Stefan Zoller

CodonPhyML: Fast Maximum Likelihood Phylogeny Estimation under Codon Substitution Models

Markov models of codon substitution naturally incorporate the structure of the genetic code and the selection intensity at the protein level, providing a more realistic representation of protein-coding sequences compared with nucleotide or amino acid models. Thus, for protein-coding genes, phylogenetic inference is expected to be more accurate under codon models. So far, phylogeny reconstruction under codon models has been elusive due to computational difficulties of dealing with high dimension matrices. Here, we present a fast maximum likelihood (ML) package for phylogenetic inference, CodonPhyML offering hundreds of different codon models, the largest variety to date, for phylogeny inference by ML. CodonPhyML is tested on simulated and real data and is shown to offer excellent speed and convergence properties. In addition, CodonPhyML includes most recent fast methods for estimating phylogenetic branch supports and provides an integral framework for models selection, including amino acid and DNA models.

Population genomic footprints of selection and associations with climate in natural populations of Arabidopsis halleri from the Alps

Natural genetic variation is essential for the adaptation of organisms to their local environment and to changing environmental conditions. Here, we examine genomewide patterns of nucleotide variation in natural populations of the outcrossing herb Arabidopsis halleri and associations with climatic variation among populations in the Alps. Using a pooled population sequencing (Pool‐Seq) approach, we discovered more than two million SNPs in five natural populations and identified highly differentiated genomic regions and SNPs using FST‐based analyses. We tested only the most strongly differentiated SNPs for associations with a nonredundant set of environmental factors using partial Mantel tests to identify topo‐climatic factors that may underlie the observed footprints of selection. Possible functions of genes showing signatures of selection were identified by Gene Ontology analysis. We found 175 genes to be highly associated with one or more of the five tested topo‐climatic factors. Of these, 23.4% had unknown functions. Genetic variation in four candidate genes was strongly associated with site water balance and solar radiation, and functional annotations were congruent with these environmental factors. Our results provide a genomewide perspective on the distribution of adaptive genetic variation in natural plant populations from a highly diverse and heterogeneous alpine environment.

Validation of SNP Allele frequencies determined by pooled next-generation sequencing in natural populations of a non-model plant species

Sequencing of pooled samples (Pool-Seq) using next-generation sequencing technologies has become increasingly popular, because it represents a rapid and cost-effective method to determine allele frequencies for single nucleotide polymorphisms (SNPs) in population pools. Validation of allele frequencies determined by Pool-Seq has been attempted using an individual genotyping approach, but these studies tend to use samples from existing model organism databases or DNA stores, and do not validate a realistic setup for sampling natural populations. Here we used pyrosequencing to validate allele frequencies determined by Pool-Seq in three natural populations of Arabidopsis halleri (Brassicaceae). The allele frequency estimates of the pooled population samples (consisting of 20 individual plant DNA samples) were determined after mapping Illumina reads to (i) the publicly available, high-quality reference genome of a closely related species (Arabidopsis thaliana) and (ii) our own de novo draft genome assembly of A. halleri. We then pyrosequenced nine selected SNPs using the same individuals from each population, resulting in a total of 540 samples. Our results show a highly significant and accurate relationship between pooled and individually determined allele frequencies, irrespective of the reference genome used. Allele frequencies differed on average by less than 4%. There was no tendency that either the Pool-Seq or the individual-based approach resulted in higher or lower estimates of allele frequencies. Moreover, the rather high coverage in the mapping to the two reference genomes, ranging from 55 to 284x, had no significant effect on the accuracy of the Pool-Seq. A resampling analysis showed that only very low coverage values (below 10-20x) would substantially reduce the precision of the method. We therefore conclude that a pooled re-sequencing approach is well suited for analyses of genetic variation in natural populations.

Transcriptome responses to aluminum stress in roots of aspen (Populus tremula)

BackgroundIonic aluminum (mainly Al3+) is rhizotoxic and can be present in acid soils at concentrations high enough to inhibit root growth. Many forest tree species grow naturally in acid soils and often tolerate high concentrations of Al. Previously, we have shown that aspen (Populus tremula) releases citrate and oxalate from roots in response to Al exposure. To obtain further insights into the root responses of aspen to Al, we investigated root gene expression at Al conditions that inhibit root growth.ResultsTreatment of the aspen roots with 500 μM Al induced a strong inhibition of root growth within 6 h of exposure time. The root growth subsequently recovered, reaching growth rates comparable to that of control plants. Changes in gene expression were determined after 6 h, 2 d, and 10 d of Al exposure. Replicated transcriptome analyses using the Affymetrix poplar genome array revealed a total of 175 significantly up-regulated and 69 down-regulated genes, of which 70% could be annotated based on Arabidopsis genome resources. Between 6 h and 2 d, the number of responsive genes strongly decreased from 202 to 26, and then the number of changes remained low. The responses after 6 h were characterized by genes involved in cell wall modification, ion transport, and oxidative stress. Two genes with prolonged induction were closely related to the Arabidopsis Al tolerance genes ALS3 (for Al sensitive 3) and MATE (for multidrug and toxin efflux protein, mediating citrate efflux). Patterns of expression in different plant organs and in response to Al indicated that the two aspen genes are homologs of the Arabidopsis ALS3 and MATE.ConclusionExposure of aspen roots to Al results in a rapid inhibition of root growth and a large change in root gene expression. The subsequent root growth recovery and the concomitant reduction in the number of responsive genes presumably reflect the success of the roots in activating Al tolerance mechanisms. The aspen genes ALS3 and MATE may be important components of these mechanisms.

A genomic approach to examine the complex evolution of laurasiatherian mammals.

Recent phylogenomic studies have failed to conclusively resolve certain branches of the placental mammalian tree, despite the evolutionary analysis of genomic data from 32 species. Previous analyses of single genes and retroposon insertion data yielded support for different phylogenetic scenarios for the most basal divergences. The results indicated that some mammalian divergences were best interpreted not as a single bifurcating tree, but as an evolutionary network. In these studies the relationships among some orders of the super-clade Laurasiatheria were poorly supported, albeit not studied in detail. Therefore, 4775 protein-coding genes (6,196,263 nucleotides) were collected and aligned in order to analyze the evolution of this clade. Additionally, over 200,000 introns were screened in silico, resulting in 32 phylogenetically informative long interspersed nuclear elements (LINE) insertion events. The present study shows that the genome evolution of Laurasiatheria may best be understood as an evolutionary network. Thus, contrary to the common expectation to resolve major evolutionary events as a bifurcating tree, genome analyses unveil complex speciation processes even in deep mammalian divergences. We exemplify this on a subset of 1159 suitable genes that have individual histories, most likely due to incomplete lineage sorting or introgression, processes that can make the genealogy of mammalian genomes complex. These unexpected results have major implications for the understanding of evolution in general, because the evolution of even some higher level taxa such as mammalian orders may sometimes not be interpreted as a simple bifurcating pattern.

Detection and genetic characterisation of a novel mycovirus in Hymenoscyphus fraxineus , the causal agent of ash dieback

The ascomycete fungus Hymenoscyphus fraxineus (synonym: Hymenoscyphuspseudoalbidus, basionym: Chalara fraxinea) is a new invasive pathogen causing severe dieback of ash trees (Fraxinus spp.) in Europe. The disease was first recorded in the 1990s in Eastern Poland and Lithuania and has meanwhile spread across the continent and to Great Britain. Mycoviruses are commonly found in all major groups of plant pathogenic fungi. Some of these viruses can cause debilitating disease or reduce virulence in their fungal host and thereby can act as biological control agents (e.g. hypoviruses in the chestnut blight fungus). The main aim of our study is to identify mycoviruses in the ash dieback pathogen and to describe their phylogenetic position. Using RNAseq we could identify a viral sequence in two fungal isolates. Amino acid sequence comparison suggests that the detected mycovirus is a putative new member of the genus Mitovirus (family Narnaviridae), and we suggest naming it Hymenoscyphus fraxineus mitovirus 1 (HfMV1). The novel mitovirus is highly variable amongst European isolates and seems to diverge rapidly. This is very promising for the future search of a strain adequate for biological control measures.

Rust fungi on Annonaceae II: the genus Dasyspora Berk. & M.A. Curtis

Dasyspora gregaria, the single species of the allegedly monotypic rust genus Dasyspora (Basidiomycota, Pucciniales), was investigated by light microscopy and DNA sequencing (ITS1–5.8S–ITS2 region, partial LSU and SSU of the nuclear rDNA, mt cytochrome oxidase subunit 3). Both methods indicated that D. gregaria is not a single species but can be split in 11 distinct taxa, each of which appear confined to a single Xylopia species (Annonaceae) host. Herein nine of these are described as new. Both the phylogenetic analyses and morphology show that the species are grouped into two main clades designated Dasyspora gregaria and D. winteri. The first comprises D. gregaria, the type species of the genus, which is restricted to X. cayennensis, two new species on X. aromatica, D. segregaria from northern South America and D. echinata from Brazil. The second clade is formed by D. winteri, recombined from Puccinia winteri on X. sericea, and the new species D. amazonica on X. amazonica, D. emarginatae on X. emarginata, D. frutescentis on X. frutescens, D. ferrugineae on X. frutescens var. ferruginea, D. guianensis on X. benthamii, D. mesoamericana on X. frutescens, and D. nitidae on X. nitida. Dasyspora frutescentis and D. mesoamericana were not clearly distinguishable by their morphology and host associations but differed from another in their sequences and geographic distributions. They are considered cryptic species. An identification key and the distributions are given for all recognized species. Along with molecular data we discuss the systematic position of Dasyspora in the Pucciniales.

Improving Phylogenetic Inference with a Semiempirical Amino Acid Substitution Model

Amino acid substitution matrices describe the rates by which amino acids are replaced during evolution. In contrast to nucleotide or codon models, amino acid substitution matrices are in general parameterless and empirically estimated, probably because there is no obvious parametrization for amino acid substitutions. Principal component analysis has previously been used to improve codon substitution models by empirically finding the most relevant parameters. Here, we apply the same method to amino acid substitution matrices, leading to a semiempirical substitution model that can adjust the transition rates to the protein sequences under investigation. Our new model almost invariably achieves the best likelihood values in large-scale comparisons with established amino acid substitution models (JTT, WAG, and LG). In particular for longer alignments, these likelihood gains are considerably larger than what could be expected from simply having more parameters. The application of our model differs from that of mixture models (such as UL2 or UL3), as we optimize one rate matrix per alignment, whereas mixture models apply the variation per alignments site. This makes our model computationally more efficient, while the performance is comparable to that of UL3. Applied to the phylogenetic problem of the origin of placental mammals, our new model and the UL3 mixed model are the only ones of the tested models that cluster Afrotheria and Xenarthra into a clade called Atlantogenata, which would be in correspondence with recent findings using more sophisticated phylogenetic methods.

MAGMA: analysis of two-channel microarrays made easy

The web application MAGMA provides a simple and intuitive interface to identify differentially expressed genes from two-channel microarray data. While the underlying algorithms are not superior to those of similar web applications, MAGMA is particularly user friendly and can be used without prior training. The user interface guides the novice user through the most typical microarray analysis workflow consisting of data upload, annotation, normalization and statistical analysis. It automatically generates R-scripts that document MAGMAs entire data processing steps, thereby allowing the user to regenerate all results in his local R installation. The implementation of MAGMA follows the model-view-controller design pattern that strictly separates the R-based statistical data processing, the web-representation and the application logic. This modular design makes the application flexible and easily extendible by experts in one of the fields: statistical microarray analysis, web design or software development. State-of-the-art Java Server Faces technology was used to generate the web interface and to perform user input processing. MAGMAs object-oriented modular framework makes it easily extendible and applicable to other fields and demonstrates that modern Java technology is also suitable for rather small and concise academic projects. MAGMA is freely available at www.magma-fgcz.uzh.ch.

Signatures of local adaptation in candidate genes of oaks ( Quercus spp.) with respect to present and future climatic conditions

Testing how populations are locally adapted and predicting their response to their future environment is of key importance in view of climate change. Landscape genomics is a powerful approach to investigate genes and environmental factors involved in local adaptation. In a pooled amplicon sequencing approach of 94 genes in 71 populations, we tested whether >3500 single nucleotide polymorphisms (SNPs) in the three most common oak species in Switzerland (Quercus petraea, Q. pubescens, Q. robur) show an association with abiotic factors related to local topography, historical climate and soil characteristics. In the analysis including all species, the most frequently associated environmental factors were those best describing the habitats of the species. In the species‐specific analyses, the most important environmental factors and associated SNPs greatly differed among species. However, we identified one SNP and seven genes that were associated with the same environmental factor across all species. We finally used regressions of allele frequencies of the most strongly associated SNPs along environmental gradients to predict the risk of nonadaptedness (RONA), which represents the average change in allele frequency at climate‐associated loci theoretically required to match future climatic conditions. RONA is considerable for some populations and species (up to 48% in single populations) and strongly differs among species. Given the long generation time of oaks, some of the required allele frequency changes might not be realistic to achieve based on standing genetic variation. Hence, future adaptedness requires gene flow or planting of individuals carrying beneficial alleles from habitats currently matching future climatic conditions.