The genotype-phenotype landscape of familial amyotrophic lateral sclerosis in Australia.

Amyotrophic lateral sclerosis (ALS) is a clinically and genetically heterogeneous fatal neurodegenerative disease. Around 10% of ALS cases are hereditary. ALS gene discoveries have provided most of our understanding of disease pathogenesis. We aimed to describe the genetic landscape of ALS in Australia by assessing 1013 Australian ALS patients for known ALS mutations by direct sequencing, whole exome sequencing or repeat primed polymerase chain reaction. Age of disease onset and disease duration were used for genotype-phenotype correlations. We report 60.8% of Australian ALS families in this cohort harbour a known ALS mutation. Hexanucleotide repeat expansions in C9orf72 accounted for 40.6% of families and 2.9% of sporadic patients. We also report ALS families with mutations in SOD1 (13.7%), FUS (2.4%), TARDBP (1.9%), UBQLN2 (.9%), OPTN (.5%), TBK1 (.5%) and CCNF (.5%). We present genotype-phenotype correlations between these genes as well as between gene mutations. Notably, C9orf72 hexanucleotide repeat expansion positive patients experienced significantly later disease onset than ALS mutation patients. Among SOD1 families, p.I114T positive patients had significantly later onset and longer survival. Our report highlights a unique spectrum of ALS gene frequencies among patients from the Australian population, and further, provides correlations between specific ALS mutations with disease onset and/or duration.

Precision of total and regional body fat estimates from dual-energy X-ray absorptiometer measurements.

OBJECTIVES: To evaluate the precision of both total %fat and all the regional %fat measures acquired from both the Lunar Prodigy and Lunar iDXA software. DESIGN: Cross-sectional study. SETTING: University-based research laboratory. PARTICIPANTS: The sample consisted of 300 individual test records from men and women who had volunteered to participate in dual-energy x-ray absorptiometer (DXA) technician precision training studies. Subjects ranged in age from 20-84 years and in body mass index from 15.7-52.0 kg.m-2. MEASUREMENTS: A total of 27 different technicians performed three total body scan measurements on 10-15 different subjects. The Lunar Prodigy and Lunar iDXA were used for 253 and 47 precision training evaluations, respectively. The regions of interest (ROI) were automatically determined by the enCORE software (autoROI) for total body, android, gynoid, trunk, legs, and arms regions and the region %fat data were used for analyses. RESULTS: The CV for total body %fat was 1.9% and 0.9% for the Prodigy and iDXA, respectively. CV's for %fat measures at regional sites ranged from 1.2-4.4% for the Prodigy measures and 0.9-2.4% for the iDXA measures. The ICC for both devices ranged from 0.990 to 0.999. CONCLUSION: Monitoring the status of body composition changes with age is gaining more clinical acceptance. Thus, it is important that practitioners use measures that are both precise and accurate. The findings from the current study add support that DXA measurements can be used with a high level of confidence for serial testing of patients.

Overexpression of Batf induces an apoptotic defect and an associated lymphoproliferative disorder in mice.

Activator protein-1 (AP-1) is a dimeric transcription factor composed of the Jun, Fos and Atf families of proteins. Batf is expressed in the immune system and participates in AP-1 dimers that modulate gene expression in response to a variety of stimuli. Transgenic (Tg) mice overexpressing human BATF in T cells were generated using the human CD2 promoter (CD2-HA (hemagglutinin antigen) - BATF). By 1 year of age, over 90% of the mice developed a lymphoproliferative disorder (LPD). The enlarged lymph nodes characteristic of this LPD contain a polyclonal accumulation of T cells with a CD4(+) bias, yet efforts to propagate these tumor cells in vitro demonstrate that they do not proliferate as well as wild-type CD4(+) T cells. Instead, the accumulation of these cells is likely due to an apoptotic defect as CD2-HA-BATF Tg T cells challenged by trophic factor withdrawal in vitro resist apoptosis and display a pro-survival pattern of Bcl-2 family protein expression. As elevated levels of Batf expression are a feature of lymphoid tumors in both humans and mice, these observations support the use of CD2-HA-BATF mice as a model for investigating the molecular details of apoptotic dysregulation in LPD.

Post-exercise cardiac troponin release is related to exercise training history.

Marathon running can increase circulating cardiac troponin above the diagnostic criteria for myocardial infarction. We determined whether prior-exercise experience (training history) might be related to the magnitude of immediate post-race troponin release following completion of the London Marathon in a group of non-elite runners. Using a prospective study design, 52 runners were recruited into either HIGH T-E (trained-experience) (n=27) or LOW T-E (n=25) groups. Cardiac troponin I (cTnI) release following race completion was compared between these 2 groups. To examine relationships between cTnI release and participant demographic and indices of prior training experience an additional 52 runners who did not meet the criteria for either the HIGH T-E or LOW T-E groups were also recruited. The combined data from all 104 runners was analysed using multivariate linear regression analysis. The results revealed a significant difference in post marathon circulating cTnI between LOW T-E runners (median: 0.11 µg/L; interquartile range [IQR]: 0.03-0.18 µg/L) and HIGH T-E runners (median: 0.03 µg/L; IQR 0.02-0.057 µg/L) (p<0.05). Average miles run per week in the last 3 years, a marker of total training experience, encompassing training volume and duration, was negatively associated with post-marathon cTnI release (p<0.001).In conclusion, exercise-induced cTnI release is strongly related to previous training experience.

Exped 2010--the challenges of an expedition doctor.

'Exped 2010', a climbing and trekking expedition to the Everest region of Nepal, marked the 150th anniversary of the Army Cadet Force, during October 2010. A team of 20 cadets aged 14-18 years together with a combination of adults, mountain leaders and the expedition doctor, completed the trek to Everest Base Camp with a small number summiting Island Peak. The junior team, consisting of younger members aged 14-16 years, were limited to sleeping at a maximum altitude of 4240m but nevertheless, achieved a trekking altitude of 5000m. Over the three week period, there were a number of challenges for the expedition doctor--integrating into the team, finding time to rest and acclimatise along with the others whilst dealing with over 35 consultations, numerous cases of traveller's diarrhoea, several helicopter evacuations and also managing cases of altitude illness both within the group and for several foreign trekkers along the way.

Deposition of Atlantic salmon Salmo salar carcasses in a Scottish upland catchment.

Radio telemetry was utilized to track 38 Atlantic salmon Salmo salar across space and time during and following their spawning run on the Conon system, Scotland. The data collected were used to assess the proportion of S. salar able to migrate successfully from an upland oligotrophic area of the catchment and the distribution of the carcasses of those fish that remained. Of these fish, 35% successfully migrated from the study area after the spawning period. The fish that remained were distributed approximately equally between riverine and lacustrine areas, but with a strong tendency to accumulate in regions of relatively slow water flows.

Energy expenditure in the Race Across America (RAAM).

Energy Expenditure was measured with doubly labelled water technique during heavy sustained exercise with an official finishing team in the Race Across America. Energy Intake was also calculated to produce an energy balance for the race. A team of 4 cyclists (Mean +/-SD age: 37+4 yr; body height: 182+8 cm; body mass: 80.8+6.6 kg) completed the race in a relay fashion. The team completed the race in 6 days 10 h and 51 min. Total mean energy expenditure was found to be 43,401 kcals (181,711 kJ) with a mean daily energy expenditure of 6,420 kcals (26,879 kJ). Total mean energy intake from all food and drink consumed was calculated at 29,506 kcals (123,536 kJ) with a mean daily energy intake of 4 918 kcals (20,591 kJ). This resulted in a total mean energy deficit of 13,878 kcals (58,104 kJ) with a mean daily energy deficit of 1,503 kcals (6,293 kJ). The high energy expenditure highlights the need for correct and practical dietary strategies and challenges nutritionists to devise high energy diets that not only contain the correct macronutrient balance, but are also palatable to the cyclists, thus encouraging a high energy intake.

Enhanced anti-HIV functional activity associated with Gag-specific CD8 T-cell responses.

Effective HIV-specific T-cell immunity requires the ability to inhibit virus replication in the infected host, but the functional characteristics of cells able to mediate this effect are not well defined. Since Gag-specific CD8 T cells have repeatedly been associated with lower viremia, we examined the influence of Gag specificity on the ability of unstimulated CD8 T cells from chronically infected persons to inhibit virus replication in autologous CD4 T cells. Persons with broad (>or=6; n = 13) or narrow (

A novel TARDBP mutation in an Australian amyotrophic lateral sclerosis kindred.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder that causes loss of motor neurons. A pathological hallmark of ALS is the presence of ubiquitinated TAR DNA binding protein (TDP-43) inclusions in the cytoplasm of affected cells. Rare pathogenic mutations within the gene TARDBP that encode TDP-43 were recently reported in ALS but their functional consequences are unknown. To further investigate the pathogenic role of TDP-43 in ALS, a mutation analysis of TARDBP was performed in an Australian cohort of 74 sporadic and 30 familial ALS cases. A novel familial ALS mutation in TDP-43 was identified that substitutes a highly conserved residue (G294V) and is predicted to disrupt the glycine rich domain in the C terminus, a region that plays a role in RNA binding and is required for the exon skipping activity of TDP-43.

Identification of novel genes and transcription factors involved in spleen, thymus and immunological development and function.

We constructed and analyzed six serial analysis of gene expression (SAGE) libraries to identify genes with previously uncharacterized roles in spleen or thymus development. A total of 625 070 tags were sequenced from the three spleen (embryonic day (E)15.5, E16.5 and adult) and three thymus (E15.5, E18.5 and adult) libraries. These tags corresponded to 83 182 tag types, which mapped unambiguously to 36 133 different genes. Genes over-represented in these libraries, compared to 115 mouse SAGE libraries (www.mouseatlas.org), included genes of known and unknown immunological or developmental relevance. The expression profiles of 11 genes with unknown roles in spleen and thymus development were validated using reverse transcription-qPCR. We further characterized the expression of one of these candidates, RIKEN cDNA 9230105E10 that encodes a murine homolog of Trim5alpha, in numerous adult tissues and immune cell types. In addition, we demonstrate that transcript levels are upregulated in response to TLR stimulation of plasmacytoid dendritic cells and macrophages. This work provides the first evidence of regulated and cell type-specific expression of this gene. In addition, these observations suggest that the SAGE libraries provide an important resource for further investigations into the molecular mechanisms regulating spleen and thymus organogenesis, as well as the development of immunological competence.

Effects on respiratory health of a reduction in air pollution from vehicle exhaust emissions.

AIMS: To determine whether residents of congested streets have a higher prevalence of respiratory symptoms than residents of nearby uncongested streets, and whether their respiratory health improves following a reduction in exposure to traffic related air pollutants. METHODS: An area was identified where certain streets were subject to air pollution from heavy road traffic, which was likely to improve following the construction of a by-pass. A respiratory survey was conducted among the residents, together with the residents of nearby uncongested streets, at baseline and again a year after the by-pass opened. Measurements were made of air pollutant concentrations in both areas on both occasions. RESULTS: Initial concentrations of PM10 and PM2.5 were substantially higher in the congested than in the uncongested streets. When the by-pass opened, the volume of heavy goods traffic fell by nearly 50%. PM10 decreased by 23% (8.0 microg/m3) in the congested streets and by 29% (3.4 microg/m3) in the uncongested streets, with similar proportionate falls in PM2.5. There were no clear or consistent differences between the residents of the two areas initially in terms of symptoms or peak flow variability. Repeat questionnaires were obtained from 165 and 283 subjects in the congested and uncongested areas respectively, and showed a tendency for most symptoms to improve in both areas. For chest symptoms, the improvement tended to be greater in the uncongested area, although the difference between the areas was not statistically significant. Rhinitis and rhinoconjunctivitis tended to improve to a greater extent in the congested streets; the difference between the areas was significant for the degree to which rhinitis interfered with daily activities. Peak flow variability tended to improve in the uncongested area. CONCLUSIONS: The by-pass reduced pollutant levels to a degree that probably alleviates rhinitis and rhinoconjunctivitis but has little effect on lower respiratory symptoms.

Phylogeography of the coronulid barnacle, Chelonibia testudinaria, from loggerhead sea turtles, Caretta caretta.

The barnacle, Chelonibia testudinaria, is a common inhabitant of the marine turtle epibiont community and plays a key role in the development of this community. Phylogeographic analysis of 79 cytochrome c oxidase I (COX1) sequences for barnacles collected from five populations found contrasting patterns of divergence for populations in the Atlantic vs. the Pacific Ocean. Our analysis indicates that the two Pacific populations, Senri Beach, Japan and Bahia Magdalena, Mexico, were not only highly divergent from the Atlantic populations but are highly divergent from one another. We suggest that barnacles from these populations may represent cryptic species. In contrast, sequence divergence was greatly reduced among barnacles collected from Wassaw Island, GA, USA, Keewaydin, FL, USA, and Kyparissia, Pèloponnésus Island, Greece. A reduction in sequence diversity at the latter site was attributed to a recent range expansion into the Mediterranean Sea. We examined historical patterns of migration among the three Atlantic and Mediterranean populations using the program migrate. This analysis indicates a high rate of migration from Keewaydin to Wassaw Island, contrasted with a much lower rate of migration in the opposite direction. The estimated migration rate from Kyparissia to Keewaydin was also non-negligible. We suggest that the association between C. testudinaria and loggerhead turtles and the patterns of turtle migration have played key roles in the expansion of the range of C. testudinaria into the Mediterranean Sea and the subsequent patterns of barnacle migration. We further propose that the difference between ocean basins, with respect to the impact of host migration on barnacle gene flow, probably stems from the fact that host-mediated dispersal in the Atlantic depends on advanced stage juveniles and adults while any host-mediated dispersal in the Pacific would have to involve early "pelagic" stage juvenile loggerheads.

Effect of opioid receptor antagonists on hypothalamic-pituitary-adrenal activity in rhesus monkeys.

Some opioid antagonists increase the release of adrenocorticotropic hormone (ACTH) and cortisol in humans and, therefore, may indicate that endogenous opioids modulate hypothalamic-pituitary-adrenal axis activity. The type of opioid receptor that may be related to these endocrine effects is unknown. The purpose of this experiment was to evaluate the ability of different opioid antagonists to increase ACTH and cortisol plasma levels in rhesus monkeys. Eight monkeys received intramuscular injections of various antagonists: 0.0032-1.0 mg/kg naltrexone, 0.1-3.2 mg/kg naltrindole (delta-selective), 0.032-0.32 mg/kg clocinnamox (mu-selective), and 1-3.2 mg/kg nor-binaltorphimine (kappa-selective). Naltrexone, 0.1-1.0 mg/kg, increased ACTH levels, whereas naltrindole and clocinnamox failed to increase ACTH levels. Nor-binaltorphimine, 1-3.2 mg/kg, increased ACTH concentrations on the day of injection, but not at a time when other assays continue to demonstrate kappa-antagonism (24 h). Cortisol concentrations generally followed the same pattern as the ACTH concentrations, but the incremental differences in cortisol release between doses were less clear. Thus, opioid modulation of ACTH and cortisol plasma levels is not clearly associated with a particular opioid receptor. Although the kappa-antagonist increased ACTH and cortisol release on the day of injection, some evidence suggests that this endocrine effect may be due to mechanisms other than those mediated by the kappa-receptor. Alternatively, the naltrexone-induced increase of ACTH and cortisol plasma levels may be caused by activity at multiple opioid receptors or some uncharacterized receptor. Finally, the increased release of ACTH and cortisol may be a response to naltrexone's aversive properties.

Ultrastructure and properties of Paecilomyces lilacinus spores.

Strains of the filamentous soil fungus Paecilomyces lilacinus are currently being developed for use as biological control agents against root-knot, cyst, and other plant-parasitic nematodes. The inoculum applied in the field consists mainly of spores. This study was undertaken to examine the size, ultrastructure, and rodlet layers of P. lilacinus spores and the effect of the culture method on structural and functional spore properties. A rodlet layer was identified on aerial spores only. Other differences noted between aerial spores and those produced in submerged culture included the size and appearance of spores and thickness of spore coat layers when examined with transmission electron microscopy. The two spore types differed in UV tolerance, with aerial spores being less sensitive to environmentally relevant UV radiation. Also, viability after drying and storage was better with the aerial spores. Both spore types exhibited similar nematophagous ability.

Bias in risk estimation: application to Down's syndrome screening.

In this paper we consider the bias associated with parametric estimation of a univariate or bivariate Gaussian density, and also the induced bias when these Gaussian densities are used to determine a likelihood ratio. Algebraic approximations are derived that accurately predict the relative biases obtained, verification being achieved by a simulation exercise. The expressions confirm that when estimating a univariate Gaussian density there are four Z-scores for which there is zero bias and that relative bias increases rapidly beyond two standard deviations from the mean. The results are then extended to determine approximate confidence intervals for both the true density and the likelihood ratio. A simulation exercise confirms that the derived 95 per cent confidence intervals have coverage that ranges from 94 to 97 per cent. The results are applied to a Down's syndrome screening programme where 95 per cent confidence intervals are established for a woman's posterior odds of carrying a Down's foetus. It is shown that patients with similar posterior odds can give rise to confidence intervals for their true posterior odds that have very different widths, thus emphasizing that not all risks are of equal quality.

Factors associated with asthma in Albania.

BACKGROUND: This study was undertaken to see whether asthma in Albania (where it is particularly uncommon) is associated with certain 'western' characteristics, and with dietary and lifestyle factors that affect its occurrence elsewhere. METHODS: A screening survey was conducted among 2653 persons aged 20-44 years. A more detailed enquiry was conducted among a random subsample plus all others whose response indicated possible asthma. Three groups were compared: 495 probable nonasthmatics ('nonsymptomatics'); 106 with asthma-like symptoms and positive skin tests ('possible allergic asthmatics'); and 116 with similar symptoms and negative skin tests ('other symptomatics'). RESULTS: The possible allergic asthmatics were less likely than the nonsymptomatics to have attended school or nursery under the age of 5 years or to have shared a bedroom with an older child before that age. Continued education after the age of 18 years also appeared to be protective. Allowing for other factors, a strong negative relation was found with fruit and vegetable consumption, and weak positive relations with prepackaged food and fizzy drinks. Symptoms in skin prick test-negative subjects were associated with personal and parental smoking habit. CONCLUSIONS: This survey provides limited evidence linking asthma with aspects of western lifestyle. It supports the hypothesis that opportunities for infection during early childhood and the consumption of fruit and vegetables protect against asthma.

Prevention of venous thromboembolism in Wales: results of a survey among general surgeons.

OBJECTIVE: To examine the current attitudes towards the prevention of venous thromboembolism among a cohort of surgeons. DESIGN: A postal survey, comprising a questionnaire covering various aspects of venous thromboembolism prophylaxis was sent to all (n=84) consultant general surgeons in Wales. RESULTS: Replies were received from 57 surgeons (68%), all of whom routinely used prophylaxis, the most frequent modalities used being heparin (100%) and graded compression stockings (79%). A combination of physical and pharmacological methods was used by over 89% of surgeons, with 60% starting prophylaxis more than two hours before operation. All surgeons continued prophylaxis after surgery, 53% until patients were mobile, 45% until they were discharged, and one surgeon continued prophylaxis for seven days after discharge. The thrombosis risk factors considered most important by surgeons when deciding about prophylaxis were (i) a previous history of venous thromboembolism, (ii) hypercoagulability, and (iii) malignancy. CONCLUSIONS: This study confirms that Welsh surgeons conform to standard methods, but also highlights some uncertainties that are present in current surgical practice. Those who responded all routinely used prophylaxis, the timing of which was variable. The main risk factors identified when considering prophylaxis were previous history of deep vein thrombosis/pulmonary embolism, hypercoagulability, and the presence of malignancy. Suggestions for future practice are made.

Interaction of morphine and naltrexone on oral ethanol self-administration in rhesus monkeys.

Opioid antagonists, such as naltrexone (NTX), reduce ethanol consumption and opioid agonists increase or decrease ethanol consumption in rats depending upon the dose. If the opioid antagonist and agonist effects on ethanol consumption are mediated by mu-opioid receptors, then NTX doses that reduce ethanol consumption should be similar to the doses necessary to antagonize the effects of opioid agonists on ethanol consumption. The purpose of these experiments was: (1) to determine whether morphine increases ethanol consumption in rhesus monkeys as it does in rodents; (2) to determine if the mu-receptor mediates the effects of morphine on ethanol consumption by conducting a pK(B) analysis using NTX; and (3) to determine if the mu-receptor also mediates the NTX-induced decreases in ethanol consumption by making comparisons between the NTX doses that affect ethanol consumption and the NTX doses that block the effects of morphine on ethanol consumption. Three male rhesus monkeys responded for 2% ethanol and water for 2 h/day on a fixed-ratio 4 schedule of reinforcement. Morphine doses as low as 0.0032mg/kg failed to increase ethanol fluid deliveries, whereas higher doses produced a dose-related decrease in ethanol fluid deliveries. Although 0.01 mg/kg NTX alone had no effect on ethanol fluid deliveries, it reduced the suppressant effects of morphine with a mu-receptor pK(B) of 8.21 (8.08-8.34). When given alone, 0.1 mg/kg NTX decreased ethanol fluid deliveries but failed to reverse the suppression caused by 1 mg/kg morphine. Therefore, monkeys may differ from rats in their response to morphine when ethanol consumption is the dependent variable. Furthermore, because the NTX dose that reduced the effects of morphine on responding for ethanol was smaller than the NTX doses that suppressed ethanol-reinforced responding when given alone, NTX may exert these two effects through different mechanisms.

Cellulose-binding modules from extracellular matrix proteins of Dictyostelium discoideum stalk and sheath.

Cellulose-binding modules (CBMs) of two extracellular matrix proteins, St15 and ShD, from the slime mold Dictyostelium discoideum were expressed in Escherichia coli. The expressed proteins were purified to > 98% purity by extracting inclusion bodies at pH 11.5 and refolding proteins at pH 7.5. The two refolded CBMs bound tightly to amorphous phosphoric acid swollen cellulose (PASC), but had a low affinity toward xylan. Neither protein exhibited cellulase activity. St15, the stalk-specific protein, had fourfold higher binding affinity toward microcrystalline cellulose (Avicel) than the sheath-specific ShD CBM. St15 is unusual in that it consists of a solitary CBM homologous to family IIa CBMs. Sequence analysis of ShD reveals three putative domains containing: (a) a C-terminal CBM homologous to family IIb CBMs; (b) a Pro/Thr-rich linker domain; and (c) a N-terminal Cys-rich domain. The biological functions and potential role of St15 and ShD in building extracellular matrices during D. discoideum development are discussed.