IMP: Imperial Metagenomics Pipeline for high-throughput sequence data

We have developed an in-house pipeline for the processing and analyses of sequence data generated during Illumina technology-based metagenomic studies of the human gut microbiota. Each component of the pipeline has been selected following comparative analysis of available tools; however, the modular nature of software facilitates replacement of any individual component with an alternative should a better tool become available in due course. The pipeline consists of quality analysis and trimming followed by taxonomic filtering of sequence data allowing reads associated with samples to be binned according to whether they represent human, prokaryotic (bacterial/archaeal), viral, parasite, fungal or plant DNA. Viral, parasite, fungal and plant DNA can be assigned to species level on a presence/absence basis, allowing – for example – identification of dietary intake of plant-based foodstuffs and their derivatives. Prokaryotic DNA is subject to taxonomic and functional analyses, with assignment to taxonomic hierarchies (kingdom, class, order, family, genus, species, strain/subspecies) and abundance determination. After de novo assembly of sequence reads, genes within samples are predicted and used to build a non-redundant catalogue of genes. From this catalogue, per-sample gene abundance can be determined after normalization of data based on gene length. Functional annotation of genes is achieved through mapping of gene clusters against KEGG proteins, and InterProScan. The pipeline is undergoing validation using the human faecal metagenomic data of Qin et al. (2014, Nature 513, 59–64). Outputs from the pipeline allow development of tools for the integration of metagenomic and metabolomic data, moving metagenomic studies beyond determination of gene richness and representation towards microbial-metabolite mapping. There is scope to improve the outputs from viral, parasite, fungal and plant DNA analyses, depending on the depth of sequencing associated with samples. The pipeline can easily be adapted for the analyses of environmental and non-human animal samples, and for use with data generated via non-Illumina sequencing platforms.

Mitochondrial sequence data and Dipsacales phylogeny: Mixed models, partitioned Bayesian analyses, and model selection

Phylogenetic analyses of chloroplast DNA sequences, morphology, and combined data have provided consistent support for many of the major branches within the angiosperm, clade Dipsacales. Here we use sequences from three mitochondrial loci to test the existing broad scale phylogeny and in an attempt to resolve several relationships that have remained uncertain. Parsimony, maximum likelihood, and Bayesian analyses of a combined mitochondrial data set recover trees broadly consistent with previous studies, although resolution and support are lower than in the largest chloroplast analyses. Combining chloroplast and mitochondrial data results in a generally well-resolved and very strongly supported topology but the previously recognized problem areas remain. To investigate why these relationships have been difficult to resolve we conducted a series of experiments using different data partitions and heterogeneous substitution models. Usually more complex modeling schemes are favored regardless of the partitions recognized but model choice had little effect on topology or support values. In contrast there are consistent but weakly supported differences in the topologies recovered from coding and non-coding matrices. These conflicts directly correspond to relationships that were poorly resolved in analyses of the full combined chloroplast-mitochondrial data set. We suggest incongruent signal has contributed to our inability to confidently resolve these problem areas. (c) 2007 Elsevier Inc. All rights reserved.

CopywriteR: DNA copy number detection from off-target sequence data.

Current methods for detection of copy number variants (CNV) and aberrations (CNA) from targeted sequencing data are based on the depth of coverage of captured exons. Accurate CNA determination is complicated by uneven genomic distribution and non-uniform capture efficiency of targeted exons. Here we present CopywriteR, which eludes these problems by exploiting 'off-target' sequence reads. CopywriteR allows for extracting uniformly distributed copy number information, can be used without reference, and can be applied to sequencing data obtained from various techniques including chromatin immunoprecipitation and target enrichment on small gene panels. CopywriteR outperforms existing methods and constitutes a widely applicable alternative to available tools.

A Petrov-Galerkin method for a singularly perturbed ordinary differential equation with non-smooth data

In this paper, a singularly perturbed ordinary differential equation with non-smooth data is considered. The numerical method is generated by means of a Petrov-Galerkin finite element method with the piecewise-exponential test function and the piecewise-linear trial function. At the discontinuous point of the coefficient, a special technique is used. The method is shown to be first-order accurate and singular perturbation parameter uniform convergence. Finally, numerical results are presented, which are in agreement with theoretical results.

Phylogeography of the pharaoh cuttle Sepia pharaonis based on partial mitochondrial 16S sequence data

The pharaoh cuttle Sepia pharaonis Ehrenberg, 1831 (Mollusca: Cephalopoda: Sepiida) is a broadly distributed species of substantial fisheries importance found from east Africa to southern Japan. Little is known about S. pharaonis phylogeography, but evidence from morphology and reproductive biology suggests that Sepia pharaonis is actually a complex of at least three species. To evaluate this possibility, we collected tissue samples from Sepia pharaonis from throughout its range. Phylogenetic analyses of partial mitochondrial 16S sequences from these samples reveal five distinct clades: a Gulf of Aden/Red Sea clade, a northern Australia clade, a Persian Gulf/Arabian Sea clade, a western Pacific clade (Gulf of Thailand and Taiwan) and an India/Andaman Sea clade. Phylogenetic analyses including several Sepia species show that S. pharaonis sensu lato may not be monophyletic. We suggest that "S. pharaonis" may consist of up to five species, but additional data will be required to fully clarify relationships within the S. pharaonis complex.

Phylogenetic relationships of graminicolous downy mildews based on cox2 sequence data

Graminicolous downy mildews (GDM) are an understudied, yet economically important, group of plant pathogens, which are one of the major constraints to poaceous crops in the tropics and subtropics. Here we present a first molecular phylogeny based on cox2 sequences comprising all genera of the GDM currently accepted, with both lasting (Graminivora, Poakatesthia, and Viennotia) and evanescent (Peronosclerospora, Sclerophthora, and Sclerospora) sporangiophores. In addition, all other downy mildew genera currently accepted, as well as a representative sample of other oomycete taxa, have been included. It was shown that all genera of the GDM have had a long, independent evolutionary history, and that the delineation between Peronosclerospora and Sclerospora is correct. Sclerophthora was found to be a particularly divergent taxon nested within a paraphyletic Phytophthora, but without support. The results confirm that the placement of Peronosclerospora and Sclerospora in the Saprolegniomycetidae is incorrect. Sclerophthora is not closely related to Pachymetra of the family Verrucalvaceae, and also does not belong to the Saprolegniomycetidae, but shows close affinities to the Peronosporaceae. In addition, all GDM are interspersed throughout the Peronosporaceae s lat., suggesting that a separate family for the Sclerosporaceae might not be justified.