Forensic ancestry analysis with two capillary electrophoresis ancestry informative marker (AIM) panels: Results of a collaborative EDNAP exercise.

There is increasing interest in forensic ancestry tests, which are part of a growing number of DNA analyses that can enhance routine profiling by obtaining additional genetic information about unidentified DNA donors. Nearly all ancestry tests use single nucleotide polymorphisms (SNPs), but these currently rely on SNaPshot single base extension chemistry that can fail to detect mixed DNA. Insertion-deletion polymorphism (Indel) tests have been developed using dye-labeled primers that allow direct capillary electrophoresis detection of PCR products (PCR-to-CE). PCR-to-CE maintains the direct relationship between input DNA and signal strength as each marker is detected with a single dye, so mixed DNA is more reliably detected. We report the results of a collaborative inter-laboratory exercise of 19 participants (15 from the EDNAP European DNA Profiling group) that assessed a 34-plex SNP test using SNaPshot and a 46-plex Indel test using PCR-to-CE. Laboratories were asked to type five samples with different ancestries and detect an additional mixed DNA sample. Statistical inference of ancestry was made by participants using the Snipper online Bayes analysis portal plus an optional PCA module that analyzes the genotype data alongside calculation of Bayes likelihood ratios. Exercise results indicated consistent genotyping performance from both tests, reaching a particularly high level of reliability for the Indel test. SNP genotyping gave 93.5% concordance (compared to the organizing laboratory's data) that rose to 97.3% excluding one laboratory with a large number of miscalled genotypes. Indel genotyping gave a higher concordance rate of 99.8% and a reduced no-call rate compared to SNP analysis. All participants detected the mixture from their Indel peak height data and successfully assigned the correct ancestry to the other samples using Snipper, with the exception of one laboratory with SNP miscalls that incorrectly assigned ancestry of two samples and did not obtain informative likelihood ratios for a third. Therefore, successful ancestry assignments were achieved by participants in 92 of 95 Snipper analyses. This exercise demonstrates that ancestry inference tests based on binary marker sets can be readily adopted by laboratories that already have well-established CE regimes in place. The Indel test proved to be easy to use and allowed all exercise participants to detect the DNA mixture as well as achieving complete and concordant profiles in nearly all cases. Lastly, two participants successfully ran parallel next-generation sequencing analyses (each using different systems) and achieved high levels of genotyping concordance using the exercise PCR primer mixes unmodified.

Randomised controlled trial of thermostatic mixer valves in reducing bath hot tap water temperature in families with young children in social housing.

OBJECTIVES: To assess the effectiveness of thermostatic mixing valves (TMVs) in reducing bath hot tap water temperature, assess acceptability of TMVs to families and impact on bath time safety practices. DESIGN: Pragmatic parallel arm randomised controlled trial. SETTING: A social housing organisation in Glasgow, Scotland, UK. PARTICIPANTS: 124 families with at least one child under 5 years. INTERVENTION: A TMV fitted by a qualified plumber and educational leaflets before and at the time of TMV fitting. MAIN OUTCOME MEASURES: Bath hot tap water temperature at 3-month and 12-month post-intervention or randomisation, acceptability, problems with TMVs and bath time safety practices. RESULTS: Intervention arm families had a significantly lower bath hot water temperature at 3-month and 12-month follow-up than families in the control arm (3 months: intervention arm median 45.0°C, control arm median 56.0°C, difference between medians, -11.0, 95% CI -14.3 to -7.7); 12 months: intervention arm median 46.0°C, control arm median 55.0°C, difference between medians -9.0, 95% CI -11.8 to -6.2) They were significantly more likely to be happy or very happy with their bath hot water temperature (RR 1.43, 95% CI 1.05 to 1.93), significantly less likely to report the temperature as being too hot (RR 0.33, 95% CI 0.16 to 0.68) and significantly less likely to report checking the temperature of every bath (RR 0.84, 95% CI 0.73 to 0.97). Seven (15%) intervention arm families reported problems with their TMV. CONCLUSIONS: TMVs and accompanying educational leaflets are effective at reducing bath hot tap water temperatures in the short and longer term and are acceptable to families. Housing providers should consider fitting TMVs in their properties and legislators should consider mandating their use in refurbishments as well as in new builds.

Automated in-vitro testing of orthopaedic implants: a case study in shoulder joint replacement.

This investigation presents the design and preliminary validation of a single station simulator with biaxial motion and loading designed to mimic the kinematics of the glenohumeral joint during arm abduction in the scapular plane. Although the design of the glenoid holder allows the glenoid component to translate in all three axes, it is primarily loaded axially, which brings it into contact with the oscillating humeral head, but is also loaded superiorly to simulate common subluxation of the humeral head. Simulating arm abduction in the scapular plane simplifies component alignment and removes the need for anterior-posterior loading, thereby creating a stable joint without the need to simulate capsular constraints. In this more physiologically accurate simulator design, the load and motion profiles influence the contact kinematics, but the wear path is ultimately determined by the conformity and constraint designed into the bearing couple. The wear data are determined and correlated with clinically retrieved glenoid components, as well as previously reported in-vitro studies, thus verifying use of the simulator in testing alternative materials and designs. The key design features, as well as the improvements proposed through this study, can be incorporated into the design of test fixtures for any other orthopaedic implant such as the hip, knee, spine, elbow, and finger.

The role of the Clinical Governance Adviser in supporting quality improvement in general dental practice: the Glasgow Quality Practice Initiative.

AIM: The purpose of this paper is to share information derived from the Glasgow Quality Practice Initiative with general dental practice teams, Dental Practice Advisers and others involved in quality improvement. METHOD: A sample of 16 general dental practices was selected from volunteers to receive assistance in working towards a Quality Practice Award. Two Clinical Governance Advisers were appointed to provide this support. DATA COLLECTED: Quantitative, qualitative and observational data were collected, and comparisons made between practices that had and had not received support. RESULTS: Selected results are presented demonstrating both the baseline position and comparisons of the 'Intervention' and 'Non-Intervention' groups. CONCLUSIONS: and recommendations Baseline levels of quality assurance were generally poor. It is asserted that the practices receiving Clinical Governance Adviser support benefited from the experience and made meaningful improvements. This has implications for the development of national policy in Scotland.

Two adult galactosaemia females with normal ovarian function and identical GALT mutations (Q188R/R333G).

We report two unrelated cases of adult galactosaemia females with normal ovarian function and Q188R/R333G mutations. Clinical history has been followed for 40 years. Biochemical finding in one patient are consistent with the presence of small amounts of galactose-1-phosphate uridyltransferase (GALT) activity, which differs from classical galactosaemia.

An improved HPLC analysis of the metabolite furoic acid in the urine of workers occupationally exposed to furfural.

An improved high-performance liquid chromatographic (HPLC) method for the analysis of the metabolite furoic acid in the urine of workers occupationally exposed to furfural is described. The procedure involved an alkaline hydrolysis step followed by solvent extraction using ethyl acetate. HPLC analysis used an acidic acetonitrile/water mobile phase with a C18 column and ultraviolet detection. The overall relative recovery of furoic acid in urine was found to be 98.8% with a relative standard deviation of 9.7%. The limit of quantitation was determined to be 0.01 mmol/L.

Negative ion chemical ionization GC/MS-MS analysis of dialkylphosphate metabolites of organophosphate pesticides in urine of non-occupationally exposed subjects.

Low level exposure to organophosphate (OP) pesticides can be determined by the measurement of dialkylphosphate (DAP) metabolites in urine. An analytical method is presented here which can measure the metabolites dimethyl phosphate (DMP), diethyl phosphate (DEP), dimethyl thiophosphate (DMTP), dimethyl dithiophosphate (DMDTP), diethyl thiophosphate (DETP), and diethyl dithiophosphate (DEDTP) at low levels. This was achieved by lyophilization of the urine, derivatization with pentafluorobenzyl bromide (PFBBr) and quantification by negative ion chemical ionization GC/MS-MS. The detection limits for the metabolites were 0.5 microg L(-1) DMP, 0.1 microg L(-1) DEP, 0.1 microg L(-1) DMTP, 0.04 microg L(-1) DMDTP, 0.04 microg L(-1) DETP and 0.02 microg L(-1) DEDTP. The RSD for the analytical method was 4-14% for the six metabolites. The method was used to monitor a group of non-occupationally exposed individuals in Sydney, Australia. The metabolites DMP, DEP, DMTP, DMDTP, DETP and DEDTP occurred in 73, 77, 96, 48, 100 and 2% of the samples with median values of 13, 3, 12, <1, 1 and 1 microg L(-1) respectively. The method is simple to use, sensitive and suitable for routine analysis of non-occupational exposure levels. These detection limits are between one and two orders of magnitude lower than those previously reported in the literature.

Results of a collaborative study of the EDNAP group regarding the reproducibility and robustness of the Y-chromosome STRs DYS19, DYS389 I and II, DYS390 and DYS393 in a PCR pentaplex format.

A collaborative exercise was carried out by the European DNA Profiling Group (EDNAP) in the frame work of the STADNAP program, i.e. standardization of DNA profiling in Europe, in order to evaluate the performance of a Y-chromosome STR pentaplex, which includes the loci DYS19, DYS389 I and II, DYS390 and DYS393 and to determine whether uniformity of results could be achieved among different European laboratories. Laboratories were asked to analyze the five Y-STRs using singleplex and multiplex conditions in three bloodstains and one mixed stain (95% female and 5% male). All the laboratories reported the same results even for the mixed stain included in the exercise. This demonstrates the reproducibility and robustness of Y-chromosome STR typing even with multiplex formats and proves the usefulness of Y-STR systems for analyzing mixed stains with a male component.A total of 930 male samples from 10 different populations from Europe were also analysed for all the loci included in the pentaplex. Eight of these ten populations also included haplotype data. As for single gene analysis, haplotype diversity was higher in Germany and Italy and lower in Western European countries and Finland. Pairwise haplotype analysis shows the Finnish departure from the rest of the populations and a relatively homogeneity in the other European populations with F(ST) estimates lower than 0.05.UPGMA analysis shows an association of Western European population (Ireland, UK, Portugal and Galicia) on the one hand and central European populations on the other.

Routine gas chromatographic determination of dialkylphosphate metabolites in the urine of workers occupationally exposed to organophosphorus insecticides.

A routine gas chromatographic (GC) method is described for the analysis of dialkylphosphate metabolites in the urine of workers occupationally exposed to organophosphorus insecticides. The procedure involves derivatizing a freeze-dried urine sample with pentafluorobenzyl bromide and then determining the metabolites using dual capillary column GC with flame photometric detection.

A physically based approach to modelling radionuclide transport in the biosphere.

Calculations of radiological risk are required to assess the safety of any potential future UK deep underground repository for intermediate-level and certain low-level solid radioactive wastes. In support of such calculations, contaminant movement and dilution in the terrestrial biosphere is investigated using the physically based modelling system SHETRAN. Two case studies are presented involving modelling of contaminants representing long-lived poorly sorbed radionuclides in the near-surface aquifers and surface waters of hypothetical catchments. The contaminants arise from diffuse sources at the base of the modelled aquifers. The catchments are characterised in terms of detailed spatial data for topography, the river network, soils and vegetation. Simulations are run for temperate and boreal climates representing possible future conditions at a repository site. Results are presented in terms of the concentration of contaminants in the aquifer, in soils and in surface waters; these are used to support the simpler models used in risk calculations.

A randomized factorial trial of reperfusion strategies and aspirin dosing in acute myocardial infarction. The DUCCS-II Investigators.

The focus of new research efforts to improve the morbidity and mortality associated with acute myocardial infarction (AMI) has turned to adjuvant agents that show promise of improving outcomes following coronary thrombolysis. We enrolled 162 patients with AMI in a randomized trial comparing front-loaded tissue-plasminogen activator (t-PA) plus weight-adjusted heparin with anisoylated plasminogen streptokinase activator complex (APSAC) without heparin as well as standard-dose (325 mg) and low-dose (81 mg) aspirin. The primary end point was an in-hospital morbidity profile; secondary end points were clinical and angiographic potency and hemorrhagic events. Selected sites performed an electrocardiographic substudy to determine the time to 50% ST-segment recovery and the time to steady state. Although the trial was terminated when the Global Utilization of Streptokinase and t-PA for Occluded Coronary Arteries-I trial showed that t-PA had a significant mortality advantage over streptokinase, important trends were evident. Patients given t-PA and heparin were better anticoagulated (p = 0.001), yet AP-SAC-treated patients had more bleeding complications. The primary end point favored t-PA (25.4% vs 31.3%), and the secondary end points were similar in both groups. In the electrocardiographic substudy, the t-PA group achieved both 50% ST-segment recovery and steady-state recovery sooner than the APSAC group. Patients taking low-dose aspirin had lower in-hospital mortality and less recurrent ischemia but more strokes than the standard-dose aspirin group. Thus, this trial demonstrated trends favoring front-loaded t-PA with weight-adjusted heparin over APSAC without heparin in the treatment of AMI. The use of low-dose aspirin did not appear to impose a loss of protection from adverse events, nor did standard-dose aspirin increase serious bleeding.

Analysis of trichloroacetic acid in the urine of workers occupationally exposed to trichloroethylene by capillary gas chromatography.

A gas chromatographic procedure is described for the determination of trichloroacetic acid in urine, the major metabolite of trichloroethylene exposure. Trichloroacetic acid was derivatised to its methyl ester with BF3/methanol reagent and then extracted into toluene and analysed by capillary gas chromatography using electron-capture detection. The response was linear in the range 0.4-100 mg/l of trichloroacetic acid in urine and showed a relative recovery of 99.6%. The procedure is suitable for monitoring occupational exposure to trichloroethylene.

Radiochemical assay of minute quantities of galactose-1-phosphate uridyltransferase activity in erythrocytes and leukocytes of galactosemia patients.

A sensitive radioisotopic method has been developed which can detect galactose-1-phosphate uridyltransferase (GALT) activity as low as 0.1% of normal control values in both erythrocytes and leukocytes. This assay utilizes carbon-14 labeled galactose-1-phosphate with high specific activity and requires removal of endogenous galactose-1-phosphate (Gal-1-P) and uridine diphosphate glucose (UDPGlc) through dialysis. Optimal exogenous UDPGlc concentration has been determined with a fixed concentration of Gal-1-P in the incubation. The rate of product, uridine diphosphate galactose (UDPGal), formation is monitored at three different times. Among 423 patients with galactosemia studied by this method, 363 patients exhibited no detectable GALT activity in their erythrocytes and 60 patients were found to have detectable erythrocyte GALT activity ranging from 0.02 to 5.0 units normal values: > 20 units). The former group of patients was designated as classic galactosemia (GG) and the latter group as galactosemia variant (GV). Leucocytes from ten patients belonging to the GG group also showed complete absence of GALT activity while leukocytes from two patients belonging to the GV group showed GALT activity at levels comparable with those found in their erythrocytes. Because there is extensive biochemical heterogeneity among galactosemia patients, we recommend that an assay with increase sensitivity be carried out on blood samples from galactosemia patients so that clinical, biochemical and molecular correlations made by different groups of investigators can be compared.

Biochemical and molecular studies of 132 patients with galactosemia.

We evaluated 132 galactosemia patients for the Q188R (glutamine-188 to arginine) mutation in the human galactose-1-phosphate uridyltransferase (GALT) gene and for GALT activity in their hemolysates by a sensitive radioisotopic method. In those without any detectable GALT activity (GG), the Q188R mutation constituted 67% of the alleles. In patients with detectable GALT activity (GV), only 16% of the alleles were accounted for by Q188R. In all patients who were homozygous for the Q188R mutation, no erythrocyte GALT activity could be demonstrated. There was an extensive variation in the amount of detectable GALT activity ranging from 0.1% to 5% of the normal values among the GV patients. There was a difference in the frequency of Q188R mutation in the GALT alleles among patients belonging to different racial and ethnic groups. In Caucasian and Hispanic patients, the frequency was not far different (64% and 58%, respectively). On the other hand, only 12% of the GALT alleles with Q188R were found in African-American patients.

Results of a survey of carrier women for the galactosemia gene.

A survey of 108 heterozygote women for the classic galactosemia gene, GALT, did not reveal that the carrier state was associated with premature ovarian failure or ovarian cancer. This survey did not support previous epidemiologic studies suggesting an increased risk for ovarian dysfunction in women with deficiency of the GALT enzyme.

Galactosemia: evaluation with MR imaging.

The cerebral findings at magnetic resonance imaging in 67 transferase-deficient galactosemic patients (36 female, 31 male; median age, 10 years) are reported. Twenty-two patients had mild cerebral atrophy, eight had cerebellar atrophy, and 11 had multiple small hyperintense lesions in the cerebral white matter on T2-weighted images. The classic galactosemic patients (those without measurable transferase activity) older than 1 year of age did not show the normal dropoff in peripheral white matter signal intensity on intermediate- and T2-weighted images. The authors postulate that this abnormal signal intensity is due to altered myelin formation secondary to the inability to make sufficient and/or normal galactocerebroside.