Assessment of motor symptoms and functional impact in prodromal and early Huntington disease.

The Functional Rating Scale Taskforce for pre-Huntington Disease (FuRST-pHD) is a multinational, multidisciplinary initiative with the goal of developing a data-driven, comprehensive, psychometrically sound, rating scale for assessing symptoms and functional ability in prodromal and early Huntington disease (HD) gene expansion carriers. The process involves input from numerous sources to identify relevant symptom domains, including HD individuals, caregivers, and experts from a variety of fields, as well as knowledge gained from the analysis of data from ongoing large-scale studies in HD using existing clinical scales. This is an iterative process in which an ongoing series of field tests in prodromal (prHD) and early HD individuals provides the team with data on which to make decisions regarding which questions should undergo further development or testing and which should be excluded. We report here the development and assessment of the first iteration of interview questions aimed to assess functional impact of motor manifestations in prHD and early HD individuals.

An item response analysis of the motor and behavioral subscales of the unified Huntington's disease rating scale in huntington disease gene expansion carriers

Although the Unified Huntington's Disease Rating Scale (UHDRS) is widely used in the assessment of Huntington disease (HD), the ability of individual items to discriminate individual differences in motor or behavioral manifestations has not been extensively studied in HD gene expansion carriers without a motor-defined clinical diagnosis (ie, prodromal-HD or prHD). To elucidate the relationship between scores on individual motor and behavioral UHDRS items and total score for each subscale, a nonparametric item response analysis was performed on retrospective data from 2 multicenter longitudinal studies. Motor and behavioral assessments were supplied for 737 prHD individuals with data from 2114 visits (PREDICT-HD) and 686 HD individuals with data from 1482 visits (REGISTRY). Option characteristic curves were generated for UHDRS subscale items in relation to their subscale score. In prHD, overall severity of motor signs was low, and participants had scores of 2 or above on very few items. In HD, motor items that assessed ocular pursuit, saccade initiation, finger tapping, tandem walking, and to a lesser extent, saccade velocity, dysarthria, tongue protrusion, pronation/supination, Luria, bradykinesia, choreas, gait, and balance on the retropulsion test were found to discriminate individual differences across a broad range of motor severity. In prHD, depressed mood, anxiety, and irritable behavior demonstrated good discriminative properties. In HD, depressed mood demonstrated a good relationship with the overall behavioral score. These data suggest that at least some UHDRS items appear to have utility across a broad range of severity, although many items demonstrate problematic features.

Calciferols and bait shyness in the laboratory rat

Rodenticides with delayed action are generally more effective than fast-acting compounds because of the phenomenon of bait shyness. Calciferols have a stop-feed effect quite soon after dosing, and physiological effects are measurable within one day of dosing. We investigated whether bait shyness might result from these fairly rapid effects in the laboratory rat. We found evidence of bait shyness following recovery from sub-lethal dosing with two forms of calciferol. Use of intubation as well as feeding showed that the response was to the bait carrier rather than to detection of calciferols per se.

Interaction with avian cells and colonisation of specific pathogen free chicks by Shiga-toxin negative Escherichia coli O157:H7 (NCTC 12900)

The prevalence of Escherichia coli O157:H7 infection in birds is low but several deliberate inoculation studies show that poultry are readily and persistently infected by this organism indicating a possible threat to public health. The mechanisms of colonisation of poultry are not understood and the aim is to establish models to study the interaction of E. coli O157:H7, at the cellular and whole animal levels. A non-toxigenic E. coli O157:H7 (NCTC 12900) was used in adherence assays with an avian epithelial cell line (Div-1) and used to inoculate 1-day-old SPF chicks. In vitro, NCTC 12900 induced micro-colonies associated with cytoskeletal arrangements and pedestal formation with intimate bacterial attachment. In the 1-day-old SPF chick, a dose of 1 x 10(5) cfu resulted in rapid and extensive colonisation of the gastrointestinal tract and transient colonisation of the liver and spleen. The number of E. coli O157:H7 organisms attained approximately 10(8) cfu/ml caecal homogenate 24 h after inoculation and approximately 10(7) cfu/ml caecal homogenate was still present at day 92. Faecal shedding persisted for 169 days, ceasing 9 days after the birds came into lay and 6% of eggs were contaminated on the eggshell. Histological analysis of tissue samples from birds dosed with 1 x 10(7) cfu gave evidence for E coli O157:H7 NCTC 12900 induced micro-colonies on the caecal mucosa, although evidence for attaching effacing lesions was equivocal. These models may be suitable to study those factors of E. coli O157:H7 that mediate persistent colonisation in avian species.

Colonization of 8-week-old conventionally reared goats by Escherichia coli O157:H7 after oral inoculation

Enterohaemorrhagic Escherichia coli O157 : H7 infections of man have been associated with consumption of unpasteurized goat's milk and direct contact with kid goats on petting farms, yet little is known about colonization of goats with this organism. To assess the contribution of flagella and intimin of E coli O157 : H7 in colonization of the goat, 8-week-old conventionally reared goats were inoculated orally in separate experiments with 1 X 10(10) c.f.u. of a non-verotoxigenic strain of E coli O157: H7 (strain NCTC 12900 Nal(r)), an aflagellate derivative (DMB1) and an intimin-deficient derivative (DMB2). At 24 In after inoculation, the three E coli O157 : H7 strains were shed at approximately 5 X 1 04 c.f.u. (g faeces)(-1) from all animals. Significantly fewer intimin-deficient bacteria were shed only on days 2 (P = 0(.)003) and 4 (P = 0(.)014), whereas from day 7 to 29 there were no differences. Tissues from three animals inoculated with wild-type E coli O157 : H7 strain NCTC 12900 Nalr were sampled at 24,48 and 96 In after inoculation and the organism was cultured from the large intestine of all three animals and from the duodenum and ileum of the animal examined at 96 h. Tissues were examined histologically but attaching-effacing (AE) lesions were not observed at any intestinal site of the animals examined at 24 or 48 In. However, the animal examined at 96 h, which had uniquely shed approximately 1 x 10(7) E coli O157: H7 (g faeces)(-1) for the preceding 3 days, showed a heavy, diffuse infection with cryptosporidia. and abundant, multifocal AE lesions in the distal colon, rectum and at the recto-anal junction. These AE lesions were confirmed by immunohistochemistry to be associated with E coli O157: H7.

Phosphorylation of the voltage-gated potassium channel Kv2.1 by AMP-activated protein kinase regulates membrane excitability

Firing of action potentials in excitable cells accelerates ATP turnover. The voltage-gated potassium channel Kv2.1 regulates action potential frequency in central neurons, whereas the ubiquitous cellular energy sensor AMP-activated protein kinase (AMPK) is activated by ATP depletion and protects cells by switching off energy-consuming processes. We show that treatment of HEK293 cells expressing Kv2.1 with the AMPK activator A-769662 caused hyperpolarizing shifts in the current-voltage relationship for channel activation and inactivation. We identified two sites (S440 and S537) directly phosphorylated on Kv2.1 by AMPK and, using phosphospecific antibodies and quantitative mass spectrometry, show that phosphorylation of both sites increased in A-769662-treated cells. Effects of A-769662 were abolished in cells expressing Kv2.1 with S440A but not with S537A substitutions, suggesting that phosphorylation of S440 was responsible for these effects. Identical shifts in voltage gating were observed after introducing into cells, via the patch pipette, recombinant AMPK rendered active but phosphatase-resistant by thiophosphorylation. Ionomycin caused changes in Kv2.1 gating very similar to those caused by A-769662 but acted via a different mechanism involving Kv2.1 dephosphorylation. In cultured rat hippocampal neurons, A-769662 caused hyperpolarizing shifts in voltage gating similar to those in HEK293 cells, effects that were abolished by intracellular dialysis with Kv2.1 antibodies. When active thiophosphorylated AMPK was introduced into cultured neurons via the patch pipette, a progressive, time-dependent decrease in the frequency of evoked action potentials was observed. Our results suggest that activation of AMPK in neurons during conditions of metabolic stress exerts a protective role by reducing neuronal excitability and thus conserving energy.

Codon and mRNA sequence optimization of microdystrophin transgenes improves expression and physiological outcome in dystrophic mdx mice following AAV2/8 gene transfer

Duchenne muscular dystrophy is a fatal muscle-wasting disorder. Lack of dystrophin compromises the integrity of the sarcolemma and results in myofibers that are highly prone to contraction-induced injury. Recombinant adenoassociated virus (rAAV)-mediated dystrophin gene transfer strategies to muscle for the treatment of Duchenne muscular dystrophy (DMD) have been limited by the small cloning capacity of rAAV vectors and high titers necessary to achieve efficient systemic gene transfer. In this study, we assess the impact of codon optimization on microdystrophin (ΔAB/R3-R18/ΔCT) expression and function in the mdx mouse and compare the function of two different configurations of codon-optimized microdystrophin genes (ΔAB/R3-R18/ΔCT and ΔR4-R23/ΔCT) under the control of a muscle-restrictive promoter (Spc5-12). Codon optimization of microdystrophin significantly increases levels of microdystrophin mRNA and protein after intramuscular and systemic administration of plasmid DNA or rAAV2/8. Physiological assessment demonstrates that codon optimization of ΔAB/R3-R18/ΔCT results in significant improvement in specific force, but does not improve resistance to eccentric contractions compared with noncodon-optimized ΔAB/ R3-R18/ΔCT. However, codon-optimized microdystrophin ΔR4-R23/ΔCT completely restored specific force generation and provided substantial protection from contraction-induced injury. These results demonstrate that codon optimization of microdystrophin under the control of a muscle-specific promoter can significantly improve expression levels such that reduced titers of rAAV vectors will be required for efficient systemic administration.

In-situ comparison of the NOy instruments flown in MOZAIC and SPURT

Two aircraft instruments for the measurement of total odd nitrogen (NOy) were compared side by side aboard a Learjet A35 in April 2003 during a campaign of the AFO2000 project SPURT (Spurengastransport in der Tropopausenregion). The instruments albeit employing the same measurement principle (gold converter and chemiluminescence) had different inlet configurations. The ECO-Physics instrument operated by ETH-Zürich in SPURT had the gold converter mounted outside the aircraft, whereas the instrument operated by FZ-Jülich in the European project MOZAIC III (Measurements of ozone, water vapour, carbon monoxide and nitrogen oxides aboard Airbus A340 in-service aircraft) employed a Rosemount probe with 80 cm of FEP-tubing connecting the inlet to the gold converter. The NOy concentrations during the flight ranged between 0.3 and 3 ppb. The two data sets were compared in a blind fashion and each team followed its normal operating procedures. On average, the measurements agreed within 7%, i.e. within the combined uncertainty of the two instruments. This puts an upper limit on potential losses of HNO3 in the Rosemount inlet of the MOZAIC instrument. Larger transient deviations were observed during periods after calibrations and when the aircraft entered the stratosphere. The time lag of the MOZAIC instrument observed in these instances is in accordance with the time constant of the MOZAIC inlet line determined in the laboratory for HNO3.

Human microbiota in health and disease

Each human body plays host to a microbial population which is both numerically vast (at around 1014 microbial cells) and phenomenally diverse (over 1,000 species). The majority of the microbial species in the gut have not been cultured but the application of culture-independent approaches for high throughput diversity and functionality analysis has allowed characterisation of the diverse microbial phylotypes present in health and disease. Studies in monozygotic twins, showing that these retain highly similar microbiota decades after birth and initial colonisation, are strongly indicative that diversity of the microbiome is host-specific and affected by the genotype. Microbial diversity in the human body is reflected in both richness and evenness. Diversity increases steeply from birth reaching its highest point in early adulthood, before declining in older age. However, in healthy subjects there appears to be a core of microbial phylotypes which remains relatively stable over time. Studies of individuals from diverse geopraphies suggest that clusters of intestinal bacterial groups tend to occur together, constituting ‘enterotypes’. So variation in intestinal microbiota is stratified rather than continuous and there may be a limited number of host/microbial states which respond differently to environmental influences. Exploration of enterotypes and functional groups may provide biomarkers for disease and insights into the potential for new treatments based on manipulation of the microbiome. In health, the microbiota interact with host defences and exist in harmonious homeostasis which can then be disturbed by invading organisms or when ‘carpet bombing’ by antibiotics occurs. In a portion of individuals with infections, the disease will resolve itself without the need for antibiotics and microbial homeostasis with the host’s defences is restored. The administration of probiotics (live microorganisms which when administered in adequate amounts confer a health benefit on the host) represents an artificial way to enhance or stimulate these natural processes. The study of innate mechanisms of antimicrobial defence on the skin, including the production of numerous antimicrobial peptides (AMPs), has shown an important role for skin commensal organisms. These organisms may produce AMPs, and also amplify the innate immune responses to pathogens by activating signalling pathways and processing host produced AMPs. Research continues into how to enhance and manipulate the role of commensal organisms on the skin. The challenges of skin infection (including diseases caused by multiply resistant organisms) and infestations remain considerable. The potential to re-colonise the skin to replace or reduce pathogens, and exploring the relationship between microbiota elsewhere and skin diseases are among a growing list of research targets. Lactobacillus species are among the best known ‘beneficial’ bacterial members of the human microbiota. Of the approximately 120 species known, about 15 are known to occur in the human vagina. These organisms have multiple properties, including the production of lactic acid, hydrogen peroxide and bacteriocins, which render the vagina inhospitable to potential pathogens. Depletion of the of the normal Lactobacillus population and overgrowth of vaginal anaerobes, accompanied by the loss of normal vaginal acidity can lead to bacterial vaginosis – the commonest cause of abnormal vaginal discharge in women. Some vaginal anaerobes are associated with the formation of vaginal biofilms which serve to act as a reservoir of organisms which persists after standard antibiotic therapy of bacterial vaginosis and may help to account for the characteristically high relapse rate in the condition. Administration of Lactobacillus species both vaginally and orally have shown beneficial effects in the treatment of bacterial vaginosis and such treatments have an excellent overall safety record. Candida albicans is a frequent coloniser of human skin and mucosal membranes, and is a normal part of the microbiota in the mouth, gut and vagina. Nevertheless Candida albicans is the most common fungal pathogen worldwide and is a leading cause of serious and often fatal nosocomial infections. What turns this organism from a commensal to a pathogen is a combination of increasing virulence in the organism and predisposing host factors that compromise immunity. There has been considerable research into the use of probiotic Lactobacillus spp. in vaginal candidiasis. Studies in reconstituted human epithelium and monolayer cell cultures have shown that L. rhamnosus GG can protect mucosa from damage caused by Candida albicans, and enhance the immune responses of mucosal surfaces. Such findings offer the promise that the use of such probiotic bacteria could provide new options for antifungal therapy. Studies of changes of the human intestinal microbiota in health and disease are complicated by its size and diversity. The Alimentary Pharmabiotic Centre in Cork (Republic of Ireland) has the mission to ‘mine microbes for mankind’ and its work illustrates the potential benefits of understanding the gut microbiota. Work undertaken at the centre includes: mapping changes in the microbiota with age; studies of the interaction between the microbiota and the gut; potential interactions between the gut microbiota and the central nervous system; the potential for probiotics to act as anti-infectives including through the production of bacteriocins; and the characterisation of interactions between gut microbiota and bile acids which have important roles as signalling molecules and in immunity. The important disease entity where the role of the gut microbiota appears to be central is the Irritable Bowel Syndrome (IBS). IBS patients show evidence of immune activation, impaired gut barrier function and abnormal gut microbiota. Studies with probiotics have shown that these organisms can exert anti-inflammatory effects in inflammatory bowel disease and may strengthen the gut barrier in IBS of the diarrhoea-predominant type. Formal randomised trials of probiotics in IBS show mixed results with limited benefit for some but not all. Studies confirm that administered probiotics can survive and temporarily colonise the gut. They can also stimulate the numbers of other lactic acid bacilli in the gut, and reduce the numbers of pathogens. However consuming live organisms is not the only way to influence gut microbiota. Dietary prebiotics are selectively fermented ingredients that can change the composition and/or activity of the gastrointestinal microbiota in beneficial ways. Dietary components that reach the colon, and are available to influence the microbiota include poorly digestible carbohydrates, such as non-starch polysaccharides, resistant starch, non-digestible oligosaccharides (NDOs) and polyphenols. Mixtures of probiotic and prebiotic ingredients that can selectively stimulate growth or activity of health promoting bacteria have been termed ‘synbiotics’. All of these approaches can influence gut microbial ecology, mainly to increase bifidobacteria and lactobacilli, but metagenomic approaches may reveal wider effects. Characterising how these changes produce physiological benefits may enable broader use of these tactics in health and disease in the future. The current status of probiotic products commercially available worldwide is less than ideal. Prevalent problems include misidentification of ingredient organisms and poor viability of probiotic microorganisms leading to inadequate shelf life. On occasions these problems mean that some commercially available products cannot be considered to meet the definition of a probiotic product. Given the potential benefits of manipulating the human microbiota for beneficial effects, there is a clear need for improved regulation of probiotics. The potential importance of the human microbiota cannot be overstated. ‘We feed our microbes, they talk to us and we benefit. We just have to understand and then exploit this.’ (Willem de Vos).

Detection and molecular characterisation of Pyrenopeziza brassicae isolates resistant to methyl benzimidazole carbamates

BACKGROUND Methyl benzimidazole carbamate (MBC) fungicides are used to control the oilseed rape pathogen Pyrenopeziza brassicae. Resistance to MBCs has been reported in P. brassicae, but the molecular mechanism(s) associated with reductions in sensitivity have not been verified in this species. Elucidation of the genetic changes responsible for resistance, hypothesised to be target-site mutations in β-tubulin, will enable resistance diagnostics and thereby inform resistance management strategies. RESULTS P. brassicae isolates were classified as sensitive, moderately resistant or resistant to MBCs. Crossing P. brassicae isolates of different MBC sensitivities indicated that resistance was conferred by a single gene. The MBC-target encoding gene β-tubulin was cloned and sequenced. Reduced MBC sensitivity of field isolates correlated with β-tubulin amino acid substitutions L240F and E198A. The highest level of MBC resistance was measured for isolates carrying E198A. Negative cross-resistance between MBCs and the fungicides diethofencarb and zoxamide was only measured in E198A isolates. PCR-RFLP was used to screen isolates for the presence of L240F and E198A. The substitutions E198G and F200Y were also detected in DNA samples from P. brassicae populations after cloning and sequencing of PCR products. The frequencies of L240F and E198A in different P. brassicae populations were quantified by pyrosequencing. There were no differences in the frequencies of these alleles between P. brassicae populations sampled from different locations or after fungicide treatment regimes. CONCLUSIONS The molecular mechanisms affecting sensitivity to MBCs in P. brassicae have been identified. Pyrosequencing assays are a powerful tool for quantifying fungicide-resistant alleles in pathogen populations.

Promoting mother–infant book sharing and infant attention and language development in an impoverished South African population: a pilot study

The low rates of child literacy in South Africa are cause for considerable concern. Research from the developed world shows that parental sharing of picture books with infants and young children is beneficial for child language and cognitive development, as well as literacy skills. We conducted a pilot study to examine whether such benefits might extend to an impoverished community in South Africa, by evaluating the impact of training mothers in book sharing with their 14–18 month old infants. Seventeen mothers received book sharing training; and 13 mothers did not, but instead received a comparison training in toy play. We assessed the mothers’ behavior during both book sharing and toy play before and after training, and we also assessed infant attention and language. Mothers receiving book sharing training engaged well with it, and they also benefited from it; thus, compared to the comparison group mothers, they became more sensitive, more facilitating, and more elaborative with their infants during book sharing, and they also became more sensitive to their infants during toy play. In addition, infants whose mothers received the book sharing training showed greater benefits than the comparison group infants in both their attention and language. Training in book sharing for families living in conditions of marked socio-economic adversity in South Africa has the potential to be of considerable benefit to child developmental progress. A large scale controlled trial is required to confirm this.

Embedding arts and humanities in the creative economy: the role of graduates in the UK

The recent change in funding structure in the UK higher education system has fuelled an animated debate about the role that arts and humanities (A&H) subjects play not only within higher education but more broadly in the society and the economy. The debate has engaged with a variety of arguments and perspectives, from the intrinsic value of A&H, to their contribution to the broader society and their economic impact, particularly in relation to the creative economy, through knowledge exchange activities. The paper argues that in the current debate very little attention has been placed on the role that A&H graduates play in the economy, through their work after graduation, and specifically in the creative economy. Using Higher Education Statistical Agency data, we analyse the performance of A&H graduates (compared with other graduates) and particularly explore how embedded they are with the creative economy and its associated industries. The results highlight a complex intersection of different subdisciplines of the A&H with the creative economy but also reveal the salary gap and unstable working conditions experienced by graduates in this field.

Ethics of care in supporting disabled forced migrants: interactions with professionals and ethical dilemmas in health and social care in the South-East of England

This article explores the interactions between disabled forced migrants with care needs and professionals and the restrictive legal, policy and practice context that health and social care professionals have to confront, based on the findings of a qualitative study with 45 participants in the South-East of England. In-depth interviews were conducted with 15 forced migrants who had diverse impairments and chronic illnesses (8 women and 7 men), 13 family caregivers and 17 support workers and strategic professionals working in social care and the third sector in Slough, Reading and London. The legal status of forced migrants significantly affects their entitlements to health and social care provision, resulting in prolonged periods of destitution for many families. National asylum support policies, difficult working relationships with UK Border Agency, higher eligibility thresholds and reduced social care budgets of local authorities were identified as significant barriers in responding to the support needs of disabled forced migrants and family caregivers. In this context, social workers experienced considerable ethical dilemmas. The research raises profound questions about the potential and limitations of health and social care policies, provision, and practice as means of protection and support in fulfilling the human rights of forced migrants with care needs.

Preliminary ethnoarchaeological research on modern animal husbandry in Bestansur, Iraqi Kurdistan: integrating animal, plant and environmental data

This paper presents preliminary results from an ethnoarchaeological study of animal husbandry in the modern village of Bestansur, situated in the lower Zagros Mountains of Iraqi Kurdistan. This research explores how modern families use and manage their livestock within the local landscape and identifies traces of this use. The aim is to provide the groundwork for future archaeological investigations focusing on the nearby Neolithic site of Bestansur. This is based on the premise that modern behaviours can suggest testable patterns for past practices within the same functional and ecological domains. Semi-structured interviews conducted with villagers from several households provided large amounts of information on modern behaviours that helped direct data collection, and which also illustrate notable shifts in practices and use of the local landscape over time. Strontium isotope analysis of modern plant material demonstrates that a measurable variation exists between the alluvial floodplain and the lower foothills, while analysis of modern dung samples shows clear variation between sheep/goat and cow dung, in terms of numbers of faecal spherulites. These results are specific to the local environment of Bestansur and can be used for evaluating and contextualising archaeological evidence as well as providing modern reference material for comparative purposes.

An in vitro study of the effect of probiotics, prebiotics and synbiotics on the elderly faecal microbiota

The use of dietary intervention in the elderly in order to beneficially modulate their gut microbiota has not been extensively studied. The influence of two probiotics (Bifidobacterium longum and Lactobacillus fermentum) and two prebiotics [isomaltooligosaccharides (IMO) and short-chain fructooligosaccharides (FOS)], individually and in synbiotic combinations (B. longum with IMO, L. fermentum with FOS) on the gut microbiota of elderly individuals was investigated using faecal batch cultures and three-stage continuous culture systems. Population changes of major bacterial groups were enumerated using fluorescent in situ hybridisation (FISH). B. longum and IMO alone significantly increased the Bifidobacterium count after 5 and 10 h of fermentation and their synbiotic combination significantly decreased the Bacteroides count after 5 h of fermentation. L. fermentum and FOS alone significantly increased the Bifidobacterium count after 10 h and 5, 10 and 24 h of fermentation respectively. B. longum with IMO as well as B. longum and IMO alone significantly increased acetic acid concentration during the fermentation in batch cultures. In the three-stage continuous culture systems, both synbiotic combinations increased the Bifidobacterium and Lactobacillus count in the third vessel representing the distal colon. In addition, the synbiotic combination of L. fermentum with scFOS resulted in a significant increase in the concentration of acetic acid. The results show that the elderly gut microbiota can be modulated in vitro with the appropriate pro-, pre- and synbiotics.