Increasing power of groupwise association test with likelihood ratio test

Sequencing studies have been discovering a numerous number of rare variants, allowing the identification of the effects of rare variants on disease susceptibility. As a method to increase the statistical power of studies on rare variants, several groupwise association tests that group rare variants in genes and detect associations between genes and diseases have been proposed. One major challenge in these methods is to determine which variants are causal in a group, and to overcome this challenge, previous methods used prior information that specifies how likely each variant is causal. Another source of information that can be used to determine causal variants is the observed data because case individuals are likely to have more causal variants than control individuals. In this article, we introduce a likelihood ratio test (LRT) that uses both data and prior information to infer which variants are causal and uses this finding to determine whether a group of variants is involved in a disease. We demonstrate through simulations that LRT achieves higher power than previous methods. We also evaluate our method on mutation screening data of the susceptibility gene for ataxia telangiectasia, and show that LRT can detect an association in real data. To increase the computational speed of our method, we show how we can decompose the computation of LRT, and propose an efficient permutation test. With this optimization, we can efficiently compute an LRT statistic and its significance at a genome-wide level. The software for our method is publicly available at http://genetics.cs.ucla.edu/rarevariants .     

Kernel-based association test

Association mapping (i.e., linkage disequilibrium mapping) is a powerful tool for positional cloning of disease genes. We propose a kernel-based association test (KBAT), which is a composite function of "P-values of single-locus association tests" and "kernel weights related to intermarker distances and/or linkage disequilibria." The KBAT is a general form of some current test statistics. This method can be applied to the study of candidate genes and can scan each chromosome using a moving average procedure. We evaluated the performance of the KBAT through simulation studies that considered evolutionary parameters, disease models, sample sizes, kernel functions, test statistics, window attributes, empirical P-value estimations, and genetic/physical maps. The results showed that the KBAT had a well-controlled false positive rate and high power compared to existing methods. In addition, the KBAT was also applied to analyze a genomewide data set from the Collaborative Study on the Genetics of Alcoholism. Important genes associated with alcoholism dependence were identified. In summary, the merits of the KBAT are multifold: the KBAT is robust against the inclusion of nuisance markers, is invariant to the map scale, and accommodates different types of genomic data, study designs, and study purposes. The proposed methods are packaged in the user-friendly software, KBAT, available at http://www.stat.sinica.edu.tw/hsinchou/genetics/association/KBAT.htm.     

Fit-climbing test: a field test for indoor rock climbing

The aim of this study was to develop an indoor rock-climbing test on an artificial wall (Fit-climbing test). Thirteen climbers (elite group [EG] = 6; recreational group [RG] = 7) performed the following tests: (a) familiarization in the Fit-climbing test, (b) the Fit-climbing test, and (c) a retest to evaluate the Fit-climbing test's reliability. Gas exchange, blood lactate concentration, handgrip strength, and heart rate were measured during the test. Oxygen uptake during the Fit-climbing test was not different between groups (EG = 8.4 ± 1.1 L; RG = 7.9 ± 1.5 L, p > 0.05). The EG performance (120 ± 7 movements) was statistically higher than the RG climbers' performance (78 ± 13 movements) during the Fit-climbing test. Consequently, the oxygen cost per movement during the Fit-climbing test of the EG was significantly lower than that of the RG (p < 0.05). Handgrip strength was higher in the EG when compared with that in the RG in both pre-Fit- and post-Fit-climbing test (p < 0.05). There were no significant differences in any other variables analyzed during the Fit-climbing test (p > 0.05). Furthermore, the performance in the Fit-climbing test presented high reliability (intraclass correlation coefficient = 0.97). Therefore, the performance during the Fit-climbing test may be an alternative to evaluate rock climbers because of its specificity and relation to oxygen cost per movement during climbing.     

The tail suspension test

The tail-suspension test is a mouse behavioral test useful in the screening of potential antidepressant drugs, and assessing of other manipulations that are expected to affect depression related behaviors. Mice are suspended by their tails with tape, in such a position that it cannot escape or hold on to nearby surfaces. During this test, typically six minutes in duration, the resulting escape oriented behaviors are quantified. The tail-suspension test is a valuable tool in drug discovery for high-throughput screening of prospective antidepressant compounds. Here, we describe the details required for implementation of this test with additional emphasis on potential problems that may occur and how to avoid them. We also offer a solution to the tail climbing behavior, a common problem that renders this test useless in some mouse strains, such as the widely used C57BL/6. Specifically, we prevent tail climbing behaviors by passing mouse tails through a small plastic cylinder prior to suspension. Finally, we detail how to manually score the behaviors that are manifested in this test.     

The acenocoumarol-carrageenin test

Rats received a single subcutaneous dose of the inflammatory agent carrageenin on the top of the skull, followed by oral acenocoumarol for 3 days; in every case, an extensive haematoma was observed on the skull on day 4. The capillary resistance on the depilated dorsal skin was significantly reduced by this combined inflammatory + anticoagulant treatment. Haematoma development was not inhibited by cortisone, non-steroidal anti-phlogistics (piroxicam, proquazone) or benzopyrone derivatives (hesperidin methylchalcone, oligomeric procyanidin). On pretreatment with vitamin K1 for 1 day, followed by daily treatment for 3 days, no haematoma was observed.     

The Avena geo-curvature test

Summary The Avena geo-curvature test is a bioassay for auxin-type growth regulators. Etiolated oat coleoptiles are used and the test is conducted in special perspex trays under diffuse daylight. The coleoptiles, with the primary leaves intact, are arranged in grooves on the trays in 4 series of 6 coleoptiles each, and cut to a size of 10 mm. The test substance is applied on an agar strip covering only the lower halves of the apical cut surfaces of each series. After a 4-hour stay in the dark in moist Petri dishes the coleoptile cylinders are shadowgraphed on a 35-mm photographic film. The curvatures are measured from an enlarged projection (10 times natural size) of the shadowgraphs.The lowest IAA concentration which can be determined is around 30 to 60 g/l, i.e. about 1 to 2 ng IAA. The concentration response curves follow a logarithmic course up to 1,000 g/l. The standard error of the mean in a test series comprising 6 coleoptiles is on an average ± 7%. The simple and quick procedures make it possible for a worker to test 60 to 80 fractions a day.Dedicated to Professor Hans Söding on the occasion of his seventieth birthday. The senior author expresses his special gratitude for having been initiated by Professor Söding in the study of auxins.present address: Dpt. of Biology Princeton Univ. Princeton, N. J. 08540, U.S.A.     

Quantitative test for Bacteroides nodosus pilus protein by an agglutination-absorption test

A pili agglutination-absorption test (PAAT) was developed for the quantitative measurement of pilus protein in cultures of Bacteroides nodosus and to quantify pili yields during purifications. The test was calibrated by recording the amount of pilus protein required to absorb a measurable amount of anti-pili antibody from antiserum. The amount of anti-pilus antibody in absorbed and unabsorbed serum was specifically measured by a Bact. nodosus K-agglutination test. The PAAT could be calibrated using any combination of crude or pure pili preparations and specific anti-pili serum or non-specific anti-Bact. nodosus serum. This had advantages over the use of radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA) techniques for measurement of pili because they require the use of highly purified reagents for calibration. The minimum quantity of pilus protein measurable by PAAT was 0.1 microgram per test which was similar in sensitivity to that reported for RIA and ELISA. The reagents used in PAAT were stable for at least six months. The amount of pilus protein per bacterium as measured by PAAT was directly proportional to the average number of pili per bacterium as measured by electron microscopy. The test was Bact. nodosus serotype specific.     

Quantitative test for Bacteroides nodosus pilus protein by an agglutination-absorption test

A pili agglutination-absorption test (PAAT) was developed for the quantitative measurement of pilus protein in cultures of Bacteroides nodosus and to quantify pili yields during purifications. The test was calibrated by recording the amount of pilus protein required to absorb a measurable amount of anti-pili antibody from antiserum. The amount of anti-pilus antibody in absorbed and unabsorbed serum was specifically measured by a Bact. nodosus K-agglutination test. The PAAT could be calibrated using any combination of crude or pure pili preparations and specific anti-pili serum or non-specific anti- Bact. nodosus serum. This had advantages over the use of radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA) techniques for measurement of pili because they require the use of highly purified reagents for calibration. The minimum quantity of pilus protein measurable by PAAT was 0.1 μg per test which was similar in sensitivity to that reported for RIA and ELISA. The reagents used in PAAT were stable for at least six months. The amount of pilus protein per bacterium as measured by PAAT was directly proportional to the average number of pili per bacterium as measured by electron microscopy. The test was Bact. nodosus serotype specific.     

The HKA test revisited: a maximum-likelihood-ratio test of the standard neutral model

We present a maximum-likelihood-ratio test of the standard neutral model, using multilocus data on polymorphism within species and divergence between species. The model is based on the Hudson-Kreitman-Aguade (HKA) test, but allows for an explicit test of selection at individual loci in a multilocus framework. We use coalescent simulations to show that the likelihood-ratio test statistic is conservative, particularly when the assumption of no recombination is violated. Application of the method to polymorphism data from 18 loci from a population of Arabidopsis lyrata provides significant evidence for a balanced polymorphism at a candidate locus thought to be linked to the centromere. The method is also applied to polymorphism data in maize, providing support for the hypothesis of directional selection on genes in the starch pathway.     

A socio-ecological test bed

A test bed simulation is described, composed of an individual-based simulation where species co-evolve on a 2D grid. This can generate, in a bottom-up manner, complex ecosystems of individuals. This test bed is designed so that the kind of ecological complexity observed is exhibited in order to be able to assess the longer-term complex dynamics that might occur if humans with different cultural characteristics are introduced. This is compared to Hubbell's ‘Neutral Theory’. A sensitivity analysis is shown. Then an example where “human agents” are introduced half way into the simulation is described. A comprehensive exploration of the impact of introducing ‘human-like’ agents is beyond the scope of this paper, but some indicative results shown, showing both the high impact humans have, but also some of the complexity of the human-ecosystem interaction.     

Comparison of agar dilution test,broth dilution test,and broth elution test for assaying susceptibility ofCandida spp

Biochemically and morphologically defined isolates ofCandida spp. were tested for their susceptibility to systemic antimycotics (5-flucytosine, ketoconazole, and miconazole). Three different susceptibility test techniques—the broth disc elution test, the agar dilution test, and the broth dilution test—were examined. The different assays were correlated with the broth dilution test at breakpoint concentrations. Reliability and practicability of the test systems described here were good, and the agreement with the broth dilution test was excellent. The elution test seems to be a valuable method for smaller laboratories, whereas the agar dilution test with antimycotic tablets is a particularly and comparatively inexpensive test for laboratories handling larger numbers of specimens.     

Influence of test duration on the sensitivity of the two-bottle choice test

The long-term two-bottle choice test is commonly used as a simple screen to examine the acceptance of taste solutions by rodents. As part of an investigation of factors influencing the sensitivity of the two-bottle choice test, we determined the extent to which test duration influenced test sensitivity. C57BL6/J and 129X1/SvJ mice received four series of eight two-bottle tests, with each test lasting 1, 2, 4 or 6 days. Each series involved sequential tests with water, 2 mM saccharin, 5 and 50 mM citric acid, 30 and 300 micro M quinine hydrochloride, 75 mM NaCl and 10% ethanol. There were significant differences between the strains in intake of saccharin, 5 and 50 mM citric acid, NaCl and ethanol in 4 and 6 day tests, but only saccharin and ethanol in 2 day tests, and 5 mM citric acid and ethanol in 1 day tests. To compare the sensitivity of the tests, we developed an analytical approach based on the comparison of deviations of individual 129X1/SvJ mice from the C57BL6/J strain mean. Our results suggest that to discriminate between strains or treatments when using 'standard' laboratory conditions and methods, 1 day tests are generally inadequate and 2 day tests are useful only if large effects are anticipated. Tests lasting 4 or 6 days are more sensitive, but conducting 6 day tests provides little additional benefit and sometimes is detrimental relative to conducting 4 day tests.     

A conservative test for multiple comparison based on highly correlated test statistics

In genetics, we often encounter a large number of highly correlated test statistics. The most famous conservative bound for multiple comparison is Bonferroni's bound, which is suitable when the test statistics are independent but not when the test statistics are highly correlated. This article proposes a new conservative bound that is easily calculated without multiple integration and is a good approximation when the test statistics are highly correlated. The performance of the proposed method is evaluated by simulation and real data analysis.     

Multivariate extensions of McNemar's test

This article considers global tests of differences between paired vectors of binomial probabilities, based on data from two dependent multivariate binary samples. Difference is defined as either an inhomogeneity in the marginal distributions or asymmetry in the joint distribution. For detecting the first type of difference, we propose a multivariate extension of McNemar's test and show that it is a generalized score test under a generalized estimating equations (GEE) approach. Univariate features such as the relationship between the Wald and score tests and the dropout of pairs with the same response carry over to the multivariate case and the test does not depend on the working correlation assumption among the components of the multivariate response. For sparse or imbalanced data, such as occurs when the number of variables is large or the proportions are close to zero, the test is best implemented using a bootstrap, and if this is computationally too complex, a permutation distribution. We apply the test to safety data for a drug, in which two doses are evaluated by comparing multiple responses by the same subjects to each one of them.     

Hypoosmotic test in equine spermatozoa

The aim of the study was to evaluate equine sperm membrane integrity using the hypoosmotic swelling (HOS) test and to correlate this test with different sperm parameters in raw and frozen thawed semen. The HOS solutions were made with fructose, sucrose, lactose and sodium citrate each at 300, 150, 100, 50 and 25 mosm. Maximum numbers of swollen spermatozoa were observed in solutions of fructose, sucrose and lactose each at 100, 50 and 25 mosm. Correlations between progressive motility, morphologically normal spermatozoa and the HOS test were r = 0.75 and r = 0.51 in raw semen and r = 0.26 and r = -0.22 in frozen-thawed semen. The correlation between HOS and percentage of intact membranes with the fluorescent stain was r = 0.32 in frozen-thawed semen. The HOS test is a simple and accessible method which could be used as a complement to routine equine semen analysis. It has the added advantages of being less susceptible to the immediate effects of cold shock and of evaluating individual spermatozoa rather than the population as a whole, as does progressive motility.     

Mantel test in population genetics

The comparison of genetic divergence or genetic distances, estimated by pairwise F_ST and related statistics, with geographical distances by Mantel test is one of the most popular approaches to evaluate spatial processes driving population structure. There have been, however, recent criticisms and discussions on the statistical performance of the Mantel test. Simultaneously, alternative frameworks for data analyses are being proposed. Here, we review the Mantel test and its variations, including Mantel correlograms and partial correlations and regressions. For illustrative purposes, we studied spatial genetic divergence among 25 populations of Dipteryx alata (“Baru”), a tree species endemic to the Cerrado, the Brazilian savannas, based on 8 microsatellite loci. We also applied alternative methods to analyze spatial patterns in this dataset, especially a multivariate generalization of Spatial Eigenfunction Analysis based on redundancy analysis. The different approaches resulted in similar estimates of the magnitude of spatial structure in the genetic data. Furthermore, the results were expected based on previous knowledge of the ecological and evolutionary processes underlying genetic variation in this species. Our review shows that a careful application and interpretation of Mantel tests, especially Mantel correlograms, can overcome some potential statistical problems and provide a simple and useful tool for multivariate analysis of spatial patterns of genetic divergence.