Linkage analyses of rheumatoid arthritis and related quantitative phenotypes: the GAW15 experience.
The group that formed on the theme of linkage analyses of rheumatoid arthritis RA and related phenotypes (Group 10) in the Genetic Analysis Workshop 15 comprised 18 sets of investigators. Two data sets were available: one was a real set provided by the North American Rheumatoid Arthritis Consortium and collaborators in Canada, France (European Consortium Of Rheumatoid Arthritis Families) and the UK; the other was a simulated data set modelled after the real data set. Whereas a majority of the investigators analyzed the RA affection status as a binary phenotype, a few contributions considered data on correlated quantitative traits such as anti-cyclic citrullinated peptide and rheumatoid factor-immunoglobulin M. The different investigators applied a wide spectrum of linkage methods. As expected, most methods could identify the human leukocyfeantigen region on chromosome 6 as a major genetic factor for RA. In addition, some novel chromosomal regions provided significant evidence of linkage in multiple contributions in the group. In this report, we discuss the different strategies explored by the different investigators with the common goal of improving the power to detect linkage.

title: A two-stage variable-stringency semiparametric method for mapping quantitative-trait loci with the use of genomewide-scan data on sib pairs.
abstract: Genomewide scans for mapping loci have proved to be extremely powerful and popular. We present a semiparametric method of mapping a quantitative-trait locus (QTL) or QTLs with the use of sib-pair data generated from a two-stage genomic scan. In a two-stage genomic scan, either the entire genome or a large portion of the genome is saturated with low-density markers at the first stage. At the second stage, the intervals that are identified as probable locations of the trait loci, by means of analysis of data from the first stage, are then saturated with higher-density markers. These data are then analyzed for fine mapping of the loci. Our statistical strategy for analysis of data from the first stage is a low-stringency method based on the rank correlation of squared trait-difference values of the sib pairs and the estimated identity-by-descent scores at the marker loci. We suggest the use of a low-stringency method at the first stage, to save on computational time and to avoid missing any marker interval that may contain the trait loci. For analysis of data from the second stage, we have developed a high-stringency nonparametric-regression approach, using the kernel-smoothing technique. Through extensive simulations, we show that this approach is more powerful than is a currently used method for mapping QTLs by use of sib pairs, particularly in the presence of dominance and epistatic effects at the trait loci.

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: 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: Identifying rare variants from exome scans: the GAW17 experience
abstract: 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: 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: 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: Activation in vitro of NF-kappa B by phosphorylation of its inhibitor I kappa B.
abstract: Nuclear factor kappa B (NF-kappa B), which was first detected by its binding to the kappa B site in the immunoglobulin kappa-gene enhancer, is important for the regulated expression of the kappa-gene and is partly responsible for the induction in appropriate cells of interleukin-2 (IL-2), IL-2 alpha receptor, beta-interferon and serum amyloid A protein. NF-kappa B is present as a nuclear DNA-binding protein in B lymphocytes and mature macrophages, but is found in the cytoplasm of many cells in a form unable to bind to DNA. The cytoplasmic form is bound to an inhibitor protein, I kappa B, from which it can be released in vitro by deoxycholate and other agents. Activation of cells by various agents, notably the phorbol esters that stimulate protein kinase C (PKC), leads to dissociation in vivo of the NF-kappa B/I kappa B complex and migration of NF-kappa B to the nucleus. Therefore, it acts as a second messenger system, transducing activation signals from the cytoplasm to the nucleus. To elucidate the mechanism of signal transfer, we have used an in vitro system in which addition of purified protein kinases to a partially purified NF-kappa B/I kappa B complex leads to the activation of the DNA-binding activity of NF-kappa B. Using gel retardation assays we found that PKC, cyclic AMP-dependent protein kinase (PKA) and a haem-regulated eIF-2 kinase (HRI) could activate NF-kappa B in vitro, whereas casein kinase II was ineffective. To determine the target for the protein kinases we purified and characterized both NF-kappa B and I kappa B and found that I kappa B is phosphorylated and inactivated in the presence of PKC and HRI but not PKA.

title: The Papworth BiVent tube: a feasibility study of a novel double-lumen endotracheal tube and bronchial blocker in human cadavers.
abstract: A novel double-lumen endotracheal tube, the Papworth BiVent tube, has been designed to allow the rapid passage of a blocker into either main bronchus, without fibreoptic endoscopic guidance.



The feasibility of lung isolation and one-lung ventilation (OLV) in human cadavers is examined, along with displacement of the bronchus blocker during head and neck movement.



Cadaveric endotracheal intubation with the Papworth BiVent tube was straightforward and comparable with intubation with a conventional single-lumen tube (SLT). Reliable lung isolation was achieved considerably faster using the Papworth BiVent tube than with a bronchoscopically guided bronchial blocker through an SLT (mean 7.75 s BiVent tube vs 128.2 s SLT). The Papworth BiVent tube also prevented displacement of the blocker from its position in the bronchus on head movement.



This study in human cadavers has shown that it is feasible to use the Papworth BiVent tube to attain rapid and secure lung isolation for OLV. Further work is required in clinical settings.

title: Whole genomic analysis reveals the porcine origin of human G9P[19] rotavirus strains Mc323 and Mc345.
abstract: The group A rotavirus (RVA) P[19] is a rare P-genotype of the RVA VP4 gene, reported so far in humans and pigs. Whole genomic analyses of P[19] strains are essential to study their origin and evolutionary patterns. To date, all the 11 genes of only two P[19] strains, RVA/Human-wt/IND/RMC321/1990/G9P[19] and RVA/Human-wt/IND/mani-97/2006/G9P[19], have been analyzed, providing evidence for their porcine origin. In the present study, the whole genomes of the first reported human P[19] strains, RVA/Human-tc/THA/Mc323/1989/G9P[19] and RVA/Human-tc/THA/Mc345/1989/G9P[19], were analyzed. Strains Mc323 and Mc345 exhibited a G9-P[19]-I5-R1-C1-M1-A8-N1-T1-E1-H1 genotype constellation. With the exception of the NSP5 gene, both the strains were closely related to each other. Most of the genes of Mc323 (VP2-4, VP6-7, NSP1-4 genes) and Mc345 (VP2-4, VP6-7 and NSP1-5 genes) appeared to be of porcine origin, whilst the exact origin of VP1 and NSP5 genes of Mc323 and VP1 gene of Mc345 could not be ascertained. Therefore, strains Mc323 and Mc345 were found to have a porcine RVA genetic backbone, and are likely of porcine origin. Taken together, our observations corroborated the hypothesis that P[19] strains might be derived from porcine RVAs, providing important insights into the origin of P[19] strains, and on interspecies transmission of RVAs.

title: Evidence for the porcine origin of equine rotavirus strain H-1.
abstract: Equine group A rotavirus (RVA) strain H-1 (RVA/Horse-tc/GBR/H-1/1975/G5P9[7]) was found to have VP4, VP6-7, NSP1 and NSP4 genes of porcine origin. In order to obtain conclusive information on the exact origin and evolution of this unusual equine strain, the remaining six genes (VP1-3, NSP2-3 and NSP5 genes) of strain H-1 were analyzed in the present study. By whole genomic analysis, strain H-1 exhibited a porcine RVA-like genotype constellation (G5-P[7]-I5-R1-C1-M1-A8-N1-T1-E1-H1), different from those of typical equine RVA strains. The VP2-3 and NSP2-3 genes of strain H-1 were found to originate from porcine RVAs. On the other hand, it was difficult to pinpoint the exact origin of the VP1 and NSP5 genes of strain H-1, though phylogenetically, these genes appeared to be possibly derived from porcine or Wa-like human strains. Taken together, at least nine (VP2-4, VP6-7 and NSP1-4 genes) of the 11 gene segments of strain H-1 were found to be of porcine origin, revealing a porcine RVA-like genetic backbone. Therefore, strain H-1 is likely a porcine RVA strain that was transmitted to horses.