Identification of genetic variants associated with Huntington's disease progression: a genome-wide association study

Background Huntington's disease is caused by a CAG repeat expansion in the huntingtin gene, HTT. Age at onset has been used as a quantitative phenotype in genetic analysis looking for Huntington's disease modifiers, but is hard to define and not always available. Therefore, we aimed to generate a novel measure of disease progression and to identify genetic markers associated with this progression measure. Methods We generated a progression score on the basis of principal component analysis of prospectively acquired longitudinal changes in motor, cognitive, and imaging measures in the 218 indivduals in the TRACK-HD cohort of Huntington's disease gene mutation carriers (data collected 2008–11). We generated a parallel progression score using data from 1773 previously genotyped participants from the European Huntington's Disease Network REGISTRY study of Huntington's disease mutation carriers (data collected 2003–13). We did a genome-wide association analyses in terms of progression for 216 TRACK-HD participants and 1773 REGISTRY participants, then a meta-analysis of these results was undertaken. Findings Longitudinal motor, cognitive, and imaging scores were correlated with each other in TRACK-HD participants, justifying use of a single, cross-domain measure of disease progression in both studies. The TRACK-HD and REGISTRY progression measures were correlated with each other (r=0·674), and with age at onset (TRACK-HD, r=0·315; REGISTRY, r=0·234). The meta-analysis of progression in TRACK-HD and REGISTRY gave a genome-wide significant signal (p=1·12 × 10−10) on chromosome 5 spanning three genes: MSH3, DHFR, and MTRNR2L2. The genes in this locus were associated with progression in TRACK-HD (MSH3 p=2·94 × 10−8 DHFR p=8·37 × 10−7 MTRNR2L2 p=2·15 × 10−9) and to a lesser extent in REGISTRY (MSH3 p=9·36 × 10−4 DHFR p=8·45 × 10−4 MTRNR2L2 p=1·20 × 10−3). The lead single nucleotide polymorphism (SNP) in TRACK-HD (rs557874766) was genome-wide significant in the meta-analysis (p=1·58 × 10−8), and encodes an aminoacid change (Pro67Ala) in MSH3. In TRACK-HD, each copy of the minor allele at this SNP was associated with a 0·4 units per year (95% CI 0·16–0·66) reduction in the rate of change of the Unified Huntington's Disease Rating Scale (UHDRS) Total Motor Score, and a reduction of 0·12 units per year (95% CI 0·06–0·18) in the rate of change of UHDRS Total Functional Capacity score. These associations remained significant after adjusting for age of onset. Interpretation The multidomain progression measure in TRACK-HD was associated with a functional variant that was genome-wide significant in our meta-analysis. The association in only 216 participants implies that the progression measure is a sensitive reflection of disease burden, that the effect size at this locus is large, or both. Knockout of Msh3 reduces somatic expansion in Huntington's disease mouse models, suggesting this mechanism as an area for future therapeutic investigation

Occurrence of pharmaceutically active compounds in surface waters of the Henares-Jarama-Tajo river system (Madrid, Spain) and a potential risk characterization

The Henares-Jarama-Tajo river system is the largest drainage basin in the Province of Madrid, Spain. This area is characterized by the presence of intensive urban and industrial activities influenced by a continental Mediterranean climate with rainfalls presenting substantial fluctuations along the different seasons. This research aimed to monitor seasonal variations in concentrations of 22 pharmaceutically active compounds (PhACs) in this river system and to establish the potential risk of sublethal effects on aquatic organisms. A total of 10 sampling sites were selected along the river system with samples collected in each of the four seasons during a year-round schedule. Most of the PhACs detected were present in sampling sites downstream in the vicinity of the most populated cities (i.e. Madrid, Guadalajara and Alcalá de Henares). Only two PhACs, fluoxetine and paraxantine, were detected in all sites regardless of the season, and showed median (± interquartile range) concentrations of 21.4 (± 31.2) ng L- 1 and 8.5 (± 5.3) ng L- 1, respectively. Other PhACs were detected with a frequency > 80% and included, caffeine, diphenylhydantoin, hydrochlorotiazide, ibuprofen, ketoprofen, diclofenac, sulfamethoxazole, atenolol, naproxen, carbamazepine and propanolol. Seasonal variations were observed with the highest concentrations in December and the lowest in September. By combining measured environmental concentrations with toxicity data (either publicly available or obtained experimentally in our laboratory), and by calculating an Maximum Risk Index (MaxRI) that each combination of PhACs should have for non exceeding the risk threshold, a high potential for long-term risk (MaxRI < 10) was estimated for most of the sampling sites and sampling dates. This research allowed the characterization of the potential risk for each of the PhACs to exert sublethal effects on aquatic organisms using acute screening methods, justifying the need for chronic data in order to refine the risk of these compounds to aquatic organisms. © 2009 Elsevier B.V. All rights reserved

Terrestrial microcosms in a feasibility study on the remediation of diesel-contaminated soils

Phytoremediation and bioremediation are site-specific processes, and feasibility studies should be performed as an initial step in scaling-up these processes. Soil microcosms can be a useful tool for investigating the efficiency of remediation alternatives under realistic conditions. In the present study, the degradation of diesel fuel in soil was studied in artificially assembled microcosms during a 180-day experiment. Microcosms consisted of soil columns seeded with a mix of Festuca arundinacea and Trifolium pratense into which earthworms were introduced and native soil microflora was used. Remediation assays were performed with different combinations of soil organisms' assemblages (micro-organisms, earthworms and plants) to determine the effects of these organisms and their interaction on diesel degradation. The microcosm system allowed for the study of the efficiency of remediation under field-relevant conditions. This system provided information about the dissipation of the pollutants as well as chemical leaching and possible toxic effects on the organisms during the remediation process. The decrease in soil hydrocarbon levels depended on the organisms' assemblage. Plants were not effective in the remediation of these soils despite their stimulation of microbial biomass. Conversely, earthworms had a beneficial impact on the dissipation of hydrocarbons that did not appear to be related to a generic improvement of plant or microbial activity. The grass species was more tolerant than the legume species to diesel fuel-contaminated soils. Leaching of hydrocarbons was negligible and independent of the organisms' assemblage. © 2011

Toxicity of the antimicrobial oxytetracycline to soil organisms in a multi-species-soil system (MS·3) and influence of manure co-addition

The effects of oxytetracycline (OTC) on soil organisms have been studied using a multi-species-soil system (MS·3). Oxytetracycline concentrations of 0.01 mg/kg, 1 mg/kg and 100 mg/kg soil were added to the 20 cm top arable soil layer, with and without horse/cow manure (0.15 g organic N/kg soil) co-addition. No mortality was observed for Eisenia foetida S. but significant effects on soil microbial enzymatic activities (phosphatase, dehydrogenase) were observed. The effects on soil microorganism were observed earlier but then recovered in systems with manure co-addition. More important, OTC related plant inhibition was observed in the manured but not in the non-manured systems. Oxytetracycline reached 0.19 and 1.85 mg/l in the leachate of the soil spiked with 1 and 100 mg OTC/kg, respectively and 0.05 and 1.14 mg/l for the same OTC concentrations in the manured systems. The results confirm that manure can modify both the fate and the effects of OTC and that the multi-species-soil systems can reproduce the conditions for a realistic effect estimation of veterinary medicines. © 2005 Elsevier B.V. All rights reserved

Ecotoxicological assessment of doxycycline in aged pig manure using multispecies soil systems

This paper assesses the ecotoxicity of the antibiotic doxycycline in aged spiked pig manure using a multispecies soil system (MS 3) covering plants, earthworms and soil microorganisms. The study reproduced realistic exposure conditions, as well as higher exposure doses covering the uncertainty factors typically employed for covering interspecies variability. MS 3, consisting of columns of natural sieved soil assembled with earthworms and seeds from three plant species, were employed. Pig manure was spiked with doxycycline (75 or 7500 μg/ml), aged for 15 days under aerobic/anaerobic conditions and added on top of the soil columns (120 ml/column, equivalent to 220 kgN/ha). Water and doxycycline free manure were used as negative controls. Doxycycline (7500 μg/ml) solution was used as a positive control. No effects on plants or earthworms were observed. Significant effects on soil phosphatase activity, indicating effects on soil microorganisms, were observed at the highest exposure dose, affecting all soil layers in the doxycycline-solution-treated MS 3 (positive control) but only the top layer in the spiked pig manure system. Chemical analysis confirmed the different behavior of doxycycline in both systems (with and without manure) and those effects were observed in soil with measured concentrations over 1 mg/kg soil. The detection of doxycycline in leachates revealed a potential mobility. Leachate concentrations were similar for doxycycline solution and spiked manure treatments. © 2003 Elsevier B.V. All rights reserved

Assessing the influence of biota on metal mobility in a multi-species soil system (MS·3)

Multi-species soil systems (MS·3) are homogeneous soil columns that allow a combined assessment of chemical fate and effects on representative soil organisms. Theoretically, the presence of organisms can modify the movement of chemicals in the soil core. This influence was studied for copper and cadmium comparing the results on MS·3 with earthworms and two plant species versus soil columns without organisms. Metals were applied on the top of the soil at three doses low (3.4 g Cu/ha + 1.7 g Cd/ha), medium (8.5 g Cu/ha + 4.3 g Cd/ha) and high (17 g Cu/ha + 8.5 g Cd/ha). Three organic compounds (pentachlorophenol, 4-chlorophenol and chlorpyrifos) were applied. Toxicity and metal levels in biota followed dose-response relationships. Results showed higher metal concentrations in the depth layers of MS·3 than in the soil columns. The effect was higher for the lower dose, where organisms were less affected, than at the higher doses, where very severe toxicity was observed, confirming the role of organisms in the enhanced mobility. Copyright © Taylor & Francis Group, LLC

Sewage sludge applied to agricultural soil Ecotoxicological effects on representative soil organisms

Application of sewage sludge to agricultural lands is a current practice in EU. European legislation permits its use when concentrations of metals in soil do not increase above the maximum permissible limits. In order to assess the fate and the effects on representative soil organisms of sewage sludge amendments on agricultural lands, a soil microcosm (multi-species soil system-MS 3) experiment was performed. The MS 3 columns were filled with spiked soil at three different doses 30, 60 and 120 t ha-1 fresh wt. Seed plants (Triticum aestivum, Vicia sativa and Brassica rapa) and earthworms (Eisenia fetida) were introduced into the systems. After a 21-d exposure period, a statistically significant increase for Cd, Cu, Zn and Hg concentrations was found for the soils treated with the highest application rate. Dose-related increase was observed for nickel concentrations in leachates. Plants and earthworm metal body burden offer much more information than metal concentrations and help to understand the potential for metal accumulation. Bioaccumulation factor (BAFplant-soil) presented a different behavior among species and large differences for BAFearthworm-soil, from control or sewage-amended soil, for Cd and Hg were found. B. rapa seed germination was reduced. Statistically significant decrease in fresh biomass was observed for T. aestivum and V. sativa at the highest application rate, whereas B. rapa biomass decreased at any application rate. Enzymatic activities (dehydrogenase and phosphatase) as well as respiration rate on soil microorganisms were enlarged

Effects of sulfachlorpyridazine in MS·3-arable land A multispecies soil system for assessing the environmental fate and effects of veterinary medicines

A multispecies soil system (MS·3) has been used to evaluate the ecological effects of veterinary pharmaceuticals in soil as a result of routine agricultural practices. Different experimental conditions were tested and the variation of the different parameters was evaluated for a final design. A protocol for the MS·3-arable land is presented here. Emergence of seedlings, plant elongation and biomass, earthworm mortality, and soil microbial enzymatic activities have been selected as toxicological endpoints for soil organisms. Toxicity tests were conducted with the leachate on aquatic organisms (in vitro fish cell lines, daphnids, and algae). The system was used for assessing the effects of the antimicrobial sulfachlorpyridazine that was tested in triplicate at concentrations of 0.01, 1, and 100 mg/kg. The chemical was mixed uniformly with a 20-cm depth soil column to resemble the distribution of manure within arable soil. Reversible and nonreversible effects on soil enzymatic activities were observed at 1 and 100 mg/kg, respectively. Earthworms were not affected. Significant reduction of plant elongation and biomass was observed at the highest concentration. Degradation and leaching contributed to the dissipation of sulfachlorpyridazine from the soil column. The undiluted leachate was highly toxic to Daphnia magna. The parent chemical was assumed responsible for the leachate toxicity although the role of mobile metabolites could not be excluded fully. No significant effects were observed for green algae Chlorella vulgaris and for the rainbow trout established cell lines RTG-2 (rainbow trout gonads) and RTL-W1 (rainbow trout liver). The MS·3 system offers a cost-effective experimental approach to measure simultaneously fate and effects of chemicals on a realistic soil system under controlled laboratory conditions. The advantages of using MS·3-effect endpoints are discussed. © 2005 SETAC