-3), and later graded into three sizes (small, medium and large). After eight months storage at 12-14degreesC, seed was maintained at 30degreesC with bimonthly monitoring of germination for fourteen months and seed damage at the end of this period. Seed quality was greatest for M-1 and was reduced by mechanical processing but not by mechanical harvesting. Large and medium seed had higher germination due to greater storage reserves but also more seed damage during mechanical processing. Immature seed from premature harvest (P-5) had poor quality especially when processed mechanically and reinforced the need for harvested seed to be physiologically mature.

Purification of post-translationally modified proteins from bacteria: homologous expression and purification of histidine-tagged pilin from Neisseria meningitidis

Until recently, glycosylation of proteins in prokaryotes was regarded as uncommon and thought to be limited to special cases such as S-layer proteins and some archeal outer membrane proteins. Now, there are an increasing number of reports of bacterial proteins that are glycosylated. Pilin of pathogenic Neisseria is one of the best characterised post-translation ally modified bacterial proteins, with four different types of modifications reported, including a novel glycosylation. Pilin monomers assemble to form pilus fibres, which are long protein filaments that protrude from the surface of bacterial cells and are key virulence factors. To aid in the investigation of these modifications, pure pilin is required. A number of pilin purification methods have been published, but none are appropriate for the routine purification of pilin from many different isolates. This study describes a novel, rapid, and simple method of pilin purification from Neisseria meningitidis C311#3, which facilitates the production of consistent quantities of pure, native pilin. A 6 x histidine tag was fused to the C-terminus of the pilin subunit structural gene, pilE, via homologous recombination placing the 6 x histidine-tagged allele in the chromosome of N. meningitidis C311#3. Pilin was purified under non-denaturing conditions via a two-step process using immobilised metal affinity chromatography (IMAC), followed by dye affinity chromatography. Analysis of the purified pilin confirmed that it retained both of the post-translational modifications examined. This novel approach may prove to be a generally applicable method for purification and analysis of post-translationally modified proteins in bacteria. (C) 2003 Elsevier Science (USA). All rights reserved.

Rabbit CYP4B1 engineered for high-level expression in Escherichia coli: ligand stabilization and processing of the N-terminus and heme prosthetic group

Modifications at the N-terminus of the rabbit CYP4B1 gene resulted in expression levels in Escherichia coli of up to 660 nmol/L. Solubilization of the enzyme from bacterial membranes led to substantial conversion to cytochrome P420 unless alpha-naphthoflavone was added as a stabilizing ligand. Mass spectrometry analysis and Edman sequencing of purified enzyme preparations revealed differential N-terminal post-translational processing of the various constructs expressed. Notably, bacterial expression of CYP4B1 produced a holoenzyme with >98.5% of its heme prosthetic group covalently linked to the protein backbone. The near fully covalently linked hernoproteins exhibited similar rates and regioselectivities of lauric acid hydroxylation to that observed previously for the partially heme processed enzyme expressed in insect cells. These studies shed new light on the consequences of covalent heme processing in CYP4B1 and provide a facile system for future mechanistic and structural studies with the enzyme. (C) 2003 Elsevier Science (USA). All rights reserved.

Fruit processing, seed viability and dormancy mechanisms of Persoonia sericea A. Cunn. ex R. Br. and P-virgata R. Br. (Proteaceae)

The morphology of the fruit and difficulties with fruit processing impose major limitations to germination of Persoonia sericea and P. virgata. The mesocarp must be removed without harming the embryo. Fermentation of fruit or manual removal of the mesocarp was effective but digestion in 32% hydrochloric acid (HCl) completely inhibited germination. The endocarp is extremely hard and therefore very difficult and time consuming to remove without damaging the seeds. The most efficient method was cracking the endocarp with pliers, followed by manual removal of seeds. Germination was completely inhibited unless at least half of the endocarp was removed. Microbial contamination of the fruit and seeds was controlled by disinfestation and germination of the seed under aseptic conditions. The results suggest that dormancy in these species is primarily due to physical restriction of the embryo by the hard endocarp.

A rapid HPLC-mass spectrometry cyclosporin method suitable for current monitoring practices

Objectives: Cyclosporin is an immunosuppressant drug with a narrow therapeutic window. Trough and 2-h post-dose blood samples are currently used for therapeutic drug monitoring in solid organ transplant recipients. The aim of the current study was to develop a rapid HPLC-tandem mass spectrometry (HPLC-MS) method for the measurement of cyclosporin in whole blood that was not only suitable for the clinical setting but also considered a reference method. Methods: Blood samples (50 mu L) were prepared by protein precipitation followed by C-18 solid-phase extraction while using d(12) cyclosporin as the internal standard. Mass spectrometric detection was by selected reaction monitoring with an electrospray interface in positive ionization mode. Results: The assay was linear from 10 to 2000 mu g/L (r(2) > 0.996, n = 9). Inter-day,analytical recovery and imprecision using whole blood quality control samples at 10, 30, 400, 1500, and 2000 mu g/L were 94.9-103.5% and

Post-crystallization treatments for improving diffraction quality of protein crystals

X-ray crystallography is the most powerful method for determining the three-dimensional structure of biological macromolecules. One of the major obstacles in the process is the production of high-quality crystals for structure determination. All too often, crystals are produced that are of poor quality and are unsuitable for diffraction studies. This review provides a compilation of post-crystallization methods that can convert poorly diffracting crystals into data-quality crystals. Protocols for annealing, dehydration, soaking and cross-linking are outlined and examples of some spectacular changes in crystal quality are provided. The protocols are easily incorporated into the structure-determination pipeline and a practical guide is provided that shows how and when to use the different post-crystallization treatments for improving crystal quality.

A microarray study of post-mortem mRNA degradation in mouse brain tissue

Background: Although there is evidence that post-mortem interval (PMI) is not a major contributor to reduced overall RNA integrity, it may differentially affect a subgroup of gene transcripts that are susceptible to PMI-related degradation. This would particularly have ramifications for microarray studies that include a broad spectrum of genes. Method: Brain tissue was removed from adult mice at 0, 6, 12, 18, 24,36 and 48 h post-mortem. RNA transcript abundance was measured by hybridising RNA from the zero time point with test RNA from each PMI time point, and differential gene expression was assessed using cDNA microarrays. Sequence and ontological analyses were performed on the group of RNA transcripts showing greater than two-fold reduction. Results: Increasing PMI was associated with decreased tissue pH and increased RNA degradation as indexed by 28S/18S ribosomal RNA ratio. Approximately 12% of mRNAs detected on the arrays displayed more than a two-fold decrease in abundance by 48 It post-mortem. An analysis of nucleotide composition provided evidence that transcripts with the AUUUA motif in the 3' untranslated region (3'UTR) were more susceptible to PMI-related RNA degradation, compared to transcripts not carrying the 3'UTR AUUUA motif. Consistent with this finding, ontological analysis showed transcription factors and elements to be over-represented in the group of transcripts susceptible to degradation. Conclusion: A subgroup of mammalian mRNA transcripts are particularly susceptible to PMI-related degradation, and as a group, they are more likely to carry the YUTR AUUUA motif. PMI should be controlled for in human and animal model post-mortem brain studies, particularly those including a broad spectrum of mRNA transcripts. (c) 2005 Elsevier B.V. All rights reserved.

An adaptive and dynamic dimensionality reduction method for high-dimensional indexing

The notorious "dimensionality curse" is a well-known phenomenon for any multi-dimensional indexes attempting to scale up to high dimensions. One well-known approach to overcome degradation in performance with respect to increasing dimensions is to reduce the dimensionality of the original dataset before constructing the index. However, identifying the correlation among the dimensions and effectively reducing them are challenging tasks. In this paper, we present an adaptive Multi-level Mahalanobis-based Dimensionality Reduction (MMDR) technique for high-dimensional indexing. Our MMDR technique has four notable features compared to existing methods. First, it discovers elliptical clusters for more effective dimensionality reduction by using only the low-dimensional subspaces. Second, data points in the different axis systems are indexed using a single B+-tree. Third, our technique is highly scalable in terms of data size and dimension. Finally, it is also dynamic and adaptive to insertions. An extensive performance study was conducted using both real and synthetic datasets, and the results show that our technique not only achieves higher precision, but also enables queries to be processed efficiently. Copyright Springer-Verlag 2005

National audit of post-operative management in spinal surgery

Background: There is some evidence from a Cochrane review that rehabilitation following spinal surgery may be beneficial. Methods: We conducted a survey of current post-operative practice amongst spinal surgeons in the United Kingdom in 2002 to determine whether such interventions are being included routinely in the post-operative management of spinal patients. The survey included all surgeons who were members of either the British Association of Spinal Surgeons ( BASS) or the Society for Back Pain Research. Data on the characteristics of each surgeon and his or her current pattern of practice and post-operative care were collected via a reply-paid postal questionnaire. Results: Usable responses were provided by 57% of the 89 surgeons included in the survey. Most surgeons (79%) had a routine post-operative management regime, but only 35% had a written set of instructions that they gave to their patients concerning this. Over half (55%) of surgeons do not send their patients for any physiotherapy after discharge, with an average of less than two sessions of treatment organised by those that refer for physiotherapy at all. Restrictions on lifting, sitting and driving showed considerable inconsistency both between surgeons and also within the recommendations given by individual surgeons. Conclusion: Demonstrable inconsistencies within and between spinal surgeons in their approaches to post-operative management can be interpreted as evidence of continuing and significant uncertainty across the sub-speciality as to what does constitute best care in these areas of practice. Conducting further large, rigorous, randomised controlled trials would be the best method for obtaining definitive answers to these questions.

Obsessive-compulsive disorder across developmental trajectory: Cognitive processing of threat in children, adolescents and adults

Background. While the cognitive theory of obsessive-compulsive disorder (OCD) is one of the most widely accepted accounts of the maintenance of the disorder in adults, no study to date has systematically evaluated the theory across children, adolescence and adults with OCD. Method. This paper investigated developmental differences in the cognitive processing of threat in a sample of children, adolescents and adults with OCD. Using an idiographic assessment approach, as well as self-report questionnaires, this study evaluated cognitive appraisals of responsibility, probability, severity, thought-action fusion (TAF), thought-suppression, self-doubt and cognitive control. It was hypothesised that there would be age related differences in reported responsibility for harm, probability of harm, severity of harm, thought suppression, TAR self-doubt and cognitive control. Results. Results of this study demonstrated that children with OCD reported experiencing fewer intrusive thoughts, which were less distressing and less uncontrollable than those experienced by adolescents and adults with OCD. Furthermore, responsibility attitudes, probability biases and thought suppression strategies were higher in adolescents and adults with OCD. Cognitive processes of TAF, perceived severity of harm, self-doubt and cognitive control were found to be comparable across age groups. Conclusions. These results suggest that the current cognitive theory of OCD needs to address developmental differences in the cognitive processing of threat. Furthermore, for a developmentally sensitive theory of OCD, further investigation is warranted into other possible age related maintenance factors. Implications of this investigation and directions for future research are discussed.

The economics of inclusion body processing

Many recombinant proteins are often over-expressed in host cells, such as Escherichia coli, and are found as insoluble and inactive protein aggregates known as inclusion bodies (IBs). Recently, a novel process for IB extraction and solubilisation, based on chemical extraction, has been reported. While this method has the potential to radically intensify traditional IB processing, the process economics of the new technique have yet to be reported. This study focuses on the evaluation of process economics for several IB processing schemes based on chemical extraction and/or traditional techniques. Simulations and economic analysis were conducted at various processing conditions using granulocyte macrophage-colony stimulating factor, expressed as IBs in E. coli, as a model protein. In most cases, IB processing schemes based on chemical extraction having a shorter downstream cascade demonstrated a competitive economic edge over the conventional route, validating the new process as an economically more viable alternative for IB processing.

Routine physiotherapy does not induce cardiorespiratory training effect post-stroke

Background and Purpose. Cardiorespiratory fitness is increasingly being recognized as an impairment requiring physiotherapy intervention after stroke. The present study seeks to investigate if routine physiotherapy treatment is capable of inducing a cardiorespiratory training effect and if stroke patients attending physiotherapy who are unable to walk experience less cardiorespiratory stress during physiotherapy when compared to those who are able to walk. Method. A descriptive, observational study, with heart rate monitoring and video-recording of physiotherapy rehabilitation, was conducted. Thirty consecutive stroke patients from a geriatric and rehabilitation unit of a tertiary metropolitan hospital, admitted for rehabilitation, and requiring physiotherapy were included in the study. The main measures of the study were duration (time) and intensity (percentage of heart rate reserve) of standing and walking activities during physiotherapy rehabilitation for non-walking and walking stroke patients. Results. Stroke patients spent an average of 21 minutes participating in standing and walking activities that were capable of inducing a cardiorespiratory training effect. Stroke patients who were able to walk spent longer in these activities during physiotherapy rehabilitation than non-walking stroke patients (p < 0.05). An average intensity of 24% heart rate reserve (HRR) during standing and walking activities was insufficient to result in a cardiorespiratory training effect, with a maximum of 35% achieved for the stroke patients able to walk and 30% for those unable to walk. Conclusions. Routine physiotherapy rehabilitation had insufficient duration and intensity to result in a cardiorespiratory training effect in our group of stroke patients. Copyright © 2006 John Wiley & Sons, Ltd.

Biologically inspired visual landmark processing for simultaneous localization and mapping

This paper illustrates a method for finding useful visual landmarks for performing simultaneous localization and mapping (SLAM). The method is based loosely on biological principles, using layers of filtering and pooling to create learned templates that correspond to different views of the environment. Rather than using a set of landmarks and reporting range and bearing to the landmark, this system maps views to poses. The challenge is to produce a system that produces the same view for small changes in robot pose, but provides different views for larger changes in pose. The method has been developed to interface with the RatSLAM system, a biologically inspired method of SLAM. The paper describes the method of learning and recalling visual landmarks in detail, and shows the performance of the visual system in real robot tests.