Identifying rare variants from exome scans: the GAW17 experience
Genetic Analysis Workshop 17 (GAW17) provided a platform for evaluating existing statistical genetic methods and for developing novel methods to analyze rare variants that modulate complex traits. In this article, we present an overview of the 1000 Genomes Project exome data and simulated phenotype data that were distributed to GAW17 participants for analyses, the different issues addressed by the participants, and the process of preparation of manuscripts resulting from the discussions during the workshop.

title: Linkage mapping of total cholesterol level in a young cohort via nonparametric regression
abstract: BackgroundCompared to model-based approaches, nonparametric methods for quantitative trait loci mapping are more robust to deviations in distributional assumptions. In this study, we modify a nonparametric regression method and the "contrast function"- based regression method to analyze total cholesterol level in the younger cohort (the offspring generation) of the Genetic Analysis Workshop 13 simulated data set.ResultsWe obtained significant evidence of linkage near four of the six non-sex-specific genes in at least 30% of the replicates.ConclusionsThe proposed nonparametric method seems to be a powerful robust alternative to distribution-based methods.

title: Integrating sibship data for mapping quantitative trait loci.
abstract: Sibship methods have been shown to be more powerful than traditional sib-pair methods in mapping quantative trait loci. We propose a statistical procedure which integrates data on sibships into a so-called 'contrast function', a natural extension of the classical squared sib-pair trait difference proposed by Haseman & Elston (1972). We also develop a combined mean and contrast function which provides more information on linkage compared to the contrast function. Our method is extended to multiple, epistatically interacting trait loci. Monte-Carlo simulations are included to compare the efficiencies of the proposed procedures with some currently used methods. An application of our proposed method is presented using data on alcohol dependence.

title: Linkage mapping of a complex trait in the New York population of the GAW14 simulated dataset: a multivariate phenotype approach
abstract: Multivariate phenotypes underlie complex traits. Thus, instead of using the end-point trait, it may be statistically more powerful to use a multivariate phenotype correlated to the end-point trait for detecting linkage. In this study, we develop a reverse regression method to analyze linkage of Kofendrerd Personality Disorder affection status in the New York population of the Genetic Analysis Workshop 14 (GAW14) simulated dataset. When we used the multivariate phenotype, we obtained significant evidence of linkage near four of the six putative loci in at least 25% of the replicates. On the other hand, the linkage analysis based on Kofendrerd Personality Disorder status as a phenotype produced significant findings only near two of the loci and in a smaller proportion of replicates.

title: A quantile-based method for association mapping of quantitative phenotypes: an application to rheumatoid arthritis phenotypes
abstract: Genetic association of population-based quantitative trait data has traditionally been analyzed using analysis of variance (ANOVA). However, violations of certain statistical assumptions may lead to false-positive association results. In this study, we have explored model-free alternatives to ANOVA using correlations between allele frequencies in the different quantile intervals of the quantitative trait and the quantile values. We performed genome-wide association scans on anti-cyclic citrullinated peptide and rheumatoid factor-immunoglobulin M, two quantitative traits correlated with rheumatoid arthritis, using the data provided in Genetic Analysis Workshop 16. Both the quantitative traits exhibited significant evidence of association on Chromosome 6, although not in the human leukocyte antigen region which is known to harbor a major gene predisposing to rheumatoid arthritis. We found that while a majority of the significant findings using the asymptotic thresholds of ANOVA was not validated using permutations, a relatively higher proportion of the significant findings using the asymptotic cut-offs of the correlation statistic were validated using permutations.

title: A novel non-parametric regression reveals linkage on chromosome 4 for the number of externalizing symptoms in sib-pairs.
abstract: In this report, we present results of a genome-wide linkage scan using as a phenotype the number of externalizing symptoms associated with alcohol use disorders. Subjects were collected by the Collaborative Study on the Genetics of Alcoholism project from families in which at least three first degree relatives were affected by alcohol dependence. We use a novel non-parametric regression method based on kernel smoothing for our analysis. We report a statistically significant linkage close to the ADH gene cluster on Chromosome 4. We also obtain evidence for epistatic interaction between a region on Chromosome 1 and one on Chromosome 15. Although alcoholism as a covariate does not have any effect on the linkage scan, it has an effect on the epistatic interaction.

title: Cloning of the human cholesteryl ester hydrolase promoter: identification of functional peroxisomal proliferator-activated receptor responsive elements.
abstract: Cholesteryl ester hydrolase (CEH) is responsible for hydrolysis of stored cholesterol esters in macrophage foam cells and release of free cholesterol for high-density lipoprotein-mediated efflux. PCR-based screening of human genomic libraries with human macrophage CEH specific primers resulted in amplification and cloning of 1.7 kb promoter sequence. Analysis of the sequence revealed a lack of consensus TATA-box but presence of a GC-rich proximal sequence, a CAAT box and several binding sites for the transcription factor Sp1. Three putative response elements for peroxisome proliferator-activated receptor (PPRE) were identified at position -176, -779, and -1316. Down-regulation of promoter activity was observed in the presence of either PPARalpha- or PPARgamma-specific ligands and introduction of a 4-point transverse mutation in the PPRE at -176 completely abolished the effect of PPAR ligands on the promoter activity. Analogous to other genes involved in macrophage cholesterol homeostasis, human CEH may also be regulated by PPAR.

title: Noninvasive electrocardiographic imaging (ECGI) of a univentricular heart with Wolff-Parkinson-White syndrome.
abstract: Electrocardiographic imaging (ECGI) is a noninvasive functional imaging modality which reconstructs epicardial potentials, electrograms, and activation and recovery maps from body-surface ECG potentials. For this purpose, up to 256 ECGs are recorded by a 256-channel body surface potential mapping (BSPM) system and the heart-torso geometry is obtained using thoracic computed tomography (CT)1,2. This technique was developed and validated extensively in normal and abnormal canine hearts3–11. More recently, ECGI was validated in humans by comparison with direct intra-operative epicardial mapping in patients undergoing open-heart surgery12. To date, ECGI has been applied in adult human subjects for the following purposes: 1. To study cardiac electrophysiology of the normal adult human heart2. 2. To image electrophysiologic responses to pacing in heart failure patients undergoing cardiac resynchronization therapy13 (pacing sites were localized with an accuracy better than 10 mm)14. 3. To guide catheter ablation of focal ventricular and atrial tachycardias15,16. 4. To image typical atrial flutter prior to catheter ablation1 and atypical atrial flutter prior to a surgical Cox-Maze procedure17. 
 
Here we describe for the first time, a case where ECGI was applied to a pediatric patient with a congenital structural heart defect. The patient had a univentricular heart and Wolff-Parkinson-White syndrome, and ECGI was used to localize the accessory pathway and help guide catheter ablation. To date, there have been no reported cases of ECGI in the pediatric population.

title: Molecular characterization of rare bovine group A rotavirus G15P[11] and G15P[21] strains from eastern India: identification of simian SA11-like VP6 genes in G15P[21] strains
abstract: During a surveillance study (November 2001–March 2005), one rare G15P[11] and two rare G15P[21] bovine group A rotavirus strains were detected in diarrhoeic calves in Eastern India. Sequence analysis of the VP8*, VP6, NSP4 and NSP5 genes of the G15P[11] strain confirmed its bovine origin. Although the NSP4 and NSP5 genes of the two G15P[21] strains were of bovine origin, their VP6 genes shared higher nucleotide and amino acid identities with simian strain SA11 (92.5–93.1% and 98.5–98.7%) than bovine strains (88.5–88.9% and 97–97.2%), and by phylogenetic analysis, exhibited clustering with SA11, distantly related to bovine strains. All these pointed towards a possible reassortment event of VP6 gene between bovine and simian (SA11-like) strains. Therefore, the present study provided molecular evidence for bovine origin of G15 strains and revealed a rare instance of genetic diversity in the bovine VP6 gene, otherwise conserved in group A rotavirus strains from cattle.

title: Full genomic analysis of a simian SA11-like G3P[2] rotavirus strain isolated from an asymptomatic infant: identification of novel VP1, VP6 and NSP4 genotypes.
abstract: We report here the full genomic analysis of a simian SA11-like G3P[2] group A rotavirus (GAR) strain, B10, isolated from an asymptomatic infant in Kenya in 1987. By nucleotide sequence identities and phylogenetic analyses, the VP7-VP4-VP2-VP3-NSP1-NSP2-NSP3-NSP5 genes of strain B10 exhibited maximum genetic relatedness to those of the different isolates of simian strain SA11, and were assigned to the G3-P[2]-C5-M5-A5-N5-T5-H5 genotypes, respectively. On the other hand, the VP1, VP6 and NSP4 genes of strain B10 did not belong to any of the established GAR genotypes, and therefore, were assigned to new genotype numbers R8, I16 and E13, respectively, by the Rotavirus Classification Working Group. These observations suggested that strain B10 might have originated from reassortment event/s involving simian SA11-like strains and GAR strains from unknown animal host species (possibly other wild animals) preceding transmission to humans. Alternatively, considering the lack of data on simian GARs, it might be also possible that the VP1, VP6 and NSP4 genes of strain B10 are those of unknown simian strains, and that strain B10 might be a typical simian strain that was directly transmitted to humans. Therefore, either hypothesis pointed towards a rare instance of possible direct transmission of GARs from an animal host (possibly a monkey or some other wild animal) to humans. This was corroborated by the presence of different species of wild animals including non-human primates, and unhygienic conditions at the sampling site. To our knowledge, the present study is the first report on the detection of a simian SA11-like G3P[2] GAR strain in humans.

title: Effects of Mn levels on resistance of Bacillus megaterium spores to heat, radiation and hydrogen peroxide.
abstract: To determine the effects of Mn levels in Bacillus megaterium sporulation and spores on spore resistance.



Bacillus megaterium was sporulated with no added MnCl(2) and up to 1 mmol l(-1) MnCl(2). The resultant spores were purified and loosely bound Mn removed, and spore Mn levels were found to vary c. 100-fold. The Mn level had no effect on spore γ-radiation resistance, but B. megaterium spores with elevated Mn levels had higher resistance to UVC radiation (as did Bacillus subtilis spores), wet and dry heat and H(2)O(2). However, levels of dipicolinic acid and the DNA-protective α/β-type small, acid-soluble spore proteins were the same in spores with high and low Mn levels.



Mn levels either in sporulation or in spores are important factors in determining levels of B. megaterium spore resistance to many agents, with the exception of γ-radiation.



The Mn level in sporulation is an important factor to consider when resistance properties of B. megaterium spores are examined, and will influence the UV resistance of B. subtilis spores, some of which are used as biological dosimeters.