Abstract Motivation Combining the results of different experiments to exhibit complex patterns or to improve statistical power is a typical aim of data integration. The starting point of the statistical analysis often comes as a set of P-values resulting from previous analyses, that need to be combined flexibly to explore complex hypotheses, while guaranteeing a low proportion of false discoveries. Results We introduce the generic concept of composed hypothesis, which corresponds to an arbitrary complex combination of simple hypotheses. We rephrase the problem of testing a composed hypothesis as a classification task and show that finding items for which the composed null hypothesis is rejected boils down to fitting a mixture model and classifying the items according to their posterior probabilities. We show that inference can be efficiently performed and provide a thorough classification rule to control for type I error. The performance and the usefulness of the approach are illustrated in simulations and on two different applications. The method is scalable, does not require any parameter tuning, and provided valuable biological insight on the considered application cases. Availability and implementation The QCH methodology is available in the qch package hosted on CRAN. Additionally, R codes to reproduce the Einkorn example are available on the personal webpage of the first author: https://www6.inrae.fr/mia-paris/Equipes/Membres/Tristan-Mary-Huard. Supplementary information Supplementary data are available at Bioinformatics online.

MOTIVATION: De novo comparative metagenomics is one of the most straightforward ways to analyze large sets of metagenomic data. Latest methods use the fraction of shared k-mers to estimate genomic similarity between read sets. However, those methods, while extremely efficient, are still limited by computational needs for practical usage outside of large computing facilities. RESULTS: We present SimkaMin, a quick comparative metagenomics tool with low disk and memory footprints, thanks to an efficient data subsampling scheme used to estimate Bray-Curtis and Jaccard dissimilarities. One billion metagenomic reads can be analyzed in <3 min, with tiny memory (1.09 GB) and disk (≈0.3 GB) requirements and without altering the quality of the downstream comparative analyses, making of SimkaMin a tool perfectly tailored for very large-scale metagenomic projects. AVAILABILITY AND IMPLEMENTATION: https://github.com/GATB/simka. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Networks provide a natural yet statistically grounded way to depict and understand how a set of entities interact. However, in many situations interactions are not directly observed and the network needs to be reconstructed based on observations collected for each entity. Our work focuses on the situation where these observations consist of counts. A typical example is the reconstruction of an ecological network based on abundance data. In this setting, the abundance of a set of species is collected in a series of samples and/or environments and we aim at inferring direct interactions between the species. The abundances at hand can be, for example, direct counts of individuals (ecology of macro-organisms) or read counts resulting from metagenomic sequencing (microbial ecology). Whatever the approach chosen to infer such a network, it has to account for the peculiaraties of the data at hand. The first, obvious one, is that the data are counts, i.e. non continuous. Also, the observed counts often vary over many orders of magnitude and are more dispersed than expected under a simple model, such as the Poisson distribution. The observed counts may also result from different sampling efforts in each sample and/or for each entity, which hampers direct comparison. Furthermore, because the network is supposed to reveal only direct interactions, it is highly desirable to account for covariates describing the environment to avoid spurious edges. Many methods of network reconstruction from count data have been proposed. In the context of microbial ecology, most methods (SparCC, REBACCA, SPIEC-EASI, gCODA, BanOCC) rely on a two-step strategy: transform the counts to pseudo Gaussian observations using simple transforms before moving back to the setting of Gaussian Graphical Models, for which state of the art methods exist to infer the network, but only in a Gaussian world. In this work, we consider instead a full-fledged probabilistic model with a latent layer where the counts follow Poisson distributions, conditional to latent (hidden) Gaussian correlated variables. In this model, known as Poisson log-normal (PLN), the dependency structure is completely captured by the latent layer and we model counts, rather than transformations thereof. To our knowledge, the PLN framework is quite new and has only been used by two other recent methods (Mint and plnDAG) to reconstruct networks from count data. In this work, we use the same mathematical framework but adopt a different optimization strategy which alleviates the whole optimization process. We also fully exploit the connection between the PLN framework and generalized linear models to account for the peculiarities of microbiological data sets. The network inference step is done as usual by adding sparsity inducing constraints on the inverse covariance matrix of the latent Gaussian vector to select only the most important interactions between species. Unlike the usual Gaussian setting, the penalized likelihood is generally not tractable in this framework. We resort instead to a variational approximation for parameter inference and solve the corresponding optimization problem by alternating a gradient descent on the variational parameters and a graphical-Lasso step on the covariance matrix. We also select the sparsity parameter using the resampling-based StARS procedure. We show that the sparse PLN approach has better performance than existing methods on simulated datasets and that it extracts relevant signal from microbial ecology datasets. We also show that the inference scales to datasets made up of hundred of species and samples, in line with other methods in the field. In short, our contributions to the field are the following: we extend the use of PLN distributions in network inference by (i) accounting for covariates and offset and thus removing some spurious edges induced by confounding factors, (ii) accounting for different sampling effort to integrate data sets from different sources and thus infer interactions between different types of organisms (e.g. bacteria - fungi), (iii) developing an inference procedure based on the iterative optimization of a well defined objective function. Our objective function is a provable lower bound of the observed likelihood and our procedure accounts for the uncertainty associated with the estimation of the latent variable, unlike the algorithm presented in Mint and plnDAG.

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BACKGROUND: Detecting local correlations in expression between neighboring genes along the genome has proved to be an effective strategy to identify possible causes of transcriptional deregulation in cancer. It has been successfully used to illustrate the role of mechanisms such as copy number variation (CNV) or epigenetic alterations as factors that may significantly alter expression in large chromosomal regions (gene silencing or gene activation). RESULTS: The identification of correlated regions requires segmenting the gene expression correlation matrix into regions of homogeneously correlated genes and assessing whether the observed local correlation is significantly higher than the background chromosomal correlation. A unified statistical framework is proposed to achieve these two tasks, where optimal segmentation is efficiently performed using dynamic programming algorithm, and detection of highly correlated regions is then achieved using an exact test procedure. We also propose a simple and efficient procedure to correct the expression signal for mechanisms already known to impact expression correlation. The performance and robustness of the proposed procedure, called SegCorr, are evaluated on simulated data. The procedure is illustrated on cancer data, where the signal is corrected for correlations caused by copy number variation. It permitted the detection of regions with high correlations linked to epigenetic marks like DNA methylation. CONCLUSIONS: SegCorr is a novel method that performs correlation matrix segmentation and applies a test procedure in order to detect highly correlated regions in gene expression.

MOTIVATION: The spatial conformation of the chromosome has a deep influence on gene regulation and expression. Hi-C technology allows the evaluation of the spatial proximity between any pair of loci along the genome. It results in a data matrix where blocks corresponding to (self-)interacting regions appear. The delimitation of such blocks is critical to better understand the spatial organization of the chromatin. From a computational point of view, it results in a 2D segmentation problem. RESULTS: We focus on the detection of cis-interacting regions, which appear to be prominent in observed data. We define a block-wise segmentation model for the detection of such regions. We prove that the maximization of the likelihood with respect to the block boundaries can be rephrased in terms of a 1D segmentation problem, for which the standard dynamic programming applies. The performance of the proposed methods is assessed by a simulation study on both synthetic and resampled data. A comparative study on public data shows good concordance with biologically confirmed regions. AVAILABILITY AND IMPLEMENTATION: The HiCseg R package is available from the Comprehensive R Archive Network and from the Web page of the corresponding author.

MOTIVATION: High-density oligonucleotide tiling array technology holds the promise of a better description of the complexity and the dynamics of transcriptional landscapes. In organisms such as bacteria and yeasts, transcription can be measured on a genome-wide scale with a resolution >25 bp. The statistical models currently used to handle these data remain however very simple, the most popular being the piecewise constant Gaussian model with a fixed number of breakpoints. RESULTS: This article describes a new methodology based on a hidden Markov model that embeds the segmentation of a continuous-valued signal in a probabilistic setting. For a computationally affordable cost, this framework (i) alleviates the difficulty of choosing a fixed number of breakpoints, and (ii) permits retrieving more information than a unique segmentation by giving access to the whole probability distribution of the transcription profile. Importantly, the model is also enriched and accounts for subtle effects such as signal 'drift' and covariates. Relevance of this framework is demonstrated on a Bacillus subtilis dataset. AVAILABILITY: A software is distributed under the GPL.

BACKGROUND: The use of current high-throughput genetic, genomic and post-genomic data leads to the simultaneous evaluation of a large number of statistical hypothesis and, at the same time, to the multiple-testing problem. As an alternative to the too conservative Family-Wise Error-Rate (FWER), the False Discovery Rate (FDR) has appeared for the last ten years as more appropriate to handle this problem. However one drawback of FDR is related to a given rejection region for the considered statistics, attributing the same value to those that are close to the boundary and those that are not. As a result, the local FDR has been recently proposed to quantify the specific probability for a given null hypothesis to be true. RESULTS: In this context we present a semi-parametric approach based on kernel estimators which is applied to different high-throughput biological data such as patterns in DNA sequences, genes expression and genome-wide association studies. CONCLUSION: The proposed method has the practical advantages, over existing approaches, to consider complex heterogeneities in the alternative hypothesis, to take into account prior information (from an expert judgment or previous studies) by allowing a semi-supervised mode, and to deal with truncated distributions such as those obtained in Monte-Carlo simulations. This method has been implemented and is available through the R package kerfdr via the CRAN or at (http://stat.genopole.cnrs.fr/software/kerfdr).

BACKGROUND: As biological networks often show complex topological features, mathematical methods are required to extract meaningful information. Clustering methods are useful in this setting, as they allow the summary of the network's topology into a small number of relevant classes. Different strategies are possible for clustering, and in this article we focus on a model-based strategy that aims at clustering nodes based on their connectivity profiles. RESULTS: We present MixNet, the first publicly available computer software that analyzes biological networks using mixture models. We apply this method to various networks such as the E. coli transcriptional regulatory network, the macaque cortex network, a foodweb network and the Buchnera aphidicola metabolic network. This method is also compared with other approaches such as module identification or hierarchical clustering. CONCLUSION: We show how MixNet can be used to extract meaningful biological information, and to give a summary of the networks topology that highlights important biological features. This approach is powerful as MixNet is adaptive to the network under study, and finds structural information without any a priori on the structure that is investigated. This makes MixNet a very powerful tool to summarize and decipher the connectivity structure of biological networks.

Compression and variable selection are two classical strategies to deal with large-dimension data sets in classification. We propose an alternative strategy, called aggregation, which consists of a clustering step of redundant variables and a compression step within each group. We develop a statistical framework to define tailored aggregation methods that can be combined with selection methods to build reliable classifiers that benefit from the information contained in redundant variables. Two algorithms are proposed for ordered and nonordered variables, respectively. Applications to the kNN and CART algorithms are presented.

BACKGROUND: Finding over- or under-represented motifs in biological sequences is now a common task in genomics. Thanks to p-value calculation for motif counts, exceptional motifs are identified and represent candidate functional motifs. The present work addresses the related question of comparing the exceptionality of one motif in two different sequences. Just comparing the motif count p-values in each sequence is indeed not sufficient to decide if this motif is significantly more exceptional in one sequence compared to the other one. A statistical test is required. RESULTS: We develop and analyze two statistical tests, an exact binomial one and an asymptotic likelihood ratio test, to decide whether the exceptionality of a given motif is equivalent or significantly different in two sequences of interest. For that purpose, motif occurrences are modeled by Poisson processes, with a special care for overlapping motifs. Both tests can take the sequence compositions into account. As an illustration, we compare the octamer exceptionalities in the Escherichia coli K-12 backbone versus variable strain-specific loops. CONCLUSION: The exact binomial test is particularly adapted for small counts. For large counts, we advise to use the likelihood ratio test which is asymptotic but strongly correlated with the exact binomial test and very simple to use.

MOTIVATION: Identifying differentially regulated genes in experiments comparing two experimental conditions is often a key step in the microarray data analysis process. Many different approaches and methodological developments have been put forward, yet the question remains open. RESULTS: Varmixt is a powerful and efficient novel methodology for this task. It is based on a flexible and realistic variance modelling strategy. It compares favourably with other popular techniques (standard t-test, SAM and Cyber-T). The relevance of the approach is demonstrated with real-world and simulated datasets. The analysis strategy was successfully applied to both a 'two-colour' cDNA microarray and an Affymetrix Genechip. Strong control of false positive and false negative rates is proven in large simulation studies. AVAILABILITY: The R package is freely available at http://www.inapg.inra.fr/ens_rech/mathinfo/recherche/mathematique/outil.html CONTACT: [email protected] SUPPLEMENTARY INFORMATION: http://www.inapg.inra.fr/ens_rech/mathinfo/recherche/mathematique/outil.html.

Following publication of this work [1], the authors became aware that Local FDR has been originally defined by Efron & al. (2001) in a mixture model framework [2].

BACKGROUND: Analysis of variance is a powerful approach to identify differentially expressed genes in a complex experimental design for microarray and macroarray data. The advantage of the anova model is the possibility to evaluate multiple sources of variation in an experiment. RESULTS: AnovArray is a package implementing ANOVA for gene expression data using SAS statistical software. The originality of the package is 1) to quantify the different sources of variation on all genes together, 2) to provide a quality control of the model, 3) to propose two models for a gene's variance estimation and to perform a correction for multiple comparisons. CONCLUSION: AnovArray is freely available at http://www-mig.jouy.inra.fr/stat/AnovArray and requires only SAS statistical software.

BACKGROUND: Microarray-CGH experiments are used to detect and map chromosomal imbalances, by hybridizing targets of genomic DNA from a test and a reference sample to sequences immobilized on a slide. These probes are genomic DNA sequences (BACs) that are mapped on the genome. The signal has a spatial coherence that can be handled by specific statistical tools. Segmentation methods seem to be a natural framework for this purpose. A CGH profile can be viewed as a succession of segments that represent homogeneous regions in the genome whose BACs share the same relative copy number on average. We model a CGH profile by a random Gaussian process whose distribution parameters are affected by abrupt changes at unknown coordinates. Two major problems arise: to determine which parameters are affected by the abrupt changes (the mean and the variance, or the mean only), and the selection of the number of segments in the profile. RESULTS: We demonstrate that existing methods for estimating the number of segments are not well adapted in the case of array CGH data, and we propose an adaptive criterion that detects previously mapped chromosomal aberrations. The performances of this method are discussed based on simulations and publicly available data sets. Then we discuss the choice of modeling for array CGH data and show that the model with a homogeneous variance is adapted to this context. CONCLUSIONS: Array CGH data analysis is an emerging field that needs appropriate statistical tools. Process segmentation and model selection provide a theoretical framework that allows precise biological interpretations. Adaptive methods for model selection give promising results concerning the estimation of the number of altered regions on the genome.

Studying the distribution of a motif along sequences may help to understand its biological function, or to detect regions of interest. A statistical model is needed to assess the significancy of the observed distribution. We propose an heterogenous compound Poisson process to model the possibility of overlap between occurrences and some heterogeneity of the sequence a priori known. The parameters estimation procedure is described and tests of homogenous sub-models are proposed. We also consider the detection of rich regions using either cumulated distances or moving intervals, via an homogenization technique. Illustrations of the method are given with applications to bacterial genomes.

BACKGROUND: Thousands of genes in a genomewide data set are tested against some null hypothesis, for detecting differentially expressed genes in microarray experiments. The expected proportion of false positive genes in a set of genes, called the False Discovery Rate (FDR), has been proposed to measure the statistical significance of this set. Various procedures exist for controlling the FDR. However the threshold (generally 5%) is arbitrary and a specific measure associated with each gene would be worthwhile. RESULTS: Using process intensity estimation methods, we define and give estimates of the local FDR, which may be considered as the probability for a gene to be a false positive. After a global assessment rule controlling the false positive error, the local FDR is a valuable guideline for deciding wether a gene is differentially expressed. The interest of the method is illustrated on three well known data sets. A R routine for computing local FDR estimates from p-values is available at http://www.inapg.fr/ens_rech/mathinfo/recherche/mathematique/outil.html. CONCLUSIONS: The local FDR associated with each gene measures the probability that it is a false positive. It gives the opportunity to compute the FDR of any given group of clones (of the same gene) or genes pertaining to the same regulation network or the same chromosomic region.

BACKGROUND: Microarray data must be normalized because they suffer from multiple biases. We have identified a source of spatial experimental variability that significantly affects data obtained with Cy3/Cy5 spotted glass arrays. It yields a periodic pattern altering both signal (Cy3/Cy5 ratio) and intensity across the array. RESULTS: Using the variogram, a geostatistical tool, we characterized the observed variability, called here the spotting effect because it most probably arises during steps in the array printing procedure. CONCLUSIONS: The spotting effect is not appropriately corrected by current normalization methods, even by those addressing spatial variability. Importantly, the spotting effect may alter differential and clustering analysis.